Addition of Bone‐Marrow Mesenchymal Stem Cells to 3D‐Printed Alginate/Gelatin Hydrogel Containing Freeze‐Dried Bone Nanoparticles Accelerates Regeneration of Critical Size Bone Defects

A 3D‐printed biodegradable hydrogel, consisting of alginate, gelatin, and freeze‐dried bone allograft nanoparticles (npFDBA), is developed as a scaffold for enhancing cell adhesion, proliferation, and osteogenic differentiation when combined with rat bone marrow mesenchymal stem cells (rBMSCs). This composite hydrogel is intended for the regeneration of critical‐sized bone defects using a rat calvaria defect model. The behavior of rBMSCs seeded onto the scaffold is evaluated through scanning electron microscope, MTT assays, and quantitative real‐time PCR. In a randomized study, thirty rats are assigned to five treatment groups: 1) rBMSCs‐loaded hydrogel, 2) rBMSCs‐loaded FDBA microparticles, 3) hydrogel alone, 4) FDBA alone, and 5) an empty defect serving as a negative control. After 8 weeks, bone regeneration is assessed using H&E, Masson's trichrome staining, and immunohistochemistry. The 3D‐printed hydrogel displays excellent adhesion, proliferation, and differentiation of rBMSCs. The rBMSCs‐loaded hydrogel exhibits comparable new bone regeneration to the rBMSCs‐loaded FDBA group, outperforming other groups with statistical significance (P‐value < 0.05). These findings are corroborated by Masson's trichrome staining and osteocalcin expression. The rBMSCs‐loaded 3D‐printed hydrogel demonstrates promising potential for significantly enhancing bone regeneration, surpassing the conventional clinical approach (FDBA). The substantial upregulation of the osteocalcin marker indicates a notable enhancement in new bone regeneration facilitated by the presence of rat bone marrow mesenchymal stem cells within the 3D printed hydrogel, particularly in critical‐sized bone defects. These results underscore the promising potential of this construct as a reliable tissue‐engineered scaffold with promising clinical applicability.