研究者データベース

須永 隆文(スナガ タカフミ)
獣医学研究院 獣医学部門 臨床獣医科学分野
助教

基本情報

所属

  • 獣医学研究院 獣医学部門 臨床獣医科学分野

職名

  • 助教

学位

  • 博士(獣医学)(2012年03月 北海道大学大学院獣医学研究科)

科研費研究者番号

  • 90649112

J-Global ID

研究キーワード

  • 獣医整形外科   

研究分野

  • ライフサイエンス / 獣医学

職歴

  • 2019年02月 - 現在 北海道大学 助教

所属学協会

  • 日本軟骨代謝学会   日本獣医麻酔外科学会   

研究活動情報

論文

  • Ekkapol Akaraphutiporn, Eugene C Bwalya, Sangho Kim, Takafumi Sunaga, Ryosuke Echigo, Masahiro Okumura
    The Journal of veterinary medical science 82 8 1209 - 1218 2020年08月28日 
    Pentosan polysulfate (PPS) is a semi-synthetic sulfated polysaccharide compound which has been shown the benefits on therapeutic treatment for osteoarthritis (OA) and has been proposed as a disease modifying osteoarthritis drugs (DMOADs). This study investigated the effects of PPS on cell proliferation, particularly in cell cycle modulation and phenotype promotion of canine articular chondrocytes (AC). Canine AC were treated with PPS (0-80 µg/ml) for 24, 48 and 72 hr. The effect of PPS on cell viability, cell proliferation and cell cycle distribution were analyzed by MTT assay, DNA quantification and flow cytometry. Chondrocyte phenotype was analyzed by quantitative real-time PCR (qPCR) and glycosaminoglycan (GAG) quantification. PPS significantly reduced AC proliferation through cell cycle modulation particularly by maintaining a significantly higher proportion of chondrocytes in the G1 phase and a significantly lower proportion in the S phase of the cell cycle in a concentration- and time-dependent manner. While the proportion of chondrocytes in G1 phase corresponded with the significant downregulation of cyclin-dependent kinase (CDK) 1 and 4. Furthermore, the study confirms that PPS promotes a chondrogenic phenotype of AC through significant upregulation of collagen type II (Col2A1) mRNA and GAG synthesis. The effect of PPS on the inhibition of chondrocyte proliferation while promoting a chondrocyte phenotype could be beneficial in the early stages of OA treatment, which transient increase in proliferative activity of chondrocytes with subsequent phenotypic shift and less productive in an essential component of extracellular matrix (ECM) is observed.
  • Ekkapol Akaraphutiporn, Takafumi Sunaga, Eugene C Bwalya, Ryosuke Echigo, Masahiro Okumura
    The Journal of veterinary medical science 82 6 793 - 803 2020年06月24日 
    This study investigated the effects of culture time on phenotype stability of canine articular chondrocytes (CACs) in non-passaged long-term monolayer culture. Third passage (P3) CACs isolated from four cartilage samples were seeded at three different initial seeding densities (0.2 × 104, 1.0 × 104 and 5.0 × 104 cells/cm2) and maintained in monolayer condition up to 8 weeks without undergoing subculture after confluence. The characteristic changes of chondrocytes during the culture period were evaluated based on the cell morphology, cell proliferation, glycosaminoglycans (GAGs) content, DNA quantification, mRNA expression and ultrastructure of chondrocytes. Chondrocytes maintained under post-confluence condition exhibited a capability to grow and proliferate up to 4 weeks. Alcian blue staining and Dimethylmethylene blue (DMMB) assay revealed that the extracellular matrix (ECM) synthesis was increased in a time-dependent manner from 2 to 8 weeks. The chondrocyte mRNA expression profile was dramatically affected by prolonged culture time, with a significant downregulation of collagen type I, whereas the expression of collagen type II, aggrecan, Sox9 and matrix metalloproteinase 13 (MMP-13) were significantly upregulated. In addition, transmission electron microscopy (TEM) result indicated dilation of rough endoplasmic reticulum (RER) in these long-term monolayer cultured chondrocytes. These findings demonstrate that the chondrocytes phenotype could be partially redifferentiated through the spontaneous redifferentiation process in long-term cultures using standard culture medium without the addition of chondrogenic supplements or tissue-culture scaffolds.
  • Atsushi Yamasaki, Yoshihiro Kunitomi, Daiki Murata, Takafumi Sunaga, Tomohide Kuramoto, Takeshi Sogawa, Kazuhiro Misumi
    Journal of orthopaedic research : official publication of the Orthopaedic Research Society 37 6 1398 - 1408 2019年06月 
