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Last Updated :2024/10/22

研究者情報

マスター

アカウント（マスター）

  • 氏名

    赤坂　司(アカサカ　ツカサ), アカサカ　ツカサ

所属（マスター）

  • 歯学研究院　口腔医学部門　口腔健康科学分野

所属（マスター）

  • 歯学研究院　口腔医学部門　口腔健康科学分野

独自項目

syllabus

  • 2021, 歯学研究概論, Research Basics in Dental Sciences, 博士後期課程, 歯学院, 歯学研究，基本的研究技法，基礎知識
  • 2021, 口腔生物学と医学, Oral Biology and Medicine, 博士後期課程, 歯学院, feeding behavior, vomiting, conditioned taste aversion (CTA), nausea, GLP-1, bacteriology, immunity, lipoprotein, salivary gland, oncogene, oral cancer, orofacial pain, tumor virus, dental materials, cementum, HIV, matrix cell biology
  • 2021, 人工生体機器研究, Developmental Research of Biomedical Devices and Artificial Organ, 博士後期課程, 歯学院, 代用臓器，バイオメカニクス，医用機器
  • 2021, 材料学的解析技法, Analysis of Dental and Biomaterials, 博士後期課程, 歯学院, 生体材料　物性評価　分析手法
  • 2021, 機能性生体材料学, Functional Biomaterials, 博士後期課程, 歯学院, 生体材料、分析化学
  • 2021, 物性歯科理工学, Functional Dental Materials and Engineering, 博士後期課程, 歯学院, 生体材料，コンポジットレジン，接着，機械的性質
  • 2021, 物性歯科理工学, Functional Dental Materials and Engineering, 博士後期課程, 歯学院, 共焦点レーザー走査型顕微鏡，原子間力顕微鏡，低真空走査型電子顕微鏡，Ｘ線走査型分析顕微鏡
  • 2021, 歯科生体材料開発研究, Developmental Research of Dental Biomaterials, 博士後期課程, 歯学院, 歯科材料，生体材料，生体適合性，ティッシュエンジニアリング，再生医学
  • 2021, 関連臨床医学, 学士課程, 歯学部, 関連臨床医学、耳鼻咽喉科学、精神医学、臨床心理学、眼科、皮膚科、産婦人科
  • 2021, 健康と社会, Health and Society, 学士課程, 全学教育, 口腔，血管，リンパ管，炎症，歯周組織，生体防御，常在細菌叢，う蝕（虫歯），歯周病，唾液, 口腔ガン，歯垢バイオフィルム
  • 2021, 健康と社会, Health and Society, 学士課程, 全学教育, 健康長寿　口腔科学　味覚　咀嚼　脳　摂食　嚥下　口腔がん　歯科インプラント　歯　歯周組織　加齢　むし歯　歯周病　老化　歯科材料　口腔細菌　義歯　喫煙
  • 2021, 健康と社会, Health and Society, 学士課程, 全学教育, 咬む（噛む），咀嚼，健康，心の健康　歯科
  • 2021, 歯科理工学実習, 学士課程, 歯学部, 歯科材料の性質、歯科材料の成形法、歯科用器械の取り扱い法
  • 2021, 臨床歯科材料学, 学士課程, 歯学部, 歯科材料、材料の組成・性質、生体反応、バイオマテリアル
  • 2021, 歯科理工学, 学士課程, 歯学部, 歯科材料、材料の組成・性質、生体反応、バイオマテリアル

researchmap

プロフィール情報

学位

  • 博士（工学）（名古屋大学）

プロフィール情報

対象リソース

業績リスト

研究キーワード

  • Cell culture   Micro/nano pattern   Nanoimprint   ナノインプリント   マイクロ・ナノパターン   歯科理工学   分化誘導   パターニング   スキャホールド   骨芽細胞   チタン   カーボンナノチューブ   糖鎖工学   Nanotechnology   Dental materials   

研究分野

  • ライフサイエンス / 生体材料学
  • ライフサイエンス / 生体医工学
  • ナノテク・材料 / ナノバイオサイエンス
  • ライフサイエンス / 口腔再生医学、歯科医用工学
  • ライフサイエンス / 補綴系歯学

経歴

  • 2011年09月 - 現在 北海道大学 歯学研究科(研究院) 准教授
  • 2011年09月 北海道大学
  • 2007年04月 - 2011年08月 - 北海道大学大学院歯科研究科 助教
  • 2007年04月 - 2011年08月 北海道大学
  • 2002年12月 - 2006年03月 北海道大学大学院歯科研究科 助手
  • 2002年12月 - 2006年03月 北海道大学
  • 2000年06月 - 2002年11月 株式会社生物有機化学研究所 研究員
  • 2000年06月 - 2002年11月 Chemical Biological Institute Researcher

学歴

  • 1997年04月 - 2000年03月   名古屋大学   工学研究科   物質制御工学
  •         - 2000年   名古屋大学
  • 1995年04月 - 1997年03月   北海道大学   大学院地球環境研究科
  • 1991年04月 - 1995年03月   明治大学   理工学部   工学化学科
  •         - 1995年   明治大学

委員歴

  • 2012年07月 - 現在   日本歯科理工学会   日本歯科理工学会 北海道・東北地方会役員 委員
  • 2012年04月 - 現在   日本歯科理工学会   研究グループ検討委員会 委員
  • 2009年02月 - 現在   ナノ・バイオメディカル学会   事務局長
  • 2011年10月 - 2016年03月   日本歯科理工学会   日本歯科理工学会 Dental Materials Journal編集委員

受賞

  • 2015年10月 日本歯科医学会 第31回「歯科医学を中心とした総合的な研究を推進する集い」優秀発表賞
     「歯表面での3D微細構造構築によるバイオミメティック次世代歯科治療へ向けて」 
    受賞者: 赤坂 司
  • 2013年10月 日本歯科理工学会 第６１回日本歯科理工学会 発表優秀賞
     「マイクロ・ナノ構造を持つパターン化バイオマテリアルの調製」 
    受賞者: 赤坂 司
  • 2008年12月 "Materials Science and Engineering C" Most Cited Author Award 2005-2008.
     Title: "Apatite formation on carbon nanotubes" T.Akasaka, F.Watari, Y.Sato, K.TohjiMaterials Science and Engineering C, 26(4), 675-678 (2006).
  • 2006年10月 日本歯科理工学会 第48回 日本歯科理工学会 研究奨励賞
     「人工複合糖質高分子によるStreptococcus mutansの増殖阻害」
  • 2006年 Molecular Biosystems Cover Gallery Winner 2005
  • 2004年10月 傾斜機能材料研究会 傾斜機能材料シンポジウム FGM奨励賞
     「糖鎖によるカーボンナノチューブの表面修飾」 
    受賞者: 赤坂 司

論文

  • Shuichi YAMAGATA, Takaaki YAMAMOTO, Yoshiaki SATO, Tsukasa AKASAKA, Yasuhiro YOSHIDA
    Dental Materials Journal 43 4 531 - 538 2024年07月25日
  • Ko Nakanishi, Tsukasa Akasaka, Hiroshi Hayashi, Kumiko Yoshihara, Teppei Nakamura, Mariko Nakamura, Bart Van Meerbeek, Yasuhiro Yoshida
    Materials (Basel, Switzerland) 17 15 2024年07月25日 
    Bioabsorbable materials have a wide range of applications, such as scaffolds for regenerative medicine and cell transplantation therapy and carriers for drug delivery systems. Therefore, although many researchers are conducting their research and development, few of them have been used in clinical practice. In addition, existing bioabsorbable materials cannot bind to the body's tissues. If bioabsorbable materials with an adhesive ability to biological tissues can be made, they can ensure the mixture remains fixed to the affected area when mixed with artificial bone or other materials. In addition, if the filling material in the bone defect is soft and uncured, resorption is rapid, which is advantageous for bone regeneration. In this paper, the development and process of a new bioabsorbable material "Phosphorylated pullulan" and its capability as a bone replacement material were demonstrated. Phosphorylated pullulan, which was developed based on the tooth adhesion theory, is the only bioabsorbable material able to adhere to bone and teeth. The phosphorylated pullulan and β-TCP mixture is a non-hardening putty. It is useful as a new resorbable bone replacement material with an adhesive ability for bone defects around implants.
  • Shintaro Nakano, Machiko Kasai, Keisuke Nakamura, Tsukasa Akasaka, Yasuhiro Yoshida, Shiho Suzuki, Yoichi Ohiro, Akira Hasebe
    Journal of Oral Biosciences 2024年05月
  • Kanako Nagamoto, Ko Nakanishi, Tsukasa Akasaka, Shigeaki Abe, Kumiko Yoshihara, Mariko Nakamura, Hiroshi Hayashi, Shinji Takemoto, Masato Tamura, Yoshimasa Kitagawa, Bart Van Meerbeek, Yasuhiro Yoshida
    Frontiers in bioengineering and biotechnology 12 1378039 - 1378039 2024年 
    Various implant surface treatment methods have been developed to achieve good osseointegration in implant treatment. However, some cases remain impossible to treat with implants because osseointegration is not obtained after implantation, and the implants fail. Thus, this study focused on phosphorylated pullulan because of its adhesiveness to titanium (Ti) and bone, high biocompatibility, and early replacement with bone. In this study, the response of bone-related cells to phosphorylated pullulan was evaluated to develop a new surface treatment method. Saos-2 (human osteosarcoma-derived osteoblast-like cells), MC3T3-E1 (mouse calvaria-derived osteoblast-like cells), and RAW264.7 (mouse macrophage-like cells) were used. In evaluating cellular responses, phosphorylated pullulan was added to the culture medium, and cell proliferation and calcification induction tests were performed. The proliferation and calcification of cells on the surface of Ti disks coated with phosphorylated pullulan were also evaluated. In addition, bone morphogenetic protein-2 (BMP-2), an osteogenic factor, was used to evaluate the role of phosphorylated pullulan as a drug carrier in inducing calcification on Ti disks. Phosphorylated pullulan tended to promote the proliferation of osteoblast-like cells and the formation of calcification on Ti disks coated with phosphorylated pullulan. Ti disks coated with phosphorylated pullulan loaded with BMP-2 enhanced calcification. Phosphorylated pullulan inhibited osteoclast-like cell formation. These results are due to the properties of phosphorylated pullulan, such as adhesiveness to titanium and drug-loading function. Therefore, phosphorylated pullulan effectively promotes bone regeneration when coated on titanium implants and is useful for developing a new surface treatment method.
  • 微細構造に応答した破骨細胞が作り出す多様なアクチン構造物
    赤坂 司, 中西 康, 吉田 靖弘
    日本歯科理工学会誌 42 Special Issue WINTER 5 - 5 (一社)日本歯科理工学会 2023年12月
  • 破骨細胞様細胞による凹凸基材に応答したアクチン構造物の形成
    赤坂 司, 宮治 裕史, 横山 敦郎, 吉田 靖弘
    日本バイオマテリアル学会大会予稿集 45回 422 - 422 日本バイオマテリアル学会 2023年10月
  • 破骨細胞様細胞による凹凸基材に応答したアクチン構造物の形成
    赤坂 司, 宮治 裕史, 横山 敦郎, 吉田 靖弘
    日本バイオマテリアル学会大会予稿集 45回 422 - 422 日本バイオマテリアル学会 2023年10月
  • Natsumi Ushijima, Tsukasa Akasaka, Shinichiro Numamoto, Tsubura Kudo, Atsuro Yokoyama
    Journal of oral biosciences 2023年06月29日 
    OBJECTIVES: We investigated the behavior of macrophages in the defined microtopography of materials. METHODS: Patterned cyclo-olefin polymer films were implanted into the femurs of seven-week-old rats. After 1 and 4 weeks, the rats were fixed with glutaraldehyde and OsO4, and their bones were observed with transmission electron microscopy (TEM). RESULTS: TEM and segmentation revealed an alternating structure in which multiple protrusions of adjacent macrophage-like cells overlapped. They were approximately 2 μm long and almost uniform in width, and were induced by the limited topography. CONCLUSION: New structures appeared between the macrophage-like cells as a result of microtopography.
  • Mohammed Katib Alruwaili, Tsutomu Sugaya, Yasuhito Morimoto, Ko Nakanishi, Tsukasa Akasaka, Yasuhiro Yoshida
    Dental materials journal 42 3 433 - 440 2023年04月07日 
    Recombinant human bone morphogenetic protein-2 (rhBMP-2) is one of the growth factors that may induce the formation of new bone. The aim was to determine the efficacy of low doses of rhBMP-2 for bone regeneration using a collagen sponge as a carrier. Three doses of rhBMP-2 (1.167, 0.117, and 0.039 mg/mL) were combined with an absorbable collagen sponge (ACS) as a delivery vehicle. The rhBMP-2/ACS implants were placed in the subcutaneous tissues of rat backs. X-ray microcomputed tomography (micro-CT) and histological analysis were used to evaluate bone formation. The samples treated with 1.167 mg/mL of rhBMP-2 showed greater bone formation than the samples treated with 0.117 mg/mL of rhBMP-2 four weeks after surgery. However, there was no evidence of bone formation in the samples that were treated with 0.039 mg/mL of rhBMP-2. It was found that rhBMP-2 was osteogenic even at one-tenth of its manufacturer's recommended concentration (1.167 mg/mL), indicating its potential for clinical use at lower concentrations.
  • 吉野 友都, 宮治 裕史, 西田 絵利香, 金本 佑生実, 浜本 朝子, 加藤 昭人, 菅谷 勉, 赤坂 司
    日本歯周病学会会誌 65 春季特別 133 - 133 (NPO)日本歯周病学会 2023年04月
  • 吉野 友都, 宮治 裕史, 西田 絵利香, 金本 佑生実, 浜本 朝子, 加藤 昭人, 菅谷 勉, 赤坂 司
    日本歯周病学会会誌 65 春季特別 133 - 133 (NPO)日本歯周病学会 2023年04月
  • 破骨細胞分化へのパターン材質と前駆細胞種の影響
    赤坂 司, 中西 康, 吉田 靖弘
    日本歯科理工学会誌 42 Special Issue81 78 - 78 (一社)日本歯科理工学会 2023年03月
  • Shuhei Otsubo, Ko Nakanishi, Kakufu Fukukawa, Ryoshun Endo, Seiichiro Yoshida, Aiko Matsumoto, Kumiko Yoshihara, Tsukasa Akasaka, Akira Hasebe, Yasuhiro Yoshida, Yoshiaki Sato
    Pharmaceutics 15 2 544 - 544 2023年02月06日 
    Although autopolymerizing resin offers numerous applications in orthodontic treatment, plaque tends to accumulate between the appliance and the mucosa, which increases the number of microorganisms present. In this study, we added cetylpyridinium chloride (CPC) loaded montmorillonite (Mont) and nanoporous silica (NPS) to autopolymerizing resin (resin-Mont, resin-NPS) and evaluated their drug release capacity, antimicrobial capacity, drug reuptake capacity, mechanical strength, and color tone for the devolvement of autopolymerizing resin with antimicrobial properties. As observed, resin-Mont and resin-NPS were capable of the sustained release of CPC for 14 d, and a higher amount of CPC was released compared to that of resin-CPC. Additionally, resin-Mont and resin-NPS could reuptake CPC. Moreover, the antimicrobial studies demonstrated that resin-Mont and resin-NPS could release effective amounts of CPC against Streptococcus mutans for 14 d and 7 d after reuptake, respectively. Compared to resin-CPC, resin-Mont exhibited a higher sustained release of CPC in all periods, both in the initial sustained release and after reuptake. However, the mechanical strength decreased with the addition of Mont and NPS, with a 36% reduction observed in flexural strength for resin-Mont and 25% for resin-NPS. The application of these results to the resin portion of the orthodontic appliances can prevent bacterial growth on the surface, as well as on the interior, of the appliances and mitigate the inflammation of the mucosa.
  • Yuto Yoshino, Hirofumi Miyaji, Erika Nishida, Yukimi Kanemoto, Asako Hamamoto, Akihito Kato, Tsutomu Sugaya, Tsukasa Akasaka
    Journal of oral biosciences 2023年01月17日 
    OBJECTIVES: Recombinant human collagen peptide (RCP) is a recombinantly created xeno-free biomaterial enriched in arginine-glycine-aspartic acid sequences with good processability whose use for regenerative medicine applications is under investigation. The biocompatibility and osteogenic ability of RCP granules combined with β-tricalcium phosphate (TCP) submicron particles (β-TCP/RCP) were recently demonstrated. In the present study, β-TCP/RCP was implanted into experimental periodontal tissue defects created in beagles to investigate its regenerative effects. METHODS: An RCP solution was lyophilized, granulated, and thermally cross-linked into particles approximately 1 mm in diameter. β-TCP dispersion (1 wt%; 500 μL) was added to 100 mg of RCP granules to form β-TCP/RCP. A three-walled intrabony defect (5 mm × 3 mm × 4 mm) was created on the mesial side of the mandibular first molar and filled with β-TCP/RCP. RESULTS: A micro-computed tomography image analysis performed at 8 weeks postoperative showed a significantly greater amount of new bone after β-TCP/RCP grafting (2.2-fold, P<0.05) than after no grafting. Histological findings showed that the transplanted β-TCP/RCP induced active bone-like tissue formation including tartaric acid-resistant acid phosphatase- and OCN-positive cells as well as bioabsorbability. Ankylosis did not occur, and periostin-positive periodontal ligament-like tissue formation was observed. Histological measurements performed at 8 weeks postoperative revealed that β-TCP/RCP implantation formed 1.7-fold more bone-like tissue and 2.1-fold more periodontal ligament-like tissue than the control condition and significantly suppressed gingival recession and epithelial downgrowth (P<0.05). CONCLUSIONS: β-TCP/RCP implantation promoted bone-like and periodontal ligament-like tissue formation, suggesting its efficacy as a periodontal tissue regenerative material.
  • 温度応答性歯科矯正用解体性接着材の開発
    春藤 彩花, 中西 康, 金子 知生, 黒川 考幸, 松本 愛子, 遠藤 諒俊, 赤坂 司, 吉田 靖弘, 佐藤 嘉晃
    日本矯正歯科学会大会プログラム・抄録集 81回 177 - 177 (公社)日本矯正歯科学会 2022年10月
  • Sayaka Nakamura, Hiroko Hoshi, Kaito Wakabayashi, Manami Seki, Makoto Watanabe, Momoka Watanabe, Hiroki Inaba, Natsumi Ushijima, Tsukasa Akasaka
    Journal of texture studies 53 5 654 - 661 2022年09月 [査読有り]
     
    Food texture is a very important factor for elderly persons, children, and patients who have difficulty swallowing. Collagen and its hydrolysis product, gelatin, are used as ingredients in foods, dietary supplements, and medical materials. In this study, we extracted atelocollagen from nonedible porcine tissues, including ear, nose, and skin, and analyzed the biophysical properties of each tissue. Extracted whole auricle collagen (AEC) showed superior springiness, while only the skin region of auricle collagen (ASC) showed superior hardness, springiness, and brittleness. Body skin collagen showed high hardness but low springiness. In a shear stress test, ASC gels showed high shear strength, and their strains coincided with hardness in a textural examination, while nose and AEC showed low maximum strains. In viscosity, the auricular collagens showed higher viscosity regardless of the region of the ear. Fibril formation in collagen from each tissue and organ varied a great deal in width and morphology. We found that the same type of collagen had a unique texture and viscosity under physiological conditions depending on the tissue or organ of extraction. The results show that the collagen extracted from each organ has a unique texture and unique possibilities to serve as an ingredient in food or supplements.
  • 金本 佑生実, 宮治 裕史, 西田 絵利香, 宮田 さほり, 眞弓 佳代子, 吉野 友都, 加藤 昭人, 菅谷 勉, 赤坂 司, 大矢根 綾子
    日本歯周病学会会誌 64 秋季特別 122 - 122 (NPO)日本歯周病学会 2022年08月
  • 宮治 裕史, 眞弓 佳代子, 金本 佑生実, 岡本 一絵, 浜本 朝子, 加藤 昭人, 菅谷 勉, 赤坂 司, 田中 佐織
    日本歯周病学会会誌 64 秋季特別 126 - 126 (NPO)日本歯周病学会 2022年08月
  • Hirofumi Miyaji, Kayoko Mayumi, Yukimi Kanemoto, Ichie Okamoto, Asako Hamamoto, Akihito Kato, Tsutomu Sugaya, Tsukasa Akasaka, Saori Tanaka
    Journal of oral biosciences 64 2 222 - 228 2022年06月 [査読有り]
     
    OBJECTIVES: Surface pre-reacted glass-ionomer (S-PRG) nanofiller, an antibacterial ion-releasing bioactive glass, has been shown to adhere to tooth surfaces and reported to improve inflammatory parameters in experimental periodontitis. In this study, cementum substrate was irrigated ultrasonically with dispersion to examine in-vitro nanofiller adhesion and antibacterial activity. Moreover, periodontal pockets in a beagle dog were ultrasonically irrigated with dispersion to assess periodontal healing. METHODS: The morphology of human cementum irrigated with S-PRG nanofiller dispersion was examined by scanning electron microscopy and energy dispersive X-ray spectrometry. The antibacterial activity of the treated cementum was tested using Actinomyces naeslundii. In addition, experimentally formed periodontal pockets in beagle dog were ultrasonically irrigated with S-PRG nanofiller dispersion. Periodontal parameters (gingival index, bleeding on probing, probing pocket depth, and clinical attachment level) were measured from baseline (0 weeks) through 12 weeks. Moreover, the effects of irrigation with S-PRG nanofiller on changes in periodontal microflora and bone healing were analyzed. RESULTS: After ultrasonic irrigation, S-PRG nanofiller adhered to the cementum and exhibited antibacterial activity. The periodontal parameters were shown to improve following ultrasonic irrigation with S-PRG nanofiller dispersion. Analysis by next-generation sequencing revealed that the ratio of red-complex species decreased in the pockets irrigated with S-PRG nanofiller dispersion. In addition, the S-PRG nanofiller showed the potential to promote bone healing. CONCLUSIONS: Ultrasonic irrigation with S-PRG nanofiller dispersion using an ultrasonic scaler system permitted delivery of the S-PRG nanofiller to the root surface, providing improved parameters in experimental periodontitis and modifying the composition of subgingival periodontal microflora.
  • Tsukasa Akasaka, Hiroshi Hayashi, Miho Tamai, Yoshitaka Yoshimura, Yoh-Ichi Tagawa, Hirofumi Miyaji, Ko Nakanishi, Yasuhiro Yoshida
    Journal of oral biosciences 64 2 237 - 244 2022年06月 [査読有り]
     
    OBJECTIVES: Osteoclasts can sense the surface topography of materials. However, it is difficult to identify the structural factors that affect osteoclast formation and its function. Furthermore, we hypothesized that the type of osteoclast precursor cells also affects osteoclastogenesis in the materials. In this study, we investigated the effects of defined micro/nanoscale patterns on osteoclastogenesis from bone marrow cells (BMCs). METHODS: Various cyclo-olefin polymer (COP) patterns were prepared using nanoimprinting. The effects of shape, size, and height of the patterns, and the wettability of the patterned surfaces on osteoclastogenesis from BMCs were evaluated in vitro. RESULTS: Osteoclast formation was promoted on pillars (diameter, 1 μm or 500 nm; height, 500 nm). Notably, osteoclastogenesis from BMCs was better promoted on hydrophobic pillars than on hydrophilic pillars. In contrast, decreased osteoclast formation was observed on the nanopillars (diameter, 100 nm; height, 200 nm). CONCLUSIONS: We demonstrated the promotion of osteoclast formation from BMCs on hydrophobic pillars with diameters of 1 μm and 500 nm. Some cellular behaviors in the patterns were dependent on the type of osteoclast precursor cells. The designed patterns are useful for designing the surface of dental implants or bone replacement materials with a controllable balance between osteoblast and osteoclast activities.
  • Tsukasa Akasaka, Miho Tamai, Yoshitaka Yoshimura, Natsumi Ushijima, Shinichiro Numamoto, Atsuro Yokoyama, Hirofumi Miyaji, Ryo Takata, Shuichi Yamagata, Yoshiaki Sato, Ko Nakanishi, Yasuhiro Yoshida
    NANO RESEARCH 2022年01月 [査読有り]
     
    The surface topography of a material can influence osteoclast activity. However, the surface structural factors that promote osteoclast activity have not yet been investigated in detail. Therefore, we investigated osteoclastogenesis by testing various defined patterns with different dimensions and shapes. The systematic patterns, made of a cyclo-olefin polymer, were prepared at a micron-, submicron-, and nano-scale with a groove, hole, or pillar shape with a 1:1 pitch ratio. RAVV264.7 cells were cultured on these patterns in the presence of the receptor activator of NF-kappa B ligand (RANKL). Osteoclast formation was induced in the order: pillar > groove >= hole. The two-dimensional factors also indicated that submicron-sized patterns strongly induced osteoclast formation. The optimal pillar dimension for osteoclast formation was 500 nm in diameter and 2 mu m in height Furthermore, we observed two types of characteristic actin structure, i.e., belt-like structures with small hollow circles and isolated ring-like structures, which formed on or around the pillars depending on size and height. Furthermore, resorption pits were observed mainly on the top of calcium phosphate-coated pillars. Thus, osteoclasts prefer convex shapes, such as pillars for differentiation and resorption. Our results indicate that osteoclastogenesis can be controlled by designing surfaces with specific morphologies.
  • Yukimi Kanemoto, Hirofumi Miyaji, Erika Nishida, Saori Miyata, Kayoko Mayumi, Yuto Yoshino, Akihito Kato, Tsutomu Sugaya, Tsukasa Akasaka, Arputharaj Joseph Nathanael, Syama Santhakumar, Ayako Oyane
    Journal of periodontal research 57 1 205 - 218 2021年11月16日 [査読有り]
     
    BACKGROUND AND OBJECTIVES: In the treatment of severe periodontal destruction, there is a strong demand for advanced scaffolds that can regenerate periodontal tissues with adequate quality and quantity. Recently, we developed a plasma- and precursor-assisted biomimetic process by which a porous collagen scaffold (CS) could be coated with low-crystalline apatite. The apatite-coated collagen scaffold (Ap-CS) promotes cellular ingrowth within the scaffold compared to CS in rat subcutaneous tissue. In the present study, the osteogenic activity of Ap-CS was characterized by cell culture and rat skull augmentation tests. In addition, the periodontal tissue reconstruction with Ap-CS in a beagle dog was compared to that with CS. METHODS: The plasma- and precursor-assisted biomimetic process was applied to CS to obtain Ap-CS with a low-crystalline apatite coating. The effects of apatite coating on the scaffold characteristics (i.e., surface morphology, water absorption, Ca release, protein adsorption, and enzymatic degradation resistance) were assessed. Cyto-compatibility and the osteogenic properties of Ap-CS and CS were assessed in vitro using preosteoblastic MC3T3-E1 cells. In addition, we performed in vivo studies to evaluate bone augmentation and periodontal tissue reconstruction with Ap-CS and CS in a rat skull and canine furcation lesion, respectively. RESULTS: As previously reported, the plasma- and precursor-assisted biomimetic process generated a low-crystalline apatite layer with a nanoporous structure that uniformly covered the Ap-CS surface. Ap-CS showed significantly higher water absorption, Ca release, lysozyme adsorption, and collagenase resistance than CS. Cell culture experiments revealed that Ap-CS was superior to CS in promoting the osteoblastic differentiation of MC3T3-E1 cells while suppressing their proliferation. Additionally, Ap-CS significantly promoted (compared to CS) the augmentation of the rat skull bone and showed the potential to regenerate alveolar bone in a dog furcation defect. CONCLUSION: Ap-CS fabricated by the plasma- and precursor-assisted biomimetic process provided superior promotion of osteogenic differentiation and bone neoformation compared to CS.
  • 岡本 一絵, 宮治 裕史, 宮田 さほり, 菅谷 勉, 蔀 佳奈子, 赤坂 司, 齋田 慧, 川崎 英也
    日本歯周病学会会誌 63 秋季特別 133 - 133 (NPO)日本歯周病学会 2021年10月 [査読無し]
  • 眞弓 佳代子, 宮治 裕史, 宮田 さほり, 西田 絵利香, 降旗 友和, 金本 佑生実, 菅谷 勉, 蔀 佳奈子, 赤坂 司
    日本歯周病学会会誌 63 秋季特別 133 - 133 (NPO)日本歯周病学会 2021年10月 [査読無し]
  • R. Endo, K. Nakanishi, Y. Bando, S. Abe, H. Maruoka, M. Nakamura, T. Akasaka, Y. Yoshida, Y. Sato
    Materials 14 19 5742 - 5742 2021年10月01日 [査読有り]
     
