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 hrGFP and ACTA2 eGFP, respectively) that identify Acta2 +/ ACTA2 + cells as they emerge in vitro in real time during iPSC-directed differentiation. We find that Acta2 hrGFP+ and ACTA2 eGFP+ 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.
In this article, Wong, Kotton, and colleagues developed murine and human iPSC lines with fluorochrome reporters for ACTA2, allowing for isolation of putative iPSC-derived smooth muscle-like populations (iPSC-SMC) with transcriptomic profiles reminiscent of immature or synthetic SMCs. The iPSC-SMCs were used to generate cell sheets with mechanical properties within physiological ranges to serve as building blocks for multi-layered tissue constructs.