Porous GelMA loaded with melatonin accelerates bone fracture healing by activating osteoblasts

Abstract

Objective(s): This study aimed to investigate the effects of melatonin at varying concentrations on the biological activities of bone marrow mesenchymal stem cells (BMSCs), as well as the therapeutic efficacy of melatonin-loaded porous gelatin methacryloyl (GelMA) in promoting bone fracture healing. Materials and Methods: BMSCs were cultured in the presence of melatonin at concentrations of 0, 0.01, 0.1, 1, and 10 μM to evaluate the dose-dependent effects of melatonin. EDU assay was performed to detect BMSC proliferation. Scanning electron microscopy (SEM) was used to characterize the morphological properties of the porous GelMA. Alkaline phosphatase (ALP) staining and alizarin red (AR) staining were conducted to assess the osteogenic differentiation potential of BMSCs. Furthermore, the expression levels of osteogenic-related proteins, including ALP, osteocalcin (OCN), and runt-related transcription factor 2 (RUNX2), were analyzed by Western blot (WB), immunofluorescence staining, and immunohistochemistry assays, respectively. Results: The results showed that the cells obtained from bone marrow met the criteria of mesenchymal stem cells (MSC). BMSC could form sunflower-like colonies and strongly expressed stem cell-related genes. In addition, BMSC showed strong ability of osteogenic, chondrogenic and adipogenic differentiation. melatonin at certain concentration (1 μmol/L) significantly promoted cell proliferation and colony-forming rate. In vitro, appropriate melatonin concentration can increase the rate of osteogenic differentiation of BMSC and promote the expression of osteo-related proteins. In vivo, the porous GelMA loaded with melatonin can accelerate bone repair of skull defects. Conclusion: Melatonin-loaded porous GelMA can effectively accelerate bone fracture healing, thereby holding promising translational potential as a novel therapeutic strategy.