Curriculum Vitaes
Profile Information
- Affiliation
- Professor, Fujita Medical Innovation Center Tokyo, Fujita Health UniversityProject Leader, Kanagawa Institute of Industrial Science and TechnologyTeam Leader, Regenerative Medicine Research Center, Keio University(Concurrent)Visiting Professor, School of Medicine
- Degree
- PhD(Keio University)
- J-GLOBAL ID
- 201701015467140157
- researchmap Member ID
- B000284421
- External link
Research Areas
2Research History
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Apr, 2025 - Present
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Apr, 2025 - Present
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Nov, 2024 - Present
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Nov, 2024 - Present
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Apr, 2024 - Present
Education
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Apr, 2008 - Mar, 2013
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Apr, 2000 - Mar, 2006
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Apr, 1997 - Mar, 2000
Committee Memberships
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Apr, 2023 - Present
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Nov, 2022 - Present
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Oct, 2022 - Present
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Aug, 2022 - Present
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May, 2022 - Present
Major Awards
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Jun, 2023
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Oct, 2021
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Nov, 2019
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Apr, 2019
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Apr, 2018
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Mar, 2017
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Mar, 2013
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Mar, 2011
Major Papers
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iScience, 27(11) 111234-111234, Nov, 2024 Peer-reviewedCorresponding author
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Advanced healthcare materials, 13(27) e2303477, Oct, 2024 Peer-reviewedCorresponding authorAbstract Here an electrical stimulation system is described for maturing microfiber‐shaped cardiac tissue (cardiac microfibers, CMFs). The system enables stable culturing of CMFs with electrical stimulation by placing the tissue between electrodes. The electrical stimulation device provides an electric field covering whole CMFs within the stimulation area and can control the beating of the cardiac microfibers. In addition, CMFs under electrical stimulation with different frequencies are examined to evaluate the maturation levels by their sarcomere lengths, electrophysiological characteristics, and gene expression. Sarcomere elongation (14% increase compared to control) is observed at day 10, and a significant upregulation of electrodynamic properties such as gap junction protein alpha 1 (GJA1) and potassium inwardly rectifying channel subfamily J member 2 (KCNJ2) (maximum fourfold increase compared to control) is observed at day 30. These results suggest that electrically stimulated cultures can accelerate the maturation of microfiber‐shaped cardiac tissues compared to those without electrical stimulation. This model will contribute to the pathological research of unexplained cardiac diseases and pharmacologic testing by stably constructing matured CMFs.
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Circulation, 150(8) 611-621, Aug 20, 2024 Peer-reviewedLead authorCorresponding authorBACKGROUND: The clinical application of human induced pluripotent stem cell-derived cardiomyocytes (CMs) for cardiac repair commenced with the epicardial delivery of engineered cardiac tissue; however, the feasibility of the direct delivery of human induced pluripotent stem cell-derived CMs into the cardiac muscle layer, which has reportedly induced electrical integration, is unclear because of concerns about poor engraftment of CMs and posttransplant arrhythmias. Thus, in this study, we prepared purified human induced pluripotent stem cell-derived cardiac spheroids (hiPSC-CSs) and investigated whether their direct injection could regenerate infarcted nonhuman primate hearts. METHODS: We performed 2 separate experiments to explore the appropriate number of human induced pluripotent stem cell-derived CMs. In the first experiment, 10 cynomolgus monkeys were subjected to myocardial infarction 2 weeks before transplantation and were designated as recipients of hiPSC-CSs containing 2×107 CMs or the vehicle. The animals were euthanized 12 weeks after transplantation for histological analysis, and cardiac function and arrhythmia were monitored during the observational period. In the second study, we repeated the equivalent transplantation study using more CMs (6×107 CMs). RESULTS: Recipients of hiPSC-CSs containing 2×107 CMs showed limited CM grafts and transient increases in fractional shortening compared with those of the vehicle (fractional shortening at 4 weeks after transplantation: 26.2±2.1%; 19.3±1.8%; P<0.05), with a low incidence of posttransplant arrhythmia. Transplantation of increased dose of CMs resulted in significantly greater engraftment and long-term contractile benefits (fractional shortening at 12 weeks after transplantation: 22.5±1.0%; 16.6±1.1%; P<0.01, left ventricular ejection fraction at 12 weeks after transplantation: 49.0±1.4%; 36.3±2.9%; P<0.01). The incidence of posttransplant arrhythmia slightly increased in recipients of hiPSC-CSs containing 6×107 CMs. CONCLUSIONS: We demonstrated that direct injection of hiPSC-CSs restores the contractile functions of injured primate hearts with an acceptable risk of posttransplant arrhythmia. Although the mechanism for the functional benefits is not fully elucidated, these findings provide a strong rationale for conducting clinical trials using the equivalent CM products.
