氧化苦参碱对人脐静脉平滑肌细胞钙化的影响及其可能机制

Effect of oxymatrine on vascular calcification of humans umbilical vein smooth muscle cells and its underlying mechanism

目的:观察氧化苦参碱(oxymatrine,OMT)对人脐静脉平滑肌细胞(humans umbilical vein smooth muscle cells,HUSMCs)钙化的影响及其可能机制。方法:以β-甘油磷酸盐诱导HUSMCs钙化,实验分为对照组、钙化组、单纯OMT组、OMT干预高、中、低剂量组。采用Von Kossa染色鉴定细胞钙化,比色法检测细胞钙含量,磷酸苯二钠法测定ALP活性,放射免疫法测定骨钙素(OC)含量,ELISA检测细胞培养液中TGF-β1和细胞内psmad2/3,smad2/3的含量变化,Western blot法检测细胞内Cbfα1蛋白的表达。结果:钙化组与对照组相比平滑肌细胞内可见大量黑色颗粒聚集,钙含量,ALP活性,OC,TGF-β1,smad2/3磷酸化和Cbfα1蛋白的含量均明显增加;OMT干预后钙化指标均下降,TGF-β1,smad2/3磷酸化和Cbfα1蛋白表达量亦明显减少,且高剂量OMT组效应强于中、低剂量组。结论:OMT可有效抑制β-甘油磷酸盐诱导的HUSMCs钙化,且OMT降低TGF-β1,smad2/3磷酸化和Cbfα1蛋白表达可能是OMT抑制HUSMCs钙化的机制之一。

Objective： To observe the effect of oxymatrine（OMT） on calcification of humans umbilical vein smooth muscle cells and its underlying mechanism.Method： Human umbilical vein smooth muscle cells（HUSMCs） were calcified by β-giycerophosphosphate（β-GP） and then divided into 6 groups： the control group,the calcification group,the pure OMT group,and lower,middle and higher-dosage OMT groups.Cell calcification were observed by Von Kossa staining,calcium content in HUSMCs were determined by the colorimetric method,the alkaline phosphatase（ALP） activity in HUSMCs were determined by phenyl diphosphate-2-sodium,the osteocalcin（OC） level in HUSMCs were determined by radioimmunossay,the transforming growth factor-β1（TGF-β1） level in HUSMC culture medium and the content changes in psmad2/3 and smad2/3 were determined by the ELISA method,and the expression of Core binding factor α 1（Cbfα1） protein in HUSMCs were determined by western blot method.Result： Compared with the control group,the calcification group showed a great number of black granules among the smooth muscle cells and significant increase in the content of calcium and OC and the activity of ALP;OMT intervention can decrease the content of calcium,OC,TGF-β1,psmad2/3 and Cbfα1 and the activity of ALP.And high-dosage OMT group had better effect than middle and low-dosage groups.Conclusion： OMT can effectively inhibit β-GP-induced HUSMC calcification and its effect on reducing TGF-β1,psmad2/3 and Cbfα1 may be one of its mechanisms in inhibiting HVSMC calcification.