不同浓度钙离子对人腹膜间皮细胞E－钙黏蛋白、成纤维细胞特异蛋白、波形蛋白表达的影响

Effect of calcium concentration on human peritoneal mesothelial cell expression of E-cadherin,FSP1 and Vimentin

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、2.25mmol/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。

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 mesothelia] 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 mmal/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.