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Cell Reports Methods, 3(12) 100666-100666, Dec, 2023 Peer-reviewedCorresponding author
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Hydrogel-Sheathed HiPSC-Derived Heart Microtissue Enables Anchor-Free Contractile Force Measurement.Advanced Science, e2301831, Oct 17, 2023 Peer-reviewedCorresponding authorIn vitro reconstruction of highly mature engineered heart tissues (EHTs) is attempted for the selection of cardiotoxic drugs suitable for individual patients before administration. Mechanical contractile force generated in the EHTs is known to be a critical indicator for evaluating the EHT response. However, measuring contractile force requires anchoring the EHT in a tailored force-sensing cell culture chamber, causing technical difficulties in the stable evaluation of contractile force in long-term culture. This paper proposes a hydrogel-sheathed human induced pluripotent stem cell (hiPSC)-derived heart microtissue (H3 M) that can provide an anchor-free contractile force measurement platform in commonly used multi-well plates. The contractile force associated with tissue formation and drug response is calculated by motion tracking and finite element analysis on the bending angle of the hydrogel sheath. From the experiment of the drug response, H3 M is an excellent drug screening platform with high sensitivity and early testing capability compared to conventionally anchored EHT. This unique platform would be useful and versatile for regenerative therapy and drug discovery research in EHT.
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Stem Cell Reports, Aug 31, 2023 Peer-reviewedLast authorCorresponding authorMonitoring cardiac differentiation and maturation from human pluripotent stem cells (hPSCs) and detecting residual undifferentiated hPSCs are indispensable for the development of cardiac regenerative therapy. MicroRNA (miRNA) is secreted from cells into the extracellular space, and its role as a biomarker is attracting attention. Here, we performed an miRNA array analysis of supernatants during the process of cardiac differentiation and maturation from hPSCs. We demonstrated that the quantification of extracellular miR-489-3p and miR-1/133a-3p levels enabled the monitoring of mesoderm and cardiac differentiation, respectively, even in clinical-grade mass culture systems. Moreover, extracellular let-7c-5p levels showed the greatest increase with cardiac maturation during long-term culture. We also verified that residual undifferentiated hPSCs in hPSC-derived cardiomyocytes (hPSC-CMs) were detectable by measuring miR-302b-3p expression, with a detection sensitivity of 0.01%. Collectively, we demonstrate that our method of seamlessly monitoring specific miRNAs secreted into the supernatant is non-destructive and effective for the quality evaluation of hPSC-CMs.
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Biomaterials, 299 122174-122174, Aug, 2023 Peer-reviewedLast authorCorresponding authorAlthough the extracellular matrix (ECM) plays essential roles in heart tissue engineering, the optimal ECM components for heart tissue organization have not previously been elucidated. Here, we focused on the main ECM component, fibrillar collagen, and analyzed the effects of collagens on heart tissue engineering, by comparing the use of porcine heart-derived collagen and other organ-derived collagens in generating engineered heart tissue (EHT). We demonstrate that heart-derived collagen induces better contraction and relaxation of human induced pluripotent stem cell-derived EHT (hiPSC-EHT) and that hiPSC-EHT with heart-derived collagen exhibit more mature profiles than those with collagens from other organs. Further, we found that collagen fibril formation and gel stiffness influence the contraction, relaxation, and maturation of hiPSC-EHT, suggesting the importance of collagen types III and type V, which are relatively abundant in the heart. Thus, we demonstrate the effectiveness of organ-specific collagens in tissue engineering and drug discovery.
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STAR Protocols, 3(2) 101360-101360, Jun 17, 2022 Peer-reviewedLast authorCorresponding authorHere we describe a protocol to obtain highly pure cardiomyocytes and neurons from human induced pluripotent stem cells (hiPSCs) via metabolic selection processes. Compared to conventional purification protocols, this approach is easier to perform and scale up and more cost-efficient. The protocol can be applied to hiPSCs and human embryonic stem cells. For complete details on the use and execution of this protocol, please refer to Tohyama et al. (2016) and Tanosaki et al. (2020).
