Matrix Stiffness of GelMA Hydrogels Regulates Lymphatic Endothelial Cells toward Enhanced Lymphangiogenesis

Lymphatic vessel regeneration is crucial for various tissue engineering strategies, particularly in resolving inflammation and restoring tissue homeostasis. In our study, we focused on investigating how hydrogel matrix stiffness influences lymphatic endothelial cells (LECs) in promoting lymphatic vessel regeneration. Gelatin methacrylate (GelMA) was chosen as our biomaterial due to its versatility in tissue engineering and biofabrication. We fabricated GelMA hydrogels at concentrations of 5, 7.5, and 15% (w/v) with corresponding Young's modulus values of 1.55 kPa (soft matrix), 12.02 kPa (medium matrix), and 48.50 kPa (stiff matrix). Among these, the 7.5% GelMA hydrogel exhibited optimal stiffness for promoting lymphangiogenesis. LECs seeded either on the hydrogel surface or within spontaneously formed a more stable lymphatic capillary network compared with other GelMA formulations. Furthermore, we investigated the enhancement of lymphangiogenesis by incorporating VEGF-C into the GelMA hydrogel, leveraging the synergistic effects of mechanical and chemical cues. Our results underscored the critical role of FAK-phosphorylation in this process; treatment with an FAK-specific inhibitor prevented the formation of tube-like structures by LECs and attenuated the expression of lymphatic markers. Overall, our findings highlight how the mechanical and chemical cues provided by GelMA hydrogels can effectively regulate LEC behavior toward enhanced lymphangiogenesis via the integrin/FAK mechanotransduction pathway. This study proposes a promising strategy for developing hydrogel-based scaffolds or bioinks tailored to promote lymphatic vessel regeneration in therapeutic applications.