A stress-reduced passaging technique improves the viability of human pluripotent cells

Kazutoshi Takahashi; Chikako Okubo; Michiko Nakamura; Mio Iwasaki; Yuka Kawahara; Tsuyoshi Tabata; Yousuke Miyamoto; Knut Woltjen; Shinya Yamanaka

doi:10.1016/j.crmeth.2021.100155

A stress-reduced passaging technique improves the viability of human pluripotent cells

Cell Rep Methods. 2022 Jan 24;2(2):100155. doi: 10.1016/j.crmeth.2021.100155. eCollection 2022 Feb 28.

Authors

Affiliations

¹ Department of Life Science Frontiers, Center for iPS Cell Research and Application, Kyoto University, Kyoto 606-8507, Japan.
² Gladstone Institute of Cardiovascular Disease, San Francisco, CA 94158, USA.
³ Department of Anatomy, University of California, San Francisco, San Francisco, CA 94143, USA.

Abstract

Xeno-free culture systems have expanded the clinical and industrial application of human pluripotent stem cells (PSCs). However, reproducibility issues, often arising from variability during passaging steps, remain. Here, we describe an improved method for the subculture of human PSCs. The revised method significantly enhances the viability of human PSCs by lowering DNA damage and apoptosis, resulting in more efficient and reproducible downstream applications such as gene editing and directed differentiation. Furthermore, the method does not alter PSC characteristics after long-term culture and attenuates the growth advantage of abnormal subpopulations. This robust passaging method minimizes experimental error and reduces the rate of PSCs failing quality control of human PSC research and application.

Keywords: DNA damage; human pluripotent stem cell; passage; reproducibility; viability; xeno-free culture.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Cell Differentiation / genetics
Humans
Pluripotent Stem Cells*
Reproducibility of Results