2023, Vol. 27 ›› Issue (10): 1507-1513

Effect of a disintegrin and metalloproteases 10 on proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells induced by steroid-induced osteonecrosis of femoral head

Fan Jun1, Wu Chenhuan2, Cheng Zhonghua2   

Abstract: BACKGROUND: Many studies have found that the pathological progression of steroid-induced osteonecrosis of the femoral head is related to abnormal proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells, in which a disintegrin and metalloproteases 10 (ADAM10) is highly involved, but the specific mechanism remains unclear.  
OBJECTIVE: To investigate the effects of ADAM10 on proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells derived from steroid-induced osteonecrosis of the femoral head patients and its possible mechanism.
METHODS: Bone marrow was harvested from proximal femur in 23 patients with steroid-induced osteonecrosis of the femoral head and 17 patients with femoral neck fracture under sterile conditions. Bone marrow mesenchymal stem cells from steroid-induced osteonecrosis of the femoral head (S-BMSCs) and femoral neck fracture (F-BMSCs) were isolated by density gradient centrifugation, and cultured to the third generation in vitro. The proliferation activity of cells in the two groups was detected by CCK-8 assay. Alizarin red staining and alkaline phosphatase staining were used to observe the osteogenic differentiation ability of cells in the two groups. qRT-PCR and western blot assay were used to detect the levels of ADAM10 mRNA and protein in cells of the two groups. S-BMSCs were infected with ADAM10 overexpression lentivirus vector, and then treated with Notch signaling pathway inhibitor DAPT. The proliferation activity of these cells was detected by CCK-8 assay. Alizarin red staining and alkaline phosphatase staining were used to observe the osteogenic differentiation of cells. qRT-PCR was used to detect the expression levels of osteogenic markers alkaline phosphatase, Runt-related transcription factor 2, and osteocalcin mRNA. The expression levels of Notch signaling pathway related proteins Notch1, NICD, and Hes1 were detected by western blot assay.  
RESULTS AND CONCLUSION: (1) The proliferation activity, osteogenic differentiation ability and the expression level of ADAM10 in S-BMSCs were significantly lower than those in F-BMSCs (P < 0.01). (2) After ADAM10 overexpression, the proliferation activity, osteogenic differentiation ability, the expression levels of alkaline phosphatase, Runt-related transcription factor 2, and osteocalcin mRNA and Notch1, NICD and Hes1 protein of S-BMSCs were significantly increased (P < 0.05). However, combined intervention with DAPT significantly inhibited the promoting effects of ADAM10 overexpression on proliferation activity and osteogenic differentiation ability of S-BMSCs, and inhibited the activation of Notch signaling pathway. (3) The results show that ADAM10 can be underexpressed in S-BMSCs, and its overexpression can enhance the proliferation and osteogenic differentiation ability of S-BMSCs, and its mechanism may be related to the activation of Notch signaling pathway.