竹原 俊幸 (タケハラ トシユキ)

  • 近畿大学病院 助教
Last Updated :2024/03/24

研究者情報

学位

  • 博士(工学)(2007年03月 近畿大学)

ホームページURL

J-Global ID

研究キーワード

  • 生殖細胞   高度生殖医療   再生医療   ES細胞   熱刺激応答   EpiSC   始原生殖細胞   低酸素   

研究分野

  • ライフサイエンス / 産婦人科学

経歴

  • 2011年  近畿大学医学部附属病院助教

所属学協会

  • 日本軟骨代謝学会   日本分子生物学会   日本細胞生物学会   日本再生医療学会   

研究活動情報

論文

  • Kensuke Toriumi; Yuta Onodera; Toshiyuki Takehara; Tatsufumi Mori; Joe Hasei; Kanae Shigi; Natsumi Iwawaki; Toshifumi Ozaki; Masao Akagi; Mahito Nakanishi; Takeshi Teramura
    iScience 26 7 106946 - 106946 2023年07月 
    Mesenchymal stem cells (MSCs) are used as a major source for cell therapy, and its application is expanding in various diseases. On the other hand, reliable method to evaluate quality and therapeutic properties of MSC is limited. In this study, we focused on TWIST1 that is a transcription factor regulating stemness of MSCs and found that the transmembrane protein LRRC15 tightly correlated with the expression of TWIST1 and useful to expect TWIST1-regulated stemness of MSCs. The LRRC15-positive MSC populations in human and mouse bone marrow tissues highly expressed stemness-associated transcription factors and therapeutic cytokines, and showed better therapeutic effect in bleomycin-induced pulmonary fibrosis model mice. This study provides evidence for the important role of TWIST1 in the MSC stemness, and for the utility of the LRRC15 protein as a marker to estimate stem cell quality in MSCs before cell transplantation.
  • Tatsufumi Mori; Masatsugu Igarashi; Yuta Onodera; Toshiyuki Takehara; Maki Itokazu; Takeshi Teramura
    Biochemical and Biophysical Research Communications 2023年04月
  • Maki Itokazu; Yuta Onodera; Tatsufumi Mori; Shinji Inoue; Kotaro Yamagishi; Akihiro Moritake; Natsumi Iwawaki; Kanae Shigi; Toshiyuki Takehara; Yuji Higashimoto; Masao Akagi; Takeshi Teramura
    The Journal of biological chemistry 298 7 102098 - 102098 2022年07月 
    Sarcopenia is an aging-associated attenuation of muscular volume and strength and is the major cause of frailty and falls in elderly individuals. The number of individuals with sarcopenia is rapidly increasing worldwide; however, little is known about the underlying mechanisms of the disease. Sarcopenia often copresents with obesity, and some patients with sarcopenia exhibit accumulation of peri-organ or intra-organ adipose tissue as ectopic fat deposition, including atrophied skeletal muscle. In this study, we showed that transplantation of the perimuscular adipose tissue (PMAT) to the hindlimb thigh muscles of young mice decreased the number of integrin α7/CD29-double positive muscular stem/progenitor cells and that the reaction was mediated by PMAT-derived exosomes. We also found that the inhibition of cell proliferation was induced by Let-7d-3p miRNA that targets HMGA2, which is an important transcription factor for stem cell self-renewal, in muscular stem/progenitor cells and the composite molecular reaction in aged adipocytes. Reduction of Let-7 miRNA repressor Lin28 A/B and activation of nuclear factor-kappa B signaling can lead to the accumulation of Let-7d-3p in the exosomes of aged PMAT. These findings suggest a novel crosstalk between adipose tissue and skeletal muscle in the development of aging-associated muscular atrophy and indicate that adipose tissue-derived miRNAs may play a key role in sarcopenia.
  • 糸数 万紀; 小野寺 勇太; 岩脇 菜摘; 信貴 香苗; 竹原 俊幸; 福田 寛二; 寺村 岳士
    移植 56 4 447 - 447 (一社)日本移植学会 2022年02月
  • Shinichiro Suzuki; Kimio Yonesaka; Takeshi Teramura; Toshiyuki Takehara; Ryoji Kato; Hitomi Sakai; Koji Haratani; Junko Tanizaki; Hisato Kawakami; Hidetoshi Hayashi; Kazuko Sakai; Kazuto Nishio; Kazuhiko Nakagawa
    Clinical cancer research : an official journal of the American Association for Cancer Research 28 2 428 - 428 2022年01月
  • Tatsufumi Mori; Yuta Onodera; Maki Itokazu; Toshiyuki Takehara; Kanae Shigi; Natsumi Iwawaki; Masao Akagi; Takeshi Teramura
    Mechanisms of ageing and development 201 111619 - 111619 2022年01月 
    Frailty of the locomotory organs has become a widespread problem in the geriatric population. The major factor leading to frailty is an age-associated decrease in muscular mass and a reduced number of muscular cells and myofibers. In aged muscular tissues, muscular satellite cells (MuSCs) are reduced due to abnormalities in their self-renewal and the induction of apoptosis. However, the molecular mechanisms connecting aging-associated physiological changes and the reduction of MuSCs are largely unknown. NIMA-related kinase 2 (Nek2), a member of the Nek family of serine/threonine kinases, was found to be downregulated in aged MuSCs/progenitors. Further, Nek2 downregulation was found to inhibit self-renewal and apoptotic cell death by activating the p53-dependent checkpoint. Attenuated NEK2 expression was also observed in the muscular tissues of elderly donors, and its function was confirmed to be conserved in humans. Overall, this study proposes a novel mechanism for inducing muscular atrophy to understand aging-associated muscular diseases.
  • 鈴木 慎一郎; 米阪 仁雄; 寺村 岳士; 竹原 俊幸; 加藤 了資; 酒井 瞳; 原谷 浩司; 谷崎 潤子; 川上 尚人; 林 秀敏; 坂井 和子; 西尾 和人; 中川 和彦
    肺癌 61 6 629 - 629 (NPO)日本肺癌学会 2021年10月
  • Shinichiro Suzuki; Kimio Yonesaka; Takeshi Teramura; Toshiyuki Takehara; Ryoji Kato; Hitomi Sakai; Koji Haratani; Junko Tanizaki; Hisato Kawakami; Hidetoshi Hayashi; Kazuko Sakai; Kazuto Nishio; Kazuhiko Nakagawa
    Clinical cancer research : an official journal of the American Association for Cancer Research 27 20 5697 - 5707 2021年08月 
    PURPOSE: Treatment with KRAS G12C inhibitors such as sotorasib can produce substantial regression of tumors in some patients with non-small cell lung cancer (NSCLC). These patients require alternative treatment after acquiring resistance to the inhibitor. The mechanisms underlying this acquired resistance are unclear. The purpose of this study was to identify the mechanisms underlying acquired sotorasib resistance, and to explore potential treatments for rescuing patients with sotorasib-resistant KRAS G12C NSCLC cells. EXPERIMENTAL DESIGN: Clones of sotorasib-sensitive KRAS G12C NSCLC H23 cells exposed to different concentrations of sotorasib were examined using whole-genomic transcriptome analysis, multiple receptor kinase phosphorylation analysis, and gene copy number evaluation. The underlying mechanisms of resistance were investigated using immunological examination, and a treatment aimed at overcoming resistance was tested in vitro and in vivo Results: Unbiased screening detected subclonal evolution of MET amplification in KRAS G12C NSCLC cells that had developed resistance to sotorasib in vitro MET knockdown using siRNA restored susceptibility to sotorasib in these resistant cells. MET activation by its amplification reinforced RAS cycling from its inactive form to its active form. In addition to RAS-mediated MEK-ERK induction, MET induced AKT activation independently of RAS. Crizotinib, a MET inhibitor, restored sensitivity to sotorasib by eliminating RAS-MEK-ERK as well as AKT signaling. MET/KRAS G12C dual inhibition led to tumor shrinkage in sotorasib-resistant xenograft mice. CONCLUSIONS: MET amplification leads to the development of resistance to KRAS G12C inhibitors in NSCLC. Dual blockade of MET and KRAS G12C could be a treatment option for MET amplified, KRAS G12C-mutated NSCLC.
  • Kei Houri; Tatsufumi Mori; Yuta Onodera; Takatoshi Tsujimoto; Toshiyuki Takehara; Shinichi Nakao; Takeshi Teramura; Kanji Fukuda
    Scientific Reports 10 1 3735 - 3735 2020年12月 [査読有り]
     
    Elevation of the levels of reactive oxygen species (ROS) is a major tissue-degenerative phenomenon involved in aging and aging-related diseases. The detailed mechanisms underlying aging-related ROS generation remain unclear. Presently, the expression of microRNA (miR)-142-5p was significantly upregulated in bone marrow mesenchymal stem cells (BMMSCs) of aged mice. Overexpression of miR-142 and subsequent observation revealed that miR-142 involved ROS accumulation through the disruption of selective autophagy for peroxisomes (pexophagy). Mechanistically, attenuation of acetyltransferase Ep300 triggered the upregulation of miR-142 in aged BMMSCs, and miR-142 targeted endothelial PAS domain protein 1 (Epas1) was identified as a regulatory protein of pexophagy. These findings support a novel molecular mechanism relating aging-associated ROS generation and organelle degradation in BMMSCs, and suggest a potential therapeutic target for aging-associated disorders that are accompanied by stem cell degeneration.
  • Takatoshi Tsujimoto; Tatsufumi Mori; Kei Houri; Yuta Onodera; Toshiyuki Takehara; Kanae Shigi; Shinichi Nakao; Takeshi Teramura; Kanji Fukuda
    Biochemical and Biophysical Research Communications 523 3 707 - 712 2020年03月 [査読有り]
     