    This study prepared glass ionomer cement (GIC) containing nanoporous silica (NPS) (GIC–NPS) at 5 wt% concentrations using 3 types of NPS with different pore and particle sizes and evaluated the differences in their cationic ion capture/release abilities and mechanical properties. The cationic water-soluble dye was used as cationic ion. The test GIC–NPS complexes captured dyes by immersion in 1 wt% dye solutions. All the GIC–NPS complexes released dyes for 28 d, and the amount of dye released from the complexes increased with decreasing pore size; however, the particle size of NPS did not affect the amount of dye released. Additionally, GIC–NPS was able to recharge the dye, and the amount of released the dye by the complexes after recharge was almost identical to the amount released on the first charge. Although not significantly different, the compressive strength of GIC–NPS was slightly greater than that of GIC without NPS regardless of the type of NPS. These results suggest that the degree of capture and release of cationic molecules, such as drugs, can be controlled by optimizing the pore size of NPS without sacrificing its mechanical strength when its content is 5 wt%.
  • Ichie Okamoto, Hirofumi Miyaji, Saori Miyata, Kanako Shitomi, Tsutomu Sugaya, Natsumi Ushijima, Tsukasa Akasaka, Satoshi Enya, Satoshi Saita, Hideya Kawasaki
    ACS OMEGA 6 13 9279 - 9290 2021年04月 [査読有り]
     
    Antibacterial photodynamic therapy (aPDT) utilizes reactive oxygen species such as singlet oxygen (O-1(2)) and free radicals via photosensitizers, which are light and light-sensitive agents, to reduce bacterial infections. It has been utilized as a treatment for dental diseases in place of antibiotic therapies. However, aPDT does not always cause the desired therapeutic effect due to the instability of organic photosensitizers and the formation of bacterial biofilms. To promote the antibacterial and antibiofilm effects of aPDT, we have proposed a lysozyme (Lys)-gold nanoclusters (Au NCs)/rose bengal (Lys-Au NCs/RB) conjugate as a novel photosensitizer. This conjugate was found to effectively impede the growth of both gram-positive and gram-negative bacteria when exposed to white light-emitting diode (LED) irradiation. The photoexcited Lys-Au NCs/RB showed significantly higher antibacterial activity than photoexcited Lys-Au NCs or RB alone. The synergistic effect is a result of the combination of Lys (an antibacterial protein) and enhanced O-1(2) generation related to resonance energy transfer (RET) in the Au NCs/RB conjugate. Photoexcited Lys-Au NCs/RB increased the effects of aPDT in a dose- and time-dependent manner. Furthermore, the photoexcited Lys-Au NCs/RB successfully decreased Streptococcus mutans biofilm formation. However, in contrast, it did not have a negative effect on the proliferation, adhesion, or spread of mammalian cells, indicating low cytotoxicity. Lys-Au NCs/RB is a novel photosensitizer with low cytotoxicity that is capable of bacterial inactivation and the suppression of biofilm formation, and could help to improve dental treatments in the future.
  • Kayoko Mayumi, Hirofumi Miyaji, Saori Miyata, Erika Nishida, Tomokazu Furihata, Yukimi Kanemoto, Tsutomu Sugaya, Kanako Shitomi, Tsukasa Akasaka
    HELIYON 7 2 2021年02月 [査読有り]
     
    Objectives: Surface pre-reacted glass-ionomer (S-PRG) fillers release antibacterial borate and fluoride ions. We fabricated nanoscale S-PRG fillers (S-PRG nanofillers) for antibacterial coating of tooth surfaces and assessed the antibacterial effects of this coating in vitro. In addition, we creating a canine model of periodontitis to evaluate the effectiveness of S-PRG nanofiller application on tooth roots and improvement of periodontal parameters.Methods: Human dentin blocks were coated with S-PRG nanofiller (average particle size: 0.48 mu m) and then characterized by scanning electron microscopy (SEM), energy dispersive X-ray spectrometer (EDX), and ion-releasing test. Antibacterial effects of dentin blocks coated with S-PRG nanofiller were examined using bacterial strains, Streptococcus mutans and Actinomyces naeslundii. Next, we created an experimental model of periodontitisin furcation of premolars of beagle dogs. Then, S-PRG nanofiller coating was applied onto exposed tooth root surfaces. Periodontal parameters, gingival index (GI), bleeding on probing (BOP), probing pocket depth (PPD), and clinical attachment level (CAL), were measured from baseline until 4 weeks. In addition, bone healing was radiographically and histologically examined.Results: SEM and EDX revealed that S-PRG nanofillers uniformly covered the dentin surface after coating. Dentin blocks coated with S-PRG nanofiller showed ion-releasing property, bacterial growth inhibition, and sterilization effects. In the experimental periodontitis model, S-PRG nanofiller coating significantly reduced clinical inflammatory parameters, such as GI (P < 0.01) and BOP (P < 0.05), compared to uncoated samples. In addition, PPD and CAL significantly decreased by S-PRG nanofiller coating (2 weeks: P < 0.05; 3 and 4 weeks: P < 0.01), suggesting the improvement of periodontitis. Micro-CT and histology revealed that bone healing of furcation defects was enhanced by S-PRG nanofiller coating.Conclusion: S-PRG nanofiller coating provides antibacterial effects to tooth surfaces and improves clinical parameters of periodontitis.
  • Erika Nishida, Hirofumi Miyaji, Kanako Shitomi, Tsutomu Sugaya, Tsukasa Akasaka
    DENTAL MATERIALS JOURNAL 40 1 157 - 164 2021年02月 [査読有り]
     
    Zinc-fluoride glass nanoparticles (Zinc-F) release several ions, such as fluoride, zinc and calcium ions, through acid-base reactions. The aim of this study was to evaluate the antibacterial and cytotoxic properties of Zinc-F. Antibacterial tests showed that a Zinc-F eluting solution significantly reduced the turbidity and colony-forming units of Streptococcus mutans and Actinomyces naeslundii, compared to that of calcium-fluoroaluminosilicate glass nanoparticles without zinc ions. In live/dead staining, Zinc-F eluate significantly decreased green-stained bacterial cells, indicating live cells, compared with the control (no application). Human dentin coated with Zinc-F showed suppressed S. mutans and A. naeslundii biofilm formation. Additionally, Zinc-F eluate showed low cytotoxic effects in osteoblastic and fibroblastic cells. Therefore, our findings suggested that Zinc-F exhibits antibacterial and biocompatible properties through multiple-ion release.
  • Tomokazu Furihata, Hirofumi Miyaji, Erika Nishida, Akihito Kato, Saori Miyata, Kanako Shitomi, Kayoko Mayumi, Yukimi Kanemoto, Tsutomu Sugaya, Tsukasa Akasaka
    JOURNAL OF BIOMEDICAL MATERIALS RESEARCH PART B-APPLIED BIOMATERIALS 108 7 3033 - 3044 2020年05月 [査読有り][通常論文]
     
    Recombinant human collagen peptide, developed based on human collagen type I, contains an arginyl-glycyl-aspartic acid (RGD)-rich motif to enhance cell behavior and is anticipated as a xeno-free polymer material for use in tissue engineering. We fabricated granules containing recombinant human collagen peptide (RCP) applied with beta-tricalcium phosphate fine particles (RCP/beta-TCP) as bone filling scaffold material and assessed the bone forming ability of RCP/beta-TCP. Recombinant peptide was thermal crosslinked and freeze-dried to prepare RCP. An aqueous dispersion of beta-TCP fine particles was added to RCP to obtain RCP/beta-TCP. Subsequently, RCP/beta-TCP were characterized using scanning electron microscopy (SEM), energy dispersive X-ray spectrometry (EDX), and cell culture assessments. Furthermore, RCP/beta-TCP were implanted into rat cranial bone defects for radiographic and histological evaluations. In SEM and EDX analyses of RCP/beta-TCP, beta-TCP particles dose-dependently covered the surface of RCP. Cell culture tests showed that RCP/beta-TCP remarkably promoted proliferation and mRNA expression of various genes, such as integrin beta 1 and osteogenic markers, of osteoblastic MC3T3-E1 cells. Histomorphometric assessment at 4 weeks showed that RCP/beta-TCP significantly promoted new skull bone formation compared to RCP (p < 0.05) and control (no application) (p < 0.01). Accordingly, these findings suggest RCP/beta-TCP possess bone forming capability and would be beneficial for bone tissue engineering therapy.
  • Oguma H, Seitoku E, Mutoh M, Yoshizawa S, Nakanishi K, Bando Y, Era Y, Kiba T, Saikaew P, Tamai M, Akasaka T, Nakamura M, Kusaka T, Yoshida Y, Sato Y, Sano H, Abe S, Alireza V, Watanabe I, Inoue S, Takada T
    Journal of nanoscience and nanotechnology 20 5 2668 - 2674 2020年05月 [査読有り][通常論文]
     
    In this study, we investigated water-dispersible surface modification for size- and shape-controlled fullerene nanoparticles (C60P) based on a condensation reaction with di-amino alkane. This modification provided for water dispersibility of C60P and the capability for secondary modification as well. The resultant C60P particles have several useful physical properties: water-dispersibility for ease of injection; fluorescence for detection and quantification; and a characteristic morphology to assist identification. These properties will widely extend the applications of these particles, especially into the biological fields of bioimaging and drug delivery.
  • Shitomi K, Miyaji H, Miyata S, Sugaya T, Ushijima N, Akasaka T, Kawasaki H
    Photodiagnosis and photodynamic therapy 30 101647 - 101647 2020年01月02日 [査読有り][通常論文]
     
    Antibacterial photodynamic therapy (a-PDT) is a promising anti-infective technique for generation of singlet oxygen (1O2) to target dental disease. However, conventional organic photosensitizers have some problems for clinical use in terms of cytotoxicity, quenching of a-PDT activity by self-dimerization, and the lack of long-term antibacterial effect. We herein propose silver nanoclusters/rose bengal nanocomposite (AgNCs/RB) as a novel photosensitizer with two primary antibacterial effects: (1) 1O2 generation by irradiated RB and (2) Ag+ ion release from AgNCs. AgNCs/RB irradiated with white light-emitting diode (LED) for a short irradiation time of 1 min significantly decreased the bacterial turbidity of Streptococcus mutans, Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans (P < 0.05). In SEM, TEM and LIVE/DEAD staining images, photoexcited AgNCs/RB reduced S. mutans colonization, destroyed the cell membrane, and increased the number of dead cells. The antibacterial efficiency of photoexcited AgNCs/RB was greater than that of AgNCs or RB alone (P < 0.05), suggesting a synergistic effect of 1O2 and Ag + ions from photoexcited AgNCs/RB. By contrast, photoexcited AgNCs/RB did not affect WST-8 and LDH activities and morphology of NIH3T3 mammalian cells, indicating low cytotoxicity. Interestingly, the antibacterial activity of AgNCs/RB on S. mutans was maintained even after the cessation of LED irradiation, indicating a long-term antibacterial effect due to released Ag+ ions. The present AgNCs/RB photosensitizers provide effective synergistic antibacterial effects for dental a-PDT via 1O2 and Ag+ ions coupled with low cytotoxicity.
  • Shuichi Yamagata, Takaaki Yamamoto, Yoshiaki Sato, Natsumi Ushijima, Tsukasa Akasaka, Yasuhiro Yoshida
    Nano Biomedicine 11 2 57 - 64 2020年 [査読有り]
     
    After orthodontic treatment has been completed, we consider fluorescent imaging to be one of the most effective solutions for secure removal of colorless residual adhesives from tooth surfaces using cutting instruments. Nanoscale Y2O3:Eu3+ particles were synthesized using the homogeneous precipitation method with two different starting concentrations of urea aqueous solutions, followed by firing at 1000°C. These particles exhibited narrow size distribution (approx. 200–300 nm) and sharp crystallinity regardless of the urea concentration. Moreover, their photoluminescence peak corresponded well with the typical 4f-4f transitions of Eu3+. The Y2O3:Eu3+ particles were almost uniformly dispersed and retained in the monomer blends and polymerized bulk bodies. Photoluminescence measurement is a valid detection method and can be useful for future studies on dispersion control. We conclude that the crystalline Y2O3:Eu3+ particles could be applicable for further development of fluorescent orthodontic adhesives.
  • Shuichi Yamagata, Takaaki Yamamoto, Yoshiaki Sato, Natsumi Ushijima, Tsukasa Akasaka, Yasuhiro Yoshida
    Nano Biomedicine 12 1 19 - 26 2020年 [査読有り]
     
    Secure removal of remnants on tooth surfaces by rotary cutting instruments after an orthodontic treatment could be guaranteed if the orthodontic adhesive used is highly visible. Fluorescent imaging is a potentially effective solution for increasing the visibility of orthodontic adhesives. In this study, Y2O3:Eu3+ nanoparticles were synthesized by a homogeneous precipitation method followed by firing at 1,000 °C. The size of most of the synthesized nanoparticles was approximately 200 nm in diameter. The photoluminescence properties of the nanoparticles were similar to that of the typical 4f-4f transition of Eu3+. Regardless of the concentration of the particle, the nanoparticles were almost uniformly dispersed and remained in their polymerized bodies without deteriorating the intrinsic flexural modulus of the dimethacrylate resin. Although the fluorescence of the 10 wt%-phosphor-containing resins was adequately recognizable to the naked eye, it was suggested that the preferred concentration of Y2O3:Eu3+ nanoparticles should be more than 10 wt% in terms of fluorescence intensity.
  • Kaga N, Akasaka T, Matsuura T, Yokoyama A, Yoshida Y
    Bio-medical materials and engineering 30 5-6 559 - 567 2020年01月 [査読有り][通常論文]
     
    BACKGROUND: Surface nanostructures in titanium (Ti) oral implants are critical for rapid osseointegration.OBJECTIVE: The purpose of this study was to evaluate the growth of osteoblast-like (Saos-2) and epithelial-like (Ca9-22) cells on nanopatterned Ti films.METHODS: Ti films with 500 nm grooves and pillars were fabricated by nanoimprinting, and seeded with Saos-2 and Ca9-22 cells. Cell viability and morphology were assessed by cell proliferation assay and scanning electron microscopy, respectively.RESULTS: As assessed after 1 hour, proliferation of Saos-2 cells was most robust on grooved films than on pillared and smooth films, in this order. These cells approximately doubled on grooved and pillared substrates in 24 hours and after 5 days, but not on smooth surfaces. In contrast, Ca9-22 cells favored smooth surfaces, followed by grooved and pillared films. Indeed, cells sparsely adhered to pillared films over 5 days of incubation (p < 0.05).CONCLUSIONS: The data show that Saos-2 and Ca9-22 cells respond differently to different nanostructures, and highlight the potential use of nanopatterns to promote bone regeneration or to prevent epithelial downgrowth at the implant-bone interface.
  • Miyaji H, Mayumi K, Miyata S, Nishida E, Shitomi K, Hamamoto A, Tanaka S, Akasaka T
    Dental materials journal 39 2 287 - 294 2019年11月28日 [査読有り][通常論文]
     
    Surface pre-reacted glass-ionomer (S-PRG) filler releases several ions, such as fluoride, borate and strontium ions, to exert bioactive effects. We fabricated an endodontic root canal sealer containing S-PRG fillers (S-PRG sealer) and then evaluated the antibacterial and anti-inflammatory properties of S-PRG sealer compared with sealer containing conventional silica fillers (silica sealer). Antibacterial tests showed that S-PRG sealer significantly reduced the turbidity of Enterococcus faecalis compared with silica sealer. Implantation of S-PRG or silica sealer blocks in rat subcutaneous tissue showed that S-PRG sealer decreased the proinflammatory response compared with silica sealer at 10 days post-implantation. In addition, immunostaining revealed that infiltration of CD68- and peroxidase-positive cells around the S-PRG sealer was significantly lower than that in silica sealer. Therefore, it was suggested that S-PRG sealer exhibits antibacterial and anti-inflammatory effects.
  • Fusion of RAW 264.7 macrophage cells on micro-scale fine pillar patterns
    J. Honma, T. Akasaka, M. Tamai, Y. Yoshimura, T. Taira, H. Miyaji, S. Yamagata, Y. Sato, Y. Yoshida
    Digest Journal of Nanomaterials and Biostructures 13 4 1123 - 1131 2018年12月 [査読有り][通常論文]
  • Aiko MATSUMOTO, Shuichi YAMAGATA, Shigeaki ABE, Tsukasa AKASAKA, Yasuhiro YOSHIDA, Junichiro IIDA
    Nano Biomedicine 10 1 3 - 8 2018年08月 [査読有り][通常論文]
  • Hirofumi MIYAJI, Shusuke MURAKAMI, Erika NISHIDA, Tsukasa AKASAKA, Bunshi FUGETSU, Junko UMEDA, Katsuyoshi KONDOH, Tadashi IIZUKA, Tsutomu SUGAYA
    Journal of Oral Tissue Engineering 15 3 123 - 130 2018年05月 [査読有り][通常論文]
  • R. Takata, T. Akasaka, M. Tamai, Y. Yoshimura, T. Taira, H. Miyaji, Y. Tagawa, S. Yamagata, J. Iida, Y. Yoshida
    Digest Journal of Nanomaterials and Biostructures 13 2 451 - 458 2018年04月 [査読有り][通常論文]
  • Nagao Keishi, Miyaji Hirofumi, Nishida Erika, Akasaka Tsukasa, Miyata Saori, Shitomi Kanako, Mayumi Kayoko, Kato Akihito, Sugaya Tsutomu
    Journal of Oral Hygiene & Health 6 1 231 - 231 OMICS International 2018年01月10日 [査読有り][通常論文]
  • Shuichi Yamagata, Takaaki Yamamoto, Yoshiaki Sato, Shigeaki Abe, Tsukasa Akasaka, Yasuhiro Yoshida
    Nano Biomedicine 10 2 53 - 60 2018年 [査読有り]
     
    We successfully prepared light-curing type fluorescent resins as orthodontic adhesives doped with tris(1,3-diphenyl-1,3-propanedionato)(1,10-phenanthroline)Eu(III) [Eu(DBM)3Phen] by a photoinduced radical polymerization method. The transparent and almost colorless appearance of the composites under natural light is favorable for an esthetic dental product. The strong luminescence under near-UV light, especially when the content of Eu(DBM)3 Phen is above 0.1 wt%, is promising for safe and complete removal of this adhesive after completing orthodontic treatments. We have also found that concentration quenching may occur around a concentration of 0.2-0.4 wt%. Based on the results obtained, in situ monomer polymerization can be applicable to Eu(DBM)3Phen-containing adhesives. Although we are still investigating this proposed system, we conclude that the Eu(DBM)3Phen-containing adhesives is a promising candidate for application to fluorescent orthodontic adhesives.
  • Shuichi Yamagata, Takaaki Yamamoto, Yoshiaki Sato, Natsumi Ushijima, Shigeaki Abe, Tsukasa Akasaka, Yasuhiro Yoshida, Junichiro Iida
    Nano Biomedicine 9 1 15 - 22 2018年 [査読有り][通常論文]
     
    Subsequent to the orthodontic treatment, removal of residual colorless adhesives from tooth surfaces by cutting instruments with the lowest possible iatrogenic damage has been a long-standing challenge. Fluorescent imaging can be one of the most effective solutions. Y2O3:Eu3+ particles were synthesized by a homogeneous precipitation method and the subsequent calcination was performed at various temperatures. The particles had a narrow size distribution (200-300 nm) and showed sharp crystallinity independent of the calcination temperatures above 800 °C. In this study, 1,000 °C is confirmed to be the optimum calcination temperature. Although the reason for the decline in emission intensity of the 1,100 °C-calcined particles is still unclear, loss of spherical shape associated with excessive crystal grain growth and simultaneous sintering to each other by firing is a possible explanation. The emission from the phosphor-containing resins, intended for use in orthodontic adhesives, is appreciable although the emission intensity of the resins was weak even if their content was 20 wt.%. The decrease in emission intensity may be attributed to the scattering by the particles or cross relaxation processes. We conclude that the crystalline Y2O3:Eu3+ particles calcined at 1,000 °C could be applicable for further development of photoluminescent orthodontic adhesives.
  • Reika Makita, Tsukasa Akasaka, Seiichi Tamagawa, Yasuhiro Yoshida, Saori Miyata, Hirofumi Miyaji, Tsutomu Sugaya
    Beilstein journal of nanotechnology 9 1735 - 1754 2018年 [査読有り][通常論文]
     
    Background: Collagen is a basic component of the periodontium and plays an important role in the function of the periodontal unit. Therefore, coating with collagen/gelatin has been applied to enable dental implants to positively interact with peri-implant tissues. Although the micro/nanoscale topography is an important property of the surface of dental implants, smaller collagen/gelatin surface patterns have not been sufficiently developed. Furthermore, only few reports on the behavior of cells on gelatin surfaces with different patterns and sizes exist. In this study, we developed micro/nanometer-scaled gelatin surfaces using genipin crosslinking, with the aim of understanding the use of patterning in surface modification of dental implants. Results: Grooves, holes, and pillars, with widths or diameters of 2 µm, 1 µm, or 500 nm were fabricated using a combination of molding and genipin crosslinking of gelatin. The stability of the different gelatin patterns could be controlled by the degree of genipin crosslinking. The gelatin patterns at 20 mM concentration of genipin and 41% crosslinking maintained a stable, patterned shape for at least 14 days in a cell culture medium. A cell morphology study showed that the cells on groves were aligned along the direction of the grooves. In contrast, the cells on pillars and holes exhibited randomly elongated filopodia. The vinculin spots of the cells were observed on the top of ridges and pillars or the upper surface of holes. The results of a cell attachment assay showed that the number of surface-attached cells increased with increasing patterning of the gelatin surface. Unlike the cell attachment assay, the results of a cell proliferation assay showed that Saos-2 cells prefer grooves with diameters of approximately 2 µm and 1 µm and pillars with diameters of 1 µm and heights of 500 nm. The number of cells on pillars with heights of 2 µm was larger than those of the other gelatin surface patterns tested. Conclusion: These data support that a detailed design of the gelatin surface pattern can control both cell attachment and proliferation of Saos-2 cells. Thus, gelatin surfaces patterned using genipin crosslinking are now an available option for biocompatible material patterning.
  • Shusuke Murakami, Hirofumi Miyaji, Erika Nishida, Kohei Kawamoto, Saori Miyata, Hiroko Takita, Tsukasa Akasaka, Bunshi Fugetsu, Toshihiko Iwanaga, Hiromi Hongo, Norio Amizuka, Tsutomu Sugaya, Masamitsu Kawanami
    Dental materials journal 36 5 573 - 583 2017年09月26日 [査読有り][通常論文]
     
    Three-dimensional collagen scaffolds coated with beta-tricalcium phosphate (β-TCP) nanoparticles reportedly exhibit good bioactivity and biodegradability. Dose effects of β-TCP nanoparticles on biocompatibility and bone forming ability were then examined. Collagen scaffold was applied with 1, 5, 10, and 25 wt% β-TCP nanoparticle dispersion and designated TCP1, TCP5, TCP10, and TCP25, respectively. Compressive strength, calcium ion release and enzyme resistance of scaffolds with β-TCP nanoparticles applied increased with β-TCP dose. TCP5 showed excellent cell-ingrowth behavior in rat subcutaneous tissue. When TCP10 was applied, osteoblastic cell proliferation and rat cranial bone augmentation were greater than for any other scaffold. The bone area of TCP10 was 7.7-fold greater than that of non-treated scaffold. In contrast, TCP25 consistently exhibited adverse biological effects. These results suggest that the application dose of β-TCP nanoparticles affects the scaffold bioproperties; consequently, the bone conductive ability of TCP10 was remarkable.
  • Saori Miyata, Hirofumi Miyaji, Hideya Kawasaki, Erika Nishida, Kanako Shitomi, Tsukasa Akasaka, Saori Tanaka, Tadashi Iizuka, Tsutomu Sugaya
    Biology, Engineering and Medicine 2 4 1 - 4 Open Access Text 2017年09月 [査読有り][通常論文]
     
    The development of photosensitizers and light sources has enabled the use of antimicrobial photodynamic therapy (aPDT) in various dental therapies. In the presentstudy, we compared the antibacterial and cytotoxic effects of Au clusters photoexcited by blue and white LED irradiation. We fabricated novel photosensitizers,captopril-protected gold (Capt-Au) clusters and lysozyme-stabilized gold (Lyz-Au) clusters, for aPDT. Au clusters were then photoexcited by two kinds of lightsources, blue high-power and white low-power light-emitting diodes (LEDs). Since white LED contains a wide spectrum of light (400–750 nm), white LED wouldbe relevant for aPDT even if using a low-power source.The turbidity and viability of Streptococcus mutans were assessed following application of Capt-Au clusters (500 μg/mL) or Lyz-Au clusters (1,000 μg/mL) photoexcitedby a blue high-power LED (1,000 mW/cm2) or white low-power LED (80 mW/cm2). In addition, the cytotoxicity of Au clusters and LED irradiation was evaluatedin NIH3T3 and MC3T3-E1 cells.Au clusters photoexcited by the white low-power LED equally decreased the turbidity and viability of S. mutans compared with blue high-power LED. However,Au clusters photoexcited by white LED irradiation caused decreased cytotoxicity in mammalian cells compared with those photoexcited by blue LED irradiation. Inconclusion, white LEDs possess biosafe properties for aPDT using Au clusters.
  • Yoshida, Yasuhiro, Inoue, Satoshi, Tamai, Miho, Abe, Shigeaki, Akasaka, Tsukasa, Yoshihara, Kumiko, Meerbeek, Bart Van
    Hokkaido Journal of Dental Science 38 86 - 98 北海道歯学会 2017年09月 [査読有り][通常論文]
     
    Recently, in dental practice, tooth reconstruction can be performed using dental adhesive technology following either a "glass-ionomer", an "etch-and-rinse" or a "self-etch" approach. Glass-ionomer cements are well-known to possess an auto-adhesive capability without requiring any surface pre-treatment. The fundamental bonding mechanism of resin-based materials to enamel and dentin is essentially based on an exchange process, in which minerals removed from the dental hard tissues are replaced by resin monomers. These resin monomers, upon polymerization become micro-mechanically interlocked in the created porosities. Besides micro-mechanical interlocking through hybridization, an additional chemical interaction between functional monomers/polymers and tooth substrate components has been found to be important. In this review, we focus on how the chemical interaction of the biomaterial-hard tissue interface can improve bond durability, especially using chemical analytical techniques.
  • Naoyuki Kaga, Tsukasa Akasaka, Rumi Horiuchi, Yasuhiro Yoshida, Atsuro Yokoyama
    Digest Journal of Nanomaterials and Biostructures 12 2 431 - 440 2017年05月 [査読有り][通常論文]
  • T. Akasaka, H. Miyaji, T. Imamura, N. Kaga, A. Yokoyama, Y. Yoshida
    DIGEST JOURNAL OF NANOMATERIALS AND BIOSTRUCTURES 12 2 281 - 292 2017年04月 [査読有り][通常論文]
     
