Development of a bovine gelatin-kappa carrageenan-based dual network biomimetic hydrogel for chondrogenic differentiation of mesenchymal stem cells

Direct stem cell delivery for cartilage tissue engineering faces significant drawbacks, including loss of cells via circulation and poor viability in a hostile microenvironment. Hence, scaffold-based approaches for stem cell delivery are gaining significant momentum. In this study, composite hydrogel films composed of gelatin and kappa-carrageenan dually crosslinked with glutaraldehyde and potassium chloride have been developed through the solvent casting method. The protein-polysaccharide composite mimics the natural extracellular matrix of native cartilage and the synergistic effects of covalent and ionic crosslinking provide mechanical strength, stability, and satisfactory biological performance. The physicochemical properties of the composite were analyzed using SEMEDS, AFM, FTIR, XPS, and XRD. Rheological analysis revealed self-healing properties of the film and mechanical analysis demonstrated the ultimate tensile strength to be 13.49 +/- 2.89 MPa, which mechanically mimics the native cartilage. The composite film remained stable for approximately 4 weeks in PBS, validating its stability. Biological assessments of the film after 7 days of culture demonstrated its long-term cytocompatibility, showing cell viability of 97.56 +/- 0.21 %, cell adhesion was observed using FESEM micrographs, and cell proliferation through Live/Dead assay. The dually crosslinked biomimetic composite films aided in chondrogenic differentiation, as confirmed using collagen II staining and TGF-beta expression studies, and hence demonstrate promising potential for cartilage tissue regeneration.