    Osteoarthritis is a major joint disease that has been extensively investigated in humans and in model animals. In this study, we examined the regeneration of articular cartilage and subchondral bone using artificial scaffold-free constructs composed of adipose tissue-derived mesenchymal stem cells (AT-MSCs) created using bio three-dimensional (3D) printing with a needle-array. Printed constructs were implanted into osteochondral defects created in the right femoral trochlear groove of six mini-pigs, using femoral defects created in the left femurs as controls. Repair within the defects was evaluated at 3 and 6 months post-implantation using computed tomography (CT) and magnetic resonance (MR) imaging. The radiolucent volume (RV, mm3 ) in the defects was calculated using multi-planar reconstruction of CT images. MR images were evaluated based on a modified 2D- MOCART (magnetic resonance observation of cartilage repair tissue) grading system. Gross and microscopic pathology were scored according to the ICRS (International Cartilage Repair Society) scale at 6 months after implantation. The percentage RV at 3 months postoperation was significantly lower in the implanted defects than in the controls, whereas total scores based on the MOCART system were significantly higher in the implanted defects as compared with the controls. Although there were no statistical differences in the gross scores, the average histological scores were significantly higher in the implanted defects than in the controls. To our knowledge, this is the first report to suggest that artificial scaffold-free 3D-printed constructs of autologous AT-MSCs can be aid in the osteochondral regeneration in pigs. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:1398-1408, 2019.
  • Atsushi Yamasaki, Takaya Omura, Daiki Murata, Minoru Kobayashi, Takafumi Sunaga, Kanichi Kusano, Yoshiharu Ueno, Tomohide Kuramoto, Seiji Hobo, Kazuhiro Misumi
    Journal of equine science 29 4 117 - 122 2018年12月 
    Synovium-derived mesenchymal stromal cells (SM-MSCs) from seven Thoroughbreds with naturally occurring intra-articular fracture proliferated to over ten million cells by the second passage. Using three experimental Thoroughbreds, columnar osteochondral defects were made arthroscopically at the bilateral distal radius. Five million allogenic SM-MSCs were implanted into the right defect, and another five million were injected into the right radio-carpal joint (implantation site). No SM-MSCs were implanted into the left defect or the same joint (control site). At 3 and 6 weeks after surgery, ten million autologous SM-MSCs were injected into the right joints. Radiolucent volumes of defects calculated by analysis of postmortem CT images 9 weeks after surgery were decreased in implanted sites compared with control sites in all horses. The average scores for ICRS gross and histopathological grading scales in implanted sites were equal to or higher than those of the controls. These results suggest that allogenic implantation and subsequent autologous injection of SM-MSCs might not obstruct subchondral bone formation in defects.
  • Y-C Lai, N Ushio, M M Rahman, Y Katanoda, K Ogihara, Y Naya, A Moriyama, T Iwanaga, Y Saitoh, T Sogawa, T Sunaga, Y Momoi, H Izumi, N Miyoshi, Y Endo, M Fujiki, H Kawaguchi, N Miura
    Veterinary and comparative oncology 16 2 288 - 296 2018年06月 
    Canine hepatocellular carcinoma (HCC) is the most common primary hepatic tumour in dogs. MicroRNA (miRNA) dysregulation has been reported in human HCC and shown to have diagnostic and prognostic value; however, there are no data on miRNA expression in canine HCC. The aim of the present study was to investigate differentially expressed miRNAs in canine HCC. Analysis of miRNA expression in canine HCC tissues and cell lines by quantitative reverse transcription PCR showed that miR-1, miR-122, let-7a, and let-7g were downregulated, whereas miR-10b and miR-21 were upregulated in canine HCC. MET is one of the target genes of miR-1. MET was upregulated in canine HCC at the gene and protein levels, and a significant correlation between the concomitant downregulation of miR-1 and upregulation of MET was observed. Fast/intermediate-proliferating canine HCC cell lines had higher MET gene and protein expression levels than the slow-proliferating cell line. These findings suggest that miRNAs are differentially expressed in canine HCC, and that the miR-1/MET pathway may be associated with canine HCC cell proliferation.
  • Daiki Murata, Atsushi Yamasaki, Shouta Matsuzaki, Takafumi Sunaga, Makoto Fujiki, Satoshi Tokunaga, Kazuhiro Misumi
    Journal of equine science 27 2 57 - 65 2016年 
    Dedifferentiated fat (DFAT) cells have been shown to be multipotent, similar to mesenchymal stem cells (MSCs). In this study, we aimed to establish and characterize equine DFAT cells. Equine adipocytes were ceiling cultured, and then dedifferentiated into DFAT cells by the seventh day of culture. The number of DFAT cells was increased to over 10 million by the fourth passage. Flow cytometry of DFAT cells showed that the cells were strongly positive for CD44, CD90, and major histocompatibility complex (MHC) class I; moderately positive for CD11a/18, CD105, and MHC class II; and negative for CD34 and CD45. Moreover, DFAT cells were positive for the expression of sex determining region Y-box 2 as a marker of multipotency. Finally, we found that DFAT cells could differentiate into osteogenic, chondrogenic, and adipogenic lineages under specific nutrient conditions. Thus, DFAT cells could have clinical applications in tissue regeneration, similar to MSCs derived from adipose tissue.
