论文部分内容阅读
目的探讨地塞米松(Dex)对马兜铃酸诱导的肾小管上皮-间充质转化(EMT)的影响及可能机制。方法体外培养的人肾小管上皮细胞(HKC)分为阴性对照组、马兜铃酸(AA,10μg/ml)作用组、Dex(100μmol/L)作用组、AA(10μg/ml)+无水乙醇(100μmol/L)组、AA(10μg/ml)+Dex(100、10、1、0.1、0.01μmol/L)共同作用组,培养细胞48h,应用激光共聚焦显微镜(CFMS)观察α-平滑肌肌动蛋白(α-SMA)、E-钙黏素(E-cadherin)的表达,采用Real-timePCR和WesternBlot方法检测α-SMA、E-cadherin、p-Smad3、转化生长因子-β1(TGF-β1)、Smad7mRNA和蛋白的表达。结果 AA作用48h后,CFMS观察发现HKC细胞E-cadherin表达减弱而α-SMA表达增强;地塞米松干预后,E-cadherin表达逐渐增强而α-SMA表达减弱,且呈剂量依赖性。Real-timePCR、WesternBlot结果均显示,与阴性对照组相比,AA组α-SMA、p-Smad3、TGF-β1mRNA和蛋白表达水平均显著增强,E-cadherin、Smad7mRNA和蛋白的表达均减弱(P<0.05)。同AA作用组比较,AA+Dex共同作用组随Dex剂量增加,α-SMA、p-Smad3、TGF-β1mRNA和蛋白的表达逐渐减弱,E-cadherin、Smad7mRNA和蛋白的表达逐渐增强(P<0.05),而无水乙醇却无此作用。结论 Dex可抑制AA诱导HKC细胞发生EMT,并同时下调EMT过程中TGF-β1和p-Smad3的表达、上调Smad7分子的表达,提示TGF-β1/Smad下调可能是Dex抑制EMT发生的机制之一。
Objective To investigate the effect of dexamethasone (Dex) on aristolochic acid induced tubular epithelial-mesenchymal transition (EMT) and its possible mechanism. Methods Human renal tubular epithelial cells (HKC) cultured in vitro were divided into four groups: negative control group, AA group (10μg / ml), Dex (100μmol / L) group, AA The cells were cultured for 48 hours in the presence of ethanol (100μmol / L), AA (10μg / ml) + Dex (100,10,1,0.1,0.01μmol / L) The expression of α-SMA and E-cadherin was detected by Real-time PCR and Western Blot. The expressions of α-SMA, E- β1), Smad7 mRNA and protein expression. Results After treated with AA for 48 hours, the expression of E-cadherin was decreased and the expression of α-SMA was increased in HKC cells after CFMS treatment. The expression of E-cadherin was gradually increased and the expression of α-SMA was decreased in a dose-dependent manner after dexamethasone treatment. The results of Real-time PCR and Western Blot showed that the mRNA and protein expressions of α-SMA, p-Smad3 and TGF-β1 in AA group were significantly increased compared with those in the negative control group, while the expressions of E-cadherin, Smad7 mRNA and protein were all decreased <0.05). Compared with AA group, the expression of α-SMA, p-Smad3, TGF-β1 mRNA and protein gradually decreased and the expression of E-cadherin, Smad7 mRNA and protein gradually increased with the increase of Dex dosage in AA + Dex group ), While anhydrous ethanol does not have this effect. Conclusions Dex can inhibit the EMT induced by AA in HKC cells and down-regulate the expression of TGF-β1 and p-Smad3 and up-regulate the expression of Smad7 in HKC cells, suggesting that down-regulation of TGF-β1 / Smad may be one of the mechanisms by which Dex inhibits EMT .