Semi-exhaustive functional screening of cell-cell interaction factors in human iPSC mini-liver

2019 Fiscal Year Final Research Report

• Project/Area Number: 18K19589
• Research Category: Grant-in-Aid for Challenging Research (Exploratory)
• Allocation Type: Multi-year Fund
• Review Section: Medium-sized Section 55:Surgery of the organs maintaining homeostasis and related fields
• Research Institution: The University of Tokyo (2019); Yokohama City University (2018)
• Principal Investigator: Sekine Keisuke 東京大学, 医科学研究所, 准教授 (00323569)
• Project Period (FY): 2018-06-29 – 2020-03-31
• Keywords: iPS細胞

Outline of Final Research Achievements

In this study, we constructed an efficient functional screening system for human iPSC mini-liver using CRISPR / CAS9 method. In order to efficiently carry out gene deletion, iCrispr-iPS cells in which a gene capable of inducing Cas9 expression by addition of doxycycline was introduced into the AAVS1 locus of human iPS cells by homologous recombination were established for a several iPSC lines. We performed a functional analysis using knockout iPSCs of genes that are thought to be involved in the residual undifferentiated cells in differentiated cell products, and succeeded in identifying the factors involved in the predisposed to remain the undifferentiated cells in differentiated cell products. In addition, we made several knockout iPSCs for urea cycle-related genes, which are also the cause of urea cycle abnormality, and performed functional verification.

Free Research Field

幹細胞生物学

Academic Significance and Societal Importance of the Research Achievements

本研究ではiPSCを用いて極めて効率よく遺伝子欠損細胞を作製し機能検証する手法を確立した。この技術を用いて、移植医療に用いる再生医療等製品における安全性の確保において極めて重要な未分化iPSCの残存に関わる因子を同定した。また、先天的な代謝性疾患の原因でもある尿素サイクル関連遺伝子について複数のノックアウトiPSCを作製し、尿素サイクルのそれぞれの酵素欠損によってバイパスされる代謝経路が異なる可能性が示唆された。今後、尿素サイクルに異常症に対する薬剤療法や代謝改善のための検証が可能になると期待される。以上のことから、本手法を用いることで様々な細胞における効率よい遺伝子機能の検証が期待される。