  • Namgil Oh, Takafumi Sunaga, Hiroki Yamazaki, Kenji Hosoya, Satoshi Takagi, Masahiro Okumura
    The Japanese journal of veterinary research 61 3 97 - 107 2013年08月 
    Cyclooxygenase (COX)-2 participates essentially in bone healing, demonstrated by COX-2 knockout mice that showed delayed fracture repair. Considerable controversy still exists on inhibitory effects of COX-2 inhibitors on bone healing in clinical cases. To assess stage-dependent effects of short-term treatment of COX-2 inhibitors on osteogenic differentiation, a canine POS osteosarcoma cell line which spontaneously differentiates into osteoblastic cell was exposed to COX-2 inhibitors such as carprofen and meloxicam for 72 hours during three different stages of osteoblast differentiation, including day 0 to 3 (pre-osteoblastic stage), day 4 to 7 (transitional stage) and day 8 to 11 (mature osteoblastic stage). As osteogenic markers, expression of alkaline phosphatase (ALP) was estimated by analysis of mRNA expression, enzymatic activity and ALP staining, and expression of osteocalcin was estimated by analysis of mRNA expression after the drug treatments. Calcified matrix formation was finally observed by von Kossa staining on day 14. Expressions of ALP showed no significant suppression by carprofen and meloxicam during all three stages. However, expressions of osteocalcin mRNA and non-calcified nodule formations were delayed by carprofen and meloxicam during transitional stage. Nevertheless, fully calcified nodule formation was observed in all experimental groups during post-medication period. These results indicate that short-term treatment of carprofen and meloxicam would reversibly suppress the differentiation of osteoblasts.
  • Takafumi Sunaga, Namgil Oh, Kenji Hosoya, Satoshi Takagi, Masahiro Okumura
    The Journal of veterinary medical science 74 6 707 - 11 2012年06月 
    Pentosan polysulfate sodium (PPS) has a heparin-like structure and is purificated from the plant of European beech wood. PPS has been used for the treatment of interstitial cystitis for human patients. Recent years, it was newly recognised that PPS reduce pain and inflammation of OA. The molecular biological mechanism of PPS to express its clinical effects is not fully understood. The purpose of the present study is to investigate a mechanism of action of PPS on inflammatory reaction of chondrocytes in vitro. It was evaluated that effects of PPS on interleukin (IL)-1β-induced phosphorylation of mitogen-actiated protein kinases (MAPKs), such as p38, extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK), nuclear translocation of nuclear factor-kappa B (NF-κB), and matrix metalloproteinase (MMP)-3 production in cultured articular chondrocytes. As a result, in the presence of PPS existence, IL-1β-induced phosphorylation of p38 and ERK were certainly inhibited, while JNK phosphorylation was not affected. Nuclear translocation of NF-κB and MMP-3 production were suppressed by PPS pretreatment prior to IL-1β stimulation. In conclusion, it is strongly suggested that PPS treatment prevents inflammatory intracellular responses induced by IL-1 β through inhibition of phosphorylation of certain MAPKs, p38 and ERK and then nuclear translocation of NF-κB in cultured chondrocytes. These PPS properties may contribute to suppressive consequence of catabolic MMP-3 synthesis. These data might translate the clinical efficacy as PPS treatment could inhibit the cartilage catabolism and related clinical symptoms of OA in dogs.
  • Takafumi Sunaga, Namgil Oh, Kenji Hosoya, Satoshi Takagi, Masahiro Okumura
    The Journal of veterinary medical science 74 6 745 - 50 2012年06月 
    Canine osteoarthritis occurs frequently and causes secondary synovitis. Administration of nonsteroidal anti-inflammatory drugs (NSAIDs) is one of the major therapeutic options for pain management of joint diseases. Tepoxalin has an unique property as an NSAIDs that suppresses both cyclooxygenase and lipoxygenase. The purpose of this study was to evaluate antiproliferative effects of tepoxalin on cultured canine synovial cells. Cytotoxic effects of tepoxalin, carprofen, meloxicam and AA-861 on cultured canine synoviocytes were evaluated by MTT colorimetric assay. Apoptosis was detected by morphological observations with Giemsa or annexin V/Hoechst 33342 staining and by the inhibition of caspase-3 activity with N-Ac-Asp-Glu-Val-Asp-CHO (Ac-DEVD-CHO). Cytotoxic effects of tepoxalin were evident in comparison with the effects of carprofen or meloxicam. The same tendency of cytotoxicity was observed when 5-lipoxygenase was inhibited by AA-861. The morphological findings and contradictory effects of Ac-DEVD-CHO with regard to the cytotoxicity proved the proapoptotic effects of tepoxalin. In conclusion, tepoxalin might control osteoarthritic synovitis by inducing apoptosis in proliferating synoviocytes, while most NSAIDs that selectively inhibit cyclooxygenase-2 most likely would not suppress synovial proliferation.

