Mineralized Hydrogels Induce Bone Regeneration in Critical Size Cranial Defects

Sequential mineralization enables the integration of minerals within the 3D structure of hydrogels. Hydrolyzed collagen‐based hydrogels are sequentially mineralized over 10 cycles. One cycle is defined as an incubation period in calcium chloride dihydrate followed by incubation in sodium phosphate dibasic dihydrate. Separate cycles are completed at 30‐minute and 24‐hour intervals. For the gels mineralized for 30 min and 24 h, the compressive moduli increases from 4.25 to 87.57 kPa and from 4.25 to 125.47 kPa, respectively, as the cycle number increases from 0 to 10. As indicated by X‐ray diffraction (XRD) and Fourier transform infrared analysis (FTIR) analyses, the minerals in the scaffolds are mainly hydroxyapatite. In vitro experiments, which measure mechanical properties, porous structure, mineral content, and gene expression are performed to evaluate the physical properties and osteoinductivity of the scaffolds. Real time‐quantitative polymerase chain reaction (RT‐qPCR) demonstrates 4–10 fold increase in the expression of BMP‐7 and osteocalcin. The in vivo subcutaneous implantation demonstrates that the scaffolds are biocompatible and 90% biodegradable. The critical size cranial defects in vivo exhibit nearly complete bone regeneration. Cycle 10 hydrogels mineralized for 24 h have a volume of 59.86 mm3 and a density of 1946.45 HU. These results demonstrate the suitability of sequentially mineralized hydrogel scaffolds for bone repair and regeneration. Gelatin‐based hydrogels are mineralized by sequential incubation in calcium chloride and soldium phosphate to obtain mineralized scaffolds. Hydroxyapatite is formed in 3D and renders the scaffolds osteoinductive. The mineralized scaffolds are then used for critical size cranial defects in rats. Bone regeneration is significantly improved by the mineralized scaffolds compared to non‐mineralized and blank controls after 12 weeks.