Fig. 3.

The osteogenesis of BMSCs cultured in macrophage-conditioned medium. The medium was prepared by incubating RAW cells with PLGA-based microspheres at 37 °C for 4 days and then aspirating the culture's supernatants. (a) The morphology of BMSCs seeded on PLGA-based microspheres for 24 h observed by SEM to evaluate surface morphology. The BMSCs attached well and even flattened on PLGA/MgO-alendronate microsphere. (b)–(f) The cell viability, proliferation, differentiation, and mineralization of BMSCs incubated with macrophage-conditioned medium at various time points. The cell viability and proliferation were analyzed by CCK-8 and BrdU incorporation assay, while the ALP activity and alizarin red tests were employed to determine osteogenic differentiation and mineralization of BMSCs. (g)–(k) The relative mRNA osteogenic levels of Col 1, ALP, OPN, OCN, BMP-2, and Runx2 of BMSCs normalized to GAPDH detected by the RT-PCR assay on days 7 and 14. The control group was defined as BMSCs cultured in DMEM without the addition of PLGA-based microspheres. * referred to a significant difference (p < 0.05); ** (p < 0.01); *** (p < 0.001). (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)