    We investigated patterning with curable dental materials via micro/nano-molding as a screening trial. We used typical curable dental materials such as a composite resin (CR), acrylic resin, varnish, adhesive resin, restoration, sealer, cement, tooth desensitizer, and gypsum with either a cyclo-olefin polymer (COP) mold or polydimethylsiloxane (PDMS) mold that had holes, pillars, and grooves with 0.5 mu m diameters and 1 mu m heights. The COP mold was suitable for patterning of holes and grooves with Bondfill SB after 2 h curing, while PDMS mold was suitable for patterning of pillars with Bondfill SB at the 0.5 mu m scale after 2 h curing. Most of the curable dental materials could be patterned using the COP or the PDMS mold. However, dental impression materials could not be patterned as required. In addition, the COP mold was suitable for many organic pillars, while the PDMS mold was suitable for inorganic pillars. Further, we achieved patterning on the curved surface of an artificial tooth with flowable CR using the COP thin film mold. We conclude that for the formation of uniform shaped patterns, it is important to cure within a suitable time and to select mold types depending on the type of curable dental material.
  • Naoyuki Kaga, Rumi Horiuchi, Atsuro Yokoyama, Tsukasa Akasaka, Yasuhiro Yoshida
    e-Journal of Surface Science and Nanotechnology 15 1 - 6 2017年02月11日 [査読有り][通常論文]
     
    Surface topography influences cell growth and differentiation. In this study, we used a nano-imprinting method to develop a titanium sheet with 500 nm-, 1 μm-, and 2 μm-wide grooved and pillared structures. We investigated the effects of the surfaces with the micro- and nano-structures on Ca9-22 cell adhesion and proliferation. Ca9-22 cells were cultured in DMEM containing 10% fetal bovine serum and counted adhered cells at 1 and 24 h post-culture. Scanning electron microscopy was used to assess cell morphology. Immunofluorescence cell staining was used to evaluate vinculin formation to observe the presence of focal contacts at 24 h. There was no difference in cell adhesion between cells cultured on a plane or groove after 1 h. However, at 24 h, the adhesion of cells cultured on the groove was reduced. In addition, the cell adhesion count on the pillar was less than that of cells cultured on a plane at both 1 hour and 24 h post-seeding. Furthermore, in the groove of the Ti sheet after 1 and 24 h, cell expansion occurred in the grooved direction. These results demonstrate that the micro and nano-grooved and pillared structures on the titanium sheet control Ca9-22 cell adhesion and orientation.
  • Miyata S, Miyaji H, Kawasaki H, Yamamoto M, Nishida E, Takita H, Akasaka T, Ushijima N, Iwanaga T, Sugaya T
    International journal of nanomedicine 12 2703 - 2716 2017年 [査読有り][通常論文]
     
    Antimicrobial photodynamic therapy (aPDT) has beneficial effects in dental treatment. We applied captopril-protected gold (Au25(Capt)18) clusters as a novel photosensitizer for aPDT. Photoexcited Au clusters under light irradiation generated singlet oxygen (1O2). Accordingly, the antimicrobial and cytotoxic effects of Au25(Capt)18 clusters under dental blue light-emitting diode (LED) irradiation were evaluated. 1O2 generation of Au25(Capt)18 clusters under blue LED irradiation (420-460 nm) was detected by a methotrexate (MTX) probe. The antimicrobial effects of photoexcited Au clusters (0, 5, 50, and 500 μg/mL) on oral bacterial cells, such as Streptococcus mutans, Aggregatibacter actinomycetemcomitans, and Porphyromonas gingivalis, were assessed by morphological observations and bacterial growth experiments. Cytotoxicity testing of Au clusters and blue LED irradiation was then performed against NIH3T3 and MC3T3-E1 cells. In addition, the biological performance of Au clusters (500 μg/mL) was compared to an organic dye photosensitizer, methylene blue (MB; 10 and 100 μg/mL). We confirmed the 1O2 generation ability of Au25(Capt)18 clusters through the fluorescence spectra of oxidized MTX. Successful application of photoexcited Au clusters to aPDT was demonstrated by dose-dependent decreases in the turbidity of oral bacterial cells. Morphological observation revealed that application of Au clusters stimulated destruction of bacterial cell walls and inhibited biofilm formation. Aggregation of Au clusters around bacterial cells was fluorescently observed. However, photoexcited Au clusters did not negatively affect the adhesion, spreading, and proliferation of mammalian cells, particularly at lower doses. In addition, application of Au clusters demonstrated significantly better cytocompatibility compared to MB. We found that a combination of Au25(Capt)18 clusters and blue LED irradiation exhibited good antimicrobial effects through 1O2 generation and biosafe characteristics, which is desirable for aPDT in dentistry.
  • Tsukasa Akasaka, Takuya Imamura, Hirofumi Miyaji, Naoyuki Kaga, Atsuro Yokoyama, Yasuhiro Yoshida
    e-Journal of Surface Science and Nanotechnology 14 225 - 230 2016年12月10日 [査読有り][通常論文]
     
    Groove patterns on the surface of implants act as an effective barrier to the apical migration of epithelial attachment, after which the grooves facilitate gingival fibroblast attachment. Cell alignment on grooves is largely influenced by the adsorbed protein type. However, cell attachment and cell alignment properties of micro/nanogrooved dental composite resins using osteoblasts and fibroblasts have not been investigated. Further, the effect of saliva-related protein adsorption has not investigated. In this study, we prepared composite resins with grooves that were 2 μm, 1 μm, and 500 nm wide and estimated the effect of pre-coating of some proteins, mainly mucin, on attachment and alignment of human gingival fibroblasts(HGF). In the cell attachment assay on mucin-coated grooves, the number of attached cells on mucin-coated planar or grooved composite resins was lower compared to that on both composite resins without pre-coating of mucin. Interestingly, the number of attached cells on grooves pre-coated with mucin was 5.7-fold higher than those on planar pre-coated with mucin. Grooves at the micro/nano level may act as a hook for floating cells during the cell attachment assay. Furthermore, the degree of cell alignment was strongly dependent on the pre-coating protein types. The cells were radially spread or round-shaped, but not have sufficient alignment on non-, mucin-, and albumin-coated grooves. Although the cells were attached on the grooves, they were not aligned along the direction of grooves. The cells on fetal bovine serum- or fibronectin-coated grooves exhibited good alignment in the groove direction, particularly on fibronectin-coated grooves. Thus, our patterning method creates an effective seal between soft tissue and dental materials to protect against microorganism invasion.
  • Kaga N, Akasaka T, Horiuchi R, Yoshida Y, Yokoyama A
    Nano Biomedicine 8 2 74 - 82 2016年12月 [査読有り][通常論文]
  • Akasaka T, Miyaji H, Kaga N, Yokoyama A, Abe A, Yoshida Y
    Nano Biomedicine 8 2 112 - 122 2016年12月 [査読有り][通常論文]
  • Shuichi YAMAGATA, Yoshiaki SATO, Takaaki YAMAMOTO, Shigeaki ABE, Tsukasa AKASAKA, Yasuhiro YOSHIDA, Junichiro IIDA
    Nano Biomedicine 8 1 35 - 40 2016年10月 [査読有り][通常論文]
     
    Orthodontic adhesives are usually colorless and transparent for aesthetic purposes. However, making such adhesives fluorescent is one of the most effective solutions to make them visible to ensure safe and complete removal after orthodontic treatments. Trivalent europium ions were doped into yttrium oxides by a homogeneous precipitation method. The particles were synthesized via a homogeneous precipitation method and had a narrow size distribution (200-300 nm) and showed sharp crystallinity independent of the starting concentration of Eu3+. The strongest emission and absorption peaks were identified at 611 nm and 396 nm, respectively. In this study, 8 mol% was confirmed to be the optimum doping concentration. The emission intensity of phosphor containing PMMA adhesives was appreciably weak even when the concentration of phosphors was 10 wt%. It may be attributed to the clustering of the particles or a multiphonon relaxation process owing to organic groups in the polymer. We conclude that the crystalline Y2O3:Eu3+ particles could be applicable for the development of fluorescent orthodontic adhesives.
  • Shuichi YAMAGATA, Takaaki YAMAMOTO, Yoshiaki SATO, Shigeaki ABE, Tsukasa AKASAKA, Yasuhiro YOSHIDA, Junichiro IIDA
    Nano Biomedicine 8 1 57 - 63 2016年10月 [査読有り][通常論文]
     
    The purpose of this study was to investigate the effect of gliding arc discharge non-thermal atmospheric pressure plasma with N2 carrier gas on surface chemical modification for adhesion improvement. The shear bond strength between the orthodontic adhesive resin and plasma-treated stainless steel surface improved slightly, although the surface free energy of the stainless steel samples increased from 73.6 mJm-2 to 112.9 mJm-2 mainly due to an increase of the dipole component (γSp) and partially due to an increase of the hydrogen bond component (γSh). This is partly because the metastability of the stainless steel surface was stabilized by immediate hydration or adsorption of water on the highly hydroxylated stainless steel surfaces. However, surface cleaning and hydroxylating efficacy of the handheld device was determined to be superior to those of other surface cleaning procedures, particularly in terms of intraoral applications. The technique may be potentially employed in establishing M-S coordinate bonds in dental precious metals and related alloys.
  • Ko Nakanishi, Shuichi Yamagata, Tsukasa Akasaka, Shigeaki Abe, Yasuhiro Yoshida, Junichiro Iida
    JOURNAL OF NANOSCIENCE AND NANOTECHNOLOGY 16 9 9292 - 9297 2016年09月 [査読有り][通常論文]
     
    Titanium orthodontic anchoring screws are being increasingly used as a source of orthodontic force. However, the incidence of screw fracture during insertion and removal is not negligible; sometimes, invasive procedures are required for removal of the screw fragments. Therefore, poly(L-lactic acid) (PLLA)/organically modified montmorillonite (OMMT) nanocomposites were fabricated via a solution intercalation method for the development of novel orthodontic anchoring screws with adequate mechanical strength and bioabsorbability. The structural features and mechanical and thermal properties of the screws were characterized, and the viability of cells exposed to OMMT was also investigated to evaluate the biological safety of the PLLA/OMMT composites. Human osteosarcoma (MG-63) and mouse osteoblastic (MC3T3-E1) cell lines were used as target cells. The (001) diffraction peaks in the X-ray diffraction patterns of samples of pure OMMT, and polymer composites containing 2 and 4 vvt% OMMT were 2 theta = 2.5, 2.3, and 2.3 degrees, indicating interlayer distances of 3.5, 3.8 and 3.8 nm, respectively. Transmission electron microscopy images of the nanocomposite with 4 wt% OMMT revealed that the interlayer distance of the compounded OMMT was irregularly enlarged and ranged from 2 to 5 nm. In addition, some of OMMT layers were exfoliated. The flexural modulus values of the nanocomposites were significantly increased compared to that for pure PLLA, according to the results of three-point transverse analyses. Differential scanning calorimetry measurements also revealed that the glass transition temperatures, melting temperatures and degree of crystallinity were nearly the same for all of the samples and, thus, independent of the quantity of added OMMT. The cell viabilities for the MG-63 and MC3T3-E1 cells (greater than 90% and 80%, respectively) were also independent of the OMMT concentration. Scanning electron microscopy images of both types of cells following OMMT exposure exhibited normal shapes. Based on these results, we conclude that PLLA/OMMT nanocomposites prepared via solution intercalation should be useful for the development of bioabsorbable orthodontic anchoring screws.
  • Shigeaki Abe, Eri Seitoku, Nobuki Lwadera, Yusuke Hamba, Shuichi Yamagata, Tsukasa Akasaka, Teruo Kusaka, Satoshi Lnoue, Yasutaka Yawaka, Junichiro Lida, Hidehiko Sano, Tetsu Yonezawa, Yasuhiro Yoshida
    JOURNAL OF BIOMEDICAL NANOTECHNOLOGY 12 3 472 - 480 2016年03月 [査読有り][通常論文]
     
    We assessed the biocompatibility of nano-sized ceramic particles with several cells types. Though these particles have less than 100 nm in diameter, they act as submicron-sized particles in saline by aggregation that was estimated using laser diffraction particle size analysis (LDS). they act as submicro-sized particles in saline by aggregation based on laser diffraction particle size analysis (LDS). Several types of cells (osteoblasts, osteosarcoma and hepatocyte cells) were exposed to these particles and their cytocompatibility was estimated. Not only the cytotoxic assay but also their static and dynamic morphology under nanoparticles exposure were investigated. The intercellular uptake of particles was determined using a confocal fluorescence microscope. The particles used in this study did not inhibit cellular activity or growth even when their concentrations were high. Only copper oxide particles caused acute cytotoxicity depending on the particle size. The cytotoxicity assay, dynamic behavior of the nanoparticle-exposed cells and their examination under a confocal fluorescence microscope suggests that the irritative reaction was induced by contact between the cells and particles, whereas eluted copper ions are not dominant factor. These results indicate that nano-sized particles used in this study have excellent biocompatibility except copper oxide ones.
  • Shigeaki Abe, Nobuki Iwadera, Mami Mutoh, Tomohiko Katsurayama, Shino Morimoto, Ko Nakanishi, Tsukasa Akasaka, Satoshi Inoue, Yasutaka Yawaka, Junichiro Iida, Yasuhiko Yoshida
    Key Engineering Materials 696 234 - 237 2016年 [査読有り][通常論文]
     
    In this study, we investigated the cytocompatibility of ceramic nanoparticles on different types of cells. All ceramics nanoparticles investigated in this study except Copper oxide (CuO) exhibited good cytocompatibility and cell viability (90% or more) even at 20 ppm concentration. In contrast, CuO nanoparticles caused cell inflammation, and their effect depended on their particle size. Confocal fluorescence microscopy measurements indicated that some particles had penetrated into the cells. These results indicate that except CuO nanoparticles, all other ceramic nanoparticles reported herein exhibited excellent cytocompatibility even for lung epithelial cells.
  • Erika NISHIDA, Hirofumi MIYAJI, Junko UMEDA, Katsuyoshi KONDOH, Hiroko TAKITA, Izumi KANAYAMA, Saori TANAKA, Akihito KATO, Bunshi FUGETSU, Tsukasa AKASAKA, Masamitsu KAWANAMI
    Nano Biomedicine 7 1 11 - 20 2015年06月 [査読有り][通常論文]
  • NAKANISHI Ko, YAMAGATA Shuichi, AKASAKA Tsukasa, ABE Shigeaki, YOSHIDA Yasuhiro, IIDA Junichiro
    Key Eng Mater 631 151 - 155 2015年 [査読無し][通常論文]
  • Li Zhao, Yong‐Hong Xu, Hongmei Qin, Shigeaki Abe, Tsukasa Akasaka, Tokuhiro Chano, Fumio Watari, Takahide Kimura, Naoki Komatsu, Xiao Chen
    ADVANCED FUNCTIONAL MATERIALS 24 34 5348 - 5357 2014年09月 [査読有り][通常論文]
     
    In the field of nanomedicine, nanoparticles with various functions are required for in vivo applications such as biomedical imaging and drug delivery. Therefore, chemical functionalization of nanoparticles has been extensively investigated. Herein, nanodiamond (ND) coated with polyglycerol (PG) and its derivatives is reported to impart good solubility in a physiological environment, a stealth nature to avoid nonspecific uptake, a targeting property to be taken up by a specific cell, and an acid-responsive drug release property to kill cancer cells. ND is first grafted with PG and the resulting ND-PG has a high solubility in physiological media. Since a large number of hydroxyl groups in PG provide scaffolds for further surface functionalization, the targeting RGD peptide and Pt-based drug are immobilized to give ND-PG-RGD, ND-PG-Pt and ND-PG-RGD-Pt. The ND with intrinsic fluorescence is also functionalized by PG and RGD to confirm cellular uptake and intracellular localization fluorescently. The results of the cell experiments indicate that PG coating shielded fND from the uptake by HeLa and U87MG cells. In contrast, fND-PG-RGD is taken up by U87MG, not HeLa cells, exhibiting high targeting efficacy. When ND-PG-RGD-Pt is applied, U87MG is selectively killed against HeLa. The multi-functional ND is a promising prodrug in targeting chemotherapy.
  • Li Zhao, Yong-Hong Xu, Tsukasa Akasaka, Shigeaki Abe, Naoki Komatsu, Fumio Watari, Xiao Chen
    BIOMATERIALS 35 20 5393 - 5406 2014年07月 [査読有り][通常論文]
     
    A successful targeted drug delivery device for cancer chemotherapy should ideally be able to avoid nonspecific uptake by nonmalignant cells, particularly the scavenging monocyte-macrophage system as well as targeting efficacy to bring the drug preferentially into tumor cells. To this purpose, we developed a platform based on detonation nanodiamond (dND) with hyperbranched polyglycerol (PG) coating (dND-PG). dND-PG was first demonstrated to evade non-specific cell uptake, particularly by macrophages (U937). RGD targeting peptide was then conjugated to dND-PG through multistep organic transformations to yield dND-PG-RGD that still evaded macrophage uptake but was preferentially taken up by targeted A549 cancer cells (expressing RGD peptide receptors). dND-PG and dND-PG-RGD showed good aqueous solubility and cytocompatibitlity. Subsequently, the anticancer agent doxorubicin (DOX) was loaded through acid-labile hydrazone linkage to yield dND-PG-DOX and dND-PG-RGD-DOX Their cellular uptake and cytotoxicity were compared against DOX in A549 cells and U937 macrophages. It was found that dND-PG-DOX uptake was substantially reduced, displaying little toxicity in either type of cells by virtue of PG coating, whereas dND-PG-RGD-DOX exerted selective toxicity to A549 cells over U937 macrophages that are otherwise highly sensitive to DOX. Finally, dND-PG was demonstrated to have little influence on U937 macrophage cell functions, except for a slight increase of TNF-alpha production in resting U937 macrophages. dND-PG is a promising drug carrier for realization of highly selective drug delivery in tumor cells through specific uptake mechanisms, with minimum uptake in and influence on macrophages. (C) 2014 Elsevier Ltd. All rights reserved.
  • Koichi IMAI, Tsubasa SHIRAI, Fumio WATARI, Tsukasa AKASAKA, Tetsunari NISHIKAWA, Tomoharu OKAMURA, Akio TANAKA, Kazuhiko SUESE, Fumiya OGAWA, Yoshitomo HONDA, Hirofumi SAWAI, Hiromasa TAKASHIMA
    Nano Biomedicine 6 1 27 - 34 2014年06月 [査読有り][通常論文]
  • Erika Nishida, Hirofumi Miyaji, Hiroko Takita, Izumi Kanayama, Maiko Tsuji, Tsukasa Akasaka, Tsutomu Sugaya, Ryuji Sakagami, Masamitsu Kawanami
    JAPANESE JOURNAL OF APPLIED PHYSICS 53 6 06JD04  2014年06月 [査読有り][通常論文]
     
    Carbon-based nanomaterials are being investigated for biomedical applications. Graphene oxide (GO), a monolayer of carbon, holds promise as a tissue engineering substrate due to its unique physicochemical properties. The aim of this study was to evaluate the effect of a GO coating on cell proliferation and differentiation in vitro. We also assessed the bioactivities of collagen scaffolds coated with different concentrations of GO in rats. The results showed that GO affects both cell proliferation and differentiation, and improves the properties of collagen scaffolds. Subcutaneous implant tests showed that low concentrations of GO scaffold enhances cell in-growth and is highly biodegradable, whereas high concentrations of GO coating resulted in adverse biological effects. Consequently, scaffolds modified with a suitable concentration of GO are useful as a bioactive material for tissue engineering. (C) 2014 The Japan Society of Applied Physics
  • Shigeaki Abe, Yusuke Hamba, Atsushi Hyono, Shuichi Yamagata, Motohiro Uo, Junichiro Iida, Takayuki Kiba, Akihiro Murayama, Tsukasa Akasaka, Fumio Watari
    JOURNAL OF THE CERAMIC SOCIETY OF JAPAN 122 1423 216 - 221 2014年03月 [査読有り][通常論文]
     
    In this study, we prepared two types of europium (Eu)-doped nanoparticle, yttoria and titania and investigated their photo-physical properties and biocompatibilities. These particles do not inhibit cell culture, even at higher concentrations. The obtained nanoparticles had a strong and narrow red photo-luminescent peak. The narrow emission was observed even when the particles were uniformly doped into a polymer film. These results suggest that they can be used for applications not only in optoelectronic devices but also in biomedical devices for bioimaging and luminescent biomedical materials. (C)2014 The Ceramic Society of Japan. All rights reserved.
  • Yoshinori Kuboki, Toshitake Furusawa, Masaaki Sato, Yongkun Sun, Hidero Unuma, Shigeaki Abe, Ryuichi Fujisawa, Tsukasa Akasaka, Fumio Watari, Hiroko Takita, Rachel Sammons
    BIO-MEDICAL MATERIALS AND ENGINEERING 24 3 1539 - 1548 2014年 [査読有り][通常論文]
     
    Based on our previous finding that a chromatography with titanium beads selectively binds phosphoproteins, including caseins, phosvitin and dentin phosphoproteins, we investigated whether bone phosphoproteins also bind to titanium. Bovine bone matrix proteins were extracted with 2 M urea/PBS after demineralization. The 2 M urea extract was directly applied to the titanium chromatography column as reported. The chromatogram showed an initial large peak at breakthrough position (non-binding fraction) and a smaller second peak eluted later (titanium-binding fraction). Both peaks were analyzed by SDS polyacrylamide gel electrophoresis. Stains-all staining which preferentially identifies phospho-proteins revealed that the first peak contained no positively stained band, while the second peak showed 4 or 5 distinctive bands indicative of bone phosphoproteins. To investigate the biological functions of the titanium-binding bone proteins (TiBP), we implanted them into calvaria of rats, combined with titanium web (TW), a highly porous titanium scaffold of thin titanium-fibers. Bone TiBP induced significantly enhanced bone formation, and new bone appeared connected directly to titanium fibers, accompanied by active blood vessel formations. Control TW alone did not induce bone formation within the titanium framework. These results demonstrate that the bone titanium-binding proteins include phosphoproteins which enhance bone formation when implanted into bone with titanium.
  • Izumi Kanayama, Hirofumi Miyaji, Hiroko Takita, Erika Nishida, Maiko Tsuji, Bunshi Fugetsu, Ling Sun, Kana Inoue, A. Sako Ibara, Tsukasa Akasaka, Tsutomu Sugaya, Masamitsu Kawanami
    INTERNATIONAL JOURNAL OF NANOMEDICINE 9 3363 - 3373 2014年 [査読有り][通常論文]
     
    Background: Graphene oxide (GO) is a single layer carbon sheet with a thickness of less than 1 nm. GO has good dispersibility due to surface modifications with numerous functional groups. Reduced graphene oxide (RGO) is produced via the reduction of GO, and has lower dispersibility. We examined the bioactivity of GO and RGO films, and collagen scaffolds coated with GO and RGO. Methods: GO and RGO films were fabricated on a culture dish. Some GO films were chemically reduced using either ascorbic acid or sodium hydrosulfite solution, resulting in preparation of RGO films. The biological properties of each film were evaluated by scanning electron microscopy (SEM), atomic force microscopy, calcium adsorption tests, and MC3T3-E1 cell seeding. Subsequently, GO-and RGO-coated collagen scaffolds were prepared and characterized by SEM and compression tests. Each scaffold was implanted into subcutaneous tissue on the backs of rats. Measurements of DNA content and cell ingrowth areas of implanted scaffolds were performed 10 days post-surgery. Results: The results show that GO and RGO possess different biological properties. Calcium adsorption and alkaline phosphatase activity were strongly enhanced by RGO, suggesting that RGO is effective for osteogenic differentiation. SEM showed that RGO-modified collagen scaffolds have rough, irregular surfaces. The compressive strengths of GO-and RGO-coated scaffolds were approximately 1.7-fold and 2.7-fold greater, respectively, when compared with the non-coated scaffold. Tissue ingrowth rate was 39% in RGO-coated scaffolds, as compared to 20% in the GO-coated scaffold and 16% in the non-coated scaffold. Conclusion: In summary, these results suggest that GO and RGO coatings provide different biological properties to collagen scaffolds, and that RGO-coated scaffolds are more bioactive than GO-coated scaffolds.
  • Naoyoshi TARUMI, Tsukasa AKASAKA, Fumio WATARI
    Nano Biomedicine 5 2 85 - 94 2013年12月 [査読有り][通常論文]
  • Yusuke HAMBA, Shuichi YAMAGATA, Tsukasa AKASAKA, Motohiro UO, Junichiro IIDA, Fumio WATARI
    Nano Biomedicine 5 2 75 - 84 2013年12月 [査読有り][通常論文]
  • Koichi IMAI, Fumio WATARI, Tsukasa AKASAKA, Kazuhiko SUESE, Fumiya OGAWA, Hirofumi SAWAI, Hiromasa TAKASHIMA
    Nano Biomedicine 5 2 104 - 108 2013年12月 [査読有り][通常論文]
  • Inoue S, Uo M, Sakairi M, Hirata E, Lee M.H, Bae T.S, Akasaka T, Watari F, Yokoyama A
    Key Engineering Materials 529-530 621 - 624 2013年11月 [査読有り][通常論文]
     
    Carbon nanotubes (CNTs) have excellent chemical, physical, and biological properties such as strong cell adhesion, protein adsorption and cell proliferation in vitro. Excellent osteocompatibility for the CNT monolith was also reported in vivo. The purpose of this study was to evaluate the effects of anodized titanium coated with multiwalled CNTs (MWCNTs) on human osteosarcoma Saos2 cells and bone tissue. Saos2 cells on CNT-Ti showed excellent proliferation with extension of cell morphology in all directions. CNT-Ti wire was implanted in the bone marrow of femurs of rats. At 2 weeks after surgery, histological investigations revealed that bone tissue attached to the surface of the CNT-Ti directly. Thus the surface modification of anodized Ti by MWCNTs can be effective for bone formation.
  • Wen Kou, Tsukasa Akasaka, Fumio Watari, Göran Sjögren
    ISRN Dentistry 2013 296727 - 296727 2013年07月 [査読有り][通常論文]
     
    THE PURPOSE OF THIS STUDY IS TO EVALUATE FUNCTIONALIZED MULTIWALLED CARBON NANOTUBES (FMWCNTS) AS A POTENTIAL COATING MATERIAL FOR DENTAL ZIRCONIA FROM A BIOLOGICAL PERSPECTIVE: its effect on cell proliferation, viability, morphology, and the attachment of an osteoblast-like cell. Osteoblast-like (Saos-2) cells were seeded on uncoated and fMWCNT-coated zirconia discs and in culture dishes that served as controls. The seeding density was 10(4) cells/cm(2), and the cells were cultured for 6 days. Cell viability, proliferation and attachment of the Saos-2 cells were studied. The results showed that Saos-2 cells were well attached to both the uncoated and the fMWCNT-coated zirconia discs. Cell viability and proliferation on the fMWCNT-coated zirconia discs were almost the same as for the control discs. Better cell attachment was seen on the fMWCNT-coated than on the uncoated zirconia discs. In conclusion, fMWCNTs seem to be a promising coating material for zirconia-based ceramic surfaces to increase the roughness and thereby enhance the osseointegration of zirconia implants.
  • Shuichi YAMAGATA, Hiroshi IWASAKI, Yusuke HAMBA, Ko NAKANISHI, Natsumi USHIJIMA, Shigeaki ABE, Tsukasa AKASAKA, Fumio WATARI, Junichiro IIDA
    Nano Biomedicine 5 1 31 - 38 2013年06月 [査読有り][通常論文]
  • M. Morikawa, Y. Kuboki, T. Akasaka, S. Abe, H. Takita, F. Watari
    Key Engineering Materials 529-530 1 615 - 620 2013年 [査読有り][通常論文]
     
