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Master

Affiliation (Master)

  • Faculty of Engineering　Mechanical and Aerospace Engineering　Human and Mechanical Systems

Affiliation (Master)

  • Faculty of Engineering　Mechanical and Aerospace Engineering　Human and Mechanical Systems

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Profile and Settings

Profile and Settings

  • Name (Japanese)

    Ohashi

  • Name (Kana)

    Toshiro

  • Name

    201301027932882821

Alternate Names

Achievement

Research Interests

  • バイオメカニクス   細胞骨格   内皮細胞   細胞力学応答   血管内皮細胞   有限要素法   微細加工技術   力学伝達経路   マイクロマシニング   細胞牽引力   平滑筋細胞   近接場光   バイオチップ   シグナル伝達   バイオMEMS   弾性係数   ソフトリソグラフィー   カルシウムイオン   FRAP   非線形弾性   マクロウェル   遊走性   細胞接着制御   流れ負荷   生物フォトン   リモデリング   動脈硬化症   マクロフルディクス   共存培養血管   遺伝子導入技術   

Research Areas

  • Life sciences / Biomaterials
  • Life sciences / Biomedical engineering
  • Informatics / Mechanics and mechatronics
  • Informatics / Robotics and intelligent systems
  • Manufacturing technology (mechanical, electrical/electronic, chemical engineering) / Machine materials and mechanics

Research Experience

  • 2009/04 - Today Hokkaido University
  • 2002/10 - 2009/03 Tohoku University Graduate School of Engineering
  • 2008/04 - 2008/10 Royal Institute of Technology Visiting researcher
  • 2004/03 - 2005/03 Queen Mary University of London Visiting researcher
  • 1994/04 - 2002/09 Tohoku University Graduate School of Engineering

Education

  •        - 1994  University of Tsukuba  Master's Program in Science and Engineering
  •        - 1991  University of Tsukuba

Committee Memberships

  • 2022/07 - Today   World Council of Biomechanics   Secretary
  • 2020/12 - Today   Engineering Academy of Japan   Fellow
  • 2018 - Today   World Council of Biomechanics   Council Member
  • 2010/04 - Today   Asian-Pacific Association for Biomechanics   Executive Board Member

Awards

  • 2008 日本機械学会賞（論文）
  • 2004 2003年度日本機械学会バイオエンジニアリング部門第12回瀬口賞
  • 2004 日本エム・イー学会第18回日本エム・イー学会秋季大会研究奨励賞
  • 2002 Outstanding Paper Award at the International Congress on Biological and Medical Engineering
  • 2002 Finalist for a Calgary Award at the IV World Congress of Biomechanics
  • 2002 日本エム・イー学会平成13年度科学新聞賞・研究奨励賞・阪本研究刊行助成賞・阿部賞
  • 2001 Award for Best Poster in the Category Cell and Molecular Mechanics
  • 2000 Outstanding paper award
  • 2000 日本機械学会賞（論文）

Published Papers

  • Mazlee Bin Mazalan, Ryota Toyohara, Toshiro Ohashi
    International Journal of Precision Engineering and Manufacturing 25 (7) 1525 - 1538 2234-7593 2024/05/27
  • Ryota Toyohara, Toshiro Ohashi
    Clinical Biomechanics 114 106233 - 106233 0268-0033 2024/04
  • Ryota TOYOHARA, Daisuke KUROSAWA, Niels HAMMER, Toshiro OHASHI
    Journal of Biomechanical Science and Engineering 2024
  • Ryota Toyohara, Niels Hammer, Toshiro Ohashi
    Bio-medical materials and engineering 2023/07/31 
    BACKGROUND: The human sacroiliac joint (SIJ) in vivo is exposed to compressive and shearing stress environment, given the joint lines are almost parallel to the direction of gravity. The SIJ supports efficient bipedal walking. Unexpected or unphysiological, repeated impacts are believed to cause joint misalignment and result in SIJ pain. In the anterior compartment of the SIJ being synovial, the articular surface presents fine irregularities, potentially restricting the motion of the joints. OBJECTIVE: To clarify how the SIJ articular surface affects the resistance of the motion under physiological loading. METHODS: SIJ surface models were created based on computed tomography data of three patients and subsequently 3D printed. Shear resistance was measured in four directions and three combined positions using a customized setup. In addition, repositionability of SIJs was investigated by unloading a shear force. RESULTS: Shear resistance of the SIJ was the highest in the inferior direction. It changed depending on the direction of the shear and the alignment position of the articular surface. CONCLUSION: SIJ articular surface morphology is likely designed to accommodate upright bipedal walking. Joint misalignment may in consequence increase the risk of subluxation.
  • Mohamad Anis Bin RAMLAN, Ryota TOYOHARA, Toshiro OHASHI
    Journal of Biomechanical Science and Engineering 18 (4) 23 - 00536 2023
  • Ryota Toyohara, Takahiro Hiramukai, Daisuke Kurosawa, Niels Hammer, Toshiro Ohashi
    Bio-medical materials and engineering 2022/12/08 
    BACKGROUND: Pain related to the sacroiliac joint (SIJ) accounts for low back pain in 15%-30% of patients. One of the most common treatment options is the use of pelvic belts. Various types of pelvic belts exist; however, the mechanisms underlying treatment and their effectiveness remain unclear to date. OBJECTIVE: To analyze stress distribution in the pelvis when a pelvic rubber belt or a padded pelvic belt is applied, to assess the effectiveness of treatment from a numerical biomechanical perspective. METHODS: The pressure distribution at the pelvic belts was measured using a device and subsequently modeled with the finite element method of a pelvis with soft tissues. The stress environment when wearing a pelvic belt in a double-leg stance was simulated. RESULTS: With the application of pelvic belts, the innominate bone rotated outward, which was termed an out-flare. This caused the SIJ to compress and cause reduction in sacrotuberous, sacrospinous, interosseous, and posterior sacroiliac ligament loading. Padded pelvic belts decreased the SIJ displacement to a greater extent than in pelvic rubber belts. CONCLUSION: Pelvic belts aid in compressing the SIJ and reduce its mobility.
  • Ryota Toyohara, Ayumi Kaneuji, Noriyuki Takano, Daisuke Kurosawa, Niels Hammer, Toshiro Ohashi
    Scientific Reports 12 (1) 14500 - 14500 2022/08/25 [Refereed]
     
    In acetabular dysplasia, the cartilaginous roof on the acetabular side does not fully cover the femoral head, which may lead to abnormal stress distribution in both the femoral head and pelvis. These stress changes may have implications to the adjacent sacroiliac joint (SIJ). The SIJ has a minimal range of motion and is closely coupled to the adjacent spine and pelvis. In consequence, the SIJ may react sensitively to changes in stress distribution at the acetabulum, with hypermobility-induced pain. The purpose of this study was to investigate the stress distribution of the SIJ in acetabular dysplasia, and to gain insight into the cause and mechanisms of hypermobility-induced pain at the SIJ. Finite element models of pre- and postoperative pelves of four patients with acetabular dysplasia were created and analyzed in double leg standing positions. The preoperative models were relatively inflare, the sacral nutation movement, SIJ cartilage equivalent stress, and the load on the surrounding ligaments decreased with increased posterior acetabular coverage. Acetabular morphology was shown to affect the SIJ, and improvement of the posterior acetabular coverage may help normalize load transmission of the pelvis and thus improve the stress environment of the SIJ in acetabular dysplasia.
  • Gloria Garoffolo, Manuel Casaburo, Francesco Amadeo, Massimo Salvi, Giacomo Bernava, Luca Piacentini, Isotta Chimenti, Germana Zaccagnini, Gesmi Milcovich, Estella Zuccolo, Marco Agrifoglio, Sara Ragazzini, Otgon Baasansuren, Claudia Cozzolino, Mattia Chiesa, Silvia Ferrari, Dario Carbonaro, Rosaria Santoro, Martina Manzoni, Loredana Casalis, Angela Raucci, Filippo Molinari, Lorenzo Menicanti, Francesca Pagano, Toshiro Ohashi, Fabio Martelli, Diana Massai, Gualtiero I. Colombo, Elisa Messina, Umberto Morbiducci, Maurizio Pesce
    Circulation Research 131 (3) 239 - 257 0009-7330 2022/07/22 [Refereed]
     
    Background: Conversion of cardiac stromal cells into myofibroblasts is typically associated with hypoxia conditions, metabolic insults, and/or inflammation, all of which are predisposing factors to cardiac fibrosis and heart failure. We hypothesized that this conversion could be also mediated by response of these cells to mechanical cues through activation of the Hippo transcriptional pathway. The objective of the present study was to assess the role of cellular/nuclear straining forces acting in myofibroblast differentiation of cardiac stromal cells under the control of YAP (yes-associated protein) transcription factor and to validate this finding using a pharmacological agent that interferes with the interactions of the YAP/TAZ (transcriptional coactivator with PDZ-binding motif) complex with their cognate transcription factors TEADs (TEA domain transcription factors), under high-strain and profibrotic stimulation. Methods: We employed high content imaging, 2-dimensional/3-dimensional culture, atomic force microscopy mapping, and molecular methods to prove the role of cell/nuclear straining in YAP-dependent fibrotic programming in a mouse model of ischemia-dependent cardiac fibrosis and in human-derived primitive cardiac stromal cells. We also tested treatment of cells with Verteporfin, a drug known to prevent the association of the YAP/TAZ complex with their cognate transcription factors TEADs. Results: Our experiments suggested that pharmacologically targeting the YAP-dependent pathway overrides the profibrotic activation of cardiac stromal cells by mechanical cues in vitro, and that this occurs even in the presence of profibrotic signaling mediated by TGF-β1 (transforming growth factor beta-1). In vivo administration of Verteporfin in mice with permanent cardiac ischemia reduced significantly fibrosis and morphometric remodeling but did not improve cardiac performance. Conclusions: Our study indicates that preventing molecular translation of mechanical cues in cardiac stromal cells reduces the impact of cardiac maladaptive remodeling with a positive effect on fibrosis.
  • Mun Kit Lai, Baasansuren Otgon, Toshiro Ohashi
    Bio-Medical Materials and Engineering 1 - 9 0959-2989 2022/04/19 [Refereed]
     
    BACKGROUND: Imaging of cells and cellular organelles has been of great interest among researchers and medical staff because it can provide useful information on cell physiology and pathology. Many researches related to collective cell migration have been established and leader cells seem to be the ones that regulate the migration, however, the identification of leader cells is very time-consuming. OBJECTIVE: This study utilized computer vision with deep learning to segment cell shape and to identify leader cells through filopodia. METHODS: Healthy Madin–Darby Canine Kidney (MDCK) cells cultured in a Polydimethylsiloxane (PDMS) microchannel device allowed collective cell migration as well as the formation of leader cells. The cells were stained, and cell images were captured to train the computer using UNet++ together with their corresponding masks created using Photoshop for automated cell segmentation. Lastly, cell shape and filopodia were filtered out using Filopodyan and FiloQuant were detected. RESULTS: The segmentation of cell shape and the identification of filopodia were successful and produced accurate results in less than one second per image. CONCLUSIONS: The proposed approach of image analysis would be a great help in the field of cell science, engineering, and diagnosis.
  • Mechanical Properties of Isolated Primary Cilia Measured by Micro-Tensile Test and Atomic Force Microscopy
    Tien-Dung Do, Jimuro Katsuyoshi, Haonai Cai, Toshiro Ohashi
    Frontiers in Bioengineeing and Biotechnology 2021 [Refereed]
  • Quantitative evaluation of the sacroiliac joint fixation in stress reduction on both sacroiliac joint cartilage and ligaments: a finite element analysis
    B. Venayre, Y. Koyama, D. Kurosawa, N. Hammer, U. Lingslebe, E. Murakami, H. Ozawa, T. Ohashi
    Clinical Biomechanics 85 105350  2021 [Refereed]
  • R. Toyohara, D. Kurosawa, N. Hammer, M. Werner, K. Honda, Y. Sekiguchi, S. Izumi, E. Murakami, H. Ozawa, T. Ohashi
    Scientific Reports 10 (1) 13683 - 13683 2020 [Refereed]
     
    The sacroiliac joint (SIJ) is burdened with variant loads. However, no methods have allowed to measure objectively how the SIJ deforms during bipedal walking. In this study, in-vivo walking conditions were replicated in a kinematic model combining the finite element method with 3D walking analysis data divided into five phases in order to visualize the load transition on the SIJ and clarify the role of the SIJ. Both models with and without inclusion of the SIJ were investigated. In models with bilateral SIJs, the displacement differed greatly between the sacrum and both hip bones on the SIJ as the boundary. The movements of the sacrum involved a nutation movement in the stance phase and a counter-nutation in the swing phase relative to the ilium. In models without SIJs, the displacement of the pelvis and loads of pelvic ligaments decreased, and the equivalent stress of the SIJs increased compared to the model with SIJs. The walking loads cause distortion of the entire pelvis, and stress concentration at the SIJ are seen due to the morphology of the pelvic ring. However, the SIJs help dissipate the resulting stresses, and the surrounding ligaments are likewise involved in load transmission.
  • Effect of Geometric Curvature on Collective Cell Migration in Tortuous Microchannel Devices
    M.B. Mazalan, M.A.B. Ramlan, J.H. Shin, T. Ohashi
    Micromachines 11 (7) 659  2020 [Refereed]
  • コラーゲン・トリペプチドの軟骨細胞に対する賦活作用
    大橋 俊朗, 山本 祥子, 松本 陽, 沼田 徳暁, 酒井 康夫, 佐藤 正明
    応用薬理 98 1 - 5 2020 [Refereed]
  • Tatsuya Seki, Katsuyoshi Jimuro, Yasushige Shingu, Satoru Wakasa, Hiroki Katoh, Tomonori Ooka, Tsuyoshi Tachibana, Suguru Kubota, Toshiro Ohashi, Yoshiro Matsui
    Journal of artificial organs : the official journal of the Japanese Society for Artificial Organs 22 (2) 126 - 133 1434-7229 2019/06 [Refereed][Not invited]
     
    Ideally, an annuloplasty ring's shape should be changed intraoperatively if mitral valve repair is unsuccessful because of a short coaptation length or systolic anterior motion. Several post-implantation adjustable rings have been developed, but they are not freely deformable and are unsuitable for asymmetric repair of the valvular annulus. We developed a novel thermally deformable mitral annuloplasty ring to address these problems and assessed the ring's mechanical properties and its effect on the mitral valve anatomy. This ring was made of polycaprolactone. Tensile and bending tests were performed to evaluate the ring's mechanical properties. The ratio of the transverse and septal-lateral length was determined as 4:3. Using 10 pig hearts, we measured the post-deformation coaptation length and minimum distance from the coaptation to the ventricular septum, which is a factor of abnormal systolic anterior motion of the mitral valve. In the mechanical tests, the ring's yield point was greater than the deformation force of the annulus in humans. In pigs with deformation from "4:3" to "4:2", the coaptation length was significantly increased in each mitral valve part. In pigs with deformation from "4:3" to "4:4", the minimum distance from the coaptation to the ventricular septum was significantly increased. Asymmetrical ring deformation increased the coaptation length only at the deformed area. In conclusion, this new thermally deformable mitral annuloplasty ring could be "order-made" to effectively change the coaptation length in all parts of the mitral valve and the distance from the coaptation to septum post-deformation via intraoperative heating.
  • S. Matsubara, T. Onodera, E. Maeda, D. Momma, M. Matsuoka, K. Homan, T. Ohashi, N. Iwasaki
    Journal of Biomechanics 94 22 - 30 2019 [Refereed]
     
    Glycosphingolipids (GSLs) are ubiquitous membrane components that play an indispensable role in maintaining chondrocyte homeostasis. To gain better insight into roles of GSLs, we studied the effects of GSL-deletion on the physiological responses of chondrocytes to mechanical stress. Mice lacking Ugcg gene (Ugcg-/-) were genetically generated to obtain GSL-deficient mice, and their chondrocytes from the joints were used for functional analyses in vitro culture experiments. The cells were seeded in a three-dimensional collagen gel and subjected to 5%, 10% or 16% cyclic tensile strain for either 3 or 24 h. The gene expressions of chondrocyte anabolic and catabolic factors, and the induction of Ca2+ signaling were analyzed. Our results revealed that chondrocytes derived from GSL-deficient mice exhibited an elevation in the expression of catabolic factors (ADAMTS-5, MMP-13) following the exposure to strain with amplitudes of 10%. Likewise, applying cyclic tensile strain with these amplitudes resulted in an increased Ca2+ oscillation ratio in chondrocytes from GSL-deficient as compared to the ratio from control mice. These results demonstrated that deletion of GSL stimulated the catabolic responses of chondrocytes to mechanical stress via the augmentation of the sensitivity to mechanical stress that may lead to the cartilage deterioration. These findings suggest that the regulation of the physiological responses of chondrocytes by GSLs could be a potential target in a therapeutic intervention in osteoarthritis.
  • 松原 新史, 小野寺 智洋, 門間 太輔, 馬場 力哉, 本谷 和俊, 上徳 善太, 宝満 健太郎, 岩崎 倫政, 大橋 俊朗, 前田 英次郎
    北海道整形災害外科学会雑誌 北海道整形災害外科学会 60 (1) 145 - 146 1343-3873 2018/08
  • Enhanced gap junction intercellular communication inhibits catabolic and pro-inflammatory responses in tenocytes against heat stress
    E. Maeda, S. Kimura, M. Yamada, M. Tashiro, T. Ohashi
    Journal of Cell Communication and Signaling 11 (4) 369 - 380 2017 [Refereed]
  • Eijiro Maeda, Haicheng Pian, Toshiro Ohashi
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 482 (4) 1170 - 1175 0006-291X 2017/01 [Refereed][Not invited]
     
    The present study has been performed on temporal changes in gap junctional intercellular communication (GJIC) between tenocytes under static tensile strain with the magnitude of 0% (no strain), 4% (physiological magnitude) or 8% (overloading magnitude) during a 24-h culture period. Tenocytes were isolated from rabbit Achilles tendon and seeded on a stretchable microgroove substrate. GJIC was evaluated as intercellular diffusion coefficient of calcein (D-GJ) using fluorescence loss in photobleaching (FLIP) protocol accompanied with a mathematical model of molecular diffusion both within the cell and between the cells. It was exhibited that the application of 4% strain for 1 h increased D-GJ significantly. The increased level was maintained for 6 h, followed by returning to the pre-strain level at 24 h. This was associated with a transient increase in connexin 43 (Cx43) gene expression and protein localisation at 1 h, suggesting the increased GJIC may have involved new synthesis of gap junctions. By contrast, the application of 8% static strain reduced DGJ to the similar or lower level from 0% strain group for 6 h, associated with inhibited Cx43 gene expression. However, Cx43 protein localisation was not changed much, and thus, there seem no direct interactions among changes in GJIC, Cx43 gene expression and Cx43 localisation. The present findings highlight the differences in mechanical regulation of GJIC between physiological and non-physiological loadings, and thus the increase or the decrease in GJIC may affect tenocyte functions in different ways. (C) 2016 Elsevier Inc. All rights reserved.
  • Toshiro Ohashi, Yoshiaki Sugaya, Naoya Sakamoto, Masaaki Sato
    Biomedical Engineering Letters 6 (1) 31 - 38 2093-985X 2016/02/01 [Refereed][Not invited]
     
    Purpose: Vascular endothelial cells (ECs) are continuously subjected to mechanical forces such as fluid shear stress, stretching and hydrostatic pressure. The effect of hydrostatic pressure on EC responses has not been fully understood compared to that of the other two stimuli. The purpose of this study is to assess mechanical responses of ECs to these mechanical stimuli. Methods: Bovine aortic ECs were exposed to hydrostatic pressure of 50, 100, and 150 mmHg and fluid shear stress of 3 Pa in simultaneous or successive fashion. Immunofluorescence staining of actin filaments and VEcadherin was then performed to observe cell morphology and cell-cell junctions, respectively Results: The results showed that ECs subjected to 50, 100, and 150 mmHg for 24 h elongated without predominant orientation and exhibited multilayered structure, whereas simultaneous application of 50 and 100 mmHg and 3 Pa for 24 h induced marked elongation and orientation of ECs parallel to the direction of flow maintaining monolayer integrity. This monolayer integrity was lost in ECs subjected to 150 mmHg together with 3 Pa. A successive application of 100 mmHg for 24 h followed by 100 mmHg and 3 Pa for 24 h, indicated that the loss of monolayer integrity due to hydrostatic pressure could not be retrieved by the following simultaneous application. Conclusions: It can be concluded that physiological shear stress of 3 Pa is dominant to physiological hydrostatic pressure up to 100 mmHg, importantly suggesting the relative contribution of physiological hydrostatic pressure and fluid shear stress to endothelial monolayer integrity.
  • Eijiro Maeda, Toshiro Ohashi
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 465 (2) 281 - 286 0006-291X 2015/09 [Refereed][Not invited]
     
    Large magnitudes of mechanical strain applied to tendon cells induce catabolic and inflammatory responses, whereas a moderate level of strain promotes anabolism. Gap junction intercellular communication (GJIC) plays an essential role in these responses, however direct regulation of GJIC by mechanical loading has not been characterised in detail. Here, we show that the GJIC between tenocytes are enhanced or inhibited depending on the magnitude of the tensile strain. The GJIC was analysed using fluorescence loss in photobleaching (FLIP), combined with a molecular diffusion model. Intercellular and intracellular transport of fluorescence tracer molecules, calcein, across multiple cells through the gap junctions was evaluated by determining the intercellular and intracellular diffusion coefficients of calcein. It was demonstrated that the intercellular diffusion coefficient was significantly higher when the cells were subjected to a physiological static tensile strain (4%) for 1 h, but significantly lower when subjected to a strain with non-physiological amplitude (8%). The intracellular diffusion coefficient was not altered by the application of static strain at any level. Connexin 43 proteins were localised within cytoplasm and at cell cell boundaries in no strained state and were also localised near cell nuclei by the 4% strain, but the localisation was reduced by the 8% strain. The findings suggest that the increase in GJIC in response to 4% strain involves opening of gap junction pores via mechanotransduction events of tenocytes, whereas the inhibition in response to 8% strain involves mechanical disruption of the junctions. (C) 2015 Elsevier Inc. All rights reserved.
  • Sorasak Danworaphong, Motonobu Tomoda, Yuki Matsumoto, Osamu Matsuda, Toshiro Ohashi, Hiromu Watanabe, Masafumi Nagayama, Kazutoshi Gohara, Paul H. Otsuka, Oliver B. Wright
    APPLIED PHYSICS LETTERS 106 (16) 0003-6951 2015/04 [Refereed][Not invited]
     
    We use picosecond ultrasonics to image animal cells in vitro-a bovine aortic endothelial cell and a mouse adipose cell-fixed to Ti-coated sapphire. Tightly focused ultrashort laser pulses generate and detect GHz acoustic pulses, allowing three-dimensional imaging (x, y, and t) of the ultrasonic propagation in the cells with similar to 1 mu m lateral and similar to 150 nm depth resolutions. Time-frequency representations of the continuous-wavelet-transform amplitude of the optical reflectivity variations inside and outside the cells show GHz Brillouin oscillations, allowing the average sound velocities of the cells and their ultrasonic attenuation to be obtained as well as the average bulk moduli. (C) 2015 AIP Publishing LLC.
  • OHASHI TOSHIRO, TASHIRO MASATAKA, MAEDA EIJIRO
    Transactions of Japanese Society for Medical and Biological Engineering 一般社団法人 日本生体医工学会 53 S234_03 - S234_03 2015 
    Core temperature of tendon is known to elevate to approximately 43°C under severe mechanical loading in horse as well as in human. This elevated temperature could be a potential etiology of tendinopathy; however, little is known about how hyperthermia affects tenocyte functions. Therefore, the present study hypothesized that such high temperature affects gap junction communication between tenocytes and upregulated tenocyte catabolism. Gap junction communication was evaluated using fluorescence loss in photobleaching (FLIP) protocol, with a mathematical model to estimate both intracellular and intercellular diffusion coefficients from FLIP result. It was exhibited that intercellular diffusion coefficient in tenocytes exposed to 43°C for 30 minutes was significantly larger than that in tenocytes exposed to 37°C. In addition, mRNA expression for catabolic markers, MMP-1 and IL-1β, was markedly upregulated but that for type I collagen was downregulated in 43°C treated group compared to 37°C treated group. Therefore, hyperthermia enhances gap junction communications and catabolic activities in tenocytes, suggesting that hyperthermia could be a trigger of inflammation response.
  • Shaoyi Chen, Akihiro Morita, Irza Sukmana, Eijiro Maeda, Toshiro Ohashi
    Journal of Biorheology 29 (1) 19 - 23 1867-0474 2015 [Refereed][Not invited]
     
    This study was performed to develop a new experimental device with a fibrin gel-microgroove structure for study of microvascularization by endothelial cells (ECs). The effects of the width of microgrooves, initial cell seeding density and a supplementation of vascular endothelial growth factor (VEGF) on in vitro microvasculaization of ECs were examined. ECs were cultured in a fibrin gel formed on a polydimethylsiloxane microgroove substrate, with the microgroove width of 50, 100, 150 and 200 μm. ECs were elongated and sprouted within the gel in all the four types of microgrooves. In addition, multicellular network by connected cell branches were frequently observed in 100- μm microgrooves. Both high initial cell density and VEGF demonstrated significant promotional effects on morphology changes. The findings indicate that microgroove structure serves as a geometrical constraint for ECs, with a promotional effect on angiogenic responses of ECs, and thus, it can be used as an experimental model in the study of in vitro vascularization.
  • Eijiro Maeda, Takehiro Tsutsumi, Nobuto Kitamura, Takayuki Kurokawa, Jian Ping Gong, Kazunori Yasuda, Toshiro Ohashi
    JOURNAL OF BIOMECHANICS 47 (13) 3408 - 3414 0021-9290 2014/10 [Refereed][Not invited]
     
    A double-network (DN) gel, which was composed of poly(2-acrylamido-2-methylpropanesulfonic acid) and poly(N,N'-dimethyl acrylamide) (PAMPS/PDMAAm), has the potential to induce chondrogenesis both in vitro and in vivo. The present study investigated the biomechanical and biological responses of chondrogenic progenitor ATDC5 cells cultured on the DN gel. ATDC5 cells were cultured on a polystyrene surface without insulin (Culture 1) and with insulin (Culture 2), and on the DN gel without insulin (Culture 3). The cultured cells were evaluated using micropipette aspiration for cell Young's modulus and qPCR for gene expression of chondrogenic and actin organization markers on days 3, 7 and 14. On day 3, the cells in Culture 3 formed nodules, in which the cells exhibited an actin cortical layer inside them, and gene expression of type-II collagen, aggrecan, and SOX9 was significantly higher in Culture 3 than Cultures 1 and 2 (p < 0.05). Young's modulus in Culture 3 was significantly higher than that in Culture 1 throughout the testing period (p < 0.05) and that in Culture 2 on day 14 (p < 0.01). There was continuous expression of actin organization markers in Culture 3. This study highlights that the cells on the DN gel increased the modulus and mRNA expression of chondrogenic markers at an earlier time point with a greater magnitude compared to those on the polystyrene surface with insulin. This study also demonstrates a possible strong interrelation among alteration of cell mechanical properties, changes in actin organization and the induction of chondrogenic differentiation. (C) 2014 Elsevier Ltd. All rights reserved.
  • E. Weibull, S. Matsui, H. Andersson-Svahn, T. Ohashi
    Journal of Biomechanical Science and Engineering The Japan Society of Mechanical Engineers 9 (1) JBSE0006 - JBSE0006 1880-9863 2014 [Refereed][Not invited]
     
    The endothelial cells lining our cardiovascular system are constantly affected by shear stress, which can alter both the morphology and biological activity of the cells. Methods to study the basic shear stress response by creating stable flow profiles on the macro scale are well established, but they do not allow the generation of controlled high precision flow profiles. The emergence of microfluidic devices has enabled well-defined individual cellular response studies on endothelial cells in scale-relevant tools. However, so far, no shear stress studies on clonal heterogeneity have been published. We have developed a novel bioassay system to study several shear stress conditions in parallel on clonal expanded single cells. The device consists of a silicon/glass microwell slide with integrated polydimethylsiloxane microchannels, which delivers shear stress to cells in a well-controlled manner using micropumps. The flow behavior of the device was numerically characterized by computational fluid dynamics analysis, which confirmed that the desired fluid-imposed shear stress was obtained. Bovine aortic endothelial cells were cultured in the microwells for 24 hours and then subjected to a fluid shear stress of up to 2.0 Pa for 6 hours. The results showed that alignment and elongation of the endothelial cells along the flow direction were dependent on the level of shear stress applied. It was demonstrated that multiple experimental conditions can be examined simultaneously within a single device and the compartmentalized structure of the microwell slide can be used to ensure physical separation of cells in individual wells. Moreover, it was shown that the device could reduce consumption of expensive reagents and enable screening of rare samples.
  • Eijiro Maeda, Megumi Sugimoto, Youtaro Kosato, Chi Tat Poon, Belinda Pingguan-Murphy, Toshiro Ohashi
    Journal of Biorheology 28 (1) 16 - 20 1867-0474 2014 [Refereed][Not invited]
     
    The present study examined expressions of type I collagen and MMP-1 mRNA of bovine tenocytes in response to elevation in intracellular tension/traction forces induced by either mechanical or chemical stimulation. Tenocytes were cultured in the following conditions: micropillars with the Young’s modulus of 75 kPa in a normal culture medium, in the presence of 1 nM calyculin A in the medium or under 4 or 8% static tensile strain in the normal medium. In all the treatments, cell traction forces were increased significantly from the levels of corresponding control tenocytes. However, these increases in traction forces were not associated with statistically significant increase in type I collagen gene expression. Because our treatments induced the increase in traction forces equidirectionally, it is speculated that highly directional increase in traction forces, associated with an elongated cell shape, is required to induce marked upregulation of type I collagen mRNA expression in tenocytes.
  • Eijiro Maeda, Yasufumi Hagiwara, James H-C. Wang, Toshiro Ohashi
    BIOMEDICAL MICRODEVICES 15 (6) 1067 - 1075 1387-2176 2013/12 [Refereed][Not invited]
     
    Tenocyte mechanotransduction has been of great interest to researchers in tendon mechanobiology and biomechanics. In vivo, tenocytes are subjected to tensile strain and fluid shear stress, but most studies of tenocyte mechanobiology have been to understand how tenocytes regulate their functions in response to tensile strain. Thus, there is still much to know about tenocyte responses to fluid shear stress, partly due to the difficulty of devising a suitable experimental set-up and understanding the exact magnitude of imposed fluid shear stress. Therefore, this study was performed to test a new experimental system, which is suitable for the application of tensile strain and fluid shear stress to tenocytes in vitro. It was experimentally and numerically confirmed that tenocytes could maintain their in situ morphology within microfabricated microgrooves; also, physiological tensile strain and a wide range of fluid shear stress magnitudes can be applied to these cells. Indeed, it was demonstrated that the combined stimulation of cyclic tensile strain and oscillatory fluid shear stress induced a greater synergetic effect on tenocyte calcium response and significantly increased the percentage of tenocyte exhibiting increases in intracellular Ca2+ concentration compared to the solo applications of these two modes of mechanical stimulation. The experimental system presented here is suitable for research of tenocyte mechanobiology, particularly mechanotransduction events, which were difficult to study using previous experimental models like explants and cell monolayers.
  • Hiroshi Ozawa, Takeo Matsumoto, Toshiro Ohashi, Masaaki Sato
    Neuroprotection and Regeneration of the Spinal Cord 61 - 74 2013/11/01 [Refereed][Not invited]
     
    Biomechanical analysis of the spinal cord is important to clarify the morphological plasticity of the spinal cord. In the present study, the elastic moduli of the gray and white matter were measured in situ by using a pipette aspiration method. The mechanical properties and function of the pia mater were investigated. Furthermore, the effect of the tensile stress on the morphology of the spinal cord was investigated. No significant difference in elastic moduli was observed between the gray and white matter of spinal cord. The elastic modulus of the pia mater was 2,300 kPa, which was 460 times higher than that of the spinal cord parenchyma. The pia mater triples the elastic modulus of the spinal cord covered by it compared with the parenchyma and increases the overall stiffness of the spinal cord. As a result of being tightly covered by the pia mater, the compressed spinal cord contains a large amount of strain energy, which enables shape restoration after decompression. The length of the spinal cord was decreased by 9.7 % after the separation to remove longitudinal tensile stress. The tensile stress along the craniocaudal axis in the spinal cord did not affect the spinal cord deformation in response to the compression, but it did affect the shape recoverability after the decompression.
  • Emilie Weibull, Shunsuke Matsui, Manabu Sakai, Helene Andersson Svahn, Toshiro Ohashi
    Biomicrofluidics 7 (6) 064115-1 - 064115-12 1932-1058 2013/11/01 [Refereed][Not invited]
     
    Understanding biomolecular gradients and their role in biological processes is essential for fully comprehending the underlying mechanisms of cells in living tissue. Conventional in vitro gradient-generating methods are unpredictable and difficult to characterize, owing to temporal and spatial fluctuations. The field of microfluidics enables complex user-defined gradients to be generated based on a detailed understanding of fluidic behavior at the μm-scale. By using microfluidic gradients created by flow, it is possible to develop rapid and dynamic stepwise concentration gradients. However, cells exposed to stepwise gradients can be perturbed by signals from neighboring cells exposed to another concentration. Hence, there is a need for a device that generates a stepwise gradient at discrete and isolated locations. Here, we present a microfluidic device for generating a stepwise concentration gradient, which utilizes a microwell slide's pre-defined compartmentalized structure to physically separate different reagent concentrations. The gradient was generated due to flow resistance in the microchannel configuration of the device, which was designed using hydraulic analogy and theoretically verified by computational fluidic dynamics simulations. The device had two reagent channels and two dilutant channels, leading to eight chambers, each containing 4 microwells. A dose-dependency assay was performed using bovine aortic endothelial cells treated with saponin. High reproducibility between experiments was confirmed by evaluating the number of living cells in a live-dead assay. Our device generates a fully mixed fluid profile using a simple microchannel configuration and could be used in various gradient studies, e.g., screening for cytostatics or antibiotics. © 2013 AIP Publishing LLC.
  • 前田 英次郎, 堤 健博, 黒川 孝幸, 北村 信人, Gong Jian Ping, 安田 和則, 大橋 俊朗
    日本整形外科学会雑誌 (公社)日本整形外科学会 87 (8) S1408 - S1408 0021-5325 2013/08
  • Eijiro Maeda, Megumi Sugimoto, Toshiro Ohashi
    JOURNAL OF BIOMECHANICS 46 (5) 991 - 997 0021-9290 2013/03 [Refereed][Not invited]
     
    Actin cytoskeletons, aggregated with myosin II generate intracellular cytoskeletal tension, which is induced to cell attaching substrate as cell traction forces. It is thought that cytoskeletal tension links closely to cell functions. The present study examined quantitative relationships between cytoskeleton tension and the balance of cell metabolism of tenocytes. Using micromachining techniques, micropillar substrates were prepared with polydimethylsiloxane, having three different values of substrate elasticity (6, 18 and 33 kPa) by changing the micropillar height. After 24 h incubation of bovine tenocytes on these micropillar substrates, cell traction forces were determined. Gene expressions for type I collagen (anabolic marker) and MMP-1 (catabolic marker) from tenocytes on micropillars were also analyzed with qPCR. In addition, effects of an inhibition of myosin II activity on tenocyte cytoskeletal tension and metabolism were examined using the inhibitor, blebbistatin. It was exhibited that cell traction forces were significantly larger in tenocytes on 33 kPa substrates compared to those on 6 kPa substrates. This was associated with significant lower expression of MMP-1 mRNA on 33 kPa substrates. Cell traction forces were decreased significantly by the supplementation of blebbistatin in a dose-dependent manner. Indeed, there were significant smaller traction forces and higher expression for MMP-1 mRNA from tenocytes treated with 10 mu M blebbistatin compared to their corresponding controls. Accordingly, tenocyte responses to substrate stiffness are associated with alterations in intracellular tension and catabolic gene expression. On the other hand, tenocyte anabolism, as measured by type I collagen mRNA expression, was not altered with substrate stiffness. (C) 2012 Elsevier Ltd. All rights reserved.
  • Shukei Sugita, Takeo Matsumoto, Toshiro Ohashi, Kiichiro Kumagai, Hiroji Akimoto, Koichi Tabayashi, Masaaki Sato
    Cardiovascular Engineering and Technology 3 (1) 41 - 51 1869-408X 2012/03 [Refereed][Not invited]
     
    Rupture properties of thoracic aortic aneurysms (TAAs) were measured in vitro in a pressure-imposed test to predict the ultimate stress of TAAs from their mechanical behavior in a physiological pressure range. Each quadrilateral (ca. 20 × 20 mm2) specimen of TAAs or porcine thoracic aortas (PTAs) was pressurized from the inner wall until rupture or up to 4500 mmHg, while its deformation was being monitored. In-plane stress σ and strain ε of the specimen were calculated using Laplace's law and deformations of the markers drawn on the specimen surface, respectively. Ultimate stress σmax and tangent elastic modulus H were determined from the σ-ε curve as its maximum stress and slope, respectively. The tangent elastic modulus H of PTA specimens tended to increase with the increase in σ, while that of TAA specimens tended to reach a plateau in a low-σ region. This tendency was confirmed by fitting a function H = Cσ(1 - exp(-σ/τσ)) to the H-σ relation of specimens: The yielding parameter τσ was significantly lower in TAAs than PTAs. Furthermore, the logarithm of the parameter τσ correlated significantly with σmax, for all specimens. These results may indicate that τσ is one of the candidate indices for rupture risk estimation. © 2011 Biomedical Engineering Society.
  • Development of a microfluidic concentration gradient generator on a microwell slide for high-throughput cell analysis
    Emilie Weibull, Shunsuke Matsui, Helene Andersson-Svahn, Toshiro Ohashi
    Proceedings of the 16th International Conference on Miniaturized Systems for Chemistry and Life Sciences, MicroTAS 2012 1573 - 1575 2012 
    The present study reports a newly developed, microfluidic concentration gradient generator that adds substantial utility to the high-throughput nature of the microwell slide that we have previously established [1, 2]. Using microfluidic dynamics, we have designed microchannels to deliver a reagent and its dilutant with precisely controlled flow volumes, generating 8 discrete steps of reagent concentration in designated microwells. As proof of concept FITC-labeled dextran and dextran-free water was used to visualize the concentration gradient. The next step will be to perform cancer drug concentration gradient experiments on an animal cell line.
  • Toshiro Ohashi, Masaaki Sato
    Single and Two-Phase Flows on Chemical and Biomedical Engineering 579 - 599 2012 [Refereed][Not invited]
     
    Vascular endothelial cells are constantly stimulated by blood flow-induced shear stress throughout the vasculature and respond by changing morphology and cytoskeletal structures as well as by modulating cell physiological functions. In particular, since endothelial cell responses to fluid shear stress have been implicated in the localization of atherosclerosis, the effects of fluid shear stress on endothelial cell morphology and functions have been exclusively studied. In fact, previous observations have given preferential localization of lipid accumulation in atherosclerosis in the arterial tree, such as branching and curved regions where blood flow is unsteady and spatially and temporally altered. So far, a lot of efforts have been made to study endothelial mechanotransduction to flow, indicating the fact that after applying fluid shear stress to endothelial cell monolayer, cells exhibit marked elongation and orientation in the direction of flow. It is now accepted that morphological changes of endothelial cells are closely associated with modulation of cell physiology and pathology. The need for experimental techniques for studying endothelial responses to flow has lead to development of different types of flow chambers. Conventional flow chambers include a cone-and-plate flow chamber and a parallel-plate flow chamber, both of which provide steady, fully-developed laminar flow. Furthermore, in order to provide non-steady flow and disturbed flow, novel flow chambers have been developed, such as a tapered channel and an obstacle-included channel. More recently, microfluidic flow chambers have emerged with a great potential for a high throughput analysis. The purpose of this review is to first summarize many types of flow chambers that apply fluid shear stress onto a monolayer of endothelial cells. Next, experimental studies on endothelial cell responses to fluid shear stress are highlighted, focusing on changes in cell morphology associated with cytoskeletal remodeling. © 2012 Bentham Science Publishers. All rights reserved.
  • Shinsuk Park, Kyehan Rhee, Toshiro Ohashi
    Journal of Biomechanical Science and Engineering 7 (4) 335  1880-9863 2012 [Refereed][Not invited]
  • Eijiro Maeda, Akito Sugawara, Justin J. Cooper-White, Toshiro Ohashi
    2011 Int. Symp. on Micro-NanoMechatronics and Human Science, Symp. on "COE for Education and Research of Micro-Nano Mechatronics", Symposium on "Hyper Bio Assembler for 3D Cellular System Innovation" 16 - 19 2012 [Refereed][Not invited]
     