    Removal of dysfunctional mitochondria is essential step to maintain normal cell physiology, and selective autophagy in mitochondria, called mitophagy, plays a critical role in quality control of mitochondria. While in several diseases and aging, disturbed mitophagy has been observed. In stem cells, accumulation of damaged mitochondria can lead to deterioration of stem cell properties. Here, we focused on miR-155-5p (miR-155), one of the most prominent miRNAs in inflammatory and aged tissues, and found that miR-155 disturbed mitophagy in mesenchymal stem cells (MSCs). As a molecular mechanism of miR-155-mediated mitophagy suppression, we found that BCL2 associated athanogene 5 (BAG5) is a direct target of miR-155. Reduction of BAG5 resulted in destabilization of PTEN-induced kinase (PINK1) and consequently disrupted mitophagy. Our study suggests a novel mechanism connecting aging and aging-associated inflammation with mitochondrial dysfunction in stem cells through a miRNA-mediated mechanism.
  • Yuta Onodera; Takeshi Teramura; Toshiyuki Takehara; Kanji Fukuda
    STEM CELLS 37 12 1595 - 1605 2019年12月 [査読有り]
  • Teramura T; Matsuda K; Takehara T; Shinohara K; Miyashita Y; Mieno Y; Mori T; Fukuda K; Suzuki K; Suemori H
    Biochemical and biophysical research communications 503 4 3114 - 3120 2018年09月 [査読有り]
  • Onodera Y; Teramura T; Takehara T; Itokazu M; Mori T; Fukuda K
    PloS one 13 10 e0204860  2018年 [査読有り]
  • Yuta Onodera; Takeshi Teramura; Toshiyuki Takehara; Kayoko Obora; Tatsufumi Mori; Kanji Fukuda
    AGING CELL 16 6 1369 - 1380 2017年12月 [査読有り]
     
    Inflammation-induced reactive oxygen species (ROS) are implicated in cellular dysfunction and an important trigger for aging- or disease-related tissue degeneration. Inflammation-induced ROS in stem cells lead to deterioration of their properties, altering tissue renewal or regeneration. Pathological ROS generation can be induced by multiple steps, and dysfunction of antioxidant systems is a major cause. The identification of the central molecule mediating the above-mentioned processes would pave the way for the development of novel therapeutics for aging, aging-related diseases, or stem cell therapies. In recent years, microRNAs (miRNAs) have been shown to play important roles in many biological reactions, including inflammation and stem cell functions. In inflammatory conditions, certain miRNAs are highly expressed and mediate some cytotoxic actions. Here, we focused on miR-155, which is one of the most prominent miRNAs in inflammation and hypothesized that miR-155 participates to inflammation-induced ROS generation in stem cells. We observed mesenchymal stem cells (MSCs) from 1.5-year-old aged mice and determined that antioxidants, Nfe2l2, Sod1, and Hmox1, were suppressed, while miR-155-5p was highly expressed. Subsequent in vitro studies demonstrated that miR-155-5p induces ROS generation by suppression of the antioxidant genes by targeting the common transcription factor C/ebp beta Moreover, this mechanism occurred during the cell transplantation process, in which ROS generation is triggering loss of transplanted stem cells. Finally, attenuation of antioxidants and ROS accumulation were partially prevented in miR-155 knockout MSCs. In conclusion, our study suggests that miR-155 is an important mediator connecting aging, inflammation, and ROS generation in stem cells.
  • Yuta Onodera; Takeshi Teramura; Toshiyuki Takehara; Kayoko Obora; Tatsufumi Mori; Kanji Fukuda
    Aging Cell 16 6 1369 - 1380 2017年12月 [査読有り]
     
    Inflammation-induced reactive oxygen species (ROS) are implicated in cellular dysfunction and an important trigger for aging- or disease-related tissue degeneration. Inflammation-induced ROS in stem cells lead to deterioration of their properties, altering tissue renewal or regeneration. Pathological ROS generation can be induced by multiple steps, and dysfunction of antioxidant systems is a major cause. The identification of the central molecule mediating the above-mentioned processes would pave the way for the development of novel therapeutics for aging, aging-related diseases, or stem cell therapies. In recent years, microRNAs (miRNAs) have been shown to play important roles in many biological reactions, including inflammation and stem cell functions. In inflammatory conditions, certain miRNAs are highly expressed and mediate some cytotoxic actions. Here, we focused on miR-155, which is one of the most prominent miRNAs in inflammation and hypothesized that miR-155 participates to inflammation-induced ROS generation in stem cells. We observed mesenchymal stem cells (MSCs) from 1.5-year-old aged mice and determined that antioxidants, Nfe2l2, Sod1, and Hmox1, were suppressed, while miR-155-5p was highly expressed. Subsequent in vitro studies demonstrated that miR-155-5p induces ROS generation by suppression of the antioxidant genes by targeting the common transcription factor C/ebpβ. Moreover, this mechanism occurred during the cell transplantation process, in which ROS generation is triggering loss of transplanted stem cells. Finally, attenuation of antioxidants and ROS accumulation were partially prevented in miR-155 knockout MSCs. In conclusion, our study suggests that miR-155 is an important mediator connecting aging, inflammation, and ROS generation in stem cells.
  • Joe Hasei; Takeshi Teramura; Toshiyuki Takehara; Yuta Onodera; Takuro Horii; Merissa Olmer; Izuho Hatada; Kanji Fukuda; Toshifumi Ozaki; Martin K. Lotz; Hiroshi Asahara
    SCIENTIFIC REPORTS 7 42990  2017年02月 [査読有り]
     
    The objective was to investigate the levels of TWIST1 in normal and OA cartilage and examine its role in regulating gene expression in chondrocytes. Human cartilage tissues and chondrocytes were obtained at autopsy from normal knee joints and from OA-affected joints at the time of total knee arthroplasty. TWIST1 expression was increased in human OA knee cartilage compared to normal knee cartilage. TWIST1 induced matrix metalloproteinase 3 (MMP3) expression without direct binding to MMP3 promoter and increased the 5-hydroxymethylcytosine (5hmC) level at the MMP3 promoter. The effect of TWIST1 on expression of TET family (TET1, 2 and 3) was measured in stable TWIST1 transfected TC28 cells, and TET1 expression was up-regulated. TWIST1 dependent upregulation of Mmp3 expression was suppressed in Tet triple KO fibroblast derived from mouse ES cells. Increased TWIST1 expression is a feature of OA-affected cartilage. We identified a novel mechanism of catabolic reaction where TWIST1 up-regulates MMP3 expression by enriching 5hmC levels at the MMP3 promoter via TET1 induction. These findings implicate TWIST1 as an important factor regulating OA related gene expression. Clarifying epigenetic mechanisms of 5hmC induced by TWIST1 is a critical molecule to understanding OA pathogenesis.
  • Obora K; Onodera Y; Takehara T; Frampton J; Hasei J; Ozaki T; Teramura T; Fukuda K
    Scientific reports 7 43604  2017年02月 [査読有り]
     
    Intracerebral inflammation resulting from injury or disease is implicated in disruption of neural regeneration and may lead to irreversible neuronal dysfunction. Analysis of inflammation-related microRNA profiles in various tissues, including the brain, has identified miR-155 among the most prominent miRNAs linked to inflammation. Here, we hypothesize that miR-155 mediates inflammation-induced suppression of neural stem cell (NSC) self-renewal. Using primary mouse NSCs and human NSCs derived from induced pluripotent stem (iPS) cells, we demonstrate that three important genes involved in NSC self-renewal (Msi1, Hes1 and Bmi1) are suppressed by miR-155. We also demonstrate that suppression of self-renewal genes is mediated by the common transcription factor C/EBP beta, which is a direct target of miR-155. Our study describes an axis linking inflammation and miR-155 to expression of genes related to NSC self-renewal, suggesting that regulation of miR-155 may hold potential as a novel therapeutic strategy for treating neuroinflammatory diseases.
  • Kohei Kitada; Akane Kizu; Takeshi Teramura; Toshiyuki Takehara; Masami Hayashi; Daisuke Tachibana; Hideki Wanibuchi; Shoji Fukushima; Masayasu Koyama; Kayo Yoshida; Takashi Morita
    ENVIRONMENTAL SCIENCE AND POLLUTION RESEARCH 22 22 18252 - 18259 2015年11月 [査読有り]
     
    A high-throughput test of cell growth inhibition was performed using mouse embryonic stem (ES) cells to assess chemical toxicities. We herein demonstrated using a 96-well culture plate approach and the MTT assay that this method was suitable for prioritization of chemicals for their cytotoxic properties. In order to categorize chemicals, we used p53 gene-modified mouse ES cells as well as wild-type ES cells. The p53 gene is a well-known tumor suppressor and controls programmed cell death (apoptosis) and cellular senescence that is triggered by DNA-damaging agents such as alkylating agents and radiation. In the present study, p53-deficient ES cells were found to be more resistant to a tumor initiator, diethylnitrosamine (DEN), than wild-type ES cells, suggesting the inhibition of apoptosis or senescence by a dysfunction in p53. Chromosome aberrations were more frequently detected in p53-deficient ES cells than in wild-type cells, indicating genomic instability due to the deletion of p53. Other tumor initiators, methyl methanesulfonate (MMS) and N-methyl-N-nitrosourea (NMU), did not reveal apparent differences in cytotoxicity between wild-type and p53-deficient ES cells. Thus, ES test system using gene-modified ES cells may be used to categorize chemicals by detecting their characteristic effects on apoptosis, genotoxic potentials as well as general cytotoxicity.
  • Toshiyuki Takehara; Takeshi Teramura; Yuta Onodera; John Frampton; Kanji Fukuda
    SCIENTIFIC REPORTS 5 14722  2015年09月 [査読有り]
     
