Insulin stimulates PI3K/AKT and cell adhesion to promote the survival of individualized human embryonic stem cells.

Insulin is present in most maintenance media for human embryonic stem cells, but little is known approximately its essential role in the cell survival of individualized cells during passage. In this report, we show that insulin suppresses caspase cleavage and apoptosis after dissociation. Insulin activates IGF receptor and PI3K/AKT cascade to promote cell survival, and its function is independent of ROCK kinase regulation. During niche reformation after passaging, insulin activates integrin that is essential for cell survival. IGF receptor co-localizes with focal adhesion complex and stimulates protein phosphorylation involved in focal adhesion formation. Insulin promotes cell spreading on matrigel-coated surfaces and suppresses myosin light chain phosphorylation. Further study showed that insulin is also required for the cell survival on E-cadherin coated surface and in suspension, indicating its essential role in cell-cell adhesion. This work highlights insulin's complex roles in signal transduction and niche re-establishment in hESCs. SIGNIFICANCE STATEMENT: This study demonstrates the essential and unique role played by insulin in cell survival and adhesion in human embryonic stem cells after individualization. The insulin regulation is independent of ROCK kinase, and is the prerequisite for the ROCK inhibitor-dependent cell survival. As the only growth factor required for cell survival in the maintenance media, insulin activates IGF1R/PI3K/AKT pathway and inhibits caspase activation. Insulin also stimulates integrin activation, and promotes cell-matrix and cell-cell adhesion. This study reveals insulin/IGF pathway as the central player in cell survival and niche re-formation during hESC passaging. © AlphaMed Press 2019.