Stanford Cardiovascular Medicine and Cardiovascular Institute, Stanford School of Medicine, Stanford University, Falk CVRC, Room CV273, MC 5406 300 Pasteur Drive, Stanford, CA, 94305, USA.
Department of Developmental and Regenerative Biology, Icahn School of Medicine at Mount Sinai, Mount Sinai, New York, NY, 10029, USA.
Stem Cell Res Ther. 2019 Jul 29;10(1):228. doi: 10.1186/s13287-019-1319-5.
Human-induced pluripotent stem cells (hiPSCs) show a great promise as a renewable source of cells with broad biomedical applications. Since insulin has been used in the maintenance of hiPSCs, in this study we explored the role of insulin in culture of these cells.
We report conditions for insulin starvation and stimulation of hiPSCs. Crystal violet staining was used to study the adhesion and proliferation of hiPSCs. Apoptosis and cell cycle assays were performed through flow cytometry. Protein arrays were used to confirm phosphorylation targets, and mRNA sequencing was used to evaluate the effect of transcriptome.
Insulin improved the seeding and proliferation of hiPSCs. We also observed an altered cell cycle profile and increase in apoptosis in hiPSCs in the absence of insulin. Furthermore, we confirmed phosphorylation of key components of insulin signaling pathway in the presence of insulin and demonstrated the significant effect of insulin on regulation of the mRNA transcriptome of hiPSCs.
Insulin is a major regulator of seeding, proliferation, phosphorylation and mRNA transcriptome in hiPSCs. Collectively, our work furthers our understanding of human pluripotency and paves the way for future studies that use hiPSCs for modeling genetic ailments affecting insulin signaling pathways.
人类诱导多能干细胞（hiPSCs）作为具有广泛生物医学应用的可再生细胞来源，具有巨大的应用潜力。由于胰岛素已被用于维持 hiPSCs，因此在本研究中我们探讨了胰岛素在这些细胞培养中的作用。
我们报告了胰岛素饥饿和刺激 hiPSCs 的条件。结晶紫染色用于研究 hiPSCs 的黏附和增殖。通过流式细胞术进行凋亡和细胞周期分析。蛋白质芯片用于确认磷酸化靶标，mRNA 测序用于评估转录组的影响。
胰岛素改善了 hiPSCs 的接种和增殖。我们还观察到在没有胰岛素的情况下，hiPSCs 的细胞周期谱发生改变，凋亡增加。此外，我们在胰岛素存在的情况下证实了胰岛素信号通路关键成分的磷酸化，并证明了胰岛素对 hiPSCs mRNA 转录组调控的显著影响。
胰岛素是 hiPSCs 接种、增殖、磷酸化和 mRNA 转录组的主要调节剂。总之，我们的工作加深了我们对人类多能性的理解，并为未来使用 hiPSCs 模拟影响胰岛素信号通路的遗传疾病的研究铺平了道路。