    The cell adhesion molecule Cadherin 2 (Cdh2) plays important roles in somatic cell adhesion, proliferation and migration. Cdh2 is also highly expressed in mouse epiblast stem cells (mEpiSCs), but its function in these cells is unknown. To understand the function of Cdh2 in mEpiSCs, we compared the expression of pluripotency-related genes in mEpiSCs and mouse embryonic stem cells (mESCs) after either Cdh2 knockdown or Cdh2 over-expression. Introduction of specific siRNA against Cdh2 led to attenuation of pluripotency-related genes. Pluripotent gene expression was not recovered by overexpression of Cdh1 following Cdh2 knockdown. Western blot analysis and co-immunoprecipitation assays revealed that Cdh2 stabilizes FGFR1 in mEpiSCs. Furthermore, stable transfection of mESCs with Cdh2 cDNA followed by FGF2 supplementation accelerated cell differentiation. Thus, Cdh2 contributes to the establishment and maintenance of FGF signaling-dependent self-renewal in mEpiSCs through stabilization of FGFR1.
  • Yuta Onodera; Takeshi Teramura; Toshiyuki Takehara; Kanji Fukuda
    FEBS OPEN BIO 5 476 - 484 2015年 [査読有り]
     
    One important pharmacological function of hyaluronic acid (HA) in chondrocytes is reduction of cellular superoxide generation and accumulation. Here we demonstrated a relationship between HA supplementation and accumulation of Nuclear factor-erythroid-2-related factor 2 (Nrf2), which is a master transcription factor in cellular redox reactions, in cultured chondrocytes derived from bovine joint cartilage. In HA-treated chondrocytes, expression of Nrf2 and its downstream genes was upregulated. In HA-treated chondrocytes, Akt was phosphorylated, and inhibition of Akt activity or suppression of HA receptors CD44 and/or RHAMM with siRNAs prevented HA-mediated Nrf2 accumulation. Furthermore, Nrf2 siRNA inhibited the HA effect on antioxidant enzymes. These results show that HA might contribute to ROS reduction through Nrf2 regulation by activating Akt. Our study suggests a new mechanism for extracellular matrix (ECM)-mediated redox systems in chondrocytes. (C) 2015 The Authors. Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies. This is an open access article under the CC BY license.
  • Yuta Onodera; Takeshi Teramura; Toshiyuki Takehara; Kanae Shigi; Kanji Fukuda
    FEBS OPEN BIO 5 492 - 501 2015年 [査読有り]
     
    Oxidative stress within the arthritis joint has been indicated to be involved in generating mediators for tissue degeneration and inflammation. COX-2 is a mediator in inflammatory action, pain and some catabolic reactions in inflamed tissues. Here, we demonstrated a direct relationship between oxidative stress and Cox-2 expression in the bovine synovial fibroblasts. Furthermore, we elucidated a novel mechanism, in which oxidative stress induced phosphorylation of MAPKs and NF-jB through TAK1 activation and resulted in increased Cox-2 and prostaglandin E2 expression. Finally, we demonstrated that ROS-induced Cox-2 expression was inhibited by supplementation of an antioxidant such as N-acetyl cysteamine and hyaluronic acid in vitro and in vivo. From these results, we conclude that oxidative stress is an important factor for generation of Cox-2 in synovial fibroblasts and thus its neutralization may be an effective strategy in palliative therapy for chronic joint diseases. (C) 2015 The Authors. Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies. This is an open access article under the CC BY license.
  • I. Tsukamoto; F. Nakamura; S. Inoue; T. Teramura; T. Takehara; Y. Onodera; M. Akagi
    OSTEOARTHRITIS AND CARTILAGE 22 Suppl. S147 - S147 2014年04月 [査読有り]
     
    Purpose: In 2013, we reported that the local renin-angiotensin system (RAS) can modulate the hypertrophic differentiation of chondrocytes activating angiotensin II type 1 receptor (AT1R) or angiotensin II type 2 receptor (AT2R). However, the details of the modulation have not been revealed. On the other hand, many researchers have already reported that the local RAS can modulate hypertrophic changes modulating the generations of reactive oxygen species (ROS) in vascular endothelial cells. The purpose of our study is to reveal whether or not the local RAS modulates the hypertrophic differentiation of chondrocytes modulating the generations of ROS.

    Methods: We cultured ATDC5 cell line under the inducing of 10 μg/ml bovine insulin for 14 days after they had reached confluence. Then, we added 1.0 μg/ml angiotensin II, 1.0 μg/ml Olmesartan and 1.0 μg/ml PD123319 to activate AT1R or AT2R separately. 6 times after the activation of the receptors, we measured the concentrations of ROS. We also administered vitamin C, vitamin E and hyaluronic acid (HA) and measured the concentrations of ROS and the expressions of type X collagen (Col.X), matrix metalloproteinase 13 (MMP13) and runt-related transcription factor (Runx2).

    Results: The generations of ROS were downregulated activating AT1R and were conversely upregulated activating AT2R. In the upregulated hypertrophic differentiation caused by activating AT2R, the expressions of Col.x, MMP13 and Runx2 and the generations of ROS were downregulated administering vitamin C, vitamin E and HA. However, in the downregulated hypertrophic differentiation caused by activating AT1R, we did not find significant changes in the the expressions of Col.x, MMP13 and Runx2 and the generations of ROS administering vitamin C, vitamin E and HA .

    Conclusions: The local RAS could modulate the hypertrophic differentiation of chondrocytes modulating the generations of ROS. Upregulation of hypertrophic differentiation of chondrocytes cased by activating AT2R could be suppressed by administering vitamin C, vitamin E and HA.
  • I. Tsukamoto; M. Akagi; S. Inoue; Y. Oda; T. Teramura; T. Takehara; Y. Onodera
    OSTEOARTHRITIS AND CARTILAGE 21 Suppl. S S130 - S130 2013年04月 [査読有り]
     
    Purpose: Recently, the local renin-angiotensin system (RAS) has attracted many researchers in many pathophysiological issues. Also in orthopedics, expression of the local RAS was found in fracture callus, bone tissues and arthritic synovium. In the last year, we reported that the local RAS expressed in chondrocytes of the epiphyseal plates of mice. The purpose of this study is to reveal immunohistological localization of the RAS components in the limb buds of mice where another physiological hypertrophic differetiation occurs and to analyze function of the local RAS in the processes of hypertrophic differentiation using ATDC5 cell line.

    Methods: The limb buds of 15-day-viviparous mice were immunostained with antibodies to angiotensinogen, angiotensinogen converting enzyme 1 (ACE1), angiotensinII type 1 receptor (AT1R) and angiotensinII type 2 receptor (AT2R). We cultured ATDC5 cell line and evaluated expression of angiotensinogen, ACE1, AT1R and AT2R during the hypertrophic term using quantitative real-time PCR and Western blot analysis. Then, we separately stimulated AT1R and AT2R in hypertrophic term using angiotensin II, olmesartan and PD123319 and evaluated expression of type X collagen using quantitative real-time PCR and Western blot analysis.

    Results: In the limb buds of mice, angiotensinogen and AT1R expressed in the resting chondrocytes, the proliferative chondrocytes and hypertrophic chondrocytes; however, ACE1 and AT2R expressed only in the hypertrophic chondrocytes. In ATDC5 cell line, angiotensinogen and AT1R expressed mainly in the proliferative term and slightly in the hypertrophic term; conversely, ACE1 and AT2R expressed mainly in the hypertrophic term and slightly in the proliferating term. In the hypertrophic term, expression of type X collagen were downregulated activating AT1R and were conversely upregulated activating AT2R.

