Altered tight junctions and fence function in NRK-52E cells induced by aristolochic acid
Altered tight junctions and fence function in NRK-52E cells induced by aristolochic acid
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DOI：
10.1177/0960327111407645
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发表时间：
2012-01-01
影响因子：
2.8
通讯作者：
Yu, Xueqing
中科院分区：
文献类型：
--
作者：
Liu, Mei;Yang, Xiao;Yu, Xueqing
马兜铃酸(AA)可在肾小管间质中积聚，引起肾脏特异性损伤。然而，AA导致肾病的机制在很大程度上仍不清楚。本研究探讨了AA-I对肾上皮细胞紧密连接(TJ)和栅栏功能的影响。将NRK-52E细胞暴露于不同浓度的AA-I作用4h或25mU AA-I作用不同时间。用四甲基偶氮唑盐比色法检测细胞存活率，流式细胞仪检测细胞凋亡率，Western印迹和免疫荧光法检测闭锁带蛋白-1(ZO-1)、E-钙粘蛋白和极性支架蛋白(Par3)的表达，跨皮细胞电阻(TEER)检测细胞膜通透性。结果发现，AA-I以浓度和时间依赖的方式降低ZO-1、E-钙粘素和PAR3的表达，并改变ZO-1和PAR3从细胞膜到细胞质的分布。在TJ蛋白表达降低的同时，TEER对AA-I处理的反应显著降低，且呈时间和浓度依赖关系。同时，AA-I处理后细胞中α-SMA的表达增加。相反，细胞存活率和凋亡率并没有随着AA-I测试剂量的增加而改变。我们的研究结果首次表明，AA-I作用于培养的肾小管上皮细胞后，TJ蛋白的表达迅速中断，提示TJ蛋白的异常表达可能参与了肾小管间质病变的发生。
Aristolochic acid (AA) can accumulate in the tubulointerstitium and cause kidney-specific injuries. However, the mechanism by which AA induces nephropathy remains largely unknown. This study explored the effect of AA-I on tight junctions (TJs), and the fence function in a renal epithelial cell (REC). NRK-52E cells were exposed to different concentrations of AA-I for 4 h or 25 mu M AA-I for different time. Cell viability was detected by MTT, cell apoptosis by flow cytometric analysis, the expression of zonula occludens-1 (ZO-1), E-cadherin and polarity scaffold (Par3) by western blot and immunofluorescence, cell membrane permeability by transepithelial electrical resistance (TEER). It was found that AA-I reduced the expression of ZO-1, E-cadherin, and Par3 in a concentration-and time-dependent fashion, and altered the distribution of ZO-1 and Par3 from cell membrane to cell plasma. In parallel to the reduced expression of TJ proteins, TEER exhibited a significant reduction in response to AA-I treatment in a time-and concentration-dependent manner. Meanwhile, alpha-SMA expression in cells was increased following AA-I treatment. In contrast, cell viability and apoptosis were unaltered with the doses of AA-I tested. Our findings show for the first time that AA-I treatment in cultured RECs induced a rapid disruption of TJ and the fence function preceding apoptosis, which indicated that aberrant expression of TJ proteins within RECs may be involved in initiating the renal tubulointerstitial disorders.