c-Src激酶对脂多糖致炎过程中occludin蛋白表达的影响

目的 探讨在脂多糖(lipopolysaccharide,LPS)处理小鼠肺泡上皮细胞(mouse lung epithelial-12,MLE-12)过程中,c-Src激酶对occhdin蛋白表达的影响. 方法 将培养的MLE-12细胞按随机数字表法分3组(每组3孔细胞):对照组(C组)、LPS实验组(L组)、LPS+c-Src激酶抑制剂PP2组(L+P组).C组不做任何处理;L组用LPS(浓度为10 mg/L)处理,处理时间为1、3、6 h;L+P组用PP2(浓度为100 μmol/L)预处理60 min,然后LPS(浓度为10 mg/L)处理6h.Western blot法和免疫荧光染色法分别检测各组不同时间点occludin蛋白的表达. 结果 Western blot发现,与C组比较:L组中6h时点occludin蛋白表达下调明显、c-Src表达明显升高,差异均有统计学意义(P＜0.05);与L组6h比较,L+P组occludin蛋白表达上调(P＜0.05).免疫荧光染色显示,occludin蛋白主要表达于细胞膜上,LPS影响occludin蛋白在膜上分布,PP2可以减轻LPS的影响. 结论 LPS导致MLE-12细胞紧密连接蛋白occludin表达下调,c-Src激酶参与其调节.

Effect of c-Src kinase on the expression of occludin during the inflammation process mediated by lipopolysaccharide

Objective To investigate the effect of c-Src kinase on the expression of the tight junction protein occludin during lipopolysaccharides(LPS) treatment on mouse lung epithelial-12(MLE-12) cells.Methods The cultured MLE-12 cells were randomly divided into 3 groups:control group(group C),LPS group (group L),LPS+c-Src kinase inhibitor(PP'2) group(group L+P).In group L,MLE-12 cells were treated with LPS (10 mg/L) for 1,3,6 h,respectively.In group L+P,MLE-12 cells were treated with c-Src kinase inhibitor PP2(100 μmol/L) for 30 min then LPS (10 mg/L) for 6h.Non-treatment was given in group C.The expression of occludin was determined in MLE-12 cells using Western blot and immunofluorescence.Results In our study,we found that compared with group C,the expression of occludin of group L was significantly down-regulated at 6 h (P＜0.05) and c-Src was upregulated (P＜0.05).The expression of occludin was significantly up-regulated in group L+P (P＜0.05) at 6 h compared with group L.Immunofluorescence showed that occludin which distributed in cytomembrane could be affected by LPS,and PP2 could alleviate the effect of LPS.And the level of occludin was increased after treated with PP2 in group L+P.Conclusions LPS leads to decreasing of the expression of occludin in MLE-12 cells,with c-Src involved in the process.