Quantifying the Vasculogenic Potential of Induced Pluripotent Stem Cell-Derived Endothelial Progenitors in Collagen Hydrogels

Abstract

Induced pluripotent stem cell-derived endothelial progenitors (iPSC-EPs) have emerged as a promising candidate cell source for patient-specific ischemic therapies. Before these cells can be appropriately deployed in a clinical setting, it is imperative to study their assembly into functional vascular networks in extracellular matrix (ECM)-mimicking, three-dimensional (3D) microenvironments. To elucidate the interactions of iPSC-EPs with the ECM, we examined how in vitro modulation of structural protein density, the presence of angiogenic growth factors, and relative proteolytic activity affected the vasculogenic potential of these progenitors, that is, their ability to self-assemble into vessel-like networks. We found that the addition of a ROCK pathway inhibitor and exogenous vascular endothelial growth factor (VEGF) are imperative for inducing robust iPSC-EP vasculogenesis in collagen hydrogels. Under these conditions, 3D vascular-like networks containing VE-cadherin-expressing lumens formed within a week of culture. To quantify this 3D vessel-like network, we developed a computational pipeline to analyze network length, connectivity, and average lumen diameter. Increasing the concentration of collagen in the hydrogels abrogated network formation and encouraged the formation of disconnected, large-diameter lumens. This phenomenon was in part related to the cells' proteolytic capacity and the hydrogels' properties, specifically hydrogel deformability and pore size. In conclusion, we demonstrate that the vasculogenic potential of iPSC-EPs is regulated by cell–matrix interactions and the matrix properties of collagen hydrogels.

Impact Statement

Our work reinforces the role of extracellular matrix (ECM) density and matrix metalloprotease activity on the formation of microvasculature from induced pluripotent stem cell (iPSC)-derived vascular cells. The cell–matrix interactions discussed in this study underscore the importance of understanding the role of mechanoregulation and matrix degradation on vasculogenesis and can potentially drive the development of ECM-mimicking angiogenic biomaterials. Furthermore, our work has broader implications concerning the response of iPSC-derived cells to the mechanics of engineered microenvironments. An understanding of these interactions will be critical to creating physiologically relevant transplantable tissue replacements.