    Cell migration plays an important role in many physiological and pathological processes such as morphogenesis, wound healing, and tumor metastasis. Although the majority of such events occur with cells moving as a group, called collective cell migration, mechanics of collective cell migrations has not been understood well compared to a single cell migration. Mechanical interactions between cells and their surroundings have been demonstrated to regulate cell migration. One of such interactions is the induction of traction forces by acto-myosin dynamics within cells to their local environment, as it has been reported that cells alter the magnitude of traction forces depending on the stiffness of attaching substrates. In connection with cell migration, it has also been demonstrated the importance of substrate stiffness during cell migration using microfabricated substrates consisting of arrays of micropillars. To understand the mechanics of collective cell migration, it is important to know how cells within a moving cell collectivity generate forces to move the collectivity forward at single cell level. Accordingly, the present study was performed to clarify the mechanics of collective cell migration by measuring traction forces exerted by mouse NIH 3T3 fibroblasts using a newly developed migration assay device. © 2011 IEEE.
  • Toshiro Ohashi, Yusaku Niida, Ryoichi Tanaka, Masaaki Sato
    PROCEEDINGS OF THE ASME SUMMER BIOENGINEERING CONFERENCE, PTS A AND B 245 - 246 2012 [Refereed][Not invited]
  • 杉田修啓, 松本健郎, 大橋俊朗, 熊谷紀一郎, 秋元弘治, 田林 晄一, 佐藤 正明
    脈管学 Japanese College of Angiology 52 277 - 283 0387-1126 2012 [Refereed][Not invited]
     
    To predict the rupture of thoracic aortic aneurysms (TAAs), the relationship between stiffness and rupture pressure was investigated in a pressure-imposed test. Each quadrilateral (ca. 20×20 mm2) specimen of human TAAs or porcine thoracic aortas (PTAs) was pressurized until rupture or up to 4500 mmHg. Tension per unit length (T) and strain (ε) of the specimen were calculated from loaded pressure (P) and deformations of the specimen. Stiffness (S) was determined as the slope of the T-ε curve. Blood pressure in vivo (Pvivo) was estimated from P, T, and diameter of the specimen. Stiffness of PTA specimens increased with an increase in Pvivo, while that of TAA specimens reached a plateau in a low-Pvivo region. To evaluate this phenomenon quantitatively, a yielding parameter (τP), i.e., Pvivo at which the stiffness S reached a plateau level, was determined by fitting an equation S=CP{1–exp(–Pvivo/τP)} to the S-Pvivo curve. The parameter τP correlated significantly with the rupture pressure in vivo. This parameter also correlated significantly with the area fraction of collagen, indicating that collagen composition caused changes in the τP in TAA. Since the yielding parameter can be obtained from clinical data, τP can be used for rupture risk estimation.
  • Analysis and techniques for controlling kinesin-driven microtubules in nanoscale transport systems
    S. Sugita, N. Sakamoto, T. Ohashi, M. Sato
    Proceedings of the Tohoku University Global Centre of Excellence Programme 144 - 150 2012 [Not refereed][Not invited]
  • Koki Oya, Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 412 (4) 678 - 682 0006-291X 2011/09 [Refereed][Not invited]
     
    Macrophages in the vessel wall of advanced abdominal aortic aneurysms (AAAs) are subjected to cyclic stretching and hypoxia because of pulsatile blood flow and intraluminal thrombi, respectively. It is possible that these conditions induce abnormal changes in macrophage functions, such as increased production of matrix metalloproteinase-2 (MMP-2), MMP-9, and inflammatory cytokines, leading to weakening of the aortic wall through excessive extracellular matrix disruption. Here we show the effects of cyclic stretching and hypoxia on the production of MMP-9 and inflammatory cytokines by macrophages. Gelatin zymography revealed that MMP-9 production by macrophages was significantly increased by 5% and 10% cyclic stretching under hypoxia (2.2% O-2). Using enzyme-linked immunosorbent assay, we also evaluated the production of 12 different inflammatory cytokines and found that there was a tendency toward higher expressions of interleukin-8 and tumor necrosis factor-alpha by macrophages subjected to 10% cyclic stretching under normoxia and hypoxia. Next, we evaluated apoptosis of smooth muscle cells (SMCs) in medium conditioned by macrophages cultured under the 2 conditions described above. SMC apoptosis increased significantly when exposed to media harvested from macrophages subjected to 10% cyclic stretching under normoxia and hypoxia. On the basis of these results, we believe that macrophages produce cytokines that induce SMC apoptosis. Our results suggest that the combination of cyclic stretching and hypoxia stimulates MMP-9 and cytokine production in macrophages, which may result in weakening of AAA walls. (C) 2011 Elsevier Inc. All rights reserved.
  • Hiroshi Ozawa, Takeo Matsumoto, Toshiro Ohashi, Masaaki Sato, Eiji Itoi
    JOURNAL OF BIOMECHANICS 44 (12) 2313 - 2315 0021-9290 2011/08 [Refereed][Not invited]
     
    The spinal cord is physiologically stretched along the craniocaudal axis, and is subjected to tensile stress. The purpose of this study was to examine the effect of the tensile stress on morphological plasticity of the spinal cord under compression and decompression condition. The C1-T2 spinal column was excised from 4 rabbits. The laminae and lateral masses were removed. After excision of surrounding structures, a small rod was placed on the spinal cord. The rod was connected with a pan of the scale balance. Varying the weight between 0 and 20 g on the other scalepan, the indentation of the rod was measured. Then, the spinal cord was cut transversely to remove longitudinal tensile stress. The samples were measured again with the same protocol at point 10 mm caudal to each pre-measured point on the spinal cord. The shape recovery rate was calculated. The length of the spinal cord decreased by 9.7% after the separation. The maximum indentation was 2.1 mm (mean) at 20 g. and did not differ between the separated and un-separated cords. The recovery rate was not significantly different between the separated and un-separated cords until 3 g. At the load under 3 g, the recovery rate after the separation was significantly lower than that before the separation. The tensile stress along the craniocaudal axis in the spinal cord did not affect the spinal cord deformation in response to the compression, but it did affect the shape recoverability after the decompression. (C) 2011 Elsevier Ltd. All rights reserved.
  • Megumi Sugimoto, Eijiro Maeda, Toshiro Ohashi
    ASME 2011 Summer Bioengineering Conference, SBC 2011 875 - 876 2011 [Refereed][Not invited]
  • Naoya Sakamoto, Kei Segawa, Makoto Kanzaki, Toshiro Ohashi, Masaaki Sato
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 398 (3) 426 - 432 0006-291X 2010/07 [Refereed][Not invited]
     
    p120-Catenin is known to play important roles in cell-cell adhesion stability by binding to cadherin and morphological changes of cells by regulating small RhoGTPase activities. Although the expression and binding states of p120-catenin are thought to dynamically change due to morphological adaptation of endothelial cells (ECs) to fluid shear stress, these dynamics remain to be explored. In the present study, we examined the time course of changes in p120-catenin expression and its binding to vascular endothelial (VE)-cadherin in ECs exposed to shear stress. Human umbilical vein ECs began to change their morphologies at 3-6 h, and became elongated and oriented to the direction of flow at 24 h after exposure to a shear stress of 1.5 Pa. Binding and co-localization of p120-catenin with VE-cadherin at the foci of cell-cell adhesions were retained in ECs during exposure to shear stress, indicating that VE-cadherin was stabilized in the plasma membrane. In contrast, cytoplasmic p120-catenin that was dissociated from VE-cadherin was transiently increased at 3-6 h after the flow onset. These results suggest that the transient increase of cytoplasmic p120-catenin may stimulate RhoGTPase activities and act as a switch for the morphological changes in ECs in response to shear stress. (C) 2010 Elsevier Inc. All rights reserved.
  • Naoya Sakamoto, Naoki Saito, Xiaobo Han, Toshiro Ohashi, Masaaki Sato
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 395 (2) 264 - 269 0006-291X 2010/04 [Refereed][Not invited]
     
    Arterial bifurcations are common sites for development of cerebral aneurysms. Although this localization of aneurysms suggests that high shear stress (SS) and high spatial SS gradient (SSG) occurring at the bifurcations may be crucial factors for endothelial dysfunction involved in aneurysm formation, the details of the relationship between the hemodynamic environment and endothelial cell (EC) responses remain unclear. In the present study, we sought morphological responses of ECs under high-SS and high-SSG conditions using a T-shaped flow chamber. Confluent ECs were exposed to SS of 2-10 Pa with SSG of up to 34 Pa/mm for 24 and 72 h. ECs exposed to SS without spatial gradient elongated and oriented to the direction of flow at 72 h through different processes depending on the magnitude of SS. In contrast, cells did not exhibit preferred orientation and elongation under the combination of SS and SSG. Unlike cells aligned to the flow by exposure to only SS, development of actin stress fibers was not observed in ECs exposed to SS with SSG. These results indicate that SSG suppresses morphological changes of ECs in response to flow. (C) 2010 Elsevier Inc. All rights reserved.
  • Hidenori Fujiwara, Yoshikatsu Saiki, Mitsuru Sato, Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato, Yasuhiko Tabata, Koichi Tabayashi
    JOURNAL OF VASCULAR SURGERY 51 (1) 194 - 202 0741-5214 2010/01 [Refereed][Not invited]
     
    Objectives: We investigated the outcomes of reinforcing anastomotic sites using (1) non biodegradable polytetrafluoroethylene (PTFE) felt, (2) biodegradable polyglycolic acid (PGA) felt, and (3) PGA felt with basic fibroblast growth factor (bFGF) in a canine descending thoracic aortic replacement model. Methods: Thirty-seven beagles underwent descending thoracic aorta replacement using a prosthetic graft with one of the above-mentioned reinforcements or no reinforcement for controls. Histologic evaluations were carried out I month and 3 months after surgery. The biomechanical strength of the anastomosis was assessed along the longitudinal axis of the aortic segments using a tensile tester. Local compliance at the anastomotic site was also evaluated in the circumferential direction. Results. The media was significantly thinner in the PTFE group than in the control group (65.8% +/- 5.1% vs 95.0% +/- 9.3% of normal thickness; P < .05). Relative to the control group, the adventitial layer was significantly thinner in the PTFE group (42.3% +/- 8.2% of control; P < .05) but significantly thicker in the PGA and the PGA + bFGF groups (117.2% +/- 11.3% and 134.1% +/- 14.2% of control, respectively; P < .05). There were more vessels in the adventitial layer in the PGA + bFGF group than in the control, PTFE, and PGA groups (29.2 +/- 2.1/mm(2) vs 13.8 +/- 0.8, 5.4 +/- 0.7, 17.0 +/- 1.3/mm(2), respectively; P < .01). There were no significant differences between the four groups in the failure force at anastomotic sites. Local compliance at the anastomotic site was higher in the PGA group than that in the PTFE group (11.6 +/- 1.6 10(-6) m(2)/N vs 5.6 +/- 1.9 10(-6) m(2)/N; P < .05). Conclusion: Reinforcement of the experimental aortic wall with PTFE felt resulted in thinning of the media and adventitia and fewer vessels at the anastomotic site. These histologic changes were not observed when biodegradable felt was used. The bFGF failed to augment the modification of the aortic wall with the exception of increased adventitial vessel number. Biomechanical strength of the anastomosis along the longitudinal axis was comparable in all four groups; however, local vascular compliance was better in the biodegradable PGA felt group. (J Vase Surg 2010;51:194-202.) Clinical Relevance: This investigation was conducted to extend our previous investigation on a biodegradable felt strip into more practical form before we proceed in a clinical application of the new, material. We hypothesized that sustaining compression of the aorta by the nonbiodegradable felt strip may cause structural derangement and local ischemia on the aortic wall, which may lead to occurrence of late postoperative false aneurysm after aortic surgery. We attempted to find a clue for preventing adverse effects of reinforcement with a conventional felt strip. We have found that biodegradable felt prevented thinning of both the media and adventitia and increased adventitial vessels with increased vascular compliance at the aortic anastomotic sites.
  • Shukei Sugita, Tatsuya Murase, Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato
    LAB ON A CHIP 10 (6) 755 - 761 1473-0197 2010 [Refereed][Not invited]
     
    Gliding microtubules (MTs) on a surface coated with kinesin biomolecular motors have been suggested for the development of nanoscale transport systems. In order to establish a sorting function for gliding MTs, events for MTs approaching micro-scale grooves were investigated. MTs longer than the width of grooves fabricated on a Si substrate bridged the grooves (bridging) and many MTs shorter than the groove width almost began to bridge, but returned to the surface that they approached from (guiding). Occurrence probabilities for the events were analyzed with focus on the geometric conditions, such as length of the MTs, width of the grooves, and the incident angle (alpha) of the MTs approaching the grooves. The occurrence probability for bridging increased with an increase in the incident angle (16%, alpha = 0-30 degrees; 51%, alpha = 30-60 degrees; 75%, alpha = 60-90 degrees), and the probability for guiding decreased with an increase in the incident angle (79%, alpha = 0-30 degrees; 55%, alpha = 30-60 degrees; 5%, alpha = 60-90 degrees). The results indicate that an incident angle of 30-60 degrees is an effective condition for MT sorting, because the bridging and guiding events can sort MTs that are longer and shorter than the groove widths, respectively. Furthermore, the occurrence probabilities of both bridging and guiding in a higher concentration of methylcellulose (0.5%) increased up to approximately 70% at incident angles of 30-60 degrees, indicating good feasibility for the development of devices for the sorting of MTs on surfaces with topographical grooves.
  • KOHATA Kengo, MAEDA Eijiro, OHASHI Toshiro
    The proceedings of the JSME annual meeting 一般社団法人日本機械学会 2010 143 - 144 2010 
    The present study hypothesized that initial geometrical shapes of cutaneous wound and the major axis of the wound to the direction of stretch could influence myofibroblast realignment observed in the maturation phase of healing process. Human dermal fibroblasts were differentiated into myofibroblasts with a treatment of TGF-β1. The cells were seeded onto a PDMS chamber either in a monolayer or in a colony of the following shapes: circle, and track fields both parallel and vertical to the stretch. Cyclic tensile stretch, with an amplitude of 20%, was provided at 1 Hz for 6 hours. Myofibroblasts seeded in the circle and the parallel track exhibited a similar response of realigning ±1:70° to the strain axis with uniformly distributed myofibroblasts in the whole area. By contrast, the cells seeded in the shape of vertical track demonstrated realignment to no specific direction. Findings supports our hypothesis, indicating that the shape and the orientation of wound influences healing processes through myofibroblasts realignment.
  • Toshiro Ohashi, Helene Andersson-Svahn
    Journal of Biomechanical Science and Engineering 5 (3) 185  1880-9863 2010 [Refereed][Not invited]
  • Effect of spatial gradient of shear stress on morphological responses of endothelial cells to flow
    N. Sakamoto, N. Saito, T. Ohashi, M. Sato
    Proceedings of 13th International Symposium of Tohoku University Global COE programme 64 - 67 2010 [Not refereed][Not invited]
  • Toshiro Ohashi, Norifumi Kameda, Shouji Nakamura, Masaaki Sato
    Journal of Biomechanical Science and Engineering 5 (3) 262 - 271 1880-9863 2010 [Refereed][Not invited]
     
    Cellular traction forces were measured by using a microfabricated substrate, particularly exploring how cytoskeletal structures such as actin filaments and microtubules contribute to traction forces. Smooth muscle cells isolated from bovine aortas were cultured and transfected with fluorescence proteins to visualize cell microstructures and then plated on a micropatterned elastomer substrate with arrays of micropillars. Cell spreading on the substrates produced deflection of micropillars which was used for estimation of cellular traction forces, and was closely associated with organization of stress fibers of actin filaments. Traction forces varied considerably among cells, showing the order of several 10 nN. After disruption of microtubules with nocodazole, traction forces significantly increased and there was no detectable change in formation of stress fibers. To inhibit the ROCK pathway, a signaling pathway of myosin light chain phosphorylation, possibly being induced by disruption of microtubules, significantly depressed the increase in traction forces after the disruption of microtubules. This result indicates that microtubules disassembly may regulate the actomyosin-based contractile system mainly through the ROCK pathway. The present study suggests that formation of stress fibers are mainly involved in cellular traction forces and a contribution of microtubules should include not only a force balance but also rather a modulator of the actomyosin contractile system in actin stress fibers. © 2010 by JSME.
  • Sara Lindstrom, Kiichiroh Mori, Toshiro Ohashi, Helene Andersson-Svahn
    ELECTROPHORESIS 30 (24) 4166 - 4171 0173-0835 2009/12 [Refereed][Not invited]
     
    Increasing awareness of the importance of cell heterogeneity in many biological and medical contexts is prompting increasing interest in systems that allow single-cell analysis rather than conventional bulk analysis (which provides average values for variables of interest from large numbers of cells). Recently, we presented a microwell chip for long-term, high-throughput single-cell analysis. The chip has proved to be useful for purposes such as screening individual cancer and stem cells for protein/gene markers. However, liquids in the wells can only be added or changed by manually rinsing the chip, or parts of it. This procedure has several well-known drawbacks - including risks of cross-contamination, large dead volumes and laboriousness - but there have been few previous attempts to integrate liquid rinsing/switching channels in "ready-to-use" systems for single-cell analysis. Here we present a microwell system designed (using flow simulations) for single-cell analysis with integrated microfluidic components (microchannels, magnetically driven micropumps and reservoirs) for supplying the cell culture wells with reagents, or rinsing, thus facilitating controlled, directed liquid handling. It can be used totally independently, since tubing is not essential. The practical utility of the integrated system has been demonstrated by culturing endothelial cells in the microwells, and successfully applying live-cell Calcein AM staining. Systems such as this combining high-density microwell chips with microfluidic components have great potential in numerous screening applications, such as exploring the important, but frequently undetected, heterogeneity in drug responses among individual cells.
  • Ichiro Yoshioka, Yoshikatsu Saiki, Azusa Ichinose, Kei Takase, Shoki Takahashi, Toshiro Ohashi, Masaaki Sato, Koichi Tabayashi
    JOURNAL OF THORACIC AND CARDIOVASCULAR SURGERY 137 (4) 957 - 962 0022-5223 2009/04 [Refereed][Not invited]
     
    Objective: Reoperative median sternotomy entails a risk of damaging the heart or great vessels. If the severity of retrosternal adhesion is accurately assessed before sternal re-entry, resternotomy-related complications can be prevented. The purpose of this investigation was to evaluate whether the severity of retrosternal adhesions can be accurately predicted by tagged cine magnetic resonance imaging. Methods: Thirteen patients who were scheduled to undergo cardiac reoperation were investigated by electrocardiography-gated tagged cine magnetic resonance imaging before sternal re-entry. With the imaging data, the severity of retrosternal adhesion was scored visually on the basis of abnormality in regional myocardial motion and discordance in the tagged signals of the sternum and the myocardium. Also, with the aid of a finite element model, strain at the surface of the right ventricle was calculated on the basis of displacement of the tags on the heart over the cardiac cycle. For comparison, the adhesion severity was scored visually at the time of redo surgery by surgeons who were blinded to the preoperative assessment. Results: The preoperative adhesion severity score, as determined visually by tagged cine magnetic resonance imaging, was correlated with the intraoperative severity score (correlation coefficient: r = 0.76, P < .01). Mean strain at the surface of the heart, as calculated preoperatively by finite element model analysis, was inversely correlated with the intraoperative adhesion severity score (r = -0.78, P < .01). Conclusion: Tagged cine magnetic resonance imaging with a finite element model can provide an accurate quantitative assessment of retrosternal adhesions before redo cardiac surgery.
  • Tsubasa S. Matsui, Shinji Deguchi, Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato
    BIORHEOLOGY 46 (5) 401 - 415 0006-355X 2009 [Refereed][Not invited]
     
    Conventional atomic force microscopy is one of the major techniques to evaluate mechanical properties of cells and subcellular components. The use of a cantilever probe for sample manipulation within the vertical plane often makes absolute positioning of the probe, subject to thermal drift, difficult. In addition, the vertical test is unable to observe changes in the sample structure responsible for mechanical behavior detected by the probe. In the present study, an alternative mechanical tester was developed that incorporated a pair of micro-needles to manipulate a sample in a project plane, allowing acquisition of the accurate probe position and entire sample image. Using a vision-based feedback control, a micro-needle driven by a piezo actuator is moved to give user-defined displacements or forces to sample. To show its usefulness and versatility, three types of viscoelastic measurements on actin stress fibers isolated from smooth muscle cells were demonstrated: strain rate-controlled tensile tests, relaxation tests and creep tests. Fluorescence imaging of the stress fibers using Qdots over the course of the measurements, obtained through multiple image detectors, was also carried out. The technique described here is useful for examining the quantitative relationship between mechanical behavior and related structural changes of biomaterials.
  • T. Ohashi, H. Ichihara, N. Sakamoto, M. Sato
    IFMBE Proceedings 23 2192 - 2193 1680-0737 2009 [Refereed][Not invited]
     
    Adherent cells are known to exert traction forces to adhere to their extracellular matrix at focal adhesions, where integrins interact with the extracellular matrix and mediate various cellular signals. Therefore, it is hypothesized that extracellular matrix-mediated integrin expression may modulate cellular traction forces, resulted in alterations in cell physiology. In this study, to test the hypothesis, traction force measurements are performed on bovine thoracic smooth muscle cells using a polydimethylsiloxane-based micropillar substrate coated with different types of extracellular matrix. The top of the micropillars was selectively coated with fibronectin (FN), vitronectin (VN), and type I collagen (COL I) at 30 and 50 μg/ml and cells were plated on the extracellular matrix-coated micropillars. For FN and VN, the cells have several spikes at the cell periphery, whereas, for COL I, the cell expresses relatively oval shape. At 30 μg/ml, traction forces were 5.5 ± 0.6, 5.8 ± 1.3 and 8.5 ± 2.2 nN for FN, VN, COL I, respectively. At 50 μg/ml, traction forces significantly increased compared to 30 μg/ml and were 12.4 ± 6.8, 13.5 ± 4.0 and 18.1± 4.7 nN for FN, VN, COL I, respectively. These results indicate that extracellular matrix may specifically modulate integrin expression followed by development of actin stress fibers, possibly leading to changes in traction forces. © 2009 International Federation of Medical and Biological Engineering.
  • Takeo Matsumoto, Tomohiro Fukui, Toshihiro Tanaka, Naoko Ikuta, Toshiro Ohashi, Kiichiro Kumagai, Hiroji Akimoto, Koichi Tabayashi, Masaaki Sato
    Journal of Biomechanical Science and Engineering 4 (4) 518 - 529 1880-9863 2009 [Refereed][Not invited]
     
    Mechanical properties of human aortic aneurysm tissues were measured with a biaxial tensile tester. Fifteen-mm-square specimens were obtained from thoracic aortic aneurysms of various origins and from undilated aortas adjacent to the aneurysms during aneurysmectomy, and were stored frozen until the measurement. Each specimen was stretched biaxially in physiological saline at room temperature at the rate of ~0.2 mm/sec. Although the ordered displacement was set equal for both directions, real strain applied to the specimens was not equibiaxial. The stress-strain curves under equibiaxial stretch were obtained by fitting measured curves with a strain energy function considering material anisotropy. Effects of freezing and ambient temperature on the mechanical properties were evaluated with porcine thoracic aortas. The mechanical properties of the frozen-stored specimens at 23oC were almost similar to those of the fresh specimens at 37oC. Elastic modulus at zero load averaged for both directions Hmi = (Hxi+Hyi)/2 was higher (P < 0.01) in the aneurysm tissues (1450 ± 250 kPa, mean ± SEM, n = 26) than in the undilated tissues (650 ± 140 kPa, n = 10). Anisotropy index K = |Hxi-Hyi|/Hmi was not significantly different between the aneurysm (20 ± 3%) and the undilated tissues (12 ± 3%) for all specimens. For the specimens whose elastic modulus Hmi was smaller than 1 MPa, however, the index K was significantly higher (P < 0.05) in the aneurysm specimens (23.1 ± 5.3%, n = 14) than the undilated tissues (9.5 ± 2.5%, n = 8). These results indicate aneurysm tissues are not only stiffer but also more anisotropic than the nonaneurysmal tissues.
  • M. Sato, N. Saito, N. Sakamoto, T. Ohashi
    IFMBE Proceedings 25 (4) 312 - 313 1680-0737 2009 [Refereed][Not invited]
     
    Arterial bifurcations are known as the common sites for development of cerebral aneurysms. Although localization of aneurysms suggests that high wall shear stress (WSS) and high wall shear stress gradient (WSSG) occurring at the bifurcations may be crucial factors for endothelial dysfunction involved in aneurysm formation, the details of the relationship between the hemodynamic environment and endothelial cell (EC) responses still remain unclear. In this study, we observed morphological responses of ECs under high WSS and high WSSG condition using a T-shaped flow chamber and under high WSS without WSSG condition using a parallel plate flow chamber to evaluate the effects of WSSG to ECs, After 24 h exposure to flow, ECs under high WSS (10 Pa) without WSSG condition oriented perpendicular to the flow, whereas ECs at high WSS (10 Pa) with WSSG condition did not cause EC alignment. After 72 h exposure to flow, ECs exposed to WSSG were not polarized whereas ECs at high WSS without WSSG condition orientated and elongated to the direction of flow. These results indicate that a WSSG may suppress orientation of ECs to the flow direction.
  • Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato
    PROCEEDINGS OF THE ASME SUMMER BIOENGINEERING CONFERENCE 2008, PTS A AND B 61 - 62 2009 [Refereed][Not invited]
  • Shulkel Sugita, Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato
    PROCEEDINGS OF THE ASME SUMMER BIOENGINEERING CONFERENCE 2008, PTS A AND B 315 - 316 2009 [Refereed][Not invited]
  • S. Sugita, N. Sakamoto, T. Ohashi, M. Sato
    Journal of Biomechanical Science and Engineering 一般社団法人 日本機械学会 4 (3) 404 - 414 2009 [Refereed][Not invited]
     
    Control of the gliding directions of kinesin-driven microtubules (MTs) in vitro has good feasibility for the development of nano-scale transport systems. A requirement for the control of transporters in these systems includes detecting the positions of gliding MTs; however, no studies have reported on the monitoring of the positions of gliding MTs. Here, we suggest an algorithm to detect tip coordinates of gliding MTs by binarization, skeletonization, and filtration of fluorescent images of MTs. The algorithm was first applied to artificially drawn segments with given lengths (10-80 pixels), widths (1-10 pixels), and curvature radii (20-120 pixels) to verify the effect of the sizes of MTs on accuracy of tip coordinates extracted by the algorithm, and error was estimated by referring to the true coordinates. The estimated errors were as small as 2 pixels in the width and were not affected by the length and the curvature radius, indicating that our algorithm is useful to extract the tips of MTs. The algorithm was subsequently applied to images of gliding MTs. Since distances from the trajectories of the MTs to the centers of gravity of the MTs (3.7 ± 2.1 pixels) were significantly larger than those to the tips (1.9 ± 0.5 pixels), the use of the tips as representative points of gliding MTs was verified. A detection method using tips of MTs, as suggested in this study, may be a useful technique for monitoring each MT in nanoscale transport systems.
  • S. Sugita, N. Sakamoto, T. Ohashi, M. Sato
    Journal of Biomechanical Science and Engineering The Japan Society of Mechanical Engineers 3 (4) 510 - 519 1880-9863 2008/12 [Refereed][Not invited]
     
    Kinesins, biomolecular motors moving along microtubules (MTs) in cells, can potentially be utilized as nano-scale transport systems with an inverted gliding assay, in which the MTs glide on a kinesin-coated surface. Although the key requirements include controls of gliding direction and velocity of MTs, the details of motility properties of MTs have not been well known. In this study, we quantitatively measured angular and gliding velocities, particularly focusing on the effects of MT length and kinesin density. The gliding assay of MTs of up to 20 μm in length was performed on a substrate coated with the kinesin density of 7.5, 38, and 75 μg/ml that resulted in the kinesin spacing of 7.8, 4.2, and 3.1 μm, respectively. The angular velocity for MTs shorter than kinesin spacings significantly decreased with increasing their length, and that for MTs longer than kinesin spacings was not affected by their length. Moreover, the angular velocity was substantially higher at lower kinesin density. These results suggest that both the number of associated kinesins with MTs and the kinesin spacings may contribute to the gliding direction. In contrast, the gliding velocity was independent of the MT length, ranging from 0.3 to 0.5 μm/s with decreasing the kinesin density. This may potentially imply the existence of an underlying mechanism with respect to the number of kinesins per the unit length of MTs. Towards development of high throughput nano-scale transport systems, long MTs and low kinesin densities would be effective for high directionality and high velocity, respectively.
  • Toshiro Ohashi, Kazuhiko Hanamura, Daisaku Azuma, Naoya Sakamoto, Masaaki Sato
    Journal of Biomechanical Science and Engineering 3 (2) 63 - 74 1880-9863 2008 [Refereed][Not invited]
     
    Cell Nuclei play a critical role in controlling gene expression and replicating DNA, and is known to deform in association with cell shape changes in response to external forces. This study dealed with morphological analysis to quantitatively assess the effect of three different mechanical stimuli including fluid shear stress, cyclic stretching, and hydrostatic pressure on nucleus morphology. Fluorescence images showed that fluid shear stress and cyclic stretching induced cell elongation and orientation very specifically to the direction of flow and stretch, respectively. In contrast, hydrostatic pressure induced cell elongation at non-preferred orientation. The nuclei were also found to deform in the same manner as that of the cells, which was, in particular, dependent on the type of mechanical stimuli, possibly suggesting the direct mechanical linkages between cell surface receptors, cytoskeletal meshworks, and nuclei. It was also shown that cytoskeletal meshworks may contribute to pre-existing strain of the nuclei. © 2008, The Japan Society of Mechanical Engineers. All rights reserved.
  • Guanbin Song, Yang Ju, Hitoshi Soyama, Toshiro Ohashi, Masaaki Sato
    MCB Molecular and Cellular Biomechanics 4 (4) 201 - 210 1556-5297 2007/12 [Refereed][Not invited]
     
    Mechanical stimulation is critical to both physiological and pathological states of living cells. Although a great deal of research has been done on biological and biochemical regulation of the behavior of bone marrow mesenchymal stem cells (MSCs), the influence of biomechanical factors on their behavior is still not fully documented. In this study, we investigated the modulation of mechanical stretch magnitude, frequency, and duration on the human marrow mesenchymal stem cells (hMSCs) proliferation by an in vitro model system using a mechanical stretch loading apparatus, and optimized the stretch regime for the proliferation of hMSCs. We applied 3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyl tetrasodium bromide (MTT) assay to estimate the overall proliferative effects of the stretch on hMSCs. We found that fibronectin coating increased adhesion to silicone chamber surface, however, it did not show significant effect on proliferation of hMSCs. A frequency of 1 Hz was more effective in stimulating hMSCs proliferation. At 1 Hz, 5% strain for 15, 30, 60 min, the significant increase of hMSCs proliferation was observed. Proliferation was enhanced at 1 Hz, 10% strain for 15, 30 min, while decreased for 60 min. At 1 Hz, 15% strain, 15 min stretch resulted in the decrease of proliferation, and 30 min and 60 min stretch showed an increased proliferation. Long time (12 and 24 h) strain application blocked the proliferation. These results indicate that mechanical stretch plays an important role in hMSCs growth and proliferation an appropriate mechanical stretch regime could be a novel approach to promoting proliferation of hMSCs in vitro. Copyright © 2007 Tech Science Press.
  • S. Deguchi, T. Ohashi, M. Sato
    Biomechanics at Micro- and Nanoscale Levels 60 - 71 2007/01/01 [Refereed][Not invited]
     
    Actin stress fibers (SFs) in vascular smooth muscle cells and endothelial cells play a critical role in transmitting intracellular forces between separate focal adhesion sites. However, quantitative studies on the berable tension level for single SFs have not yet appeared. Here, we estimated magnitude of preexisting tension in SFs based on measurements of their preexisting stretching strain and tensile properties. Cultured cells expressing fluorescentlylabeled actin were treated with detergents to extract the acin bundles. One end of an individual SF was then dislodged from the substrate by using a microneedle, resulting in a shortening of the SF due to a release of preexisting tension. Tensile tests of the isolated single SFs were conducted with a pair of cantilevers to measure the force required for stretching it up to the original length that corresponds to preexisting tension. The magnitude of the preexisting tension, ~10 nN on average, was comparable in magnitude to previously reported data on the cell traction force generated by living adherent cells at focal adhesion sites to keep cell integrity. The Young’s modulus of the isolated SFs was estimated to be ~300-1500 kPa from the tensile tests. These data will be fundamental in considering the intracellular force transmission mechanism in vascular cells.
  • Toshiro Ohashi, Shouji Nakamura, Naoya Sakamoto, Masaaki Sato
    2007 INTERNATIONAL SYMPOSIUM ON MICRO-NANO MECHATRONICS AND HUMAN SCIENCE, VOLS 1 AND 2 400 - + 2474-378X 2007 [Refereed][Not invited]
     
    This study addresses a technique to explore mechanical role of intracellular structures by using a triton cytoskeleton model (TC model) for traction force measurements. Inhibition of signaling pathways of myosin light chain phosphorylation, possibly being induced by disruption of mictotubules, is also performed. Traction forces for the TC model significantly decreased compared to control. In contrast, traction forces significantly increased from 10.3 +/- 2.5 nN to 13.3 +/- 3.7 nN after treatment of nocodazole, which is well consistent with previous studies. From these results, not only cytoskeletal structures but also other cellular components such as cytoplasm should be involved in cell mechanics. Separate fluorescence studies showed that microtubules disruption induced myosin light chain phosphorylation. Exposure to Y27632 showed that traction forces decreased by 80% compared to control within 15 min and the following treatment with nocodazole showed only 40% recovery from the priori decreased forces. This result indicates that microtubules disassembly may modulate the actomyosin mechanism leading to the increase in traction forces, mainly through the ROCK pathway. It can be concluded that contribution of microtubules should include not only a force balance but also a modulator of the actomyosin-based contractile system.
  • S. Deguchi, K. Maeda, T. Ohashi, M. Sato
    Journal of Biomechanics 39 (7) 1362  0021-9290 2006 [Refereed][Not invited]
  • Shinji Deguchi, Toshiro Ohashi, Masaaki Sato
    FUTURE MEDICAL ENGINEERING BASED ON BIONANOTECHNOLOGY, PROCEEDINGS 71 - + 2006 [Refereed][Not invited]
     
    Intracellular tension transmission through basal actin stress fibers (SFs) running between separate focal adhesion sites has been proposed to play a role in sensing extracellular forces to adapt the adherent cell structure. However, quantitative descriptions of tension level in SFs are still poorly studied. Here, magnitude of preexisting tension in single SFs of endothelial cells was evaluated by in vitro tensile test. Single SFs were isolated from the cells with a combination of low ionic-strength solution and detergent extractions. A tensile test was conducted with a pair of micro-cantilevers driven by a piezo-electric actuator to measure the mechanical properties of the isolated single SF and the force required for stretching it up to its original length, yielding an estimate of the preexisting tension. The Young's modulus was estimated similar to 290 kPa assuming that the specimen was homogeneous. The magnitude of the preexisting tension, 4 nN on average, was comparable to previously reported data of the traction force generated by adherent cells at single adhesion sites to keep cell integrity. These data will contribute to quantitative understanding of intracellular stress transmission mechanism in adherent cells.
  • Kazushi Ito, Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato
    FUTURE MEDICAL ENGINEERING BASED ON BIONANOTECHNOLOGY, PROCEEDINGS 103 - + 2006 [Refereed][Not invited]
     
    In vivo, endothelial cells are exposed to varying frequencies of pulsatile flow. However, little is known of how their frequency affects cell morphology and function. In this study, the effects of frequency of pulsatile flow on morphology and 03 integrin expression of bovine aortic endothelial cells (BAEC) were investigated. BAEC were exposed to pulsatile flow at frequencies of 0.5, 1.0 and 1.5 Hz for 12 h using a newly developed flow-system. BAEC at 0.5 and 1.0 Hz elongated and oriented with the direction of flow and exhibited developed stress fibers parallel to the cell major axis, and these morphological changes were similar to these of BAEC under steady flow conditions. BAEC showed elongation but a random orientation after exposure to 1.5 Hz pulsatile flow. Fluorescent staining revealed that the expression of alpha(v)beta(3) integrin at 1.0 Hz was significantly higher at the central regions of cells than at the upstream regions. In contrast, at 0.5 and 1.5 Hz, the expression of alpha(v)beta(3) integrin was significantly higher at the upstream regions than at the central regions. These findings suggest that frequency of pulsatile flow play important roles in morphological changes and alpha(v)beta(3) integrin expression in EC.
  • Toshiro Ohashi, Naoya Sakamoto, Masaaki Sato
    FUTURE MEDICAL ENGINEERING BASED ON BIONANOTECHNOLOGY, PROCEEDINGS 23 - + 2006 [Refereed][Not invited]
     
    Vascular endothelial cells are an interface between blood vessel walls and blood flow, and are widely known to play important roles in physiological functions of the walls. Since endothelial cell responses to fluid shear stress have been implicated in the localization of atherosclerosis, the effect of shear stress on endothelial cell morphology and functions has been exclusively studied. After applying fluid shear stress, cultured endothelial cells show marked elongation and orientation in the direction of flow. In addition, thick stress fibers of actin filaments appear and align along the cell long axis. In contrast, although endothelial cells are always exposed to hydrostatic pressure in vivo, little is known of how hydrostatic pressure affects endothelial cell morphology and functions. We have recently reported that endothelial cells exposed to hydrostatic pressure exhibited marked elongation and orientation with the random direction, together with development of centrally located, thick stress fibers. Pressured endothelial cells also exhibited multilayered structure unlike under control conditions. These results indicate that endothelial cells may respond very specifically to the type of imposed mechanical stimuli including fluid shear stress and hydrostatic pressure. The purpose of this study is to summarize endothelial cell responses to both fluid flow and hydrostatic pressure, focusing on the changes in cell shape and cytoskeletal structure.
  • Katherine Baria, Yoshiaki Sugaya, Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato
    FUTURE MEDICAL ENGINEERING BASED ON BIONANOTECHNOLOGY, PROCEEDINGS 61 - + 2006 [Refereed][Not invited]
     
    The subcellular location of beta-catenin and N-cadherin in cultured bovine endothelial cells (ECs) exposed to hydrostatic pressure was investigated. ECs were exposed to physiological blood pressure under a hydrostatic head of culture medium for 24 hours. Pressured ECs exhibited changes in morphology and proliferation. Immunofluorescent localization of beta-catenin and N-cadherin indicated changes in their distribution in pressured compared to control cells. beta-catenin was located at the cell membrane in control cells but in the cytoplasm and nucleus of pressured cells. In contrast, N-cadherin was associated with the cell membrane in pressure cells. Changes in the location of these two proteins in ECs have previously been associated with increase proliferation and changes in cell morphology. Therefore the hydrostatic pressure induced changes in these may be mediated, in part, by beta-catenin and N-cadherin.
  • Tsubasa Matsui, Shinji Deguchi, Toshiro Ohashi, Masaaki Sato
    FUTURE MEDICAL ENGINEERING BASED ON BIONANOTECHNOLOGY, PROCEEDINGS 129 - + 2006 [Refereed][Not invited]
     
    As only a few studies have been reported so far, mechanical properties of stress fibers (SF) have poorly been understood, mainly due to difficulties with experimental technique. A previous study in our laboratory has described development of a micro-tensile tester to measure tensile properties of single SF chemically isolated from vascular smooth muscle cells (VSMC). Tensile tests were performed at a single strain rate of 0.02 s(-1). In this study, the strain rate dependency of tensile properties of SF was investigated. The results showed that the initial stiffness of SF at strain rate of 0.01 s(-1), 0.05 s(-1) and 0.1 s(-1) were 0.010 +/- 0.004 N/m, 0.006 +/- 0.006 N/m, 0.028 +/- 0.0 11 N/m, respectively. Averaged force-strain relationships were found to be almost linear in the physiological strain range of 0.0-0.4 for 0.01 s(-1) and 0.05 s(-1). These linear relationships were consistent with a previous report at a strain rate of 0.02 s(-1). In contrast, the force-strain relationship was nonlinear for the 0.1 s(-1) strain rate. The reason for this difference is unclear, possibly due to a threshold in strain rate between 0.0 1 s(-1) and 0.1 s(-1). Mechanical properties of SF may be strain rate dependent, particularly having a threshold in the linearity of the force-strain relationships.
  • Toshiro Ohashi, Junichi Yamazaki, Norifumi Kameda, Masaaki Sato
    2006 IEEE INTERNATIONAL SYMPOSIUM ON MICRO-NANOMECHATRONICS AND HUMAN SCIENCE 140 - + 2474-378X 2006 [Refereed][Not invited]
     
