Enhanced efficiency of β cell differentiation from hESCs by changing ECM architecture

Abstract

Pancreatic beta cells derived from human embryonic stem cells (hESCs) are a promising cell source for diabetes treatment. The changes in the extracellular matrix (ECM) control the cell fate of stem cells. Thus, the combination of extracellular signal and intrinsic signaling has been developed as a method to increase differentiation efficiency from hESCs to various lineage. However, attempts to improve the differentiation efficiency into beta cells through modification of the extracellular matrix (ECM) are still insufficient. This study investigated whether the use of Polydimethylsiloxane (PDMS) was advantageous as a substrate to differentiate hESCs into pancreatic endocrine cells. Here, I found that matrigel, an ECM widely used for stem cell culture, was coated in different ways, such as mash or film-like structure on TCPS and PDMS depending on the underlying substrate. Film-like structure of Matrigel coated on PDMS (Matrigel_PDMS) provided higher cellular anchorage between cell-ECM compared to mash-like structure of Matrigel coated on TCPS (Matrigel _TCPS). On the Matrigel_PDMS expression of pancreatic endodermal markers was significantly higher during early pancreatic development of hESCs compared to the Matrigel_TCPS. Furthermore, pancreatic islet-like organoids (PIOs) derived on the Matrigel_PDMS exhibited a higher level of insulin secretion than those on the Matrigel_TCPS. In this study, I confirmed that increment of focal adhesion in hESCs cultured on Matrigel_PDMS during early pancreatic development than those cultured on Matrigel_TCPS. Increased focal adhesion induced activation of YAP/TEF1 in pancreatic endoderm cells, ultimately promoting the transcriptional activities of pancreatic endoderm-associated genes. Interestingly, YAP activation was induced in pancreatic endoderm cells via the integrin α3-FAK-CDC42-PP1A signaling. In this study, I demonstrate that the unique microstructure on Matrigel_PDMS promotes focal adhesions, and then activates YAP via Hippo independent signaling pathway, thereby leading to maturation of pancreatic endocrine development of hESCs in vitro