摘要
目的:验证紧密连接蛋白ZO-1、Claudin-5和Occludin保护脓毒症肠屏障功能与血管扩张刺激磷蛋白(vasodilator-stimulated phosphoprotein,VASP)相关.方法:盲肠结扎穿孔术构建脓毒症小鼠模型,实验分为:对照组、假手术组、模型组和VASP磷酸化(p-VASP)激动剂Bucladesine组.ELISA检测血清中二胺氧化酶(DAO)和D-乳酸(D-LA)含量.HE染色观察小肠黏膜组织形态.TUNEL染色检测小肠上皮细胞凋亡.Western blotting检测Bax、Bcl-2、p-VASP、ZO-1、Occludin和claudin-5蛋白表达水平.结果:与对照组和假手术组相比,模型组中小肠绒毛数量减少、绒毛断裂、TUNEL阳性细胞增加、DAO和D-LA含量升高(P＜0.05),Bax蛋白、p-VASP蛋白表达水平升高(P＜0.05),ZO-1、Occludin和claudin-5蛋白、Bcl-2蛋白表达水平降低(P＜0.05).与模型组相比,Bucladesine组中小肠绒毛数量增加、绒毛断裂减少、TUNEL阳性细胞减少、DAO和D-LA含量降低(P＜0.05),p-VASP蛋白表达水平降低(P＜0.05),ZO-1、Occludin和claudin-5蛋白表达水平升高(P＜0.05).结论:抑制脓毒症小鼠p-VASP后小肠上皮屏障功能恢复,这与上调紧密连接蛋白表达水平有关.
Abstract
Objective:To verify that the tight junction proteins ZO-1,Claudin-5 and Occludin in protecting intestinal barrier function in sepsis were associated with vasodilator-stimulated phosphoprotein(VASP).Methods:A mouse model of sepsis was constructed by cecum ligation perforation,and the rats were divided into the control group,sham-operated group,model group and Bucladesine of VASP phosphorylation agonist group.The serum levels of diamine oxidase(DAO)and D-lactic acid(D-LA)were measured by ELISA.HE staining was performed to observe the histomorphology of the small intestinal mucosa.TUNEL staining was used to detect the apoptosis of small intestinal epithelial cells.Bax,Bcl-2,p-VASP,ZO-1,Occludin and claudin-5 protein expression levels were detected by Western blotting.Results:Compared with the control and sham-operated groups,the number of small intestinal villi reduced,the villi broken,the TUNEL-positive cells increased,the levels of DAO and D-LA significantly increased(P＜0.05),the levels of p-VASP protein expression significantly increased(P＜0.05),and the levels of ZO-1,Occludin and claudin-5 protein expression significantly decreased in the model group(P＜0.05).Compared with the model group,the number of small intestinal villi increased,the villi breakage decreased,the TUNEL-positive cells decreased,the DAO and D-LA content significantly decreased(P＜0.05),the p-VASP and Bax protein expression levels significantly decreased(P＜0.05),and the expression levels of ZO-1,Occludin,claudin-5 and Bcl-2 protein increased in the Bucladesine group(P＜0.05).Conclusions:The function of small intestinal epithelial barrier recovers after inhibiting the p-VASP in sepsis mice,which is related to up-regulation of tight junction protein expression.
维普
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