    For application of carbon nanotubes (CNTs) as biomaterials, it is important to clarify the interaction between CNTs and proteins, which may affect on cell proliferation and differentiation. In this study, the adsorption behavior was investigated for representative proteins, bovine serum albumin, lysozyme and fetal bovine serum by using chromatography system. It was also aimed whether the different treatment conditions of CNTs affected the adsorption of proteins. CNTs used for this study had a straight shape and about 70nm in a diameter from SEM observation. There was not much difference between untreated and treated CNTs from SEM images. In chromatography experiments, all the proteins of albumin, lysozyme and serum were eluted immediately after injected to the column of untreated CNTs. Second elution appeared after buffer was changed from phosphate saline buffer to 25mM sodium hydroxide (NaOH). The same tendency was confirmed for CNTs with only acid treatment. However the eluted peaks became remarkably smaller after the column was changed to CNTs with thermal and twice acid treatments. These results indicated that treatment conditions for CNTs affected the adsorption behavior of proteins. © (2013) Trans Tech Publications, Switzerland.
  • Nobuki Iwadera, Shigeaki Abe, Tsukasa Akasaka, Yasutaka Yawaka, Fumio Watari
    BIOCERAMICS 24 529-530 374 - + 2013年 [査読有り][通常論文]
     
    Cytotoxicity and cell behavior to micro / nano particles of TiO2 and CuO was evaluated using the viability measurement and time-lapse observation. After cultured, osteoblastic cells MC3T3-E1 were exposed to particles. After 24 hour exposure, their morphology was observed using a SEM and the viability was measured. Cells exposed to TiO2 indicated no or very low decrease of viability. The results were independent of the particle size. On the other hand, the viability of cells exposed to CuO decreased with the concentration, and showed the size dependence. The nano-sized CuO indicated higher toxicity compared with micro-sized one. Dynamic behavior of cells exposed to nano particles, was succeeded to observe in a time-lapse method for 24 hours. The observation showed that the cells exposed to CuO became dead after forming a spherical shape. This is consistent with the image taken by SEM. Time-lapse observation made it possible to see the dynamic reaction process from cell contact to particles at first, the following cell activity response and finally to cell death, which revealed a considerably different morphology from the static cell observed after fixation by conventional method.
  • Shigeaki Abe, Yusaku Hamba, Nobuki Iwadera, Tsukasa Akasaka, Shuichi Yamagata, Yasutaka Yawaka, Junichiro Iida, Motohiro Uo, Tetsu Yonezawa, Fumio Watari
    BIOCERAMICS 24 529-530 625 - + 2013年 [査読有り][通常論文]
     
    We succeeded in determination the biodistribution of several nano-sized particles administered to mice through the tail vein. After administration, these particles were observed in the lung, liver and spleen. The distribution behaviors depend upon not only chemical species but also the particles size. To estimate their cytocompatibility, these particles were exposed to osteoblastic cell at several concentrations. When the concentration reached at 10 ppm, their viability remained at 80% or more even nano-sized particle contained rare earth element. Only CuO particles indicated the viability decrease. The effect depended on the particle size. These results suggested that the chemical species played a dominant key in the biodistribution and biocompatibility of nanoparticles compared with the size-effect.
  • Tsukasa Akasaka, Shigeaki Abe, Fumio Watari
    Key Engineering Materials 529-530 1 379 - 384 2013年 [査読無し][通常論文]
     
    The reports on cytotoxic studies of carbon nanotubes (CNTs) increased exponentially. In the present study, we investigate murine macrophage RAW264.7 cell response for the CNTs immobilized on a polystyrene substrate. We prepared CNT-coated dishes, and estimate the interaction of RAW264.7 cells with CNTs by cell adhesion, proliferation assay, and measurement of TNF-α production. As a result, the highest cell adhesion and proliferation was observed on a commercially cell culture polystyrene dish, while CNT-coated dish indicate slightly lower activity of them. Moreover, amount of production of TNF-α on the CNT-coated dishes was considerable lower than that in the case of lipopolysaccharide (LPS) addition as a control. These results indicated that CNT-coated dishes could not show strong cytotoxicity for RAW264.7 cells in vitro. © (2013) Trans Tech Publications, Switzerland.
  • T. Akasaka, Fumio Watari
    Hokkaido Journal of Dental Science 32 2 189 - 191 2013年 [査読有り][通常論文]
  • Shigeaki Abe, Chika Koyama, Mami Mutoh, Tsukasa Akasaka, Motohiro Uo, Fumio Watari
    APPLIED SURFACE SCIENCE 262 125 - 128 2012年12月 [査読有り][通常論文]
     
    In this study, we investigated the biodistribution of platinum (Pt) microparticles in mice. The particles were administered through the tail vein, and then the biodistribution behavior was observed using inductively coupled plasma-atomic emission spectroscopy (ICP) and scanning X-ray analytical microscopy (XSAM). The administered particles quickly reached the spleen, liver, and lung at a constant ratio, and the ratio remained constant for four weeks. We also estimated the correlation between XSAM and ICP measurement. The relative ratio of XSAM intensity showed strong correlation with the relative ratio of Pt concentration in organs. (c) 2012 Elsevier B.V. All rights reserved.
  • Shigeaki Abe, Nobuki Iwadera, Mitsue Esaki, Ken-Ichi Aoyama, Tsukasa Akasaka, Motohiro Uo, Manabu Morita, Yasutaka Yawaka, Fumio Watari
    APPLIED SURFACE SCIENCE 262 248 - 252 2012年12月 [査読有り][通常論文]
     
    We investigated morphologies, size distributions and elemental compositions of dental debris formed by cutting/grinding teeth or dental alloys. The average size of debris formed by cutting/grinding dental alloy was around 100 mu m and that of teeth was 20 mu m. The debris formed by grinding with diamond or carborundum point had isotropic irregular shape, while the debris formed by cutting with carbide bar had characteristic lathe-cut shape. The elemental analysis indicated that the debris formed by grinding dental alloy with carborundum point consisted of not only the particles of the alloy but also the particles of Si compounds with the size of around 10 mu m. The particles of Si compounds would be formed by abrasion of the grinding instrument (carborundum, SiC). Similarly, the debris formed by grinding with diamond point also contained submicro-sized particles consisting of C compounds. The results indicate that the morphology and composition of dental debris are varied depending on the combination between the workpiece and the cutting/grinding materials and that the dental debris consist of both the workpiece and the cutting/grinding materials in some combination. In addition, some of the debris of tooth had the size less than 2 mu m, which has a potential to induce inflammation. Though the inflammation can be expected at low level, it is required to investigate the details in future. (c) 2012 Elsevier B.V. All rights reserved.
  • Kazunori FURUHASHI, Tsukasa AKASAKA, Yoshimasa KITAGAWA, Fumio WATARI
    Nano Biomedicine 4 2 76 - 84 2012年12月 [査読有り][通常論文]
     
    The use of dental implants currently entails problems associated with lack of adhesion to the oral mucosa at the point of abutment, which differs from that of natural teeth. Usually, the experimental evaluation of adhesion between test implant materials and soft tissue requires animal experiments. In this study, to facilitate easier evaluation, adhesion experiments were performed using a three-dimensional (3D) in vitro epidermal cell culture model. The implant materials examined were nano hydroxyapatite (nHAp) coated polyethylene terephthalate (PET), uncoated PET, zirconia, and titanium. The nHAp particles exhibited high cell adhesion, and nHAp coated PET exhibited high tissue-adhesion in the 3D epidermal cell culture in vitro. Zirconia exhibited substantial tissue adhesion to the 3D epithelial tissue, but titanium exhibited little adhesion. This 3D in vitro epidermal cell culture model could be used instead of animal experiments to facilitate easier evaluation of adhesion between tissue and materials, and it could be useful in screening tests for the development of tissue adhesive materials and processing conditions.
  • Shuichi Yamagata, Yusuke Hamba, Tsukasa Akasaka, Natsumi Ushijima, Motohiro Uo, Junichiro Iida, Fumio Watari
    APPLIED SURFACE SCIENCE 262 56 - 59 2012年12月 [査読有り][通常論文]
     
    Transparent poly(methyl methacrylate)/organically modified montmorillonite (PMMA/OMMT) nanocomposites were fabricated using a solution intercalation method. Two grades of OMMT modified with quaternary alkylammonium ions and containing different amounts of organic matter, NZ70 and NX, were used. X-ray diffraction patterns showed that the peaks regarded as a (0 0 1) d-spacing and a second peak were shifted toward lower 2 theta values, implying the expansion or the partial exfoliation of the silicate layers, respectively. TEM images showed that the silicate platelets were largely well dispersed independent of the hydrophobicity, although some aggregates were observed. The flexural modulus of the PMMA/OMMT nanocomposites increased favorably with an increasing amount of OMMT of either grade. However, only the NX displayed comparable strain at breakage to that of the PMMA. These results suggest that the highly hydrophobic grade NX shows great promise for use in nanocomposites made via solution intercalation. (c) 2012 Elsevier B.V. All rights reserved.
  • Eri Hirata, Tsukasa Akasaka, Motohiro Uo, Hiroko Takita, Fumio Watari, Atsuro Yokoyama
    APPLIED SURFACE SCIENCE 262 24 - 27 2012年12月 [査読有り][通常論文]
     
    The surface of a polylactic acid (PLLA) was coated multiwalled carbon nanotubes (MWCNTs) in order to improve the surface properties. In addition, its surface characteristics and cell culturing properties were examined. Whole surface of PLLA was homogeneously covered by MWCNTs maintained a unique tubular structure. MWCNT-coated PLLA showed remarkable higher wettability than uncoated PLLA. Human osteosarcoma cell line (Saos2) adhered well on the CNT-coated PLLA whereas there are few cells attached on the uncoated PLLA at 2 h after seeding. The number of the cells on uncoated PLLA was still smaller than on the MWCNT-coated PLLA at 1 and 3 days. Moreover, The DNA content in the cells attached to the MWCNT-coated PLLA was significantly higher than that on the uncoated PLLA (p < 0.05) at 1 and 3 days. There was no significant difference between the scaffolds for ALP activity normalized by DNA content at both term (p > 0.1). Therefore MWCNT-coating on PLLA improved the surface wettability and initial cell attachment at early stage. (c) 2012 Elsevier B.V. All rights reserved.
  • Yuhe Zhu, Wei Wang, Xingya Jia, Tsukasa Akasaka, Susan Liao, Fumio Watari
    APPLIED SURFACE SCIENCE 262 156 - 158 2012年12月 [査読有り][通常論文]
     
    Deposition of titanium carbide (TiC) layer on titanium (Ti) surface has been demonstrated by an ion-enhanced triode plasma chemical vapor deposition (CVD) method using a TiCl4 + CH4 + H-2 gas mixture. Physical and mechanical properties of the deposited TiC film on Ti were investigated to examine its potential application as an abrasion resistant implant material. X-ray diffraction (XRD) showed that the specimen was consisted of TiC and Ti. Carbide layer of about 6 mu m thickness was observed on the cross section of the specimen by scanning electron microscopy (SEM). The Vickers hardness of surface carbide was more than 2000, which confirmed its high abrasion resistance. (c) 2012 Elsevier B.V. All rights reserved.
  • Parvin Begum, Refi Ikhtiari, Bunshi Fugetsu, Makoto Matsuoka, Tsukasa Akasaka, Fumio Watari
    APPLIED SURFACE SCIENCE 262 120 - 124 2012年12月 [査読有り][通常論文]
     
    Abundant experimental data have shown that multi-walled carbon nanotubes (MWNTs) are toxic to plants, but the potential impacts of exposure remain unclear. The objective of the present study was to evaluate possible phytotoxicity of MWNTs at 0, 20, 200, 1000, and 2000 mg/L with red spinach, lettuce, rice, cucumber, chili, lady's finger, and soybean based on root and shoot growth, cell death, and electrolyte leakage at the seedling stage. After 15 days of hydroponic culture, the root and shoot lengths of red spinach, lettuce, and cucumber were significantly reduced following exposure to 1000 mg/L and 2000 mg/L MWNTs. Similar toxic effects occurred regarding cell death and electrolyte leakage. Red spinach and lettuce were most sensitive to MWNTs, followed by rice and cucumber. Very little or no toxic effects were observed for chili, lady's finger, and soybean. (c) 2012 Elsevier B.V. All rights reserved.
  • Koichi Imai, Tsukasa Akasaka, Fumio Watari, Akito Tanoue, Kazuaki Nakamura, Kazuhiko Suese, Hiromasa Takashima, Tetsunari Nishikawa, Akio Tanaka, Shoji Takeda
    APPLIED SURFACE SCIENCE 258 22 8444 - 8447 2012年09月 [査読有り][通常論文]
     
    The effects of nanomaterials on human reproduction and development remain unknown. The risks of nanomaterials for future generations should be elucidated. Thus, it is important to establish an experimental method to accurately examine embryotoxicity. We previously investigated the myocardial cell differentiation of ES-D3 cells using monolayer (SWCNTs) and multilayer (MWCNTs) nanocarbon tubes. As a result, in spite of having the same carbon composition, the effects on the cell differentiation levels differed between the tubes. We investigated their cell differentiation and cytotoxic effects on EL M3 and ES-R1-EGFP B2/EGFP cells, which require feeder cells. As a result, myocardial pulse rates differed between the presence of SWCNTs and MWCNTs even when feeder cells existed between the samples and cells. The different surface structures of SWCNTs and MWCNTs may have influenced ES cell differentiation. (C) 2012 Elsevier B. V. All rights reserved.
  • Shigeaki ABE, Minoru YOHDA, Aki NISHIYAMA, Mitsunori NAKATANI, Yuhei FUJII, Arisa KUSUHARA, Emi YAMATOYA, Tomoki ISHIDA, Tsukasa AKASAKA, Motohiro UO, Fumio WATARI, Daisuke HAYASHI, Tomoya TAKADA
    Nano Biomedicine 4 1 29 - 34 2012年06月 [査読有り][通常論文]
  • Mai MORIKAWA, Yoshinori KUBOKI, Tsukasa AKASAKA, Sigeaki ABE, Fumio WATARI
    Nano Biomedicine 4 1 35 - 41 2012年06月 [査読有り][通常論文]
  • Shuichi YAMAGATA, Yusuke HAMBA, Ko NAKANISHI, Shigeaki ABE, Tsukasa AKASAKA, Natsumi USHIJIMA, Motohiro UO, Junichiro IIDA, Fumio WATARI
    Nano Biomedicine 4 1 11 - 17 2012年06月 [査読有り][通常論文]
  • Makoto Matsuoka, Tsukasa Akasaka, Yasunori Totsuka, Fumio Watari
    MATERIALS SCIENCE & ENGINEERING C-MATERIALS FOR BIOLOGICAL APPLICATIONS 32 3 574 - 580 2012年04月 [査読有り][通常論文]
     
    Artificial cell scaffolds that support cell adhesion, growth, and organization need to be fabricated for various purposes. Recently, there have been increasing reports of cell patterning using electrical fields. We fabricated scaffolds consisting of silicone sheets coated with single-walled (SW) or multi-walled (MW) carbon nanotubes (CNTs) and evaluated their electrical properties and biocompatibility. We also performed cell alignment with dielectrophoresis using CNT-coated sheets as electrodes. Silicone coated with 10 mu g/cm(2) SWCNTs exhibited the least sheet resistance (0.8 k Omega/sq); its conductivity was maintained even after 100 stretching cycles. CNT coating also improved cell adhesion and proliferation. When an electric field was applied to the cell suspension introduced on the CNT-coated scaffold, the cells became aligned in a pearl-chain pattern. These results indicate that CNT coating not only provides electro-conductivity but also promotes cell adhesion to the silicone scaffold: cells seeded on the scaffold can be organized using electricity. These findings demonstrate that CNT-coated silicone can be useful as a biocompatible scaffold. (C) 2011 Elsevier B.V. All rights reserved.
  • Shigeaki Abe, Nobuki Iwadera, Takashi Narushima, Yoshiki Uchida, Motohiro Uo, Tsukasa Akasaka, Yasutaka Yawaka, Fumio Watari, Tetsu Yonezawa
    e-Journal of Surface Science and Nanotechnology 10 33 - 37 2012年03月17日 [査読有り][通常論文]
     
    In this study, we investigated the biocompatibility of gelatin-coated copper nanoparticles. To estimate their cytotoxicity, the coated copper nanoparticles were exposed to osteoblastic cells. The cell proliferation remained above 80% even when the particles concentration increased. When uncoated copper oxide nanoparticles were exposed to the cells, the proliferation ratio rapidly decreased with the concentration reaching 20% under the same conditions. To determine their biodistribution, the nanoparticles were administered to mice through their tail veins. The particles were subsequently found in some organs using an energy-dispersed X-ray spectrometer and inductively coupled plasma-atomic emission spectroscopy. The polymer-coated nanoparticles were observed in the lung, liver and spleen. They were also detected in the urine at higher concentrations than the copper oxide nanoparticles. Thus, the polymer coating is expected to improve biocompatibility by virtue of the excellent cytocompatibility and acceleration of the excretion process. © 2012 The Surface Science Society of Japan.
  • S. Abe, A. Sakaki, T. Narushima, Y. Uchida, T. Akasaka, M. Uo, T. Yonezawa, F. Watari
    Key Engneering Materials 493-494 752 - 756 2012年 [査読無し][通常論文]
  • Yoshinori Kuboki, Toshitake Furusawa, Masaaki Sato, Yongkun Sun, Hidero Unuma, Ryuichi Fujisawa, Shigeaki Abe, Tsukasa Akasaka, Fumio Watari, Hiroko Takita, Rachel Sammons
    BIO-MEDICAL MATERIALS AND ENGINEERING 22 5 283 - 288 2012年 [査読有り][通常論文]
     
    The biochemical mechanism behind the strong binding between titanium and living bone has not been fully elucidated, in spite of worldwide clinical application of this phenomenon. We hypothesized that one of the core mechanisms may reside in the interaction between certain proteins in the host tissues and the implanted titanium. To verify the interaction between titanium and proteins, we chose the technique of chromatography in that titanium spherical beads (45 mu m) were packed into a column to obtain a bed volume of 16 x 50 mm, which was eluted with phosphate buffered saline (PBS) and a straight gradient system made by using PBS and 25 mM NaOH. Fetal calf serum, albumin, lysozyme, casein, phosvitin and dentin phosphoprotein (phosphophoryn) were applied to the column. Most part of albumin and lysozyme eluted with the breakthrough peak, indicating practically no affinity to titanium. Fetal bovine serum also eluted mostly as the breakthrough peak, but distinct retained peak was observed. On the other hand, alpha-casein, phosvitin and phosphophoryn exhibited a distinct retained peak separated from the breakthrough peak. We proposed that phosphate groups (phosphoserines) in the major phosphoproteins, alpha-casein, phosvitin and phosphophoryn may be involved in the binding of these proteins with titanium.
  • Shigeaki Abe, Sachiko Itoh, Daisuke Hayashi, Toshihisa Kobayashi, Takayuki Kiba, Tsukasa Akasaka, Motohiro Uo, Yasutaka Yawaka, Shin-Ichiro Sato, Fumio Watari, Tomoya Takada
    JOURNAL OF NANOSCIENCE AND NANOTECHNOLOGY 12 1 700 - 706 2012年01月 [査読有り][通常論文]
     
    In this study, we prepared two-types of water-dispersible carbon nanotubes (CNTs) and investigated their biodistribution in mice as well as bio-/cyto-compatibility. After administration, their organs were excised at various post-injection times, then observed using both optical and transmission electron microscopy (TEM). The color of the liver and lung markedly darkened, suggesting that administered CNTs reached these organs. By TEM observation, the CNTs were found in the liver and lung. They were observed even in the kidney and spleen, though their distributions in those organs were very low compared with that in liver and lung. Therefore, most of the administered CNTs would be accumulated in the liver or lung. However, the time profile of the body weight of CNT-administered mice was close to that of control mice. In addition, we estimated the cytocompatibility of the water-dispersible CNTs for hepatocytes. According to a TNF-alpha assay of the cells cultured with CNTs, the expression level was almost the same as that of the control. These results suggested that the water-dispersible CNTs have good bio-/cyto-compatibility under this condition.
  • Shigeaki ABE, Akiko SASAKI, Nobuki IWADERA, Tsukasa AKASAKA, Motohiro UO, FumioI WATARI
    Nano Biomedicine 3 2 300 - 305 2011年11月 [査読有り][通常論文]
  • Shuichi YAMAGATA, Yusuke HAMBA, Tsukasa AKASAKA, Motohiro UO, Junichiro IIDA, Fumio WATARI
    Nano Biomedicine 3 1 217 - 223 ナノ・バイオメディカル学会 2011年08月 [査読有り][通常論文]
     
    Transparent poly (methyl methacrylate)/organically modified montmorillonite (PMMA/OMMT) nanocomposites were fabricated using a solution intercalation method. OMMT was modified with quaternary alkylammonium ions. The haze values, which are the ratios of diffuse transmittance to total light transmittance via the nanocomposites, gradually increased with an increasing amount of OMMT; however, the deterioration of total light transmittance was maintained within a narrow range. X-ray diffraction patterns showed that a peak regarded as a (001) d-spacing or a second peak corresponding to (002) plane was shifted toward lower 2θ values indicating the penetration of the PMMA polymer chains into the interlayer regions of OMMT. Board-shaped specimens used for the flexural tests were fabricated by compression molding at 230°C. The specimens were sufficiently transparent for their use as esthetic orthodontic wires. The flexural modulus of the PMMA/OMMT nanocomposites increased favorably with an increasing amount of OMMT. It is reasonable to consider that the addition of OMMT to the nanocomposites effectively improves the stiffness of PMMA without seriously deteriorating its flexibility.
  • Shigeaki ABE, Katsutoshi NAKAYAMA, Hirohisa KOBAYASHI, Takayuki KIBA, Tsukasa AKASAKA, Shin-ichiro SATO, Motohiro UO, Fumio WATARI, Tomoya TAKADA
    Nano Biomedicine 3 1 208 - 216 ナノ・バイオメディカル学会 2011年08月 [査読有り][通常論文]
     
    There is considerable investigation into the applications of carbon nanotubes because of their valuable properties. In this study, our goal was to develop several carbon nanotube (CNT)-coated substrates through an amide-bonding process. We prepared polycarboxylated multi-walled CNTs via a carboxylation reaction and then CNT derivatives were tethered to amino groups presented on a titanium, gold or glass substrate in the presence of a condensation agent. The obtained substrates were characterized with a Raman microscope, an atomic force microscope (AFM) and a scanning electron microscope (SEM). Raman mapping images indicated that the CNTs existed on the substrate surface homogeneously, and AFM and SEM observation revealed that the CNTs coating the surfaces had a three-dimensional network nanostructure. The CNTs remained on the surface even after ultrasonication treatment in organic solvent. However, they were removed from the substrate surface after soaking in NaOH aqueous solution. This result suggested that the CNTs were tethered to the substrate through amide bonding.
  • Koichi IMAI, Tsukasa AKASAKA, Fumio WATARI, Akito TANOUE, Kazuaki NAKAMURA, Kazuhiko SUESE, Hiromasa TAKASHIMA, Tetsunari NISHIKAWA, Akio TANAKA, Shoji TAKEDA
    Nano Biomedicine 3 1 224 - 230 ナノ・バイオメディカル学会 2011年08月 [査読有り][通常論文]
     
    Nano and submicron titanium dioxides are widely used as ingredients of sun screen. Thus, its reproductive and developmental toxicities in humans are of concern. We used two kinds of nano titanium dioxides, involving different production methods and surface coatings. The embryotoxicities of these nano titanium dioxides were determined based on inhibitory effects on the cardiomyocyte differentiation of mouse Embryonic Stem cells. As ultrafine titanium dioxide, titanium dioxide (particle diameter, 0.03-0.05 μm) produced by the sintering method (T1) and that (particle diame-ter, 0.01-0.02 μm) produced by the wet method (T2) were used. T2 may cause little developmental toxicity because it has no potent inhibitory effects on cardiomyocyte differentiation. However, T1 should be further investigated.
  • Eri Hirata, Motohiro Uo, Hiroko Takita, Tsukasa Akasaka, Fumio Watari, Atsuro Yokoyama
    CARBON 49 10 3284 - 3291 2011年08月 [査読有り][通常論文]
     
    Surface coating treatment with multiwalled carbon nanotubes (MWCNTs) was applied to 3D collagen scaffold for bone tissue engineering. Rat primary osteoblasts (ROBs) were cultured on an MWCNT-coated collagen sponge (MWCNT-coated sponge) in a 3D dynamic flow cell culture system and differentiation markers were measured. Alkaline phosphatase activity at 1 day, and calcium and osteopontin contents of the MWCNT-coated sponges at 7 days were significantly higher than those of uncoated sponges. ROBs on the MWCNT-coated sponge differentiated earlier than on the uncoated sponge. In addition, the tissue response to the MWCNT-coated sponge was evaluated. Slight inflammation was observed around MWCNTs at 7 and 28 days after implantation in subcutaneous tissue. Significantly more bone formation was observed around the MWCNT-coated sponges than around the uncoated sponges and new bone attached to MWCNTs directly at 28 and 56 days after implantation in the femur. Moreover, at 28 days after implantation of the MWCNT-coated sponge with osteoblasts cultured for 1 day, bone tissues were successfully formed in the pores according to its honeycomb structure. Therefore, MWCNT coating appears to be effective for bone tissue engineering. (C) 2011 Elsevier Ltd. All rights reserved.
  • Tsukasa Akasaka, Atsuro Yokoyama, Makoto Matsuoka, Takeshi Hashimoto, Fumio Watari
    CARBON 49 7 2287 - 2299 2011年06月 [査読有り][通常論文]
     
    Induced pluripotent stem (iPS) cells have attracted worldwide interest. However, there have been only a few studies investigating effective culture substrates for feeder-free culturing for the maintenance of iPS cells. In this study, we cultured mouse iPS cells under feeder-free conditions on carbon nanotube (CNT)-coated dishes and then evaluated the colony morphology and differentiation state of the cells on the dishes. After 5 d of cultivation in a medium containing 15% fetal bovine serum (FBS) and leukemia inhibitory factor (LIF), the colonies on thick films of multi-walled CNTs (MWCNTs) were observed to be hemiround; further, the cells expressed early undifferentiation markers. On the other hand, the colonies on a cell culture polystyrene dish and a collagen-coated polystyrene dish showed indistinct outline and spread well, and most spreading cells only weakly expressed early undifferentiation markers. These results indicate that the thick films of MWCNTs could maintain hemiround colonies and undifferentiated state of mouse iPS cells cultured under feeder-free conditions. (C) 2011 Elsevier Ltd. All rights reserved.
  • Motohiro Uo, Tsukasa Akasaka, Fumio Watari, Yoshinori Sato, Kazuyuki Tohji
    DENTAL MATERIALS JOURNAL 30 3 245 - 263 2011年05月 [査読有り][通常論文]
     
    After the discovery of fullerene and carbon nanotubes, various carbon nanomaterials were discovered or synthesized. The carbon nanomaterials have remarkable properties, different from bulk materials with the same chemical composition, and are therefore useful for industrial applications. However, the toxicity of nanomaterials may also differ from that of the bulk materials; this difference poses a concern. The physical similarity of nanomaterials to asbestos has led to evaluations for toxicity by many researchers using various methods. In this review, we compile and compare the toxicity evaluations of each carbon nanomaterial.
  • Shigeaki Abe, Kosuke Ishikawa, Atsushi Hyono, Hirohisa Kobayashi, Takayuki Kiba, Tsukasa Akasaka, Motohiro Uo, Yasutaka Yawaka, Shin-Ichiro Sato, Tetsu Yonezawa, Fumio Watari
    e-Journal of Surface Science and Nanotechnology 9 80 - 84 2011年03月05日 [査読有り][通常論文]
     
