Simultaneous binding of bFGF to both FGFR and integrin maintains properties of primed human induced pluripotent stem cells

Introduction: Basic fibroblast growth factor (bFGF, FGF2) and integrin a6b1 are important for maintaining the pluripotency of human pluripotent stem cells (hPSCs). Although bFGF-integrin binding contributes to biofunctions in cancer cells, the relationship in hPSCs remains unclear.Methods: To investigate the relationship between bFGF and integrin in human induced pluripotent stem cells (hiPSCs), we generated recombinant human bFGF wild-type and mutant proteins, that do not bind to integrin, FGFR, or both. We then cultured hiPSCs with these recombinant bFGF proteins. To evaluate the abilities of recombinant bFGF proteins in maintaining hPSC properties, pluripotent markers, ERK activity, and focal adhesion structure were analyzed through flow cytometry, immunofluorescence (IF), and immunoblotting (IB).Result: We identified an interaction between bFGF and integrin a6b1 in vitro and in hiPSCs. The integrin non-binding mutant was incapable of inducing the hPSC properties, such as proliferation, ERK activity, and large focal adhesions at the edges of hiPSC colonies. Signaling induced by bFGF-FGFR binding was essential during the first 24 h after cell seeding for maintaining the properties of hPSCs, followed by a shift towards intracellular signaling via the bFGF-integrin interaction. The mixture of the two bFGF mutants also failed to maintain hPSC properties, indicating that bFGF binds to both FGFR and integrin.Conclusion: Our study demonstrates that the integrin-bFGF-FGFR ternary complex maintains the prop-erties of hPSCs via intracellular signaling, providing insights into the functional crosstalk between bFGF and integrins in hiPSCs. (c) 2023, The Japanese Society for Regenerative Medicine. Production and hosting by Elsevier B.V.