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作 者:丁召[1] 陈知水[1] 陈曦林[1] 郭晖[1] 宫念樵[1]
机构地区:[1]华中科技大学同济医学院附属同济医院器官移植研究所,武汉430030
出 处:《中华病理学杂志》2009年第7期462-465,共4页Chinese Journal of Pathology
基 金:国家自然科学基金(30300324)
摘 要:目的探讨结缔组织生长因子(CTGF)对肾小管上皮细胞向间充质细胞转化的影响。方法体外培养的人肾小管上皮细胞(HK-2),按是否用CTGF处理将其分为实验组和对照组。直接免疫荧光法、间接免疫化学检测培养72h后肾小管上皮细胞E—cadherin、波形蛋白、α-平滑肌肌动蛋白(α—SMA)、细胞外信号调节激酶2(ERK2)的表达;Western blot方法在72h检测E—cadherin、波形蛋白和ERK2的表达;Boyden小室在1、3、5d检测细胞迁移能力的变化。结果(1)实验组上皮细胞表型标志蛋白E-cadherin随着时间的延长而表达递减,波形蛋白表达渐增,ERK2表达上调;(2)在第1天,实验组与对照组细胞迁移能力差异无统计学意义,第3天实验组迁移至滤膜下面细胞多于对照组,第5天实验组细胞迁移数明显高于对照组[(45.0±1.1):(14.0±1.2),P〈0.05]。结论在体外,人肾小管上皮细胞由CTGF刺激表现出向间充质细胞转化的特性,并且ERK2信号转导通路可能参与了对这一过程的调控效应。Objective To investigate the role of connective tissue growth factor (CTGF) in epithelial mesenchymal transition of HK-2 cells in vitro. Methods HK-2 ceils were randomly divided into two groups: ( 1 ) control group including cells cultured in DMEM medium supplemented with 10% fetal bovine serum only; and (2) experimental group including ceils cultured in DMEM medium supplemented with 10% fetal bovine serum and recombinant CTGF at a final concentration of 5 μg/L. The cells were collected at 72 h time points. Direct immunofluorescence staining and immunohistochemistry were used to evaluate the E-cadherin, Vimentin, α-SMA and ERK2 in cells. Western-blotting was used to detect the E-cadherin, Vimentin and ERK2 protein expression. Boyden Chamber was used to detect the migration of tubular endothelium at 1 d, 3 d and 5 d. Results There were less E-cadherin but more Vimentin expressed in cells of the experimental group. The presence of α-SMA was detected at 48 h with peak at 72 h in the cells of the experimental group. On the first day, the cellular migration in the two groups showed no difference. However, after 3 days, the transformed cells migrated surpassed the control group with peak at the5th day [(45.0±1.1): (14.0±1.2), P〈0.05)]. Conclusion Connective tissue growth factor induces mesenchymal transformation of HK-2 cells, in which the ERK2 signaling pathway may play an important role.
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