Nanoparticle-based hybrid scaffolds for deciphering the role of multimodal cues in cardiac tissue engineering

Myocardial microenvironment plays a decisive role in guiding the function and fate of cardiomyocytes, and engineering this extracellular niche holds great promise for cardiac tissue regeneration. Platforms utilizing hybrid hydrogels containing various types of conductive nanoparticles have been a critical tool for constructing engineered cardiac tissues with outstanding mechanical integrity and improved electrophysiological properties. However, there has been no attempt to directly compare the efficacy of these hybrid hydrogels and decipher the mechanisms behind how these platforms differentially regulate cardiomyocyte behavior. Here, we employed gelatin methacryloyl (GelMA) hydrogels containing three different types of carbon-based nanoparticles: carbon nanotubes (CNTs), graphene oxide (GO), and reduced GO (rGO), to investigate the influence of these hybrid scaffolds on the structural organization and functionality of cardiomyocytes. Using immunofluorescent staining for assessing cellular organization and proliferation, we showed that electrically conductive scaffolds (CNT- and rGO-GelMA compared to relatively nonconductive GO-GelMA) played a significant role in promoting desirable morphology of cardiomyocytes and elevated the expression of functional cardiac markers, while maintaining their viability. Electrophysiological analysis revealed that these engineered cardiac tissues showed distinct cardiomyocyte phenotypes and different levels of maturity based on the substrate (CNT-GelMA: ventricular-like, GO-GelMA: atrial-like, and rGO-GelMA: ventricular/atrial mixed phenotypes). Through analysis of gene-expression patterns, we uncovered that the engineered cardiac tissues matured on CNT-GelMA and native cardiac tissues showed comparable expression levels of maturation markers. Furthermore, we demonstrated that engineered cardiac tissues matured on CNT-GelMA have increased functionality through integrin-mediated mechanotransduction (via YAP/TAZ) in contrast to cardiomyocytes cultured on rGO-GelMA