摘要
目的 了解肾小管上皮-间充质细胞转化（EMT）过程中肾小管上皮细胞迁移能力的动态变化及对细胞周期的影响.方法 通过体外培养大鼠肾小管上皮细胞（NRK-52E）,转化生长因子（TGF）-β1（5μg/L）诱导肾小管上皮细胞向间充质细胞转化,利用Transwell小室法以及采用流式细胞术分别观察EMT过程中细胞迁移能力的动态变化规律和细胞周期变化.结果 1.TGF-β1成功诱导EMT：TGF-β1（5μg/L）干预后,NRK-52E细胞发生形态学改变,表现为细胞排列松散,部分细胞胞体伸长,呈梭形改变；Western blot发现干预后的NRK-52E细胞上皮标志物E-cadherin表达明显下调（P＜0.05）,间质细胞标志物α-平滑肌肌动蛋白（α-SMA）表达增强（P＜0.05）,且两者均呈时间依赖性变化.2.TGF-β1体外诱导大鼠肾小管上皮细胞EMT过程中,TGF-β1干预后12 h细胞迁移能力明显增强,且随着时间的推移细胞迁移能力增强更为显著（与空白对照组比较,P＜0.01）.3.EMT过程中,细胞周期S期的比例随时间变化并未发生明显改变（与空白对照组比较,P＞0.05）.结论 TGF-β1体外诱导NRK-52E发生EMT过程中其细胞迁移能力不断增强,而细胞周期并未发生明显改变.
Objective To evaluate the dynamic changes of cell mobility of renal tubular epithelial cells in the course of epithelial-mesenchymal transition（EMT） and their effect on cell cycle.Methods NRK-52E cells were cultured in vitro and treated with 5 μg/L transforming growth factor（TGF）-β1 to induce EMT.The cell mobility was assessed by using Transwell chamber assay and flow cytometry （FCW） after being treated with TGF-β1 for 4 h,8 h,12 h,24 h and 48 h.The proliferative cell cycle of NRK-52E cells were evaluated by using the FCW.Results 1.EMT was successfully induced by TGF-β1.After being treated by TGF-β1 （5 μg/L）,the morphological changes of NRK-52E cells were found with loose cell arrangement and elongated fusiform change in cells body.Meanwhile,after getting treated by TGF-β1,the expressions of E-cadherin protein（epithelial marker） of NRK-52E cells were significantly decreased with time-dependent （P ＜ 0.05）,while the expressions of α-smooth musle actin （α-SMA） （mesenchymal cell marker）were significantly increased with time-dependent （P ＜ 0.05）.2.The Transwell chamber assay showed that compared with the control group,the cell mobility in the group treated with TGF-β1 was significantly enhanced from 12 h after getting treated with TGF-β1 （P ＜ 0.01）.3.The proliferative cell cycle of NRK-52E cells showed no significant difference after being treated with TGF-β1 （P ＞ 0.05）.Conclusions The migration ability of the NRK-52E cells are increased incessantly in the course of EMT,which is induced by TGF-β1 without the influence of cell proliferation in vitro.
出处
《中华实用儿科临床杂志》
CAS
CSCD
北大核心
2013年第17期1302-1305,共4页
Chinese Journal of Applied Clinical Pediatrics
基金
国家自然科学基金（30871177）
上海市科委科研项目（114119a1700）
