ROLE OF MBT FAMILY GENES IN STABILITY OF UNDIFFERENTIATED STATE OF HUMAN PLURIPOTENT STEM CELLS

• Project/Area Number: 15K01280
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Biomedical engineering/Biomaterial science and engineering
• Research Institution: Shinshu University
• Principal Investigator: Daihachiro Tomotsune 信州大学, 学術研究院医学系, 助教 (80283802)
• Research Collaborator: Yue Fengming ; Sasaki Katsunori ; Hirashima Kanji ; Fujii Masako ; Nakamura Shunsuke
• Project Period (FY): 2015-04-01 – 2019-03-31
• Project Status: Completed (Fiscal Year 2018)
• Budget Amount: ¥4,550,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥1,050,000); Fiscal Year 2017: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000); Fiscal Year 2016: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2015: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
• Keywords: 多能性幹細胞 / iPS細胞 / 未分化維持 / ヒト多能性幹細胞 / ヒトiPS細胞 / エピジェネティック / ポリコーム / ポリコーム遺伝子群 / 未分化状態

Outline of Final Research Achievements

Maintenance of undifferentiated state is a critical requirement for regenerative medicine using pluripotent stem cells. In this study, we created the spontaneous differentiation by removing MEF cells, and then analyzed global changes of gene expression during the spontaneous differentiation using DNA microarray. As results of the microarray analysis using four human pluripotent stem cell lines, we found that some polycomb group genes were commonly up- regulated during the collapse of undifferentiated state. Then we analyzed function of the spontaneous differentiation-induced polycomb group genes in pluripotent stem cells by CRISPR-Cas9 mediated knockout. These knockout cells showed tendency to differentiate more frequently than control. Therefore these polycomb group proteins may function as defense against the spontaneous differentiation. To evaluate the proposed function, we analyzed cell lines over-expressing the polycomb genes. The results supported the hypothesis.

Academic Significance and Societal Importance of the Research Achievements

ヒトiPS細胞などの多能性幹細胞は、様々な細胞に分化する能力を持つため、疾患のある組織を健康な組織で置き換える再生医療において、その細胞供給源になると期待されている。多分化能を保持するためには、未だ分化してない未分化状態である必要があるため、未分化維持機構の解明は医学的な重要性がある。本研究では、ポリコーム遺伝子の二つ、L3MBTL1とFMBT2は未分化状態が不安定になる際に発現して、未分化に戻す役割を果たしていることが示された。これは分化が正しい方向にのみ進むことを補助しているとも言えるもので、再生医療だけでなく、分化研究においても意義深い。

Research Products

• [Journal Article] Simple Isolation of Pancreatic Progenitor Cells from Human Induced Pluripotent Stem Cells Using the ALDEFLUOR. (2017)
  • Author(s): Nakamura, S., Takizawa-Shirasawa S., Yokoyama T., Tomotsune, D.*, Sasaki,K.
  • Journal Title: Stem Cell Adv Res Ther Volume: 1
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Enrichment of Pluripotent Stem Cell-Derived Hepatocyte-Like Cells by Ammonia Treatment. (2016)
  • Author(s): Tomotsune, D.*, Hirashima K., Fujii M., Yue F., Matsumoto K., Takizawa-Shirasawa S., Yokoyama T., Sasaki K.
  • Journal Title: PLOS ONE Volume: 111
  • NAID: 120007113477
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Presentation] ROLE OF POLYCOMB GROUP GENES IN STABILITY OF UNDIFFERENTIATED STATE OF HUMAN PLURIPOTENT STEM CELLS (2018)
  • Author(s): Tomotsune, D. Yoshitome, A., Yue,F., Hirashima,K., Takizawa-Shirasawa, S., Yokoyama, T., Sasaki, K.
  • Organizer: The 16th ISSCR Annual Meeting
  • Int'l Joint Research
• [Presentation] 網羅的遺伝子解析を用いた低コストな幹細胞誘導法開発の試み (2017)
  • Author(s): 友常 大八郎、平島 寛司、吉留 明子、藤井 昌子、滝澤 佐季子、横山 忠幸、松本 健、岳 鳳鳴、佐々木 克典
  • Organizer: 第16回 日本再生医療学会総会
  • Place of Presentation: 仙台、仙台国際センター
  • Year and Date: 2017-03-07
• [Presentation] Enrichment of pluripotent stem cell derived hepatocyte-like cells by lanford medium and low-molecular compound (2017)
  • Author(s): Tomotsune D, Yoshitome A, Hirashima K, Fujii M, Yue F, Matsumoto K, Takizawa-Shirasawa S, Yokoyama T, Nakamura S, Sakai S, Sasaki K.
  • Organizer: The 14th ISSCR Annual Meeting
  • Int'l Joint Research
• [Presentation] ANALYSIS AND COMPARISON OF EMBRYOID BODY FORMATION FROM HUMAN PLURIPOTENT STEM CELLS (2015)
  • Author(s): Tomotsune, D. Mogi, A. Yue, F. Matsushima, A. Sakai, Y. Takizawa-Shirasawa, S. Yokoyama, T. isashi Shimizu, H. Yoshitome, A. Nakamura, S. Sakai, S. Hirashima, K. Fujita, D. Sasaki, K
  • Organizer: 国際幹細胞学会（ISSCR）
  • Place of Presentation: ストックホルム
  • Year and Date: 2015-06-24
  • Int'l Joint Research
• [Remarks] 信州大学医学部組織発生学教室
  • URL: http://www.shinshu-u.ac.jp/faculty/medicine/chair/i-1kaibo/
• [Remarks] 信州大学バイオメディカル研究所
  • URL: http://www.shinshu-u.ac.jp/institution/ibs/
• [Remarks] 信州大学医学部 組織発生学教室
  • URL: http://www.shinshu-u.ac.jp/faculty/medicine/chair/i-1kaibo/
• [Remarks] 信州大学 バイオメディカル研究所
  • URL: http://www.shinshu-u.ac.jp/institution/ibs/