    Conclusions: The local RAS also expresses in the limb buds of mice. In the hypertrophic term of the chondrocyte differentiation, activating AT1R decelerates the hypertrophic differentiation; conversely, activating AT2R accelerates the hypertrophic differentiation. Therefore, the local RAS might regulate the chondrocyte hypertrophic differentiation.
  • Takeshi Teramura; Hironobu Sugimoto; John Frampton; Yuta Kida; Miho Nakano; Makiko Kawakami; Hiroki Izumi; Naoto Fukunaga; Yuta Onodera; Toshiyuki Takehara; Kanji Fukuda; Yoshihiko Hosoi
    STEM CELLS AND DEVELOPMENT 22 6 928 - 938 2013年03月 [査読有り]
     
    In mammalian ovaries, many immature follicles remain after the dominant follicles undergo ovulation. Here we report the successful production of rabbit embryonic stem cells (ESCs) from oocytes produced by in vitro culture of immature follicles and subsequent in vitro maturation treatment. In total, we obtained 53 blastocysts from oocytes that received intracytoplasmic sperm injection followed by in vitro culture. Although only weak expression of POU5f1 was observed in the inner cell masses of in-vitro-cultured follicle-derived embryos, repeated careful cloning enabled establishment of 3 stable ESC lines. These ESC lines displayed the morphological characteristics of primed pluripotent stem cells. The ESC lines also expressed the pluripotent markers Nanog, POU5f1, and Sox2. Further, these ESCs could be differentiated into each of the 3 different germ layers both in vitro and in vivo. These results demonstrate that immature follicles from rabbits can be used to generate ESCs. Moreover, the use of rabbit oocytes as a cell source provides an experimental system that closely matches human reproductive and stem cell physiology.
  • Takeshi Teramura; Yuta Onodera; Toshiyuki Takehara; John Frampton; Toshiki Matsuoka; Syunsuke Ito; Koichi Nakagawa; Yoshihisa Miki; Yoshihiko Hosoi; Chiaki Hamanishi; Kanji Fukuda
    Cell Transplantation 22 2 309 - 329 2013年 [査読有り]
     
    Embryonic stem cells (ESCs) have the potential to be used as an unlimited cell source for cell transplantation therapy, as well as for studying mechanisms of disease and early mammalian development. However, applications involving ESCs have been limited by the lack of reliable differentiation methods in many cases. Mesenchymal stem cells (MSCs) have also emerged as a promising cell source, but as suggested in recent studies, these cells display limited potential for proliferation and differentiation, thereby limiting their usefulness in the clinic and in the laboratory. Unfortunately, effective methods for induction of MSCs from pluripotent stem cells have not been established, and the development of such methods remains a major challenge facing stem cell biologists. Oxygen concentration is one of the most important factors regulating tissue development. It has profound effects on cell metabolism and physiology and can strongly influence stem cell fate. Here we demonstrate that severe low O2 concentrations (1%) can function as a selective pressure for removing undifferentiated pluripotent cells during the induction of MSCs from rabbit ESCs (rESCs) and that MSCs induced under severe hypoxic conditions function as normal MSCs that is, they repopulate after cloning, express specific markers (vimentin, CD29, CD90, CD105, and CD140a) and differentiate into adipocytes, osteoblasts, and chondrocytes. Furthermore, we demonstrate that these cells can contribute to cartilage regeneration in an in vivo rabbit model for joint cartilage injury. These results support the notion that exposing ESCs to severe hypoxic conditions during differentiation can be used as a strategy for the preparation of functional MSCs from ESCs. © 2013 Cognizant Comm. Corp.
  • Ichiro Tsukamoto; Shinji Inoue; Takeshi Teramura; Toshiyuki Takehara; Kazuhiro Ohtani; Masao Akagi
    FEBS Open Bio 3 1 279 - 284 2013年01月 [査読有り]
  • Koichi Nakagawa; Takeshi Teramura; Toshiyuki Takehara; Yuta Onodera; Chiaki Hamanishi; Masao Akagi; Kanji Fukuda
    INFLAMMATION RESEARCH 61 10 1093 - 1100 2012年10月 [査読有り]
     
    Excessive mechanical stress on the cartilage causes the degradation of the matrix, leading to the osteoarthritis (OA). Matrix metalloproteinases 13 (MMP13) is a major catalytic enzyme in OA and p38 plays an important role in its induction. However, precise pathway inducing p38 activation has not been elucidated. We hypothesized here that the small GTPase Rho and its effector ROCK might function in upper part of the mechanical stress-induced matrix degeneration pathway. Bovine metacarpal phalangeal articular cartilage explants were loaded with 1 MPa dynamic compression for 6 h with or without a ROCK specific inhibitor Y27632 or/and a p38 specific inhibitor SB202190. Then p38 phosphorylation and MMP13 expression were assessed by western blot or/and quantitative RT-PCR. Rho-activity was measured by pull-down assay using glutathione S-transferase fusion protein of Rho binding domain. Cyclic compression caused Rho activation, p38 phosphorylation and MMP13 expression. Both Y27632 and SB202190 were found to block the mechanical stress-enhanced p38 phosphorylation and subsequent MMP13 expression. The present results show that p38 phosphorylation and MMP13 expression are regulated by Rho/ROCK activation, and support the potential novel pathway that Rho/ROCK is in the upper part of the mechanical stress-induced matrix degeneration cascade in cartilage comprised of p38 and MMP13.
  • Toshiyuki Takehara; Takeshi Teramura; Yuta Onodera; Chiaki Hamanishi; Kanji Fukuda
    STEM CELLS AND DEVELOPMENT 21 8 1239 - 1249 2012年05月 [査読有り]
     
    Recently, an additional type of pluripotent stem cell-line derived from mouse embryos has been established and termed epiblast stem cell (EpiSC), and is expected to be an important tool for studying the mechanisms of maintenance of pluripotency since they depend on basic fibroblast growth factor-MAPK and Activin A-Smad2/3 signaling to maintain pluripotency, unlike mouse embryonic stem cells (ESCs). Further, because of the similarities between mouse EpiSCs and human ESCs, EpiSCs are expected to be effective experimental models for human stem cell therapy. Recently, study for conversion from ESC state to EpiSC state or reversion from EpiSC state to ESC state has attracted interest since these techniques may lead to increasing the potential of pluripotent stem cells and our knowledge about their developmental status. In the present study, we find that a low oxygen concentration in culture environment accelerated, improved, and stabilized the EpiSC state of the converted cells from the ESC state using Oct4 Delta PE-GFP transgenic ESCs. Induced EpiSCs (iEpiSCs) in hypoxia possess closer gene expression patterns to native EpiSCs, and bisulfite sequences for the promoter regions of Stella and Oct4 genes have elucidated that the iEpiSC gain EpiSC-specific methylation patterns in hypoxia. Our data provide evidence that oxygen concentration is an important factor for establishment of the EpiSC-specific state.
  • I. Tsukamoto; M. Akagi; S. Inoue; T. Teramura; T. Takehara; Y. Onodera; K. Hashimoto; C. Hamanishi
    OSTEOARTHRITIS AND CARTILAGE 20 Suppl. 1 S151 - S151 2012年04月 [査読有り]
     
    Purpose: Recently, the local renin-angiotensin system (RAS) has attracted many researchers in many pathophysiological issues. In Orthopedics, expression of local RAS was found in bone tissues, fracture callus and arthritic synovium. The purpose of this study is to reveal immunohistological localization of the RAS components in the epiphyseal plates of mice and to analyze function of the local RAS in the processes of hypertrophic differentiation using ATDC5 chondroprogenitor cells.

    Methods: The epiphyseal plates of 8-week-old mice was immunostained with antibodies to angiotensinogen, angiotensinogen converting enzyme 1 (ACE1), angiotensinII type 1 receptor (AT1R) and angiotensinII type 2 receptor (AT2R). We cultured ATDC5 in long term and evaluated the expression of angiotensinogen, ACE1, AT1R, AT2R and type 2 collagen (COL2) by real- time PCR and Western blot analysis.

    Results: In the epiphyseal plates of mice, angiotensinogen and AT1R expressed in the resting chondrocytes, the proliferative chondrocytes and hypertrophic chondrocytes; however, ACE1 and AT2R expressed only in the hypertrophic chondrocytes. In ATDC5 chondroprogenitor cells, the local RAS components expressed both in proliferative and hypertrophic differentiating stages.

    Conclusions: The local RAS expresses in the epiphyseal plates of mice and might play an important role in the process of hypertrophic differentiation.
  • Takeshi Teramura; Toshiyuki Takehara; Yuta Onodera; Koichi Nakagawa; Chiaki Hamanishi; Kanji Fukuda
    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 417 2 836 - 841 2012年01月 [査読有り]
     
    Mechanical stimulation has been shown to regulate the proliferation and differentiation of stem cells. However, the effects of the mechanical stress on the sternness or related molecular mechanisms have not been well determined. Pluripotent stem cells such as embryonic stem (ES) cells and induced pluripotent stem (iPS) cells are used as good materials for cell transplantation therapy and research of mammalian development, since they can self-renew infinitely and differentiate into various cell lineages. Here we demonstrated that the mechanical stimulation to human iPS cells altered alignment of actin fibers and expressions of the pluripotent related genes Nanog, POU5f1 and Sox2. In the mechanically stimulated iPS cells, small GTPase Rho was activated and interestingly, ART phosphorylation was decreased. Inhibition of Rho-associated kinase ROCK recovered the ART phosphorylation and the gene expressions. These results clearly suggested that the Rho/ROCK is a potent primary effector of mechanical stress in the pluripotent stem cells and it participates to pluripotency-related signaling cascades as an upper stream regulator. (C) 2011 Elsevier Inc. All rights reserved.
  • ストレスに対するヒアルロン酸の軟骨保護作用
    中川 晃一; 寺村 岳士; 竹原 俊幸; 小野寺 勇太; 福田 寛二; 濱西 千秋
    近畿大学医学雑誌 36 3-4 14A - 14A 近畿大学医学会 2011年12月
  • 小野寺 勇太; 寺村 岳士; 竹原 俊幸; 福田 寛二
    日本プロテオーム学会大会要旨集 2011 100 - 100 日本プロテオーム学会(日本ヒトプロテオーム機構) 2011年
  • Naoto Fukunaga; Takeshi Teramura; Yuta Onodera; Toshiyuki Takehara; Kanji Fukuda; Yoshihiko Hosoi
    CELLULAR REPROGRAMMING 12 4 369 - 376 2010年08月 [査読有り]
     
