Advancing cardiac tissue engineering via multicellular complexity

Abstract

Here I advance the field of tissue engineering through the additions of epicardial (EPC) and epicardial-derived cells to 2D and 3D in vitro ventricular models derived solely from human induced pluripotent stem cells (hiPSC). Through the singular addition of EPCs, EPCs undergoing EMT into epicardial-derived cells (EPC->DC), and fully differentiated epicardial-derived fibroblasts (EPD-FB) I was able to advance our understanding of the effect each of these cells on driving the maturation of hiPSC-derived cardiomyocytes (CM). First, I determined that EPCs and EPC->DC both spur CM proliferation while simultaneously promoting electrochemical maturation with no changes in cellular or tissue level force generation. In the presence of EPCs, CM maintain expression of fetal-like myofilament isoforms while the EPC->DCs had the opposite effect on CM, driving myofilament maturation in a subpopulation of CM. Next, to promote force generation in parallel with electrochemical maturation spurred by EPCs, fully differentiated EPD-FBs were added to engineered heart tissues (EHTs). These tissues showed robust electrochemical maturation, force generation, as well as more mature myofilament isoforms. Intrigued by this finding, we explored the effects of EPD-FBs alone on CM phenotype in 2D and 3D co-cultures. EPD-FBs served to drive aspects of CM maturation that EPCs did not such as CM alignment, hypertrophy, multinucleation, and tissue level force generation. However, the electrochemical functionality lagged behind the EPC co-cultures. These results suggest that the developmentally informed addition of cardiac multicellularity drives advanced cardiac tissue functionality via a sequential transition from CM proliferation to maturation. These studies benefitted from a new cohort of hiPSC-lines that I reprogrammed from the aLVCF of three female and three male donors such that outcomes could be validated on multiple lines including those of male and female biologic sex. Going forward this cohort will serve as a valuable resource for evaluating the functional and genetic differences between female and male hiPSC- derived cell types in health and disease. Indeed, I have already used this cohort to identify global differences between male and female hiPSC-CM, even in the absence of hormonal signaling.