淫羊藿苷对大鼠血管平滑肌细胞钙化的抑制作用

Inhibition of Icariin on Calcification in Rat Vascular Smooth Muscle Cells

目的:研究淫羊藿苷(Icariin)对大鼠血管平滑肌细胞(VSMCs)钙化的抑制作用。方法:组织贴壁法培养大鼠胸主动脉VSMCs,取5-8代细胞分为对照组、钙化模型组和淫羊藿苷干预A、B、C组。对照组用DMEM培养基培养;钙化模型组用含10mmol/Lβ-甘油磷酸盐(β-GP)的DMEM培养基培养;干预A、B、C组均以钙化模型组为基础分别与10-6 mol/L、10-5 mol/L和10-4 mol/L Icariin的DMEM培养基共培养,茜素红-S染色法鉴定各组细胞钙化情况,硝基苯酚法测定各组细胞碱性磷酸酶(ALP)活性,RT-PCR检测各组细胞内骨保护素(OPG)和核因子κB(NF-κB)受体活化因子配体(RANKL)的mRNA水平,用Western blotting检测各组细胞OPG和RANKL的蛋白表达。结果:与对照组比较,钙化模型组VSMCs发生细胞钙化,并形成红色结节,ALP活性显著升高(P<0.01),OPG、RANKL mRNA和蛋白表达均增加(P<0.01);与钙化模型组比较,干预A、B、C组细胞钙化减少,ALP活性减低(P<0.01),OPG、RANKL mRNA和蛋白表达均下调(P<0.01),尤以干预B组效果最显著。结论:淫羊藿苷可能通过降低ALP活性及OPG、RANKL表达来抑制动脉钙化。

Objective:To investigate the effect of Icariin on calcification in rat vascular smooth muscle cells(VSMCs).Method:Primary VSMCs were obtained from rat thoracic aorta by plant method.The passage 5-8cells were divided into control group,calcification group and intervention group A,B and C,the control group was cultured by DMEM;the calcification group was treated with DMEM contained 10mmol/Lβ-glyeerophosphat(β-GP);the intervention group was cultured with 10-6 mol/L,10-5 mol/L and 10-4 mol/L Icariin respectively on the basis of calcification group.All groups were identified by Alizarin Red S stain,Alkaline phosphatase(ALP)activity was measured by p-Nitrophenyl phosphate,the mRNA expression of OPG and RANKL were determined by RT-PCR,Western blotting was used to identify the protein of OPG and RANKL.Results:Red nodules,the activity of ALP,the mRNA expression of OPG and RANKL,the protein of OPG and RANKL were greatly increased in the calcification group as compared with control group(P<0.01);compared with calcification group,the calcification in intervention groups were attenuated and ALP activity,mRNA and protein of OPG and RANKL were decreased significantly(P<0.01),especially in the intervention group B.Conclusion:Icariin may inhibit calcification of rat VSMCs by reducing the activity of ALP and the expression of OPG and RANKL.