    Development of scaffolds consisting of micro-/nano-sized materials have attracted a great deal of attention for their potential use in tissue engineering tools. We prepared a cell culture scaffold of carbon nanotubes, which is a typical bio-inert nanomaterial, and then investigated the surface morphology and properties. A three-dimensional nano-level network structure was observed using a scanning electron microscope and an atomic force microscope. The scaffold also exhibited excellent protein absorption. In order to apply the obtained scaffold to the cultivation of osteoblast cells, the cytocompatibility was comparable to that of a conventional cell culture dish. © 2011 The Surface Science Society of Japan.
  • Shigeaki Abe, Nobuki Iwadera, Mami Mutoh, Chika Koyama, Mitsue Esaki, Tsukasa Akasaka, Motohiro Uo, Manabu Morita, Yoshinori Kuboki, Koichi Haneda, Yasutaka Yawaka, Fumio Uchida, Tetsu Yonezawa, Fumio Watari
    Bioceramics Development and Applications 1 D110134  2011年01月 [査読有り][通常論文]
  • Shigeaki Abe, Katsutoshi Nakayama, Daisuke Hayashi, Tsukasa Akasaka, Motohiro Uo, Fumio Watari, Tomoya Takada
    9TH INTERNATIONAL CONFERENCE ON NANO-MOLECULAR ELECTRONICS 14 147 - 151 2011年 [査読有り][通常論文]
     
    There is considerable investigation into the applications of carbon nanotubes because of their valuable properties. In this study, we succeeded in poly-carboxylation of multi-walled carbon nanotubes (CNTs) tethered to a glass substrate through covalent bonding. By causing a carboxylation reaction using a peroxide derivative, we were able to observe many carboxyl groups on the surfaces of CNTs. We characterized the obtained CNTs derivatives using Infrared and Raman spectroscopy. The CNTs reacted directly with glass substrates that presented amino groups, and the tethered CNTs were not removed from the substrate by sonication, suggesting the formation of a strong bond between the CNTs and substrate. Nanostructures in the shape of curled individual strings were observed using a scanning electron microscope and an atomic force microscope. The transparency of substrates was more than 97% in visible regions. (C) 2010 Published by Elsevier B.V.
  • Shigeaki Abe, Nobuki Iwadera, Mitsue Esaki, Ikuhiro Kida, Mami Mutoh, Tsukasa Akasaka, Motohiro Uo, Yasutaka Yawaka, Manabu Morita, Koichi Haneda, Tetsu Yonezawa, Fumio Watari
    IOP Conference Series: Materials Science and Engineering 18 13 192013  2011年 [査読有り][通常論文]
     
    Nano-sized materials have received much attention lately, both in terms of their multiple applications and their biocompatibility. From both viewpoints, understanding the biodistribution of administered nano-materials is very important. In this study, we succeeded in visualizing the biodistribution of administered nano-materials using a scanning X-ray analytical microscope and magnetic resonance imaging method. Quantitative observation was carried out by inductively coupled plasma - atomic emission spectroscopy. We observed that the administered nano-particles accumulated in the liver, lung and spleen of mice. To estimate their cytocompatibility, the nano-particles were exposed to human liver cells. The results suggested that the micro-/ nano- particles have good cytocompatibility, except for copper oxide nano-particles. © 2011 Ceramic Society of Japan.
  • Kosuke Ishikawa, Tsukasa Akasaka, Shigeaki Abe, Yasutaka Yawaka, Masaya Suzuki, Fumio Watari
    Bioceramics Development and Applications 1 D110133  2011年01月 [査読有り][通常論文]
  • Hiromi KITAHARA, Yoshinori KUBOKI, Hiroko TAKITA, Tsukasa AKASAKA, Fumio WATARI, Nobuo INOUE
    Nano Biomedicine 2 2 81 - 92 ナノ・バイオメディカル学会 2010年12月 [査読有り][通常論文]
     
    Cell proliferations on carbon nanotube scaffolds were investigated for the embryonic stem cells (ES cells) of mouse and the bone marrow-derived mesenchymal stem cells (MSC) of rat. Although ES cells were well attached and proliferated on gelatin-coated plastic dishes, they did not attach on the CNT-coated scaffold, and formed the spheroids growing in the culture medium. For MSC, they attached on the CNT-coated scaffold and grew to form mineralized particles as on the plastic dishes of control. The growth rate of the MSC was lower than control but showed the higher expression of alkaline phosphatase activity per cell in 2 weeks and degree of mineralization per cell for 3 and 4 weeks.
  • Makoto Matsuoka, Tsukasa Akasaka, Yasunori Totsuka, Fumio Watari
    MATERIALS SCIENCE AND ENGINEERING B-ADVANCED FUNCTIONAL SOLID-STATE MATERIALS 173 1-3 182 - 186 2010年10月 [査読有り][通常論文]
     
    In recent years, carbon nanotubes (CNTs) have been considered potential biomedical materials because of their unique character. The aim of this study was to investigate the response of a human osteoblast-like cell line - Saos-2 - on single-walled CNTs (SWCNTs) and multi-walled CNTs (MWCNTs). The surface of a culture dish was coated with CNTs, and Saos-2 cells were cultured for three days. Cell morphology, viability, alkaline phosphatase (ALP) activity, adhesion, and vinculin expression were evaluated. The result showed high cell viability and strong adhesion to MWCNTs. Saos-2 cultured on MWCNTs exhibited vinculin expression throughout the cell body. while the cells attached to SWCNTs and glass were mostly limited to their periphery. Our results suggest that CNT coatings promote cell activity and adhesiveness. These findings indicate that MWCNTs could be used as surface coating materials to promote cell adhesion. (C) 2009 Elsevier B.V. All rights reserved.
  • Tsukasa Akasaka, Makoto Matsuoka, Takeshi Hashimoto, Shigeaki Abe, Motohiro Uo, Fumio Watari
    MATERIALS SCIENCE AND ENGINEERING B-ADVANCED FUNCTIONAL SOLID-STATE MATERIALS 173 1-3 187 - 190 2010年10月 [査読有り][通常論文]
     
    Dental caries are mainly associated with oral pathogens, and Streptococcus mutans is a primary cariogenic organism. Many methods have been established to eliminate S. mutans from the oral cavity. This study aimed to evaluate the effect of carbon nanotube (CNT)/agar composites irradiated with near-infrared (NIR) light on S. mutans, as a potential photothermal antimicrobial nanotherapy. A colony-forming unit assay clearly showed that CNT/agar composites attain bactericidal activity after NIR light irradiation; this bactericidal activity is higher than that of graphite (GP)/agar and activated carbon (AC)/agar composites. Furthermore, it was observed that longer irradiation times immobilized S. mutans in the CNT/agar composite. (C) 2010 Elsevier B.V. All rights reserved.
  • Shigeaki Abe, Takayuki Kiba, Kiyotada Hosokawa, Satoru Nitobe, Takashi Hirota, Hirohisa Kobayashi, Tsukasa Akasaka, Motohiro Uo, Yoshinori Kuboki, Shin-Ichiro Sato, Fumio Watari, Iosif D. Rosca
    JOURNAL OF ELECTRON SPECTROSCOPY AND RELATED PHENOMENA 181 2-3 181 - 185 2010年08月 [査読有り][通常論文]
     
    We prepared fluorescent coumarin dye-doped poly (acrylic acid) microparticles, which are well known as a biodegradable polyester, and the photophysical properties were characterized by scanning electron microscope, atomic force microscope and spectroscopic investigation. Spherical particles with diameters ranging from 0.5 to a few mu m were obtained. Based on spectroscopic investigation, the internal environment was close to that of a polar solvent such as methanol, and the dyes were dispersed without aggregation inside the particles. The obtained particles were administered to a mouse through the tail vein, and the biodistribution was then observed after some organs were excited at 1-day and 1-week post-injection. The particles were accumulated ill the organs, especially in the lung and spleen. After injection, the particles were trapped temporally in the lung, and then seemed to be transported to other organs by blood circulation. This tendency is similar to the biodistribution of TiO(2) microparticles that we have reported previously. (C) 2010 Elsevier B.V. All Fights reserved.
  • Yoshinori Kuboki, Takamitu Koshikawa, Hiroko Takita, Ryuichi Fujisawa, Min-ho Lee, Shigeaki Abe, Tsukasa Akasaka, Motohiro Uo, Fumio Watari, Rachel Sammons
    DENTAL MATERIALS JOURNAL 29 4 369 - 373 2010年07月 [査読有り][通常論文]
     
    Chromatography technology was employed to clarify the mechanism of interaction between multi-wall carbon nanotubes (MWCNT) and proteins. A column (16x100 mm) was packed with purified MWCNT, and various proteins were eluted with phosphate buffered saline (PBS) with and without gradient systems. It was found that albumin in bovine serum was eluted immediately from the column without any adsorption to MWCNT. Conversely, the non-albumin proteins, including a protein of 85 kDa molecular mass and a group of proteins with molecular masses higher than 115 kDa, exhibited considerably high affinity towards MWCNT. A sample of pure bovine serum albumin was also eluted immediately from the column, while lysozyme did not elute as a peak with PBS, but eluted with 0.6 M NaCl. Fundamentally, carbon nanotubes are devoid of any electrical charge. Therefore, other forces including the hydrogen bonds, hydrophilic interactions, and van der Waals forces were most probably responsible for the differential elution behaviors. In conclusion, this chromatographic method provided a simple and direct analysis of the interactions between carbon nanotubes and the various proteins.
  • Shigeaki Abe, Ikuhiro Kida, Mitsue Esaki, Nobuki Iwadera, Mami Mutoh, Chika Koyama, Tsukasa Akasaka, Motohiro Uo, Yoshinori Kuboki, Manabu Morita, Yoshinori Sato, Koichi Haneda, Tetsu Yonezawa, Balachandran Jeyadevan, Kazuyuki Tohji, Fumio Watari
    JOURNAL OF THE CERAMIC SOCIETY OF JAPAN 118 1378 525 - 529 2010年06月 [査読有り][通常論文]
     
    Internal distribution of several nanoparticles in mice has been investigated using scanning X-ray analytical microscopy, magnetic resonance imaging and inductively coupled plasma atomic emission spectroscopy. According dynamic laser scattering analysis, the actual particle size of obtained metal oxide was similar to that of metal particles. The estimated diameters were ca. one micrometer. After administration through the tail vein of the mice, metal particles quickly reached some organs. The distribution ratio reached a constant value and was then maintained. On the other hand, the metal oxide particles were first localized in the spleen and lung. The concentration in the lung was decreased with post-injection time. This result suggested that the particles were temporally trapped in the lung then removed to other organs. The behaviors between metal and metal oxide were quite different even when those particles had a similar actual particle size. Therefore, the distribution behavior of particles depended on the chemical species. (C) 2010 The Ceramic Society of Japan All rights reserved.
  • Koichi IMAI, Kazuhiko SUESE, Tsukasa AKASAKA, Fumio WATARI, Shoji TAKEDA
    Nano Biomedicine 2 1 47 - 51 ナノ・バイオメディカル学会 2010年06月 [査読有り][通常論文]
     
    We examined the influence of culture methods with mono- and multilayer nanocarbon tubes (SWCNT and MWCNT, respectively) on the differentiation of mouse embryonic stem (ES) cells using a cellular differentiation technique for the embryonic stem cell test (EST). On differentiation assay, beating was noted in 72, 35, and 60% of the cells in the control, SWCNT, and MWCNT groups, respectively. In addition, there was no influence on cytotoxicity based on the LDH level. In this study, the MWCNT did not influence the differentiation of ES cells, but the SWCNT markedly inhibited it, confirming that the differentiation of ES cells depends on the material surface structure.
  • Eri Hirata, Motohiro Uo, Yoshinobu Nodasaka, Hiroko Takita, Natsumi Ushijima, Tsukasa Akasaka, Fumio Watari, Atsuro Yokoyama
    JOURNAL OF BIOMEDICAL MATERIALS RESEARCH PART B-APPLIED BIOMATERIALS 93B 2 544 - 550 2010年05月 [査読有り][通常論文]
     
    The cell adhesion in a multiwalled carbon nanotube-coated collagen sponge (MWCNT-coated sponge) was investigated Immediately after seeding, the cells adhered to the inner surface of the MWCNT-coated sponge and a significantly larger number of cells were observed there than for a pure collagen sponge used as control On the MWCNT-coated sponge, the cells appeared favorable adhesion and spread in the early stages in the center part of the sponge which cells rarely attached without MWCNT-coating It was suggested that the physical structure of MWCNTs was effective for initial adhesion of cells from the result of serum-free culture MWCNT-coating makes the material a suitable three-dimensional scaffold for cell culturing, as opposed to other scaffold systems where such an effect is not seen (C) 2010 Wiley Periodicals, Inc J Biomed Mater Res Part B Appl Biomater 93B 544-550, 2010
  • Tsukasa Akasaka, Atsuro Yokoyama, Makoto Matsuoka, Takeshi Hashimoto, Fumio Watari
    MATERIALS SCIENCE & ENGINEERING C-MATERIALS FOR BIOLOGICAL APPLICATIONS 30 3 391 - 399 2010年04月 [査読有り][通常論文]
     
    One strategy used for the regeneration of bone is the development of cell culture substrates and scaffolds that can control osteoblast proliferation and differentiation. In recent investigations, carbon nanotubes (CNTs) have been utilized as scaffolds for osteoblastic cell cultures; however, there are only a few reports describing the proliferation of osteoblastic cells on thin CNT films; in particular, the effects of serum concentration on cell proliferation have not been studied. In the present study, we prepared culture dishes with homogeneous thin or thick films of non-modified CNTs and examined the effect of serum concentrations on human osteoblastic cells (Saos-2) proliferation in these culture dishes. We demonstrated that the ratio of cell proliferation was strongly affected by the concentration of serum. Interestingly, single-walled carbon nanotube (SWNT) thin films were found to be the most effective substrate for the proliferation of Saos-2 cells in low concentrations of serum. Thus, thin SWNT films may be used as an effective biomaterial for the culture of Saos-2 cells in low serum concentrations. (C) 2009 Elsevier B.V. All rights reserved.
  • Kosuke Ishikawa, Tsukasa Akasaka, Yasutaka Yawaka, Fumio Watari
    JOURNAL OF BIOMEDICAL NANOTECHNOLOGY 6 1 59 - 65 2010年02月 [査読有り][通常論文]
     
    Imogolite is a naturally occurring aluminosilicate clay mineral with a nanotube structure of 2 nm diameter and 60 nm-2 mu m length. In this study, synthetic imogolite nanotubes were applied to cell culture and the properties as a scaffold were compared with those of a conventional culture dish and a carbon nanotube (CNT) scaffold. The surface characteristics of the imogolite scaffold were drastically changed with the amount of coated imogolite on the dish. With the increase of imogolite concentration, the surface morphology of the scaffolds changed from an island-like shape in random orientation to a self-organized fiber texture aligned in the same direction, and finally full coverage in a random orientation with plural layers. Silicon releasability of the scaffolds was increased with the amount of coated imogolite. Mouse osteoblast-like cells (MC3T3-E1) cultured on imogolite showed a flat-form morphology and a cytoskeleton developed in all directions in contrast to a spindle-shaped morphology developed in one direction on cell culture dishes and CNT scaffolds. The proliferation and osteoblastic functions in ALP activity and mineralization showed similar increasing tendency. The largest values were obtained from the scaffold with a high concentration of imogolite. The results indicated good biocompatibility between the cells and imogolite scaffold and the enhancement effect of proliferation and osteoblastic differentiation.
  • F. Watari, T. Akasaka, K. Ishikawa, M. Matsuoka, E. Hirata, N. Terada, A. Yokoyama, M. Uo, S. Itoh, Y. Yawaka, M. Suzuki, N. Takashi, Y. Totsuka, Y. Kitagawa, S. Abe, I. D. Rosca, Y. Kuboki, Y. Bando
    MULTISCALE, MULTIFUNCTIONAL AND FUNCTIONALLY GRADED MATERIALS 631-632 181 - + 2010年 [査読有り][通常論文]
     
    Carbon nanotubes (CNT) and their derivatives with different structure and compositions have unique features. In the present study, cell proliferation was performed on various nanotubes such as single walled CNTs, multiwalled CNTs and imogolite which is nanotubes of aluminosilicate. SEM observation of the growth of osteoblast-like cells cultured on CNTs showed the morphology fully developed for the whole direction, which was different from that extended to the one direction on the usual scaffold. Numerous filopodia were grown from cell edge, extended far long and combined with CNT meshwork. Apatite precipitation in simulated body fluid, affinity for proteins and saccharides, and nanosize meshwork structure with large porosity would be the properties responsible for these cell adhesion and growth. Imogolite showed the similar properties to CNTs. Nanotubes could be the favorable materials for biomedical applications.
  • Shuichi YAMAGATA, Tsukasa AKASAKA, Motohiro UO, Natsumi USHIJIMA, Yoshinobu NODASAKA, Junichiro IIDA, Fumio WATARI
    Nano Biomedicine 1 2 151 - 158 2009年12月 [査読有り][通常論文]
  • Kosuke ISHIKAWA, Tsukasa AKASAKA, Yoshinobu NODASAKA, Natsumi USHIJIMA, Masayuki KAGA, Shigeaki ABE, Motohiro UO, Yasutaka YAWAKA, Masaya SUZUKI, Fumio WATARI
    Nano Biomedicine 1 2 109 - 120 2009年12月 [査読有り][通常論文]
  • Shigeaki ABE, Daisuke HAYASHI, Tsukasa AkASAKA, Motohiro UO, Yoshinori KUBOKI, Fumio WATARI, Tomoya TAKADA
    Nano Biomedicine 1 2 143 - 150 2009年12月 [査読有り][通常論文]
  • Xiaoming Li, Hong Gao, Motohiro Uo, Yoshinori Sato, Tsukasa Akasaka, Qingling Feng, Fuzhai Cui, Xinhui Liu, Fumio Watari
    JOURNAL OF BIOMEDICAL MATERIALS RESEARCH PART A 91A 1 132 - 139 2009年10月 [査読有り][通常論文]
     
    Carbon nanotubes (CNTs) have been shown to affect cell behavior. But how and why the CNTs affect potential differentiation of the attached cells has not been largely known. In this study, multiwalled carbon nanotubes (MWNTs) and graphite (GP) were pressed as compacts. Higher ability of CNTs to adsorb proteins, compared with GP, was shown. Myoblastic mouse cells (C2C12) were cultured and the cell responses to the two kinds of compacts were compared in vitro. Meanwhile, we used cell culture on the culture plate as a control. During the conventional culture, significantly better cell attachment, proliferation, and differentiation of cells on the MWNTs were found. To confirm the hypothesis that the larger amount of protein adsorbed on the CNTs was crucial for this, we made the compacts adsorb more proteins in culture medium with 50% fetal bovine serum (FBS) before cell culture. With the adsorption of the proteins in advance, the increments of the total-protein/DNA and alkaline phosphatase (ALP)/DNA for the MWNTs was respectively as about 11 times and 18 times as the increments of those for GP and the control at both day 4 and day 7. Therefore, the CNTs might induce cellular functions by adsorbing more proteins, which indicated that the CNTs might be a candidate for scaffold material for tissue engineering. (C) 2008 Wiley Periodicals, Inc. J Biomed Mater Res 91A: 132-139, 2009
  • Shigeaki Abe, Chika Koyama, Motohiro Uo, Tsukasa Akasaka, Yoshinobu Kuboki, Fumio Watari
    JOURNAL OF NANOSCIENCE AND NANOTECHNOLOGY 9 8 4988 - 4991 2009年08月 [査読有り][通常論文]
     
    Micro- and nano-sized materials have received much attention regarding their biocompatibility and toxicity. To understand the influence of such materials on animals, it is very important to determine their internal distribution behavior. In this study, the biodistributions of Pt and TiO(2) micro- and nano-sized particles in mice were estimated and visualized by X-ray scanning analytical microscopy and inductively coupled plasma-atomic emission spectroscopy. We also determined the effect of particle size, difference between metal and oxide, and time dependence for the distributions, because the biodistribution depends upon both the chemical character of materials and the size of particles. The results of the present study indicated that the difference in chemical character had a greater effect than did particle size. We predict that X-ray scanning analytical microscopy will be a useful method for studying biodistribution of micro- and nano-sized particles, because this method requires no labeling or treatment of the target particles.
  • Xiaoming Li, Xinhui Liu, Wei Dong, Qingling Feng, Fuzhai Cui, Motohiro Uo, Tsukasa Akasaka, Fumio Watari
    Journal of Biomedical Materials Research Part B: Applied Biomaterials 90B 2 503 - 509 2009年08月 [査読有り][通常論文]
     
    In this study, the previously reported porous three-dimensional poly(L-lactic acid) (PLLA) scaffolds reinforced by the chitin fibers (PLLA/CF) with and without the link were evaluated in vitro. Firstly, pH value of the phosphate buffered saline lixiviums of the PLLA/CF with different content of the chitin fibers was measured to get an appropriate content of the chitin fibers in the PLLA/CF. Then, the cell functions (attachment, proliferation, alkaline phosphatase per unit cell, total protein per unit cell, and osteonectin, osteopontin, and osteocalcin gene expression) of human osteoblast-like cells (SaOS2) cultured on the PLLA/CF with the link, PLLA/CF without the link and PLLA scaffold were compared. The results showed that the link treatment did not significantly influence the pH value of the lixiviums of the scaffolds, 30% volume content might be an appropriate content of the chitin fibers in PLLA/CF scaffold to keep the pH value of the lixiviums of the scaffolds between 7.0 and 7.2 during the lixiviation time of 16 weeks, the PLLA/CF scaffold was significantly better for the attachment, proliferation, differentiation, and mineralization of the osteoblast than PLLA, the link treatment did not significantly influence these cells activities, which further suggested that PLLA/CF with the link treatment might be an appropriate scaffold for tissue engineering. (C) 2009 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 90B: 503-509, 2009
  • Eri Hirata, Motohiro Uo, Hiroko Takita, Tsukasa Akasaka, Fumio Watari, Atsuro Yokoyama
    JOURNAL OF BIOMEDICAL MATERIALS RESEARCH PART B-APPLIED BIOMATERIALS 90B 2 629 - 634 2009年08月 [査読有り][通常論文]
     
    Carbon nanotubes (CNTs) have attractive biochemical properties such as strong cell adhesion and protein absorption, which are very useful for a cell cultivation scaffold. In this study, We prepared a multiwalled carbon nanotube-coated collagen sponge (MWCNT-coated sponge) to improve the surface properties of the collagen sponge, and its cell culturing properties Were examined. The suface of the collagen sponge was homogeneously coated with MWCNTs by dispersion. MC3T3-E1 cells were cultured on and inside the MWCNT-coated sponge. The DNA content on the MWCNT-coated sponge after 1 week of culture was significantly higher than on an uncoated collagen sponge (p < 0.05). There was no significant difference between the estimated ALP activity normalized by DNA quantity on the MWCNT-coated sponge and that on the uncoated collagen sponge which is well known as one of the best scaffolds for cell cultivation. In addition, the MWCNT-coated surface shows strong cell adhesion. Therefore, the MWCNT-coated collagen sponge is expected to be a useful 3D scaffold for cell cultivation. (C) 2009 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 90B: 629-634, 2009
  • F. Watari, T. Akasaka, Xiaoming Li, M. Uo, A. Yokoyama
    Frontiers of Materials Science in China 3 2 169 - 173 2009年06月 [査読有り][通常論文]
     
    Carbon nanotubes (CNT) have a unique structure and feature. In the present study, cell proliferation was performed on the scaffolds of single-walled CNTs (SWCNT), multiwalled CNTs (MWCNT), and on graphite, one of the representative isomorphs of pure carbon, for the sake of comparison. Scanning electron microscopy observation of the growth of osteoblast-like cells (Saos2) cultured on CNTs showed the morphology fully developed for the whole direction, which is different from that extended to one direction on the usual scaffold. Numerous filopodia were grown from cell edge, extended far long and combined with the CNT meshwork. CNTs showed the affinity for collagen and proteins. Proliferated cell numbers are largest on SWCNTs, followed by MWCNTs, and are very low on graphite. This is in good agreement with the sequence in the results of the adsorbed amount of proteins and expression of alkaline phosphatase activity for these scaffolds. The adsorption of proteins would be one of the most influential factors to make a contrast difference in cell attachment and proliferation between graphite and CNTs, both of which are isomorphs of carbon and composed of similar graphene sheet crystal structure. In addition, the nanosize meshwork structure with large porosity is another property responsible for the excellent cell adhesion and growth on CNTs. CNTs could be the favorable materials for biomedical applications. © 2009 Higher Education Press and Springer-Verlag GmbH.
  • Shigeaki ABE, Tetsu YONEZAWA, Tsukasa AKASAKA, Motohiro UO, Fumio UCHIDA, Fumio WATARI
    Nano Biomedicine 1 1 70 - 74 2009年06月 [査読有り][通常論文]
  • Fumio Watari, Noriyuki Takashi, Atsuro Yokoyama, Motohiro Uo, Tsukasa Akasaka, Yoshinori Sato, Shigeaki Abe, Yasunori Totsuka, Kazuyuki Tohji
    JOURNAL OF THE ROYAL SOCIETY INTERFACE 6 3 S371 - S388 2009年06月 [査読有り][通常論文]
     
    Nanosizing effects of materials on biological organisms was investigated by biochemical cell functional tests, cell proliferation and animal implantation testing. The increase in specific surface area causes the enhancement of ionic dissolution and serious toxicity for soluble, stimulative materials. This effect originates solely from materials and enhances the same functions as those in a macroscopic size as a catalyst. There are other effects that become prominent, especially for non-soluble, biocompatible materials such as Ti. Particle size dependence showed the critical size for the transition of behaviour is at approximately 100 mm, 10 mm and 200 nm. This effect has its origin in the biological interaction process between both particles and cells/tissue. Expression of superoxide anions, cytokines tumour necrosis factor-alpha and interleukin-1 beta from neutrophils was increased with the decrease in particle size and especially pronounced below 10 mm, inducing phagocytosis to cells and inflammation of tissue, although inductively coupled plasma chemical analysis showed no dissolution from Ti particles. Below 200 nm, stimulus decreases, then particles invade into the internal body through the respiratory or digestive systems and diffuse inside the body. Although macroscopic hydroxyapatite, which exhibits excellent osteoconductivity, is not replaced with natural bone, nanoapatite composites induce both phagocytosis of composites by osteoclasts and new bone formation by osteoblasts when implanted in bone defects. The progress of this bioreaction results in the conversion of functions to bone substitution. Although macroscopic graphite is non-cell adhesive, carbon nanotubes (CNTs) are cell adhesive. The adsorption of proteins and nano-meshwork structure contribute to the excellent cell adhesion and growth on CNTs. Non-actuation of the immune system except for a few innate immunity processes gives the non-specific nature to the particle bioreaction and restricts reaction to the size-sensitive phagocytosis. Materials larger than cell size, approximately 10 mm, behave inertly, but those smaller become biointeractive and induce the intrinsic functions of living organisms. This bioreaction process causes the conversion of functions such as from biocompatibility to stimulus in Ti-abraded particles, from non-bone substitutional to bone substitutional in nanoapatite and from non-cell adhesive to cell adhesive CNTs. The insensitive nature permits nanoparticles that are less than 200 nm to slip through body defence systems and invade directly into the internal body.
  • Xiaoming Li, Hong Gao, Motohiro Uo, Yoshinori Sato, Tsukasa Akasaka, Shigeaki Abe, Qingling Feng, Fuzhai Cui, Fumio Watari
    BIOMEDICAL MATERIALS 4 1 015995  2009年02月 [査読有り][通常論文]
     
