Discovery of retinoic acid receptor agonists as proliferators of cardiac progenitor cells through a phenotypic screening approach

Abstract

Abstract Identification of small molecules with the potential to selectively proliferate cardiac progenitor cells (CPCs) will aid our understanding of the signaling pathways and mechanisms involved and could ultimately provide tools for regenerative therapies for the treatment of post-MI cardiac dysfunction. We have used an in vitro human induced pluripotent stem cell-derived CPC model to screen a 10,000-compound library containing molecules representing different target classes and compounds reported to modulate the phenotype of stem or primary cells. The primary readout of this phenotypic screen was proliferation as measured by nuclear count. We identified retinoic acid receptor (RAR) agonists as potent proliferators of CPCs. The CPCs retained their progenitor phenotype following proliferation and the identified RAR agonists did not proliferate human cardiac fibroblasts, the major cell type in the heart. In addition, the RAR agonists were able to proliferate an independent source of CPCs, HuES6. The RAR agonists had a time-of-differentiation-dependent effect on the HuES6-derived CPCs. At 4 days of differentiation, treatment with retinoic acid induced differentiation of the CPCs to atrial cells. However, after 5 days of differentiation treatment with RAR agonists led to an inhibition of terminal differentiation to cardiomyocytes and enhanced the proliferation of the cells. RAR agonists, at least transiently, enhance the proliferation of human CPCs, at the expense of terminal cardiac differentiation. How this mechanism translates in vivo to activate endogenous CPCs and whether enhancing proliferation of these rare progenitor cells is sufficient to enhance cardiac repair remains to be investigated. Significance statement Progenitor cells are rare and difficult to isolate, and little is known about the molecular mechanisms required for their proliferation and differentiation. Identification of small molecules that selectively proliferate cardiac progenitor cells will aid in the understanding of the signaling mechanisms involved and could provide tools for regenerative therapies for the treatment of cardiac dysfunction. A phenotypic screen identified retinoic acid receptor agonists as potent proliferators of induced pluripotent stem cell-derived cardiac progenitor cells. The agonists did not proliferate human cardiac fibroblasts, the major cell type in the heart but did proliferate human pluripotent stem cells, HuES6, in a time-of-differentiation-dependent manner.