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STAR Protocols, 3(2) 101341-101341, Jun 17, 2022 Peer-reviewedLast authorCorresponding authorWe describe a protocol for the efficient culture of human pluripotent stem cells (hPSCs) by supplementing conventional culture medium with L-tryptophan (TRP). TRP is an essential amino acid that is widely available at an affordable cost, thereby allowing cost-effective proliferation of hPSCs compared to using a conventional medium alone. Here, we describe the steps for enhanced proliferation of hPSCs from dermal fibroblasts or peripheral blood cells, but the protocol can be applied to any hPSCs. For complete details on the use and execution of this protocol, please refer to Someya et al. (2021).
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iScience, 24(2) 102090-102090, Feb 19, 2021 Peer-reviewedCorresponding authorHuman pluripotent stem cells (hPSCs) have a unique metabolic signature for maintenance of pluripotency, self-renewal, and survival. Although hPSCs could be potentially used in regenerative medicine, the prohibitive cost associated with large-scale cell culture presents a major barrier to the clinical application of hPSC. Moreover, without a fully characterized metabolic signature, hPSC culture conditions are not optimized. Here, we performed detailed amino acid profiling and found that tryptophan (TRP) plays a key role in the proliferation with maintenance of pluripotency. In addition, metabolome analyses revealed that intra- and extracellular kynurenine (KYN) is decreased under TRP-supplemented conditions, whereas N-formylkynurenine (NFK), the upstream metabolite of KYN, is increased thereby contributing to proliferation promotion. Taken together, we demonstrate that TRP is indispensable for survival and proliferation of hPSCs. A deeper understanding of TRP metabolism will enable cost-effective large-scale production of hPSCs, leading to advances in regenerative medicine.
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iScience, 23(9) 101535-101535, Sep 25, 2020 Peer-reviewedCorresponding authorThe role of lipid metabolism in human pluripotent stem cells (hPSCs) is poorly understood. We have used large-scale targeted proteomics to demonstrate that undifferentiated hPSCs express different fatty acid (FA) biosynthesis-related enzymes, including ATP citrate lyase and FA synthase (FASN), than those expressed in hPSC-derived cardiomyocytes (hPSC-CMs). Detailed lipid profiling revealed that inhibition of FASN resulted in significant reduction of sphingolipids and phosphatidylcholine (PC); moreover, we found that PC was the key metabolite for cell survival in hPSCs. Inhibition of FASN induced cell death in undifferentiated hPSCs via mitochondria-mediated apoptosis; however, it did not affect cell survival in hPSC-CMs, neurons, or hepatocytes as there was no significant reduction of PC. Furthermore, we did not observe tumor formation following transplantation of FASN inhibitor-treated cells. Our findings demonstrate the importance of de novo FA synthesis in the survival of undifferentiated hPSCs and suggest applications for FASN inhibition in regenerative medicine.
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Cell stem cell, 23(3) 382-395, Sep 6, 2018 Peer-reviewedThe mesoderm arises from pluripotent epiblasts and differentiates into multiple lineages; however, the underlying molecular mechanisms are unclear. Tbx6 is enriched in the paraxial mesoderm and is implicated in somite formation, but its function in other mesoderms remains elusive. Here, using direct reprogramming-based screening, single-cell RNA-seq in mouse embryos, and directed cardiac differentiation in pluripotent stem cells (PSCs), we demonstrated that Tbx6 induces nascent mesoderm from PSCs and determines cardiovascular and somite lineage specification via its temporal expression. Tbx6 knockout in mouse PSCs using CRISPR/Cas9 technology inhibited mesoderm and cardiovascular differentiation, whereas transient Tbx6 expression induced mesoderm and cardiovascular specification from mouse and human PSCs via direct upregulation of Mesp1, repression of Sox2, and activation of BMP/Nodal/Wnt signaling. Notably, prolonged Tbx6 expression suppressed cardiac differentiation and induced somite lineages, including skeletal muscle and chondrocytes. Thus, Tbx6 is critical for mesoderm induction and subsequent lineage diversification.