    Recently, several research groups have shown that germ cells can be produced in vitro from pluripotent embryonic stem cells (ESCs). In the mouse, live births of offspring using germ cells induced from ESCs in vitro have been reported. Furthermore, some efficient methods for inducing the useful number of germ cells from ESCs have also been developed. On the other hand, in primates, despite the appearances of germ cell-like cells including meiotic cells were observed by spontaneous differentiation or introducing transgenes, it has not been determined whether fully functional germ cells can be derived from ESCs. To elucidate the property for the germ cells induced from primate ESCs, specification of the promoting factors for the germ cell development and improving the efficiency of germ cell derivation are essential. Leukemia inhibitory factor (LIF) has been reported as one of the important factors for mouse primordial germ cell (PGC) survival in vitro. However, the effects of LIF on germ cell formation from pluripotent cells of primates have not been examined. The aim of this study is to determine whether LIF addition can improve in vitro germ cell production from cynomolgus monkey ESCs (cyESCs). After 8 days of differentiation, LIF added culture induced dome-shaped germ cell colonies as indicated by the intense expression of alkaline phosphatase activity (ALP). These cells also demonstrate high-level expression of the germ cell-marker VASA, OCT-4, and BLIMP-1, and show SSEA-1 expression that supports their early stage germ cell identity. Finally, we observed that adding LIF to differentiating cultures inhibited meiotic gene expressions and increased the percentage of ALP-positive cells, and demonstrate that the addition of LIF to differentiation media increases differentiation of early germ cells from the cyESCs.
  • Yuta Onodera; Takeshi Teramura; Madoka Ozawa; Toshiyuki Takehara; Tasuku Mitani; Masayuki Anzai; Norimasa Sagawa; Chiaki Hamanishi; Yoshihiko Hosoi; Kanji Fukuda
    Theriogenology 74 1 135 - 145 2010年07月 [査読有り]
     
    Recent studies have illustrated multiple differentiation potentials of embryonic stem cells (ESCs), derived from parthenogenetic embryos, to various kinds of cells (all three embryonic germ layers). However, differentiation diversity of the parthenogenetic ESCs (PgESCs) in vivo remains to be elucidated. In the present study, we established mouse PgESC-lines and observed their contribution diversity in vivo by producing chimeric mice using embryos possessing single nucleotide polymorphisms of mitochondrial DNA (mtDNA) as hosts. Based on southern blot analysis using specific probes to detect the SNPs on mtDNA, PgESC-derived mtDNA were contained in many organs such as brain, lung, and heart of the chimeric mouse. We concluded that PgESCs contributed to various internal organs in vivo, and that they were also stably maintained in adult animals. © 2010 Elsevier Inc.
  • 小野寺 勇太; 寺村 岳士; 竹原 俊幸; 福田 寛二
    日本プロテオーム学会大会要旨集 2010 88 - 88 日本プロテオーム学会(日本ヒトプロテオーム機構) 2010年
  • Toshiyuki Takehara; Takeshi Teramura; Yuta Onodera; Satoshi Kishigami; Kazuya Matsumoto; Kazuhiro Saeki; Kanji Fukuda; Yoshihiko Hosoi
    STEM CELLS AND DEVELOPMENT 18 10 1433 - 1440 2009年12月 [査読有り]
     
    Neural stem cells (NSCs) are tissue-specific stem cells with self-renewal potential in brain, and are committed cells of the central nervous system. Recently, some reports have suggested the possibility of the NSCs to differentiate into non-CNS mesodermal derivatives, such as blood cells and skeletal muscle cells. Here we isolated NSCs as neurospheres from a neonatal mouse brain using serum replacement medium, and demonstrated that the stem cell population expressing pluripotent-related genes such as Oct-4, Sox-2, and Nanog possess multiple differentiation potentials to ectodermal, mesodermal, and endodermal lineages, that is, some neural cells, beating cardiomyocytes, adipocytes, and insulin-producing cells. The results of the present study partly provide further evidence for multiple differentiation properties of NSCs and suggest common characteristics between NSCs and other pluripotent stem cells.
  • Takeshi Teramura; Yuta Onodera; Hideki Murakami; Syunsuke Ito; Toshihiro Mihara; Toshiyuki Takehara; Hiromi Kato; Tasuku Mitani; Masayuki Anzai; Kazuya Matsumoto; Kazuhiro Saeki; Kanji Fukuda; Norimasa Sagawa; Yoshihiko Hosoi
    JOURNAL OF REPRODUCTION AND DEVELOPMENT 55 3 283 - 292 2009年06月 [査読有り]
     
    The embryos of some rodents and primates can precede early development without the process of fertilization; however, they cease to develop after implantation because of restricted expressions of imprinting genes. Asexually developed embryos are classified into parthenote/gynogenote and androgenote by their genomic origins. Embryonic stem cells (ESCs) derived from asexual origins have also been reported. To date, ESCs derived from parthenogenetic embryos (PgESCs) have been established in some species, including humans, and the possibility to be alternative sources for autologous cell transplantation in regenerative medicine has been proposed. However, some developmental characteristics, which might be important for therapeutic applications, such as multiple differentiation capacity and transplantability of the ESCs of androgenetic origin (AgESCs) are uncertain. Here, we induced differentiation of mouse AgESCs and observed derivation of neural cells, cardiomyocytes and hepatocytes in vitro. Following differentiated embryoid body (EB) transplantation in various mouse strains including the strain of origin, we found that the EBs Could engraft in theoretically MHC-matched strains. Our results indicate that AgESCs possess at least two important characteristics, multiple differentiation properties in vitro and transplantability after differentiation, and suggest that they can also serve as a source of histocompatible, tissues for transplantation.
  • Toshiyuki Takehara; Takeshi Teramura; Yuta Onodera; Ryo Kakegawa; Naoto Fukunaga; Makoto Takenoshita; Norimasa Sagawa; Kanji Fukuda; Yoshihiko Hosoi
    MOLECULAR HUMAN REPRODUCTION 14 11 627 - 634 2008年11月 [査読有り]
     
    Non-human primates are suitable models for preclinical research aimed at cell-replacement therapies. Recently, it has been reported that Rho-associated kinase inhibitor Y-27632 markedly reduced dissociation-induced apoptosis of human embryonic stem (hES) cells, and is expected as a novel supplement for hES cell maintenance or differentiation inductions; however, the effects of the chemical are still to be determined in model animals. Here, we demonstrated the effect of Y-27632 on cynomolgus monkey ES (cyES) cells. Also, in cyES cells, Y-27632 treatment dramatically improved the efficiency of colony formation from single cells without affecting the pluripotent state and karyotype. Y-27632 supplementation was also effective for feeder-free culture and differentiation induction. Neural stem cells directly induced from cyES cells could give rise to neurons, astrocytes and dopamine producing cells. The present result not only suggests that the chemical was effective for improving the culture system of primate ES cells, but also the similarity between cyES and hES cells regarding the reactions to the chemical, which might be further evidence that cyES cells are superior models for hES cells.
  • Ryo Kakegawa; Takeshi Teramura; Toshiyuki Takehara; Masayuki Anzai; Tasuku Mitani; Kazuya Matsumoto; Kazuhiro Saeki; Norimasa Sagawa; Kanji Fukuda; Yoshihiko Hosoi
    JOURNAL OF REPRODUCTION AND DEVELOPMENT 54 5 352 - 357 2008年10月 [査読有り]
     
    Primordial germ cells (PGCs) are embryonic precursors of the gametes of adult animals and are considered stem cells of the germline. Since their proliferation in vitro correlates well with the schedule of developmental changes in vivo, they might be interesting research tools for genomic imprinting, germ-cell tumors and fertility. Furthermore, once primordial germ cells are separated and placed on a feeder layer with cytokines, they become Cultured pluripotent cell lines called embryonic germ (EG) cells. EG cells share several important characteristics with embryonic stem (ES) cells as they call also contribute to the germ line of chimeras. To investigate the characteristics of PGCs and establish rabbit EG (rEG) cells, we cultured rabbit PGCs (rPGCs) in vitro with various combinations of leukemia inhibitory factor (LIF), basic fibroblast growth factor (bFGF) and forskolin oil inactivated mouse embryonic fibroblast (MEF) feeder layers. The present study found PGC proliferation in early cultures and induction of rEG-like colonies. These cells expressed pluripotent markers, such as alkaline phosphatase activity, OCT-4, Sox-2 and SSEA-1, in the undifferentiated state; however, the cells did not develop into a teratoma when injected into the kidney capsules of SCID mice, although the restricted differentiation potentials to neural cells were determined via embryoid body formation. From these characteristics and further characterization of the germ stem cell markers Vasa, SCP-1 and SCP-3, we suggested that these were hybrid cells with characteristics somewhere between PGC and EG cells.
  • Miyuri Kawasumi; Masayuki Anzai; Toshiyuki Takehara; Tasuku Mitani; Hiromi Kato; Kazuhiro Saeki; Akira Iritani; Kazuya Matsumoto; Yoshihiko Hosoi
    Journal of Reproduction and Development 53 3 615 - 622 2007年06月 [査読有り]
     