    Cellular traction forces are known to play an important role in the interactions between cells and their substrates. Although several studies have estimated traction forces using microfabrication techniques, little is known of how intracellular structures contribute to traction forces. In this study, microfabricated substrates with different arrays of microposts were used to estimate traction forces of smooth muscle cells, particularly exploring the contribution of cytoskeletal structures to traction forces. Smooth muscle cells isolated from bovine aortas were transfected with fluorescent protein conjugates, and then plated on an elastomer substrate with arrays of microposts (3 pin in diameter, 10 pin in height, 6 or 10 pin in spacing). The total area of the microposts arrays was designed to be 30 pin x 30 pin to control cell area. A BrdU incorporation assay revealed that cell proliferation significantly decreased when the spacing of microposts was increased from 6 to 10 mu m. A significant increase in traction forces from 15.2 +/- 1.4 to 22.4 +/- 1.8 nN was also found by increasing the spacing of the microposts, although less organization of stress fibers was observed. This result indicates that, for the 10 pin spacing, actin-myosin interactions may be more activated, producing higher forces per microposts. Moreover, when microtubules were disrupted, using 10 mu g/ml nocodazole, a 30% increase in traction forces was observed in cells plated on both sets of microposts. Therefore, this study proposes that the contraction forces of stress fibers are mainly involved in cellular traction forces, and that microtubules, directly or indirectly, contribute to the contraction forces of stress fibers.
  • 藤原 英記, 小田, 克彦, 齋木, 佳克, 吉岡, 一朗, 河津, 聡, 佐藤, 真一, 坂元, 尚哉, 大橋, 俊朗, 佐藤, 正明, 田畑, 泰彦, 田林 晄一
    The Japanese Journal of THORACIC AND CARDIOVASCULAR SURGERY シュプリンガー・ジャパン(株) 53 (Suppl.II) 395 - 395 1344-4964 2005/09 [Not refereed][Not invited]
  • Hiroshi Ozawa, Takeo Matsumoto, Toshiro Ohashi, Masaaki Sato, Shoichi Kokubun
    Journal of neurosurgery. Spine 1 (1) 122 - 7 1547-5654 2004/07 
    OBJECT: The pia mater has received little attention regarding its function in the deformation of the spinal cord under compression. In this study the mechanical properties and function of the spinal pia mater were investigated using three methods. METHODS: Spinal cord segments were excised from rabbits. The elastic modulus of the pia mater was measured by performing a tensile test using specimens with the pia mater intact and ones with the pia mater stripped off. The stiffness of the spinal cord was examined by performing a compression test with specimens containing an intact pia mater and ones with a pia mater that was incised at both sides. The cross-sectional area and circumference of the spinal cord were measured on axial views of magnetic resonance images in patients with cervical disc herniations before and after surgery. The pia mater had an elastic modulus of 2300 kPa, which was 460 times higher than that of spinal cord parenchyma. By covering the parenchyma, it tripled the overall elastic modulus of the spinal cord. The pia mater increased the stiffness of the spinal cord and enhanced its shape recovery after removal of the compression. The cross-sectional area of the spinal cord increased after surgery, whereas the circumference of the spinal cord changed little. CONCLUSIONS: The pia mater firmly covers the spinal cord and has a high elastic modulus; it therefore provides a constraint on the spinal cord surface. It prevents elongation of the circumference and produces a large strain energy that is responsible for shape restoration following decompression.
  • Finite element analysis of sheared endothelial cells using structural optimization method
    S. Seo, T. Ohashi, T. Matsumoto, M. Sato
    Proceedings of The First International Symposium on Advanced Fluid Information 143 - 146 2001 [Not refereed][Not invited]
  • T Ohashi, H Sugawara, T Matsumoto, M Sato
    JSME INTERNATIONAL JOURNAL SERIES C-MECHANICAL SYSTEMS MACHINE ELEMENTS AND MANUFACTURING 43 (4) 780 - 786 1340-8062 2000/12 [Refereed][Not invited]
     
    Intracellular stress distributions in cultured endothelial cells exposed to fluid shear stress were studied using finite-element analysis, in which cell surface geometries were measured by atomic force microscopy. After applying shear stress, the endothelial cells showed marked elongation and orientation in the flow direction. Finite-element analysis indicated that wall shear stress in the central region around the nucleus was significantly higher than those in the upstream and downstream sides for both control, statically cultured cells, and sheared cells, but the intracellular stress concentrations between the central regions and the other two sides reduced with fluid flow. Fluorescent images showed that stress fibers of F-actin bundles were mainly formed in the central portion of the cells. These results indicate that endothelial cells may change their cytoskeletal structures to increase their stiffness and reduce stress concentrations in the cells.
  • An application of the pipette technique to the measurement of local mechanical properties of aortic walls
    M. Sato, T. Matsumoto, T. Ohashi
    Proceedings of Toyota International Symposium on Human Life Support Biomechanics 147 - 154 2000 [Not refereed][Not invited]
  • T Ohashi, Y Kato, T Matsumoto, M Sato
    JSME INTERNATIONAL JOURNAL SERIES C-MECHANICAL SYSTEMS MACHINE ELEMENTS AND MANUFACTURING 42 (3) 568 - 573 1340-8062 1999/09 [Refereed][Not invited]
     
    Local elastic moduli in the wall of porcine thoracic aortas were measured in the axial, circumferential, and radial directions by the pipette aspiration method. The local elastic moduli in the three directions were found to be independent of wall position. Then, histological analysis using color classification based on Mahalanobis' generalized distance was performed to quantitatively assess the correlation between elastic moduli and histology. Porcine thoracic aortas had a typical layered structure. The area fraction values, i.e,, the percentage area of three structural components, elastin, collagen fibers and smooth muscle cells, were almost uniform in the wall. On the other hand, the moduli in bovine aortas reported previously decreased significantly from the inner to the outer sides of the wall,The histological structure was not homogeneous, that is, large clusters of smooth muscle cells existed in the outer side. These results suggest that the intramural distributions of elastic moduli are closely correlated with the histological structure.
  • Local elastic moduli of aortic walls measured by pipette aspiration technique
    T. Ohashi, T. Matsumoto, T. Aoki, M. Sato
    Proceedings of 4th China-Japan-USA-Singapore Conference on Biomechanics 115 - 118 1995 [Not refereed][Not invited]

MISC

  • 有限要素法による直立二足歩行時の仙腸関節の動きと負荷変化の解析
    黒澤 大輔, 豊原 涼太, Hammer Niels, Uwe Lingslebe, 本田 啓太, 関口 雄介, 出江 紳一, 村上 栄一, 小澤 浩司, 大橋 俊朗  Journal of Spine Research  11-  (3)  456  -456  2020/03
  • TOYOHARA Ryota, KUROSAWA Daisuke, HAMMER Niels, WERNER Michael, HONDA Keita, SEKIGUCHI Yusuke, IZUMI Shin-Ichi, MURAKAMI Eiichi, OZAWA Hiroshi, OHASHI Toshiro  The Proceedings of the JSME Conference on Frontiers in Bioengineering  2020.31-  1C14  2020  [Not refereed]
  • 軟骨細胞の力学的ストレス応答におけるスフィンゴ糖脂質の機能解析
    松原 新史, 小野寺 智洋, 門間 太輔, 馬場 力哉, 本谷 和俊, 上徳 善太, 宝満 健太郎, 岩崎 倫政, 大橋 俊朗, 前田 英次郎  北海道整形災害外科学会雑誌  60-  (1)  145  -146  2018/08  [Not refereed][Not invited]
  • 軟骨細胞の力学的ストレス応答におけるスフィンゴ糖脂質の機能解析
    松原 新史, 小野寺 智洋, 門間 太輔, 馬場 力哉, 本谷 和俊, 上徳 善太, 宝満 健太郎, 岩崎 倫政, 前田 英次郎, 大橋 俊朗  北海道整形災害外科学会雑誌  59-  (2)  253  -254  2018/03  [Not refereed][Not invited]
  • 松原新史, 小野寺智洋, 前田英次郎, 門間太輔, 馬場力哉, 本谷俊彦, 上徳善太, 宝満健太郎, 大橋俊朗, 大橋俊朗, 岩崎倫政  日本整形外科学会雑誌  91-  (8)  S1774  -S1774  2017/08/31  [Not refereed][Not invited]
  • 軟骨細胞の力学的ストレス応答におけるスフィンゴ糖脂質の機能解析
    松原新史, 小野寺智洋, 前田英次郎, 門間太輔, 馬場力哉, 本谷和俊, 上徳善太, 宝満健太郎, 大橋俊朗, 岩崎倫政  第30回日本軟骨代謝学会  120-  2017/03  [Not refereed][Not invited]
  • Depletion Of Glycosphingolipids Induces The Excessive Response Of Chondrocytes Under Mechanical Stress Condition.
    Shinji Matsubara, Tomohiro Onodera, Eijiro Maeda, Daisuke Momma, Masatake Matsuoka, Rikiya Baba, Kazutoshi Hontani, Zenta Joutoku, Kentarou Homan Homan, Toshiro Ohashi, Norimasa Iwasaki  The 63th Annual Meeting of Orthopaedic Research Society (ORS)  2017/03  [Not refereed][Not invited]
  • PIAN Haicheng, MAEDA Eijiro, OHASHI Toshiro  バイオエンジニアリング講演会講演論文集  2016-  (28)  "2D15  -1"-"2D15-4"  2016/01/09  
    Gap junction communication is known to play essential roles in tenocyte functions in response to mechanical loading. In our previous study, it was exhibited that an application of static tensile strain with a physiological amplitude for 1h promoted gap junction communication in tenocytes while overloading strain suppressed gap junction communication. However, temporal regulation of gap junction communication during an extended static loading has not been characterized. Accordingly, the present study tested a hypothesis that gap junction communication is regulated in a manner depending on the duration of mechanical loading. Gap junction communication was investigated using a fluorescence loss in photobleaching (FLIP) technique. It was demonstrated that the level of gap junction communication was fluctuated under physiological 4% static strain during 24h period, while it was gradually increased under 8% static strain.
  • MAEDA Eijiro, TASHIRO Masataka, OHASHI Toshiro  バイオエンジニアリング講演会講演論文集  2016-  (28)  "2D33  -1"-"2D33-5"  2016/01/09  
    Tendinopathy has been one of major health concerns in orthopaedic surgery, not only for active athletes but also for others like office workers. Although the exact etiology of tendinopathy has yet to be elucidated, it has been demonstrated that tendon core temperature elevated up to 43 to 45 degree Celsius when humans or racehorses are subjected to extensive exercises (harsh running). A long exposure to such high temperature could induce cell death. However, effects of the exposure to the high temperature for relatively short terms (simulating running activity) on tenocyte functions are still not understood. Therefore, the present study was performed to investigate how tenocyte anabolism and catabolism are altered by a short thermal stimulation. In addition, the present study tested a hypothesis that such alterations of tenocyte functions are dependent in the presence of gap junction intercellular communication (GJIC). It was demonstrated that tenocyte GJIC was not significantly altered by a 30 minutes exposure to 43 degree. However, catabolic and inflammatory gene expressions were significantly upregulated following a 24 h post-heating culture period. With the presence of GJ blocker, upregulation of catabolic genes by the thermal stimulation was inhibited, although inflammatory gene expression was further enhanced. These results may indicate that GJIC in part contribute to alterations in tenocyte functions in response to thermal stimulation in a complicated manner.
  • 佐藤 岳彦, 大橋 俊朗, 川野 聡恭, 白樫 了  日本機械学會誌  118-  (1165)  758  -758  2015/12/05
  • Ohashi Toshiro, Hagiwara Yasufumi, Wang James H-C, Maeda Eijiro  Abstracts of ATEM : International Conference on Advanced Technology in Experimental Mechanics : Asian Conference on Experimental Mechanics  2015-  (14)  255  -255  2015/10/04
  • MAEDA Eijiro, Muramatsu Tomomi, Ohashi Toshiro  Mechanical Engineering Congress, Japan  2015-  "J0220101  -1"-"J0220101-4"  2015/09/13  
    Tendon is known to remodel its structure and alter mechanical properties in response to changes in its mechanical environment. Tendon cells, tenocytes, are thought to play significant roles in tendon tissue remodelling, by responding to mechanical stress. Like other adherent cells, tenocytes exert traction forces to their substrate when the cells attach to the substrate. Cell traction forces are thought to reflect the level of intracellular tension generated in actin cytoskeletons. It has been shown that intracellular tension level is a regulatory factor of tenocyte catabolism. Accordingly, the present study hypothesized that tenocyte response to interleukin-1β is dependent on the level of intracellular tension. Tenocytes, isolated from rabbit Achilles tendons, were seeded microfabricated, elastic micropillar substrates, and were subjected to the treatment of IL-1β at four different concentrations. It was demonstrated that tenocyte traction force level was dependent on the elastic modulus of micropillar substrates. In addition, cells on softer substrates were more prone to the stimulation of the catabolic agent, exhibiting a greater change in cell morphology. These results suggest that tenocytes with a low level of intracellular tension, which implies that the cells are catabolic, are stimulated by catabolic/inflammatory agents and further enhance their catabolism.
  • NAKAJIMA Toshiaki, MAEDA Eijiro, MATSUBARA Shinji, MOMMA Daisuke, ONODERA Tomohiro, IWASAKI Norimasa, OHASHI Toshiro  バイオエンジニアリング講演会講演論文集  2015-  (27)  129  -130  2015/01/08
  • TASHIRO Masataka, MAEDA Eijiro, OHASHI Toshiro  バイオエンジニアリング講演会講演論文集  2015-  (27)  101  -102  2015/01/08
  • MAEDA Eijiro, OHASHI Toshiro  Mechanical Engineering Congress, Japan  2014-  "J0270204  -1"-"J0270204-4"  2014/09/07  
    Gap junctions have been known to be involved in regulations of cell functions. It is also suggested that gap junction intercellular communications (GJICs) between tenocytes play an essential role in collagen synthesis in response to mechanical loadings. However, there were little knowledge on how GJICs are mechanically regulated and how changes in GJIC affect tenocyte functions. Accordingly, the present study has proposed a new numerical model to estimate intra- and intercellular diffusivity, based on one-dimensional diffusion theory, by fitting the model to data obtained from FLIP experiments, which visualises intercellular mass transport using a confocal laser microscope. Tenocytes were seeded within microgrooves integrated in a custom-made PDMS device, and were loaded with calcein-AM. FLIP experiment consisted of a series of instantaneous 100% power laser irradiation provided to one of tenocytes in a series every 2.6 second for 99 times, and fluorescence decay profiles of the targeted tenocyte and its neighbouring cells were obtained. Using the numerical model, intracellular and intercellular diffusion coefficient of tenocytes under static culture was estimated to be 25.3 and 1.06 μm^2/sec, respectively. The latter was significantly increased by an application of physiological, 4% tensile strain to the tenocytes, whereas that was significantly reduced by excessive, 8% tensile strain to the cells. Therefore, it can be concluded that physiological mechanical loading enhances intercellular communications, whereas non-physiological, overloading disrupt them.
  • OHASHI Toshiro, NAKATA Yu  M&M材料力学カンファレンス  2014-  "OS0612  -1"-"OS0612-2"  2014/07/19  
    Since articular chondrocytes are in vivo exposed to complex mechanical environment, it is important to study whether chondrocyte responses to such mechanical environment are closely related to articular cartilage diseases such as osteoarthritis. In this study, chondrocytes are exposed to an impact loading with drop tower impact testing and the associated cell deformation is examined based on actin filament structures. Cell aspect ratio, an index of cell deformation, was significantly decreased after impact loading, exhibiting a circular shape for the control group and an ellipsoidal shape for the impact groups. This finding suggests that chondrocytes can be deformed by impact loading, possibly leading to an alternation in cell physiological functions.
  • TSUCHIYA Hitoshi, MAEDA Eijiro, KUROKAWA Takayuki, GONG Jian Ping, KITAMURA Nobuto, YASUDA Kazunori, OHASHI Toshiro  バイオエンジニアリング講演会講演論文集  2014-  (26)  481  -482  2014/01/10
  • MAEDA Eijiro, SUGIMOTO Megumi, KOSATO Yotaro, OHASHI Toshiro  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2013-  (24)  57  -58  2013/10/31
  • OHASHI Toshiro, SUGAWARA Akito, Justin Cooper-White  M&M材料力学カンファレンス  2013-  "OS0712  -1"-"OS0712-2"  2013/10/12  
    Cell migration plays an important role in many physiological and pathological events, however, the mechanism of collective cell migration has not been well understood. This study performs traction force microscopy on migrating NIH 3T3 fibroblasts using a multichanel device. The device consists of a reservoir for cell culture and 24 microchannels with arrays of micropillars for traction force microscopy. Here, the effect of anisotropic properties in stiffness of substrate on collective cell migration is tested with ellipsoidal cross section of micropillars. The results showed that cells migrated into the microchannels and produced higher traction forces in the direction of major axis of micropillars. This finding suggests that cell can sense the stiffness of substrate and produce traction forces depending on the stiffness, resulting in difference in cell migration rate.
  • OHASHI Toshiro, SUGAWARA Akito, COOPER-WHITE Justin  Mechanical Engineering Congress, Japan  2013-  "J026036  -1"-"J026036-3"  2013/09/08  
    Cell migration plays an important role in many physiological and pathological processes such as morphogenesis, wound healing, and tumor metastasis. Although the majority of such events occur with cells moving as a group, called collective cell migration, the mechanism of collective cell migrations has not been well understood. This study performs traction force microscopy on migrating NIH 3T3 fibroblasts by using a microfabricated device to clarify the mechanism of collective cell migration. The device consists of a reservoir for establishing a confluent cell monolayer and 24 microchannels with arrays of micropillars that are used for traction force microscopy. Here the effect of anisotropic properties in stiffness of substrate on cell migration is tested with ellipsoidal cross section of micropillars. Cells produced higher traction forces in the direction of major axis of micropillars. Moreover, cells migrated faster when the major axis of micropillars is parallel to the direction of microchannels. These results suggest that cell can sense the stiffness of substrate and produce traction forces depending on the stiffness, leading to modulation of cell migration rate.
  • 大橋 俊朗, 亀田 憲史, 中村 昌司, 佐藤 正明  Journal of the Japan Society of Mechanical Engineers  116-  (1134)  289  -289  2013/05/05
  • 前田英次郎, 堤健博, 黒川孝幸, 北村信人, GONG Jian Ping, 安田和則, 大橋俊朗  バイオエンジニアリング講演会講演論文集  25th-  (25)  221  -222  2013/01/08  [Not refereed][Not invited]
  • Sugimoto Megumi, Maeda Eijiro, Ohashi Toshiro  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2012-  (23)  125  -126  2012/10/04
  • Niida Yusaku, Sato Masaaki, Ohashi Toshiro  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2012-  (23)  117  -118  2012/10/04
  • Sakai Manabu, Weibull Emilie, Andersson-Svahn Helene, Ohashi Toshiro  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2012-  (23)  151  -152  2012/10/04
  • NIIDA Yusaku, SATO Masaaki, OHASHI Toshiro  Mechanical Engineering Congress, Japan  2012-  "J028034  -1"-"J028034-3"  2012/09/09  
    Vascular endothelial cells (ECs) are lining the entire vascular trees and constantly exposed to blood flow. ECs demonstrate elongation and orientation in response to blood flow and physiological functions of ECs are known to be affected by fluid shear stress. Adherent cells such as ECs are known to generate contractile forces to their substrate through focal adhesion sites as traction forces. Cell traction forces reveal not only intracellular mechanics but also physiological state. In order to monitor changes of traction forces of ECs in response to fluid flow, we have newly developed a flow-imposed device, in which a substrate with arrays of microfabricated elastomeric pillars is integrated at the bottom of a parallel plate flow chamber. The present study was performed to characterize spatio-temporal distributions of traction forces of ECs undergoing remodeling process to fluid shear stress and to examine the relationship between the changes of cell traction forces and cell morphology.
  • SUGIMOTO Megumi, MAEDA Eijiro, OHASHI Toshiro  Mechanical Engineering Congress, Japan  2012-  "J028023  -1"-"J028023-4"  2012/09/09  
    Tendon is subjected to dynamic mechanical loading in vivo and thus effects of cyclic tensile loading on tenocyte metabolism have been well characterized. However, the relationship between cell metabolism and cytoskeletal tension has not been studied in detail. Elastic micropillars made from silicone elastomer (PDMS) have been proposed to measure cellular traction forces which reflect cytoskeletal tension. Micropillar substrates with three different stiffness were prepared by changing the height of the pillars. First, to examine the effects of the substrate stiffness, tenocytes isolated from bovine tendons were seeded on the micropillar substrates. At the end of 24 hrs incubation period, cellular traction forces were determined. Real-time qPCR was also performed to examine the expressions of type I collagen (anabolic gene) and MMP-1 (catabolic gene) mRNA from tenocytes. In addition, to examine the role of acto-myosin contractility, tenocytes were treated with myosin II inhibitor, blebbistatin at 20 hrs. It was found that there were significant increases in cellular traction forces with increasing stiffness of micropillar substrates. Although there were no significant differences in the expression level of type I collagen among the three substrates, significant increases were observed in the expression level of MMP-1 with decreasing in stiffness of substrate. Moreover, traction forces decreased and the expression level of MMP-1 increased with the treatment of blebbistatin. This may indicate that intracellular cytoskeletal tension, driven by acto-myosin contractility, could strongly influence tenocyte catabolism rather than anabolism.
  • SUGITA Shukei, MATSUMOTO Takeo, OHASHI Toshiro, KUMAGAI Kiichiro, AKIMOTO Hiroji, TABAYASHI Koichi, SATO Masaaki  東海支部総会講演会講演論文集  2012-  (61)  "702  -1"-"702-2"  2012/03/15
  • SUGAWARA Akito, MAEDA Eijiro, COOPER-WHITE Justin John, OHASHI Toshiro  バイオエンジニアリング講演会講演論文集  2012-  (24)  "7C41  -1"-"7C41-2"  2012/01/06
  • MATSUI Shunsuke, MAEDA Eijiro, WEIBULL Emilie, ANDERSSON-SVAHN Helene, OHASHI Toshiro  バイオエンジニアリング講演会講演論文集  2012-  (24)  "7C22  -1"-"7C22-2"  2012/01/06
  • NIIDA Yusaku, MAEDA Eijiro, SATO Masaaki, OHASHI Toshiro  バイオエンジニアリング講演会講演論文集  2012-  (24)  "7C42  -1"-"7C42-2"  2012/01/06
  • SUGIMOTO Megumi, MAEDA Eijiro, OHASHI Toshiro  バイオエンジニアリング講演会講演論文集  2012-  (24)  "7F41  -1"-"7F41-2"  2012/01/06
  • HAGIWARA Yasufumi, MAEDA Eijiro, WANG James H.C., OHASHI Toshiro  バイオエンジニアリング講演会講演論文集  2012-  (24)  "7E42  -1"-"7E42-2"  2012/01/06
  • 堤健博, 前田英次郎, 北村信人, 安田和則, 大橋俊朗  バイオエンジニアリング講演会講演論文集  24th (CD-ROM)-  (24)  ROMBUNNO.7F42  -1"-"7F42-2"  2012/01/06  [Not refereed][Not invited]
  • SUGAWARA Akito, OHASHI Toshiro  Mechanical Engineering Congress, Japan  2011-  "S022031  -1"-"S022031-4"  2011/09/11  
    Cell migration is essential for a variety of biological and pathological processes such as wound healing, inflammation and tumor metastasis. However, a mechanical field within a group of multiple cells during a collective migration has not been characterized well. In this study, a polydimethylsiloxane-made multichannel device was fabricated using photolithography and soft lithography, and was used to monitor traction forces generated by cells during migration. The device consists of a reservoir for establishing a confluent cell monolayer, attached with 24 microchannels for cell migration. Four patterns of micropillars were designed on the bottom surface of the microchannels to measure cell traction forces during migration on four combinations of substrate stiffness and topography. Collective cell migration from the reservoir into the microchannels was initiated when the channels were backfilled with culture medium. Migration rate of 5.7 pm/h was measured in the microchannel equipped with micropillars whose dimensions were 6.5 pm in height, 2 pm in diameter and 7 pm in spacing. In detail, cells on the moving front of the migration, leading cells, generated the traction forces toward the backward and the maximum magnitude of 14 nN was measured at their front side. Traction forces generated by the cells behind the leading cells directed backward at both the front and the rear sides. However, traction forces generated by the cells behind the second row directed random directions with smaller magnitudes compared to those on the front and the second lines. It is assumed that cells on the front line generated a large magnitude of traction forces and migrated actively as single cells, pulling adjacent cells forward, whereas the movement of cells after third row was restricted by mechanical linkages between their neighboring cells, generating a small magnitude of traction forces.
  • SUGIMOTO Megumi, MAEDA Eijiro, OHASHI Toshiro  Mechanical Engineering Congress, Japan  2011-  "S022024  -1"-"S022024-4"  2011/09/11  
    Tendon is subjected to dynamic mechanical loading in vivo. Effects of cyclic tensile loading on tenocyte metabolism have been characterized well. However, the relationship between cell metabolism and internal tension within cytoskeleton has not been studied in detail. Elastic micropillars made from silicone elastomer (PDMS) have been proposed to measure cellular traction forces which reflect cytoskeletal tension. Micropillar substrates with three different elastic moduli were prepared by changing the height of the pillars. After tenocytes were seeded on each substrate for 24 hours, traction forces were determined. Real-time qPCR was also performed to examine the expressions of type I collagen (anabolic gene) and MMP-1 (catabolic gene) mRNA from tenocytes on these substrates. It was found that there were significant increases in cellular traction forces with the elastic modulus of micropillar substrates. Although there were no significant differences in the expression level of type I collagen among three substrates, increases were observed in the expression level of MMP-1 with decreases in the elastic modulus of substrate. This may indicate that intracellular cytoskeletal tension strongly influences tenocyte catabolism rather than anabolism.
  • NIIDA Yusaku, OHASHI Toshiro  Mechanical Engineering Congress, Japan  2011-  "S022021  -1"-"S022021-3"  2011/09/11  
    This paper presents development of a technique to control cell adhesion using an array of magnetic microbeads. Using MEMS techniques, we fabricated an array of microholes (3 gm in diameter and 2 gm in depth) on the surface of a glass substrate, to trap magnetic microbeads (2.8 gm in diameter) in the microholes. The glass substrate was integrated with a PDMS-made device and a suspension of magnetic microbeads was then introduced into the device by a syringe pump. The magnetic microbeads were coated with cell adhesive ligands prior to the loading. A permanent magnet was placed under the substrate and was used to direct microbeads into the microholes, followed by an introduction of cell suspension to the device. After a 24 hours incubation, it was observed that cells attached only to the magnetic microbeads, but not to the other parts of the glass surface. The current method of controlling cell adhesion would be useful to develop advanced cell culture system.
  • MATSUI Shunsuke, OHASHI Toshiro  Mechanical Engineering Congress, Japan  2011-  "S022035  -1"-"S022035-4"  2011/09/11  
    Single cell analysis has been of a great interest in bioengineering as it can highlight heterogeneity of individual cells in medical screening tests compared to conventional bulk analysis. We have been developing a novel bioassay system designed for single cell analysis. The device mainly consists of a silicon/glass microwell slide and a PDMS microchannel, and enables to supply reagents to microwells in a well-controlled manner using micropumps. In this study, bovine aortic endothelial cells were cultured in the microwells for 48 h and then subjected to fluid shear stress at up to 2 Pa for 6 h. A result showed that realignment and elongation of endothelial cells to the flow direction were dependent on the level of shear stress. It was confirmed that the device could reduce consumption of expensive reagents and precious all samples. Moreover, unlike conventional bulk devices, multiple experimental conditions can be examined at the same time within a single device. It can be concluded that the device has a potential to be applied to multi-purpose single cell tests.
  • HAGIWARA Yasuhumi, MAEDA Eijiro, Wang James H-C, OHASHI Toshiro  M&M材料力学カンファレンス  2011-  "OS1105  -1"-"OS1105-2"  2011/07/16  
    Tendon is subjected to continuous mechanical loading, which imposes cyclic tensile strain and interstitial fluid shear stress to tenocytes. Although a number of studies have characterized mechanical responses of tenocytes to cyclic tensile strain, only few studies have been performed to examine tenocyte responses to fluid shear stress. The present study proposes a newly fabricated cell culture device using MEMS technologies, which enables to apply cyclic tensile strain or fluid shear stress, or both simultaneously, to tenocytes seeded onto microgrooves. It was confirmed that tenocytes exhibited a similar morphology as observed in vivo. Numerical analysis of flow patterns within the device demonstrated that a wide range of fluid shear stress, including a previously predicted physiological value, could be applied to cells. In summary, the present study developed a novel experimental model, which will reveal further details of tenocyte mechanotransduction events.
  • HAGIWARA Yasufumi, OHASHI Toshiro  バイオエンジニアリング講演会講演論文集  2010-  (23)  583  -584  2011/01/07
  • TANAKA Ryoichi, SATO Masaaki, OHASHI Toshiro  バイオエンジニアリング講演会講演論文集  2010-  (23)  153  -154  2011/01/07  [Not refereed][Not invited]
  • OHASHI Toshiro, KOHATA Kengo  M&M材料力学カンファレンス  2010-  379  -380  2010/10/09  [Not refereed][Not invited]
     
    In this study, it is hypothesized that initial geometrical shapes of cutaneous wound and the major axis of the wound to the direction of stretch could influence myofibroblast realignment observed in the maturation phase of healing process. Firstly, human dermal fibroblasts were differentiated into myofibroblasts with a treatment of TGF-β1. The cells were then seeded onto a PDMS chamber either in a monolayer or in a colony of the following shapes: circle, and track fields both parallel and vertical to the stretch. For mechanical tests, cyclic tensile stretch, with an amplitude of 20%, was pr...
  • Sugita Shukei, Matsumoto Takeo, Ohashi Toshiro, Kumagai Kiichiro, Akimoto Hiroji, Tabayashi Koichi, Sato Masaaki  The proceedings of the JSME annual meeting  2010-  (5)  51  -52  2010/09/04  [Not refereed][Not invited]
     
    Mechanical properties of the thoracic aortic aneurysm (TAA) tissues were measured in a pressure-imposed test along with histological analysis to estimate their rupture risk. Quadrilateral specimens obtained from TAA were sandwiched with plates having a hole and pressurized from one side of the hole to bulge into the other side until rupture. Intramural stress and strain of each specimen were calculated from its deformation-loading pressure relationship to obtain its ultimate strength and tangent modulus. Histological analysis was then performed to obtain the area fraction of histological co...
  • Oya Koki, Sakamoto Naoya, Ohashi Toshiro, Sato Masaaki  バイオエンジニアリング講演会講演論文集  2009-  (22)  22  -22  2010/01/08  [Not refereed][Not invited]
  • SETOWAKI Sota, SUGITA Shukei, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2009-  (20)  95  -96  2009/11/06  [Not refereed][Not invited]
  • 渡辺 祥伍, 大橋 俊朗, 坂元 尚哉, 佐藤 正明  運動療法と物理療法 = The Journal of physical medicine  20-  (2)  145  -145  2009/06/11  [Not refereed][Not invited]
  • 大橋 俊朗, 花村 和彦, 我妻 大策, 坂元 尚哉, 佐藤 正明  Journal of the Japan Society of Mechanical Engineers  112-  (1086)  352  -353  2009/05/05  [Not refereed][Not invited]
  • Saito Naoki, Sakamoto Naoya, Ohashi Toshiro, Sato Masaaki  講演論文集  2009-  (44)  20  -21  2009/03/13  [Not refereed][Not invited]
  • HANAMURA Kazuhiko, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  講演論文集  2009-  (44)  176  -177  2009/03/13  [Not refereed][Not invited]
  • ISODA Takumi, UEKI Yousuke, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  講演論文集  2009-  (44)  178  -179  2009/03/13  [Not refereed][Not invited]
  • Y. Ueki, N. Sakamoto, T. Ohashi, M. Sato  EXPERIMENTAL MECHANICS  49-  (1)  125  -134  2009/02  [Not refereed][Not invited]
     
    It has been well established that mechanical stimuli including fluid shear stress and cyclic stretch play a key role in endothelial cell (EC) remodeling. However, in contrast to global remodeling to these mechanical stimuli, little is known of how local mechanical forces are transmitted through cells to induce cell remodeling leading to alteration in cell functions. In this study, we demonstrated that EC remodeling can be exerted by local tension generated in a neighboring EC. In this technique, a glass microneedle was used to apply local stretch in an EC in confluent monolayer and the resulting tension is transmitted to a neighboring EC across intercellular junctions. Local stretch induced reorientation and elongation of ECs parallel to the direction of stretch associated with reorganization of stress fibers. In addition, recruitment of Src homology 2-containing tyrosine phosphatase-2, binding to intercellular adhesion molecules platelet-endothelial cellular adhesion molecules-1, was selectively observed at the force-transmitted intercellular junctions after application of local stretch. These findings suggest that intercellular junctions can not only transmit but also sense local forces, and are potentially involved in EC mechanotransduction pathways.
  • Mori Kiichiroh, Ohashi Toshiro, Lindstrom Sara, Svahn Helene Andersson, Sato Masaaki  バイオエンジニアリング講演会講演論文集  2008-  (21)  97  -98  2009/01/22  [Not refereed][Not invited]
  • MATSUI Tsubasa, DEGUCHI Shinji, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2008-  (21)  129  -130  2009/01/22  [Not refereed][Not invited]
  • Saito Naoki, Sakamoto Naoya, Ohashi Toshiro, Sato Masaaki  バイオエンジニアリング講演会講演論文集  2008-  (21)  293  -294  2009/01/22  [Not refereed][Not invited]
  • Maruta Toshiyuki, Sakamoto Naoya, Ohashi Toshiro, Sato Masaaki  バイオエンジニアリング講演会講演論文集  2008-  (21)  295  -296  2009/01/22  [Not refereed][Not invited]
  • Oya Koki, Sakamoto Naoya, Ohashi Toshiro, Sato Masaaki  バイオエンジニアリング講演会講演論文集  2008-  (21)  297  -298  2009/01/22  [Not refereed][Not invited]
  • In vitro measurement of viscoelastic properties of actin bundles using micro-tensile tester with visual feedback control
    Matsui, T, Deguchi, S, Sakamoto, N, Ohashi, T, Sato, M  4th Asian Pacific Conference on Biomechanics  2009  [Not refereed][Not invited]
  • Development of microneedle-based viscoelastic test system for actin bundles
    Matsui, T, Deguchi, S, Sakamoto, N, Ohashi, T, Sato, M  9th International Symposium of Tohoku University Global COE Program  2009  [Not refereed][Not invited]
  • OHASHI Toshiro, NAKAMURA Shinji, SAKAMOTO Naoya, SATO Masaaki  運動療法と物理療法 = The Journal of physical medicine  19-  (3)  176  -183  2008/11/30  [Not refereed][Not invited]
  • 大橋 俊朗  日本機械学會誌  111-  (1077)  650  -650  2008/08/05  [Not refereed][Not invited]
  • 田中 英一, 大橋 俊朗, 大島 まり, 田中 英一, 田中 志信, 川野 聡恭  日本機械学會誌  111-  (1077)  650  -651  2008/08/05  [Not refereed][Not invited]
  • SUGITA Shukei, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  The proceedings of the JSME annual meeting  2008-  (8)  265  -266  2008/08/02  [Not refereed][Not invited]
     
    Microtubules (MTs) gliding on the kinesin-coated surface in vitro have a good feasibility for development of micro-scale transport systems. The key requirements in the development of the system include an image analysis to detect and recognize gliding MTs. In this study, we present an algorithm for detecting the position of the leading tips of gliding MTs. The algorithm operates on binarized fluorescent images of MTs and extracts the tips of MTs with a kernel. For verification, the algorithm was applied to a sample segment, and the distance between extracted coordinates with the algorithm a...
  • SAKAMOTO Naoya, SEGAWA Kei, OHASHI Toshiro, SATO Masaaki  Transactions of Visualization Society of Japan  28-  (1)  323  -324  2008/07/01  [Not refereed][Not invited]
  • 出口 真次, 大橋 俊朗, 佐藤 正明  Journal of the Japan Society of Mechanical Engineers  111-  (1074)  372  -373  2008/05/05  [Not refereed][Not invited]
  • 大橋 俊朗  Reports of Toyoda Physical and Chemical Research Institute.  (61)  131  -135  2008/05  [Not refereed][Not invited]
  • 熊谷 紀一郎, 本吉 直孝, 田林 晄一, 大橋 俊朗, 佐藤 正明  日本外科学会雑誌  109-  (2)  562  -562  2008/04/25  [Not refereed][Not invited]
  • Makoto Nishimura, Mirei Chiba, Toshiro Ohashi, Masaaki Sato, Yoshiyuki Shimizu, Kaoru Igarashi, Hideo Mitani  AMERICAN JOURNAL OF ORTHODONTICS AND DENTOFACIAL ORTHOPEDICS  133-  (4)  572  -583  2008/04  [Not refereed][Not invited]
     
    Introduction: Accelerating the speed of orthodontic tooth movement should contribute to the shortening of the treatment period. This would be beneficial because long treatment times are a negative aspect of orthodontic treatment. In this study, we evaluated the effects of mechanical stimulation by resonance vibration on tooth movement, and we showed the cellular and molecular mechanisms of periodontal ligament responses. Methods: The maxillary first molars of 6-week-old male Wistar rats were moved to the buccal side by using an expansive spring for 21 days (n = 6, control group), and the amount of tooth movement was measured. Additional vibrational stimulation (60 Hz, 1.0 m/s(2)) was applied to the first molars by using a loading vibration system for 8 minutes on days 0, 7, and 14 during orthodontic tooth movement (n = 6, experimental group). The animals were killed under anesthesia, and each maxilla was dissected. The specimens were fixed, decalcified, and embedded in paraffin. Sections were used for immunohistochemical analysis of receptor activator of NF kappa B ligand (RANKL) expression. The number of osteoclasts in the alveolar bone was counted by using TRAP staining, and the amount of root resorption was measured in sections stained with hematoxylin and eosin. Results: The average resonance frequency of the maxillary first molar was 61.02 +/- 8.38 Hz. Tooth movement in the experimental group was significantly greater than in the control group (P <. 05). Enhanced RANKL expression was observed at fibroblasts and osteoclasts in the periodontal ligament of the experimental group on day 3. The number of osteoclasts in the experimental group was significantly increased over the control group on day 8 (P <. 05). Histologically, there were no pathological findings in either group or significant differences in the amount of root resorption between the 2 groups. Conclusions: The application of resonance vibration might accelerate orthodontic tooth movement via enhanced RANKL expression in the periodontal ligament without additional damage to periodontal tissues such as root resorption.
  • Makoto Nishimura, Mirei Chiba, Toshiro Ohashi, Masaaki Sato, Yoshiyuki Shimizu, Kaoru Igarashi, Hideo Mitani  AMERICAN JOURNAL OF ORTHODONTICS AND DENTOFACIAL ORTHOPEDICS  133-  (4)  572  -583  2008/04  [Not refereed][Not invited]
     
    Introduction: Accelerating the speed of orthodontic tooth movement should contribute to the shortening of the treatment period. This would be beneficial because long treatment times are a negative aspect of orthodontic treatment. In this study, we evaluated the effects of mechanical stimulation by resonance vibration on tooth movement, and we showed the cellular and molecular mechanisms of periodontal ligament responses. Methods: The maxillary first molars of 6-week-old male Wistar rats were moved to the buccal side by using an expansive spring for 21 days (n = 6, control group), and the amount of tooth movement was measured. Additional vibrational stimulation (60 Hz, 1.0 m/s(2)) was applied to the first molars by using a loading vibration system for 8 minutes on days 0, 7, and 14 during orthodontic tooth movement (n = 6, experimental group). The animals were killed under anesthesia, and each maxilla was dissected. The specimens were fixed, decalcified, and embedded in paraffin. Sections were used for immunohistochemical analysis of receptor activator of NF kappa B ligand (RANKL) expression. The number of osteoclasts in the alveolar bone was counted by using TRAP staining, and the amount of root resorption was measured in sections stained with hematoxylin and eosin. Results: The average resonance frequency of the maxillary first molar was 61.02 +/- 8.38 Hz. Tooth movement in the experimental group was significantly greater than in the control group (P <. 05). Enhanced RANKL expression was observed at fibroblasts and osteoclasts in the periodontal ligament of the experimental group on day 3. The number of osteoclasts in the experimental group was significantly increased over the control group on day 8 (P <. 05). Histologically, there were no pathological findings in either group or significant differences in the amount of root resorption between the 2 groups. Conclusions: The application of resonance vibration might accelerate orthodontic tooth movement via enhanced RANKL expression in the periodontal ligament without additional damage to periodontal tissues such as root resorption.
  • Makoto Nishimura, Mirei Chiba, Toshiro Ohashi, Masaaki Sato, Yoshiyuki Shimizu, Kaoru Igarashi, Hideo Mitani  AMERICAN JOURNAL OF ORTHODONTICS AND DENTOFACIAL ORTHOPEDICS  133-  (4)  572  -583  2008/04  [Not refereed][Not invited]
     
