The effect of nano-natural biomaterial on stimulation of RADMSCs toward osteogenesis: an in vitro study

Abstract In recent years, natural biomaterials have been the focal point of most biomedical investigations. To prepare the Ostrich Eggshell/Hydroxyapatite (OE/HA) powder scaffolds, OE powder was immersed in 1% Triton X-100 for 24 h and then autoclaved twice. Medical grade HA was combined with pulverized OE at different ratios to yield three different OE/HA powder scaffolds. The resultant scaffolds underwent characterization experiments using different techniques, including FTIR, XRD, radiography, and SEM analyses. Rabbit adipose-derived mesenchymal stem cells (RADMSCs) were isolated from adipose tissues and characterized by flow cytometry. A direct contact test was performed for the cytotoxicity test using L929 cells. Cell adhesion was validated using SEM imaging, viability was assessed using DAPI and Annexin V/PI staining, and osteogenic differentiation was investigated using the ALP assay. The cytotoxicity assay using L929 cells verified that the OE/HA powder scaffold is safe and further functional experiments could be carried out. Behavioral assays, radiography, surface markers, surface topography analyses, and viability tests yielded promising results confirming cellular acceptance of the scaffold. In the present study, we provided evidence that the prepared OE/HA scaffold is not only cytocompatible but also can enhance mesenchymal stem cell adhesion, growth, and osteogenic differentiation in vitro.