GelMA hydrogels guide lineage-specific self-assembly of endothelial and interstitial cells into valve-mimetic tissue...

Heart valve diseases cause progressive loss of function and remain a leading contributor to cardiovascular morbidity. Tissue-engineered heart valves (TEHVs) offer a potential solution, but reproducing the extracellular matrix architecture and mechanical microenvironment of native leaflets remains a critical challenge. Gelatin methacryloyl (GelMA) hydrogels provide tunable structural and mechanical properties, offering a promising platform. However, their ability to direct lineage-specific tissue reconstruction remains unclear. Here, we investigated the responses of human umbilical vein endothelial cells (HuVECs) and immortalized human aortic valve interstitial cells (iHAVICs) cultured either on the surface (2D) or encapsulated within (3D) GelMA hydrogels. Both cell types maintained high viability across dimensional culture conditions (2D and 3D). In 2D, HuVECs proliferated and assembled into highly interconnected vascular plexuses containing luminal structures, while iHAVICs formed spongy, mesh-like frameworks. In 3D, HuVECs developed complex, spatially interconnected vascular-like networks, and iHAVICs transitioned from spherical to spindle morphologies, aligning into linear and fibrous skeletons. These findings establish that GelMA substrates direct cell-type-specific morphogenesis, yielding vascular networks from HuVECs and valve-like stromal architectures from iHAVICs in a dimensionality-dependent manner. This study demonstrates the feasibility of using GelMA hydrogels to support distinct, tissue-relevant architectures, providing a basis for future strategies in TEHVs.