Fig. 4

Establishment of Apelin overexpression hBMSC cell line and effects of Apelin on osteogenic differentiation of hBMSCs. a–d Verification of Apelin overexpression in hBMSCs. Scale bars, 100 μm. e–g Relative expression of osteo-specific proteins (RUNX2 and COL1A1) was significantly upregulated on day 3 of osteogenesis. The mRNA expression levels were normalized to that of 18S ribosomal RNA. h, i Staining detection of ALP activity on day 3 of osteogenesis. Scale bars, 500 μm. j, k Alizarin Red S staining on day 10. Alizarin Red S-stained area was determined by measuring the absorbance at 560 nm. Scale bars, 500 μm. Data are expressed as mean ± SD. Reactions were performed in triplicate. *P < 0.05 and **P < 0.01 compared with the control group