    Introduction: Accelerating the speed of orthodontic tooth movement should contribute to the shortening of the treatment period. This would be beneficial because long treatment times are a negative aspect of orthodontic treatment. In this study, we evaluated the effects of mechanical stimulation by resonance vibration on tooth movement, and we showed the cellular and molecular mechanisms of periodontal ligament responses. Methods: The maxillary first molars of 6-week-old male Wistar rats were moved to the buccal side by using an expansive spring for 21 days (n = 6, control group), and the amount of tooth movement was measured. Additional vibrational stimulation (60 Hz, 1.0 m/s(2)) was applied to the first molars by using a loading vibration system for 8 minutes on days 0, 7, and 14 during orthodontic tooth movement (n = 6, experimental group). The animals were killed under anesthesia, and each maxilla was dissected. The specimens were fixed, decalcified, and embedded in paraffin. Sections were used for immunohistochemical analysis of receptor activator of NF kappa B ligand (RANKL) expression. The number of osteoclasts in the alveolar bone was counted by using TRAP staining, and the amount of root resorption was measured in sections stained with hematoxylin and eosin. Results: The average resonance frequency of the maxillary first molar was 61.02 +/- 8.38 Hz. Tooth movement in the experimental group was significantly greater than in the control group (P <. 05). Enhanced RANKL expression was observed at fibroblasts and osteoclasts in the periodontal ligament of the experimental group on day 3. The number of osteoclasts in the experimental group was significantly increased over the control group on day 8 (P <. 05). Histologically, there were no pathological findings in either group or significant differences in the amount of root resorption between the 2 groups. Conclusions: The application of resonance vibration might accelerate orthodontic tooth movement via enhanced RANKL expression in the periodontal ligament without additional damage to periodontal tissues such as root resorption.
  • Makoto Nishimura, Mirei Chiba, Toshiro Ohashi, Masaaki Sato, Yoshiyuki Shimizu, Kaoru Igarashi, Hideo Mitani  AMERICAN JOURNAL OF ORTHODONTICS AND DENTOFACIAL ORTHOPEDICS  133-  (4)  572  -583  2008/04  [Not refereed][Not invited]
     
    Introduction: Accelerating the speed of orthodontic tooth movement should contribute to the shortening of the treatment period. This would be beneficial because long treatment times are a negative aspect of orthodontic treatment. In this study, we evaluated the effects of mechanical stimulation by resonance vibration on tooth movement, and we showed the cellular and molecular mechanisms of periodontal ligament responses. Methods: The maxillary first molars of 6-week-old male Wistar rats were moved to the buccal side by using an expansive spring for 21 days (n = 6, control group), and the amount of tooth movement was measured. Additional vibrational stimulation (60 Hz, 1.0 m/s(2)) was applied to the first molars by using a loading vibration system for 8 minutes on days 0, 7, and 14 during orthodontic tooth movement (n = 6, experimental group). The animals were killed under anesthesia, and each maxilla was dissected. The specimens were fixed, decalcified, and embedded in paraffin. Sections were used for immunohistochemical analysis of receptor activator of NF kappa B ligand (RANKL) expression. The number of osteoclasts in the alveolar bone was counted by using TRAP staining, and the amount of root resorption was measured in sections stained with hematoxylin and eosin. Results: The average resonance frequency of the maxillary first molar was 61.02 +/- 8.38 Hz. Tooth movement in the experimental group was significantly greater than in the control group (P <. 05). Enhanced RANKL expression was observed at fibroblasts and osteoclasts in the periodontal ligament of the experimental group on day 3. The number of osteoclasts in the experimental group was significantly increased over the control group on day 8 (P <. 05). Histologically, there were no pathological findings in either group or significant differences in the amount of root resorption between the 2 groups. Conclusions: The application of resonance vibration might accelerate orthodontic tooth movement via enhanced RANKL expression in the periodontal ligament without additional damage to periodontal tissues such as root resorption.
  • 大橋 俊朗  生体医工学 : 日本エム・イー学会誌  46-  (1)  127  -127  2008/02/10  [Not refereed][Not invited]
  • Ito Kazushi, HANAMURA Kazuhiko, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2007-  (20)  109  -110  2008/01/24  [Not refereed][Not invited]
  • MATSUI Tsubasa, DEGUCHI Shinji, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2007-  (20)  105  -106  2008/01/24  [Not refereed][Not invited]
  • Sakamoto Naoya, Mochimaru Takato, Ohashi Toshiro, Sato Masaaki  バイオエンジニアリング講演会講演論文集  2007-  (20)  91  -92  2008/01/24  [Not refereed][Not invited]
  • KIUCHI Takuya, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2007-  (20)  89  -90  2008/01/24  [Not refereed][Not invited]
  • SUGITA Shukei, SETOWAKI Sota, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2007-  (20)  317  -318  2008/01/24  [Not refereed][Not invited]
  • MURASE Tatsuya, SUGITA Shukei, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2007-  (20)  315  -316  2008/01/24  [Not refereed][Not invited]
  • ICHIHARA Hirokazu, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2007-  (20)  309  -310  2008/01/24  [Not refereed][Not invited]
  • Y. Ueki, N. Sakamoto, T. Ohashi, M. Sato  BIORHEOLOGY  45-  (1-2)  96  -96  2008  [Not refereed][Not invited]
  • N. Sakamoto, T. Ohashi, M. Sato  The 13th International Conference on Biomedical Engineering  2008  [Not refereed][Not invited]
  • In vitro viscoelastic test apparatus for cytoskeletal bundles
    Matsui, T, Deguchi, S, Sakamoto, N, Ohashi, T, Sato, M  Proceedings of GPBE/NUS-Tohoku Graduate Student Conference in Bioengineering  2008  [Not refereed][Not invited]
  • A dynamic force-strain control system for isolated actin bundles
    Matsui, T, Deguchi, S, Sakamoto, N, Ohashi, T, Sato, M  Proceedings of Tohoku-NUS Student Joint Symposium  2008  [Not refereed][Not invited]
  • Intercellular junctions-mediated morphological responses of vascular endothelial cells induced by local stretch
    Experimental Mechanics  2008  [Not refereed][Not invited]
  • Intercellular junctions-mediated morphological responses of vascular endothelial cells induced by local stretch
    Experimental Mechanics  2008  [Not refereed][Not invited]
  • OHASHI Toshiro, KURODA Ryosuke, Knight Martin, Bader Dan, SATO Masaaki  The proceedings of the JSME annual meeting  2007-  (5)  223  -224  2007/09/07  [Not refereed][Not invited]
     
    Articular chondrocytes in vivo are continuously exposed to severe mechanical conditions and are supposed to exhibit reorganization of cytoskeletons, possibly altering mechanical properties of cells. In this study, the effect of local deformation on mechanical properties of chondrocytes was investigated using a micropipette aspiration technique, particularly focusing on actin remodeling. Repeat aspiration experiments showed that change in the Young's modulus of chondrocytes significantly depends on the aspiration rate. Ca^<2+> blocking experiments revealed that local deformation induces Ca^<...
  • Shinya Masuda, Yoshikatsu Saiki, Satoshi Kawatsu, Ichiro Yoshioka, Hidenori Fujiwara, Shunsuke Kawamoto, Sadahiro Sai, Atsushi Iguchi, Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato, Koichi Tabayashi  JOURNAL OF THORACIC AND CARDIOVASCULAR SURGERY  134-  (3)  723  -730  2007/09  [Not refereed][Not invited]
     
    Objectives: We created a new vascular clip designed for aortic surgery. The purposes of this investigation were to examine surgical applicability in a clinically relevant aortic replacement model and to assess biomechanical strength of the clipped anastomosis and serial histologic changes in the clipped anastomotic site. Methods: Twenty-one beagles underwent descending thoracic aortic replacement. Distal anastomosis was performed with the new clips, mimicking the cuffed anastomosis technique, and proximal anastomosis was carried out by conventional suture anastomosis. Tissue specimens of the anastomotic sites were harvested at 1, 3, 6, and 12 months postoperatively for examination. Results: There was no significant difference in the time required to carry out clip anastomosis ( 12.2 +/- 1.3 minutes) and suture anastomosis ( 13.7 +/- 0.9 minutes; P = .38). Neither type of anastomotic site was disrupted by raising the intraluminal pressure to 280 mm Hg. Microscopically, the areas of aortic wall compressed by vascular clips appeared as hyalinized areas adjacent to surrounding collagen fibers, with no significant infiltration of inflammatory cells. Identical histologic changes were observed at the site of the sutured anastomosis. The media at the clipped anastomosis site was significantly thinner than that at the sutured anastomosis site at 1 month after the operation. However, there was no significant difference in the thickness of the media at 3 months. Conclusions: The new vascular clips were effective in this clinically relevant model, with appropriate biomechanical strength, and the anastomotic sites underwent similar histologic changes to those observed after suture anastomosis.
  • Shinya Masuda, Yoshikatsu Saiki, Satoshi Kawatsu, Ichiro Yoshioka, Hidenori Fujiwara, Shunsuke Kawamoto, Sadahiro Sai, Atsushi Iguchi, Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato, Koichi Tabayashi  Journal of Thoracic and Cardiovascular Surgery  134-  (3)  723  -730  2007/09  [Not refereed][Not invited]
     
    Objectives: We created a new vascular clip designed for aortic surgery. The purposes of this investigation were to examine surgical applicability in a clinically relevant aortic replacement model and to assess biomechanical strength of the clipped anastomosis and serial histologic changes in the clipped anastomotic site. Methods: Twenty-one beagles underwent descending thoracic aortic replacement. Distal anastomosis was performed with the new clips, mimicking the cuffed anastomosis technique, and proximal anastomosis was carried out by conventional suture anastomosis. Tissue specimens of the anastomotic sites were harvested at 1, 3, 6, and 12 months postoperatively for examination. Results: There was no significant difference in the time required to carry out clip anastomosis (12.2 ± 1.3 minutes) and suture anastomosis (13.7 ± 0.9 minutes P = .38). Neither type of anastomotic site was disrupted by raising the intraluminal pressure to 280 mm Hg. Microscopically, the areas of aortic wall compressed by vascular clips appeared as hyalinized areas adjacent to surrounding collagen fibers, with no significant infiltration of inflammatory cells. Identical histologic changes were observed at the site of the sutured anastomosis. The media at the clipped anastomosis site was significantly thinner than that at the sutured anastomosis site at 1 month after the operation. However, there was no significant difference in the thickness of the media at 3 months. Conclusions: The new vascular clips were effective in this clinically relevant model, with appropriate biomechanical strength, and the anastomotic sites underwent similar histologic changes to those observed after suture anastomosis. © 2007 The American Association for Thoracic Surgery.
  • 大橋 俊朗, 亀田 憲史, 佐藤 正明  日本平滑筋学会雑誌  11-  (1)  "J  -21"  2007/06/26  [Not refereed][Not invited]
  • 大橋 俊朗  生体医工学 : 日本エム・イー学会誌  45-  (2)  186  -187  2007/06/10  [Not refereed][Not invited]
  • 大橋 俊朗, 黄 文敬, 坂元 尚哉, 佐藤 正明  運動療法と物理療法 = The Journal of physical medicine  18-  (2)  89  -89  2007/06/07  [Not refereed][Not invited]
  • Masaaki Sato, Kenichi Suzuki, Yosuke Ueki, Toshiro Ohashi  ACTA BIOMATERIALIA  3-  (3)  311  -319  2007/05  [Not refereed][Not invited]
     
    The surface topography and local elastic moduli of endothelial cells exposed to shear stress were measured using atomic force microscopy. Bovine aortic endothelial cells were exposed to shear stress of 2 Pa for 6, 12 or 24 h. In addition, a confocal laser-scanning microscope used in conjunction with the atomic force microscope was used to observe the actin filament structure of these endothelial cells to elucidate the relationship between mechanical properties and cytoskeletal structure. The elastic modulus, calculated using the Hertz model, was measured at 50 x 50 points at I put intervals within 40 min. For endothelial cells sheared for 6 h and 12 It, the elastic modulus at the upstream region was found to be higher than that at the downstream region. For endothelial cells sheared for 24 h, the elastic modulus at both the upstream and downstream regions increased. Fluorescent images showed thick, elongated actin filaments oriented in the direction of flow at the ventral surface of the cells. In the middle plane of the cells, actin filaments developed around the nucleus, while in the upper plane, short, thick actin filaments were observed but thick stress fibers were not present. The high elastic modulus came from the stress fibers. These results indicate that the higher elastic modulus observed in the upstream and downstream regions of sheared endothelial cells is mainly due to the development of stress fibers at the ventral surface and middle plane of the cell. (C) 2006 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.
  • Masaaki Sato, Kenichi Suzuki, Yosuke Ueki, Toshiro Ohashi  ACTA BIOMATERIALIA  3-  (3)  311  -319  2007/05  [Not refereed][Not invited]
     
    The surface topography and local elastic moduli of endothelial cells exposed to shear stress were measured using atomic force microscopy. Bovine aortic endothelial cells were exposed to shear stress of 2 Pa for 6, 12 or 24 h. In addition, a confocal laser-scanning microscope used in conjunction with the atomic force microscope was used to observe the actin filament structure of these endothelial cells to elucidate the relationship between mechanical properties and cytoskeletal structure. The elastic modulus, calculated using the Hertz model, was measured at 50 x 50 points at I put intervals within 40 min. For endothelial cells sheared for 6 h and 12 It, the elastic modulus at the upstream region was found to be higher than that at the downstream region. For endothelial cells sheared for 24 h, the elastic modulus at both the upstream and downstream regions increased. Fluorescent images showed thick, elongated actin filaments oriented in the direction of flow at the ventral surface of the cells. In the middle plane of the cells, actin filaments developed around the nucleus, while in the upper plane, short, thick actin filaments were observed but thick stress fibers were not present. The high elastic modulus came from the stress fibers. These results indicate that the higher elastic modulus observed in the upstream and downstream regions of sheared endothelial cells is mainly due to the development of stress fibers at the ventral surface and middle plane of the cell. (C) 2006 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.
  • Masaaki Sato, Kenichi Suzuki, Yosuke Ueki, Toshiro Ohashi  ACTA BIOMATERIALIA  3-  (3)  311  -319  2007/05  [Not refereed][Not invited]
     
    The surface topography and local elastic moduli of endothelial cells exposed to shear stress were measured using atomic force microscopy. Bovine aortic endothelial cells were exposed to shear stress of 2 Pa for 6, 12 or 24 h. In addition, a confocal laser-scanning microscope used in conjunction with the atomic force microscope was used to observe the actin filament structure of these endothelial cells to elucidate the relationship between mechanical properties and cytoskeletal structure. The elastic modulus, calculated using the Hertz model, was measured at 50 x 50 points at I put intervals within 40 min. For endothelial cells sheared for 6 h and 12 It, the elastic modulus at the upstream region was found to be higher than that at the downstream region. For endothelial cells sheared for 24 h, the elastic modulus at both the upstream and downstream regions increased. Fluorescent images showed thick, elongated actin filaments oriented in the direction of flow at the ventral surface of the cells. In the middle plane of the cells, actin filaments developed around the nucleus, while in the upper plane, short, thick actin filaments were observed but thick stress fibers were not present. The high elastic modulus came from the stress fibers. These results indicate that the higher elastic modulus observed in the upstream and downstream regions of sheared endothelial cells is mainly due to the development of stress fibers at the ventral surface and middle plane of the cell. (C) 2006 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.
  • Masaaki Sato, Kenichi Suzuki, Yosuke Ueki, Toshiro Ohashi  ACTA BIOMATERIALIA  3-  (3)  311  -319  2007/05  [Not refereed][Not invited]
     
    The surface topography and local elastic moduli of endothelial cells exposed to shear stress were measured using atomic force microscopy. Bovine aortic endothelial cells were exposed to shear stress of 2 Pa for 6, 12 or 24 h. In addition, a confocal laser-scanning microscope used in conjunction with the atomic force microscope was used to observe the actin filament structure of these endothelial cells to elucidate the relationship between mechanical properties and cytoskeletal structure. The elastic modulus, calculated using the Hertz model, was measured at 50 x 50 points at I put intervals within 40 min. For endothelial cells sheared for 6 h and 12 It, the elastic modulus at the upstream region was found to be higher than that at the downstream region. For endothelial cells sheared for 24 h, the elastic modulus at both the upstream and downstream regions increased. Fluorescent images showed thick, elongated actin filaments oriented in the direction of flow at the ventral surface of the cells. In the middle plane of the cells, actin filaments developed around the nucleus, while in the upper plane, short, thick actin filaments were observed but thick stress fibers were not present. The high elastic modulus came from the stress fibers. These results indicate that the higher elastic modulus observed in the upstream and downstream regions of sheared endothelial cells is mainly due to the development of stress fibers at the ventral surface and middle plane of the cell. (C) 2006 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.
  • UEKI Yosuke, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2006-  (19)  46  -47  2007/01/06  [Not refereed][Not invited]
  • SEGAWA Kei, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2006-  (19)  202  -203  2007/01/06  [Not refereed][Not invited]
  • Ito Kazushi, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2006-  (19)  370  -371  2007/01/06  [Not refereed][Not invited]
  • NAKAMURA Shouji, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2006-  (19)  372  -373  2007/01/06  [Not refereed][Not invited]
  • KURODA Ryosuke, OHASHI Toshiro, Knight Martin, Bader Dan, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2006-  (19)  386  -387  2007/01/06  [Not refereed][Not invited]
  • YAMAZAKI Yoshimasa, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2006-  (19)  388  -389  2007/01/06  [Not refereed][Not invited]
  • Effect of fluid shear stress on smooth muscle cell migration and permeability of endothelial monolayer in endothelial-smooth muscle cells cocultured model
    N. Sakamoto, T. Ohashi, M. Sato  Proceedings of the Final Symposium of the Tohoku University 21st Century Center of Excellence Program  163  -172  2007  [Not refereed][Not invited]
  • Mechanical properties of subcellular structural components isolated from cultured vascular cells
    S Deguchi, T Ohashi, M Sato  Proc. 9th International Symposium of Future Medical Engineering Based on Bio-nanotechnology  2007  [Not refereed][Not invited]
  • Stress-relaxation properties of stress fibers isolated from smooth muscle cells
    Matsui, T, Deguchi, S, Sakamoto, N, Ohashi, T, Sato, M  Proceedings of 9th International Symposium of Future Medical Engineering Based on Bio-nanotechnology  2007  [Not refereed][Not invited]
  • Modifying anastomotic site in thoracic aortic surgery by using biodegradable felt strips with or without basic fibroblast growth factor
    H. Fujiwara, Y. Saiki, K. Oda, S. Kawatsu, I. Yoshioka, N. Sakamoto, T. Ohashi, M. Sato, Y. Tabata, K. Tabayashi  The 8th Annual International Symposium on Advances in Understanding Aortic Diseases（ 2007.10.13－14．Tokyo）  2007  [Refereed][Not invited]
  • Influence of Pulsatile Flow on Integrin Expression and Morphology of Vascular Endothelial Cells
    Technology and Health Care  15-  91  -101  2007  [Not refereed][Not invited]
  • Effect of Mechanical Environment of Focal Adhesions on Remodeling of Endothelial Cells Subjected to Cyclic Stretch Using Microstructured Substrate
    Journal of Robotics and Mechatronics  2007  [Not refereed][Not invited]
  • Toshiro Ohashi, Masanori Masuda, Takeo Matsumoto, Masaaki Sato  CLINICAL HEMORHEOLOGY AND MICROCIRCULATION  37-  (1-2)  37  -46  2007  [Not refereed][Not invited]
     
    We have developed a novel uniaxial cyclic stretching technique to apply a ventral nonuniform strain to cells. In this system cells are grown on a glass-embedded silicone substrate instead of the commonly used uniform substrates. This unique substrate has been developed to give a strain gradient of 0.2%/mu m across each individual cell. Bovine aortic endothelial cells (BAECs) were cyclically stretched up to a maximum strain of 50% at 0.5 Hz for 30 min or 3 hours, focusing on the effect of the ventral strain gradient on local cell remodeling. After the experiments, BAECs were fixed and stained with rhodamine-phalloidin to observe actin filament structure. BAECs showed local development of stress fibers and localization of cell nuclei at regions exposed to higher strain. This result suggests that BAECs may sense ventral nonuniform strain and remodel cytoskeletal structure accordingly followed by the movement of cell nuclei.
  • Toshiro Ohashi, Masanori Masuda, Takeo Matsumoto, Masaaki Sato  CLINICAL HEMORHEOLOGY AND MICROCIRCULATION  37-  (1-2)  37  -46  2007  [Not refereed][Not invited]
     
    We have developed a novel uniaxial cyclic stretching technique to apply a ventral nonuniform strain to cells. In this system cells are grown on a glass-embedded silicone substrate instead of the commonly used uniform substrates. This unique substrate has been developed to give a strain gradient of 0.2%/mu m across each individual cell. Bovine aortic endothelial cells (BAECs) were cyclically stretched up to a maximum strain of 50% at 0.5 Hz for 30 min or 3 hours, focusing on the effect of the ventral strain gradient on local cell remodeling. After the experiments, BAECs were fixed and stained with rhodamine-phalloidin to observe actin filament structure. BAECs showed local development of stress fibers and localization of cell nuclei at regions exposed to higher strain. This result suggests that BAECs may sense ventral nonuniform strain and remodel cytoskeletal structure accordingly followed by the movement of cell nuclei.
  • Journal of Biomechanics  2007  [Not refereed][Not invited]
  • Estimation of Mechanical Role of Intracellular Cellular Structures in Smooth Muscle Cells by Using Traction Force Measurements
    Proceedings of The 11th International Conference on Miniaturized Systems for Chemistry and Life Sciences  2007  [Not refereed][Not invited]
  • Influence of Pulsatile Flow on Integrin Expression and Morphology of Vascular Endothelial Cells
    Technology and Health Care  15-  91  -101  2007  [Not refereed][Not invited]
  • Effects of Frequency of Pulsatile Flow on Morphology and Integrin Expression of Vascular Endothelial Cells
    Technology and Health Care  15-  91  -101  2007  [Not refereed][Not invited]
  • Adhesive Force of Human Hepatoma HepG2 Cells to Endothelial Cells and Expression of E-selectin
    Molecular & Cellular Biomechanics  2007  [Not refereed][Not invited]
  • Estimation of Mechanical Role of Intracellular Cellular Structures in Smooth Muscle Cells by Using Traction Force Measurements
    Proceedings of The 11th International Conference on Miniaturized Systems for Chemistry and Life Sciences  2007  [Not refereed][Not invited]
  • Effects of Frequency of Pulsatile Flow on Morphology and Integrin Expression of Vascular Endothelial Cells
    Technology and Health Care  15-  91  -101  2007  [Not refereed][Not invited]
  • Effect of Mechanical Environment of Focal Adhesions on Remodeling of Endothelial Cells Subjected to Cyclic Stretch Using Microstructured Substrate
    Journal of Robotics and Mechatronics  2007  [Not refereed][Not invited]
  • Toshiro Ohashi, Yoshiaki Sugaya, Naoya Sakamoto, Masaaki Sato  JOURNAL OF BIOMECHANICS  40-  (11)  2399  -2405  2007  [Not refereed][Not invited]
     
    Bovine aortic endothelial cells (BAECs) were exposed to hydrostatic pressures of 50, 100, and 150 mmHg and changes in morphology and expression of vascular enclothelial (VE)-cadherin were studied. After exposure to hydrostatic pressure, BAECs exhibited elongated and tortuous shape without predominant orientation, together with the development of centrally located, thick stress fibers. Pressured BAECs also exhibited a multilayered structure unlike those under control conditions and showed a significant increase in proliferation compared with control cells. Western blot analysis demonstrated that protein level of VE-cadherin were significantly lower under pressure conditions than under control conditions. Inhibition of VE-cadherin expression, using an antibody to VE-cadherin, induced the formation of numerous randomly distributed intercellular gaps, elongated and tortuous shapes, and multilayering. These responses were similar to those of pressured BAECs. The exposure of BAECs to hydrostatic pressure may therefore downregulate the expression of VE-cadherin, resulting in loss of contact inhibition followed by increased proliferation and formation of a multilayered structure. (c) 2007 Elsevier Ltd. All rights reserved.
  • Adhesive Force of Human Hepatoma HepG2 Cells to Endothelial Cells and Expression of E-selectin
    Molecular & Cellular Biomechanics  2007  [Not refereed][Not invited]
  • MATSUI Tsubasa, OHASHI Toshiro, ENDO Hideaki, SASAKI Tomohumi, ITO Hidemi, SATO Masaaki  Journal of JSEM  6-  (4)  423  -427  2006/12/25  [Not refereed][Not invited]
  • Sato Masaaki, Sugaya Yoshiaki, Sakamoto Naoya, Ohashi Toshiro  Biophysics  46-  (2)  S131  2006/10/01  [Not refereed][Not invited]
  • OHASHI Toshiro, SATO Yoshikazu, SASAKI Tomofumi, ITOH Hidemi, SATO Masaaki  Journal of JSEM  6-  (3)  269  -274  2006/09/25  [Not refereed][Not invited]
     
    This paper describes the feasibility of using finite element analysis to predict stress distributions in mandibular bone treated with a distal-extension partial denture, compared with conventional photoelastic method. A finite element model was constructed based on the CT scan data of the three-dimensional photoelastic model, which was anatomically realistic replica of a patient mandibular bone with teeth. A vertical loading was applied to a specially designed metal pyramid mounted on the first molar region to mimic functional movements of the partial denture. The numerical results showed that higher stress was observed in the apical and distal sides of the abutment teeth, which was the similar tendency to the experimental results obtained by the photoelastic method. Moreover, stress distributions depended on the variety of loadings applied to the different point of the pyramid. The present analysis may give a better prediction of stress distributions in patient mandibular bone than the photoelastic method.
  • MATSUI Tsubasa, OHASHI Toshiro, DEGUCHI Shinji, SATO Masaaki  The proceedings of the JSME annual meeting  2006-  (5)  227  -228  2006/09/15  [Not refereed][Not invited]
     
    In our previous study, tensile properties of single stress fibers (SF) chemically isolated from vascular smooth muscle cells have been investigated. Tensile tests were performed at a single strain rate of 0.02s^<-1>. In this study, the strain rate dependency of tensile properties of SF was investigated. The results showed that averaged force-strain relationships were found to be almost linear in the physio-logical strain range of 0.0-0.4 for 0.01s^<-1> and 0.05s^<-1>. In contrast, the force-strain relationship was nonlinear for the 0.1s^<-1> strain rate. The reason for this difference is un...
  • OHASHI Toshiro, YAMAZAKI Junichi, SATO Masaaki  The proceedings of the JSME annual meeting  2006-  (5)  269  -270  2006/09/15  [Not refereed][Not invited]
     
    Although cellular traction forces are believed to play an important role in the interactions between cells and their substrates, little is known of the effect of mechanical environment on the magnitude and direction of traction forces and hence how intracellular structures contribute to traction forces. In this study, microfabricated substrates, array of micropillars with different spacing, were used to estimate cellular traction forces of smooth muscle cells, particularly exploring the contribution of microtubules to traction forces. A significant increase in traction forces from 15.2±1.4 ...
  • OHASHI Toshiro, SEGAWA Kei, SAKAMOTO Naoya, SATO Masaaki  Transactions of the Japanese Society for Medical and Biological Engineering : BME  44-  (3)  454  -459  2006/09/10  [Not refereed][Not invited]
     
    To study endothelial cell remodeling in response to hydrostatic pressure, human umbilical vein endothelial cells (HUVECs) were exposed to hydrostatic pressure of 25 and 100 mmHg and changes in morphology and expression of VE-cadherin were studied. After exposure to hydrostatic pressure, HUVECs exhibited elongated and tortuous shape without predominant orientation, together with development of centrally located, thick stress fibers. Particularly, HUVEs exposed to 25 mmHg exhibited more tortuous shape compared to those both under control conditions and 100 mmHg. HUVECs did not exhibit a multi...
  • 大橋 俊朗  B & R  20-  (3)  110  -112  2006/09  [Not refereed][Not invited]
  • 伊藤 一志, 坂元 尚哉, 大橋 俊朗, 佐藤 正明  生体医工学 : 日本エム・イー学会誌  44-  (2)  319  -319  2006/06/10  [Not refereed][Not invited]
  • YAMAZAKI Yoshimasa, SASAKI Tomohumi, ITO Hidemi, OHASHI Toshiro, SATO Masaaki  Journal of JSEM  6-  (1)  19  -23  2006/03/25  [Not refereed][Not invited]
  • MATSUI Tsubasa, DEGUCHI Shinji, OHASHI Toshiro, SATO Masaaki  講演論文集  2006-  (41)  145  -146  2006/03/14  [Not refereed][Not invited]
  • YAMAZAKI Junichi, OHASHI Toshiro, SATO Masaaki  講演論文集  2006-  (41)  147  -148  2006/03/14  [Not refereed][Not invited]
  • IDO Hideto, KURODA Ryosuke, OHASHI Toshiro, SATO Masaaki  講演論文集  2006-  (41)  193  -194  2006/03/14  [Not refereed][Not invited]
  • USHIYAMA Takatomo, OHASHI Toshiro, SATO Masaaki  講演論文集  2006-  (41)  197  -198  2006/03/14  [Not refereed][Not invited]
  • Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato  Annals of Biomedical Engineering  34-  (3)  408  -415  2006/03  [Not refereed][Not invited]
     
    Migration of smooth muscle cells (SMCs) in hyperplasia is thought to have a correlation with blood flow conditions. In this study, the effect of shear stress applied to endothelial cells (ECs) on SMC migration was examined using a newly designed EC-SMC coculture model (CM), in which bovine SMCs and ECs were separated by a collagen layer and a membrane filter. After exposing the CM to shear stresses of 0.5, 1.0, or 1.5 Pa for 48 h, the number of SMCs migrating into the collagen layer was counted. Under static conditions, the migration of SMCs in the CM increased compared with SMCs cultured alone. Shear stress of 1.5 Pa significantly suppressed the SMC migration (p < 0.05) compared with the static CM. Media conditioned with the CM exposed to shear stress of 1.0 Pa (p < 0.05) and 1.5 Pa (p < 0.005) exhibited reduction in activated matrix metalloproteinase-2 (MMP-2) compared with the static CM, as analyzed by zymography. Addition of an inhibitor of nitric oxide (NO) synthase, N ω-nitro-L-arginine methyle ester, to the media inhibited the effect of 1.5 Pa shear stress on SMC migration but MMP-2 activity was unaffected. These results suggest that physiological shear stress has protective roles in atherosclerogenesis. © 2006 Biomedical Engineering Society.
  • Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato  Annals of Biomedical Engineering  34-  (3)  408  -415  2006/03  [Not refereed][Not invited]
     
    Migration of smooth muscle cells (SMCs) in hyperplasia is thought to have a correlation with blood flow conditions. In this study, the effect of shear stress applied to endothelial cells (ECs) on SMC migration was examined using a newly designed EC-SMC coculture model (CM), in which bovine SMCs and ECs were separated by a collagen layer and a membrane filter. After exposing the CM to shear stresses of 0.5, 1.0, or 1.5 Pa for 48 h, the number of SMCs migrating into the collagen layer was counted. Under static conditions, the migration of SMCs in the CM increased compared with SMCs cultured alone. Shear stress of 1.5 Pa significantly suppressed the SMC migration (p < 0.05) compared with the static CM. Media conditioned with the CM exposed to shear stress of 1.0 Pa (p < 0.05) and 1.5 Pa (p < 0.005) exhibited reduction in activated matrix metalloproteinase-2 (MMP-2) compared with the static CM, as analyzed by zymography. Addition of an inhibitor of nitric oxide (NO) synthase, N ω-nitro-L-arginine methyle ester, to the media inhibited the effect of 1.5 Pa shear stress on SMC migration but MMP-2 activity was unaffected. These results suggest that physiological shear stress has protective roles in atherosclerogenesis. © 2006 Biomedical Engineering Society.
  • H Fujiwara, K Oda, Y Saiki, N Sakamoto, T Ohashi, M Sato, Y Tabata, K Tabayashi  JOURNAL OF VASCULAR SURGERY  43-  (2)  349  -356  2006/02  [Not refereed][Not invited]
     
    Objectives: Wrapping methods have been widely used to reinforce the anastomotic site in vascular surgery; however, postoperative changes in the aortic wall wrapped by nonbiodegradable felt have not been well characterized. The purposes of this investigation are to elucidate the sequelae of wrapping with nonbiodegradable felt on the aortic wall and to modify those changes by using biodegradable felt with or without basic fibroblast growth factor (bFGF). Methods. The descending thoracic aortas of 15 beagles were wrapped with three different materials: nonbiodegradable polytetrafluoroethylene (PTFE) felt, biodegradable polyglycol acid (PGA) material, and PGA with 100 mu g bFGF (n = 5 in each group). The descending thoracic aorta was resected after 3 months. The thickness of the aortic wall, vessel density in the media and the adventitia, and the wall strength were assessed. Untreated native aortic wall served as a normal control. Results: The thickness of the media of the PTFE group was lower than that of the PGA + bFGF group (66% :+/- 5% vs 85% +/- 6% of control, P <.05). The adventitia-media ratio in the PTFE group decreased compared with controls (59.1% of normal, P < 0.05), whereas those in the PGA and PGA + bFGF groups increased (172.1% and 189.6% of normal, respectively, P <.01). The collagen-smooth muscle ratio in the media was higher in the PTFE group than in the controls (0.14 +/- 0.02 vs 0.07 +/- 0.01, P <.01). The number of vessels in the adventitia was higher in the PGA + bFGF group than those in PTFE or PGA groups (29.6 +/- 2.5/mm(2) vs 6.4 +/- 0.8/mm(2), 19.0 +/- 1.1/mm(2), p <.01). The PGA + bFGF group demonstrated larger failure force than the PTFE group (4.0 +/- 0.3 kgf vs 1.6 +/- 0.3 kgf, P <.01). The failure stress in the PGA and PGA + bFGF groups was larger than that in PTFE group (PTFE:PGA + bFGF = 5.3 +/- 0.9 X 10(2) kPa:11.7 +/- 1.7 x 10(2) kPa, P <.01; PTFE:PGA = 5.3 +/- 0.9 x 10(2) kPa:11.2 +/- 1.2 X 10(2) kPa, P <.05). Conclusion: The aortic wall wrapped with nonbiodegradable PTFE felt showed a reduced thickness and diminished vessels in the adventitia. Biodegradable felt (PGA), with or without bFGF, modified these histologic changes. The vessel-rich thickened adventitia, after wrapping by PGA with bFGF, was associated with increased aortic wall strength.(J Vasc Surg 2006;43:349-56.) Clinical Relevance: This investigation was conducted in an attempt to elucidate mechanisms underlying the occurrence of late postoperative false aneurysm after aortic surgery. We hypothesized that sustaining compression of the aorta by the felt strip may cause structural derangement and local ischemia on the aortic wall. We used a simple wrapping of the aorta with a felt strip rather than a felt strip at anastomotic sites to simplify the experimental model and to exclude confounding factors brought by technical inconsistency on the surgical anastomosis. We further attempted to find a clue for preventing adverse effects of wrapping with a conventional felt strip. Practically, we pursued a possible application of a biodegradable felt strip to aortic wrapping in our experimental model before we proceed in a clinical application of the new material.
  • Hidenori Fujiwara, Katsuhiko Oda, Yoshikatsu Saiki, Naoya Sakamoto, Toshiro Ohashi, Masaaki Sato, Yasuhiko Tabata, Koichi Tabayashi  Journal of Vascular Surgery  43-  (2)  349  -356  2006/02  [Not refereed][Not invited]
     
    Objectives: Wrapping methods have been widely used to reinforce the anastomotic site in vascular surgery however, postoperative changes in the aortic wall wrapped by nonbiodegradable felt have not been well characterized. The purposes of this investigation are to elucidate the sequelae of wrapping with nonbiodegradable felt on the aortic wall and to modify those changes by using biodegradable felt with or without basic fibroblast growth factor (bFGF). Methods: The descending thoracic aortas of 15 beagles were wrapped with three different materials: nonbiodegradable polytetrafluoroethylene (PTFE) felt, biodegradable polyglycol acid (PGA) material, and PGA with 100 μg bFGF (n = 5 in each group). The descending thoracic aorta was resected after 3 months. The thickness of the aortic wall, vessel density in the media and the adventitia, and the wall strength were assessed. Untreated native aortic wall served as a normal control. Results: The thickness of the media of the PTFE group was lower than that of the PGA + bFGF group (66% ± 5% vs 85% ± 6% of control, P < .05). The adventitia-media ratio in the PTFE group decreased compared with controls (59.1% of normal, P < 0.05), whereas those in the PGA and PGA + bFGF groups increased (172.1% and 189.6% of normal, respectively, P < .01). The collagen-smooth muscle ratio in the media was higher in the PTFE group than in the controls (0.14 ± 0.02 vs 0.07 ± 0.01, P < .01). The number of vessels in the adventitia was higher in the PGA + bFGF group than those in PTFE or PGA groups (29.6 ± 2.5/mm 2 vs 6.4 ± 0.8/mm 2, 19.0 ± 1.1/mm 2, P < .01). The PGA + bFGF group demonstrated larger failure force than the PTFE group (4.0 ± 0.3 kgf vs 1.6 ± 0.3 kgf, P < .01). The failure stress in the PGA and PGA + bFGF groups was larger than that in PTFE group (PTFE:PGA + bFGF = 5.3 ± 0.9 ×10 2 kPa:11.7 ± 1.7 × 10 2 kPa, P < .01 PTFE:PGA = 5.3 ± 0.9 × 10 2 kPa:11.2 ± 1.2 × 10 2 kPa, P < .05). Conclusion: The aortic wall wrapped with nonbiodegradable PTFE felt showed a reduced thickness and diminished vessels in the adventitia. Biodegradable felt (PGA), with or without bFGF, modified these histologic changes. The vessel-rich thickened adventitia, after wrapping by PGA with bFGF, was associated with increased aortic wall strength. Copyright © 2006 by The Society for Vascular Surgery.
  • MATSUI Tsubasa, DEGUCHI Shinji, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2005-  (18)  41  -42  2006/01/12  [Not refereed][Not invited]
  • OHASHI Toshiro, KAMEDA Norifumi, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2005-  (18)  43  -44  2006/01/12  [Not refereed][Not invited]
  • IDO Hideto, KURODA Ryosuke, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2005-  (18)  55  -56  2006/01/12  [Not refereed][Not invited]
  • OI Masaki, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2005-  (18)  211  -212  2006/01/12  [Not refereed][Not invited]
  • YAMAZAKI Junichi, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2005-  (18)  217  -218  2006/01/12  [Not refereed][Not invited]
  • USHIYAMA Takatomo, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2005-  (18)  283  -284  2006/01/12  [Not refereed][Not invited]
  • S Deguchi, K Maeda, T Ohashi, M Sato  JOURNAL OF BIOMECHANICS  39-  (7)  1362  -1362  2006  [Not refereed][Not invited]
  • Intracellular force balance in actin stress fiber network
    S Deguchi, T Ohashi, M Sato  Proc. 5th World Congress of Biomechanics  2006  [Not refereed][Not invited]
  • Biomechanical properties and roles of stress fibers in mechanotransduction
    M Sato, S Deguchi, T Matsui, T Ohashi  Proc. 5th World Congress of Biomechanics  2006  [Not refereed][Not invited]
  • Viscoelastic properties and locally heterogeneous deformation of stress fibers isolated from smooth muscle cells
    Matsui, T, Deguchi, S, Sakamoto, N, Ohashi, T, Sato, M  Proceedings of the 2nd Tohoku-NUS Joint Symposium on the Future Nano-medicine and Bioengineering in the East Asian Region  2006  [Not refereed][Not invited]
  • Shinji Deguchi, Toshiro Ohashi, Masaaki Sato  JOURNAL OF BIOMECHANICS  39-  (14)  2603  -2610  2006  [Not refereed][Not invited]
     
