Interpenetrating Polymer Network HA/Alg-RGD Hydrogel: An Equilibrium of Macroscopic Stability and Microscopic Adaptability for 3D Cell Growth and Vascularization

Two-dimensional (2D) cell culture methods dominate the current research. However, the inherent responsiveness of cells to their native three-dimensional (3D) microenvironment necessitates a paradigm shift toward the development of advanced hydrogels that faithfully mimic the intricacies of the extracellular matrix (ECM) and enable continuous cell-ECM interactions. To address the constraints of traditional static hydrogel networks that impede effective cell-matrix and cell-cell interactions, and to tackle the inherent stability issues associated with dynamically cross-linked hydrogels, which have become a pressing concern. Herein, we present an interpenetrating polymer network (IPN) hydrogel (HA/Alg-RGD hydrogel) that combines a physically cross-linked network between alginate and calcium ions (Alg-Ca²⁺) for the enhanced cell growth adaptability with a chemically cross-linked hyaluronic acid (HA) network to ensure macroscopic stability during cell culture. The incorporation of arginine-glycine-aspartic peptide modified alginate (Alg-RGD) further facilitates cell adhesion and improves the cell-hydrogel interaction. Notably, this IPN hydrogel demonstrates mechanical stability and enables cell spreading and growth within its structural framework. Leveraging the reversible characteristics of the ionically cross-linked Alg-Ca²⁺ network within IPN hydrogels, we demonstrate the feasibility of the gelatin sacrificial solution for 3D printing purposes within the hydrogel matrix. Subsequent UV-induced covalent cross-linking enables the fabrication of vascularized microfluidic channels within the resulting construct. Our results demonstrate endothelial cell spreading and spontaneous cell sprouting within the hydrogel matrix, thus highlighting the efficacy of this IPN hydrogel system in facilitating 3D cell growth. Additionally, our study emphasizes the potential of 3D printed constructs as a promising approach for vascularization in tissue engineering. The importance of RGD peptides in promoting favorable cell-hydrogel scaffold interactions is also highlighted, emphasizing their critical role in optimizing biomaterial-cell interfaces.