Controlling Chemical and Mechanical Cues to Augment Endothelial Fate from Human Induced Pluripotent Stem Cells

The efficient differentiation of functional endothelial cells (ECs) from human induced pluripotent stem cells (hiPSCs) is pivotal for their therapeutic potential. Aims to increase differentiation efficiency relies on the presentation of physiologically relevant cues including controlling the mechanical micro-environment. While several studies have demonstrated the effects of substrate stiffness on the differentiation efficiency of mesenchymal stem cells towards adipo/osteogeneic linages, less is known on the effects of substrate rigidity of hiPSCs differentiating towards vascular lineages.

Here we investigated whether mechanical dosing of hiPSCs during early stages of mesoderm induction biases endothelial differentiation. We identified the gene expression of the connective tissue growth factor (CTGF) and localization of the Yes-associated protein (YAP) when hiPSCs are cultured on polydimethylsiloxane (PDMS) (Young’s Modulus E ~ 3kPa or 1.7 MPa) or polystyrene substrates (E ~ 3 GPa). We found that mechanically dosing hiPSCs on 1.7 MPa substrates in mesoderm inducing media, increased Brachyury (T) expression when compared to culture on 3 kPa PDMS or tissue culture plates. In addition, when these mechanically dosed cells were continually differentiated towards a vascular phenotype, we found that cells primed on 1.7 MPa consistently lead to a robust endothelial phenotype. We conclude that differentiating hiPSCs are mechanosensitive through YAP and CTGF activation, leading to enhanced T expression during mesoderm induction, and robust endothelial differentiation.