Enhancing Osteogenic Potential in Bone Tissue Engineering: Optimizing Pore Size in Alginate–Gelatin Composite Hydrogels

Bone tissue engineering relies on crucial scaffolds for tissue formation and stem cell differentiation. A composite scaffold of alginate‐gelatin effectively supports these processes. This study aims to design a porous alginate‐gelatin hydrogel and assess pore size effects on cell behavior, focusing on morphology, adhesion, and proliferation in distinct osteogenic environments. Hydrogels are prepared using various alginate‐gelatin concentrations: 4% alginate and 6% gelatin (4A6G) or 3% alginate and 5% gelatin (3A5G), cross‐linked with 2% CaCl2. Pore size optimization employs simple freezing and thawing cycles. Scanning electron microscopy reveals varying pore sizes: 340 µm ± 30 µm for 4A6G and 635 µm ± 25 µm for 3A5G. Stiffness measurements indicate significant differences: ≈26.3 kPa ± 0.6 KPa for 4A6G and 21.6 kPa ± 0.2 KPa for 3A5G. Cell interaction studies demonstrate higher adhesion and proliferation rates in larger‐pored hydrogels. Evaluation of bone tissue formation, including RT‐PCR, ALP activity, and ARS staining, reveal superior osteogenic potential in the 3A5G hydrogel compared to 4A6G. In conclusion, the 3A5G hydrogel (3% alginate and 5% gelatin) holds promise for bone tissue regeneration due to its biodegradability and favorable bone‐forming properties.