Generation of a Purified iPSC-Derived Smooth Muscle-like Population for Cell Sheet Engineering.

Induced pluripotent stem cells (iPSCs) provide a potential source for the derivation of smooth muscle cells (SMCs); however, current approaches are limited by the production of heterogeneous cell types and a paucity of tools or markers for tracking and purifying candidate SMCs. Here, we develop murine and human iPSC lines carrying fluorochrome reporters (Acta2 and ACTA2, respectively) that identify Acta2/ACTA2 cells as they emerge in vitro in real time during iPSC-directed differentiation. We find that Acta2 and ACTA2 cells can be sorted to purity and are enriched in markers characteristic of an immature or synthetic SMC. We characterize the resulting GFP populations through global transcriptomic profiling and functional studies, including the capacity to form engineered cell sheets. We conclude that these reporter lines allow for generation of sortable, live iPSC-derived Acta2/ACTA2 cells highly enriched in smooth muscle lineages for basic developmental studies, tissue engineering, or future clinical regenerative applications.