    Stress fibers (SFs), a contractile bundle of actin filaments, play a critical role in mechanotransduction in adherent cells; yet, the mechanical properties of SFs are poorly understood. Here, we measured tensile properties of single SFs by in vitro manipulation with cantilevers. SFs were isolated from cultured vascular smooth muscle cells with a combination of low ionic-strength extraction and detergent extraction and were stretched until breaking. The breaking force and the Young's modulus (assuming that SFs were isotropic) were, on average, 377 nN and 1.45 MPa, which were approximately 600-fold greater and three orders of magnitude lower, respectively, than those of actin filaments reported previously. Strain-induced stiffening was observed in the force-strain curve. We also found that the extracted SFs shortened to approximately 80% of the original length in an ATP-independent manner after they were dislodged from the substrate, suggesting that SFs had preexisting strain in the cytoplasm. The force required for stretching the single SFs from the zero-stress length back to the original length was approximately 10 nN, which was comparable with the traction force level applied by adherent cells at single adhesion sites to maintain cell integrity. These results suggest that SFs can bear intracellular stresses that may affect overall cell mechanical properties and will impact interpretation of intracellular stress distribution and force-transmission mechanism in adherent cells. (c) 2005 Elsevier Ltd. All rights reserved.
  • Evaluation of tension in actin bundle of endothelial cells based on preexisting strain and tensile properties measurements
    Molecular & Cellular Biomechanics  2006  [Not refereed][Not invited]
  • T. Ohashi, M. Hagiwara, D. L. Bader, M. M. Knight  BIORHEOLOGY  43-  (3-4)  201  -214  2006  [Not refereed][Not invited]
     
    The present study utilised pipette aspiration and simultaneous confocal microscopy to test the hypothesis that chondrocyte deformation is associated with distortion of intracellular organelles and activation of calcium signalling. Aspiration pressure was applied to isolated articular chondrocytes in increments of 2 cm of water every 60 seconds up to a maximum of 10 cm of water. At each pressure increment, confocal microscopy was used to visualise the mitochondria and nucleus labelled with JC-1 and Syto-16, respectively. To investigate intracellular calcium signalling, separate cells were labelled with Fluo 4, rapidly aspirated to 5 cm of water and then imaged for 5 minutes at a tare pressure of 0.1 cm of water. Partial cell aspiration was associated with distortion of the mitochondrial network, elongation of the nucleus and movement towards the pipette mouth. Treatment with cytochalasin D or nocodazole produced an increase in cell aspiration indicating that both the actin microfilaments and microtubules provide mechanical integrity to the cell. When the data was normalised to account for the increased cell deformation, both actin microfilaments and microtubules were shown to be necessary for strain transfer to the intracellular organelles. Mitochondria and nucleus deformation may both be involved in chondrocyte mechanotransduction as well as cellular and intracellular mechanics. In addition, pipette aspiration induced intracellular calcium signalling which may also form part of a mechanotransduction pathway. Alternatively calcium mobilisation may serve to modify actin polymerisation, thereby changing cell mechanics and membrane rigidity in order to facilitate localised cell deformation. These findings have important implications for our understanding of cell mechanics and mechanotransduction as well as interpretation and modelling of pipette aspiration data.
  • Shinji Deguchi, Toshiro Ohashi, Masaaki Sato  Journal of Biomechanics  39-  (14)  2603  -2610  2006  [Not refereed][Not invited]
     
    Stress fibers (SFs), a contractile bundle of actin filaments, play a critical role in mechanotransduction in adherent cells yet, the mechanical properties of SFs are poorly understood. Here, we measured tensile properties of single SFs by in vitro manipulation with cantilevers. SFs were isolated from cultured vascular smooth muscle cells with a combination of low ionic-strength extraction and detergent extraction and were stretched until breaking. The breaking force and the Young's modulus (assuming that SFs were isotropic) were, on average, 377 nN and 1.45 MPa, which were approximately 600-fold greater and three orders of magnitude lower, respectively, than those of actin filaments reported previously. Strain-induced stiffening was observed in the force-strain curve. We also found that the extracted SFs shortened to approximately 80% of the original length in an ATP-independent manner after they were dislodged from the substrate, suggesting that SFs had preexisting strain in the cytoplasm. The force required for stretching the single SFs from the zero-stress length back to the original length was approximately 10 nN, which was comparable with the traction force level applied by adherent cells at single adhesion sites to maintain cell integrity. These results suggest that SFs can bear intracellular stresses that may affect overall cell mechanical properties and will impact interpretation of intracellular stress distribution and force-transmission mechanism in adherent cells. © 2005 Elsevier Ltd. All rights reserved.
  • Intracellular stress transmission through actin stress fiber network in adherent vascular cells
    Molecular & Cellular Biomechanics  2006  [Not refereed][Not invited]
  • T. Ohashi, M. Hagiwara, D. L. Bader, M. M. Knight  BIORHEOLOGY  43-  (3-4)  201  -214  2006  [Not refereed][Not invited]
     
    The present study utilised pipette aspiration and simultaneous confocal microscopy to test the hypothesis that chondrocyte deformation is associated with distortion of intracellular organelles and activation of calcium signalling. Aspiration pressure was applied to isolated articular chondrocytes in increments of 2 cm of water every 60 seconds up to a maximum of 10 cm of water. At each pressure increment, confocal microscopy was used to visualise the mitochondria and nucleus labelled with JC-1 and Syto-16, respectively. To investigate intracellular calcium signalling, separate cells were labelled with Fluo 4, rapidly aspirated to 5 cm of water and then imaged for 5 minutes at a tare pressure of 0.1 cm of water. Partial cell aspiration was associated with distortion of the mitochondrial network, elongation of the nucleus and movement towards the pipette mouth. Treatment with cytochalasin D or nocodazole produced an increase in cell aspiration indicating that both the actin microfilaments and microtubules provide mechanical integrity to the cell. When the data was normalised to account for the increased cell deformation, both actin microfilaments and microtubules were shown to be necessary for strain transfer to the intracellular organelles. Mitochondria and nucleus deformation may both be involved in chondrocyte mechanotransduction as well as cellular and intracellular mechanics. In addition, pipette aspiration induced intracellular calcium signalling which may also form part of a mechanotransduction pathway. Alternatively calcium mobilisation may serve to modify actin polymerisation, thereby changing cell mechanics and membrane rigidity in order to facilitate localised cell deformation. These findings have important implications for our understanding of cell mechanics and mechanotransduction as well as interpretation and modelling of pipette aspiration data.
  • Transendothelial Migration of Leukocytes in Cocultured Vessel Model Exposed to Shear Stress
    Proceedings of the BMES Annual Fall Meeting  2006  [Not refereed][Not invited]
  • Annals of Biomedical Engineering  34-  (3)  408  -415  2006  [Not refereed][Not invited]
  • 細胞バイオメカニクスにおける近年の進歩
    日本バイオレオロジー学会誌  20-  (3)  12  -14  2006  [Not refereed][Not invited]
  • Evaluation of tension in actin bundle of endothelial cells based on preexisting strain and tensile properties measurements
    Molecular & Cellular Biomechanics  2006  [Not refereed][Not invited]
  • SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  Annals of Biomedical Engineering  34-  (3)  408  -415  2006  [Not refereed][Not invited]
  • SAKAMOTO Naoya, OI Masaki, OHASHI Toshiro, SATO Masaaki  JBSE  1-  (1)  224  -233  2006  [Not refereed][Not invited]
     
    This study investigated nitric oxide (NO) production and phenotype changes of smooth muscle cells (SMC) in a cocultured model (CM) exposed to fluid shear stress. The CM was composed of human umbilical endothelial cells (EC) and SMC, a collagen layer, and a porous membrane. After exposing the CM to shear stress of 1.5 Pa for 24 hours, α-smooth muscle actin (α-SMA) expression of SMC and NO production in culture media were examined. Under static conditions, α-SMA expression in the CM was significantly lower than that of a SMC monocultured model (SMC model). After exposure to shear stress, NO production in the CM increased compared to that in the static CM, and there was no significant difference in α-SMA expression between the CM and SMC model. These results suggest that EC may regulate phenotype changes of cocultured SMC, and NO may be one of the factors which induce dedifferentiation of SMC.
  • Shinji Deguchi, Toshiro Ohashi, Masaaki Sato  Journal of Biomechanics  39-  (14)  2603  -2610  2006  [Not refereed][Not invited]
     
    Stress fibers (SFs), a contractile bundle of actin filaments, play a critical role in mechanotransduction in adherent cells yet, the mechanical properties of SFs are poorly understood. Here, we measured tensile properties of single SFs by in vitro manipulation with cantilevers. SFs were isolated from cultured vascular smooth muscle cells with a combination of low ionic-strength extraction and detergent extraction and were stretched until breaking. The breaking force and the Young's modulus (assuming that SFs were isotropic) were, on average, 377 nN and 1.45 MPa, which were approximately 600-fold greater and three orders of magnitude lower, respectively, than those of actin filaments reported previously. Strain-induced stiffening was observed in the force-strain curve. We also found that the extracted SFs shortened to approximately 80% of the original length in an ATP-independent manner after they were dislodged from the substrate, suggesting that SFs had preexisting strain in the cytoplasm. The force required for stretching the single SFs from the zero-stress length back to the original length was approximately 10 nN, which was comparable with the traction force level applied by adherent cells at single adhesion sites to maintain cell integrity. These results suggest that SFs can bear intracellular stresses that may affect overall cell mechanical properties and will impact interpretation of intracellular stress distribution and force-transmission mechanism in adherent cells. © 2005 Elsevier Ltd. All rights reserved.
  • Intracellular stress transmission through actin stress fiber network in adherent vascular cells
    Molecular & Cellular Biomechanics  2006  [Not refereed][Not invited]
  • SEM像立体解析による歯根破折の破壊力学的検討
    日本実験力学会誌  6-  (4)  77  -81  2006  [Not refereed][Not invited]
  • SAKAMOTO Naoya, OI Masaki, OHASHI Toshiro, SATO Masaaki  Journal of Biomechanical Science and Engineering  1-  (1)  224  -233  2006  [Not refereed][Not invited]
     
    This study investigated nitric oxide (NO) production and phenotype changes of smooth muscle cells (SMC) in a cocultured model (CM) exposed to fluid shear stress. The CM was composed of human umbilical endothelial cells (EC) and SMC, a collagen layer, and a porous membrane. After exposing the CM to shear stress of 1.5 Pa for 24 hours, α-smooth muscle actin (α-SMA) expression of SMC and NO production in culture media were examined. Under static conditions, α-SMA expression in the CM was significantly lower than that of a SMC monocultured model (SMC model). After exposure to shear stress, NO production in the CM increased compared to that in the static CM, and there was no significant difference in α-SMA expression between the CM and SMC model. These results suggest that EC may regulate phenotype changes of cocultured SMC, and NO may be one of the factors which induce dedifferentiation of SMC.
  • Transendothelial Migration of Leukocytes in Cocultured Vessel Model Exposed to Shear Stress
    Proceedings of the BMES Annual Fall Meeting  2006  [Not refereed][Not invited]
  • 大橋 俊朗  人工臓器  34-  (3)  184  -186  2005/12/15  [Not refereed][Not invited]
  • S. Deguchi, T. Ohashi, M. Sato  MCB Molecular and Cellular Biomechanics  2-  (4)  205  -216  2005/12  [Not refereed][Not invited]
     
    Intracellular stress transmission through subcellular structural components has been proposed to affect activation of localized mechano-sensing sites such as focal adhesions in adherent cells. Previous studies reported that physiological extracellular forces produced heterogeneous spatial distributions of cytoplasmic strain. However, mechanical signaling pathway involved in intracellular force transmission through basal actin stress fibers (SFs), a mechano-responsive cytoskeletal structure, remains elusive. In the present study, we investigated force balance within the basal SFs of cultured smooth muscle cells and endothelial cells by (i) removing the cell membrane and cytoplasmic constituents except for materials physically attaching to the substrate (i.e., SF-focal adhesion complexities) or (ii) dislodging either mechanically or chemically the cell processes of the cells expressing fluorescent proteins-labeled actin and focal adhesions in order to examine stress-release-induced deformation of the basal SFs. The result showed that a removal of mechanical restrictions for SFs resulted in a decrease in the length of the remaining SFs, which means SFs bear tension. In addition, a release of the preexisting tension in a single SF was transmitted to another SF physically linked to the former, but not transmitted to the other ones physically independent of the former, suggesting that the prestress is balanced in tensed SF networks. These results support a hypothesis regarding cell structural architecture that physiological extracellular forces can produce in the basal SF network a directional intracellular stress or strain distribution. Therefore, consideration of the coexistence of the directional stretching strain along the axial direction of SFs and the heterogeneous strain in the other cytoplasmic region will be essential for understanding intracellular stress transmission in the adherent cells. Copyright © 2000 Tech Science Press.
  • S. Deguchi, T. Ohashi, M. Sato  MCB Molecular and Cellular Biomechanics  2-  (4)  205  -216  2005/12  [Not refereed][Not invited]
     
    Intracellular stress transmission through subcellular structural components has been proposed to affect activation of localized mechano-sensing sites such as focal adhesions in adherent cells. Previous studies reported that physiological extracellular forces produced heterogeneous spatial distributions of cytoplasmic strain. However, mechanical signaling pathway involved in intracellular force transmission through basal actin stress fibers (SFs), a mechano-responsive cytoskeletal structure, remains elusive. In the present study, we investigated force balance within the basal SFs of cultured smooth muscle cells and endothelial cells by (i) removing the cell membrane and cytoplasmic constituents except for materials physically attaching to the substrate (i.e., SF-focal adhesion complexities) or (ii) dislodging either mechanically or chemically the cell processes of the cells expressing fluorescent proteins-labeled actin and focal adhesions in order to examine stress-release-induced deformation of the basal SFs. The result showed that a removal of mechanical restrictions for SFs resulted in a decrease in the length of the remaining SFs, which means SFs bear tension. In addition, a release of the preexisting tension in a single SF was transmitted to another SF physically linked to the former, but not transmitted to the other ones physically independent of the former, suggesting that the prestress is balanced in tensed SF networks. These results support a hypothesis regarding cell structural architecture that physiological extracellular forces can produce in the basal SF network a directional intracellular stress or strain distribution. Therefore, consideration of the coexistence of the directional stretching strain along the axial direction of SFs and the heterogeneous strain in the other cytoplasmic region will be essential for understanding intracellular stress transmission in the adherent cells. Copyright © 2000 Tech Science Press.
  • 伊藤 一志, 坂元 尚哉, 大橋 俊朗  日本バイオレオロジ-学会誌  19-  (4)  164  -168  2005/12  [Not refereed][Not invited]
  • S Deguchi, T Ohashi, M Sato  JSME INTERNATIONAL JOURNAL SERIES C-MECHANICAL SYSTEMS MACHINE ELEMENTS AND MANUFACTURING  48-  (4)  396  -402  2005/12  [Not refereed][Not invited]
     
    A tensile test system for isolated cytoskeletal filaments, which enables to control strain rate, was newly designed. A pair of piezo-driven cantilevers were used to manipulate the specimen and to measure tensile load from the deflection of one of the cantilevers. The displacements of the cantilevers were optically and electrically detected. The specimen strain, determined from the cantilever displacements, was used as a feedback signal. We proposed a servo-system for strain rate control in which a desired path for the strain transition was designated. The path was chosen as a triangular-shape waveform against time, along which the strain rate is kept constant. We measured tensile properties of a single stress fiber isolated from a smooth muscle cell with this system to obtain a stretching stiffness of 45 nN per strain. Performance evaluation and the tensile test demonstrated that the system enabled to carry out strain rate-controlled tensile test.
  • UEKI Yosuke, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2005-  (16)  1  -2  2005/11/08  [Not refereed][Not invited]
  • Ito Kazushi, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2005-  (16)  3  -4  2005/11/08  [Not refereed][Not invited]
  • KURODA Ryosuke, IDO Hideto, OHASHI Toshiro, SATO Masaaki  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2005-  (16)  5  -6  2005/11/08  [Not refereed][Not invited]
  • T Ohashi, H Abe, T Matsumoto, M Sato  JOURNAL OF BIOMECHANICS  38-  (11)  2248  -2256  2005/11  [Not refereed][Not invited]
     
    A pipette aspiration technique was proposed for the measurement of nonlinear mechanical properties of arteries under biaxial stretching. A cross-shaped specimen of porcine thoracic aorta whose principal axes corresponded with the axial and circumferential directions of the aortic walls was excised. The intraluminal surface of the specimen was aspirated with a circular cross-sectioned glass pipette while the specimen was stretching in the axial and circumferential directions in 10% increments. The elastic modulus agreed with the incremental elastic modulus obtained through a conventional pressure-diameter test of the same specimen to within an error of 30% at a circumferential stretch ratio below 1.3 and an axial stretch ratio of 1.0, 1.1 or 1.2, which represent lower range of physiological stretch ratios for the porcine aorta. A rectangular cross-sectioned pipette was utilized to measure anisotropic properties of the specimen under biaxial stretching. When aspirated with such a pipette, the specimens' elastic properties along the length of the rectangular pipette cross section can be neglected. The elastic modulus was found to increase rapidly when the specimen was stretched in the direction of the pipette's width. Thus, pipette aspiration should have many advantages such as well measurement of the local nonlinear and anisotropic mechanical properties of blood vessel walls. (c) 2004 Elsevier Ltd. All rights reserved.
  • T Ohashi, H Abe, T Matsumoto, M Sato  JOURNAL OF BIOMECHANICS  38-  (11)  2248  -2256  2005/11  [Not refereed][Not invited]
     
    A pipette aspiration technique was proposed for the measurement of nonlinear mechanical properties of arteries under biaxial stretching. A cross-shaped specimen of porcine thoracic aorta whose principal axes corresponded with the axial and circumferential directions of the aortic walls was excised. The intraluminal surface of the specimen was aspirated with a circular cross-sectioned glass pipette while the specimen was stretching in the axial and circumferential directions in 10% increments. The elastic modulus agreed with the incremental elastic modulus obtained through a conventional pressure-diameter test of the same specimen to within an error of 30% at a circumferential stretch ratio below 1.3 and an axial stretch ratio of 1.0, 1.1 or 1.2, which represent lower range of physiological stretch ratios for the porcine aorta. A rectangular cross-sectioned pipette was utilized to measure anisotropic properties of the specimen under biaxial stretching. When aspirated with such a pipette, the specimens' elastic properties along the length of the rectangular pipette cross section can be neglected. The elastic modulus was found to increase rapidly when the specimen was stretched in the direction of the pipette's width. Thus, pipette aspiration should have many advantages such as well measurement of the local nonlinear and anisotropic mechanical properties of blood vessel walls. (c) 2004 Elsevier Ltd. All rights reserved.
  • OHASHI Toshiro, KUMAGAI Yoshiharu, MOCHIZUKI Naoki, SATO Masaaki  The proceedings of the JSME annual meeting  2005-  (6)  113  -114  2005/09/18  [Not refereed][Not invited]
     
    Temporal behavior of cytoskeletons such as actin filaments, and microtubules and FAT (Focal Adhesion Targeting) domain was observed in sheared endothelial cells using gene transfection technique to understand their mechanical roles in endothelial cell remodeling. Plasmid encoding enhanced GFP-actin, GFP-tubulin and RFP-FAT domain were transfected into bovine aortic endothelial cells using a liposomal method. After gene transfection, cells were exposed to shear stress of 2 Pa using a parallel plate flow chamber and temporally observed by fluorescence microscopy. During the process of cell el...
  • OHASHI Toshiro  日本機械学会東北支部秋季講演会講演論文集  2005-  (41)  29  -30  2005/09/09  [Not refereed][Not invited]
  • S. Deguchi, T. Ohashi, M. Sato  MCB Molecular and Cellular Biomechanics  2-  (3)  125  -133  2005/09  [Not refereed][Not invited]
     
    Actin bundles in vascular endothelial cells (ECs) play a critical role in transmitting intracellular forces between separate focal adhesion sites. However, quantitative descriptions of tension level in single actin bundles in a physiological condition are still poorly studied. Here, we evaluated magnitude of preexisting tension in a single actin bundle of ECs on the basis of measurements of its preexisting stretching strain and tensile properties. Cultured ECs expressing fluorescently-labeled actin were treated with detergents to extract acin bundles. One end of an actin bundle was then dislodged from the substrate by using a microneedle, resulting in a shortening of the actin bundle due to a release of preexisting tension. Assuming the shortened actin bundle reached its non-stress state, preexisting stretching strain was determined to be 0.24 on average. A tensile test of the dislodged single acin bundle was conducted with a pair of cantilevers to measure the force required for stretching it up to the original length, yielding an estimate of preexisting tension in the actin bundle. The magnitude of the preexisting tension, 4 nN on average, was comparable to previously reported data of the traction force generated by adherent cells at single adhesion sites to keep cell integrity. The Young's modulus of the isolated actin bundle was estimated to be ∼300 kPa from the tensile tests together with evaluation of average diameter of the isolated actin bundle based on transmission electron microscopy. These data will contribute to better understanding of intracellular stress transmission mechanism in ECs. Copyright © 2005 Tech Science Press.
  • S. Deguchi, T. Ohashi, M. Sato  MCB Molecular and Cellular Biomechanics  2-  (3)  125  -133  2005/09  [Not refereed][Not invited]
     
    Actin bundles in vascular endothelial cells (ECs) play a critical role in transmitting intracellular forces between separate focal adhesion sites. However, quantitative descriptions of tension level in single actin bundles in a physiological condition are still poorly studied. Here, we evaluated magnitude of preexisting tension in a single actin bundle of ECs on the basis of measurements of its preexisting stretching strain and tensile properties. Cultured ECs expressing fluorescently-labeled actin were treated with detergents to extract acin bundles. One end of an actin bundle was then dislodged from the substrate by using a microneedle, resulting in a shortening of the actin bundle due to a release of preexisting tension. Assuming the shortened actin bundle reached its non-stress state, preexisting stretching strain was determined to be 0.24 on average. A tensile test of the dislodged single acin bundle was conducted with a pair of cantilevers to measure the force required for stretching it up to the original length, yielding an estimate of preexisting tension in the actin bundle. The magnitude of the preexisting tension, 4 nN on average, was comparable to previously reported data of the traction force generated by adherent cells at single adhesion sites to keep cell integrity. The Young's modulus of the isolated actin bundle was estimated to be ∼300 kPa from the tensile tests together with evaluation of average diameter of the isolated actin bundle based on transmission electron microscopy. These data will contribute to better understanding of intracellular stress transmission mechanism in ECs. Copyright © 2005 Tech Science Press.
  • S Deguchi, K Maeda, T Ohashi, M Sato  JOURNAL OF BIOMECHANICS  38-  (9)  1751  -1759  2005/09  [Not refereed][Not invited]
     
    The mechanical contribution of nucleus in adherent cells to bearing intracellular stresses remains unclear. In this paper, the effects of fluid shear stress on morphology and elastic properties of endothelial nuclei were investigated. The morphological observation suggested that the nuclei in the cytoplasm were being vertically compressed under static conditions, whereas they were elongated and more compressed with a fluid shear stress of 2 Pa (20 dyn/cm(2)) onto the cell. The elongated nuclei remained the shape even after they were isolated from the cells. The micropipette aspiration technique on the isolated nuclei revealed that the elastic modulus of elongated nuclei, 0.62 +/- 0.15 kPa (n = 13, mean +/- SD), was significantly higher than that of control nuclei, 0.42 +/- 0.12 kPa (n = 11), suggesting that the nuclei remodeled their structure due to the shear stress. Based of these results and a transmission electron microscopy, a possibility of the nucleus as an intracellular compression-bearing organelle was proposed, which will impact interpretation of stress distribution in adherent cells. (C) 2005 Elsevier Ltd. All rights reserved.
  • S Deguchi, K Maeda, T Ohashi, M Sato  JOURNAL OF BIOMECHANICS  38-  (9)  1751  -1759  2005/09  [Not refereed][Not invited]
     
    The mechanical contribution of nucleus in adherent cells to bearing intracellular stresses remains unclear. In this paper, the effects of fluid shear stress on morphology and elastic properties of endothelial nuclei were investigated. The morphological observation suggested that the nuclei in the cytoplasm were being vertically compressed under static conditions, whereas they were elongated and more compressed with a fluid shear stress of 2 Pa (20 dyn/cm(2)) onto the cell. The elongated nuclei remained the shape even after they were isolated from the cells. The micropipette aspiration technique on the isolated nuclei revealed that the elastic modulus of elongated nuclei, 0.62 +/- 0.15 kPa (n = 13, mean +/- SD), was significantly higher than that of control nuclei, 0.42 +/- 0.12 kPa (n = 11), suggesting that the nuclei remodeled their structure due to the shear stress. Based of these results and a transmission electron microscopy, a possibility of the nucleus as an intracellular compression-bearing organelle was proposed, which will impact interpretation of stress distribution in adherent cells. (C) 2005 Elsevier Ltd. All rights reserved.
  • Toshiro Ohashi, Masaaki Sato  Fluid Dynamics Research  37-  (1-2)  40  -59  2005/07  [Not refereed][Not invited]
     
    Vascular endothelial cells are an interface between blood vessel walls and blood flow, and play important roles in physiological functions. Since endothelial cell responses to fluid shear stress have been implicated in the localization of atherosclerosis, the effect of shear stress on endothelial cell morphology and functions has been exclusively studied. After applying fluid shear stress, cultured endothelial cells show marked elongation and orientation in the flow direction. In addition, thick stress fibers of actin filaments appear and align along the cell long axis. Thus, the endothelial cell morphology is closely related to the cytoskeletal structure. The purpose of this review is to summarize endothelial cell responses to fluid flow which have been studied, focusing on the changes in cell shape and cytoskeletal structure. Numerical studies to simulate local flow field at the cellular level and the resulting intracellular stresses are also reviewed. © 2005 Published by The Japan Society of Fluid Mechanics and Elsevier B.V. All rights reserved.
  • T Ohashi, M Sato  FLUID DYNAMICS RESEARCH  37-  (1-2)  40  -59  2005/07  [Not refereed][Not invited]
     
    Vascular endothelial cells are an interface between blood vessel walls and blood flow, and play important roles in physiological functions. Since endothelial cell responses to fluid shear stress have been implicated in the localization of atherosclerosis, the effect of shear stress on endothelial cell morphology and functions has been exclusively studied. After applying fluid shear stress, cultured endothelial cells show marked elongation and orientation in the flow direction. In addition, thick stress fibers of actin filaments appear and align along the cell long axis. Thus, the endothelial cell morphology is closely related to the cytoskeletal structure. The purpose of this review is to summarize endothelial cell responses to fluid flow which have been studied, focusing on the changes in cell shape and cytoskeletal structure. Numerical studies to simulate local flow field at the cellular level and the resulting intracellular stresses are also reviewed. (C) 2005 Published by The Japan Society of Fluid Mechanics and Elsevier B.V. All rights reserved.
  • 伊藤 一志, 坂元 尚哉, 大橋 俊朗, 佐藤 正明  日本バイオレオロジー学会年会抄録集  28-  26  -26  2005/06/30  [Not refereed][Not invited]
  • 大井 正樹, 坂元 尚哉, 大橋 俊朗, 佐藤 正明  日本バイオレオロジー学会年会抄録集  28-  38  -38  2005/06/30  [Not refereed][Not invited]
  • 大橋 俊朗, 亀田 憲史, 佐藤 正明  日本バイオレオロジー学会年会抄録集  28-  44  -44  2005/06/30  [Not refereed][Not invited]
  • 佐藤 正明, 菅谷 嘉晃, 坂元 尚哉, 大橋 俊朗  日本バイオレオロジー学会年会抄録集  28-  52  -52  2005/06/30  [Not refereed][Not invited]
  • 大井 正樹, 坂元 尚哉, 大橋 俊朗, 佐藤 正明  生体医工学 : 日本エム・イー学会誌  43-  (2)  328  -329  2005/06/10  [Not refereed][Not invited]
  • 井戸 秀人, 黒田 亮典, 大橋 俊朗, 佐藤 正明  生体医工学 : 日本エム・イー学会誌  43-  (2)  329  -329  2005/06/10  [Not refereed][Not invited]
  • KUMAGAI Yoshiharu, OHASHI Toshiro, MOCHIZUKI Naoki, SATO Masaaki  The ... JSME Bioengineering Conference and Seminar  2004-  (17)  23  -24  2005/01/21  [Not refereed][Not invited]
  • Ito Kazushi, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  The ... JSME Bioengineering Conference and Seminar  2004-  (17)  259  -260  2005/01/21  [Not refereed][Not invited]
  • Mechanical properties of isolated stress fibers and nuclei to understand mechanical responses of endothelial cells
    M Sato, S Deguchi, T Ohashi  Proc. 6th Asian-Pacific Conference on Medical and Biological Engineering  2005  [Not refereed][Not invited]
  • Mechanical properties of stress fiber and nucleus to understand intracellular stress transmission
    S Deguchi, T Ohashi, M Sato  Proc. The Second Japan-Switzerland Workshop on Biomechanics  2005  [Not refereed][Not invited]
  • T Fukui, T Matsumoto, T Tanaka, T Ohashi, K Kumagai, H Akimoto, K Tabayashi, M Sato  BIO-MEDICAL MATERIALS AND ENGINEERING  15-  (4)  295  -305  2005  [Not refereed][Not invited]
     
    To investigate the mechanism of aneurysm rupture, it is necessary to examine the mechanical properties of aneurysm tissues in vivo. A new approach to evaluate in vivo mechanical properties of aortic aneurysmal tissues has been proposed in this study. The shape of the aneurysm was modeled as a sphere, and equi-biaxial stress in the in vivo state was estimated from the diameter and the wall thickness of each aneurysm and mean blood pressure of each patient. The mechanical properties of the aneurysm at the in vivo stress were estimated from its in vitro biaxial tensile properties. There were no significant correlations among maximum diameter D, wall thickness t, and mean infinitesimal strain in the in vivo state epsilon(m). This indicates the wall deformation during aneurysm development was not elastic but plastic. The mean incremental elastic modulus H-m, an index of tissue stiffness, had a significant positive correlation with elastic modulus anisotropy index K-H. This indicates the aneurysmal wall got more anisotropic in vivo as it becomes stiffer.
  • T Fukui, T Matsumoto, T Tanaka, T Ohashi, K Kumagai, H Akimoto, K Tabayashi, M Sato  BIO-MEDICAL MATERIALS AND ENGINEERING  15-  (4)  295  -305  2005  [Not refereed][Not invited]
     
    To investigate the mechanism of aneurysm rupture, it is necessary to examine the mechanical properties of aneurysm tissues in vivo. A new approach to evaluate in vivo mechanical properties of aortic aneurysmal tissues has been proposed in this study. The shape of the aneurysm was modeled as a sphere, and equi-biaxial stress in the in vivo state was estimated from the diameter and the wall thickness of each aneurysm and mean blood pressure of each patient. The mechanical properties of the aneurysm at the in vivo stress were estimated from its in vitro biaxial tensile properties. There were no significant correlations among maximum diameter D, wall thickness t, and mean infinitesimal strain in the in vivo state epsilon(m). This indicates the wall deformation during aneurysm development was not elastic but plastic. The mean incremental elastic modulus H-m, an index of tissue stiffness, had a significant positive correlation with elastic modulus anisotropy index K-H. This indicates the aneurysmal wall got more anisotropic in vivo as it becomes stiffer.
  • Multilayered Phenomena of Cultured Bovine Aortic Endothelial Cells by Exposure to Hydrostatic Pressure
    Proceedings of the International Federation for Medical and Biological Engineering  2005  [Not refereed][Not invited]
  • Subcellular Distribution of Traction Forces of Smooth Muscle Cells Using Micropatterned Substrates
    Proceedings of the 12th International Congress of Biorheology  2005  [Not refereed][Not invited]
  • Multilayered Phenomena of Cultured Bovine Aortic Endothelial Cells by Exposure to Hydrostatic Pressure
    Proceedings of the International Federation for Medical and Biological Engineering  2005  [Not refereed][Not invited]
  • Adhesive Property of Leukocytes to Endothelial Cells in Cocultured Model Exposed to Fluid Shear Stress
    Proceedings of the ASME 2005 Summer Bioengineering Conference  2005  [Not refereed][Not invited]
  • BIORHEOLOGY  42-  (1-2)  37  -38  2005  [Not refereed][Not invited]
  • Contributions of Cytoskeletal Structures to Traction Forces Generated by Smooth Muscle Cells Using Microfabricated Substrate
    Proceedings of the International Federation for Medical and Biological Engineering  2005  [Not refereed][Not invited]
  • Tensile Properties of Single Stress Fibers Characterized by In Vitro Manipulation
    Proceedings of the 12th International Congress of Biorheology  2005  [Not refereed][Not invited]
  • Subcellular Distribution of Traction Forces of Smooth Muscle Cells Using Micropatterned Substrates
    Proceedings of the 12th International Congress of Biorheology  2005  [Not refereed][Not invited]
  • M Sato, T Ohashi  BIORHEOLOGY  42-  (6)  421  -441  2005  [Not refereed][Not invited]
     
    Vascular endothelial cells are located at the innermost layer of the blood vessel wall and are always exposed to three different mechanical forces: shear stress due to blood flow, hydrostatic pressure due to blood pressure and cyclic stretch due to vessel deformation. It is well known that endothelial cells respond to these mechanical forces and change their shapes, cytoskeletal structures and functions. In this review, we would like to mainly focus on the effects of shear stress and hydrostatic pressure on endothelial cell morphology. After applying fluid shear stress, cultured endothelial cells show marked elongation and orientation in the flow direction. In addition, thick stress fibers of actin filaments appear and align along the cell long axis. Thus, endothelial cell morphology is closely related to the cytoskeletal structure. Further, the dynamic course of the morphological changes is shown and the related events such as changes in mechanical stiffness and functions are also summarized. When endothelial cells were exposed to hydrostatic pressure, they exhibited a marked elongation and orientation in a random direction, together with development of centrally located, thick stress fibers. Pressured endothelial cells also exhibited a multilayered structure with less expression of VE-cadherin unlike under control conditions. Simultaneous loading of hydrostatic pressure and shear stress inhibited endothelial cell multilayering and induced elongation and orientation of endothelial cells with well-developed VE-cadherin in a monolayer, which suggests that for a better understanding of vascular endothelial cell responses one has to take into consideration the combination of the different mechanical forces such as exist under in vivo mechanical conditions.
  • Journal of Biomechanics  39-  (14)  2603  -2610  2005  [Not refereed][Not invited]
  • Biorheological Views of Endothelial Cell Responses to Mechanical Stimuli
    Biorheology  42-  (6)  421  -441  2005  [Not refereed][Not invited]
  • Contributions of Cytoskeletal Structures to Traction Forces Generated by Smooth Muscle Cells Using Microfabricated Substrate
    Proceedings of the International Federation for Medical and Biological Engineering  2005  [Not refereed][Not invited]
  • Adhesive Property of Leukocytes to Endothelial Cells in Cocultured Model Exposed to Fluid Shear Stress
    Proceedings of the ASME 2005 Summer Bioengineering Conference  2005  [Not refereed][Not invited]
  • N Sakamoto, T Ohashi, M Sato  JSME INTERNATIONAL JOURNAL SERIES C-MECHANICAL SYSTEMS MACHINE ELEMENTS AND MANUFACTURING  47-  (4)  992  -999  2004/12  [Not refereed][Not invited]
     
    Effect of fluid shear stress on permeability of endothelial monolayer was investigated using an endothelial cell (EC)-smooth muscle cell (SMC) cocultured model (CM). Permeability of ECs to bovine serum albumin was measured after exposure to shear stress of 1.5 Pa for 48 hours. Morphology and VE-cadherin expression of ECs in CM was almost same as of ECs cultured alone (monocultured model, MM). Under static condition, EC permeability was 5.1 +/- 3.0 x 10(-6) cm/sec (mean +/- SD) in MM and 6.5 +/- 3.4 x 10(-6) cm/sec in CM. After exposure to shear stress, EC permeability in CM (2.2 +/- 1.9 x 10(-6) cm/sec, p < 0.05) significantly decreased compared with the static model. However, EC permeability in MM (3.9 +/- 3.2 x 10(-6) cm/sec) did not significantly change compared with static cultured condition. These results suggested that cellular interactions between ECs and SMCs have important influences on EC permeability.
  • IDO Hideto, OHASHI Toshiro, SATO Masaaki  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2004-  (15)  17  -18  2004/11/04  [Not refereed][Not invited]
  • OI Masaki, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2004-  (15)  21  -22  2004/11/04  [Not refereed][Not invited]
  • OHASHI Toshiro, SEO Shinji, MATSUMOTO Takeo, SATO Masaaki  Transactions of the Japan Society of Mechanical Engineers. A  70-  (697)  1262  -1268  2004/09/25  [Not refereed][Not invited]
     
    A finite element analysis using structural optimization method was performed to simulate the remodeling of bovine aortic endothelial cells (BAECs) under flow condition. BAECs showed marked elongation and aligned in the flow direction by exposing to a steady shear stress of 2 Pa for 24h. An atomic force microscope (AFM) system was used to obtain cell surface topography, showing that the cell height decreased significantly from 2.8±1.0μm to 1.4±0.5μm with exposure to fluid flow. The fluorescent images of cells stained by rhodamine-phalloidin showed that control cells exhibited dense periphera...
  • Deguchi Shinji, Ohashi Toshiro, Sato Masaaki  The proceedings of the JSME annual meeting  2004-  (5)  33  -34  2004/09/04  [Not refereed][Not invited]
     
    In this paper, mechanical architecture of adherent cells such as endothelial cells was discussed based on experimental evaluation of mechanical properties of subcellular structural components. Tensile properties of single stress fibers, bundles of actin filaments, in endothelial cells were obtained with in vitro micromanipulation. Preexisting tension in the stress fibers was then evaluated from the tensile properties and preexisting strain. Physiological tension level of the stress fibers was found to be 1-10 nN order of magnitude, which was "comparable to that of the traction force applied...
  • SUGAYA Yoshiaki, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  The proceedings of the JSME annual meeting  2004-  (5)  111  -112  2004/09/04  [Not refereed][Not invited]
     
    Effect of hydrostatic pressure on morphology and expression of VE-cadherin of cultured bovine endothelial cells (ECs) was investigated. After confluent ECs were exposed to hydrostatic pressure of 0, 50, 100 and 150 mmHg for 24 hours, F-actin filaments and VE-cadherin of ECs were stained. Statically cultured ECs formed a cobblestone pattern of contact-inhibited cells with thin, short F-actin filaments. VE-cadherin was uniformly distributed at the periphery of cells. In contrast, ECs exposed to hydrostatic pressure exhibited marked elongation and random orientation, together with development ...
  • H Ozawa, T Matsumoto, T Ohashi, M Sato, S Kokubun  JOURNAL OF NEUROSURGERY-SPINE  1-  (1)  122  -127  2004/07  [Not refereed][Not invited]
     
    Object. The pia mater has received little attention regarding its function in the deformation of the spinal cord under compression. In this study the mechanical properties and function of the spinal pia mater were investigated using three methods. Methods. Spinal cord segments were excised from rabbits. The elastic modulus of the pia mater was measured by performing a tensile test using specimens with the pia mater intact and ones with the pia mater stripped off. The stiffness of the spinal cord was examined by performing a compression test with specimens containing an intact pia mater and ones with a pia mater that was incised at both sides. The cross-sectional area and circumference of the spinal cord were measured on axial views of magnetic resonance images in patients with cervical disc herniations before and after surgery. The pia mater had an elastic modulus of 2300 kPa, which was 460 times higher than that of spinal cord parenchyma. By covering the parenchyma, it tripled the overall elastic modulus of the spinal cord. The pia mater increased the stiffness of the spinal cord and enhanced its shape recovery after removal of the compression. The cross-sectional area of the spinal cord increased after surgery, whereas the circumference of the spinal cord changed little. Conclusions. The pia mater firmly covers the spinal cord and has a high elastic modulus; it therefore provides a constraint on the spinal cord surface. It prevents elongation of the circumference and produces a large strain energy that is responsible for shape restoration following decompression.
  • H Ozawa, T Matsumoto, T Ohashi, M Sato, S Kokubun  JOURNAL OF NEUROSURGERY-SPINE  1-  (1)  122  -127  2004/07  [Not refereed][Not invited]
     
