摘要
目的:研究垂盆草(SSB)提取物对马兜铃酸(AA)诱导的肾小管上皮细胞表型转化和胶原累积的作用,并初步探讨可能的分子机制。方法:将体外培养的大鼠肾小管上皮细胞(NRK-52E)分为:(1)溶剂对照组:未加入SSB和AA;(2)AA损伤组:只加AA,浓度为1~100 mg/L;(3)SSB提取物干预组:在加入10 mg/L AA基础上,同时加入SSB提取物(10~2 000 mg/L)。细胞培养24 h后,酶联免疫吸附试验检测转化生长因子β1(TGF-β1)含量;细胞免疫荧光染色检测肌成纤维细胞标志物α-平滑肌肌动蛋白(α-SMA)、上皮细胞标志物E-cadherin和基质成分III型胶原的表达;实时荧光定量PCR检测α-SMA、E-cadherin、骨形成蛋白7(BMP-7)和I型胶原mRNA的表达。结果:AA可诱导NRK-52E细胞呈现纤维化样改变;1 mg/L和10 mg/L AA不仅可增加基质成分I型和III型胶原的表达量,同时也可促进α-SMA表达,抑制E-cadherin的表达。用SSB提取物干预后,AA所致的纤维化改变明显减轻;SSB提取物下调了α-SMA、I型和III型胶原的表达,并且促进E-cadherin和BMP-7的表达。此外,SSB提取物也抑制TGF-β1的分泌,并呈浓度依赖性。结论:应用SSB提取物干预可明显降低AA所致的肾纤维化效应,可能的机制为SSB提取物降低TGF-β1的表达,抑制小管上皮细胞的表型转化,进而抑制胶原的累积。
AIM： To investigate the effect of Sedum sarmentosum Bunge （SSB） extract on epithelial-mesenchy- mal transition （EMT） and collagen accumulation induced by aristolochic acid （AA） in renal tubular epithelial cells. METHODS： Rat renal tubular epithelial NRK-52E cells were randomly divided into 3 groups, including control group （only treated with solvent）, AA group （treated with AA at concentrations ranging from 1 to 100 mg/L） and SSB group （treated with AA at a concentration of 10 mg/L plus SSB extract at concentrations ranging from 10 to 2 000 mg/L）. After cultured for 24 h, the morphology of the NRK-52E cells was observed under inverted phase-contrast microscope. The level of transforming growth factor 13~ （TGF-~I） in the culture supernatant was measured by ELISA. Immunofluorescent analysis was performed to detect the expression of epithelial marker c^-smooth muscle actin （ ct-SMA）, mesenchymal marker E-cadherin, and extracellu- lar cell matrix component type III collagen. The mRNA expression of E-cadherin, c^-SMA, bone morphogenetic protein 7 （BMP-7） and type I collagen was also quantified by real-time PCR. RESULTS： Fibrosis-like reaction observed under mi- croscope was obviously increased in AA-treated NRK-52E cells, and aggravated as the increase in the concentration of AA. AA at concentrations of 1 and 10 mg/L increased the expression of α-SMA, type I and type III collagens, and decreased the expression of E-cadherin. With SSB extract treatment, fibrosis in NRK-52E cells was alleviated, accompanied with the decreasing expression of α-SMA, type I and type III collagen, and the enhancing expression of E-cadherin and BMP-7. Mo-reover, SSB extract down-regulated TGF-β level in a concentration-dependent manner. CONCLUSION： AA-induced fi- brosis-like reaction in renal tubular epithelial cells is reduced by the treatment with SSB extract. The possible mechanism is that SSB extract decreases TGF-β1 level, and inhibits renal EMT and collagen accumulation induced by AA.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2013年第12期2172-2178,共7页
Chinese Journal of Pathophysiology
基金
浙江省自然科学基金资助项目(No.LQ12H050001
No.LY12H050004)
温州市科技计划(No.Y20110028
No.Y20110072)
关键词
垂盆草
马兜铃酸
转化生长因子β1
上皮-间充质转化
胶原
Sedum sarmentosum Bunge
Aristolochic acid
Transforming growth factorβ1
Epithelial-mesen-chymal transition
Collagen
