Fig. 4

Impaired osteogenesis in MD-MSCs. a Representative images of ALP activity in WT- and MD-MSCs during OB differentiation. ALP activity was observed as red granules. D, days after osteogenesis induction. b Representative images of alizarin red S staining in WT- and MD-MSCs during OB differentiation. Alizarin red S staining presented as red granules. c Representative images of Von Kossa staining in WT- and MD-MSCs during OB differentiation. Von Kossa staining was observed as black dots. Scale bars = 500 μm. d Relative expression of osteogenic genes RUNX2, OPN, and OCN in MD-MSCs during osteogenesis. The data are presented as the mean ± SE (n = 3). e, f Effects of ATP7A knock-down on osteogenesis in WT-MSCs. e Representative images of ALP activity, alizarin red S staining, and Von Kossa staining after transfection of siRNAs targeting ATP7A gene. Scramble siRNA (si-SCR) was also transfected in WT-MSCs as a control. f Relative expression of RUNX2, OPN, and OCN after ATP7A knockdown. The data are represented as the mean ± SE (n = 3). *p < 0.05, **p < 0.01. ALP alkaline phosphatase, D day of differentiation, MD1/2 Menkes disease patient 1/2, WT wild type