Extracellular matrix-mediated differentiation of human embryonic stem cells: differentiation to insulin-secreting beta cells.

Stem cells have tremendous potential for treating various human diseases. Protocols have been established to differentiate stem cells into specific lineages through the provision of signals in the form of growth factors, cytokines, or small molecules. Herein we investigate an alternative strategy for directed differentiation of human embryonic stem cells (hESCs)--extracellular-matrix (ECM) mediated differentiation. Decellularized ECM and conditioned media from the appropriate committed cell lines are used to differentiate stem cells to the required phenotype. Applying this strategy to differentiate hESCs to pancreatic beta cells, we have obtained functional cells that secreted insulin in a glucose-responsive manner, and were able to recover normoglycemia in a streptozotocin (STZ)-induced diabetic mouse model. ECM-mediated differentiation was also demonstrated to be effective for the differentiation of hESCs into kidney tubule cells and cardiomyocytes. Gene expression studies suggested the involvement of integrins and catenins in the beta cell differentiation process; in particular, α1, αv, and β1 integrins, and β-catenin showed the highest upregulation. To further elucidate the biochemical and mechanical cues that have led to effective hESC differentiation to beta cells, we have employed an artificial system that allowed for variation of matrix stiffness and combination of individual ECM proteins at various ratios. The differentiation response of hESCs to the native ECM could be approximated by optimizing this system.