Fig. 5

BMP4 signaling involved in regulation of hBMSC osteogenesis after DEPTOR knockdown. a Hierarchical clustering indicated BMP4 was differentially expressed (P < 0.05) in DEPTOR knockdown hBMSCs. b Relative mRNA expression of BMP4 measured by qRT-PCR in DEPTOR knockdown hBMSCs. GAPDH used for normalization. c Western blot analysis of BMP4 and its downstream Smad expression in DEPTOR knockdown hBMSCs. GAPDH utilized as internal control. d–f BMP antagonist Noggin (500 ng·mL^− 1) administered to DEPTOR knockdown and NC hBMSCs. d ALP staining on day 7 and ARS staining on day 14 after osteogenic induction. e, f ALP activity on day 7 and AZR quantification on day 14 after osteogenic induction. Data presented as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001 (n = 3 independent experiments). ALP alkaline phosphatase, ARS/AZR Alizarin red S, BMP4 bone morphogenetic protein 4, DEPTOR DEP domain containing mTOR interacting protein, GAPDH glyceraldehyde 3-phosphate dehydrogenase, shDEPTOR #1/shDEPTOR #2 lentivirus expressing shRNAs targeting DEPTOR, shNC scrambled control, sh short hairpin