Decellularized muscle-derived hydrogels support in vitro cardiac microtissue fabrication

Cardiovascular research has considerably benefited from in vitro models of cardiac tissue. Two important elements of these constructs, cardiac cells and the extracellular matrix (ECM), play essential roles that mimic the structural and functional aspects of myocardium. Here, we compared decellularized ECM from cardiac muscle (D-CM), skeletal muscle (D-SM), aorta (D-Ao), liver (D-Liv), small intestine submucosa (D-SIS), and human umbilical cord (D-hUC) in terms of their biocompatibility and potential for differentiation of human embryonic stem cell-derived cardiac progenitor cells (hESC-derived CPCs) to cardiovascular lineage cells. The decellularization procedure successfully removed resident cells of the tissues but preserved ECM components such as laminin and fibronectin, which was identified by histological studies of decellularized tissue (D-tissues) and immunostaining. Encapsulation of hESC-derived CPCs and human umbilical vein endothelial cells within hydrogels that were obtained from all decellularized tissues did not induce cytotoxicity after 10 days of culture. Upregulation of cardiac specific genes, cTNT and αMHC, as well as the presence of cTNT⁺ cardiomyocytes were also observed in CPCs cultured on D-CM, D-SM, D-Liv, and D-SIS, which showed their support for cardiogenic differentiation. However, D-CM provided substantially higher expression of cardiac markers compared to the other D-tissues. The endothelial and smooth muscle specific genes, CD31 and PDGFRα, were upregulated in cells cultured on D-Ao and D-hUC, which reflected their support for vascular lineage cell differentiation. In conclusion, it might be imperative to use decellularized tissue of muscle origins in combination with naturally derived vascular tissues to generate in vitro vascularized human cardiac microtissues.