    Object. The pia mater has received little attention regarding its function in the deformation of the spinal cord under compression. In this study the mechanical properties and function of the spinal pia mater were investigated using three methods. Methods. Spinal cord segments were excised from rabbits. The elastic modulus of the pia mater was measured by performing a tensile test using specimens with the pia mater intact and ones with the pia mater stripped off. The stiffness of the spinal cord was examined by performing a compression test with specimens containing an intact pia mater and ones with a pia mater that was incised at both sides. The cross-sectional area and circumference of the spinal cord were measured on axial views of magnetic resonance images in patients with cervical disc herniations before and after surgery. The pia mater had an elastic modulus of 2300 kPa, which was 460 times higher than that of spinal cord parenchyma. By covering the parenchyma, it tripled the overall elastic modulus of the spinal cord. The pia mater increased the stiffness of the spinal cord and enhanced its shape recovery after removal of the compression. The cross-sectional area of the spinal cord increased after surgery, whereas the circumference of the spinal cord changed little. Conclusions. The pia mater firmly covers the spinal cord and has a high elastic modulus; it therefore provides a constraint on the spinal cord surface. It prevents elongation of the circumference and produces a large strain energy that is responsible for shape restoration following decompression.
  • 坂元 尚哉, 大橋 俊朗, 佐藤 正明  日本バイオレオロジー学会誌(B&R) = Journal of Japanese Society of Biorheology  18-  40  -40  2004/05/24  [Not refereed][Not invited]
  • Deguchi Shinji, Ohashi Toshiro, Sato Masaaki  Proceedings of the ... Asian Pacific Conference on Biomechanics : emerging science and technology in biomechanics  2004-  (1)  77  -78  2004/03/25  [Not refereed][Not invited]
  • Sakamoto Naoya, Ohashi Toshiro, Sato Masaaki  Proceedings of the ... Asian Pacific Conference on Biomechanics : emerging science and technology in biomechanics  2004-  (1)  83  -84  2004/03/25  [Not refereed][Not invited]
  • Deguchi Shinji, Ohashi Toshiro, Sato Masaaki  Proceedings of the ... Asian Pacific Conference on Biomechanics : emerging science and technology in biomechanics  2004-  (1)  221  -222  2004/03/25  [Not refereed][Not invited]
  • SUZUKI Kenichi, OHASHI Toshiro, SATO Masaaki  講演論文集  2004-  (39)  186  -187  2004/03/13  [Not refereed][Not invited]
  • IKEDA Takeshi, OHASHI Toshiro, SATO Masaaki  講演論文集  2004-  (39)  188  -189  2004/03/13  [Not refereed][Not invited]
  • DEGUCHI Shinji, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2004-  (16)  27  -28  2004/01/21  [Not refereed][Not invited]
  • Sakamoto Naoya, Ohashi Toshiro, Sato Masaaki  バイオエンジニアリング講演会講演論文集  2004-  (16)  31  -32  2004/01/21  [Not refereed][Not invited]
  • Measurement of preexisting strain of stress fibers in smooth muscle cells
    S Deguchi, T Ohashi, M Sato  Proc. The 43rd Annual Conference of Japanese Society for Medical and Biological Engineering  2004  [Not refereed][Not invited]
  • Morphological Responses of Aortic Endothelial Cells to Physiological Hydrostatic Pressure
    Proceedings of the14th European Society of Biomechanics Conference  2004  [Not refereed][Not invited]
  • Morphology and Expression of VE-cadherin of Bovine Endothelial Cells Exposed to Hydrostatic Pressure
    Proceedings of the BMES Annual Fall Meeting  2004  [Not refereed][Not invited]
  • Mechanical Properties of a Single Stress Fiber Isolated from Cultured Smooth Muscle Cells
    Proceedings of the BMES Annual Fall Meeting  2004  [Not refereed][Not invited]
  • Morphological Responses of Aortic Endothelial Cells to Physiological Hydrostatic Pressure
    Proceedings of the14th European Society of Biomechanics Conference  2004  [Not refereed][Not invited]
  • Mechanical Properties of a Single Stress Fiber Isolated from Cultured Smooth Muscle Cells
    Proceedings of the BMES Annual Fall Meeting  2004  [Not refereed][Not invited]
  • Morphology and Expression of VE-cadherin of Bovine Endothelial Cells Exposed to Hydrostatic Pressure
    Proceedings of the BMES Annual Fall Meeting  2004  [Not refereed][Not invited]
  • Y Sugaya, N Sakamoto, T Ohashi, M Sato  JSME INTERNATIONAL JOURNAL SERIES C-MECHANICAL SYSTEMS MACHINE ELEMENTS AND MANUFACTURING  46-  (4)  1248  -1255  2003/12  [Not refereed][Not invited]
     
    Morphological responses of cultured bovine endothelial cells (ECs) exposed to hydrostatic pressure were investigated. ECs were exposed to physiological blood pressure under a hydrostatic head of culture medium for 24 hours. Pressured ECs exhibited marked elongation and orientation with the random direction,, together with development of centrally located, thick stress fibers. Pressured ECs also exhibited multilayered structure unlike under control conditions. The area and the shape index value significantly decreased after exposure to hydrostatic pressure, which were in good agreement with the results from conventional flow-imposed experiments. In contrast, a tortuosity index, which was newly introduced to represent cell shape tortuosity, significantly increased for pressured ECs, while sheared ECs had no difference in turtuosity index from control. In addition, pressured ECs aligned with no predominant direction, while sheared ECs aligned in the flow direction. These results indicate that ECs can respond very specifically to the type of imposed mechanical stimuli such as hydrostatic pressure and fluid shear stress.
  • T Ohashi, S Sugita, T Matsumoto, K Kumagai, H Akimoto, K Tabayashi, M Sato  JSME INTERNATIONAL JOURNAL SERIES C-MECHANICAL SYSTEMS MACHINE ELEMENTS AND MANUFACTURING  46-  (4)  1290  -1296  2003/12  [Not refereed][Not invited]
     
    It is expected to be clinically useful to know the mechanical properties of human aortic aneurysms in assessing the potential for aneurysm rupture. For this purpose, a newly designed experimental setup was fabricated to measure the rupture properties of blood vessel walls. A square specimen of porcine thoracic aortas is inflated by air pressure at a rate of 10 mmHg/s (approximate to 1.3 MPa/s) until rupture occurs. Mean breaking stress was 1.8+/-0.4 MPa (mean+/-SD) for the specimens proximal to the heart and 2.3+/-0.8 MPa for the distal specimens, which are not significantly different to those values obtained longitudinally from conventional tensile tests. Moreover, the local breaking stretch ratio in the longitudinal direction was significantly higher at the ruptured site (2.7+/-0.5) than at the unruptured site (2.2+/-0.4). This testing system for studying the rupture properties of aortic walls is expected to be applicable to aortic aneurysms. Experimental verification of the present technique for the homogeneous, isotropic material is also presented.
  • SUGAYA Yoshiaki, SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2003-  (14)  3  -4  2003/09/17  [Not refereed][Not invited]
  • ARUGA Noriko, OHASHI Toshiro, SAKAMOTO Naoya, SATO Masaaki  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2003-  (14)  17  -18  2003/09/17  [Not refereed][Not invited]
  • DEGUCHI Shinji, OHASHI Toshiro, SATO Masaaki  Proceedings of the ... JSME Conference on Frontiers in Bioengineering  2003-  (14)  25  -26  2003/09/17  [Not refereed][Not invited]
  • 佐藤 正明, 山本 嗣雅, 出口 真次, 望月 直樹, 大橋 俊朗  年次大会講演資料集  0-  (3)  325  -326  2003/08/05  [Not refereed][Not invited]
  • DEGUCHI Shinji, MAEDA Kenjiro, OHASHI Toshiro, SATO Masaaki  The proceedings of the JSME annual meeting  2003-  (5)  29  -30  2003/08/05  [Not refereed][Not invited]
     
    The changes in morphology of isolated nuclei nuclei from endothelial cells were investigated before and after exposure to fluid flow. In addition, the Young's modulus of nuclei was determined with pipette aspiration technique. In fluorescence observation, the nuclei in the cells were initially found compressed vertically and the isolated nuclei remained almost rounded shape under static conditions. In contrast, the nuclei before and after isolation showed elongated shape under sheared conditions. These results showed that the compressed nuclei were elongated by fluid flow in the same fashio...
  • OHASHI Toshiro, YAMAMOTO Tsugumasa, MOCHIZUKI Naoki, SATO Masaaki  The proceedings of the JSME annual meeting  2003-  (5)  33  -34  2003/08/05  [Not refereed][Not invited]
     
    Dynamic behavior of actin filaments and focal adhesion targeting (FAT) was observed in endothelial cells during exposure to fluid flow. Cultured bovine aortic endothelial cells were co-transfected with GFP-actin and RFP-FAT using a liposomal method. Fluid shear stress of 2Pa was then applied to confluent endothlial cell monolayer using a parallel-plate flow chamber. During experiments, fluorescence images were observed every 10min up to 50min by a laser scanning microscope. Before exposure to shear stress, a number of small FAT clusters were observed and the distal tip of actin filaments wa...
  • SATO Yoshiyuki, SASAKI Tomofumi, ITOH Hidemi, OHASHI Toshiro, SATO Masaaki  The proceedings of the JSME annual meeting  2003-  (7)  7  -8  2003/08/05  [Not refereed][Not invited]
     
    Finite element analysis was performed on mandibular bone with distal extension removable partial denture (RPD) to assess the effects of RPD on stress distributions in mandibular bone. A finite element model was generated using X-ray CT data on the plaster model having the same morphology as the mandibular model, in which the mechanical properties of mandibular bone, natural teeth and RPD were assigned. In the analyses, a vertical loading was applied to the first molar to approximate occlusal loading of teeth. The analytical results showed the high stress concentrations in the distal and api...
  • OHASHI Toshiro, SATO Masaaki  Biophysics  43-  (3)  136  -141  2003/05/25  [Not refereed][Not invited]
     
    Morphology and local mechanical properties of sheared endothelial cells were measured by using atomic force microscope. After applying a steady shear stress of 2 Pa for 6 hours, bovine aortic endothelial cells showed marked elongation and aligned in the flow direction. The peak cell height decreased significantly with exposure to fluid flow. The fluorescent images showed that control cells exhibited dense peripheral bands of F-actin filaments, while sheared cells exhibited F-actin stress fibers which were thick and centrally located parallel to the flow direction. Elastic modulus estimated ...
  • YAMAMOTO Tsugumasa, OHASHI Toshiro, MOCHIZUKI Naoki, SATO Masaaki  講演論文集  2003-  (38)  68  -69  2003/03/15  [Not refereed][Not invited]
  • 出口 真次, 大橋 俊朗, 佐藤 正明  生体医工学 : 日本エム・イー学会誌  41-  (1)  69  -69  2003/03/10  [Not refereed][Not invited]
  • DEGUCHI Shinji, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2003-  (15)  11  -12  2003/01/20  [Not refereed][Not invited]
  • YAMAMOTO Tsugumasa, OHASHI Toshiro, MOCHIZUKI Naoki, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2003-  (15)  217  -218  2003/01/20  [Not refereed][Not invited]
  • SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2003-  (15)  225  -226  2003/01/20  [Not refereed][Not invited]
  • 大橋 俊朗, 佐藤 正明  BME : bio medical engineering  17-  (1)  124  -125  2003/01/10  [Not refereed][Not invited]
  • Remodeling of endothelial nucleus in response to fluid flow
    S Deguchi, K Maeda, T Ohashi, M Sato  Proc. The First US National Symposium on Frontiers in Biomechanics  2003  [Not refereed][Not invited]
  • Toshiro Ohashi, Naoya Sakamoto, Akiyo Iwao, Masaaki Sato  Technology and Health Care  11-  (4)  263  -274  2003  [Not refereed][Not invited]
     
    The effects of oxygen gas tensions and hydrostatic pressure on intracellular calcium, [Ca2+]i, response to a flow stimulus in endothelial cells were investigated. Cultured bovine aortic endothelial cells (BAECs) were exposed to a hydrostatic pressure of 100 mmHg under low oxygen gas tensions and were subsequently subjected to a 1 minute mechanical stimulation of fluid shear stress of 20 dynes/cm2. The [Ca2+]i response in BAECs was measured using a fluorescent indicator, Calcium Green-1. The maximum intensity for low oxygen tension was significantly lower than that for normal oxygen tension, which provides evidence that low oxygen tension regulates cellular functions downward. Moreover, preloading of hydrostatic pressure also reduced the increases in [Ca2+]i. These results suggest that BAECs in venous system, where oxygen tension and hydrostatic pressure are lower than those in arterial system, may be less sensitive to fluid flow. A separate observation showed that low oxygen tension did not significantly affect the cell morphology. In constrast, BAECs exposed to hydrostatic pressure showed marked elongation with no predominant orientation and the F-actin filament distributions were rearranged, indicating centrally located thick stress fibers. For better understanding of endothelial cell physiology, it is very important to elucidate the effect of oxygen gas tensions together with mechanical environment on endothelial cell responses.
  • Dynamic Behavior of Focal Adhesion-Associated Proteins of Cultured Endothelial Cells in Shear Flow Conditions
    Proceedings of the World Congress on Medical Physics and Biomedical Engineering  2003  [Not refereed][Not invited]
  • Endothelial Cells Exposed to Fluid Shear Stress Regulate Migration of Cocultured Smooth Muscle Cells
    Proceedings of the 2003 ASME Summer Bioengineering Conference  2003  [Not refereed][Not invited]
  • Single Stress Fibers Isolated from Vascular Smooth Muscle Cells Possess Surprisingly High Extensibility
    Proceedings of the 2003 ASME Summer Bioengineering Conference  2003  [Not refereed][Not invited]
  • Temporal Observation of Dynamic Process of Focal Adhesion-Associated Proteins in Sheared Endothelial Cells
    Proceedings of the 2003 ASME Summer Bioengineering Conference  2003  [Not refereed][Not invited]
  • Dynamic Behavior of Focal Adhesion-Associated Proteins of Cultured Endothelial Cells in Shear Flow Conditions
    Proceedings of the World Congress on Medical Physics and Biomedical Engineering  2003  [Not refereed][Not invited]
  • Mechanical Properties Differ between Single Stress Fibers and Actin Filaments
    Proceedings of the World Congress on Medical Physics and Biomedical Engineering  2003  [Not refereed][Not invited]
  • Mechanical Properties Differ between Single Stress Fibers and Actin Filaments
    Proceedings of the World Congress on Medical Physics and Biomedical Engineering  2003  [Not refereed][Not invited]
  • Oxygen Tension Modulates Ca2+ Response to Flow Stimulus in Endothelial Cells Exposed to Hydrostatic Pressure
    Technology and Health Care  11-  (4)  263  -274  2003  [Not refereed][Not invited]
  • Single Stress Fibers Isolated from Vascular Smooth Muscle Cells Possess Surprisingly High Extensibility
    Proceedings of the 2003 ASME Summer Bioengineering Conference  2003  [Not refereed][Not invited]
  • Temporal Observation of Dynamic Process of Focal Adhesion-Associated Proteins in Sheared Endothelial Cells
    Proceedings of the 2003 ASME Summer Bioengineering Conference  2003  [Not refereed][Not invited]
  • Evaluation of Rupture Properties of Human Thoracic Aortic Aneurysms Using a Pressure-Imposed Test
    Proceedings of the World Congress on Medical Physics and Biomedical Engineering  2003  [Not refereed][Not invited]
  • Evaluation of Rupture Properties of Human Thoracic Aortic Aneurysms Using a Pressure-Imposed Test
    Proceedings of the World Congress on Medical Physics and Biomedical Engineering  2003  [Not refereed][Not invited]
  • Endothelial Cells Exposed to Fluid Shear Stress Regulate Migration of Cocultured Smooth Muscle Cells
    Proceedings of the 2003 ASME Summer Bioengineering Conference  2003  [Not refereed][Not invited]
  • Takeo Matsumoto, Hironobu Abe, Toshiro Ohashi, Yoko Kato, Masaaki Sato  Physiological Measurement  23-  (4)  635  -648  2002/11  [Not refereed][Not invited]
     
    Changes in mechanical properties of arteries during atherogenesis remain controversial. One of the reasons could be that they have been evaluated with parameters measured in a whole vessel, although the lesions are localized. The local elastic modulus of atherosclerotic lesions was measured by the pipette aspiration method in thoracic aortas of rabbits fed a cholesterol diet for 8, 16, 24 and 28 weeks. The global elastic modulus of the whole aorta was measured by the pressure-diameter test. The local modulus decreased from that of the normal tissue in 8 weeks and then increased during the cholesterol feeding period. The global modulus did not change until 24 weeks and increased by 28 weeks. Histological observation revealed that the initial soft lesion was mainly composed of foam cells, and the stiffening accompanied first the appearance of smooth muscle cells in the top layer of the hyperplastic intima and then calcification in its bottom layer. The global elastic modulus did not change until marked calcification occurred in the tissue. These results suggest that change in mechanical properties of atherosclerotic lesion is not simple and has a close correlation with its histology. Assessment of local mechanical properties is important for studying mechanical properties of atherosclerotic arteries.
  • Takeo Matsumoto, Hironobu Abe, Toshiro Ohashi, Yoko Kato, Masaaki Sato  Physiological Measurement  23-  (4)  635  -648  2002/11  [Not refereed][Not invited]
     
    Changes in mechanical properties of arteries during atherogenesis remain controversial. One of the reasons could be that they have been evaluated with parameters measured in a whole vessel, although the lesions are localized. The local elastic modulus of atherosclerotic lesions was measured by the pipette aspiration method in thoracic aortas of rabbits fed a cholesterol diet for 8, 16, 24 and 28 weeks. The global elastic modulus of the whole aorta was measured by the pressure-diameter test. The local modulus decreased from that of the normal tissue in 8 weeks and then increased during the cholesterol feeding period. The global modulus did not change until 24 weeks and increased by 28 weeks. Histological observation revealed that the initial soft lesion was mainly composed of foam cells, and the stiffening accompanied first the appearance of smooth muscle cells in the top layer of the hyperplastic intima and then calcification in its bottom layer. The global elastic modulus did not change until marked calcification occurred in the tissue. These results suggest that change in mechanical properties of atherosclerotic lesion is not simple and has a close correlation with its histology. Assessment of local mechanical properties is important for studying mechanical properties of atherosclerotic arteries.
  • OHASHI Toshiro, SUGITA Syukei, MATSUMOTO Takeo, KUMAGAI Kiichiro, AKIMOTO Hiroji, TABAYASHI Koichi, SATO Masaaki  The proceedings of the JSME annual meeting  2002-  (1)  121  -122  2002/09/20  [Not refereed][Not invited]
     
    A newly designed experimental setup was fabricated to measure rupture properties of aortic aneurysms. Square specimen obtained from human thoracic aortic aneurysms was inflated by air pressure at a rate of 10 mmHg/s until rupture occurs. Deformation process was observed through two CCD cameras and a high-speed camera. Mean breaking stress and strain were estimated from the specimen geometry considering incompressibility, resulting in 0.97±0.39Mpa (mean±SD) and 0.64±0.10,respectively. Moreover, significant difference was found between mean breaking stress and area fraction of collagen fibers...
  • OHASHI Toshiro, IKEDA Takeshi, MATSUMOTO Takeo, SATO Masaaki  The proceedings of the JSME annual meeting  2002-  (6)  75  -76  2002/09/20  [Not refereed][Not invited]
     
    To determine the effect of extracellular matrix on chondrocytes, mechanical properties and production of type II collagen of chondrocytes were measured. Isolated bovine articular chondrocytes were cultured on a culture dish for 3 weeks and then embedded in a type I collagen gels. After 3-4 weeks, the chondrocytes were excised from the constructs and a pipette aspiration test was performed. The Young's modulus significantly increased when the chondrocytes were embedded in the collagen gels, which corresponded to the development in cytoskeletal structure. In addition, type II collagen was obs...
  • OHASHI Toshiro, SEO Shinji, ISHII Yasuaki, ISHIKAWA Yasushi, MATSUMOTO Takeo, SATO Masaaki  生体・生理工学シンポジウム論文集  17-  15  -18  2002/09/09  [Not refereed][Not invited]
  • SATO Yoshiyuki, OHASHI Toshiro, SASAKI Tomofumi, ITOH Hidemi, TAKAHASHI Shoki, MATSUMOTO Takeo, SATO Masaaki  The ... JSME Bioengineering Conference and Seminar  2002-  (13)  3  -4  2002/09/04  [Not refereed][Not invited]
  • IKEDA Takeshi, OHASHI Toshiro, SATO Masaaki  The ... JSME Bioengineering Conference and Seminar  2002-  (13)  13  -14  2002/09/04  [Not refereed][Not invited]
  • SAKAMOTO Naoya, OHASHI Toshiro, MATSUMOTO Takeo, SATO Masaaki  The ... JSME Bioengineering Conference and Seminar  2002-  (13)  93  -94  2002/09/04  [Not refereed][Not invited]
  • U Takeda, A Utani, JH Wu, E Adachi, H Koseki, M Taniguchi, T Matsumoto, T Ohashi, M Sato, H Shinkai  JOURNAL OF INVESTIGATIVE DERMATOLOGY  119-  (3)  678  -683  2002/09  [Not refereed][Not invited]
     
    Gene targeting of a member of small leucine-rich repeat proteoglycans demonstrates that collagen fibrillogenesis is mediated by a set of extracellular matrix components, which interact with collagen. Collagen-associated protein dermatopontin knockout mice were generated in order to analyze the biologic involvement of dermatopontin in the formation of collagen fibrils. Although dermatopontin-null mice did not exhibit any obvious anatomical abnormality, skin elasticity was increased. Skin tensile tests revealed that the initial elastic modulus was 57% lower in dermatopontin-null mice than in wild-type mice, and that maximum tensile strength was similar. Remarkably, light microscopy study showed a significant decrease in the relative thickness of the dermis in dermatopontin-null mice compared with wild-type mice (45.2 +/- 3.09% and 57.8 +/- 4.25%, respectively). The skin collagen content was 40% lower in dermatopontin-null than in wild-type mice. Collagen fibrils in dermatopontin-null mice showed a great variety in diameter and irregular contours under the electron microscope. These data indicate that dermatopontin plays a critical role in elasticity of skin and collagen accumulation attributed to collagen fibrillogenesis in vivo.
  • U Takeda, A Utani, JH Wu, E Adachi, H Koseki, M Taniguchi, T Matsumoto, T Ohashi, M Sato, H Shinkai  JOURNAL OF INVESTIGATIVE DERMATOLOGY  119-  (3)  678  -683  2002/09  [Not refereed][Not invited]
     
    Gene targeting of a member of small leucine-rich repeat proteoglycans demonstrates that collagen fibrillogenesis is mediated by a set of extracellular matrix components, which interact with collagen. Collagen-associated protein dermatopontin knockout mice were generated in order to analyze the biologic involvement of dermatopontin in the formation of collagen fibrils. Although dermatopontin-null mice did not exhibit any obvious anatomical abnormality, skin elasticity was increased. Skin tensile tests revealed that the initial elastic modulus was 57% lower in dermatopontin-null mice than in wild-type mice, and that maximum tensile strength was similar. Remarkably, light microscopy study showed a significant decrease in the relative thickness of the dermis in dermatopontin-null mice compared with wild-type mice (45.2 +/- 3.09% and 57.8 +/- 4.25%, respectively). The skin collagen content was 40% lower in dermatopontin-null than in wild-type mice. Collagen fibrils in dermatopontin-null mice showed a great variety in diameter and irregular contours under the electron microscope. These data indicate that dermatopontin plays a critical role in elasticity of skin and collagen accumulation attributed to collagen fibrillogenesis in vivo.
  • SATO Yoshiyuki, OHASHI Toshiro, ITOH Hidemi, SASAKI Tomofumi, SATO Masaaki, TAKAHASHI Shoki  Proceedings of JSEM  2-  78  -80  2002/08/05  [Not refereed][Not invited]
  • Ohashi Toshiro, Tsuyuki Kei, Sato Masaaki  日本バイオレオロジー学会誌  25-  (10)  24  -28  2002/05/13  [Not refereed][Not invited]
     
    Actin filament dynamics in endothelial cells was observed during exposure to fluid shear stress. A plasmid encoding enhanced green fluorescent protein (GFP) fused to the amino terminus of actin was transfected into cultured bovine aortic endothelial cells (BAECs) using a liposomal method. This functional GFP-actin construct enabled observation of actin filaments in BAECs. BAECs were exposed to fluid shear stress of 2 Pa using a parallel plate flow chamber and dynamic behavior of actin filaments in BAECs expressing stable GFP-actin was observed under a time lapse fluorescence microscopy. Cytoskeletal reorganization during exposure to shear stress preceded changes in cell shape. In a typical example, dense peripheral bands located upstream side in a cell moved toward the nucleus and fine network structure of actin filaments was partly destroyed within 60 min. After 180 min, actin filaments started to form thick bundles in the whole region of the cell, followed by cell elongation to the flow direction. Changes in cell shape and cytoskeletal structure seemed to be finished around 300 min. The GFP-actin expressed in endothelial cells should reveal detailed moving process of actin filaments in endothelial cells under mechanical stimuli.
  • SUGITA Syukei, OHASHI Toshiro, MATSUMOTO Takeo, KUMAGAI Kiichiro, AKIMOTO Hiroji, TABAYASHI Koichi, SATO Masaaki  講演論文集  2002-  (37)  66  -67  2002/03/16  [Not refereed][Not invited]
  • TSUYUKI Kei, OHASHI Toshiro, SATO Masaaki  講演論文集  2002-  (37)  182  -183  2002/03/16  [Not refereed][Not invited]
  • SUGITA Syukei, OHASHI Toshiro, MATSUMOTO Takeo, KUMAGAI Kiichiro, AKIMOTO Hiroji, TABAYASHI Koichi, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2002-  (14)  123  -124  2002/03/01  [Not refereed][Not invited]
  • TSUYUKI Kei, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2002-  (14)  129  -130  2002/03/01  [Not refereed][Not invited]
  • SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2002-  (14)  253  -254  2002/03/01  [Not refereed][Not invited]
  • T Ohashi, Y Ishii, Y Ishikawa, T Matsumoto, M Sato  BIO-MEDICAL MATERIALS AND ENGINEERING  12-  (3)  319  -327  2002  [Not refereed][Not invited]
     
    Local mechanical properties were measured for bovine endothelial cells exposed to shear stress using an atomic force microscopy (AFM), and the AFM indentations were simulated using a finite element method (FEM) to determine the elastic modulus. After exposure to shear stress, the endothelial cells showed marked elongation and orientation in the flow direction, together with significant decrease in the peak cell height. The applied force-indentation depth curve was obtained at seven different locations on the major axis of the cell surface and quantitatively expressed by the quadratic equation. The elastic modulus was determined by comparison of the experimental and numerical results. The modulus using our FEM model significantly became higher from 12.2+/-4.2 to 18.7+/-5.7 kPa with exposure to shear stress. Fluorescent images showed that stress fibers of F-actin bundles were mainly formed in the central portion of the sheared cells. The significant increase in the modulus may be due to this remodeling of cytoskeletal structure. The moduli using the Hertz model are 0.87+/-0.23 and 1.75+/-0.43 kPa for control and sheared endothelial cells respectively. This difference can be attributable to the differences in approximation functions to determine the elastic modulus. The elastic modulus would contribute a better understanding of local mechanical properties of the cells.
  • T Ohashi, Y Ishii, Y Ishikawa, T Matsumoto, M Sato  BIO-MEDICAL MATERIALS AND ENGINEERING  12-  (3)  319  -327  2002  [Not refereed][Not invited]
     
    Local mechanical properties were measured for bovine endothelial cells exposed to shear stress using an atomic force microscopy (AFM), and the AFM indentations were simulated using a finite element method (FEM) to determine the elastic modulus. After exposure to shear stress, the endothelial cells showed marked elongation and orientation in the flow direction, together with significant decrease in the peak cell height. The applied force-indentation depth curve was obtained at seven different locations on the major axis of the cell surface and quantitatively expressed by the quadratic equation. The elastic modulus was determined by comparison of the experimental and numerical results. The modulus using our FEM model significantly became higher from 12.2+/-4.2 to 18.7+/-5.7 kPa with exposure to shear stress. Fluorescent images showed that stress fibers of F-actin bundles were mainly formed in the central portion of the sheared cells. The significant increase in the modulus may be due to this remodeling of cytoskeletal structure. The moduli using the Hertz model are 0.87+/-0.23 and 1.75+/-0.43 kPa for control and sheared endothelial cells respectively. This difference can be attributable to the differences in approximation functions to determine the elastic modulus. The elastic modulus would contribute a better understanding of local mechanical properties of the cells.
  • Changes in Local Mechanical Properties and Actin Filament Distribution in Endothelial Cells Exposed to Shear Stress
    Proceedings of the 11th International Congress of Biorheology  2002  [Not refereed][Not invited]
  • Rupture Properties of Thoracic Aortic Aneurysms Using Pressure-Imposed Experiment
    Proceedings of the IV World Congress of Biomechanics  2002  [Not refereed][Not invited]
  • Toshiro Ohashi, Y. Ishii, Y. Ishikawa, T. Matsumoto, M. Sato  Bio-Medical Materials and Engineering  12-  (3)  319  -327  2002  [Not refereed][Not invited]
     
    Local mechanical properties were measured for bovine endothelial cells exposed to shear stress using an atomic force microscopy (AFM), and the AFM indentations were simulated using a finite element method (FEM) to determine the elastic modulus. After exposure to shear stress, the endothelial cells showed marked elongation and orientation in the flow direction, together with significant decrease in the peak cell height. The applied force-indentation depth curve was obtained at seven different locations on the major axis of the cell surface and quantitatively expressed by the quadratic equation. The elastic modulus was determined by comparison of the experimental and numerical results. The modulus using our FEM model significantly became higher from 12.2 ± 4.2 to 18.7 ± 5.7 kPa with exposure to shear stress. Fluorescent images showed that stress fibers of F-actin bundles were mainly formed in the central portion of the sheared cells. The significant increase in the modulus may be due to this remodeling of cytoskeletal structure. The moduli using the Hertz model are 0.87 ± 0.23 and 1.75 ± 0.43 kPa for control and sheared endothelial cells respectively. This difference can be attributable to the differences in approximation functions to determine the elastic modulus. The elastic modulus would contribute a better understanding of local mechanical properties of the cells.
  • Effect of Magnetic Field on Nitric Oxide Synthesis of Cultured Endothelial Cells
    International Journal Applied Electromagnetics and Mechanics  14-  317  -322  2002  [Not refereed][Not invited]
  • Toshiro Ohashi, Y. Ishii, Y. Ishikawa, T. Matsumoto, M. Sato  Bio-Medical Materials and Engineering  12-  (3)  319  -327  2002  [Not refereed][Not invited]
     
    Local mechanical properties were measured for bovine endothelial cells exposed to shear stress using an atomic force microscopy (AFM), and the AFM indentations were simulated using a finite element method (FEM) to determine the elastic modulus. After exposure to shear stress, the endothelial cells showed marked elongation and orientation in the flow direction, together with significant decrease in the peak cell height. The applied force-indentation depth curve was obtained at seven different locations on the major axis of the cell surface and quantitatively expressed by the quadratic equation. The elastic modulus was determined by comparison of the experimental and numerical results. The modulus using our FEM model significantly became higher from 12.2 ± 4.2 to 18.7 ± 5.7 kPa with exposure to shear stress. Fluorescent images showed that stress fibers of F-actin bundles were mainly formed in the central portion of the sheared cells. The significant increase in the modulus may be due to this remodeling of cytoskeletal structure. The moduli using the Hertz model are 0.87 ± 0.23 and 1.75 ± 0.43 kPa for control and sheared endothelial cells respectively. This difference can be attributable to the differences in approximation functions to determine the elastic modulus. The elastic modulus would contribute a better understanding of local mechanical properties of the cells.
  • Measurement of Rupture Properties of Thoracic Aortic Aneurysms Using Pressure-Imposed Test
    Proceedings of the International Congress on Biological and Medical Engineering  2002  [Not refereed][Not invited]
  • Biaxial Tensile Properties of Aortic Aneurysm Tissues under Equibiaxial Stress
    Proceedings of the IV World Congress of Biomechanics  2002  [Not refereed][Not invited]
  • Effect of Magnetic Field on Nitric Oxide Synthesis of Cultured Endothelial Cells
    International Journal Applied Electromagnetics and Mechanics  14-  317  -322  2002  [Not refereed][Not invited]
  • Structural Optimization Analysis of Endothelial Cell Remodeling to Fluid Flow
    Proceedings of the 2nd Joint EMBS & BMES conference  2002  [Not refereed][Not invited]
  • Biaxial Tensile Properties of Aortic Aneurysm Tissues under Equibiaxial Stress
    Proceedings of the IV World Congress of Biomechanics  2002  [Not refereed][Not invited]
  • Dynamic Behavior of Focal Adhesion - Associated Proteins of Endothelial cells in The Early Stage of Shear Stress Exposure
    Proceedings of the International Congress on Biological and Medical Engineering  2002  [Not refereed][Not invited]
  • Dynamic Behavior of Focal Adhesion - Associated Proteins of Endothelial cells in The Early Stage of Shear Stress Exposure
    Proceedings of the International Congress on Biological and Medical Engineering  2002  [Not refereed][Not invited]
  • Observation of Actin Filament Movement and Remodeling Process of Endothelial Cells During Exposure to Shear Stress
    Proceedings of the International Congress on Biological and Medical Engineering  2002  [Not refereed][Not invited]
  • Measurement of Rupture Properties of Thoracic Aortic Aneurysms Using Pressure-Imposed Test
    Proceedings of the International Congress on Biological and Medical Engineering  2002  [Not refereed][Not invited]
  • Dynamic Behavior of Actin Filaments in Cultured Endothelial Cells During Exposure to Shear Stress
    Proceedings of the IV World Congress of Biomechanics  2002  [Not refereed][Not invited]
  • Observation of Actin Filament Movement and Remodeling Process of Endothelial Cells During Exposure to Shear Stress
    Proceedings of the International Congress on Biological and Medical Engineering  2002  [Not refereed][Not invited]
  • Structural Optimization Analysis of Endothelial Cell Remodeling to Fluid Flow
    Proceedings of the 2nd Joint EMBS & BMES conference  2002  [Not refereed][Not invited]
  • Rupture Properties of Thoracic Aortic Aneurysms Using Pressure-Imposed Experiment
    Proceedings of the IV World Congress of Biomechanics  2002  [Not refereed][Not invited]
  • Fluid Shear Stress Affects Migration of Vascular Smooth Muscle Cells Cocultured with Endothelial Cells
    Proceedings of the International Congress on Biological and Medical Engineering  2002  [Not refereed][Not invited]
  • DL Bader, T Ohashi, MM Knight, DA Lee, M Sato  BIORHEOLOGY  39-  (1-2)  69  -78  2002  [Not refereed][Not invited]
     
    This paper presents a series of techniques, which examine the deformation characteristics of bovine articular chondrocytes. The direct contact approach employs well established methodology, involving AFM and micropipette aspiration, to yield structural properties of local regions of isolated chondrocytes. The former technique yields a non-linear response with increased structural stiffness in a central location on a projected image of the chondrocyte. A simple viscoelastic model can be used with data from the micropipette aspiration technique to yield a mean value of Young's modulus, which is similar to that recently reported (Jones et al., 1999). An indirect approach is also described, involving the response of chondrocytes seeded within compressed agarose constructs. For 1% agarose constructs, the resulting cell strain, yields a gross cell modulus of 2.7 kPa. The study highlights the difficulties in establishing unique mechanical parameters, which reflect the deformation behaviour of articular chondrocytes.
  • Fluid Shear Stress Affects Migration of Vascular Smooth Muscle Cells Cocultured with Endothelial Cells
    Proceedings of the International Congress on Biological and Medical Engineering  2002  [Not refereed][Not invited]
  • DL Bader, T Ohashi, MM Knight, DA Lee, M Sato  BIORHEOLOGY  39-  (1-2)  69  -78  2002  [Not refereed][Not invited]
     
    This paper presents a series of techniques, which examine the deformation characteristics of bovine articular chondrocytes. The direct contact approach employs well established methodology, involving AFM and micropipette aspiration, to yield structural properties of local regions of isolated chondrocytes. The former technique yields a non-linear response with increased structural stiffness in a central location on a projected image of the chondrocyte. A simple viscoelastic model can be used with data from the micropipette aspiration technique to yield a mean value of Young's modulus, which is similar to that recently reported (Jones et al., 1999). An indirect approach is also described, involving the response of chondrocytes seeded within compressed agarose constructs. For 1% agarose constructs, the resulting cell strain, yields a gross cell modulus of 2.7 kPa. The study highlights the difficulties in establishing unique mechanical parameters, which reflect the deformation behaviour of articular chondrocytes.
  • Changes in Local Mechanical Properties and Actin Filament Distribution in Endothelial Cells Exposed to Shear Stress
    Proceedings of the 11th International Congress of Biorheology  2002  [Not refereed][Not invited]
  • Dynamic Behavior of Actin Filaments in Cultured Endothelial Cells During Exposure to Shear Stress
    Proceedings of the IV World Congress of Biomechanics  2002  [Not refereed][Not invited]
  • Y Saijo, T Ohashi, H Sasaki, M Sato, CS Jorgensen, SI Nitta  ANNALS OF BIOMEDICAL ENGINEERING  29-  (12)  1048  -1053  2001/12  [Not refereed][Not invited]
     
    Scanning acoustic microscopy (SAM) was equipped to assess the acoustic properties of normal and atherosclerotic coronary arteries. The SAM image in the atherosclerotic lesion clearly demonstrated that the sound speed was higher than that in the normal intima, and that the variation of elasticity was found within the fibrous cap of the plaque. Young's elastic modulus of each region was calculated and the finite element analysis was applied to derive the stress distribution in these arterial walls. In a case of normal coronary artery, the stress was dominant in the intima and the distribution was rather homogeneous and in a case of atherosclerosis, high stress was concentrated to the relatively soft lesion in the fibrous cap overlying lipid pool. SAM provides information on the physical properties, which cannot be obtained by the optical microscope. The results would help in understanding the pathological features of atherosclerosis. (C) 2001 Biomedical Engineering Society.
  • Y Saijo, T Ohashi, H Sasaki, M Sato, CS Jorgensen, SI Nitta  ANNALS OF BIOMEDICAL ENGINEERING  29-  (12)  1048  -1053  2001/12  [Not refereed][Not invited]
     
    Scanning acoustic microscopy (SAM) was equipped to assess the acoustic properties of normal and atherosclerotic coronary arteries. The SAM image in the atherosclerotic lesion clearly demonstrated that the sound speed was higher than that in the normal intima, and that the variation of elasticity was found within the fibrous cap of the plaque. Young's elastic modulus of each region was calculated and the finite element analysis was applied to derive the stress distribution in these arterial walls. In a case of normal coronary artery, the stress was dominant in the intima and the distribution was rather homogeneous and in a case of atherosclerosis, high stress was concentrated to the relatively soft lesion in the fibrous cap overlying lipid pool. SAM provides information on the physical properties, which cannot be obtained by the optical microscope. The results would help in understanding the pathological features of atherosclerosis. (C) 2001 Biomedical Engineering Society.
  • TSUYUKI Kei, OHASHI Toshiro, SATO Masaaki  The ... JSME Bioengineering Conference and Seminar  2001-  (12)  85  -86  2001/10/26  [Not refereed][Not invited]
  • MASUDA Masanori, OHASHI Toshiro, MATSUMOTO Takeo, SATO Masaaki  The ... JSME Bioengineering Conference and Seminar  2001-  (12)  131  -132  2001/10/26  [Not refereed][Not invited]
  • YAMAMOTO Tsugumasa, OHASHI Toshiro, SATO Masaaki  The ... JSME Bioengineering Conference and Seminar  2001-  (12)  133  -134  2001/10/26  [Not refereed][Not invited]
  • Fukui Tomohiro, Matsumoto Takeo, Kumagai Kiichiro, Akimoto Hiroji, Tabayashi Koichi, Ohashi Toshiro, Sato Masaaki  The ... JSME Bioengineering Conference and Seminar  2001-  (12)  195  -196  2001/10/26  [Not refereed][Not invited]
  • H Ozawa, T Matsumoto, T Ohashi, M Sato, S Kokubun  JOURNAL OF NEUROSURGERY  95-  (2)  221  -224  2001/10  [Not refereed][Not invited]
     
    Object. Although the gray matter of the spinal cord has been thought to be softer than the white matter, there is no evidence to support this belief. Because the spinal cord is extremely soft, it has been difficult to measure the mechanical properties of the gray and white matter. The modulis of elasticity of the gray and white matter were measured in situ by using a pipette aspiration method. Method. The spinal cord specimens were excised from Japanese white rabbits. Specimens were cut to display the surfaces of axial, frontal, and sagittal sections. The surfaces of the gray and white matter were aspirated using a 0.8-mm-inner-diameter glass pipette while monitoring with a video microscope, and the deformed length in the pipette was measured on a monitor. In each case the modulus of elasticity was calculated by comparing the relationship between the aspiration pressure and aspirated volume of the specimen with that determined by finite element analysis. The moduli of elasticity of the gray and white matter were 3.4 +/- 1.4 kPa (mean +/- standard deviation) and 3.4 +/- 0.9 kPa in the axial section, 3 +/- 0.3 kPa and 3.5 +/- 0.5 kPa in the frontal section, and 3.5 +/- 0.9 kPa and 2.8 +/- 0.4 kPa in the sagittal section, respectively. Conclusions. No significant difference in modulus of elasticity was shown between the gray and white matter of the spinal in sections made in various directions.
  • H Ozawa, T Matsumoto, T Ohashi, M Sato, S Kokubun  JOURNAL OF NEUROSURGERY  95-  (2)  221  -224  2001/10  [Not refereed][Not invited]
     