    The majority of somatic cell nuclear transferred (SCNT) embryos die before or after implantation. Many studies have focused on morphological remodeling of the donor nucleus and its associated cytoskeletal structures in the early events of nuclear transfer. However, little is known about the 2-cell stage of SCNT embryos after the first division. In this study, we compared the morphological status of chromosomal division during the 1-cell stage to the 2-cell stage in SCNT embryos with that in intracytoplasmic sperm injection (ICSI) embryos. The microtubules and cytoplasmic asters, which are related to chromatin segregation, disappeared at the pronuclear stage, although formation of the first mitotic spindle was normal in both the SCNT and ICSI embryos. However, nuclear fragmentation was observed in 30% of the 2-cell SCNT embryos and 12% of the 2-cell ICSI embryos. Nuclear fragmentation was present in both blastomeres of these embryos. No apoptotic DNA fragmentation was observed in TdT-mediated dUTP-biotin Nick End Labeling (TUNEL) assays for either the SCNT or ICSI embryos. In both the SCNT and ICSI embryos, the distribution of chromosomes in the first mitotic spindle was disturbed during the process of division from the 1-cell stage to the 2-cell stage. These results suggest that loss of SCNT embryos just before or after implantation may be due to an abnormal chromosome distribution at the 2-cell stage.
  • Takeshi Teramura; Toshiyuki Takehara; Nobuyuki Kawata; Nahoko Fujinami; Tasuku Mitani; Makoto Takenoshita; Kazuya Matsumoto; Kazuhiro Saeki; Akira Iritani; Norimasa Sagawa; Yoshihiko Hosoi
    Cloning and Stem Cells 9 2 144 - 156 2007年 [査読有り]
     
    Embryonic stem cells (ESCs) of nonhuman primates are important for research into human gametogenesis because of similarities between the embryos and fetuses of nonhuman primates and those of humans. Recently, the formation of germ cells from mouse ESCs in vitro has been reported. In this study, we established cynomolgus monkey ES cell lines (cyESCs) and attempted to induce their differentiation into germ cells to obtain further information on the development of primate germ cells by observing the markers specific to germ cells. Three cyESCs were newly established and confirmed to be pluripotent. When the cells are induced to differentiate, the transcripts of Vasa and some meiotic markers were expressed. VASA protein accumulated in differentiated cell clumps and VASA-positive cells gathered in clumps as the number of differentiation days increased. In the later stages, VASA-positive clumps coexpressed OCT-4, suggesting that these cells might correspond to early gonocytes at the postmigration stage. Furthermore, meiosis-specific gene expression was also observed. These results demonstrate that cyESCs can differentiate to developing germ cells such as primordial germ cells (PGCs) or more developed gonocytes in our differentiation systems, and may be a suitable model for studying the mechanisms of primate germ cell development. © Mary Ann Liebert, Inc.
  • Takeshi Teramura; Toshiyuki Takehara; Naoko Kishi; Toshihiro Mihara; Nobuyuki Kawata; Hiroki Takeuchi; Makoto Takenoshita; Kazuya Matsumoto; Kazuhiro Saeki; Akira Iritani; Norimasa Sagawa; Yoshihiko Hosoi
    Cloning and Stem Cells 9 4 485 - 494 2007年 [査読有り]
     
    Embryonic stem cells (ESCs) are a good material for the study of mammalian development, production of genetically modified animals, and drug discovery because they proliferate infinitely while maintaining a multilinege differentiation potency and a normal karyotype. However, ethical considerations limit the use of human embryos for the establishment of ESCs. Recently, ESCs have been produced from blastomeres divided by biopsy in mice and humans. The method is expected to be less controversial because it does not destroy the embryo. However, no one has yet produced both a pup and an ESC from a single embryo. Here, we describe the production of individual/ESC pairs from each of three embryos out of 20 attempts, and is thus considered efficient. Blastomere-derived ESC could differentiate some types of tissues and contribute to chimera mouse. These results show that each blastomere at two-cell stage possesses pluripotency and separated blastomeres maintain viability to develop to a pup or pluripotent ESC. © 2007 Mary Ann Liebert, Inc.

講演・口頭発表等

  • 副甲状腺組織再生を目指した多能性幹細胞由来副甲状腺細胞の誘導の試み  [通常講演]
    竹原俊幸; 小野寺勇太; 森 樹史; 寺村岳士
    第22回日本再生医療学会総会 2023年03月 口頭発表(一般)
  • βcateninに着目した新規ヒトナイーブ型多能性幹細胞の作出  [通常講演]
    竹原 俊幸
    第21回日本再生医療学会総会 2022年03月 口頭発表(一般)
  • 多能性維持機構におけるβ catenin の役割とそのパートナー分子の探索
    竹原俊幸
    第20回日本再生医療学会総会 2021年03月 ポスター発表
  • 竹原俊幸; 福田寛二; 小野寺勇太; 寺村岳士
    ISSCR 2020年06月 ポスター発表
  • 異なる多能性状態:Naïve型及びPrimed型に対するßcateninの役割  [通常講演]
    竹原俊幸; 小野寺勇太; 寺村岳士; 福田寛二
    第19回日本再生医療学会総会 2020年03月 ポスター発表
  • p62変異型筋萎縮性側索硬化症iPS細胞を用いた疾患機序解明の試み  [通常講演]
    竹原俊幸
    第18回日本再生医療学会総会 2019年03月 口頭発表(一般)
  • N-cadherin supports FGFR1 stability and subsequent activation of MEK/ERK dependent pluripotency on mouse epiblast stem cell  [通常講演]
    竹原俊幸
    第70回日本細胞生物発生生物合同大会2018 2018年06月 ポスター発表
  • 多能性状態に着目したマウスES細胞から生殖細胞誘導の試み  [通常講演]
    竹原俊幸
    第17回日本再生医療学会総会 2018年03月 口頭発表(一般)
  • 間葉系幹細胞におけるmiR155とROS産生の関係性  [通常講演]
    竹原俊幸; 小野寺勇太; 寺村岳士; 福田寛二
    第31回日本軟骨代謝学会 2018年03月 口頭発表(一般)
  • β-cateninを標的とした未分化性制御による基底状態多能性幹細胞獲得の試み  [通常講演]
    竹原俊幸; 寺村岳士; 小野寺勇太; 福田寛二
    第32回日本整形外科学会基礎学術集会 2017年10月 口頭発表(一般)
  • ヒトiPS細胞の自動評価および近赤外線レーザーによる分化細胞の除去と未分化維持のおける有効性の検討  [通常講演]
    寺村岳士; 松田浩一; 竹原俊幸; 小野寺勇太; 福田寛二; 鈴木孝一; 末盛博文
    第16回日本再生医療学会 2017年03月 口頭発表(一般)
  • miR-155はC/EBPβを介して神経幹細胞の未分化能を抑制する  [通常講演]
    竹原俊幸小野寺勇太; 寺村岳士; 竹原俊幸; 福田寛二
    第16回日本再生医療学会 2017年03月 口頭発表(一般)
  • TGFb-activated kinase (TAK1)は間葉系幹細胞の増殖に必要である  [通常講演]
    小野寺勇太; 寺村岳士; 竹原俊幸; 福田寛二
    第30回日本軟骨代謝学会 2017年03月 口頭発表(一般)
  • Twist1は間葉系幹細胞(MSC)の自己複製に重要である  [通常講演]
    竹原俊幸; 寺村岳士; 小野寺勇太; 福田寛二
    第30回日本軟骨代謝学会 2017年03月 口頭発表(一般)
  • β-catenin経路に着目した新規iPS細胞誘導技術の開発  [通常講演]
    竹原俊幸; 寺村岳士; 小野寺勇太; 福田寛二
    第16回日本再生医療学会 2017年03月 ポスター発表
  • miR-155は神経幹細胞の未分化能を抑制する  [通常講演]
    大洞佳代子; 小野寺勇太; 寺村岳士; 竹原俊幸; 長谷井嬢; 福田寛二
    第31回 日本整形外科学会基礎学術集会 2016年10月 ポスター発表
  • TWIST1による5-ヒドロキシメチルシトシン(5hmC)ステータス変動が引き起こすOAにおける新規エピジェネティクスの解明  [通常講演]
    長谷井嬢; 寺村岳士; 竹原俊幸; 小野寺勇太; 福田寛二; Martin Lotz; 浅原弘嗣; 尾﨑敏文
    第31回 日本整形外科学会基礎学術集会 2016年10月 口頭発表(一般)
  • TGF b-activated kinase(TAK1)は間葉系幹細胞のStemnessの維持に重要である  [通常講演]
    小野寺勇太; 寺村岳士; 竹原俊幸; 大洞佳代子; 長谷井嬢; 福田寛二
    第31回 日本整形外科学会基礎学術集会 2016年10月 ポスター発表
  • TAK1抑制によるマウスES細胞の多分可能性の検討  [通常講演]
    小野寺勇太; 寺村岳士; 竹原俊幸; 福田寛二
    第15回日本再生医療学会 2016年03月 口頭発表(一般)
  • 多能性転写因子nanogはTfamの抑制を介してミトコンドリアの制御に関与する  [通常講演]
    寺村岳士; 小野寺勇太; 竹原俊幸; 福田寛二
    第15回日本再生医療学会 2016年03月 口頭発表(一般)
  • The activation of CBP/beta-catenin signal pathway is involved in the transition from primed- to naïve state on mice pluripotent stem cells
    Toshiyuki Takehara; Takeshi Teramura; Yuta Onodera; Kanji Fukuda
    International Society for Stem Cell Reseach Annual Meeting 2015
  • CADHERIN-SWITCHING IS IMPORTANT FOR PLURIPOTENT STEM CELL BETWEEN Naïve AND PRIMED STATE
    Toshiyuki Takehara; Takeshi Teramura; Yuta Onodera; Kanji Fukuda
    International Society for Stem Cell Reseach Annual Meeting 2014
  • INDUCTION OF DIAFFERENTION INTO PRIMORDIAL GERM CELL FROM EPOBLAST STEM CELL FROM EROM MOUSE EMBRYONIC STEM CELL
    Toshiyuki Takehara; Takeshi Teramura; Yuta Onodera; Kanji Fukuda
    11th Intemational Society for Stem Cell Research,Annual Meeting
  • Differention reversibility between mous embryonic stem cells and epiblast stem cells  [通常講演]
    Toshiyuki Takehara; Takeshi Teramura; Yuta Onodera; Kanji Fukuda
    10th Intemational Society for Stem Cell Research,Annual Meeting 2012年06月
  • Detection of Oct4 Expression Subfraction in Neural Stem Cells from the Brain of a Newborn Mouse
    Toshiyuki Takehara; Takeshi Teramura; Yuta Onodera; Kayo Yoshida; Satosi Kisigami; Kazuhiro Saeki; Kazuya Matsumoto; Akira Iritani; Takashi Morita; Norimasa Sagawa; Yoshihiko Hosoi
    International Society for Stem Cell Reseach Annual Meeting 2008 2008年

