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目的:细胞水平观察不同浓度的辛伐他汀对骨髓间充质干细胞(bone marrow-drived mesenchymal stem cells,BMSCs)的增殖及旁分泌功能的影响并探讨其可能的机制。方法:采用全骨髓贴壁培养法培养SD大鼠的骨髓间充质干细胞,取3代对数生长期细胞,无血清培养12h,不同剂量的辛伐他汀(0、0.001、0.010、0.100、1.000μmol/L)与MSCs共培养24h,采用CCK-8试剂盒检测各浓度组BMSCs增殖情况,通过半定量RT-PCR检测不同浓度的辛伐他汀诱导BMSCs中血管内皮生长因子(vascular endothelial growth factor,VEGF)和肝细胞生长因子(hepatocyte growth factor,HGF)的mRNA表达。采用Western blot检测PAkt、AKt的表达。结果:辛伐他汀在一定浓度范围内(0.001～0.100μmol/L)显著提高了BMSCs增殖和VEGF和HGF的mRNA表达水平(与对照组比较,P<0.01)。辛伐他汀浓度在0.010μmol/L时对BMSCs的增殖和分泌功能的影响最为显著,随着药物浓度的加大,BMSCs的上述功能呈下降趋势。与对照组比较,1.000μmol/L辛伐他汀对BMSCs无显著的促增殖作用,但却明显上调了VEGF和HGFmRNA的表达。0.010μmol/L辛伐他汀组pAKt表达及pAKt/AKt比值显著高于对照组(P<0.01)。结论:辛伐他汀在一定浓度范围内对骨髓间充质干细胞的增殖及分泌功能有促进作用,其机制可能与Akt信号通路有关。
OBJECTIVE: To observe the effects of different concentrations of simvastatin on the proliferation and paracrine function of bone marrow-derived mesenchymal stem cells (BMSCs) at the cellular level and to explore its possible mechanism. METHODS: Bone marrow mesenchymal stem cells of SD rats were cultured with whole bone marrow adherent culture. The cells in 3 logarithmic growth phase were cultured in serum-free medium for 12 h. Different doses of simvastatin (0, 0.001, 0.010, 0.100, 1.000 μmol / L) were co-cultured with MSCs for 24 h. The proliferation of BMSCs was detected by CCK-8 kit. The levels of vascular endothelial growth factor (VEGF) in BMSCs induced by different concentrations of simvastatin were detected by semi-quantitative RT- VEGF) and hepatocyte growth factor (HGF) mRNA expression. Western blot was used to detect the expression of PAkt and AKt. Results: Simvastatin significantly increased the proliferation of BMSCs and the mRNA expression of VEGF and HGF in a certain concentration range (0.001 ~ 0.100μmol / L) (P <0.01 compared with the control group). The effect of simvastatin on the proliferation and secretion of BMSCs was the most significant when the concentration of simvastatin was 0.010μmol / L. With the increase of drug concentration, the above function of BMSCs showed a decreasing trend. Compared with the control group, 1.000μmol / L simvastatin did not significantly promote the proliferation of BMSCs, but significantly upregulated the expression of VEGF and HGF mRNA. The pAKt expression and the ratio of pAKt / AKt in 0.010μmol / L simvastatin group were significantly higher than those in control group (P <0.01). Conclusion: Simvastatin can promote the proliferation and secretion of bone marrow mesenchymal stem cells in a certain concentration range, which may be related to the Akt signaling pathway.