generation of functional CX26 gap junction forming cell derived from mouse iPS cells and its application to hereditary deafness

2019 Fiscal Year Final Research Report

• Project/Area Number: 17K16948
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Multi-year Fund
• Research Field: Otorhinolaryngology
• Research Institution: Juntendo University
• Principal Investigator: Fukunaga Ichiro 順天堂大学, 医学(系)研究科(研究院), 助教 (20746581)
• Project Period (FY): 2017-04-01 – 2020-03-31
• Keywords: GJB2 / 遺伝性難聴 / ES/iPS細胞 / 分化誘導 / 鳥類胚

Outline of Final Research Achievements

Mutation of the Gap Junction Beta 2 gene (GJB2) encoding connexin 26 (CX26) is the most frequent cause of hereditary deafness worldwide. In this study, we generated functional CX26 gap junction-forming cell (iCX26GJC) from mouse ESC/iPSC by using 3D and 2D culture. In 3D culture, iCX6GJC were observed in a part of the aggregate. Aggregates were subcultured in 2D culture, and proliferation of iCX26GJCs were observed. To investigate whether the iCx26GJC were functional, we performed scrape-loading assay and Ca2+ imaging. In the iCX26GJC culture, we observed that Lucifer yellow diffused beyond the wounded parental cells. Furthermore, spontaneous Ca2+ signaling activity was observed. Finally, we found that combination with growth factor and inhibitor has been shown to result in greater production of isolatable CX26-expressing region and higher Gjb2 mRNA levels, thereby increasing the yield of highly purified iCX26GJCs.

Free Research Field

発生生物学

Academic Significance and Societal Importance of the Research Achievements

これまで、ES/iPS細胞から感覚細胞への分化誘導法は複数報告されているが、CX26ギャップ結合を構築する蝸牛支持細胞を作製した報告はなかった。CX26をコードするGJB2遺伝子は世界最大の遺伝性難聴の原因遺伝子であり、疾患の対象となる細胞は蝸牛支持細胞などの非感覚細胞である。本研究の成果は、GJB2変異難聴を含めた、非感覚細胞を対象とする難聴の病態の解明や、治療法の開発への応用が期待される。