紧密连接ZO-1蛋白在缺血性脑卒中肠黏膜改变的意义

The role of protein ZO-1 in intestine barrier changes in ischemic stroke

目的研究缺血性脑卒中时是否存在肠黏膜屏障改变以及紧密连接蛋白Zonula Occluden-1(ZO-1)在其中的可能机制。方法杂犬20只,随机分为实验组和对照组,每组10只。实验组用双硅柱法制作缺血性脑卒中模型。实验前、实验后0.5、3、6、12、24h行血浆二胺氧化酶(DAO)活性和D-乳酸(D-lac)浓度检测;光镜下对肠黏膜病理形态进行观察;免疫组化法分析肠黏膜ZO-1蛋白的表达。结果和对照组比较,实验组24hDAO、12h和24hD-lac明显增高(24h DAO:0.5512±0.1057比0.3822±0.0789;12hD-lac:6.2249±1.2683比4.3342±0.7900;24hD-lac:8.4581±1.7416比4.4766±0.9486;P<0.01);实验组ZO-1蛋白平均光密度(OD)明显降低(0.1989±0.0118比0.2224±0.0150;P<0.01)。相关和回归分析显示,血浆24hDAO与24h、12h、6hD-lac呈正相关关系(γ分别为0.787、0.8117、0.862,P<0.01);ZO-1蛋白OD与24hDAO、24hD-Lac呈负相关(γ分别为-0.8385、-0.7088,P=0.002、0.022),光镜下观察,实验组存在小肠病理损伤,对照组小肠结构正常。结论血浆DAO活性和D-lac能有效反映缺血性脑卒中时肠屏障损伤。肠黏膜紧密连接蛋白ZO-1在缺血性脑卒中时表达降低,这是肠屏障功能障碍的重要分子机制之一。

Objective To observe intestine barrier changes in ischemic stroke and explore the possible role of protein ZO-1 in the pathogenesis.Methods Twentymongrel dogs were randomly divided into 2 groups with 10 in each.Double silicone cylinders measuring 1.1mm in diameter and 8mm in length were placed into their internal carotid arteries in all dogs of group A.And group B served as controls,with sham operation.Blood samples were taken at 6 time points：before operation （0h）,0.5h,3h,6h,12h and 24h after operation.The concentration of plasma D-lactate and the activity of diamine oxidase （DAO） in plasma were measured at these 6 time points.Light microscopic examination was performed for morphological measurement of intestinal epithelial cell.Immunohistochemistry was used to analyze the changes of protein ZO-1 localization at tight junction of intestine epithelial cells.Results The results of DAO at 24h and the results of D-lactate at 12h,24h increased significantly in all dogs of group A（24h DAO：0.5512±0.1057 to 0.3822±0.0789;12hD-lac：6.2249±1.2683 to 4.3342±0.7900;24hD-lac：8.4581±1.7416 to 4.4766±0.9486;P0.01）,the Zo-1 protein level in group A was significantly lower（0.1989±0.0118 to 0.2224±0.0150;P0.01）.There was positive correlation between the activity of DAO at 24h and the level of D-lac at 24h,12h,6h in plasma（γ：0.787,0.8117,0.862,P0.01）.There was negative correlation between the level of ZO-1 and DAO,D-Lac at 24h（γ：-0.8385,-0.7088,P =0.002,0.022）.Under light microscopy,intestinal mucousal injuries were noted in group A,and no evident changes occurred in group B.Conclusions In ischemic stroke,the activity of DAO and concentration of D-lac in plasma changes correspondingly with the destruction of intestinal mucosal function,so detection of these changesmay sensitively reflect the destruction of intestinal mucosal function early.Destruction of tight junction is one of the molecular mechanisms of intestine barrier dysfunction in ischemic stroke.