その他活動・業績

  • ロジックで学ぶ 犬と猫の臨床テクニック(第22回) 皮膚のデブライドメント
    須永 隆文 CAP: Companion Animal Practice 34 (9) 132 -135 2019年09月
  • 滅菌と消毒 手術部位感染症を予防するために
    須永 隆文 日本獣医麻酔外科学雑誌 50 (Suppl.1) 66 -67 2019年06月
  • 【実践 創傷管理】咬傷、刺し傷などの処置 深い傷のある患部の治療と管理
    須永 隆文 CLINIC NOTE 13 (2) 8 -15 2017年02月
  • 症例から学ぶ鑑別診断(第29回) 吐出を呈した症例
    須永 隆文 SA Medicine 18 (2) 54 -60 2016年04月
  • 須永 隆文, 高木 哲, 小川 修治, 細谷 謙次, 奥村 正裕 日本獣医師会雑誌 66 (1) 57 -60 2013年01月

教育活動情報

主要な担当授業

  • 伴侶動物獣医療実習Ⅰ
    開講年度 : 2020年
    課程区分 : 学士課程
    開講学部 : 獣医学部
    キーワード : 伴侶動物、内科、外科、検査、診断、治療
  • 伴侶動物獣医療実習Ⅱ
    開講年度 : 2020年
    課程区分 : 学士課程
    開講学部 : 獣医学部
    キーワード : 伴侶動物、内科、外科、検査、診断、治療
  • 総合獣医療実習
    開講年度 : 2020年
    課程区分 : 学士課程
    開講学部 : 獣医学部
    キーワード : 臨床病理、画像診断、細胞診、病理解剖、麻酔
  • 夜間・救急獣医療実習Ⅰ
    開講年度 : 2020年
    課程区分 : 学士課程
    開講学部 : 獣医学部
    キーワード : 夜間・休日診療、救急処置


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