    Osteogenic maturation of the osteoblast is crucial for bone formation. In this study, multi-walled carbon nanotubes (MWCNTs) and graphite (GP) were pressed as compacts. The greater ability of carbon nanotubes to adsorb proteins, compared with graphite, was shown. Human osteoblast-like SaoS2 cells were cultured and the cell response to the two kinds of compacts was compared in vitro. Meanwhile, we used cell culture on the culture plate as a control. Assays for osteonectin, osteopontin and osteocalcin gene expression, total protein (TP) amount, alkaline phosphatase activity (ALP) and DNA of cells cultured on the samples were done. During the conventional culture, significantly higher osteonectin, osteopontin and osteocalcin gene expression level, ALP/DNA and TP/DNA on carbon nanotubes were found. To confirm the hypothesis that the larger amount of specific proteins adsorbed on the carbon nanotubes was crucial for this, the compacts were pre-soaked in culture medium having additional recombinant human bone morphogenetic protein-2 (rhBMP-2) before cell culture. Compared with GP, osteonectin, osteopontin and osteocalcin gene expression level, ALP/DNA and TP/DNA of the cells tested increased more on the MWCNTs after the compacts were pre-soaked in the culture medium with rhBMP-2. The results indicated that the carbon nanotubes might induce osteogenic maturation of the osteoblast by adsorbing more specific proteins.
  • Tsukasa Akasaka, Fumio Watari
    ACTA BIOMATERIALIA 5 2 607 - 612 2009年02月 [査読有り][通常論文]
     
    Capture of bacteria with flexible carbon nanotubes (CNTs) was done in vitro. Bundles of single-walled carbon nanotubes (SWCNTs) or multi-walled carbon nanotubes (MWCNTs) was mixed with Streptococcus mutans. Precipitation assays and colony-forming unit formation assays showed free S. mutans in the solution was significantly decreased by the addition of the CNTs. Observation of the precipitate by scanning electron microscopy showed bacterial adhesion to CNTs. It has been shown that CNTs of different diameters have significantly different effects on the precipitation efficiency, and the manners in which they capture the cells are different. We found that MWCNTs (diameter of approximately 30 nm) had the highest precipitation efficiency, which was attributable to both their adequate dispersibility and aggregation activity. From observations by scanning electron microscopy, bundles of SWCNTs and thin MWCNTs (diameter of approximately 30 nm), which were moderately flexible, were easily wound around the curved surface of S. mutans. Bare CNTs having high adhesive ability could be useful as biomaterials, e.g., as tools for the elimination of oral pathogens at the nano-level. (c) 2008 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.
  • Tsukasa Akasaka, Atsuro Yokoyama, Makoto Matsuoka, Takeshi Hashimoto, Shigeaki Abe, Motohiro Uo, Fumio Watari
    BIO-MEDICAL MATERIALS AND ENGINEERING 19 2-3 147 - 153 2009年 [査読有り][通常論文]
     
    Carbon nanotubes (CNTs) exhibit excellent cell proliferation properties, which can serve as a scaffold for cell culturing. However, there are only a few reports on adhesion of osteoblast-like cells to a CNT sheet. In this study, we investigated adhesion of osteoblast-like cells to single-walled carbon nanotube (SWNT) and multi-walled carbon nanotube (MWNT) sheets and compared these adhesions with that on a cell culture polystyrene dish by using a cell adhesion test and a scanning electron microscope. The MWNT sheets exhibited faster adhesion of cells at an initial stage than SWNT sheets and cell culture polystyrene dish. The number of attached cells on the MWNT sheets seemed to be greater than on SWNT sheets and cell culture polystyrene. Moreover, the MWNT sheets exhibited both high speed and good capacity for cell adhesion. However, the surface of the MWNT sheets was such that it facilitated cell adherence but hindered the spreading of the attached cells. Interestingly, cell adhesion to CNT sheets was significantly influenced by pre-coating with serum. These results indicate that CNT sheets would play an important role in adsorption of serum proteins, which would consequently facilitate cell adhesion, and that the MWNT sheets have a high cell adhesiveness.
  • Makoto Matsuoka, Tsukasa Akasaka, Takeshi Hashimoto, Yasunori Totsuka, Fumio Watari
    BIO-MEDICAL MATERIALS AND ENGINEERING 19 2-3 155 - 162 2009年 [査読有り][通常論文]
     
    Silicone rubbers are widely used as tissue implants because of their flexibility and chemical stability. However, they have limited cellular adhesiveness and may cause problems in the long term. In this study, a coating of carbon nanotubes (CNTs) was applied to silicone rubber to improve its cellular adhesiveness. Scanning electron micrograph of this coating revealed that CNTs had formed a densely packed meshwork; the R-a values and protein adsorption capacity were enhanced. Although the contact angle did not change after coating, it decreased after immersion into a culture medium. After cultivation for 6 d, while Saos-2 cells were hardly observed on untreated silicone, the cells proliferated on CNT-coated silicone. Thus, CNT coating might be a simple and effective solution to problems associated with silicone implants.
  • Tsukasa Akasaka, Keiko Nakata, Motohiro Uo, Fumio Watari
    BIO-MEDICAL MATERIALS AND ENGINEERING 19 2-3 179 - 185 2009年 [査読有り][通常論文]
     
    Recent studies have shown that carbon nanotubes (CNTs) can be used as biomedical materials because of their unique properties. CNTs effect nucleation of hydroxyapatite, because of which considerable interest has been generated regarding the use of CNTs in dentistry. However, there are only a few reports on the use of CNTs as dental materials. In this study, we investigated the changes induced in the surfaces of tooth slices by the application of a coating of CNTs by observing CNT-coated tooth slices both macroscopically as well as under a scanning electron microscope. Further, we investigated the effect of CNT coating on the tensile bond strength of dentin adhesives. CNTs adhered easily to the tooth surfaces when tooth slices were suspended in a CNT-dispersed solution. Interestingly, it was observed that CNTs selectively adhered to the surfaces of dentin and cementum, possibly by adhering to their exposed collagen fibers. In addition, the CNT coating did not affect the tensile bond strength of dentin adhesives. These results indicate that coating of the teeth with CNTs can be a possible application of CNTs as dental materials.
  • F. Watari, S. Abe, I. D. Rosca, A. Yokoyama, M. Uo, T. Akasaka, N. Takashi, Y. Totsuka, E. Hirata, M. Matsuoka, K. Ishikawa, S. Itoh, Y. Yawaka
    BIOCERAMICS 21 396-398 569 - 572 2009年 [査読有り][通常論文]
     
    Nanoparticles may invade directly into the internal body through the respiratory or digestive system and diffuse inside body. The behavior of nanoparticles in the internal body is also essential to comprehend for the realization of DDS. Thus it is necessary to reveal the internal dynamics for the proper treatments and biomedical applications of nanoparticles. In the present study the plural methods with different principles such as X-ray scanning analytical microscope (XSAM), MRI and Fluorescent microscopy were applied to enable the observation of the internal diffusion of micro/nanoparticles in the (1) whole body level, (2) inner organ level and (3) tissue and intracellular level. Chemical analysis was also done by ICP-AES for organs and compared with the results of XSAM mapping.
  • Motohiro Uo, Tomoka Hasegawa, Tsukasa Akasaka, Isao Tanaka, Fuminori Munekane, Mamoru Omori, Hisamichi Kimura, Reiko Nakatomi, Kohei Soga, Yasuo Kogo, Fumio Watari
    BIO-MEDICAL MATERIALS AND ENGINEERING 19 1 11 - 17 2009年 [査読有り][通常論文]
     
    Three types of multiwalled carbon nanotube (MWCNT) monoliths without any binders were obtained by spark plasma sintering (SPS) treatment at 2000 degrees C under 80 MPa sintering pressure. Three MWCNTs with different diameters: thin (empty set20-30 nm, CNT Co., Ltd., Korea), thick (empty set100 nm, Nano Carbon Technologies Co., Ltd., Japan) and spherical thin (empty set20-30 nm, granulated diameter = 1-3 mu m, Shimizu Corporation, Japan) were employed for SPS. SEM observation confirmed that these materials maintained the nanosized tube microstructure of raw CNT powder after SPS treatment. The densest monolith was prepared with the spherical MWCNTs. The mechanical properties of this material were estimated by the dynamic hardness test. The elastic modulus of the monolith did not depend on the difference of MWCNTs, but the hardness of spherical MWCNTs was higher than that of thick MWCNTs. The high density and hardness of the spherical MWCNTs were caused by the high packing density during the SPS process because of its spherical granulation. Thus, the spherical MWCNTs were most useful for the MWCNT monolith preparation with the SPS process and its application as a bone substitute material and a bone tissue engineering scaffold material was suggested.
  • Shigeaki Abe, Ikuhiro Kida, Mitsue Esaki, Tsukasa Akasaka, Motohiro Uo, Toshiaki Hosono, Yoshinori Sato, Balachandran Jeyadevan, Yoshinori Kuboki, Manabu Morita, Kazuyuki Tohji, Fumio Watari
    BIO-MEDICAL MATERIALS AND ENGINEERING 19 2-3 213 - 220 2009年 [査読有り][通常論文]
     
    Nano-sized particles have received much attention in view of their varied application in a wide range of fields. For example, magnetite (Fe(3)O(4)) nanoparticles have been investigated for various medical applications. In this study, we visualized the distribution of administered magnetic nanoparticles in mice using both X-ray scanning analytical microscopy (XSAM) and magnetic resonance imaging (MRI). After administration, the nanoparticles were rapidly dispersed via the blood circulation, and reached the liver, kidney and spleen. Using the XSAM and MRI methods in a complementary fashion, the biodistribution of nano-sized magnetite particles was successfully visualized.
  • Shigeaki Abe, Chila Koyama, Mitsue Esaki, Tsukasa Akasaka, Motohiro Uo, Yoshinori Kuboki, Manabu Morita, Fumio Watari
    BIO-MEDICAL MATERIALS AND ENGINEERING 19 2-3 221 - 229 2009年 [査読有り][通常論文]
     
    We observed the internal diffusion behavior of inorganic micro/nano particles through oral administration. By oral exposure, the fed particles were absorbed through the digestive system then reached some organs after internal diffusion in the body. For example, TiO(2) particles fed to mice were detected in the lung, liver, and spleen after 10 days of feeding. Whereas, the absorption efficiency was extremely low compared with intravenous injection. In a comparison of the simple amount of administration, oral exposure required 10(2) times or more amount by intravenous injection for detection by an X-ray scanning analytical microscope. During dental treatment, micro/nano particles from tooth or dental materials would generate in the oral cavity, and some of the particles had a possibility to be swallowed, absorbed through the digestive system, and then diffuse into the body. However, our results suggest that biocompatible microparticles that are naturally taken orally affect animals only rarely because of the low absorption efficiency.
  • Michiko Terada, Shigeaki Abe, Tsukasa Akasaka, Motohiro Uo, Yoshimasa Kitagawa, Fumio Watari
    BIO-MEDICAL MATERIALS AND ENGINEERING 19 1 45 - 52 2009年 [査読有り][通常論文]
     
    Carbon nanotubes (CNTs) have excellent chemical durability, mechanical strength and electrical properties. Therefore, there is interest in CNTs for not only electrical and mechanical applications, but also biological and medical applications. We coated titanium, a common material for dental implants, with multiwalled carbon nanotubes (MWCNTs). First, titanium was aminated and covered with collagen. Then, the carboxylated MWCNTs were coated onto the collagen attached to the titanium plate. The collagen-coated titanium plate had a homogeneous MWCNT coating, which showed strong attachment to the titanium surface as a thin layer. The surface roughness was significantly increased with the MWCNT coating. MC3T3-E1 cells were cultured on the MWCNT-coated Ti plate, and showed good cell proliferation and strong cell adhesion. Therefore, the MWCNT coating for titanium could be useful for improvement of cell adhesion on titanium implants.
  • Michiko Terada, Shigeaki Abe, Tsukasa Akasaka, Motohiro Uo, Yoshimasa Kitagawa, Fumio Watari
    Dental Materials Journal 28 1 82 - 88 2009年 [査読有り][通常論文]
     
    Carbon nanotubes (CNTs) are one of the most interesting nanomaterials because of their excellent characteristics. In this study, a transparent CNTs coating for cell culture dishes was developed and its properties for cell culture were estimated. Carboxylated multiwalled carbon nanotubes (MWCNTs) were dispersed in aqueous sodium cholate solution and applied on a collagen type I-coated cell culture dish (cover glass). The dish surface was homogeneously covered by MWCNTs without aggregation. The MWCNT-coated dish was slightly gray and had good transparency, so conventional optical microscopic observation of the cells on the MWCNT-coated dish was possible. Rat osteoblast-like cells cultured on the MWCNT-coated dish showed slightly lower viability and proliferation compared to the collagen-coated dish. The cell adhesion on the MWCNT-coated dish was much higher than that on the collagen-coated dish. Therefore, MWCNT-coating for dishes will be a useful new material for cell culture.
  • Motohiro Uo, Kiyotaka Asakura, Kazuchika Tamura, Yasunori Totsuka, Shigeaki Abe, Tsukasa Akasaka, Fumio Watari
    CHEMISTRY LETTERS 37 9 958 - 959 2008年09月 [査読有り][通常論文]
     
    Ti, a Ni-Ti alloy, and stainless steel (SUS304) were implanted in rat soft tissues for 6 months. The chemical states of the component elements contained in the tissues surrounding those metallic implants were analyzed by fluorescence X-ray absorption line structure (XAFS). Titanium oxide and a nickel aquo complex, which Would be Caused by the erosion of Ti and Ni, were detected in the Surrounding tissues. The concentrations of Ti and Ni in the tissues were 1-3 and 300 ppm, respectively. Thus, the dissolved conditions and concentrations of those practical metallic materials in vivo were successfully confirmed.
  • Wei Wang, Atsuro Yokoyama, Susan Liao, Mamoru Omori, Yuhe Zhu, Motohiro Uo, Tsukasa Akasaka, Fumio Watari
    MATERIALS SCIENCE & ENGINEERING C-BIOMIMETIC AND SUPRAMOLECULAR SYSTEMS 28 7 1082 - 1086 2008年08月 [査読有り][通常論文]
     
    A carbon nanotube (CNT) monolith without any binders was obtained by spark plasma sintering (SPS) treatment at 1100 degrees C under 40 MPa pressure. Transmission electron microscope results confirmed that this material maintained the nanosized tube microstructure of raw CNT powder after SPS treatment. The density and mechanical properties of this material were similar to cancellous bone. The material was implanted in subcutaneous tissue and femurs of rats and tissue samples were analyzed by histopathology at I and 4 weeks after surgery. Although some foreign body giant cells were seen around the CNTs, no severe inflammatory response such as necrosis was observed, and CNT implants were surrounded by newly formed bone in the femur. The study provides the first in vivo testing evidence that pure bulk carbon nanotubes are not a strongly inflammatory substance and have no toxicity for bone regeneration. Our study is the first successful experiment to consolidate CNTs without binders, and may provide an effective method for CNT monolith synthesis, as well as demonstrating that a binderless carbon nanotube material with a strength matching that of bone could be a candidate bone Substitute material and a bone tissue engineering scaffold material. (c) 2007 Elsevier B.V. All rights reserved.
  • Fumio Watari, Kazuchika Tamura, Atsruro Yokoyama, Kenichiro Shibata, Tsukasa Akasaka, Bunshi Fugetsu, Kiyotaka Asakura, Motohiro Uo, Yasunori Totsuka, Yoshinori Sato, Kazuyuki Tohji
    Biological Aspects and Structure Formation 127 - 144 2008年03月 [査読有り][通常論文]
     
    ISBN: 9783527619443
  • Determination of administered organic/inorganic particles in mice
    Shigeaki Abe, Chika Koyama, Mitsue Esaki, Ikuhiro Kida, Iosif Rosca, Tsukasa Akasaka, Motohiro Uo, Manabu Morita, Yoshinori Sato, Balachandran Jeyadevan, Yoshinori Kuboki, Kazuyuki Tohji, Fumio Watari
    8th World Biomaterials Congress 2008 1 460  2008年
  • Tsukasa Akasaka, Fumio Watari
    FULLERENES NANOTUBES AND CARBON NANOSTRUCTURES 16 2 114 - 125 2008年 [査読有り][通常論文]
     
    We have demonstrated that multi-walled carbon nanotubes (MWNTs) coated with a carbohydrate-carrying polymer for use as biological recognition signals can be easily prepared by a non-covalent method via hydrophobic interactions. Fluorescence observation by confocal laser scanning microscopy showed that the carbohydrate-carrying polymers were densely localized around the MWNTs. To evaluate biological recognition affinity, interactions of the MWNTs with lectins were examined by binding tests. The resultant MWNTs were found to acquire a selective binding affinity to the corresponding lectin without a non-specific interaction. On the other hand, bare MWNTs non-specifically interacted with lectins. These results showed that the MWNTs coated with a carbohydrate-carrying polymer have biological recognition signals. Modification of carbon nanotubes with various carbohydrate chains will be a useful protocol for molecular designs of biomaterials, nanoarchitecture and biosensors.
  • Fumio Watari, Shigeaki Abe, Chika Koyama, Atsuro Yokoyama, Tukasa Akasaka, Motohiro Uo, Makoto Matsuoka, Yasunori Totsuka, Mitsue Esaki, Manabu Morita, Tetsu Yonezawa
    JOURNAL OF THE CERAMIC SOCIETY OF JAPAN 116 1349 1 - 5 2008年01月 [査読有り][通常論文]
     
    To clarify the effect of micro/nanosizing of materials onto biological organism, the particle size dependence of reaction of cells and tissue as investigated by both biochemical cell functional test and animal implantation test. Especially for nanoparticles the behavior of invasion and internal diffusion inside body was visualized using an XSAM (X-ray Scanning Analytical Microscope). The increase of specific surface area is usually counted as nanosizing effect which causes the enhancement of chemical reactivity and therefore toxicity of materials such as carcinogenicity found in 500 nm Ni particles for the long term implantation in the soft tissue of rat. Even biocompatible materials such as Ti and TiO2 shows stimulus with the decrease of particle size. They cause phagocytosis to cells and inflammation to tissue when the size of particles is below 3 mu m. For the size below 50 nm, they may invade into the internal body through the respiratory or digestive system and diffuse inside body. After compulsory exposure test of 30 nm TiO2 particles through the respiratory system, the Ti mapping by XSAM showed the internal diffusion inside the whole body. Nanoparticles injected from caudal vein diffused with time course to lung, liver and spleen. The uptake of 30 nm TiO2 particles through the digestive system and diffusion into these organs was also confirmed. These phenomena observed in biocompatible or bioinert materials are the nonspecific, physical particle and shape effects which occur independent of materials. Nanoparticles might be the objects whose existence has not been assumed by the living body defense system.
  • Susan Liao, Furnio Watari, Yuhe Zhu, Motohiro Uo, Tsukasa Akasaka, Wei Wang, Guofu Xu, Fuzhai Cui
    DENTAL MATERIALS 23 9 1120 - 1128 2007年09月 [査読有り][通常論文]
     
    Objective. The purpose of this paper was to investigate the in vitro biodegradation of a guided tissue regeneration composite membrane, nano-carbonated hydroxyapatite/collagen/ poly(lactic - co- glycolic acid) (nCHAC/PLGA). Especially for periodontal therapy, the functional graded material (FGM) nCHAC/PLGA membrane was prepared that consisted of three layers with 8 wt% nCHAC + PLGA/4 wt% nCHAC + PLGA/PLGA, where one face of the membrane is porous, thereby allowing cell growth thereon and the opposite face of the membrane smooth, thereby inhibiting cell adhesion. Methods. For evaluation, in vitro degradation specimens of nCHAC/PLGA were immersed into artificial saliva solution at 37 degrees C for 1, 2, 4, 8 and 12 weeks to detect the weight loss over the period, and set pure PLGA membrane as control to compare the degraded behaviors. pH value and calcium concentration of the residual solution were measured, and morphology change was investigated by scanning electron microscopy (SEM). Results. During the experimental period in vitro, the whole shape of the membrane could be kept for 4 weeks, after that it became powder at between 8 and 12 weeks. The results demonstrated that weight loss increased continuously with a reduction in mass of 23.1% after 4 weeks and 88% after 12 week for the nCHAC/PLGA three FGM layers composite membrane. The calcium concentration in the residual solution showed a significant increase after 4 weeks, which referred to the nano-carbonated hydroxyapatite degradation. Moreover, the pH value in the solution of the nCHAC/PLGA membrane was a little higher than that of the pure PLGA membrane, which demonstrated the possible neutralization effect from nCHAC composite for the acid outcome of PLGA in the solution. The pore structure of 8 wt% nCHAC + PLGA was enlarged on the porous surface, while the nonporous surface of pure PLGA also showed a small porous structure after increased time. Significance. Degradation of the composite membrane is appropriate for practical periodontal repair. Moreover, the new mineral formation on the surface of the composite membrane referred to the possible positive effect in vivo for new bone tissue regeneration. (C) 2006 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.
  • Naofumi Aoki, Tsukasa Akasaka, Fumio Watari, Atsuro Yokoyama
    DENTAL MATERIALS JOURNAL 26 2 178 - 185 2007年03月 [査読有り][通常論文]
     
    To investigate the dependence of blocompatibility of carbon materials on crystal structure with the aim of developing biomedical applications, single-(SW) and multi-walled (MW) carbon nanotubes (CNTs) were employed as scaffolds for cell culture and compared with graphite (GP). SaOS2 cells were used to investigate the properties and response of osteoblast-like cells. Polycarbonate membranes (PC) coated with CNTs by vacuum filtration formed a meshwork nanostructure. Cells grown on CNTs greatly extended in all directions. In terms of cell proliferation, alkaline phosphatase (ALP) activity, and protein adsorption on the substrates, CNTs showed better results than PC and GP. SW showed the best cell proliferation and total ALP. These favorable results might be attributed to the structure of CNTs and the affinity of CNTs toward proteins, thereby suggesting that CNTs could be potential scaffold materials for cell culture.
  • Motohiro Uo, Kiyotaka Asakura, Atsuro Yokoyama, Makoto Ishikawa, Kazuchika Tamura, Yasunori Totsuka, Tsukasa Akasaka, Fumio Watari
    DENTAL MATERIALS JOURNAL 26 2 268 - 273 2007年03月 [査読有り][通常論文]
     
    Tissues contacting Ti dental implants were subjected to X-ray absorption fine structure (XAFS) analysis to examine the chemical state of Ti transferred from the placed implant into the surrounding tissue. Nine tissues that contacted pure Ti cover screws for several months were excised in a second surgery whereby healing abutments were set. Six tissues that surrounded implants retrieved due to their failure were also excised. Ti distributions in the excised specimens were confirmed by X-ray scanning analytical microscopy (XSAM), and the specimens were subjected to fluorescence XAFS analysis to determine the chemical states of the low concentrations of Ti in the tissues surrounding Ti dental implants. Ti mostly existed in the metallic state and was considered to be debris derived from the abrasion of implant pieces during implant surgery. Oxidized forms of Ti, such as anatase and rutile, were also detected in a few specimens-and existed in either a pure state or mixed state with metallic Ti. It was concluded that the existence of Ti in the tissue did not cause implant failure. Moreover, the usefulness of XAFS for analysis of the chemical states of rarely contained elements in biological tissue was demonstrated.
  • Motohiro Uo, Hanako Kachi, Tsukasa Akasaka, Fumio Watari, Yoshinori Sato, Kenichi Motomiya, Kazuyuki Tohji
    FULLERENES NANOTUBES AND CARBON NANOSTRUCTURES 15 4 303 - 309 2007年 [査読有り][通常論文]
     
    Rare earth elements ( Y, La, Ce, Nd, Gd and Dy) encapsulated by carbon nanocapsules ( CNCs) were synthesized and their purity and air oxidation stability were estimated. The purity was estimated as the rare earth carbide content. Gd- and Dy-encapsulating CNCs had higher than 30 wt% and others 15 to 20 wt%. Encapsulated rare earth carbide was oxidized by heating in air at 400 degrees C or higher. This suggested oxidation damage to the graphene capsules of CNCs.
  • Tsukasa Akasaka
    the journal of Dental Engineering 161 161 35 - 36 (一社)日本歯科理工学会 2007年 [査読無し][通常論文]
  • Naofumi Aoki, Atsuro Yokoyama, Yoshinobu Nodasaka, Tsukasa Akasaka, Motohiro Uo, Yoshinori Sato, Kazuyuki Tohji, Fumio Watari
    Journal of Bionanoscience 1 1 14 - 16 2007年 [査読有り][通常論文]
  • Naofumi Aoki, Atsuro Yokoyama, Yoshinobu Nodasaka, Tsukasa Akasaka, Motohiro Uo, Yoshinori Sato, Kazuyuki Tohji, Fumio Watari
    CHEMISTRY LETTERS 35 5 508 - 509 2006年05月 [査読無し][通常論文]
     
    The morphology of cells cultured on carbon nanotube (CNTs) scaffolds was investigated using a confocal laser scanning microscope (CLSM) and a scanning electron microscope (SEM) and it was shown that the cells extended strikingly in all directions and numerous filopodia extended far from the cells.
  • T Akasaka, F Watari, Y Sato, K Tohji
    MATERIALS SCIENCE & ENGINEERING C-BIOMIMETIC AND SUPRAMOLECULAR SYSTEMS 26 4 675 - 678 2006年05月 [査読有り][通常論文]
     
    Apatite coating on carbon-nanotubes (CNTs) was done with a biomimetic coating method. The multi-walled CNTs (MWNTs) of curled shape with about 30 nm in diameter were immersed for 2 weeks in the simulated body fluid. Observation by scanning electron microscopy (SEM) showed the formation of apatite on the MWNTs surface. The clusters of spherules consisting of needle-shaped apatite crystallites were massively grown on the aggregated MWNTs. The crystallites of 100 nm in width and 200-500 nm in length were grown perpendicularly to the longitudinal direction and radially originating from a common center of a single MWNT. Thus, the architecture of crystalline apatite at nano-scale levels could be produced by simple method and the MWNT may be acting as core for initial crystallization of apatite. (c) 2005 Elsevier B.V. All rights reserved.
  • Y. Sato, M. Ohtsubo, B. Jeyadevan, K. Tohji, K. Motomiya, R. Hatakeyama, G. Yamamoto, M. Omori, T. Hashida, K. Tamura, T. Akasaka, M. Uo, A. Yokoyama, F. Watari
    Physical Review B 73 144103-1 - 144103-11 2006年04月 [査読有り][通常論文]
  • S Liao, K Tamura, YH Zhu, W Wang, M Uo, T Akasaka, FZ Cui, F Watari
    JOURNAL OF BIOMEDICAL MATERIALS RESEARCH PART A 76A 4 820 - 825 2006年03月 [査読無し][通常論文]
     
    The impact of biodegraded nano-hydroxyapatite/collagen (nHAC) composite and nano-hydroxyapatite/ collagen/poly(L-lactic acid) (nHAC/PLA) scaffold composite on neutrophils reaction was evaluated in vitro. Neutrophils were separated from human peripheral blood of healthy subjects. The nHAC and nHAC/PLA materials were immersed in the D-Hanks' Balanced Salt Solution (DHBSS) for I day, 7 days and 2, 4, 8 weeks (37 degrees C) as testing solution, which mixed with the neutrophils for 1 h. Both of the nHAC and nHAC/PLA materials were shown the same cell Survival rate as blank control, but the lactate dehydrogenase (LDH) and tumor necrosis factor alpha (TNF-alpha) released from the neutrophils were increased significantly after the 2 weeks in nHAC sample. The possible reason relied on the high concentration of calcium due to the quick biodegradation of the nHAC material. Before 2 weeks, the LDH value of nHAC/PLA is higher than that of nHAC sample that corresponded to the initial PLA degradation in vitro. This study provided the biocompatibility test of neutrophils other than common methods, such as osteoblastic cells for biomimetic materials. Moreover, it demonstrated the calcium concentration stimulating effect for cytokine release from neutrophils. (c) 2005 Wiley Periodicals, Inc.
  • Functions of Sugar chains in oral cavity
    Tsukasa Akasaka, Fumio Watari
    The journal of dental engineering 159 26 - 28 2006年 [査読無し][通常論文]
  • Glyco-technology and Dentistry
    Tsukasa Akasaka, Motohiro Uo, Fumio Watari
    Hokkaido Journal of Dental Science 27 1 41 - 44 2006年 [査読無し][通常論文]
  • Naofurni Aoki, Atsuro Yokoyama, Yoshinobu Nodasaka, Tsukasa Akasaka, Motohiro Uo, Yoshinori Sato, Kazuyuki Tohji, Furnio Watari
    JOURNAL OF BIOMEDICAL NANOTECHNOLOGY 1 4 402 - 405 2005年12月 [査読有り][通常論文]
     
