Objective:To investigate the possible mechanism of fibrosis in peritoneal membrane during peritoneal dialysis by determining whether epithelial-mesenchymal transition (EMT) is caused by calcium with diverse in human peritoneal mesothelial cells. Methodology:An human peritoneal mesothelial cell line (HMrSV5) was used in this vitro system. HMrSV5 was incubated with calcium (0. 25、0.75、1.25、1.75、2. 25 mmol/L) and control ( 0 mmol/L) for 48 hours. Then the cells were collected and the expression of E-cadherin, fibroblast specific protein ( FSP1 ), and vimentin was detected by Western blot and indrect immunofluorescence staining. Results:There was nearly no expression of E-cadherin, and low expression of FSP1 and vimentin in control. After 48 hours treatment with different concentration of calcium, the expression of E-cadherin was increased in a dose dependent manner at first, until reach the peak in 1.25 mmol/L, then decreased gradually. While the expression of vimentin and FSP1 both increased notably in 1.75 mmol/L and were up-regulated dose dependently. The indirect immunofluorescence staining revealed the similar result,fluorescence intensity of vimentin and FSP1 was significantly increased in high calcium compared with that in low calcium concentration. Conclusion:Diversity of calcium ion may induce the alterations of E-cadherin, FSP1 and vimentin in human peritoneal mesothelial cells, which suggests that calcium ion as an important factor regulate EMT of peritoneal mesothelial cells.%目的:观察钙浓度变化对人腹膜间皮细胞上皮细胞-间充质细胞转分化的影响,探讨腹膜透析中腹膜纤维化的可能机制.方法:以人腹膜间皮细胞永生化细胞株(HMrSV5)为研究对象,予不同浓度钙(0.25、0.75、1.25、1.75、2.25 mmol/L)处理细胞48h,以不含钙处理组为对照.采用蛋白免疫印迹和间接免疫荧光染色方法检测E-钙黏蛋白(E-cadherin)、成纤维细胞特异蛋白(fibroblast specific protein,FSP1)和波形蛋白(Vimentin)的表达.结果:不含钙组HMrSV5细胞基本不表达E-cadherin,极低水平表达Vimentin和FSP1;而予含不同浓度钙离子处理48h后,HMrSV5中E-cadherin先呈剂量依赖性上调,1.25mmol/L组至顶峰,再呈剂量依赖性下降;而Vimentin和FSP1均在1.75mmol/L组HMrSV5中明显升高(P<0.05)并呈剂量依赖性上调.间接免疫荧光染色结果也显示,高钙组Vimentin和FSP1荧光强度比低钙组显著增强. 结论:钙离子浓度的变化能诱导人腹膜间皮细胞E-cadherin、FSP1、Vimentin蛋白表达发生变化,提示钙离子可能是参与调控间皮细胞间充质转分化的一个重要因素.
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