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Stem Cell Reports, 9(5) 1406-1414, Nov 14, 2017 Peer-reviewedLead authorCardiac regenerative therapies utilizing human induced pluripotent stem cells (hiPSCs) are hampered by ineffective large-scale culture. hiPSCs were cultured in multilayer culture plates (CPs) with active gas ventilation (AGV), resulting in stable proliferation and pluripotency. Seeding of 1 × 106 hiPSCs per layer yielded 7.2 × 108 hiPSCs in 4-layer CPs and 1.7 × 109 hiPSCs in 10-layer CPs with pluripotency. hiPSCs were sequentially differentiated into cardiomyocytes (CMs) in a two-dimensional (2D) differentiation protocol. The efficiency of cardiac differentiation using 10-layer CPs with AGV was 66%-87%. Approximately 6.2-7.0 × 108 cells (4-layer) and 1.5-2.8 × 109 cells (10-layer) were obtained with AGV. After metabolic purification with glucose- and glutamine-depleted and lactate-supplemented media, a massive amount of purified CMs was prepared. Here, we present a scalable 2D culture system using multilayer CPs with AGV for hiPSC-derived CMs, which will facilitate clinical applications for severe heart failure in the near future.
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Circulation Research, 120(10) 1558-1560, May 12, 2017 Peer-reviewedLead authorCorresponding authorA clinical study on human-induced pluripotent stem cells (hiPSCs) is underway in the ophthalmic field, and patients with advanced heart failure will be among the next targets for cell transplantation with hiPSCs. Although many approaches for production of hiPSC-derived cardiac myocytes (hiPSC-CMs) have been developed, numerous hurdles must be overcome to achieve safe and effective cardiac regenerative therapy (CRT).
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Cell Metabolism, 23(4) 663-74, Apr 12, 2016 Peer-reviewedLead authorHuman pluripotent stem cells (hPSCs) are uniquely dependent on aerobic glycolysis to generate ATP. However, the importance of oxidative phosphorylation (OXPHOS) has not been elucidated. Detailed amino acid profiling has revealed that glutamine is indispensable for the survival of hPSCs. Under glucose- and glutamine-depleted conditions, hPSCs quickly died due to the loss of ATP. Metabolome analyses showed that hPSCs oxidized pyruvate poorly and that glutamine was the main energy source for OXPHOS. hPSCs were unable to utilize pyruvate-derived citrate due to negligible expression of aconitase 2 (ACO2) and isocitrate dehydrogenase 2/3 (IDH2/3) and high expression of ATP-citrate lyase. Cardiomyocytes with mature mitochondria were not able to survive without glucose and glutamine, although they were able to use lactate to synthesize pyruvate and glutamate. This distinguishing feature of hPSC metabolism allows preparation of clinical-grade cell sources free of undifferentiated hPSCs, which prevents tumor formation during stem cell therapy.
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Cell Stem Cell, 12(1) 127-37, Jan 3, 2013 Peer-reviewedLead authorHeart disease remains a major cause of death despite advances in medical technology. Heart-regenerative therapy that uses pluripotent stem cells (PSCs) is a potentially promising strategy for patients with heart disease, but the inability to generate highly purified cardiomyocytes in sufficient quantities has been a barrier to realizing this potential. Here, we report a nongenetic method for mass-producing cardiomyocytes from mouse and human PSC derivatives that is based on the marked biochemical differences in glucose and lactate metabolism between cardiomyocytes and noncardiomyocytes, including undifferentiated cells. We cultured PSC derivatives with glucose-depleted culture medium containing abundant lactate and found that only cardiomyocytes survived. Using this approach, we obtained cardiomyocytes of up to 99% purity that did not form tumors after transplantation. We believe that our technological method broadens the range of potential applications for purified PSC-derived cardiomyocytes and could facilitate progress toward PSC-based cardiac regenerative therapy.
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Cell Stem Cell, 7(1) 11-4, Jul 2, 2010 Peer-reviewed
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Nature methods, 7(1) 61-6, Jan, 2010 Peer-reviewedSeveral applications of pluripotent stem cell (PSC)-derived cardiomyocytes require elimination of undifferentiated cells. A major limitation for cardiomyocyte purification is the lack of easy and specific cell marking techniques. We found that a fluorescent dye that labels mitochondria, tetramethylrhodamine methyl ester perchlorate, could be used to selectively mark embryonic and neonatal rat cardiomyocytes, as well as mouse, marmoset and human PSC-derived cardiomyocytes, and that the cells could subsequently be enriched (>99% purity) by fluorescence-activated cell sorting. Purified cardiomyocytes transplanted into testes did not induce teratoma formation. Moreover, aggregate formation of PSC-derived cardiomyocytes through homophilic cell-cell adhesion improved their survival in the immunodeficient mouse heart. Our approaches will aid in the future success of using PSC-derived cardiomyocytes for basic and clinical applications.
Misc.