    The developments of nanoscale substance such as carbon nanotubes (CNTs) for medical applications have attracted a great deal of attention. In the present study, an attempt was made to development of CNTs as scaffolds for cell culture. CNT scaffolds were formed on polycarbonate membranes by vacuum filtration and cell proliferation and morphology were investigated using a scanning electron microscopy. Osteoblast cells (bone-forming cell) on CNTs showed excellent proliferation with extension of cell morphology in all directions. Numerous filopodia were extended from cells toward the inside of fibrous aggregates of CNTs and tightly bound to scaffolds. These results suggest that CNTs can be used as scaffolds with excellent affinity for cell adhesion.
  • S Liao, W Wang, M Uo, S Ohkawa, T Akasaka, K Tamura, FZ Cui, F Watari
    BIOMATERIALS 26 36 7564 - 7571 2005年12月 [査読有り][通常論文]
     
    Functional graded materials (FGM) provided us one new concept for guided tissue regeneration (GTR) membrane design with graded component and graded structure where one face of the membrane is porous thereby allowing cell growth thereon and the opposite face of the membrane is smooth, thereby inhibiting cell adhesion in periodontal therapy. The goal of the present study was to develop a three-layered graded membrane, with one face of 8% nano-carbonated hydroxyapatite/collagen/poly(lactic-co-glycolic acid) (nCHAC/PLGA) porous membrane, the opposite face of pure PLGA non-porous membrane, the middle layer of 4% nCHAC/ PLGA as the transition through layer-by-layer casting method. Then the three layers were combined well with each other with flexibility and enough high mechanical strength as membrane because the three layers all contained PLGA polymer that can be easily used for practical medical application. This high biocompatibility and osteoconductivity of this biodegraded composite membrane was enhanced by the nCHAC addition, for the same component and nano-level crystal size with natural bone tissue. The osteoblastic MC3T3-E1 cells were cultured on the three-layered composite membrane, the primary result shows the positive response compared with pure PLGA membrane. (c) 2005 Elsevier Ltd. All rights reserved.
  • ID Rosca, F Watari, M Uo, T Akaska
    CARBON 43 15 3124 - 3131 2005年12月 [査読無し][通常論文]
     
    The oxidation of MWCNTs in nitric acid was monitored using sample weight, Raman spectrum. solubility. morphology and alignment. The influence of the acid concentration, temperature and oxidation duration on the monitored parameters was assessed. A new method, based on optical microscopy is proposed for the determination of MWCNT solubility in concentrated aqueous-suspensions. The investigations revealed that the Solubility is determined not only by the functional groups on the MWCNT, but also by the functionalized amorphous carbon generated during the digestion of the nanotubes, High solubility (20-40 mg/ml) is obtained only after prolonged exposure (24-48 h) in concentrated acid (>= 60%). But in these conditions 60-90% of the MWCNTs are lost. Furthermore the MWCNTs are strongly fragmented and covered by amorphous carbon after 48 h of oxidation. It was found that the solubility correlates well with the area ratio of the G and D bands from tile Raman spectrum, SEM examination of the MWCNT films showed extended alignment after 24 h of oxidation. (C) 2005 Elsevier Ltd. All rights reserved.
  • Size separation of carbon nanotubes for biomedical application
    Kazuyuki Tohji, Yoshinori Sato, Atsuro Yokoyama, Ken-ichiro Shibata, Shin-ichi Ogino, Kazuchika Tamura, Tsukasa Akasaka, Motohiro Uo, Kenichi Motomiya, Balachandran Jeyadevan, Rikizo. Hatakeyama, Fumio Watari
    Proceedings of International Symposium on EcoTopia Science 2005 1 273 - 276 2005年08月08日 [査読有り][通常論文]
  • Water-soluble hat-stacked carbon nanofibers for biomedical applications
    Yoshinori Sato, Ken-ichiro Shibata, Fugetsu Bunshi, Atsuro Yokoyama, Shin-ichi Ogino, Kazuchika Tamura, Tsukasa Akasaka, Motohiro Uo, Kenichi Motomiya, Balachandran Jeyadevan, Rikizo. Hatakeyama, Fumio Watari, Kazuyuki Tohji
    Proceedings of International Symposium on EcoTopia Science 2005 1 277 - 280 2005年08月08日 [査読有り][通常論文]
  • Y Sato, K Shibata, H Kataoka, S Ogino, F Bunshi, A Yokoyama, K Tamura, T Akasaka, M Uo, K Motomiya, B Jeyadevan, R Hatakeyama, F Watari, K Tohji
    MOLECULAR BIOSYSTEMS 1 2 142 - 145 2005年07月 [査読有り][通常論文]
     
    Water-soluble H-CNFs modified with a carboxyl group possessed the ability to induce TNF-alpha, whereas CHAPS-treated H-CNFs possessed significantly greater activity and were also found to activate NF-kappa B reporter activity, to a significantly greater level than H-CNFs; furthermore the functional group modified or coated on the surface of H-CNFs was a significant cytotoxic factor that affected cell activation.
  • Y Sato, A Yokoyama, K Shibata, Y Akimoto, S Ogino, Y Nodasaka, T Kohgo, K Tamura, T Akasaka, M Uo, K Motomiya, B Jeyadevan, M Ishiguro, R Hatakeyama, F Watari, K Tohji
    MOLECULAR BIOSYSTEMS 1 2 176 - 182 2005年07月 [査読有り][通常論文]
     
    Carbon nanotubes (CNTs) are single- or multi-cylindrical graphene structures that possess diameters of a few nanometers, while the length can be up to a few micrometers. These could have unusual toxicological properties, in that they share intermediate morphological characteristics of both fibers and nanoparticles. To date, no detailed study has been carried out to determine the effect of length on CNT cytotoxicity. In this paper, we investigated the activation of the human acute monocytic leukemia cell line THP-1 in vitro and the response in subcutaneous tissue in vivo to CNTs of different lengths. We used 220 nm and 825 nm-long CNT samples for testing, referred to as "220-CNTs" and "825-CNTs", respectively. 220-CNTs and 825-CNTs induced human monocytes in vitro, although the activity was significantly lower than that of microbial lipopeptide and lipopolysaccharide, and no activity appeared following variation in the length of CNTs. On the other hand, the degree of inflammatory response in subcutaneous tissue in rats around the 220-CNTs was slight in comparison with that around the 825-CNTs. These results indicated that the degree of inflammation around 825-CNTs was stronger than that around 220-CNTs since macrophages could envelop 220-CNTs more readily than 825-CNTs. However, no severe inflammatory response such as necrosis, degeneration or neutrophil infiltration in vivo was observed around both CNTs examined throughout the experimental period.
  • M Uo, K Asakura, A Yokoyama, K Tamura, Y Totsuka, T Akasaka, F Watari
    CHEMISTRY LETTERS 34 6 776 - 777 2005年06月 [査読有り][通常論文]
     
    X-ray absorption fine structure (XAFS) analysis was applied for the analysis of titanium (Ti) in the human oral mucosa in contact with Ti dental implants from two different patients. Metallic Ti and an oxide (anatase) were detected. The metallic Ti was distributed in particulate form, which would be debris formed by the abrasion during the operation of implantation. The possibility of using XAFS for the analysis of the chemical state of rarely contained elements in biological tissue was suggested.
  • T Akasaka, F Watari
    CHEMISTRY LETTERS 34 6 826 - 827 2005年06月 [査読有り][通常論文]
     
    Calcium phosphate (Ca-P) coating on carbon nanotubes (CNTs) was done with a biomimetic coating method. The multi-walled CNTs (MWNTs) were immersed for 2 weeks in the simulated body fluid. Observation by scanning electron microscopy showed that needle shape of Ca-P crystallites at nano-scale levels were massively grown on MWNTs. Thus the nano-architecture of crystalline Ca-P on MWNT surface could be produced by simple method and the MWNT may be acting as core for initial crystallization.
  • Preparation and Biocompatibility of Sintered Carbon Nanotube Disk
    M. Ohtsubo, Y. Sato, B. Jeyadevan, K. Tohji, R. Hatakeyama, G. Yamamoto, M. Ohmori, T. Hashida, K. Tamura, T. Akasaka, M. Uo, A. Yokoyama, F. Watari
    Fullerenes and Nanotubes –Materials for the New Chemical Frontier-Fullerenes (The Electrochemical Society, Inc.) PV2004-12 243 - 248 2005年03月01日 [査読無し][招待有り]
  • A Yokoyama, Y Sato, Y Nodasaka, S Yamamoto, T Kawasaki, M Shindoh, T Kohgo, T Akasaka, M Uo, F Watari, K Tohji
    NANO LETTERS 5 1 157 - 161 2005年01月 [査読有り][通常論文]
     
    The tissue response to hat-stacked carbon nanofibers (H-CNFs) was evaluated. H-CNFs were implanted in the subcutaneous tissue of rats. Histological and ultrastructural investigations were carried out by transmission electron microscopy. Although many macrophages and foreign body giant cells were seen around H-CNFs, no severe inflammatory response such as necrosis was observed. Some H-CNFs were observed in lysosomal vacuoles of phagocytes. These results showed that H-CNFs were not strong prophlogistic substances and were englobed in vivo.
  • W. Wang, A. Yokoyama, M. Omori, S. Liao, Y. Zhu, T. Akasaka, M. Uo, F. Watari
    International Journal of Oral and Maxillofacial Surgery 34 1 83  2005年 [査読有り][通常論文]
  • B Fugetsu, S Satoh, T Shiba, T Mizutani, YB Lin, N Terui, Y Nodasaka, K Sasa, K Shimizu, R Akasaka, M Shindoh, KI Shibata, A Yokoyama, M Mori, K Tanaka, Y Sato, K Tohji, S Tanaka, N Nishi, F Watari
    ENVIRONMENTAL SCIENCE & TECHNOLOGY 38 24 6890 - 6896 2004年12月 [査読有り][通常論文]
     
    Multiwalled carbon nanotubes (MWCNTs) were used as the active elements for the first time for affinity-based elimination of ionic dyes. MWCNTs were encapsulated in cross-linked alginate (ALG) microvesicles using Ba2+ as the bridging ion. The Ba2+-alginate matrix constitutes a cage which holds the physically trapped MWCNTs. The cage carries negative charges on its surface. The cage restricts the access of anions of large molecular weight, such as humic acids, because of electrostatic repulsion. The cage also restricts the access of colloids of large size, because of size exclusion. Ionic dyes partition into the cage and then are captured by MWCNTs probably on the basis of van der Waals interactions occurring between the hexagonally arrayed carbon atoms in the graphite sheet of MWCNTs and the aromatic backbones of the dyes. As a result of these interactions the target species, namely, the ionic dyes, are eliminated efficiently by the MWCNTs of Ba2+-ALG/MWCNT composite adsorbents. The adsorptive capacities for elimination of acridine orange, ethidium bromide, eosin bluish, and orange G (the model species used for this study) were found as high as 0.44, 0.43, 0.33, and 0.31 mumol, respectively, for 1.0 mg of the caged MWCNTs. Adsorptive experiments with carbon nanofibers and activated carbons as the adsorbents were also performed. The MWCNT-based adsorbents provided the best capability for the affinity-based elimination of these targeted species. Biocompatibility experiments performed in vitro and in vivo provided promising results, suggesting potential applications of the caged MWCNTs in in situ environmental remediation.
  • K Iwasaki, S Ohkawa, JD Rosca, M Uo, T Akasaka, F Watari
    DENTAL MATERIALS JOURNAL 23 4 593 - 599 2004年12月 [査読有り][通常論文]
     
    The distortion of laser welded titanium plates was assessed for different operating conditions of the laser welding device, and with different welding parameters (in terms of weld point and prewelding). In this study, Nd : YAG laser welding device was used to join the titanium plates. The results showed that distortion increased stepwise after each welding point along the welding zone (one-side welding), but decreased consecutively as the welding proceeded on the second side of the weld (two-side welding). In the case of one-side welding, the dependence of distortion on current and spot diameter presented maxima - due to changes in the welding pool characteristics. For two-side weld the same parameters exercised little influence on its distortion recovery, due to the effect of solidified weld pools from the first side. Current and spot diameter determined the weld pool, which in turn regulated distortion based on shrinkage. Four-point prewelding significantly decreased the final distortion for both one- and two-side welds. Alternating two-side welding of prewelded assembly showed lower distortion than a classic two-side weld.
  • MK Yamada, M Uo, S Ohkawa, T Akasaka, F Watari
    JOURNAL OF BIOMEDICAL MATERIALS RESEARCH PART B-APPLIED BIOMATERIALS 71B 1 7 - 15 2004年10月 [査読有り][通常論文]
     
    A three-dimensional analyzer installed in a scanning electron microscope was used to evaluate the morphology and surface roughness using noncontact profilometry. Observations were carried out on the enamel and dentin surface irradiated by three different lasers: Nd:YAG (wavelength 1.06 mum), Er:YAG (2.94 mum), and CO2 (10.6 mum). Spectroscopic analysis was done by Raman spectroscopy for nonirradiated and laser-irradiated surfaces. The lasers were applied perpendicularly to vertically sectioned and polished human extracted caries-free molars. The tooth was sectioned at each cavity for cross-section analysis after laser irradiation. Irradiation by Nd:YAG and CO2 lasers of the enamel surface showed an opaque white color, different from dentin where the surface turned black. The Er:YAG laser induced no changes in color of the dentin. Numerous cracks associated with thermal stress were observed in the CO2 laser-irradiated dentin. Noncontact surface profile analysis of Er:YAG laser-irradiated enamel and dentin showed the deepest cavities, and direct cross-sectional observations of them showed similar cavity outlines. The CO2 laser-irradiated dentin had the least surface roughness. Raman spectroscopic analysis showed that fluorescence from the laser-irradiated tooth was generally greater than from nonirradiated teeth. Bands in dentin attributed to organic collagen matrix were lost after Nd:YAG and CO2 laser irradiation, and a broad peak due to amorphous carbon appeared. The Er:YAG laser-irradiated dentin showed no sign of a carbon band and had more suitable results for dental ablation. Noncontact surface profile analysis was effective to evaluate the structural change in the tooth in the microarea of study after laser irradiation. (C) 2004 Wiley Periodicals, Inc.
  • K Iwasaki, S Ohkawa, M Uo, T Akasaka, F Watari
    MATERIALS TRANSACTIONS 45 4 1140 - 1146 2004年04月 [査読無し][通常論文]
     
    The performance of laser welding of Ti and two dental precious alloys (Ag-Pd-Au, Au-Pt-Ag) was investigated by mechanical testing and microscopic observation, using butt joint as design. The laser irradiation was done using a commercial Nd:YAG dental laser-welding device in a single pulse mode with output currents 150, 200, 250, 300 A, spot diameters 0.6, 0.9, 1.2 mm, and pulse duration 10 ms. The average welding fracture strength of the dissimilar metals were 108.9 and 137.2 MPa for Ti and Ag-Pd-Au alloy, and Au-Pt-Ag alloy, respectively. The average welding fracture strength of the same metals were 594.9. 648.8 and 312.9 MPa for Ti, Ag-Pd-Au alloy and Au-Pt-Ag alloy. The hardness increased in weld zone, compared with the base metals. Penetration depths were affected by the welding conditions such as the output currents and spot diameters. The welding cracks and porosity were observed in microstructures of the welds. Mapping by EPMA showed the remarkable heterogeneity of the component metals concerned. The welding cracks, porosities and granular precipitates similar to metallic compounds in the weld zones were suggested as the cause for lower welding fracture strength in the dissimilar welds, compared with the similar metals.
  • MK Yamada, M Uo, S Ohkawa, T Akasaka, A Watari
    MATERIALS TRANSACTIONS 45 4 1033 - 1040 2004年04月 [査読無し][通常論文]
     
    A newly developed scanning electron microscope (SEM) installed with a 3D analyzer and a confocal laser scanning microscope (CLSM) were used to evaluate the images and surface profiles of enamel and dentin after CO2 laser irradiation. The surface roughness (Ra) was measured and the results were correlated with a stylus profilometer. Raman analysis was done and the laser irradiation effect after acid etching was also examined. Human intact extracted teeth were vertically sectioned. The laser was applied perpendicularly to non-etched and 35% phosphoric acid-etched teeth. The enamel resulted in a crater-like surface. The material was removed in the places where hydroxyapatite crystallites ran parallel to the irradiated surface. Non-etched dentin showed little change after irradiation, with some cracks mostly in the peritubular dentin; whereas in acid-etched dentin there was evaporation of collagen and melting of hydroxyapatite. The SEM and CLSM showed similar profiles and different image contrast. Ra levels obtained by the SEM and CLSM were similar to that obtained with the stylus profilometer. Raman analysis showed that hands of organic collagen matrix on dentin were lost and broad peaks due to carbon were observed. Both the SEM and CLSM provided non-contact evaluation of structural changes of teeth by laser irradiation through surface analysis in selected microareas, which was not possible using the stylus profilometer. Dentin showed more structural changes on the acid-etched surface by irradiation than on the nonetched one.
  • K Tamura, N Takashi, T Akasaka, ID Roska, M Uo, Y Totsuka, F Watari
    BIOCERAMICS, VOL 16 254-2 919 - 922 2004年 [査読有り][通常論文]
     
    The cytotoxicity of micro/nano particles in Ti, TiO and carbon nanotube was investigated by in vitro biochemical analyses using human neutrophils. The particles smaller and larger than the neutrophils were used to determine the relationship between cell and particle size with respect to cytotoxicity. As the particle size decreased, the cell survival rate was decreased and, with the good corresponding relation to this, the value of lactate dehydrogenase (LDH), which is the indication of cell disruption, was increased. The release of superoxide anion showed the increasing tendency. Proinflammatory cytokines were detected distinctly for 3 mum or smaller particles and very little in more than 10 mum, which is closely related to the phagocytosis by neutrophils. ICP elemental analysis showed that the dissolution from Ti particles was below detection limit. Micro and nano particles stimulated the cell reactions according to the results of the human neutrophil functional tests. As the particle size was smaller, the inflammation was pronounced. The fine particles less than 3 mum caused distinctly the inflammation in the surrounding tissue. All these results indicated that the cytotoxicity was induced due to the physical size effect of particles, which is different from the ionic dissolution effect. The clinical phenomenon confirmed the result obtained in vitro cell tests. The neutrophils stimulated by fine particles may cause the inflammatory cascade and harm the surrounding tissue.
  • Preparation of carbon nanotubes for biomedical applications: Size-separation and their biocompatibilities
    K. Tohji, Y. Akimoto, Y. Sato, B. Jeyadevan, K. Tamura, T. Akasaka, M. Uo, A. Yokoyama, K. - I. Shibata, F. Watari
    Proceedings - Electrochemical Society PV 2004-12 178 - 183 2004年 [査読有り][通常論文]
  • Y. Sato, M. Ohtsubo, B. Jeyadevan, K. Tohji, K. Motomiya, R. Hatakeyama, G. Yamamoto, M. Omori, T. Hashida, K. Tamura, T. Akasaka, M. Uo, A. Yokoyama, F. Watari
    Proc. SPIE 623 - 627 2004年01月 [査読有り][通常論文]
  • Y. Sato, Y. Akimoto, B. Jeyadevan, K. Motomiya, R. Hatakeyama, K. Tamura, T. Akasaka, M. Uo, A. Yokoyama, K. Shibata, F. Watari, K. Tohji
    Proc. SPIE 13 - 17 2004年01月 [査読有り][通常論文]
  • Kazunori Matsuura, Tsukasa Akasaka, Miki Hibino, Kazukiyo Kobayashi
    Chem.Lett 28 3 247 - 248 2003年05月 [査読有り][通常論文]
  • 微粒子と生体反応
    亘理文夫, 田村一央, 高師則行, 宇尾基弘, 赤坂 司, 戸塚靖典, 古月文志, 佐藤義倫, 田路和幸
    ナノ学会会報 2 1 34 - 37 2003年 [査読有り][通常論文]
  • 赤坂司, 前川宜彦, 西村紳一郎
    Cellul Commun 9 1 13 - 17 2002年03月 [査読有り][通常論文]
  • T Akasaka, K Matsuura, K Kobayashi
    BIOCONJUGATE CHEMISTRY 12 5 776 - 785 2001年09月 [査読有り][通常論文]
     
    Block-type oligonucleotide-glycopolymer conjugates bearing alpha -mannosides and beta -galactosides were prepared by coupling 5 ' -thiol-modified oligonucleotides with iodoacetamidated glycopolymers that were synthesized by telomerization. The conjugates minimally affected the DNA conformation and melting behavior of the duplex. Their self-organization via hybridization with the half-sliding complementary oligonucleotides produced graft-type conjugates or macromolecular gapped DNA duplexes grafted with glycopolymers at regular intervals, which was confirmed using size exclusion chromatography and electrophoresis. The binding affinity of block-type and self-organized graft-type conjugates to lectins was investigated using fluorometry. The affinity of the graft-type duplex assembly bearing mannosides to Con A was approximately 2 times stronger than that of block-type single-stranded or double-stranded conjugates with full complementary oligonucleotides. The organization strategy of DNA-glycopolymer conjugates might be useful for constructing novel glyco-clusters and also for developing a new methodology for gene therapy.
  • K Matsuura, T Akasaka, M Hibino, K Kobayashi
    BIOCONJUGATE CHEMISTRY 11 2 202 - 211 2000年03月 [査読有り][通常論文]
     
    Stable and lectin-recognizable DNA-carbohydrate conjugates were prepared by diazo coupling of lactose and cellobiose derivatives to fragmented salmon testes DNA. The diazo coupling is suggested to take place selectively to guanine bases since the amount of lactose moiety introduced was directly proportional to the G content of various DNAs with different G contents. According to the CD spectra, the conjugates bearing carbohydrate less than 25% content kept a typical B-type conformation similar to native DNA. The conjugates possessed higher melting temperature and stronger nuclease resistance both to exo- and endonucleases than native DNA. Gel shift assay and fluorescence binding assay showed that the DNA-lactose conjugates were specifically bound to galactose-specific lectin RCA(120) with strong binding affinity (K-a = 10(4)-10(5) M-1) due to glycoside cluster effect. This facile method will be a useful protocol of molecular design for cell-targeted gene therapy.
  • T Akasaka, K Matsuura, N Emi, K Kobayashi
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 260 2 323 - 328 1999年07月 [査読有り][通常論文]
     
    Oligosaccharide-plasmid DNA conjugates were synthesized simply and effectively via the diazocoupling method. Plasmids (pUC19, pTRI-beta-actin, and pEGFP-C1) were treated with an N-beta-lactoside-substituted diazonium salt to yield diazocoupling products with degree of substitutions of 2.5-3.1 mol% of overall nucleobases. The lactose-pUC19 conjugate was found to resist restriction enzymes more strongly than the nonconjugated plasmid DNA and to acquire a strong binding affinity to galactose-specific lectin RCA(120). The diazocoupling modification of pTRI-beta-actin plasmid DNA little influenced in vitro transcription with T7 RNA polymerase. When lactose-pEGFP-C1 conjugate was transfected to baby hamster kidney (BHK) cells by means of cationic lipids, transduced gene was expressed in BHK cells similarly with the nonconjugated pEGFP-C1. The modification of plasmid DNA with carbohydrate enhanced the resistance to restriction enzymes and developed a strong binding affinity to galactose-specific lectin. (C) 1999 Academic Press.
  • T Yoshida, T Akasaka, Y Choi, K Hattori, B Yu, T Mimura, Y Kaneko, H Nakashima, E Aragaki, M Premanathan, N Yamamoto, T Uryu
    JOURNAL OF POLYMER SCIENCE PART A-POLYMER CHEMISTRY 37 6 789 - 800 1999年03月 [査読有り][通常論文]
     
    Anti-HIV (human immunodeficiency virus) active polymethacrylates having pendant sulfated oligosaccharides were synthesized, and the relationship between structures and biological activities of the polymethacrylates was examined. Acetylated 1-O-methacryloyl maltoheptaoside (MA-AcM7) was polymerized with AIBN as an initiator to give polymethacrylates having a pendant acetylated maltoheptaose in every repeating unit, poly(MA-AcM7)s. After hydroxyl groups were recovered by deacetylation, the polymethacrylates having maltoheptaose units, poly(MA-M7)s, were sulfated to give polymethacrylates having sulfated maltoheptaose side-chains, poly(MA-SM7)s, with degrees of sulfation of 1.1 to 2.7 (maximum, 3.0). These polymethacrylates including sulfated oligosaccharides exhibited low anti-HIV activities represented by the 50% protecting concentration (EC50) in the range of 15-62 mu g/mL and low blood anticoagulant activities around 10 unit/mg (standard dextran sulfate, 22.7 unit/mg). The anti-HIV activity increased with increasing degree of sulfation to reach EC50 of 15-16 mu g/mL. In addition, copolymerization of MA-AcM7 with methyl methacrylate (MMA) and subsequent sulfation gave polymethacrylates consisting of various proportions of highly sulfated maltoheptaose and MMA units. It was revealed that the anti-HIV activity increased with decreasing proportion of the sulfated oligosaccharide moiety and that a copolymethacrylate having 22 mol % of sulfated maltoheptaose units (DS = 3.0) had a high anti-HIV activity in the EC50 of 0.3 mu g/mL. The blood anticoagulant activity increased slightly from 9 to 18 unit/mg with decreasing proportion of the sulfated maltoheptaose units. These results suggested that the biological activities were influenced strongly by the spatial distance between sulfated oligosaccharide substituents in the polymethacrylate main chain. Distinction and conformation of the oligosaccharide side chains also played an important role. (C) 1999 John Wiley & Sons, Inc.

