Establishment of iPSC disease models with mitochondrial DNA mutations using genome editing technology.

2023 Fiscal Year Final Research Report

• Project/Area Number: 21K06848
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Review Section: Basic Section 48040:Medical biochemistry-related
• Research Institution: Fujita Health University
• Principal Investigator: Yahata Naoki 藤田医科大学, 医学部, 講師 (60450607)
• Co-Investigator(Kenkyū-buntansha): 大熊 真人 藤田医科大学, 医学部, 講師 (50329710) ; 中嶋 和紀 岐阜大学, 糖鎖生命コア研究所, 准教授 (10442998)
• Project Period (FY): 2021-04-01 – 2024-03-31
• Keywords: mtDNA / iPS細胞 / TALEN / ミトコンドリア

Outline of Final Research Achievements

In this study, we aimed to clarify the cellular phenotypes of skeletal myocytes and neuronal cells differentiated from MELAS-iPSCs with mtDNA mutation in order to elucidate the pathophysiological mechanism of mitochondrial diseases.
Skeletal myocytes were differentiated from MELAS-iPSCs containing various proportions of mutant mtDNA. Metabolic profiles in the culture supernatants were analyzed using a LC-MS, revealing differences in their profiles depending on the presence or absence of mutant mtDNA. We also generated iN-iPSCs in which the expression of NGN2 could be induced by the addition of Doxycycline (Dox). Neuronal cells capable of generating action potentials could be obtained from iN-iPSCs with mutant mtDNA. We tried to create iPSCs in which mtDNA mutation could be introduced in a drug-dependent manner using the latest base editing tool. By adding Dox, we could introduce a point mutation into the ND4 gene in normal iPSCs.

Free Research Field

幹細胞創薬生命科学

Academic Significance and Societal Importance of the Research Achievements

本研究では、分化誘導が難しい、ミトコンドリア病の原因となる変異mtDNAを有するiPS細胞から、骨格筋細胞および神経細胞への分化誘導に成功した。さらに、塩基編集技術を用いることで、正常iPS細胞に変異mtDNAを導入することに成功した。本研究で作製したこれらのiPS細胞モデルは、今後ミトコンドリア病の病態解析、さらには治療法の開発に寄与すると考えられる。