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再生医療, 23(2) 78-83, May, 2024慢性心不全は進行性の左室機能障害であり,心不全に対する従来型の医療も近年進歩してきたが,その予後は依然として不良でありさらなる治療の改善が求められている。心筋修復を目指した細胞治療は,心不全治療の新たな手法になり得る可能性を秘めており,期待されている。成体組織幹細胞(骨髄由来単核球細胞,間葉系幹細胞など)を用いた第一世代細胞治療は安全性が確認されている一方,細胞の生着率が低く,成熟心筋細胞を形成する能力や既存の心筋細胞との電気的結合能は限定的であった。また,二重盲検の臨床試験でみられた心機能の改善は極めて小さく,それは主にパラクライン効果に起因することが示唆された。次世代の細胞治療として,ヒト胚性幹細胞やヒト人工多能性幹細胞(hiPSC)由来の心筋細胞が期待されている。これらの心筋細胞移植治療については,霊長類を用いた前臨床試験において良好な生着および周囲のホスト心筋細胞との電気生理学的結合が観察されている。細胞培養技術の進歩により,エネルギー代謝の相違を利用した心筋細胞の純化精製法により未分化細胞や非心筋細胞を除去することにより,奇形腫形成リスクが著しく低下した。また,細胞デリバリー技術の進歩により移植心筋細胞の生着率が向上するとともに,ホスト心筋との結合が観察されるようになった。現在ヒト心不全への同種hiPSC由来心筋細胞移植の臨床試験が開始されており,限られた選択肢しかない重症心不全患者にとって希望をもたらす可能性を秘めている。(著者抄録)
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カレントテラピー, 42(1) 20-25, Jan, 2024ヒト人工多能性幹細胞(ヒトiPS細胞)は再生医療や病態解明,創薬研究における有用なツールとして用いられている.心臓領域において,ヒトiPS細胞由来心筋細胞(ヒトiPS心筋細胞)を用いた移植治療は,将来的に心臓移植の代替療法になり得ると期待が高まっているが,そのためには克服すべき課題は多く存在する.特に「安全性の高い高品質ヒトiPS心筋細胞を作製する」ことは難題である.また,世界中にヒトiPS心筋細胞を届けるためには,安価かつ大量にヒトiPS心筋細胞を作製することも重要な課題である.これらの難題を乗り越え,重症心不全患者を対象にヒトiPS心筋細胞を用いた細胞移植治療は開始されており,安全性や有効性の検証が行われている.本稿では,この難題をどのように解決してきたか,ヒトiPS細胞やヒトiPS心筋細胞の代謝特性に着目して紹介する.(著者抄録)
Presentations
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KSSCR 2024 Annual Meeting, Aug 30, 2024 Invited
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Cardiovascular Bioengineering 2023 (NIH), May 29, 2023 Invited
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Oct 12, 2022 Invited
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東京医科歯科大学主催 TMDU Innovation Park BBセミナー, Jan 12, 2022 Invited
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第38回国際心臓研究学会日本部会(ISHR), Dec 10, 2021 Invited
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培養細胞が拓く未来~Nova Adbanced Cell Science Seminar2021~再生・細胞医療セッション, Sep 17, 2021 Invited
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Keio-Stanford Webinar, May 29, 2021 Invited
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PSConf 2021 International Symposium and Workshop on Development of hPSCs for Clinical Application, Apr 23, 2021 Invited
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Mar 28, 2021 Invited
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The 84th Annual Scientific Meeting of the Japanese Circulation Society Invited
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The 53rd Lake Kawaguchi Conference of Cardiology, Jul 21, 2019 Invited
Teaching Experience
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Apr, 2021 - Present
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Sep, 2019 - Mar, 2024
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Dec, 2022 - Mar, 2023
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Apr, 2020 - Nov, 2021
Professional Memberships
8Research Projects
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再生・細胞医療・遺伝子治療実現加速化プログラム, 日本医療研究開発機構, Jun, 2024 - Mar, 2026
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科学研究費助成事業, 日本学術振興会, Apr, 2023 - Mar, 2026
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科学研究費助成事業, 日本学術振興会, Apr, 2022 - Mar, 2025
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戦略研究シーズ育成事業プロジェクトリーダー, 神奈川県立産業総合研究所(KISTEC), Apr, 2023 - Mar, 2025
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再生・細胞医療・遺伝子治療実現加速化プログラム(基礎応用研究課題), 日本医療研究開発機構, Jul, 2022 - Mar, 2025