MISC

書籍等出版物

  • 応用物理学会・ナノインプリント技術研究会 (担当:分担執筆範囲:13.6 マイクロ・ナノパターンの歯科バイオ応用)
    オーム社 2019年11月 (ISBN: 9784274224485) xiii, 781p
  • スタンダード歯科理工学 第７版
    赤坂 司 (担当:分担執筆範囲:15 接着・合着用材料)
    学建書院 2019年03月 (ISBN: 9784762456145)
  • スタンダード歯科理工学 第６版
    赤坂 司 (担当:分担執筆範囲:１４ 接着・合着用材料)
    2016年03月
  • 動物細胞培養法
    赤坂 司 (担当:分担執筆範囲:第２章、第３節「カーボンナノチューブ薄膜上での細胞培養」)
    株式会社技術情報協会 2014年

所属学協会

  • 代用臓器・再生医学研究会   日本バイオマテリアル学会   ナノ・バイオメディカル学会   日本歯科理工学会   Japanese Society of Carbohydrate Research   

共同研究・競争的資金等の研究課題

  • 日本学術振興会：科学研究費助成事業
    研究期間 : 2024年04月 -2027年03月 
    代表者 : 赤坂 司, 横山 敦郎, 吉村 善隆, 宮治 裕史, 牛島 夏未, 星 淡子
  • 日本学術振興会：科学研究費助成事業
    研究期間 : 2023年04月 -2027年03月 
    代表者 : 赤坂 司, 横山 敦郎, 吉村 善隆, 宮治 裕史, 牛島 夏未, 星 淡子
  • 日本学術振興会：科学研究費助成事業
    研究期間 : 2023年04月 -2026年03月 
    代表者 : 吉田 靖弘, 赤坂 司
  • 歯の表面に貼り付けて使うセメント質と歯根膜の形状と成分を疑似した人工歯根膜の開発
    日本学術振興会：科学研究費助成事業 挑戦的研究(萌芽)
    研究期間 : 2022年06月 -2025年03月 
    代表者 : 赤坂 司
  • 日本学術振興会：科学研究費助成事業 基盤研究(C)
    研究期間 : 2022年04月 -2025年03月 
    代表者 : 西田 絵利香, 宮治 裕史, 大矢根 綾子, 中村 真紀, 赤坂 司
     
    アパタイト2層構造化による２段階薬剤放出性第４世代スキャフォールド構築のため，初年度はまず1層目の構築・評価を行った． 1層目として，アパタイトの難溶化（溶解性低下）と抗菌性の期待される生体微量元素（フッ素）を種々の濃度（0～1000 uM）でCaP過飽和溶液に添加し，スキャフォールド上にフッ素含有アパタイト層を作製した． その結果，スキャフォールド内部および外部表面への均一なフッ素含有アパタイト層の生成を確認し，過飽和溶液へのフッ素添加濃度の増加により生成するアパタイト層のフッ素置換率（F/P比で評価）向上が認められた． 成膜後のスキャフォールドを生食に浸漬して溶解性等を評価したところ，過飽和溶液へのフッ素添加濃度の増加に伴い，生食中へのアパタイトの溶解量（7日後）は減少した．アパタイト結晶にフッ化物イオンが取り込まれることにより，層の難溶化したと考えられる，また，生食中へのフッ素溶出量も増加した． 以上より，フッ化物イオンをバイオミメティックコーティング法によりアパタイト層に取り込むことに成功した．フッ化物濃度の増加とともにアパタイト層の溶解度が低下し，減少した．
  • 日本学術振興会：科学研究費助成事業 基盤研究(C)
    研究期間 : 2020年04月 -2023年03月 
    代表者 : 吉田 靖弘, 赤坂 司, 玉井 美保
     
    本課題では幹細胞として胚性幹細胞（ES細胞）および乳歯歯髄幹細胞（SHED）に注目し、各種微細パターン上での細胞挙動を検討した。マウスES細胞をマイクロパターン上で培養し、三胚葉の分化マーカーを免疫染色した。多くのパターンでは外胚葉＞内胚葉＞中胚葉の順で多かったが、2μm以下のパターンの一部では内胚葉への分化の割合が少し上昇した。次に、SHEDのマイクロパターン上での細胞挙動を検討した。細胞接着では付着数に差がなかったが、細胞増殖や石灰化では特定のパターンにて促進が観察された。以上より、パターンのデザインにより幹細胞の挙動を制御できる可能性が示唆された。
  • 日本学術振興会：科学研究費助成事業 基盤研究(B)
    研究期間 : 2019年04月 -2023年03月 
    代表者 : 赤坂 司, 横山 敦郎, 吉村 善隆, 牛島 夏未, 宮治 裕史, 星 淡子, 加我 公行, 玉井 美保
  • 日本学術振興会：科学研究費助成事業 基盤研究(B)
    研究期間 : 2019年04月 -2022年03月 
    代表者 : 横山 敦郎, 赤坂 司, 山本 悟, 平田 恵理, 高野 勇太, 湯田坂 雅子, 岡崎 俊也
     
    本年度の研究の概要を以下に示す． 1.骨再生誘導法への単層カーボンナノチューブ膜の応用 単層カーボンナノチューブ(SWCNTs)とヒアルロン酸を用いてGuided Bone Regeneration (GBR)用膜(SWCNT膜)を開発し，in vivoにおける周囲組織の反応と骨形成を観察した．また，in vitroにおけるSWCNT膜上でマウス頭蓋冠骨芽細胞様細胞(MC3T3-E1)，マウス胎児線維芽細胞(NIH/3T3)を培養し，それぞれの細胞増殖について検討した．10週齢雄性Wistar 系ラットの頭蓋冠に直径７mmの骨欠損部を形成後，SWCNT膜で被覆した．術後８週で周囲組織とともに摘出し，マイクロCTを用いて観察するとともに，非脱灰標本を製作して組織学的に検索した．SWCNT膜で被膜した骨欠損部には骨形成を示す不透過象が観察された．SWCNT膜周囲には炎症反応は観察されず，膜上面は薄い線維性結合組織で被覆されていた．膜の下面には，コントロール群(非被覆群)に比較して活発な新生骨の形成が認められた．NIH/3T3の増殖率は，MC3T3-E1の増殖率と比較して低い値を示したことから，線維芽細胞の増殖を抑制する可能性があることが推察された．以上の結果より，今回製作したCNT膜は，遮蔽性を有し，起炎性が低く，骨形成を促進することが示された． 2.光照射によって薬剤を徐放させるカーボンナノホーンの開発 カーボンナノホーン（CNHs）を, ミノサイクリン（MC）を分散媒として水溶液中に分散させると，分散液中ではMCがCNHsに付着していることが明らかになった．一方で，このMC/CNHsに近赤外光を照射し，薬剤の除放量や静菌作用について確認したところ，光照射による薬剤の徐放量は微量であった．この理由として，CNHsに対してMCが強力に付着していることが推測された．
  • 歯科応用へ向けた骨芽細胞機能評価スクリーニング用マイクロ・ナノパターンの開発
    物質・デバイス領域共同研究拠点：2020年度 物質・デバイス領域共同研究拠点
    研究期間 : 2020年03月 -2022年03月 
    代表者 : 赤坂 司, 伊東 駿也, 中川 勝, 宮内 昭浩
  • 生体応用へ向けた細胞機能評価スクリーニング用マイクロ・ナノパターンの試作
    生体医歯工学共同研究拠点：2019年度 生体医歯工学共同研究拠点共同研究プロジェクト
    研究期間 : 2019年06月 -2022年03月 
    代表者 : 赤坂 司, 中川 勝, 伊東 駿也, 宮内 昭浩
  • 歯表面で歯医者自身が直接構築できる微細パターンによる歯周病治療法の開発
    文部科学省：科研費 挑戦的研究（萌芽）
    研究期間 : 2018年04月 -2021年03月 
    代表者 : 赤坂 司
  • 日本学術振興会：科学研究費助成事業 基盤研究(B)
    研究期間 : 2016年04月 -2019年03月 
    代表者 : 横山 敦郎, 赤坂 司, 山本 悟, 平田 恵理, 佐藤 義倫, 東野 史裕, 湯田坂 雅子, 髙田 紗理, 前田 由佳利, 木村 貞仁
     
    本研究では、カーボンナノマテリアル(CNMs)の臨床応用を視野に、カーボンナノホーン(CNHs)の生体内分解性の観察を行い、CNHsへの抗生剤（ミノサイクリン）やグロースファクター(骨形成タンパク)の担持ならびに徐放性制御とその効果を確認した。さらに、CNHsを表面に修飾したチタンを開発した。また、局所に埋入したカーボンナノチューブ(CNTs)の体内動態を追跡し、そのほとんどが局所に留まることを明らかにした。以上の結果、使用用途に応じて多機能化したスマートナノバイオマテリアルの創製と口腔領域への応用の可能性が示唆された。
  • マイクロブラシ状コラーゲンからなる人工歯根膜を持つインプラントの開発
    公益財団法人鈴木謙三記念医科学応用研究財団：平成２９年度調査研究助成金
    研究期間 : 2017年12月 -2018年11月 
    代表者 : 赤坂 司
  • 日本学術振興会：科学研究費助成事業 挑戦的萌芽研究
    研究期間 : 2014年04月 -2017年03月 
    代表者 : 横山 敦郎, 赤坂 司, 山本 悟, 宮治 裕史
     
    熱ナノインプリント法を用いてマイクロ・ナノパターン（Groove構造およびPillar構造）を持つゼラチンシート、チタンシートを作製し、骨芽細胞、線維芽細胞、上皮細胞を培養し、細胞付着挙動について検討した。骨芽細胞、線維芽細胞では、マイクロ・ナノ構造は、接着班の数や細胞接着数を増加させたが、上皮細胞では、特にPillar構造では細胞の接着を抑制した。 以上の結果から、マイクロ・ナノパターン形状は、骨芽細胞、線維芽細胞および上皮細胞における細胞接着、配向性に影響を与えることが示された。
  • 歯肉退縮を治療するバイオミメティック次世代歯科治療の開発
    （公）寿原記念財団：研究助成
    研究期間 : 2016年04月 -2017年03月 
    代表者 : 赤坂 司
  • 文部科学省：科学研究費補助金（基盤研究Ｃ）
    研究期間 : 2013年04月 -2017年03月 
    代表者 : 赤坂 司
  • 日本学術振興会：科学研究費助成事業 基盤研究(B)
    研究期間 : 2013年04月 -2016年03月 
    代表者 : 横山 敦郎, 赤坂 司, 佐藤 義倫, 山本 悟, 滝田 裕子, 湯田坂 雅子, 坂口 紀史, 芝 清隆, 平田 恵理, 進藤 正信
     
    カーボンナノ物質が骨形成に与える影響の解明を目的としたカーボンナノホーン(CNHs)が骨髄間質細胞に与える影響の検討，デンタルインプラント開発を目的としたカーボンナノチューブ(CNTs)とCNHsのチタン表面修飾ならびに開発した材料についての細胞培養と動物実験，さらにCNHsの生体材料としての安全性解明のためのCNHsを局所埋入後の体内移行について検討を行った．その結果、CNHsはOncostatin Mを介して骨髄間質細胞の骨芽細胞への分化を促進すること，CNTsおよびCNHsの表面修飾により骨形成が促進されること，CNHsは局所埋入した場合は他臓器には蓄積されないことが明らかとなった．
  • 歯肉退縮を止めるための北海道産材料による歯科用マイクロ・ナノパターンシートの開発
    秋山記念生命科学振興財団：研究助成<一般>
    研究期間 : 2014年10月 -2015年03月 
    代表者 : 赤坂 司
  • 歯肉退縮を防止する歯表面でのマイクロ・ナノパターン直接形成による新規治療法の開発
    科学技術振興機構(JST)：A-STEP 探索タイプ
    研究期間 : 2014年04月 -2015年03月 
    代表者 : 赤坂 司
  • マイクロ・ナノパターン化バイオマテリアル上での細胞培養
    ナノテクノロジープラットフォーム：平成25年度研究設備の試行的利用
    研究期間 : 2013年04月 -2014年03月 
    代表者 : 赤坂 司
  • 電気刺激可能なナノ表面構造による細胞機能の制御
    武田科学振興財団：医学系研究奨励
    研究期間 : 2011年04月 -2014年03月 
    代表者 : 赤坂 司
  • 日本学術振興会：科学研究費助成事業 特別研究員奨励費
    研究期間 : 2012年 -2013年 
    代表者 : 赤坂 司, 亘理 文夫, CHEN X.
     
    抗癌剤を長間使用すると多剤耐性が発現し、次第に抗癌剤の薬効が減衰する。抗癌剤の作用を保持し癌化学療法の治療効果を上げるには、この多剤耐性を抑制する必要がある。本研究では、代表的なナノマテリアルであるカーボンナノチューブ(CNT)の糖鎖・蛋白質吸着性、細胞膜透過性というユニークな特性に着目し、多剤耐性原因因子の吸着・制御による抗癌治療の高効率化を目的とした。今年度は、2種類のナノカーボン、(1)CNTと(2)ナノダイアモンド(ND)について抗癌薬剤への応用研究を行い、それぞれ新しい進展が得られた。 (1)CNTの多剤耐性抑制効果 : 癌細胞から薬剤を細胞外へ早期に排出させ、抗癌剤の細胞内滞留時間を低減し薬効を減じ、多剤耐性の原因となる因子として細胞膜のポンピング作用を司る蛋白質P-グリコプロテイン(PGP)や多剤耐性関連蛋白質(MRP)がある。CNTを投与すると透過型電子顕微鏡(TEM)観察から細胞内取込が確認され、投与量とともに、m-RNA, PGP総蛋白量、免疫染色のいずれのレベルでもこれらの蛋白量の低下が認められた。これはCNTの抗癌剤の多剤耐性抑制効果のメカニズムの直接的な解明である。 (2)NDのDDS応用のためのステルス性付与 : 現在、開発中のドラッグデリバリーシステム(DDS)においてはターゲット癌組織に至る前に、マクロファージ等の免疫細胞により異物と見なされ捕捉貧食されてれしまうという問題点がある。この問題をいかに回避するかというのは現時点における課題であるが、今回薬剤担体として用いたナノダイアモンドについて表面をポリグリセロール(PG)でコーティングすることにより、異物と認識されず貪食を回避できるステルス性を獲得することが示された。これはDDSの臨床応用にきわめて重要である。
  • 文部科学省：科学研究費補助金(若手研究(B))
    研究期間 : 2010年 -2012年 
    代表者 : 赤坂 司
     
    iPS細胞の機能制御のため、カーボンナノチューブ(CNT)電極を持ったチャンバーを作製した。CNTコートは種類やコート量を制御することによりiPS細胞の未分化培養ができる可能性が示された。インクジェットプリンターでCNTインクを使い繰り返しパターンを印刷することにより、十分な導電性を持ったCNT電極パターンを作製できた。得られたCNT電極により誘電泳動を利用してiPS細胞の配列が可能となった。
  • 文部科学省：科学研究費補助金(基盤研究(B))
    研究期間 : 2010年 -2012年 
    代表者 : 横山 敦郎, 赤坂 司, 佐藤 義倫, 山本 悟, 安田 元昭, 滝田 裕子
     
    b-FGFを共有結合させることにより高機能化した多層カーボンナノチューブおよびスタチンを空間内に担持することにより高機能化したカーボンナノホーンを開発し、動物実験にてその骨形成を検討した。高機能化したカーボンナノマテリアルを用いることにより、通常のカーボンナノマテリアルに比較してより多くの骨形成が観察された。またカーボンナノマテリアルの生体内における長期的な経過についても動物実験によりその生体適合性を確認した。
  • 文部科学省：科学研究費補助金(基盤研究(C))
    研究期間 : 2010年 -2012年 
    代表者 : 加我 正行, 橋本 正則, 赤坂 司
     
    機能性ガラスであるS-PRG緻粒子を添加したシーラント材と充填材料(グラアイオノマーセメントとレジン)から溶出するアルカリ性イオンの種類と溶出量を測定し、それらの酸緩衝能を調べた。硬化した試料を乳酸溶液に浸漬するとイオン溶出効果が発現した。歯質がう蝕になる環境は、う蝕原因菌の酸産生によりpHが5.5以下になるとハイドロキシアパタイトが分解される。ここでアルカリ性イオンを放出する材料により脱灰が抑制され、再石灰誘導が生じる。この作用は材料は本研究で使用したバイオグラス、MTA等にもみられた。これら材料の配合比率を検索しながら再石灰化誘導の実験を行った。象牙質切片と微粒子を添加した実験材料との間に10μmのスペースを作り両端を固定して、人工唾液と蒸留水中に37℃で長期間保存し、象牙質切片表面上の析出物、結晶成分のSEM観察とエックス線回折とレーザーラマン分光光度計による同定を行った。試料から溶出するアルカリ性イオンや抗菌性のイオン、燐酸アパタイトの構成成分のカルシウムが再石灰化誘導に関与してる様子がわかり、人口唾液に浸漬した場合に析出物が6ヶ月目でわずかに観察された。SEM観察によりフラットな円盤状象牙質表面に針状の結晶が観察された。EDXによる析出物の同定から主に燐酸カルシウムと判断できた。長期的な展望に立った研究が必要である。実験試料とヒト抜去歯の接着強度の測定を微小引張試験(マイクロテンサイル)方法で行った。S-PRGを含有したレジンは歯質との接着方法の工夫により安定した接着強度が得られた。
  • 文部科学省：科学研究費補助金(挑戦的萌芽研究)
    研究期間 : 2009年 -2011年 
    代表者 : 宇尾 基弘, 赤坂 司, 阿部 薫明
     
    カーボンナノチューブ(CNTs)は直径=数～数十nmの単結晶に近い繊維状材料であり、格子欠陥が少ないことから材料の理論強度に近い機械的特性を有するため、構造材料としての応用も期待されている。本研究ではCNTsの固化体や複合材料の作製と物性評価を行い、歯科・生体材料への応用を検討した。CNTsの焼結では放電プラズマ焼結により緻密な焼結体が得られ、骨に近い弾性係数を示すことが判明した。またCNTs/コラーゲン複合膜では柔軟で強度のある膜を作成する方法を見いだした。
  • iPS細胞の未分化培養に有効な北海道産材料を用いた培養基材の探索
    ノーステック財団：研究開発助成事業 （タレント補助金）
    研究期間 : 2009年10月 -2010年03月 
    代表者 : 赤坂 司
  • シリコーンゴム剤における細胞培養機能の評価
    全国中小企業団体中央会：ものづくり中小企業製品開発等支援補助金
    研究期間 : 2009年04月 -2010年03月 
    代表者 : 赤坂 司
  • カーボンナノチューブ透明導電性薄膜での近赤外照射による殺菌
    ノーステック財団：研究開発助成事業 （タレント補助金）
    研究期間 : 2008年10月 -2009年03月 
    代表者 : 赤坂 司
  • カーボンナノチューブコート細胞培養用ディッシュの新たな機能探索
    北海道大学：総長室重点配分経費 （若手研究者自立支援）
    研究期間 : 2008年07月 -2009年03月 
    代表者 : 赤坂 司
  • 単層CNTを利用した新規細胞培養方法の開発
    科学技術振興機構(JST)：産学共同シーズイノベーション化事業（顕在化ステージ）
    研究期間 : 2008年04月 -2009年03月 
    代表者 : 赤坂 司
  • 文部科学省：科学研究費補助金(基盤研究(B))
    研究期間 : 2007年 -2009年 
    代表者 : 横山 敦郎, 赤坂 司, 佐藤 義倫, 山本 悟, 安田 元昭, 進藤 正信, 東野 史裕
     
    カーボンナノチューブ(CNTs)を用いた顎骨再建を目的として、本研究ではハニカム構造を持つコラーゲンスポンジ表面にCNTsをコーティングし、細胞培養用三次元スキャホールドを開発した。CNTsコーティングにより、骨芽細胞のスキャホールドへの初期接着は促進され、特にスキャホールド深部での細胞の接着、伸展に有効であることが示された。また、CNTsコーティングスキャホールドを骨内に埋入したところ、良好な骨伝導性が認められたことから顎骨再生への応用の可能性が示唆された。
  • 文部科学省：科学研究費補助金(基盤研究(B))
    研究期間 : 2006年 -2009年 
    代表者 : 宇尾 基弘, 朝倉 清高, 亘理 文夫, 横山 敦朗, 赤坂 司, 阿部 薫明
     
    生体組織中の一部に微量存在する溶出微量金属元素や微小異物などをXAFS法により状態分析を行い、チタン、ニッケル-チタン合金、クロム基合金などの生体内での挙動を明らかにした。歯科・生体材料周囲に存在するチタンは多くの場合、金属状態であり埋入時の摩耗に由来すると推測されたが、一部で二酸化チタンとして存在するものもあり、生体内での溶出・再析出によると考えられた。またニッケル-チタン合金などからのニッケルがイオン状態で溶出していることも確認され、溶出Niの生体への為害性が懸念された。さらに生体内のSeなどの微量元素の状態分析や試作X線微小光学系を用いた微小部XAFS分析についても検討を行い、XAFS法の生物試料分析への有用性を見出した。
  • カーボンナノチューブによるミュータンス菌の除菌および殺菌効果
    ノーステック財団：基盤的研究開発育成事業補助金 （若手研究補助金）
    研究期間 : 2007年10月 -2008年03月 
    代表者 : 赤坂 司
  • 人工複合糖質高分子およびレクチンを用いた口腔内細菌が認識する糖鎖構造の解明
    秋山記念生命科学振興財団：研究助成 （奨励助成）
    研究期間 : 2006年08月 -2007年03月 
    代表者 : 赤坂 司
  • 文部科学省：科学研究費補助金(萌芽研究)
    研究期間 : 2005年 -2006年 
    代表者 : 横山 敦郎, 赤坂 司, 進藤 正信, 山本 悟
     
    本年度においては、多層カーボンナノチューブ(MWCNTs)を吸着することにより作製したスキャホールド上で骨髄細胞をデキサメサゾン(Dex)で骨芽細胞に誘導し、遺伝子発現に及ぼすMWCNTsの影響を検索するとともに、Dex添加と非添加の条件で骨髄細胞を培養したスキャホールドを、細胞を付着させた状態でラットの皮下組織に埋入した。 ラット骨髄細胞のDex誘導後1、3および7日の遺伝子の発現を、MWCNTsを吸着したスキャホールドで培養した場合とポリスチレンディッシュ上で培養した場合について、マイクロアレイを用いて比較検討した。全期間を通してMWCNTs固着スキャホールドで、インターロイキンなどの炎症に関与する遺伝子の発現が強く認められ、1、3日では、Matrix Gra ProteinやInsulin like Growth Factorなどの骨に関連する遺伝子の発現が、また7日ではインテグリンなどの接着に関与する遺伝子の発現が強く認められた。この結果から、MWCNTsは起炎性を有するが、骨髄細胞をより速く骨芽細胞に分化・誘導する可能性が示唆された。骨髄細胞をMWCNTs上でDexを添加して2週間培養した後、ラット皮下組織に6週間埋入すると、骨組織様の構造が認められたが、Dexを添加せずに培養した場合は観察されなかった。これらのことから、Dexを添加しMWCNTs上で骨髄細胞を培養すると骨芽細胞に分化し、皮下組織内で骨組織を形成する可能性が示唆された。 以上の結果から、MWCNTsをスキャホールドに用いると、骨髄細胞を強固にスキャホールドに付着した状態で骨芽細胞への分化を促進し、さらにこのスキャホールドは軟組織内で骨組織を形成する可能性が示された。
  • 文部科学省：科学研究費補助金(若手研究(B))
    研究期間 : 2004年 -2006年 
    代表者 : 赤坂 司
     
    歯周病の原因である歯周病細菌の歯への定着を阻害する歯科用糖鎖ナノ材料の開発を目的とし、カーボンナノチューブやその糖鎖修飾体とし、効率的に歯周病細菌へ結合する歯科用ナノ糖鎖材料を開発する。 糖鎖自体の効果をより明白とするため、ポリスチレンディッシュに様々な糖鎖構造を持つ人工複合糖質高分子を吸着させ、基盤表面へのStreptococcus mutans(S.mutans)の吸着を検討した。その結果、ラクトースやキトビオース構造に対して付着し、さらにキトビオース構造は、S.mutansの増殖を抑制することが明らかとなった。 カーボンナノチューブによる口腔内細菌の捕捉を効率化するため、カーボンナノチューブの二次的な形態(直鎖状および球状)による菌体吸着の影響を検討した結果、両方の形態にて細菌捕捉が効率的に起こるものの、直鎖状のカーボンナノチューブの方がより効率的に菌体を吸着することが分かった。また、カーボンナノチューブにより吸着された細菌は引き続き培養を行うと細菌は増殖する事より、カーボンナノチューブによる相互作用自体は殺菌的ではなく吸着による除菌的効果であることが分かった。 以上の結果より、付着能や増殖抑制能を持つ糖鎖により修飾された直鎖状のカーボンナノチューブ網による細菌の捕捉が効率的であると考えられる。今後、カーボンナノチューブ自体に条件付与にて殺菌能の発現などの更なる効率化・機能付与化が望まれる。
  • 文部科学省：科学研究費補助金(基盤研究(B))
    研究期間 : 2004年 -2006年 
    代表者 : 横山 敦郎, 安田 元昭, 赤坂 司, 進藤 正信, 堀内 留美
     
    本研究においては,カーボンナノチューブ(CNTs)を用いて顎骨再生を図ることを目的とし,CNTsの生体適合性の解明,CNTsを用いた骨芽細胞培養用スキャホールドの作製と細胞培養,および骨補填材としてのCNTs固化体の生体適合性の検討を行った. CNTsをラット皮下組織に埋入し,2年間経時的に組織学的,超微細構造学的に検索した.埋入初期では,周囲に肉芽組織が観察され,一部のCNTsはマクロファージに貪食されており,2年後においては軽度の肉芽腫性炎が認められた.透過型電子顕微鏡による観察では,埋入初期においては細胞内でCNTsの強い凝集が認められたが,経時的に凝集は弱くなる傾向を示した.また,CNTsの特有なチューブ状の構造には,ほとんど変化が認められなかった.これらの結果から,CNTsの起炎性は低く,生体内で安定であることが明らかとなった. CNTsを脱イオン水に分散し,ポリカーボネート膜(PC膜)で吸引ろ過することにより,細胞培養用のスキャホールドを作製した.このスキャホールド上で骨芽細胞様細胞を培養すると,細胞の形態は扁平であり,増殖はPC膜より速く,アルカリフォスファターゼ活性も高い結果が得られた.このスキャホールドをGBR膜として,ラット頭頂骨に作製した骨欠損部に応用すると,コントロールであるPC膜に比較し,骨修復は速い傾向を示した. CNTsをプラズマ放電焼結法により作製したCNTs固化体は,骨に近似した機械的性質を有しており,皮下組織および大腿骨に埋入したところ,優れた生体適合性を示し,骨補填材としての可能性が示された. 以上の結果から,CNTsを用いた顎骨再生療法の可能性が示唆された.
  • Biological applications of carbon nanotubes
    Funded Research
    研究期間 : 2002年 -2004年
  • 文部科学省：科学研究費補助金(基盤研究(B))
    研究期間 : 2002年 -2003年 
    代表者 : 亘理 文夫, 高師 則行, 戸塚 靖則, 宇尾 基弘, 横山 敦郎, 赤坂 司
     
    [目的]粉液法が多用される歯科材料や人工関節骨頭摺動部で発生する摩耗粉等、生体適合性に及ぼす微粒子のサイズ依存性を生化学的手法を導入し細胞機能性試験と組織埋入試験で調べた。 [方法]粒度に分布幅を持つ粒子群から、沈降法により0.5,3,10μmを、限外濾過法により300nm以下の単分散粒子を抽出し、各材料間で同一に揃えた。 [サイズ依存性]ヒト好中球、歯根膜細胞、ラットマクロファージを用いた細胞機能性試験ではTi,Fe,Ni(各500nm;3,10,50,150μm),TiO_2(30,60,300,500nm;2μm),ポリ乳酸(1,13μm)の微粒子径が小さくなるほど細胞生存率が低下し、LDH、活性酸素、サイトカインTNF-α,IL-1β,IL-8,M-CSFの放出は増加した。特に細胞より小さな10μm以下では貧食を誘発し著しく増大した。その刺激性は細胞内毒素に比べると1/100〜1/1000程度オーダーが低い。 [形状依存性]0.5,3,10μmの形状異方性の小さい塊状(平均径)と針状(長径)TiO_2粒子を比較すると、細胞生存率、LDH、活性酸素、IL-1βからは針状のほうが刺激性が高く、TNF-αからは細胞と同等サイズの3,10μmでは刺激性、貧食能が形状に影響を受けるが、細胞よりはるかに小さな0.5μmでは影響が小さい結果が得られた。 [材料依存性]ともに生体親和性で、ICP元素分析でイオン溶出が無視できることを確認したTiと溶出するFeはよく似たサイズ依存性を示した。Niは細胞生存率が低めで、活性酸素、サイトカイン放出量が多く、顕微鏡で細胞の破壊が観察され、材料の為害性の影響と考えられる。 [結論]動物埋入試験でも約100μm以上のTi粒子は生体親和性を示すが、50μm以下では炎症性反応を惹起し、10μm以下になると貧食を誘発し、長期間強い炎症を引き起こす。マクロサイズの生体適合性には通常、溶解イオンとして発現する材質依存的な化学的効果が支配的であるが、μm〜nmになるとbioactive,bioinert材料でも、微粒子と細胞・組織の相対的なサイズ関係に由来する、材質非特異的な物理的サイズ効果が顕在化する。

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