Alsobaie Sarah, Alsobaie Tamador, Mantalaris Sakis
Department of Clinical Laboratory Science, King Saud University, Riyadh, 11451, Saudi Arabia.
Biological Systems Engineering Laboratory, Department of Chemical Engineering, Imperial College London, London, SW7 2AZ, UK.
Stem Cells Cloning. 2022 Jul 2;15:43-52. doi: 10.2147/SCCAA.S365776. eCollection 2022.
High-efficacy single-cell cloning of human-induced pluripotent cells (IPSCs) remains a major challenge. The development of a culture method that supports single-cell passaging while maintaining reproducibility, homogeneity, scalability, and cell expansion to clinically relevant numbers is necessary for clinical application.
To address this issue, we combined the use of the rho-associated protein kinase (ROCK) inhibitor Y-27632 and hypoxic conditions in culture to produce a novel, efficient single-cell culture method for human IPSCs and embryonic stem cells.
Through immunocytochemistry, alkaline phosphatase assays, and flow cytometry, we demonstrated that our method enabled high single-cell proliferation while maintaining self-renewal and pluripotency abilities.
We showed the beneficial effect of the interaction between hypoxia and ROCK inhibition in regulating cell proliferation, pluripotency, and single-cell survival of pluripotent cells.
人类诱导多能干细胞（iPSC）的高效单细胞克隆仍然是一项重大挑战。开发一种支持单细胞传代同时保持可重复性、同质性、可扩展性以及将细胞扩增至临床相关数量的培养方法对于临床应用而言是必要的。
为解决这一问题，我们在培养过程中联合使用了Rho相关蛋白激酶（ROCK）抑制剂Y-27632和低氧条件，以产生一种用于人类iPSC和胚胎干细胞的新型高效单细胞培养方法。
通过免疫细胞化学、碱性磷酸酶检测和流式细胞术，我们证明我们的方法能够实现高单细胞增殖，同时保持自我更新和多能性能力。
我们展示了低氧与ROCK抑制之间的相互作用在调节多能细胞的细胞增殖、多能性和单细胞存活方面的有益效果。
