Fig. 3.

Osteogenesis in vitro. (A) The proliferation of rBMSCs tested by CCK8. rBMSCs showed good viability and maintained the potential to continue to proliferate. (B) ALP activity determined by ALP kit. It indicated that doped BSNs promoted the expression of ALP, especially for BSNs doped with copper. (C–E) The expression of osteogenesis-related genes including BMP2 (C), COL1A1 (D) and RUNX2 (E) assessed by RT-PCR. All the osteogenesis-related genes were upregulated under the effects of BSNs, and the effect of BSNs-Cu-Mn was particularly significant. (F) ALP staining and ARS staining on day 21; scale bars: 100 μm. It showed BSNs doped with copper significantly promoted the osteogenesis of rBMSCs in vitro. (G) Quantified positive area of ALP and ARS staining. (n = 3, P < 0.05, one-way ANOVA was used for statistical analysis.)