    Object. Although the gray matter of the spinal cord has been thought to be softer than the white matter, there is no evidence to support this belief. Because the spinal cord is extremely soft, it has been difficult to measure the mechanical properties of the gray and white matter. The modulis of elasticity of the gray and white matter were measured in situ by using a pipette aspiration method. Method. The spinal cord specimens were excised from Japanese white rabbits. Specimens were cut to display the surfaces of axial, frontal, and sagittal sections. The surfaces of the gray and white matter were aspirated using a 0.8-mm-inner-diameter glass pipette while monitoring with a video microscope, and the deformed length in the pipette was measured on a monitor. In each case the modulus of elasticity was calculated by comparing the relationship between the aspiration pressure and aspirated volume of the specimen with that determined by finite element analysis. The moduli of elasticity of the gray and white matter were 3.4 +/- 1.4 kPa (mean +/- standard deviation) and 3.4 +/- 0.9 kPa in the axial section, 3 +/- 0.3 kPa and 3.5 +/- 0.5 kPa in the frontal section, and 3.5 +/- 0.9 kPa and 2.8 +/- 0.4 kPa in the sagittal section, respectively. Conclusions. No significant difference in modulus of elasticity was shown between the gray and white matter of the spinal in sections made in various directions.
  • NAGANO Yujiro, MATSUMOTO Takeo, KAWAWA Takuo, OHASHI Toshiro, SATO Masaaki  講演論文集  2001-  15  -16  2001/09/21  [Not refereed][Not invited]
  • OHASHI Toshiro, SEO Shinji, MATSUMOTO Takeo, SATO Masaaki  The proceedings of the JSME annual meeting  0-  (1)  43  -44  2001/08/22  [Not refereed][Not invited]
     
    A finite element analysis using structural optimization method was performed to simulate the remodeling of bovine aortic endothelial cells (BAECs), in which cell surface geometries were measured by an atomic force microscope (AFM) system. After applying a steady shear stress of 2 Pa for 24 hours, BAECs showed marked elongation and aligned in the flow direction. In the AFM measurements, the peak cell height decreased significantly from 2.8 ± 1.0 μm to 1.4 ± 0.5 μm with exposure to fluid flow. The fluorescent images showed that control cells exhibited dense peripheral bands of F-actin filamen...
  • SAKAMOTO Naoya, OHASHI Toshiro, IWAO Akiyo, SATO Masaaki  The proceedings of the JSME annual meeting  0-  (1)  99  -100  2001/08/22  [Not refereed][Not invited]
     
    Intracellular calcium response to a flow stimulus and morphological changes in sheared endothelial cells were investigated under arterial and venous gas conditions. Prior to the flow stimulus experiments, the endothelial cells were exposed to shear stress of 2 Pa for 6 hours using a parallel-plate flow chamber. Maximum fluorescent intensity of intracellular calcium for venous gas condition was significantly lower than that for arterial gas condition, which provides evidence that low oxygen tensions downregulate cellular physiological functions. However, preloading of shear stress prevented ...
  • MATSUMOTO Takeo, NAGANO Yujiro, KAWAWA Takuo, OHASHI Toshiro, SATO Masaaki  Proceedings of the ... annual meeting of JSME/MMD  2001-  35  -36  2001/07/19  [Not refereed][Not invited]
  • 佐藤 正明, 露木 啓, 大橋 俊朗  日本バイオレオロジー学会年会抄録集  24-  37  -37  2001/05/28  [Not refereed][Not invited]
  • Structural Optimization Method of Sheared Endothelial Cells Using Coupled Fluid-Structural Analysis
    大橋俊朗, 清尾慎司, 松本健郎, 佐藤正明  日本数値流体力学会雑誌  9-  (4)  99  -105  2001/04  [Not refereed][Not invited]
  • OHASHI Toshiro, MATSUMOTO Takeo, SATO Masaaki  Conference on Information, Intelligence and Precision Equipment : IIP  2001-  72  -73  2001/03/23  [Not refereed][Not invited]
  • SEO Shinji, OHASHI Toshiro, MATSUMOTO Takeo, SATO Masaaki  講演論文集  2001-  (36)  52  -53  2001/03/10  [Not refereed][Not invited]
  • KATO Y, MATSUMOTO T, OHASHI T, SATO M  脈管学  41-  (1)  23  -29  2001/01/25  [Not refereed][Not invited]
  • OHASHI Toshiro, FUJITA Manabu, MASUDA Masanori, MATSUMOTO Takeo, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2001-  (13)  10  -11  2001/01/15  [Not refereed][Not invited]
  • SAKAMOTO Naoya, OHASHI Toshiro, MATSUMOTO Takeo, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2001-  (13)  176  -177  2001/01/15  [Not refereed][Not invited]
  • SEO Shinji, OHASHI Toshiro, MATSUMOTO Takeo, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2001-  (13)  200  -201  2001/01/15  [Not refereed][Not invited]
  • MATSUMOTO Takeo, KAWAWA Takuo, NAGANO Yujiro, OHASHI Toshiro, SATO Masaaki  バイオエンジニアリング講演会講演論文集  2001-  (13)  228  -229  2001/01/15  [Not refereed][Not invited]
  • Effect of Magnetic Field on Nitric Oxide Synthesis of Cultured Endothelial Cells
    International Journal Applied Electromagnetics and Mechanics  14-  317  -322  2001  [Not refereed][Not invited]
  • Analysis of Local Mechanical Properties of Sheared Endothelial Cells Measured by Atomic Force Microscopy
    Proceedings of the ASME 2001 Summer Bioengineering Conference  2001  [Not refereed][Not invited]
  • Analysis of Local Mechanical Properties of Sheared Endothelial Cells Measured by Atomic Force Microscopy
    Proceedings of the ASME 2001 Summer Bioengineering Conference  2001  [Not refereed][Not invited]
  • 流体ー構造連成解析による流れ負荷培養内皮細胞の構造最適化シミュレーション
    日本数値流体力学会誌特集「CFD関連分野における連成シミュレーション」  9-  (4)  99  -105  2001  [Not refereed][Not invited]
  • N Sakamoto, T Ohashi, M Sato  INTERNATIONAL JOURNAL OF APPLIED ELECTROMAGNETICS AND MECHANICS  14-  (1-4)  317  -322  2001  [Not refereed][Not invited]
     
    The effect of magnetic field on nitric oxide (NO) production of cultured human umbilical vein endothelial cells (HUVECs) was investigated. Static magnetic fields (30 mT, 300 mT) and an oscillating magnetic field (30 mT, 20 Hz) were applied to HUVECs for 0.5, 1 and 6 hours and intracellular NO was observed using a fluorescent indicator. There was no effect of the static magnetic field on the NO production of the HUVECs. In contrast, the NO production increased with applying the oscillating magnetic field for 0.5 hour. However, such an increase of NO production was not observed for 1 and 6 hours. We performed a separate experiment to clarify the mechanism of NO synthesis by using an inhibitor of calcium-independent activation of NO synthase. The inhibitor suppressed the increase of NO production caused by the oscillating magnetic field. It can be concluded that oscillating magnetic field may activate calcium-independent pathway of activation for NO synthase.
  • OHASHI Toshiro, SEO Shinji, SATO Masaaki  The Computational Mechanics Conference  2000-  (13)  219  -220  2000/11/27  [Not refereed][Not invited]
  • 大橋 俊朗, 松本 健郎, 佐藤 正明  BME : bio medical engineering  14-  (10)  21  -24  2000/10/10  [Not refereed][Not invited]
  • SEO Shinji, OHASHI Toshiro, ISHII Yasuaki, SATO Masaaki  The proceedings of the JSME annual meeting  2000-  (2)  253  -254  2000/07/31  [Not refereed][Not invited]
  • SAKAMOTO Naoya, OHASHI Toshiro, SATO Masaaki  The proceedings of the JSME annual meeting  2000-  (2)  269  -270  2000/07/31  [Not refereed][Not invited]
  • OHASHI Toshiro, ISHII Yasuaki, SATO Masaaki  The proceedings of the JSME annual meeting  2000-  (2)  273  -274  2000/07/31  [Not refereed][Not invited]
  • Ohashi Toshiro, Seo Shinji, Sato Masaaki  Proceedings, ... meeting of Japan Society of Fluid Mechanics  2000-  259  -260  2000/07/25  [Not refereed][Not invited]
     
    In this paper, intracellular stress distributions in cultured endothelial cells exposed to fluid shear stress were studied using coupled field finite element analysis, in which cell surface geometries are measured by an atomic force microscope (AFM). After application of 6 h exposure to shear stress of 2 Pa, the endothelial cells showed marked elongation and orientation in the flow direction. A three-dimensional finite element model was generated on the basis of the cell surface geometry measured by the AFM. A fluid flow-structural analysis indicated that the endothelial cells may change th...
  • 坂元 尚哉, 大橋 俊朗, 佐藤 正明  運動療法と物理療法 = The Journal of physical medicine  11-  (3)  244  -244  2000/06/08  [Not refereed][Not invited]
  • 大橋 俊朗, 安倍 裕宣, 松本 健郎, 青木 隆平, 佐藤 正明  日本機械学會誌  103-  (978)  281  -281  2000/05/05  [Not refereed][Not invited]
  • MATSUMOTO Takeo, OHASHI Toshiro, SATO Masaaki, OZAWA Hiroshi, KOKUBUN Shoichi  運動療法と物理療法 = The Journal of physical medicine  11-  (1)  2  -10  2000/04/30  [Not refereed][Not invited]
  • OZAWA Hiroshi, KOKUBUN Shoichi, MATSUMOTO Takeo, OHASHI Toshiro, SATO Masaaki  運動療法と物理療法 = The Journal of physical medicine  11-  (1)  11  -16  2000/04/30  [Not refereed][Not invited]
  • IWAO Akiyo, OHASHI Toshiro, SATO Masaaki  講演論文集  2000-  (35)  198  -199  2000/03/11  [Not refereed][Not invited]
  • Changes in Local Mechanical Stiffness of Cultured Endothelial Cells by Exposure to Shear Stress
    Proceedings of the 12th Conference of the European Society of Biomechanics  2000  [Not refereed][Not invited]
  • Oscillating Magnetic Field Affects NO Synthesis of Cultured Endothelial Cell
    Proceedings of the 10th International Conference on Biomedical Engineering  2000  [Not refereed][Not invited]
  • Oscillating Magnetic Field Affects NO Synthesis of Cultured Endothelial Cell
    Proceedings of the 10th International Conference on Biomedical Engineering  2000  [Not refereed][Not invited]
  • Changes in Local Mechanical Stiffness of Cultured Endothelial Cells by Exposure to Shear Stress
    Proceedings of the 12th Conference of the European Society of Biomechanics  2000  [Not refereed][Not invited]
  • The Mechanical Response of Individual Chondrocytes
    Proceedings of the 12th Conference of the European Society of Biomechanics  2000  [Not refereed][Not invited]
  • The Mechanical Response of Individual Chondrocytes
    Proceedings of the 12th Conference of the European Society of Biomechanics  2000  [Not refereed][Not invited]
  • T. Ohashi, S. Sugawara, T. Matsumoto, M. Sato  JSME International Journal, Series C: Mechanical Systems, Machine Elements and Manufacturing  43-  (4)  780  -786  2000  [Not refereed][Not invited]
     
    Intracellular stress distributions in cultured endothelial cells exposed to fluid shear stress were studied using finite-element analysis, in which cell surface geometries were measured by atomic force microscopy. After applying shear stress, the endothelial cells showed marked elongation and orientation in the flow direction. Finite-element analysis indicated that wall shear stress in the central region around the nucleus was significantly higher than those in the upstream and downstream sides for both control, statically cultured cells, and sheared cells, but the intracellular stress concentrations between the central regions and the other two sides reduced with fluid flow. Fluorescent images showed that stress fibers of F-actin bundles were mainly formed in the central portion of the cells. These results indicate that endothelial cells may change their cytoskeletal structures to increase their stiffness and reduce stress concentrations in the cells.
  • Kato Yoko, Matsumoto Takeo, Ohashi Toshiro, Sato Masaaki  IEICE technical report.  99-  (191)  1  -5  1999/07/16  [Not refereed][Not invited]
     
    It has been suggested that local elastic moduli of soft biological tissues have a close correlation with their histology. We have developed an image analysis method in which structural components in histological images are classified based on their color tones to obtain their area fractions, and studied the relation between local elastic moduli of tissues and their histology quantitatively. Histological analysis was carried out of bovine thoracic aortas and atherosclerotic legions in rabbit thoracic aortas, whose local elastic moduli had been measured with a pipette aspiration technique. Th...
  • OHASHI Toshiro, MATSUMOTO Takeo, AOKI Takahira, SATO Masaaki  計算工学講演会論文集  4-  (2)  1059  -1060  1999/05/26  [Not refereed][Not invited]
  • 3-D Stress Analysis in Cultured Endothelial Cells Exposed to Fluid Shear Stress
    Proceedings of the 1999 ASME International Mechanical Engineering Congress and Exposition  1999  [Not refereed][Not invited]
  • 3-D Stress Distribution in Cultured Endothelial Cells Exposed to Shear Stress
    Proceedings of the First Joint Meeting of BMES & EMBS  1999  [Not refereed][Not invited]
  • 3-D Stress Distribution in Cultured Endothelial Cells Exposed to Shear Stress
    Proceedings of the First Joint Meeting of BMES & EMBS  1999  [Not refereed][Not invited]
  • Local Elastic Modulus of Atherosclerotic Lesions of Rabbit Thoracic Aortas and Its Relation to Histology
    Proceedings of the First Joint Meeting of BMES & EMBS  1999  [Not refereed][Not invited]
  • Toshiro Ohashi, Mitsunobu Yoshida, Kazuhiro Hane  IEEJ Transactions on Sensors and Micromachines  119-  (42591)  424  -429  1999  [Not refereed][Not invited]
     
    A novel optically addressed spatial light modulator for optical information processing is proposed. The modulator is integrated with micromirror, on which magnetic material is electroplated, photodiode and electric coil. A phtotocurrent will flow in the electric coil and the mirror supported by four cantilevers is actuated by the generated magnetic forces. This movable mirror is placed parallel to another fixed mirror, which structure is called a Fabry-Perot interferometer configuration. Theoretical analysis is performed to estimate the electromagnetic forces acting on the mirror and the displacement of the mirror. Numerical simulations by three-dimensional coupled magneto-structural finite element analysis are also performed, resulting in good agreement with theoretical results. In addition, it is shown that the proposed modulator is effective for pattern recognition. © 1999, The Institute of Electrical Engineers of Japan. All rights reserved.
  • Toshiro Ohashi, Takayuki Hirano, Minoru Sasaki, Kazuhiro Hane, Atsushi Ieki, Keiji Matsui, Masayuki Nashiki  IEEJ Transactions on Sensors and Micromachines  119-  (2)  86  -93  1999  [Not refereed][Not invited]
     
    In this paper, a novel optical encoder having pitch-modulated photodiode array as an index scale has been proposed for integrating the gratings with the detectors. The pitch-modulated photodiode array, which obtains four phase-shifted signals simultaneously, is specially designed to reduce higher harmonics distortions of displacement signal and then fabricated by using integrated circuit process technology. In a preliminary study, the effect of the sensitivity distribution of the photodiode on the measurement has been evaluated. In the encoder experiment, it is shown that the 3rd and 5th order distortions for the pitch-modulated photodiode array are reduced by approximately a factor of more than 10, and are almost independent of the air gap between the main scale and the index scale. The proposed encoder is effective for obtaining precise positioning. © 1999, The Institute of Electrical Engineers of Japan. All rights reserved.
  • Toshiro Ohashi, Mitsunobu Yoshida, Kazuhiro Hane  IEEJ Transactions on Sensors and Micromachines  119-  (42591)  424  -429  1999  [Not refereed][Not invited]
     
    A novel optically addressed spatial light modulator for optical information processing is proposed. The modulator is integrated with micromirror, on which magnetic material is electroplated, photodiode and electric coil. A phtotocurrent will flow in the electric coil and the mirror supported by four cantilevers is actuated by the generated magnetic forces. This movable mirror is placed parallel to another fixed mirror, which structure is called a Fabry-Perot interferometer configuration. Theoretical analysis is performed to estimate the electromagnetic forces acting on the mirror and the displacement of the mirror. Numerical simulations by three-dimensional coupled magneto-structural finite element analysis are also performed, resulting in good agreement with theoretical results. In addition, it is shown that the proposed modulator is effective for pattern recognition. © 1999, The Institute of Electrical Engineers of Japan. All rights reserved.
  • 3-D Stress Analysis in Cultured Endothelial Cells Exposed to Fluid Shear Stress
    Proceedings of the 1999 ASME International Mechanical Engineering Congress and Exposition  1999  [Not refereed][Not invited]
  • Local Elastic Modulus of Atherosclerotic Lesions of Rabbit Thoracic Aortas and Its Relation to Histology
    Proceedings of the First Joint Meeting of BMES & EMBS  1999  [Not refereed][Not invited]
  • Toshiro Ohashi, Takayuki Hirano, Minoru Sasaki, Kazuhiro Hane, Atsushi Ieki, Keiji Matsui, Masayuki Nashiki  IEEJ Transactions on Sensors and Micromachines  119-  (2)  86  -93  1999  [Not refereed][Not invited]
     
    In this paper, a novel optical encoder having pitch-modulated photodiode array as an index scale has been proposed for integrating the gratings with the detectors. The pitch-modulated photodiode array, which obtains four phase-shifted signals simultaneously, is specially designed to reduce higher harmonics distortions of displacement signal and then fabricated by using integrated circuit process technology. In a preliminary study, the effect of the sensitivity distribution of the photodiode on the measurement has been evaluated. In the encoder experiment, it is shown that the 3rd and 5th order distortions for the pitch-modulated photodiode array are reduced by approximately a factor of more than 10, and are almost independent of the air gap between the main scale and the index scale. The proposed encoder is effective for obtaining precise positioning. © 1999, The Institute of Electrical Engineers of Japan. All rights reserved.
  • Toshiro Ohashi, Yoko Kato, Takeo Matsumoto, Masaaki Sato  JSME International Journal, Series C: Mechanical Systems, Machine Elements and Manufacturing  42-  (3)  568  -573  1999  [Not refereed][Not invited]
     
    Local elastic moduli in the wall of porcine thoracic aortas were measured in the axial, circumferential, and radial directions by the pipette aspiration method. The local elastic moduli in the three directions were found to be independent of wall position. Then, histological analysis using color classification based on Mahalanobis' generalized distance was performed to quantitatively assess the correlation between elastic moduli and histology. Porcine thoracic aortas had a typical layered structure. The area fraction values, i.e., the percentage area of three structural components, elastin, collagen fibers and smooth muscle cells, were almost uniform in the wall. On the other hand, the moduli in bovine aortas reported previously decreased significantly from the inner to the outer sides of the wall. The histological structure was not homogeneous, that is, large clusters of smooth muscle cells existed in the outer side. These results suggest that the intramural distributions of elastic moduli are closely correlated with the histological structure.
  • TANAKA Kotaro, OHASHI Toshiro, GOTO Kazuya, HANE Kazuhiro  シンポジウム電磁力関連のダイナミックス講演論文集  10-  121  -122  1998/10/23  [Not refereed][Not invited]
  • YOSHIDA Mitsunobu, OHASHI Toshiro, HANE Kazuhiro  シンポジウム電磁力関連のダイナミックス講演論文集  10-  123  -124  1998/10/23  [Not refereed][Not invited]
  • OZAWA H  日本整形外科學會雜誌 = The Journal of the Japanese Orthopaedic Association  71-  (8)  S1400  1997/08/25  [Not refereed][Not invited]
  • T Aoki, T Ohashi, T Matsumoto, M Sato  ANNALS OF BIOMEDICAL ENGINEERING  25-  (3)  581  -587  1997/05  [Not refereed][Not invited]
     
    A simple method of identifying the initial slope of the stress-strain curve (i.e., Young's modulus of the soft tissue) by introducing the pipette aspiration technique is presented. The tissue was assumed to be isotropic and macroscopically homogeneous. Numerical simulations by the linear finite element analysis were performed for the axisymmetric model to survey the effects of friction at the tissue-pipette contact boundary, pipette cross-sectional geometry, relative size of the specimen to the pipette, and the layered inhomogeneity of the specimen tissue. The friction at the contact region had little effect on the measurement of Young's modulus. The configuration of the pipette was shown to affect the measurement for small pipette wall thickness. The measurement also depended on the relative size of the specimen to the pipette for relatively small specimens. The extent of the region contributing to the measurement was roughly twice the inside radius of the pipette. In this region, the maximum stress did not exceed the level of the aspiration pressure, with only minor exceptional locations. Calculation of strain energy components indicated that the major contributions to the deformation under pipette aspiration were by the normal extension and shear deformation in pipette axial direction. Experimental verification of the present method for the isotropic, homogeneous artificial material is also presented.
  • T Aoki, T Ohashi, T Matsumoto, M Sato  ANNALS OF BIOMEDICAL ENGINEERING  25-  (3)  581  -587  1997/05  [Not refereed][Not invited]
     
    A simple method of identifying the initial slope of the stress-strain curve (i.e., Young's modulus of the soft tissue) by introducing the pipette aspiration technique is presented. The tissue was assumed to be isotropic and macroscopically homogeneous. Numerical simulations by the linear finite element analysis were performed for the axisymmetric model to survey the effects of friction at the tissue-pipette contact boundary, pipette cross-sectional geometry, relative size of the specimen to the pipette, and the layered inhomogeneity of the specimen tissue. The friction at the contact region had little effect on the measurement of Young's modulus. The configuration of the pipette was shown to affect the measurement for small pipette wall thickness. The measurement also depended on the relative size of the specimen to the pipette for relatively small specimens. The extent of the region contributing to the measurement was roughly twice the inside radius of the pipette. In this region, the maximum stress did not exceed the level of the aspiration pressure, with only minor exceptional locations. Calculation of strain energy components indicated that the major contributions to the deformation under pipette aspiration were by the normal extension and shear deformation in pipette axial direction. Experimental verification of the present method for the isotropic, homogeneous artificial material is also presented.
  • T Aoki, T Ohashi, T Matsumoto, M Sato  ANNALS OF BIOMEDICAL ENGINEERING  25-  (3)  581  -587  1997/05  [Not refereed][Not invited]
     
    A simple method of identifying the initial slope of the stress-strain curve (i.e., Young's modulus of the soft tissue) by introducing the pipette aspiration technique is presented. The tissue was assumed to be isotropic and macroscopically homogeneous. Numerical simulations by the linear finite element analysis were performed for the axisymmetric model to survey the effects of friction at the tissue-pipette contact boundary, pipette cross-sectional geometry, relative size of the specimen to the pipette, and the layered inhomogeneity of the specimen tissue. The friction at the contact region had little effect on the measurement of Young's modulus. The configuration of the pipette was shown to affect the measurement for small pipette wall thickness. The measurement also depended on the relative size of the specimen to the pipette for relatively small specimens. The extent of the region contributing to the measurement was roughly twice the inside radius of the pipette. In this region, the maximum stress did not exceed the level of the aspiration pressure, with only minor exceptional locations. Calculation of strain energy components indicated that the major contributions to the deformation under pipette aspiration were by the normal extension and shear deformation in pipette axial direction. Experimental verification of the present method for the isotropic, homogeneous artificial material is also presented.
  • OHASHI Toshiro, ABE Hironobu, MATSUMOTO Takeo, AOKI Takahira, SATO Masaaki  Transactions of the Japan Society of Mechanical Engineers. C  63-  (607)  867  -874  1997/03/25  [Not refereed][Not invited]
     
    A pipette aspiration technique has been developed to clarify the local anisotropic stiffness of soft biological tissues. This technique provides a simple and practical method of measuring an initial elastic modulus by comparing the slope of the pressure-deformation curve obtained from a finite element analysis with that obtained by experiment using a pipette to aspirate the tissue surface. In previous studies, linear numerical simulations were performed for circular cross-sectional pipettes using an axisymmetric model that is assumed to be isotropic and homogeneous. In this paper, the use o...
  • Intramural Distribution of Local Elastic Moduli in Bovine Thoracic Aorta Measured by Pipette Aspiration Method
    Cellular Engineering  2-  (1)  12  -18  1997  [Not refereed][Not invited]
  • Intramural distribution of local elastic moduli in bovine thoracic aorta measured by pipette aspiration method
    Cellular Engineering  2-  (1)  12  -18  1997  [Not refereed][Not invited]
  • Intramural distribution of local elastic moduli in bovine thoracic aorta measured by pipette aspiration method
    Cellular Engineering  2-  (1)  12  -18  1997  [Not refereed][Not invited]
  • Takahira Aoki, Toshiro Ohashi, Takeo Matsumoto, Masaaki Sato  Annals of Biomedical Engineering  25-  (3)  581  -587  1997  [Not refereed][Not invited]
     
    A simple method of identifying the initial slope of the stress-strain curve (i.e., Young's modulus of the soft tissue) by introducing the pipette aspiration technique is presented. The tissue was assumed to be isotropic and macroscopically homogeneous. Numerical simulations by the linear finite element analysis were performed for the axisymmetric model to survey the effects of friction at the tissue-pipette contact boundary, pipette cross-sectional geometry, relative size of the specimen to the pipette, and the layered inhomogeneity of the specimen tissue. The friction at the contact region had little effect on the measurement of Young's modulus. The configuration of the pipette was shown to affect the measurement for small pipette wall thickness. The measurement also depended on the relative size of the specimen to the pipette for relatively small specimens. The extent of the region contributing to the measurement was roughly twice the inside radius of the pipette. In this region, the maximum stress did not exceed the level of the aspiration pressure, with only minor exceptional locations. Calculation of strain energy components indicated that the major contributions to the deformation under pipette aspiration were by the normal extension and shear deformation in pipette axial direction. Experimental verification of the present method for the isotropic, homogeneous artificial material is also presented.
  • Intramural Distribution of Local Elastic Moduli in Bovine Thoracic Aorta Measured by Pipette Aspiration Method
    Cellular Engineering  2-  (1)  12  -18  1997  [Not refereed][Not invited]
  • Local Mechanical Properties in Aortic Walls Measured by Pipette Technique
    Proceedings of the World Congress on Medical Physics and Biomedical Engineering  1997  [Not refereed][Not invited]
  • Local Mechanical Properties in Aortic Walls Measured by Pipette Technique
    Proceedings of the World Congress on Medical Physics and Biomedical Engineering  1997  [Not refereed][Not invited]
  • OZAWA H  日本整形外科學會雜誌 = The Journal of the Japanese Orthopaedic Association  70-  (8)  S1563  1996/08/25  [Not refereed][Not invited]
  • Local Elastic Moduli of Bovine and Porcine Aortic Walls Measured with Pipette Aspiration Technique
    1995 Advances in Bioengineering  1995  [Not refereed][Not invited]
  • Local Elastic Moduli of Bovine and Porcine Aortic Walls Measured with Pipette Aspiration Technique
    1995 Advances in Bioengineering  1995  [Not refereed][Not invited]
  • Application of the Pipette Aspiration Technique to the Measurement of Local Elastic Moduli of Cholesterol-fed Rabbit Aortas
    Theoretical and Applied Mechanics  43-  233  -238  1994  [Not refereed][Not invited]
  • Application of the Pipette Aspiration Technique to the Measurement of Local Elastic Moduli of Cholesterol-fed Rabbit Aortas
    Theoretical and Applied Mechanics  43-  233  -238  1994  [Not refereed][Not invited]
  • Local Mechanical Properties of Cholesterol-fed Rabbit Aortae Measured by the Pipette Aspiration Technique
    Proceedings of the 2nd World Congress of Biomechanics  1994  [Not refereed][Not invited]
  • Local Mechanical Properties of Cholesterol-fed Rabbit Aortae Measured by the Pipette Aspiration Technique
    Proceedings of the 2nd World Congress of Biomechanics  1994  [Not refereed][Not invited]
  • Finite Element Analysis of Local Deformation of Soft Tissues Aspirated by Pipette
    Proceedings of the 2nd World Congress of Biomechanics  1994  [Not refereed][Not invited]
  • Finite Element Analysis of Local Deformation of Soft Tissues Aspirated by Pipette
    Proceedings of the 2nd World Congress of Biomechanics  1994  [Not refereed][Not invited]

Books etc

  • ナノメデイシンの基礎と最先端
    株式会社オーム社 2008
  • MEME/NEMS工学大全
    株式会社テクノシステム 2008
  • Future Medical Engineering Based on Bionanotechnology
    Imperial College Press 2007
  • Future Medical Engineering Based on Bionanotechnology
    Imperial College Press 2007
  • Human Biomechanics and Injury Prevention
    Springer-Verlag 2000
  • Human Biomechanics and Injury Prevention
    Springer-Verlag 2000
  • Data Book on Mechanical Properties of Living Cells, Tissues, and Organs
    Springer-Verlag 1996
  • Data Book on Mechanical Properties of Living Cells, Tissues, and Organs
    Springer-Verlag 1996

Association Memberships

  • 日本機械学会バイオエンジニアリング部門国際委員会   第46回日本生体医工学会大会実行委員会   ASME Journal of Biomechanical Engineering   日本機械学会東北支部第41期総会・講演会実行委員会   The Second Japan-Switzerland Workshop on Biomechanics   日本機械学会論文集編集委員会   日本機械学会第14回バイオフロンティア講演会実行委員会   日本機械学会バイオエンジニアリング部門広報委員会   日本エム・イー学会東北支部（現：日本生体医工学会）   日本バイオレオロジー学会   日本機械学会第19回バイオエンジニアリング講演会実行委員会   日本エム・イー学会東北支部   日本機械学会バイオエンジニアリング部門   日本機械学会バイオエンジニアリング部門運営委員会   日本機械学会第13回バイオエンジニアリング講演会実行委員会   日本実験力学会   日本生体医工学会   Japanese Society for Medical and Biological Engineering   Acta Biomaterialia   第28回日本バイオレオロジー学会年会実行委員会   日本生体医工学会バイオメカニクス研究会   日本機械学会   The Japan Society for Experimental Mechanics   The 1st International Symposium on Future Medical Engineering based on Bio-nanotechnology   Japanese Society of Biorheology   Japan Society of Mechanical Engineers   

Works

  • マトリックスデバイスによる細胞の力学伝達経路の解明
    2006 -2007
  • 最適な腰椎牽引療法を目指した有限要素法解析
    2007
  • 生体内微小循環動態評価に関する研究
    2006 -2006
  • Bio-MEMSによる細胞操作に関する研究
    2006
  • コラーゲン・トリペプチド（CTP）の軟骨細胞および血管内皮細胞に対する機能性評価
    2004
  • 最適なステント設計のための有限要素法解析
    2003 -2003
  • ラミン欠損核の力学特性に関する研究
    2002 -2002
  • Resiliometerを用いた皮膚力学特性の計測に関する有限要素法解析
    2002 -2002
  • ヒト繊維芽細胞の力学特性に関する研究
    2000 -2000
  • 軟骨細胞の力学応答機構に関する研究
    1998

Research Projects

  • 日本学術振興会：科学研究費助成事業
    Date (from‐to) : 2023/04 -2026/03 
    Author : 大橋 俊朗, 豊原 涼太
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Fund for the Promotion of Joint International Research (Fostering Joint International Research (B))
    Date (from‐to) : 2019/10 -2023/03 
    Author : 大橋 俊朗, 東藤 正浩, 山田 悟史
     
    細胞は周囲の力学環境に適応して自らの形態および機能を変化させることが知られている．これは細胞が力学刺激を感知・伝達し生化学的信号に変換しているためでありこの機構はメカノトランスダクションと呼ばれている．例えば，血管内皮細胞は血流に応答して血流方向に伸長・配向することから盛んに研究が行われてきたがメカノトランスダクション機構の全容は明らかではない．本申請課題の目的は，磁気ナノビーズ技術を高度に駆使して流れ刺激に対する内皮細胞の力学伝達経路の推定するものである．近年では，内皮細胞表面に突出する直径0.2ミクロン，長さ数ミクロン～十数ミクロン程度の線状のPrimary ciliumが力学刺激の感知に深く寄与していることが指摘されている． 以上の背景の下，磁気ナノビーズ技術およびマイクロフルイディクス技術により内皮細胞に力学刺激を負荷する部位を空間的に制御し，内皮細胞の流れ負荷応答のメカニズムを新しい視点で探求するものである．研究代表者の大橋がこれまでに従事してきた流れ負荷内皮細胞実験技術，マイクロフルイディクス技術および海外共同研究者のProf. Ibarraより新たに導入する磁気ナノビーズ技術を高度に融合させ，海外共同研究者のProf. Longとともに流れ負荷刺激を模擬して磁気ナノビーズにより力学刺激部位を空間的に制御することにより，流れ負荷によるメカノトランスダクション機構を検討するものである．今年度は磁気ナノビーズを人工細胞内に導入し生細胞と共培養する実験系の確立に着手してきた．外部磁場を負荷することで人工細胞を駆動させることができるため細胞間力を任意に調整することが可能であり，細胞間の力学情報伝達経路を探求するものである．また，Primary ciliumの変形から力学特性を計測できる可能性が示唆されたため，Primary ciliumの力学特性計測という新しい研究展開も図った．
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)
    Date (from‐to) : 2018/04 -2021/03 
    Author : Ohashi Toshiro
     
    The purpose of this study is to measure mechanical properties of endothelial primary cilia with the diameter of ca. 200 nm and the length of several micron existing on the cell surface and to identify them as a mechanosensor. We fabricated an in-house micro-tensile tester to measure mechanical properties of primary cilia and successfully obtained their Young's modulus. In addition, when calcium experiments were performed on the isolated primary cilia by tensile stimulation, an increase in calcium expression was observed in the primary cilia, indicating that calcium channels existing on the surface of primary cilia could activate the entry of calcium ion.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)
    Date (from‐to) : 2016/04 -2019/03 
    Author : Kajiwara Itsuro
     
    This study developed a health monitoring system for detecting damage on structures. An approach for detecting a hole drilled on a pipe based on laser plasma acoustic excitation and acoustic measurement is included in the results of this study. Non-contact acoustic impulse excitation can be realized by laser-induced plasma and a microphone is used for measuring the time response of acoustic pressure. In this study, we focus on the detection of the hole on the pipe. The time response of acoustic pressure changes due to reflection of acoustic wave caused at the hole drilled on the pipe. The position of the hole is identified by applying continuous wavelet transform to measured time response data with/without the hole. This study demonstrated the effectiveness of the present damage detection method based on the acoustic excitation using the laser-induced plasma.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Challenging Exploratory Research
    Date (from‐to) : 2013 -2015 
    Author : Ohashi Toshiro, Cooper-White Justin
     
    Cell migration is known to play an important role in a number of physiological events in living body such as morphogenesis, wound healing, and tumor metastasis. It is therefore important to study the mechanism of cell migration from the viewpoint of mechanics. The objective of this study is to newly design and fabricate a multichannel device integrated with micropillar technology to evaluate cell traction forces during cell migration. The multichannel device has been successfully fabricated, which allows to modulate cell migration rate by changing the stiffness of micropillars and by giving the micropillars anisotropic properties.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (A)
    Date (from‐to) : 2011/04 -2014/03 
    Author : YASUDA Kazunori, OMIYA Yoshihiro, OHASHI Toshiro, KUROKAWA Takayuki, KITAMURA Nobuto, TANAKA Shinya, SEMBA Shingo
     
    This study discovered several important phenomena and their molecular mechanisms during chondrogenic differentiation and regeneration of mesenchymal cells induced by the PAMPS/PDMAAm gel or soluble proteins:(1) ATP oscillation, which depends on oscillation of both the mitochondrial respiration and the glycolysis, was induced by calcium ion oscillation. The ATP oscillation does not control cell differentiation but controls cell aggregation and tissue formation via controlling secretion of soluble proteins. (2) Young's modulus of a chondrogenic precursor cell increased to a significantly greater degree during differentiation induced by the gel than during differentiation induced by Insulin. (3) Both the BMP-Smad signaling pathway and the IGF-depending signaling pathway were specifically activated during the differentiation induced by DN gel in comparison with that induced by Insulin. (4) The proteomics analysis showed that the gel has a function as a reservoir of soluble factors.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)
    Date (from‐to) : 2012 -2014 
    Author : OHASHI Toshiro, MAEDA Eijiro, ANDERSSON-SVAHN Helene, COOPER-WHITE Justin
     
    In this project, MEMS devices that allow to realize measurements and control of physical environment of cells were developed. The developed MEMS devices are for experiments of cell responses to mechanical environment, cell responses to chemical environment and cell migration. For the experiment of cell mechanical responses, it was confirmed to apply different level of fluid shear stress to endothelial cells cultured in a microwell to perform flow experiments. For the experiment of cell chemical responses, it was confirmed to apply different concentration of Saponin solution to endothelial cells to perform live/dead cell assay. For cell migration assay, mechanical characterization of fibroblast migration could be performed by using a micropillar array.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Challenging Exploratory Research
    Date (from‐to) : 2010 -2012 
    Author : OHASHI Toshiro, MAEDA Eijiro
     
    This paper presents development of a technique to control cell adhesion usingan array of magnetic microbeads. Using MEMS techniques, we fabricated an array of microholes (3 μmin diameter and 2 μm in depth) on the surface of a glass substrate, to trap magnetic microbeads (2.8 μmin diameter) in the microholes. The magnetic microbeads were coated with fibronectin prior to theloading. A permanent magnet was placed under the substrate and was used to trap microbeads into themicroholes. After 24 hours incubation, it was observed that cells attached only to the magneticmicrobeads, but not to the other parts of the glass surface.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Specially Promoted Research
    Date (from‐to) : 2008 -2012 
    Author : SATO Masaaki, OHASHI Toshiro, KANNZAKI Makoto, DEGUCHI Shinji, SAKAMOTO Naoya, ADACHI Taiji, OGURA Toshihiko, OHASHI Kazumasa
     
    There are many cells sensing mechanical forces in our body and the cells control the functions of tissues and organs to be adapted to mechanical environment. However, the mechanisms were not elucidated yet. In this study, we mainly focused on vascular endothelial cells, osteocytes and myocytes. It became clear that cell-cell and focal adhesions and cytoskeletons such as stress fiber are working as mechanosensors and some proteins in cytoplasm were identified as signal messengers. Mechanical forces were also playing important roles in organogenesis such as valve formation in cardiovascular system.
  • Ministry of Education, Culture, Sports, Science and Technology：Grants-in-Aid for Scientific Research(基盤研究(B))
    Date (from‐to) : 2008 -2010 
    Author : Toshiro OHASHI, 佐藤 正明, 坂元 尚哉, Eijiro MAEDA, Masaaki SATO, Naoya SAKAMOTO
     
    Since most laboratory bioassays used for cell analysis are designed to measure average effects on cell population, it is difficult to study characteristics of individual cells such as patient's tumor cell samples in a high throughput manner. Recently, a novel microwell slide that allows several thousands of single cells to be addressed to an exact position, cultured for weeks, and treated separately in high throughput has been developed. In this project we have proposed a bioassay system consisting of an active microfluidic device integrated on the microwell plate for improved control of, a...
  • 文部科学省：科学研究費補助金(特定領域研究)
    Date (from‐to) : 2008 -2008 
    Author : 大橋 俊朗, 佐藤 正明, 坂元 尚哉
     
    生体内において細胞は力学環境に曝されている. 例えば, 血管内皮細胞や平滑筋細胞には血流によるせん断応力や血管拡張に伴う伸展刺激が作用している. 細胞はこのような周囲の力学環境に適応して自らの形態および物質産生等の機能を変化させることが知られている. これは細胞が力学刺激を感知・伝達し生化学的信号に変換しているためであり力学情報伝達(mechanotransduction)と呼ばれている. 申請者らはこれまでに, 力学情報伝達の経路の一つに細胞骨格が深く関与していると考え力学刺激に対する内皮細胞や平滑筋細胞の骨格構造変化を観察してきた. このような中, 申請者らはマイクロピラーを有する基質の上に平滑筋細胞を培養し, マイクロピラーの撓み細胞牽引力を計測することで細胞内の力学情報伝達経路を探る研究に本格的に着手した. 例えば, ガラスのマイクロニードルで細胞辺縁部の特定のマイクロピラーを撓ませると, ストレスファイバーによって接続されていた他端のマイクロピラーが撓むことから細胞骨格を介した力学伝達が実現されていることを確認した. 研究を遂行していく中で, この技術を応用し任意のマイクロピラーを能動的に動かすことができれば特定の焦点接着斑にのみ選択的に力学刺激を負荷できるのではないかと考えた. そこで本研究課題では, 従来の研究をさらに発展させ, 磁性ビーズを埋入したマイクロピラ...
  • 日本学術振興会：科学研究費助成事業 萌芽研究
    Date (from‐to) : 2007 -2008 
    Author : 佐藤 正明, 大橋 俊朗, 坂元 尚哉
     
