维生素K_2对大鼠血管平滑肌细胞钙化及Toll样受体2和4表达的影响

Effect of vitamin K_2 on calcification of vascular smooth muscle cells and expression of Toll-like receptors 2 and 4 in rats

目的探讨维生素K_2对血管平滑肌细胞(VSMC)钙化及Toll样受体(TLR)2和TLR4表达的影响。方法体外培养A7r5VSMC分为正常组(基础培养液),钙化组(加入10mmol/Lβ磷酸甘油),干预组(钙化基础上加入维生素K_210-6 mmol/L),各组均干预14d。Von Kossa染色观察VSMC钙化发生情况;检测细胞钙含量和碱性磷酸酶(ALP)活性;实时定量PCR及Western blot检测TLR2和TLR4mRNA和蛋白表达水平。结果钙化组细胞钙含量及ALP活性较正常组分别增加11.5倍和9.3倍(P<0.01);干预组细胞钙含量及ALP活性较钙化组分别减少1.5倍和2.3倍(P<0.01)。与正常组比较,钙化组细胞TLR2、TLR4蛋白及mRNA表达升高;与钙化组比较,干预组细胞TLR2、TLR4蛋白及mRNA表达降低(P<0.05,P<0.01)。结论维生素K_2抑制细胞钙化的作用可能与TLR2和TLR4表达的下调有关。

Objective To study the effect of vitamin K_2 on calcification of vascular smooth muscle cells（VSMC）and expression of Toll-like receptors（TLR）2and 4in rats.Methods VSMC cultured in vitro were divided into normal contrl group,calcified group and vitamin K_2 treatment group.The calcification of VSMC was detected with Von Kossa staining and identified by measuring the serum calcium level and alkaline phosphatase（ALP）activity.Expressions of TLR 2and 4mRNA and protein were detected by PCR and Western blot respectively.Results The serum calcium level and ALP activity were 11.5-fold and 9.3-fold higher in calcified group than in normal control group（P0.01）,and 1.5-fold and 2.3-fold lower in vitamin K_2 treatment group than in calcified group（P0.01）.The expression levels of TLR 2and 4protein and mRNA were significantly higher in calcified group than in normal control group and significantly lower in vitamin K_2 treatment group than in calcified group（P0.05,P0.01）.Conclusion Vitamin K_2 inhibits the calcification of VSMC by downregulating the expression of TLR 2and 4.