The effect of nitroprusside-induced chondrocyte apoptosis on the osteogenic differentiation of bone marrow mesenchymal stem cells

Mou Zhefei; Chen Jiaru; Chen Mangmang; Lyu Yangxun; Ni Yueping; Ye Xiuyun

doi:10.3969/j.issn.1672-8157.2025.03.044

Abstract：Objective To observe the effect of sodium nitroprusside-induced chondrocyte apoptosis on the osteogenic differ-entiation ability of bone marrow mesenchymal stem cells(BMSC)and analyze the regulatory role of the MSR1/PI3K/Akt pathway in this process.Methods Primary mouse chondrocytes and mouse BMSCs were isolated and cultured.Sodium nitroprusside was used to construct a chondrocyte apoptosis model,while the supernatant of apoptotic chondrocytes,along with dexamethasone,vita-min C phosphate and β-glycerophosphate,was used to treat BMSCs to explore its effect on the osteogenic differentiation ability of BMSCs.The cell counting Kit-8 method was used to assess chondrocyte viability;Western blot,RT-qPCR and ELISA were employed to detect apoptosis-related indicators,including Bcl-2,Bax and Caspase-3 expression.Additionally,osteogenesis-related genes such as alkaline phosphatase,osteopontin and osteocalcin,and key pathway genes like MSR1,PI3K and Akt were measured in BMSCs,including their expression and phosphorylation levels.Alizarin red staining was used to observe the osteogenic differentiation of BM-SCs.Results Sodium nitroprusside at 1.0 mmol·L-1 could inhibit the viability of chondrocytes(1.0 mmol·L-1:(81.13±3.79)vs con-trol:(100.00±7.97)),while upregulating the expression of Bcl-2-associated X protein(Bax)and caspase-3 at both mRNA and protein levels,and inhibiting the expression of Bcl-2 at the prot-ein level(Bcl-2-associated X protein:1.0 mmol·L-1:(1.82±0.16)vs control:(1.00±0.09);caspase-3:1.0 mmol·L-1:(1.95±0.12)vs control:(1.00±0.06);Bcl-2:1.0 mmol·L-1:(0.64±0.04)vs control:(1.00±0.07))and mRNA level(Bcl-2-associated X protein:1.0 mmol·L-1:(3.19±0.20)vs control:(1.00±0.05);caspase-3:1.0 mmol·L-1:(4.00±0.20)vs control:(1.01±0.12);Bcl-2:1.0 mmol·L-1:(0.51±0.11)vs control:(1.00±0.10))(t=3.700,18.316,22.278,7.737,12.264,5.709,7.734,P<0.05).The supernatant from apoptotic chondrocytes could promote the osteogenic differentiation of BMSCs,upreg-ulating the expression of osteogenic-related genes such as alkaline phosphatase(osteogenic induction+supernatant:(3.02±0.14)vs control:(1.00±0.08)),osteopontin(osteogenic induction+supernatant:(2.92±0.28)vs control:(1.00±0.11))and osteocalcin(osteo-genic induction+supernatant:(3.32±0.33)vs control:(1.00±0.10),as well as increasing the positive rate of Alizarin red staining(t=21.698,11.054,11.654,P<0.05).Mechanistic studies suggested that the promotion of osteogenic differentiation in BMSCs by the supernatant from apoptotic chondrocytes was related to the activation of the MSR1/PI3K/Akt pathway.Conclusion Sodium nitroprusside-induced chondrocyte apoptosis promotes osteogenic differentiation in BMSCs and activates the MSR1/PI3K/Akt pathway during the process of osteogenic differentiation.