MISC

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 基盤研究(C)
    研究期間 : 2022年04月 -2027年03月 
    代表者 : 小野寺 勇太; 寺村 岳士; 竹原 俊幸
  • 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 : 2023年04月 -2026年03月 
    代表者 : 寺村 岳士; 舘野浩章; 村川泰裕; 小野寺勇太; 竹原俊幸
  • 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 : 平野 牧人; 竹原 俊幸
  • 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 : 2021年04月 -2024年03月 
    代表者 : 森 樹史; 寺村 岳士; 竹原 俊幸
     
    間葉系幹細胞(Mesenchymal stem cell : MSC、stromal stem cell)は骨髄や脂肪などから分離される組織幹細胞であり、優れたサイトカイン産生能力と分化能を有する事から様々な疾患に対する再生医療材料としての期待が高まっている。一方で、多能性幹細胞とは異なり、ドナー年齢や細胞の継代数などにより劣化しやすく、維持・供給が安定しないという課題がある。また、品質を評価するマーカーが存在しないため、供給された細胞の品質がばらつくという問題も抱えている。申請者らは、転写因子Twist1がMSCの幹細胞性に極めて重要な分子であり、MSCの性能を評価する上で有用であることを発見している。体内において、MSCは骨髄、脂肪組織、筋組織等の血管周囲や間質に見られる。体外培養時と同様、体内においてもMSCは加齢や炎症などストレスの影響を受けるため、若齢患者から得られるMSCと高齢患者から得られるMSCでは性質、品質が全く異なることがわかってきている。一方で、体内に存在するMSCは、存在場所、ストレス蓄積量、それまでの分裂回数など複数の要素によってその品質が非常にヘテロな状態になっており、たとえ高齢ドナーであっても適した細胞マーカーに基づき評価分別できれば、より高い治療効果を安定的に得られると考えられる。本研究では、同知見を有用な医療技術として発展させる為、Twist1を操作しヒトMSCの作製・評価を目的として行う。
  • 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 : 2021年04月 -2024年03月 
    代表者 : 竹原 俊幸
     
    間葉系幹細胞(Mesenchymal Stem Cell:MSC)を用いた再生医療が次々と進められているが、 その細胞供給には根本的な問題がある。新たなMSCの供給源としてヒト多能性幹細胞からの分化誘導が進められているが、普及には至っていない。その理由として、ヒトMSCの発生機序、詳細な性質と正しい分類、由来組織や時期、培養方法による性質の変化など細胞そのものに対する基本的理解が進んでいないことにある。一方で、ヒトの発生解析は、倫理的にも技術的にも実施困難である。本研究では、ヒトNaive型多能性幹細胞を出発点とした試験管内における初期胚発生モデルGastruloidの構築と、これを用いた間葉系幹細胞の発生機序の解明を行う。Gastruloidは三次元的にヒト初期胚発生を模した培養技術であり、現存する発生モデルとしては最もin vivoに近い。本研究では、発生初期がブラックボックスに覆われるヒト間葉系原基について、ヒトNaive型多能性幹細胞より誘導したGastruloidを用い、発生機序の解明と新しい分化誘導法の確立を試みる。 令和3年度では、本研究の重要なツールであるGastruloid誘導法の確立を試みた。すでにいくつかのGastruloid法は報告されているが、確立されたとはいえない。そこで、Wnt/beta catenin経路の活性化あるいは不活性化による誘導法のどちらが適しているか検討した。すると、マウス多能性幹細胞ではどちらの方法も可能であったが、ヒト多能性幹細胞では、Wnt/beta catenin経路の阻害ではGastruloidの誘導ができなかった。これは、多能性幹細胞の状態が大きく影響していることが示唆される。正常な発生過程を体外で作り出すためには多能性幹細胞の状態を考慮する必要があると考えられる。
  • 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 : 2020年04月 -2023年03月 
    代表者 : 寺村 岳士; 味八木 茂; 小野寺 勇太; 村川 泰裕; 竹原 俊幸
     
    高齢化が急速に進行する中で、加齢性組織変性・疾患とその主因の一つである幹細胞老化を理解し方策を講じることは重要な課題である。筋骨格組織における幹細胞としては筋衛星細胞(Muscular Satellite Cell: MuSC)や間葉系幹細胞(Mesenchymal stem cell: MSC)が知られており、加齢に伴う細胞数の減少、自己複製能や分化能の低下が知られている。一方、老化研究においては体外培養下での培養細胞の表現系が個体の老化を反映しないため、転写解析、エピジェネティック解析を始め、モデル研究から得られた知見については明確な結論が得られている機序は限られていた。本研究では、ATACseq、NET-CAGEseq、IP質量分析といったオミックス解析を駆使しながら、幹細胞老化の本質となるメカニズムを明らかにすることを目的とする。さらに、新しい遺伝子治療技術SRVなどを取り入れ、加齢性に減少したクロマチン修飾因子、転写活性化因子の補完をはじめとする老化形質の制御法の開発を試みる。 本年度は、老化モデルマウスよりFACSで分離した筋MuSC、骨髄MSCを用いてトランスクリプトーム解析を行った。その結果、加齢MuSCで著しく発現が低下する分子としてNek2を抽出した。モデル細胞を用いた検討において、Nek2がG2期制御と非対称分裂の制御によって幹細胞の自己複製/分化の制御に関わっており、老化筋組織ではその調節が破綻している可能性を見出した。 また、昨年度までに申請者らが報告した炎症メディエターMAP3KであるTAK1と相互作用する分子をIP質量分析にて検索し、核内タンパク質Lyarを同定した。Lyarは老化骨髄MSCで発現が低下しており、トランスクリプトーム解析の結果、Lyarの機能は脂肪分化制御にあることを発見している。モデル細胞を用いた解析により、Lyarの減少は脂肪分化の促進をもたらすことが明らかとなり、老化骨髄でみられる脂肪組織蓄積を引き起こしうる新たな分子機序が示された。
  • SCA8関連筋萎縮性側索硬化症に対する治療薬の検証
    国立研究開発法人日本医療研究開発機構:創薬支援推進事業・創薬総合支援事業
    研究期間 : 2022年04月 -2023年03月 
    代表者 : 平野牧人; 竹原俊幸
  • 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 : 2019年04月 -2022年03月 
    代表者 : 平野 牧人; 竹原 俊幸
     
    本研究は、ATXN8OS関連ALSの病態に関連する蛋白の同定とiPS細胞由来の運動ニューロンモデルの構築と治療法開発を目指す。さらに、ブレインバンクから検体を提供いただき、病理組織も用いて結果を検証する。まず、病理的にALSと確認されている患者の遺伝子のスクリーニングを東京都健康長寿医療センター高齢者ブレインバンクと共同研究にて実施した。予定は最低30例であったが、49例提供していただき、全例で検査を施行した。その結果1例でATXN8OS変異陽性者が同定された。次にATXN8OS変異陽性患者の人工多能性幹 (iPS)細胞の樹立については、1例では既にiPS細胞の樹立は終了している。最近の研究でATXN8OS遺伝子変異陽性ALSがさらに2例同定された。その2症例の線維芽細胞を樹立し、現在iPS細胞を作製中である。また、既に樹立しているATXN8OS変異陽性患者のiPS細胞の運動ニューロンへの誘導を行い、運動ニューロンへ誘導することができ、現在、形態的、生化学的な性質を解析中である。さらに、iPS細胞由来運動ニューロンの染色とrepeat-associated non-AUG (RAN) 翻訳の確認を行っていくために、ポリセリンに続くアミノ酸配列、およびポリアラニンに対するポリクローナル抗体を作成し、ELISA標的抗原ペプチドへの反応は得られた。また、RAN翻訳にて生じるポリアミノ酸と関連する蛋白の同定のために、GFP融合の正常長ATXN8OSを発現するプラスミドベクターを構築した。iPS細胞由来運動ニューロンへの治療介入のため、3種類のsiRNAと3種類のアンチセンス核酸を設計・作製し、現在線維芽細胞において発現抑制が可能かの予備実験を行っている。
  • 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 : 2019年04月 -2022年03月 
    代表者 : 小野寺 勇太; 寺村 岳士; 竹原 俊幸
     
