梗阻性黄疸大鼠肠黏膜上皮紧密连接蛋白和MLCK的研究

目的:探讨梗阻性黄疸肠黏膜屏障破坏的机制。方法:建立梗阻性黄疸大鼠的动物模型,分别于胆管结扎10 d和20 d后，采用免疫组化、Western blot方法检测末端回肠黏膜的紧密连接蛋白成员ZO-1和Occludin与肌球蛋白轻链激酶（MLCK）的分布和表达。结果:正常回肠黏膜层ZO-1和Occludin的分布相似，主要位于上皮细胞的边缘，细胞膜顶端，沿绒毛下方均匀连续分布；MLCK主要分布在细胞浆内。梗阻性黄疸时ZO-1和Occludin分布不均, 染色变淡，线条模糊，边缘粗糙有毛刺状突起；MLCK分布散乱，染色稀疏。与对照组相比20 d组和10 d组的ZO-1，Occludin，MLCK数量明显减少，强阳性表达率分别从70.0 %，80.0 %，70.0 %降至10 d组的28.6 %，28.6 %，28.6 % (均P＜0.05)和20 d组的ZO-1从10 d 组的28.6 %降至14.3 %, 35.7 %, 21.4 % (均P＜0.05)。20 d 组的ZO-1下降较10 d组更为明显（P<0.05）, 而Occludin和MLCK变化不明显。Western blot检测的结果与之相似。结论:梗阻性黄疸时大鼠小肠黏膜上皮ZO-1，Occludin，MLCK分布紊乱，表达下降，小肠黏膜上皮屏障的完整性破坏。

The study on tight junction proteins and myosin light chain kinase in obstructive jaundice in rats

Abstract:Objective:To elucidate the mechanisms of disruption of intestinal mucosa barrier in obstructive jaundice.Methods :The obstructive jaundice model of rats was set up.At 10 d and 20 d after operation,immunohistochemistry and Western-blot techniques were used to examine the distribution and expression of tight junction proteins (ZO-1,Occludin) and myosin light chain kinase（MLCK）in intestinal mucosa.Results:In normal control groups, the staining of ZO-1 and occludin was predominantly localized to the margins of the epithelial cells and the apex of the cell membrane, and displayed a continuous and uniform distribution along the under surface of the villae. MLCK was distributed mainly in cytoplasm.In obstructive jaundice groups, ZO-1 and occludin staining appeared discontinuous and vague,with rough edges and spiculate processes.The staining of MLCK was also discontinuous and scanty.The strong positive express ratio of ZO-1,Occludin and MLCK were obviously lower in two experiment group than those in the control group all P＜0.05). ZO-1 was significantly decreased in 20 d group than that in 10 d group（14.3 % vs28.6 %，P＜0.05), but no significant difference in occludin and MLCK staining between 10 d group and 20 d group.Similar outcomes were obtained by quantitative analysis of Western blot images.Conclusions:Obstructive jaundice rats have distribution of ZO-1,occludin and MLCK in ileum mucous membrane epithelium, and is associated with breakdown of the integrity of the intestinal barrier.