硫酸吲哚酚对大鼠主动脉平滑肌细胞增生影响的机制研究

Effect of indoxyl sulfate on proliferation of rat vascular smooth muscle cells

目的：探讨硫酸吲哚酚（indoxyl sulfate,IS）对大鼠血管平滑肌细胞（vascular smooth muscle cell,VSMC）增生的影响以及这种变化和氧化应激的关系。方法实验均分为8组：正常组、IS 100μM组、IS 300μM组、IS 500μM组、辛伐他汀10μM＋正常组、辛伐他汀10μM＋IS 100μM组、辛伐他汀10μM＋IS 300μM组、辛伐他汀10μM＋IS 500μM组。采用WST-1法检测VSMC增殖情况；硫代巴比妥酸法检测培养基中丙二醛（malondialdehyde,MDA）含量；ELISA方法测定晚期氧化蛋白产物（advanced oxidation protein products,AOPP）。结果与正常组比较,IS 300μM组和 IS 500μM 组上清液的 WST-1（OD 值）[（1.55±0.27）比（1.18±0.25）与（1.73±0.30）比（1.18±0.25）,P〈0.01]含量、MDA含量[（2.60±0.47）μg/ml比（1.59±0.21）μg/ml与（2.82±0.54）μg/ml比（1.59±0.21）μg/ml,P〈0.01]和 AOPP 含量[（67.94±8.58）μmol/L 比（54.97±8.46）μmol/L与（72.09±9.49）μmol/L比（54.97±8.46）μmol/L,P〈0.01]均明显增高。与同浓度 IS 组相比,辛伐他汀10μM＋IS 300μM 组和辛伐他汀10μM＋IS 500μM 组上清液 WST-1（OD 值）[（1.22±0.24）比（1.55±0.27）与（1.32±0.30）比（1.73±0.30）,P〈0.05]、MDA[（1.64±0.38）μg/mL 比（2.60±0.47）μg/ml 与（1.70±0.40）μg/ml 比（2.82±0.54）μg/ml,P〈0.01]含量和 AOPP 含量[（55.56±7.41）μmol/L比（67.94±8.58）μmol/L 与（55.54±6.80）μmol/L 比（72.09±9.49）μmol/L,P〈0.05]均明显降低。大鼠 VSMC 上清液的 WST-1含量与 MDA 含量呈正相关（r=0.621,P〈0.01）,大鼠VSMC上清液的WST-1含量与AOPP含量呈正相关（r=0.581,P〈0.01）。结论 IS可浓度依赖性地促进大鼠 VSMC增生,其作用可能与 IS增加 VSMC的氧化应激有关；辛伐他汀可抑制此作用。

Objective To investigate the effect of indoxyl sulfate （IS）on proliferation of rat vascular smooth muscle cells （VSMCs）and the relationship between oxidative stress and this effect. Methods Cultured rat VSMCs were divided into normal group,100μM IS treatment group,300μM IS treatment group,500μM IS treatment group,simvastatin10μM＋normal saline group,simvastatin 10μM＋100μM IS treatment group,simvastatin 10μM＋300μM IS treatment group and simvastatin 10μM＋500μM IS treatment group.Calcium deposition in VSMCs was measured by BCA.The cell proliferation in VSMCs was detected by WST-1 method.Themalondialdehyde （MDA）content was de-termined by thiobarbituric acid method.Advanced oxidation protein products （AOPP）content was ex-amined by ELISA.Results As compared with normal group,the WST-1 contents in the supernatant of 300μM IS treatment group and 500μM IS treatment group [A values of （1.55±0.27）and （1.73 ±0.30）vs.（1.18±0.25）（P〈0.01）],MDA contents [（2.60±0.47）and （2.82±0.54）μg/mL vs.（1.59±0.21）μg/mL （P〈0.01）],and AOPP contents [（67.94±8.58）and （72.09±9.49）μmol/L vs.（54.97±8.46）μmol/L （P〈0.01）]were significantly increased.As compared with the same con-centration IS treatment group,the WST-1 contents in the supernatant of simvastatin 10μM＋300μM IS treatment group and simvastatin 10μM＋500μM IS treatment group [A values of （1.22±0.24） vs.（1.55 ±0.27）（P〈0.05）and （1.32 ±0.30）vs.（1.73 ±0.30）（P〈0.05）],MDA contents [（1.64±0.38）vs.（2.60±0.47）μg/mL （P〈0.01）and （1.70±0.40）vs.（2.82±0.54）μg/mL （P〈0.01）],and AOPP contents [（55.56±7.41）vs.（67.94±8.58）μmol/L （P〈0.05）and （55.54 ±6.80）vs.（72.09±9.49）μmol/L （P〈0.05）]were significantly decreased.There was significantly positive correlation between the WST-1 contents and MDA contents （r=0.621,P〈0.01）,and be-tween the WST-1 contents and AOPP contents （r=0.581,P〈0.01）.Conclusions IS may promote proliferation of rat VSMCs in a concentration-dependent manner,which may be possibly related with the increased oxidative stress induced by IS.Simvastatin may inhibit this effect.