    間葉系幹細胞(MSC)は現在の再生医療における中心的な細胞材料の一つである。良好な臨床効果が複数報告されているが、MSCの幹細胞性維持機構はほとんど分かっていない。我々はMAP3K の1つでTgfβシグナルを伝達するTgfβ-activated kinase(Tak1)を欠くMSCが殆ど増殖出来ないという新しい知見を得た。本研究ではTak1とHippo pathwayとの連関を中心にMSCの増殖制御機構に迫り、Tak1阻害による静止期同期を応用した新たな移植用細胞調整法の可能性を検討する。 Tak1は、皮膚や造血幹細胞、肝細胞、神経堤細胞の維持に必須で、Tak1阻害が初期胚や多能性幹細胞の未分化性を上昇させること、筋衛星細胞の自己複製に必須であること等の報告がある。 先行研究から、MSC / マウスへのTak1阻害は●分化能力に影響せず、●細胞周期の停止、抗ストレス遺伝子等の発現が上昇、●解糖系代謝に傾倒し、●骨髄内MSCの細胞数および有意な体重減少が生じるが投与中止後は回復する(Tak1阻害は不可逆的である)ことが明らかとなっている。 これらの結果を踏まえ、細胞増殖や器官サイズの制御を担うHippo Pathwayの重要な因子であるYap1 / Tazとの関係を明らかにする事を試みた。Tak1がYap1 / Tazと結合することで核内移行し、細胞増殖を促進している知見を得た。また、Tak1阻害によりストレス耐性が亢進していることからマウス髄腔内細胞移植を実施し、移植時の課題である細胞死や生着数改善に寄与するか検討した。Tak1阻害細胞は移植直後において有意に生存数が多く、幹細胞性を保持し、術後一か月後においても生着細胞が多く観察された。 以上の結果は「Stem cells」に掲載された(doi: 10.1002/stem.3083.)。
  • 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 : 2017年04月 -2020年03月 
    代表者 : 竹原 俊幸; 寺村 岳士; 福田 寛二; 末盛 博文
     
    ヒトiPS細胞を臨床応用するには、より能力が高くまた安定した基底状態へ導く必要がある。本研究では、βcatenin分子に着目し、その修飾状態を変化させることで遺伝子組換えを経ず、また単一の因子の制御によって基底状態ヒトiPS細胞の獲得が可能であった。本研究成果は、多能性ネットワークの理解およびiPS細胞のさらなる利用価値を高める結果となる。また、βcateninは細胞の癌化や細胞死に強く関連することから、本研究により得られた新しいβcateninの機能は未知の細胞現象における一つの重要な知見となりうる。
  • TWIST1による幹細胞性の誘導とメカニズムの解明
    公益財団法人中冨健康科学振興財団:研究助成金
    研究期間 : 2019年04月 -2020年03月 
    代表者 : 竹原俊幸
  • 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 : 2016年04月 -2019年03月 
    代表者 : 福田 寛二; 小野寺 勇太; 寺村 岳士; 丹羽 淳子; 高橋 英夫; 竹原 俊幸
     
    本研究では、生体内において酸化ストレス除去を担う分子経路のマスター転写因子であるNrf2に着目し、運動器において最も含有量の多い幹細胞である間葉系幹細胞においてその発現調節機序と機能を明らかにすることを目的に研究を行った。加齢組織においては慢性炎症、あるいは転写因子Notch1の減少によりmiR-155が誘導され、C/ebpβの発現が抑制されることでNrf2の発現抑制が生じ、抗ストレス経路の破綻が生じることが明らかとなった。Nrf2には幹細胞の自己複製、エピジェネティック修飾の維持に関わる機能を有することも示唆されたことから、Nrf2は運動機能維持を目指した標的分子になりうると考えられた。
  • 日本学術振興会:科学研究費助成事業 若手研究(B)
    研究期間 : 2016年04月 -2019年03月 
    代表者 : 小野寺 勇太; 寺村 岳士; 竹原 俊幸
     
    骨髄組織由来の間葉系幹細胞(BMMSC)は、創傷治癒や組織再生を目的とした移植治療の重要な細胞ソースとして注目されている。しかし、BMMSCの未分化能を制御する分子機構には未知な点が多い。TAK1に着目しその機能と介入対象としての可能性を検討した。 TAK1阻害剤(5zox)濃度依存的にBMMSCの増殖能は低下した。また、Fucci-BMMSCの解析およびトランスクリプトーム解析により、TAK1の抑制はBMMSCに静止期を誘導することが明らかとなった。TAK1阻害剤が静止期同調をもたらすことを利用し、BMMSCの骨髄髄腔内移植を実施し、定着率の向上に有効であった。
  • 日本学術振興会:科学研究費助成事業 若手研究(B)
    研究期間 : 2014年04月 -2017年03月 
    代表者 : 竹原 俊幸
     
    本研究では、ドナー細胞の性質・種類に問われないiPS細胞の誘導及び培養法の開発を試みた。実験には様々な生理現象に関わっており、また種保存性が高いβ-cateninおよび転写コアクティベータCBPに着目した。CBP/β-catenin経路の活性化によってマウスだけでなく、ヒト細胞においても多能性幹細胞へのリプログラムを促進することを明らかにした。また、マウス多能性幹細胞の未分化維持に対して、CBP/β-cateninの活性化は基底状態を強く正に制御することが観察された。以上のことから、CBP/β-catenin経路の活性化は、再現性が高いかつ効率的なiPS細胞の誘導及び培養法であると考えられる。
  • 日本学術振興会:科学研究費助成事業 若手研究(B)
    研究期間 : 2011年 -2013年 
    代表者 : 竹原 俊幸
     
    ストレス因子に着目し、生体内の環境を模した培養環境を作り出し、ES細胞から半数体生殖細胞の誘導とその評価を試みた。本研究では体内における生殖細胞の発生過程を模範するため、①ES細胞からエピブラスト幹細胞(EpiSC)の誘導②始原生殖細胞の誘導という2段階の誘導方法の検討を行った。得られた誘導細胞は、始原生殖細胞特異的なマーカーを発現していた。さらに、始原生殖細胞で特徴的に生じるインプリンティング遺伝子の脱メチル化も認められた。また、これらの誘導細胞を不妊雄マウス精巣内に移植すると精子様細胞の出現が認められた。以上より、ES細胞から生殖細胞の誘導が可能であるという結論が得られた。
  • 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 : 1998年 -1999年 
    代表者 : 竹原 俊幸; 長野 正史; 井手 久満
     
    骨形成蛋白質(Bone morphogenic protein:BMP)は骨基質中に存在し、異所性骨形成を誘導する蛋白性因子として分離、同定された。BMPの生物学的機能としては、強力な骨形成誘導因子であるのみならず、胚形成や形態形成、アポトーシス、細胞分化、走化性などに関与し、多機能性を有する重要な蛋白質である。そのBMPの受容体としては、2つのI型受容体(BMPR-IA,-IB)とひとつのII型受容体(BMPR-II)が知られている。一方、前立腺がんは高率に骨転移をきたし、その多くが特徴的な骨硬化像を示す。本研究において、我々はBMP/BMPR信号伝達系の前立腺がんにおける生物学的関与を検討してきた。その過程において、BMPの受容体のひとつであるヒトBMPR-IBのcDNAをクローニングした。また、FISH法、Radiation Hybrid Mapping法を用いて、BMPR-IAは10q22.36に、BMPR-IBは4q23-24に位置することを示した。BMP-2の前立腺がんにおける増殖効果としては、アンドロゲン感受性の前立腺がん細胞株LNCaPにおいて、BMP-2はアンドロゲン存在下では増殖抑制効果を示したが、アンドロゲン非存在下では増殖促進効果を示した。またこのとき、BMP受容体中、BMPR-IBのみが、アンドロゲンによりその発現が増加していた。以上の結果から、BMPの増殖効果において、BMPR-IA、-IBが前立腺がん細胞において、それぞれ増殖促進、抑制効果と相反する増殖制御信号を伝達する可能性が示された。

その他

  • 2023年04月 - 2026年03月  ヒト多能性幹細胞由来 Gasruloid(人工擬似胚)を用いた副甲状腺組織の理解と再生の試み 
    近畿大学 21世紀研究開発奨励金 【共同研究助成金】(KD2307) 研究代表者:竹原俊幸 共同研究者:東本有司,寺村岳士,森 樹史
  • 2018年04月 - 2018年04月  展示動物からの細胞・配偶子を用いた生殖・発生工学による新規生物多様性保全技術の開発と動物園・水族館との統合研究連携モデルの構築 
    近畿大学学内研究助成金 21世紀研究開発奨励金(KD201801) 研究代表者:三谷 匤 分担者:竹原俊幸
  • 2016年04月 - 2016年04月  「古(いにしえ)の生物情報」の知的探求から絶滅動物の再生へ 
    近畿大学学内研究助成金 21世紀研究開発奨励金(KD10) 研究代表者:細井美彦 分担者:竹原俊幸
  • 2016年04月 - 2016年04月  樹立困難な動物種におけるiPS細胞誘導技術の開発 
    近畿大学学内研究助成金 奨励研究助成金(SR19) 研究代表者:竹原俊幸
  • 2014年04月 - 2014年04月  マンモスおよび希少動物の再生に向けて:異種間核移植法とiPS細胞を介した配偶子作成法の確立 
    近畿大学学内研究助成金 21世紀研究開発奨励金(KD10) 研究代表者:細井美彦 分担者:竹原俊幸
  • 2011年04月 - 2011年04月  マンモスおよび希少動物の再生に向けて:異種間核移植法とiPS細胞を介した配偶子作製法の検討 
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