    本研究の目的は,分子スイッチをコートした磁気ビーズを細胞内に導入し,これを外部から磁場を印可することによって任意に操作し,細胞の仮足形成を任意に操作し,細胞運動を制御するところにあった。研究第一段階として,細胞内での磁気ビーズ操作に必要な磁気ピンセット装置の開発を行った。より自由度の高い操作を実現する為,電磁石駆動型の装置を開発し,電源のON/OFFによって磁場のコントロールが可能な系とした。また,持続的な操作を可能とするため,電磁コイルの冷却装置を実装した。第二段階として,細胞内に食作用によって導入した磁気ビーズを,実際に細胞内で空間的に操作することを試みた。細胞内に導入された複数個の磁気ビーズを任意の位置(細胞辺縁部)へと集積させることに成功した。また,共焦点レーザー顕微鏡を用いたリアルタイム三次元観察によって,ビーズ操作が細胞膜内で行われていることも確認された。次に,分子スイッチをコートしか磁気ビーズを作製し,細胞内への導入を試みた。当初は細胞のエンドサイトーシス作用などを用いる予定であったが,この手法では磁気ビーズが細胞内でエンドソームなどのリン脂質に包まれた状態となってしまう為,分子スイッチが機能しなかった。そこで,物理的外力によって細胞内に微粒子を撃ち込むパーティクルガン装置によって磁気ビーズの導入を試みた。磁気ビーズを本手法によって導入すること自体には成功したものの,本来,当該装置は植物細胞に金やタングステンなどの比重の大きい微粒子を撃ち込む装置であり,本研究では動物細胞および比重の軽い磁気ビーズを用いることから,実験条件が大きく異なるため,これまでのところ細胞に与えるダメージを十分低減させて導入するまでには至っておらず,導入条件を検討中である。
  • 文部科学省：科学研究費補助金(萌芽研究)
    Date (from‐to) : 2006 -2008 
    Author : 大橋 俊朗, 佐藤 正明, 坂元 尚哉
     
    細胞の力学情報伝達は細胞の生理・病理に深く関わることからそのメカニズムを知ることは重要であるが, 力学刺激が実際に細胞内でどのように伝達されているか, またそれによって機能がどのように修飾を受けるのかに関しては未知の部分が多い. 本研究では, 微細加工技術により細胞底面に発生する力を計測できる実験系を構築し, 基質形状により細胞の形態を制御すること, さらに力学刺激が細胞骨格を伝達する経路を求めることを目的とする. 微細加工技術を用いて細胞を培養するマトリックス上にPDMSによるマイクロピラーアレイ(高さ10μm, 直径3μm, ピラー間ピッチ8μm)を形成し, マイクロピラーのたわみから細胞底面で発生する牽引力を計測するものである. 本年度においては, 前年度に引き続き細胞形態制御実験, マイクロピペットによる力学環境操作実験, 細胞骨格の破壊実験を行い, データの蓄積を試みた. 細胞形態制御実験では, ピラー間ピッチを6, 8, 10μmと変化させ, 細胞はピラー間ピッチに依存して牽引力を変化させることを示した. マイクロピペットによる力学環境操作実験ではより精度良くマイクロピラーによる局所力学刺激負荷を行うため磁気駆動式マイクロピラーの製作にとりかかっているところである. 細胞骨格の破壊実験では, アクチンフィラメント, 微小管, 中間径フィラメントそれぞれの破壊により...
  • 文部科学省：科学研究費補助金(特定領域研究)
    Date (from‐to) : 2006 -2007 
    Author : 大橋 俊朗, 佐藤 正明, 坂元 尚哉
     
    細胞の力学情報伝達は細胞・組織の生理・病理に深く関わることからそのメカニズムを知ることは非常に重要であるが,力学刺激が実際に細胞内でどのように伝達されているか,またそれによって機能がどのように修飾を受けるのかに関しては未知の部分が多い.本研究では,マイクロマシニング技術により細胞底面に発生する力を直接計測できる実験系を構築し,力学刺激が細胞骨格を伝達する経路およびその大きさを定量的に求めること,さらにこの実験系により機能制御の実現可能性を探ることを目的とする.微細加工技術を用いて細胞を培養するマトリックス(細胞外基質)上にPDMSによるマイクロピラーアレイ(高さ10μm,直径3μm,マイクロピラー間ピッチ8μm)を形成し,マイクロピラーのたわみから細胞底面で発生する牽引力を計測するものである.本年度において,先端径が2〜3μm程度のガラスのマイクロピペットをマイクロマニピュレータで操作し,特定のマイクロピラーを強制的にたわませる実験を行った.その結果,たわませたマイクロピラーに接続しているストレスファイバの他端のマイクロピラーが連動してたわむ様子が観察された.すなわち,ストレスファイバを通して一つの焦点接着斑からもう一つの焦点接着斑に力学刺激が伝達されていることが明らかとなった.また,ノコダゾール処理により微小管を破壊したところ牽引力は有意に増大した.一方,微小管破壊によりス...
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (A)
    Date (from‐to) : 2005 -2007 
    Author : SATO Masaaki, OHASHI Toshiro, DEGUCHI Shinji, SAKAMOTO Naoya
     
    1) Evaluation of mechanical property of stress fibers Mechanical properties of stress fibers extracted from smooth muscle cells (SMCs) and endothelial cells (ECs) were measured with a microneedle-based tensile test system developed in this study. Strain-induced stiffening was observed in the force-strain curve. We also found that the extracted SFs shortened to approximately 80% of the original length after they were dislodged from the substrate, suggesting that SFs had preexisting strain in the cytoplasm. The force required for stretching the single SFs from the zero-stress length back to the original length was approximately 10 nN for SMCs and 4.1 nN for ECs. 2) Measurement of traction forces of cells using microfabricated micropost arrays A mold of silicon substrates with arrays of micro-needle-like posts (3μm in diameter, 10μm in height, and 8μm intervals) was fabricated to estimate cellular traction forces using microfabrication techniques. SMCs spreading on the substrates produced deflection of the posts and was associated with organization of stress fibers of actin filaments. Traction forces varied considerably among cells, showing an average of approximately 12 nN. 3) Cyclic stretch test with microsubstrates We applied cyclic stretch to ECs using a microstructured substrate with arrays of micropost, on which cells were selectively stretched between FAs but FA-substrate contact area were hardly stretched. After exposure ECs to cyclic stretch for 3h, cells on both a flat and the micropost substrates aligned perpendicular to the direction of stretch. Stress fibers oriented about 60° to the stretch direction for the flat substrate at 3h, while stress fibers on the micropost substrate oriented approximately 90° to the stretch direction after 6h stretching. These results suggest that strain in FA-substrate contact area may have impact on rates of reorienation of stress fibers in ECs in response to cyclic stretch.
  • 文部科学省：科学研究費補助金(若手研究(A))
    Date (from‐to) : 2005 -2007 
    Author : 大橋 俊朗
     
    細胞の力学情報伝達機構を知ることは細胞の生理・病理を知る上で非常に重要である.力学刺激が実際に細胞内でどのように伝達されているかに関してはその計測の困難さからほとんど解明されていない.本研究では細胞内で力学刺激が伝達していく過程を明らかにするため,ピペット吸引法とFRAP技術を組み合わせて力学刺激に対する細胞骨格構造の変化を蛍光画像として追跡し画像解析により細胞内の力学環境を定量的に把握することを目的とする.またセカンドメッセンジャーの一つであるカルシウムイオン濃度の変化を同時にイメージングすることにより骨格構造変化との相関関係を調べる.本年度においては,FRAP技術を用いてピペット吸引時の軟骨細胞におけるアクチン分子の拡散の様子を詳細に調べた.その結果,ピペット先端において吸引直後はアクチンコーティカルレイヤーが分解され,その後,アクチンの再重合が促進されていることがわかった.また,このプロセスはカルシウム依存でありカルシウムが存在しない場合には,アクチンの脱重合が引き起こされにくいことが示唆された.さらに,ピペット吸引時において力学刺激が細胞内をどのように伝達していくのかを観察するために,ミトコンドリアの染色画像より画像相関法を用いて細胞内ひずみ分布を得た.その結果,吸引直後は細胞周囲のアクチンコーティカルレイヤーを介して細胞全体に力学刺激が伝達される様子が明らかとなった...
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research on Priority Areas
    Date (from‐to) : 2003 -2007 
    Author : WADA Hiroshi, ADACHI Taiji, MASTUMOTO Takeo, USHIDA Takashi, TANAKA Masao, OHASHI Toshiro
     
    本研究では平成15年度から18年度まで実施した特定領域研究「マイクロ・ナノバイオメカニクスの開拓」の成果を学術図書として取りまとめた.本特定領域研究の目的を以下に記す.生命現象の本質的な多様性は,遺伝子のみではなく,細胞をめぐるマイクロあるいはナノスケールの構造と遺伝情報との相互作用によって担われており,その中でも,力学的相互作用の意義は極めて大きい.従って,力学的環境と生命機能の関係を解明することができれば,細胞及び組織の構造と機能をコントロールする手段を見いだせる可能性が高い.また,これを用いることによって,必要な構造と機能をもつ細胞と組織を創成することが可能となる.そこで,力学を基礎とし,マイクロ領域である細胞レベル及びナノ領域である生体高分子や細胞内小器官レベルの力学現象を解明し,これに基づいて人工材料の新たな設計指針の開発と創製を行うと共に,生命体の機能と構造を創成ないし再構成する技術を開発してきた.日本の生体工学・バイオメカニクス研究を牽引する研究者を各計画研究の代表者として集め,バイオメカニクス的視点を強く打ち出し,細胞や生体組織の再構築との関連性ならびに基礎科学から臨床応用までの広い視点をもった研究を展開し,世界一級の成果を上げて来たので,これら成果を取りまとめた.具体的には以下を行った. 1. 第10回全体会議(平成19年10月,東京)ならびに第11回全体会議,(平成20年1月,東京)を開催し,研究グループの研究成果に関する情報交換を行った. 2. 本領域の4つの班毎に,研究成果を取りまとめた. 3. 4班の成果を統合し,全体にわたった構成を4班の代表者4名により取りまとめた. 4. 出版社との打ち合わせを行い,全体の原稿を完成した. 5. 印刷原稿の校正を各研究代表者が行った. 6. 印刷製本を業者に委託し,学術図書を完成させ,研究成果を広く公開した.
  • 日本学術振興会：科学研究費助成事業 萌芽研究
    Date (from‐to) : 2005 -2006 
    Author : 佐藤 正明, 大橋 俊朗, 出口 真次, 坂元 尚哉
     
    細胞の形態変化と機能を理解する上で,主要な構成要素であるストレスファイバの力学環境に対する応答性,すなわち力学特性を把握しておくことは大変重要である.最終年度である本年度は,前年度に開発・確立した装置およびストレスファイバ単離手法を用いて以下の2つの研究を行った. 1.ストレスファイバの粘弾性特性の計測 前年度確立した単離方法に従って平滑筋細胞から単離したストレスファイバの引張試験を行い,ひずみ速度が引張特性に与える影響を調べた.ひずみ速度を0.01s^<-1>,0.05s^<-1>,0.1s^<-1>と設定しストレスファイバの引張特性計測を行ったところ,0.01s^<-1>と0.05s^<-1>における張力ーひずみ関係はほぼ線形で両者に顕著な差は見られなかった.一方,生理的なひずみ速度と考えられる0.1s^<-1>では上に凸の非線形な関係が得られた.またストレスファイバの変形初期の強度を示す初期スティフネスに関しても0.1s^<-1>では0.01s^<-1>および0.05s^<-1>に比べ大きくなる傾向が得られた.これらの結果から,ストレフファイバの引張特性にはひずみ速度依存性の閾値が存在する可能性が考えられた. 2.ストレスファイバの局所ひずみ計測 量子ドットを用いてストレスファイバを斑点状に可視化し,これをマーカーとして伸展時におけるストレフファイバ内の局所挙動を観察した.その結果,ストレフファイバは一様に伸展しているのではなくその挙動は局所で異なり,部位によって収縮を示す場合があることが明らかになった.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research on Priority Areas
    Date (from‐to) : 2003 -2006 
    Author : SATO Masaaki, OHASHI Toshiro, DEGUCHI Shinji, SAKAMOTO Naoya
     
    The goal of this project is to study dynamics of FAT domain in focal adhesions and associated actin filaments in order to understand remodeling process of adherent cells in response to external mechanical stimuli. In particular, newly developed experimental techniques here include a time-course observation of dynamics of FAT domain and actin filaments under application of fluid flow and a stretching system using an elastic membrane with nonuniform surface structure. First, we have developed an experimental system which enables to observe dynamics of cells using RFP-conjugated FAT-domain vector and GFP-conjugated actin vector. The result showed that actin filament structure was reconfigured in a close association with appearance/disappearance of FAT domain. Second, PDMS membrane with an array of micropillars has been fabricated by using soft lithography technique to locally apply stretching between focal adhesions. Endothelial cells formed focal adhesion complexes at peripheries of the top of the micropillars and developed stress fibers, thick actin bundles, perpendicular to the direction of stretch. Finally, we have developed a micro-tensile tester to measure mechanical properties of stress fibers, which would be critical for understanding of intracellular stress transfer. Tensile tests were, for the first time, carried out on stress fibers chemically isolated from endothelial cells, providing a load-displacement relationship. It was found that the load-displacement curve was nonlinear with high extensibility. We have successfully studied important roles of cytoskeletons and focal adhesions of mechanically stimulated cells in intracellular force transmission mechanism.
  • 日本学術振興会：科学研究費助成事業 萌芽研究
    Date (from‐to) : 2003 -2004 
    Author : 佐藤 正明, 坂元 尚哉, 大橋 俊朗
     
    細胞は細胞外基質と焦点接着斑と呼ばれるタンパク質の塊状部位によって接触を実現し形態を維持していると考えられている。従って力学的刺激あるいは化学的刺激に対する細胞の応答を知る上で焦点接触力を計測することは重要である。本研究では,シリコンの微細加工技術を用いて微小なセンサを開発し細胞の焦点接触力を測定することを目的とする。最終年度である今年度は,初年度に目指していたピエゾ抵抗型センサ方式を変更し,マイクロポストのたわみから接触力(牽引力)を推定する方式を検討した。まず,アレイ状マイクロポストのデザインを起こしレチクルを製作した。次にシリコンウェハにレチクルのデザインを露光転写した。ウェハをエッチングしレジストを剥離して型を製作した。続いて型にシリコーンポリマーを流し込みカバーガラスを載せた後,加温硬化させた。硬化後,型からカバーガラスごとシリコーンポリマーをはがし細胞外基質を完成させた。マイクロポストのサイズはおよそ直径3μm,高さ10μm,中心間距離8μmである。試料として,当初の予定であった内皮細胞よりも骨格構造により特徴のある平滑筋細胞を用いた。製作した基質上に平滑筋細胞を培養しマイクロポストのたわみを画像処理によって求めた。焦点接着斑の位置を確認し,かつアクチンフィラメントとの関係を把握するために,RFP-FATとGFP-アクチンのベクターを細胞に導入した。その結果,推定された平滑筋細胞の牽引力は11.8nNであり,最大値は81.3nNであった。また,それストレスファイバの分布から推測される力の合成ベクトルと牽引力のベクトルの方向は明らかに一致していた。このことからストレスファイバの発達は細胞の牽引力に支配的であることが考えられた。以上のように,本研究ではマイクロポストを有する細胞外基質デバイスを製作し,細胞底面に発生するミクロな力学環境を解明することに成功した。
  • Ministry of Education, Culture, Sports, Science and Technology：Grants-in-Aid for Scientific Research(基盤研究(B))
    Date (from‐to) : 2003 -2004 
    Author : 大橋 俊朗, Naoya SAKAMOTO, 坂元 尚哉, Masaaki SATO
     
    This project investigated the mechanism of development of atherosclerosis using endothelial-smooth muscle cell (EC-SMC) cocultured blood vessel model and obtained the following results.1.Migration of SMCs in the cocultured blood vessel models increased about 1.8 times higher than that in the model constructed with SMCs only. Exposure the cocultured model to fluid shear stress resulted in suppression of SMC migration. The migration of SMCs was not decreased by exposure to shear stress with medium containing an inhibitor of nitric oxide (NO) synthase. Theses results indicate that EC derived N...
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research
    Date (from‐to) : 2002 -2004 
    Author : SATO Masaaki, HANE Kazuhiro, MATSUMOTO Takao, OHASHI Toshiro, SAKAMOTO Naoya, DEGUCHI Shinji
     
    In order to elucidate the response mechanisms of endothelial cells to mechanical stimuli, dynamic changes in actin filaments and integrins were precisely observed using confocal laser scanning microscopy. Shear stress, membrane tension of substrate and hydrostatic pressure as mechanical stimuli were applied to cultured endothelial cells and the following conclusions were obtained. 1.GFP-actin and RFP-FAT (focal adhesion targetting) vectors were transfected into cultured endothelial cells that were used to impose to shear stress. Actin filament was suggested to sense the tension and occur the dynamic changes as the first step. Secondly the lamellipodia were formed. During this process, it was suggested that the movement and allocation of integrins are possible to be controlled through microtubules. 2.Endothelial cells were cultured on silicone membrane in which a cover glass was immersed to form heterogeneous strain distribution. A cell was exposed to non-uniform strain and showed that distribution of actin filaments and the location of nucleus was dependent on the strain distribution. It was suggested that the cell is able to sense mechanical stress and respond so as to keep the mechanical balance in the cell. 3.We have reported the cultured endothelial cells respond to the hydrostatic pressure and show multilayering, cell elongation with no predominant orientation and characteristic distribution of actin filaments. To elucidate the mechanisms, we have focused on expression and distribution of VE-cadherin as an important protein to adhere the cells. It was suggested that the hydrostatic pressure has important effect on the expression of VE-cadherin.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research
    Date (from‐to) : 2001 -2003 
    Author : MATSUMOTO Takeo, NAGAYAMA Kazuki, OHASHI Toshiro, SATO Masaaki
     
    Vascular smooth muscle cells not only change blood vessel diameter through their contraction and relaxation, but also remodel blood vessel wall by producing extracellular matrices such as collagen. Their contractile properties and matrix synthesis change in response to the mechanical stimuli applied to them : For example, their contractile velocity increases with repetitive contraction, and the energy necessary to maintain their isometric contraction decreases when the cells are kept contracted. Thus, the smooth muscle cells have interesting characteristics as a functional material. In this three-year study, we have examined the mechanical properties and the contractile properties of the vascular smooth muscle cells in a single cell level to clarify their relationship with the mechanical stimuli. In the first year, we found that the elastic modulus of the smooth muscle cells isolated from the hypertensive rat thoracic aorta was about one half of that of the normotensive aorta. In the second year, we found that the elastic modulus of the smooth muscle cells obtained from the rabbit thoracic aorta increased by the factor of 10 when they got contracted. In the last year, we examined the relationship between the tensile properties of the cell and the amount of their intracellular actin filament : We first measured the tensile properties of the smooth muscle cells freshly isolated from the normotensive rat thoracic aortas (FSMCs) and those of the cultured cells of the same origin (Passage 4-9, CSMCs). Then, we stained the cells with rhodamine-phalloidin for actin filaments to know the difference in the actin filament amount between the FSMCs and the CSMCs. The initial elastic modulus of the cells was about 11 kPa (N=8) for FSMCs and about 2.6 kPa (N=10) for CSMCs. The actin filaments in the FSMCs were abundant in the cytosol and it was hard to distinguish each filaments, while in the CSMCs, the filaments were sparsely distributed and stress fibers are clearly distinguishable, indicating that the decrease in the actin filament amount in the CSMCs. Such difference in the actin filament amount may cause the difference in the mechanical properties of the cells. We should analyze quantitatively the relationship between the actin filament amount and the mechanical properties in the next study.
  • 文部科学省：科学研究費補助金(奨励研究(A), 若手研究(B))
    Date (from‐to) : 2001 -2002 
    Author : 大橋 俊朗
     
    本研究では流れ負荷培養内皮細胞のアクチンフィラメントが再構築される過程をリアルタイムで観察し,同時に有限要素法解析を行い細胞の力学応答のメカニズムを解明することを目的とする.平成14年度では以下の成果を得た.昨年度用いたGFP-アクチン融合ベクタに加えて,本年度はRFP-FAT融合ベクタを遺伝子導入した.FATとはFocal Adhesion Targetingの略称であり,これにより細胞底面の焦点接着斑の挙動を観察できるため,アクチンフィラメント挙動と併せて内皮細胞の力学応答をより詳細に観察できる.これらの遺伝子を培養ウシ内皮細胞に遺伝子導入し,流れによるせん断応力を5時間にわたって負荷し,共焦点レーザ顕微鏡によりアクチンフィラメントおよび焦点接着斑の挙動を経時的に観察した。その結果,接着斑は流れの方向へ配向し,アクチンフィラメントの収縮に伴い移動する様子が観察された.また,焦点接着斑の間を連結するようにアクチンフィラメントが新たに形成されていく様子も観された.以上のことより,接着斑とアクチンフィラメントの相互関係が流れ負荷内皮細胞の形態変化に深く関与していることが示された.また,細胞骨格構造を模擬した有限要素モデルを構築した.成長の構成則に代表される構造最適化のアルゴリズムを導入して計算を行った結果,実験結果と同様に流れ負荷後,細胞の高さは減少傾向を示し。流れ負荷に伴い流...
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research
    Date (from‐to) : 2000 -2002 
    Author : MATSUMOTO Takeo, OHASHI Toshiro, SAIO Masaaki
     
    A cell rotation system has been developed to observe the 3D miciostructure of cells precisely from their images obtained from various view points. In this system, a specimen cell is held with a micropipette by aspirating the cellular surface gently under an upright microscope with water immersion objectives, and is rotated around the axis of the micropipette which is set perpendicular to the optical axis of the microscope. The pipette was mounted on a 3D electric micromanipulator and the position of the pipette tip was controlled based on its image taken with a CCD video camera mounted on the microscope. The pipette tip was coated with black ink and the tip position in the focusing (X-Y) plane was determined by binarization of the tip image. The tip position signal was fed to a comparator controlling the micromanipulators in X and Y direction. The pipette tip was kept focused with a PID controller maintaining the binarized area of the pipette tip minimum by changing the Z position of the pipette. Cultured bovine thoracic aortic smooth muscle cells were stained with SYTO13 for their nucleus and DiI for their cellular membrane and attached to the micropipette tip. Pan-focus images were constructed from optical sections across a cell at every 9° from 0° to 18° . The three-dimensional images of the cell nucleus and the cell membrane were obtained from these pan-focus images. These reconstructed images indicated that the nucleus of the cell detacher from the substrate wirth trypsin was round in shape, while that of the cell fixed with formalin on the substrate was elongated and flattened. The present method for the reconstruction of the cell microstructure with the cell rotation system would be useful for quantitative 3D analysis of the cell morphology.
  • 日本学術振興会：科学研究費助成事業
    Date (from‐to) : 2000 -2001 
    Author : 佐藤 正明, 大橋 俊朗, 松本 健郎
     
    魚類や円口類(ヤツメウナギなど)の体の両側にある側線は音を感じるセンサであることはよく知られている.また,このセンサは魚などのまわりの水の動きに反応しているのではないかという考えが提案されているが,このような観点からの研究例はほとんどない.もし,水の動きを感じることができるとすれば,流れのせん断応力あるいはせん断速度を計測可能なセンサとして利用可能になる.そこで,本研究では魚などの側線に存在する感覚細胞を取り出して培養し,流れ負荷の状態で魚の感覚細胞の特性を明らかにし,その応用についても議論することを目的とする. これまで報告されている感覚細胞を使った研究ではカジカが利用されているが,本研究ではカジカが入手困難のため金魚を使い,次の結果を得た. 1.金魚の側線感覚細胞の存在する部位から組織を切り出し,Ca^<2+>,Mg^<2+>を含まないハンクス液にコラゲナーゼとエラスターゼを含む溶液を作製して,組織を処理した.種々の濃度の組み合わせにより検討した結果,最適の濃度を見いだし,側線感覚細胞を単離することができた.単離後,細胞活性をトリパンブルーにより観察したところ,特に強い傷害も受けていないことを確認した. 2.感覚細胞を通常の細胞培養液中にて培養を試みたところ,2〜3日の培養は可能であるが,その後は急速に細胞活性を失っていくことを確認した. 3.感覚細胞をゲル中に包埋してせん断応力を負荷する装置にセットし,流れ負荷をかける実験を試みたが,細胞の活動度を電極を用いて定量的に評価する計測までには至らなかった.今後はこの点を中心に検討を進める予定である. 4. せん断応力負荷時の側線細胞の応答機構について有限要素法を用いて解析し,細胞内の変形と応力分布を得た.今後細胞内構造と応力分布の関係を検討する予定である.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research
    Date (from‐to) : 2000 -2001 
    Author : SATO Masaaki, OHASHI Toshiro, MATSUMOTO Takeo
     
    Under flow condition endothelial cells are elongated and oriented to flow direction depending upon the level of shear stress and the duration of exposure. It is well known that cytoskeletal components, especially F-actin filaments, are playing important roles in the process of adaptation of cell shape to mechanical environment. In this project We have mainly focued two topics and obtained the results shown below. 1. Dynamic behavior of actin filament in living cells We observed dynamic behavior of actin filaments in living cultured endothelial cells during exposure to shear stress. To do this, a vector of a green fluorescent protein (GFP)/actin fusion was introduced into cells using a lipofectoamine. Dynamic behavior of actin filament in an endothelial cell after introducing cytochalasin D was observed through CCD camera under an inverted fluorescent microscope. In different experiments, the endothelial cells were exposed to fluid shear stress of 2 Pa in a parallel plate flow chamber and the cytoskeletal structure was observed. Actin filaments located first at peripheral regions seemed to change the alignment after flow exposure and aligned to a long axis of the cell, which was not oriented to the flow direction yet, but already elongated. 2. Stress analysis in a cell exposed to shear stress A fluid flow-structural analysis, i.e. a coupled field analysis, was performed to simulate three dimensional stress distribution in endothelial cells exposed to shear stress. The three-dimensional finite element model was generated on the basis of the cell surface geometry measured by an atomic force microscopy. The model consisted of a fluid element and a solid element representing the flow field and the endothelial cells, respectively. Analytical results on stress distribution in the cell showed that high compressive stress appeared both in the upstream side and the downstream side. These results may indicate that the stress distributions in the cells have close correlation with the F-actin distributions.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research
    Date (from‐to) : 1999 -2000 
    Author : MATSUMOTO Takeo, OHASHI Toshiro, SATO Masaaki
     
    To reveal the effects of mechanical factors on the initiation and progression of atherosclerosis, it is important to know the stress and strain distribution in the atherosclerotic lesions as well as the effects of blood flow on the endothelial cells. In this 2-year project, we experimentally induced atherosclerotic lesions in the rabbit thoracic aortas and studied the mechanical environment of the atherosclerotic lesions by measuring their local mechanical properties and local defomation of the aortas during inflation. We first measured local elastic modulus by a pipette aspiration method and studied the relationship between the modulus and the histology quantitatively to find that the atherosclerotic tissue became softer than the normal tissue at the initial stage of the lesion, then became stiffer with the appearance of the smooth muscle cells and calcified tissues. Next, we measured the deformation of the tubular specimen of the aortas during inflation as follows : We made four fine needles penetrate through the wall at an angular interval of about 90゜ as markers and observed the displacement of the needles during inflation from an end of the specimen. Local deformations of the wall were obtained at four circumferential segments held with the two adjacent needles. Strain referring to the stress-free state was obtained for each segment. The strain was not uniform among the segments in the atherosclerotic aorta. The difference among the segments tended to increase with the progression of the disease. Similar measurement performed for human coronary arteries indicated that the segment with thicker plaque had smaller strain. These results indicate that atherosclerotic arteries show marked heterogeneity. Local measurement is indispensable to reveal the mechanical environment of atherosclerotic lesions.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research
    Date (from‐to) : 1998 -2000 
    Author : HANE Kazuhiro, SASAKI Minoru, ESASHI Masayoshi
     
    We proposed a fabrication technique for sub-wavelength grating using electron-beam drawing machine and fast atom beam etching machine. On the surface of nonconductive optical glass, 150nm period surface gratings were fabricated. The etching of the nonconductive surface was carried out by using advantageously the neutral fast atom beam instead of ions in the conventional etching plasma. Without charging the surface, the deep etched profile with high aspect ratio was obtained. Using the proposed method, the reflection from the optical glass surface was suppressed considerably. A program for calculating the rigorous electro-magnetic field in sub-wavelength grating was developed. It was revealed that the reflection properties were influenced very much by the period and aspect ratio of the gratings. Using porous alumina membrane as a mask, the sub-wavelength gratings having 100nm period and the aspect ratio of 6 were fabricated on Si substrate. In the wavelength region from 370nm to 800nm, the superior properties for anti-reflection were obtained. Moreover, sub-wavelength gratings were also fabricated on the surface of the gallium arsenic light emission diode. Using the etching property of GaAs, tapered profile of surface grating was fabricated and a increase of the light emission was successfully obtained. In addition, the combinations between the surface gratings and micro-actuators were also investigated.
  • Cell structure and mechanobiology
    Date (from‐to) : 2000
  • 細胞の構造とメカノバイオロジーに関する研究
    Date (from‐to) : 2000
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research
    Date (from‐to) : 1998 -1999 
    Author : SATO Masaaki, HAYASHI Yoshiaki, OHASHI Toshiro, MATSUMOTO Takeo, KATAOKA Noriyuki
     
    The atomic force microscope (AFM) system was originally developed in combination with an inverted confocal laser scanning microscope to simultaneously measure mechanical stiffness and to observe microstructure of cultured cell. To detect an indentation depth of the cantilever, a cantilever moving system was introduced. A special specimen holder was made to hold a commercially available culture dish. The movements of the cantilever and the specimen holder were controlled by a personal computer. This system was applied to statically cultured and shear stress exposed endothelial cells, and the following results were obtained. 1. Observation of actin filaments and measurement of three dimensional configuration were performed for a fixed endothelial cell. 2. Input and output (I/O) ports of culture medium were newly designed to use a culture dish for flow exposure experiments. Morphology and mechanical properties of cultured endothelial cells were measured using the AFM system. Endothelial cells cultured at static condition had a polygonal shape and more soft mechanical properties around a nucleus than those at peripheral regions. The stiffness of the endothelial cells exposed to shear stress of 2 Pa became higher with the duration time of exposure. Cell shape became elongated to the flow direction and the location of a nucleus moved to downstream side by shear flow. 3. A fluid flow-structural analysis was performed. The three-dimensional finite element model was generated on the basis of the cell surface geometry measured by the AFM. The model consisted of a fluid element and a solid element representing the flow field and the endothelial cells, respectively. Analytical results on stress distribution in the cell showed that high compressive stress appeared mainly in the upstream side. This result may indicate that the stress distributions in the cells have close correlation with the F-actin distributions.
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research
    Date (from‐to) : 1998 -1999 
    Author : SATO Masaaki, SADAHIRO Mitsuaki, MATSUMOTO Takeo, TABAYASHI Koichi, AKIMOTO Hiroji, OHASHI Toshiro
     
    The aim of this project are to obtain mechanical properties of aneurysmal wall in vitro, to noninvasively measure the three dimensional shape of aneurysms for patients from MRI images, and to make models to predict the dangerous regions for aneurysm failure from the motion and stress analyses of aneurysmal wall. The following results were obtained. 1. The specimen excised form the aneurysmal region of patients was used for the biaxial tensile testing. The machine was newly developed in our laboratory. The diseased specimen from aneurysm was stiffer than normal tissues. The specimens with the higher elastic moduli were the higher anisotropic mechanical properties. 2. Three dimensional finite element model was successfully composed from MRI images for human thoracic aortas. In this model, the aortic wall was assumed to be thin membrane, isotropic, large linear deformation between diastolic and systolic pressures. The stress concentration was found in the frontal and dosal sides of aortic arch where aneurysm formation is frequently observed. Further, the location and shapes of the aneurysm had important and significant effects on the highest stress values and the stress distribution. 3. The strain and stress were calculated around the aneurysm of patients by measuring the deformation of aortic wall in MRI images. The results showed us that the strain was large around the frontal walls of aortic arch in healthy normal person. In aneurysmal patients, the large deformation to the outer wall side around the aneurysms was observed. According to the present results, we will develop the new system to rapidly construct the finite element model from the MRI images and study more precise mechanical properties of aneurysmal walls.
  • 文部科学省：科学研究費補助金(奨励研究(A))
    Date (from‐to) : 1998 -1999 
    Author : 大橋 俊朗
     
    近年,生物フォトンと呼ばれる超微弱光が注目されてきた.この生物フォトンが計測できれば新たな生体情報として非常に有用である.そこで,本研究では生物フォトンを効率よく検出するために微細加工技術を用いて微小なプローブを開発することを目的とする.この計測は近接場顕微鏡の原理を応用したもので,生体表面上のフォトンを微小プローブによって散乱させ,プローブ先端からフォトンを集めるものである.まずシリコンの微細加工技術により微小プローブを製作するために,有限要素法解析(FEM)により片持ち梁およびねじれ梁の振動特性を理論的に解析した.その結果,ねじれ梁は高次のモードを排除できるが片持ち梁の方が同程度の形状を有する場合,共振周波数は高い値を示した.次にフォトリソグラフィー技術により単結晶シリコンの微小プローブを製作した.製作した微小プローブの周波数特性はLC共振回路による容量センサおよび発信器から構成される実験装置によって測定を行ない,FEM解析結果とよい一致を示すことを確認した.この微小プローブを原子間力顕微鏡(AFM)のプローブとして用いれば細胞の表面形状とともにフォトンが検出できる.そこで第一段階として実際にウシ大動脈由来血管内皮細胞を用いて細胞表面形状を測定した.また,微弱光を検出するためのフォトダイオードの製作を併せて行なった.n型のシリコン基板にp型のボロンイオンを打ち込むことでp...
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research
    Date (from‐to) : 1997 -1999 
    Author : MATSUMOTO Takeo, KATAOKA Noriyuki, OHASHI Toshiro, SATO Masaaki
     
    In this 3-year study, we have developed a micro tensile tester to measure tensile properties of a single isolated cell and to observe change in intracellular structure during stretching simultaneously. Both ends of the cell were aspirated with glass micropipettes (3-15mm in I.D.) coated with cellular adhesive (Cell-Tak), and one pipette was moved with an electrical manipulator (MMS-77, Shimadzu) to stretch the cell under a microscope (BX50WI, Olympus). The force applied to the cell was measured by the deflection of a cantilever part of the other pipette. To obtain its clear image, the cell was observed with water immersion objectives under the upright microscope. To make cell manipulation easier, the stage of the microscope was fixed and the objectives were moved to adjust the focus. Bovine aortic smooth muscle cells (BASMs) obtained by the explant method (P6-7) and rat aortic smooth muscle cells (RASMs) obtained by the enzymatic dispersion method were successfully stretched with the tester under a physiological salt solution at 37℃ to obtain their tensile properties. Tensile tests of BASMs at various stretch rate show that initial elastic modulus of the cell have a significant positive correlation with the strain rate over 4 %/s. Elastic modulus of BASMs and RASMs were about 3 kPa and about 12kPa, respectively under the strain rate of 0.2-4 %/s. Due to the difference in the cell preparation method, BASMs and RASMs are considered to show synthetic and contractile phenotype, respectively. Contractile cells contain much more contractile proteins than synthetic cells. Contractile proteins may be stiffer than cytoplasm. Contractile cells may thus be stiffer than synthetic cells. The difference in the mechanical properties of BASMs and RASMs in the present study may be due to the difference in the phenotype of the two cell specimens.
  • 日本学術振興会：科学研究費助成事業
    Date (from‐to) : 1998 -1998 
    Author : 羽根 一博, 大橋 俊朗, 佐々木 実
     
    非伝播性のエバネッセント波から伝播性の光に変換するために、近接場顕微鏡においては尖ったファイバープローブや微細加工プローブが用いられる。よく用いられるファイバープローブに比較して、微細加工プローブの特徴はプローブに多機能を付加できるSiデバイスとの集積化が可能であることである。SiO_xN_y,Si_3N_4やSiO_2などの材料が従来、導波路材料として用いられている。一方、アモルファスシリコン(α-Si)は結晶Siデバイスとの製造互換性をもった興味ある材料であるが、導波路の材料としてはあまり用いられてこなかった。α-Siは0.82μm以上の波長の光に対して吸収が激減し、ほとんど透明になる。一方、結晶Siは1.1μm以上の赤外光において透明である。0.82μm-1.1μmの波長領域に対してα-Siは透明媒質として働き、結晶Siは吸収材料として働く。また、この領域には高出力GaAlAs半導体レーザの波長が含まれている。α-Siが高い屈折率(3.6)を持つことから微小領域に光を閉じこめることが期待でき、クラッドの材料として多くの選択が可能となる。これらのことより、本研究ではα-Siを用いて導波路型カンチレバーを製作した。 半導体レーザを全反射角でプリズムに入射し発生させたエバネッセント波により、プリズム表面に近付けたα-Siカンチレバーを照射し、その様子を光学顕微鏡により観察した。反りのためカンチレバー中を伝播する光がカンチレバーの中央から漏れ出ているのが見られた。広いカンチレバーの方が先端が三角型のカンチレバーよりも明るい光出力が得られた。α-Siカンチレバーからの光の放射を観測することで基本的な導波特性を確認した。次にSiフォトダイオードや半導体レーザなどとの集積化を試みる。
  • 細胞内力学伝達機構の解明
    Date (from‐to) : 1998
  • Japan Society for the Promotion of Science：Grants-in-Aid for Scientific Research
    Date (from‐to) : 1995 -1997 
    Author : SATO Masaaki, HELLUMS J David, GIRARD Peggy R, NEREM Robert M, KATAOKA Noriyuki, MATSUMOTO Takeo
     
    As this year was the last term of this project, we examined our results obtained for the past two years form global point of view, and decided to develop some new experimental methods to overcome methodological problems in the previous studies. 1. DEVELOPMENT OF A NEW TECHNIQUE TO OBSERVE ACTIN FILAMENTS IN ALIVE CELLS Immunofluorescence technique which has been utilized widely to observe intracellular actin filaments requires fixation of the cells, and it was vey difficult to observe actin filament dynamics in intact cells. To overcome this problem, we have introduced a combined gene of alpha-actinin, a bridging element of actin filaments and GFP (Green Fluorescent Protein) into vascular endothelial cells. Cells introduced such genes have autofluorescence in their alpha-actinin, and thus their actin filaments. We successfully observed actin filament dynamics in cells under static and shear-exposed conditions. 2. DEVELOPMENT OF A FLOW-LOADING SYSTEM FOR ENDOTHELIAL CELLS ON ARTERIAL INTRALUMINAL SURFACE There have been a lot of contradictions between results obtained from animal experiments and that based on cultured endothelial cells. To establish a new method compensating shortcomings of both experiments, we have developed a new flow-loading system to impose shear stress on the endothelial surface of an arterial segment, in which a piece of a cut-opened arterial segment is laid flat to expose its intraluminal surface to flow in a parallel plate chamber. We applied 1 Pa of shear stress to rabbit thoracic aortas perpendicularly to their vessel axis, and found that most of the endothelial cells changed their alignment parallel to the direction of flow in 72 hours. We concluded that this system can be widely used for the future studies on the mechanical response of vascular endothelial cells to shear stress.
  • 微細加工デバイスを用いた細胞機能の計測
    Date (from‐to) : 1997
  • 文部科学省：科学研究費補助金(奨励研究(A))
    Date (from‐to) : 1996 -1996 
    Author : 大橋 俊朗
     
    1.はじめに 血管壁の大変形時の弾性特性を知るための新たな測定法として、二軸引張装置により予めひずみを与えた状態でピペット吸引を行うことで試料の大変形時の弾性係数を推定する方法を提案した。また、内圧負荷試験を併せて行い両試験法による結果を比較・検討した。2.方法 ブタ胸部下行大動脈(外径約20mm、壁厚約2mm)から40x40mm程度の十字型試料を、主軸を血管壁の軸および円周方向に一致させながら十字型金型で打ち抜いた。試料を内腔面を上にして自作の二軸引張装置に固定し、周囲を室温の生理食塩水で満たした。ネジ送りステージを調節して軸および円周方向の伸び比を1.0〜1.5程度の範囲で0.1ずつ変化させながら、内径800μmのガラスピペットで試料表面を吸引した。ピペット内の試料の変形量はビデオマイクロスコープを通してモニタ上で二値化して求めた。一方、軸および円周方向の試料のひずみはCCDカメラで撮影し、試料表面の標線を二値化して求めた。弾性率Eは実験から得られた吸引圧-変形量関係を有限要素法解析から得られた同様の関係と比較することで求めた。また、別途比較のため同一個体の円筒状試料に対して従来法の内圧負荷試験を行い増分弾性係数H_<θθ>を求めた。3.結果および考察 軸方向伸長比λ_zが一定の場合、Eは円周方向伸長比λ_θが増加するに伴い単調に増加した。一方H_<θθ>は指数関数的に増...

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  • 局所弾性率測定装置
    特願平 11-185490

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