S-亚硝基谷胱甘肽对小鼠肠急性缺血再灌注损伤后肠屏障功能的影响

The effect of S-Nitrosoglutathione on the intestinal epithelial barrier after acute ischemia reperfusion injury

目的探讨S-亚硝基谷胱甘肽（GSNO）预处理对小鼠肠急性缺血再灌注（I/R）所致的肠黏膜屏障损伤的影响。方法6~8周龄雄性C57BL/6小鼠24只,分为Sham组、I/R组、I/R＋GSNO组,每组8只。采用夹闭肠系膜上动脉30min,再灌注6h作为小鼠肠I/R模型。苏木素-伊红（HE）染色观察小肠组织形态学的变化;免疫组织化学观察肠上皮细胞间紧密连接蛋白claudin-1的表达情况;Western blot检测claudin-1蛋白水平变化。结果 HE染色显示,与Sham组比较,I/R组肠黏膜明显肿胀,绒毛部分脱落、变粗,绒毛倒伏、断裂,而I/R＋GSNO组肠黏膜无明显肿胀,绒毛完整,形态接近正常;免疫组织化学观察显示I/R刺激可造成肠上皮表面claudin-1连续性明显破坏,阳性表达率明显降低,I/R＋GSNO组肠上皮claudin-1连续性明显恢复,阳性染色显著增加;肠上皮claudin-1蛋白定量分析显示：与Sham组比较,I/R组及I/R＋GSNO小肠上皮紧密连接蛋白claudin-1表达分别下降32.5%,13.8%（P〈0.05）;与I/R组比较,I/R＋GSNO组小鼠肠上皮紧密连接蛋白claudin-1表达上调27.8%（P〈0.05）。结论小肠急性I/R刺激可造成明显的肠黏膜损伤及肠上皮间紧密连接的显著破坏;GSNO预处理可通过上调claudin-1表达,有效缓解I/R对肠黏膜结构与肠上皮屏障功能的损伤。

Objective To investigate the effect of S-Nitrosoglutathione（GSNO）on acute ischemia reperfusion（I/R）induced intestinal barrier function lesion in a mouse model.Methods Twenty-four 6-8-year-old C57BL/6mice were divided into 3groups,8for each：（1）the sham group;（2）the I/R group;（3）the I/R＋GSNO group.The mouse intestine I/R model was established by the occlusion of the superior mesenteric artery temporarily followed by reperfusion for 6h.Histological changes in the small intestine were observed after HE staining;the expression of claudin-1protein in the intestine epithelium was assessed by immunohistochemistry staining as well as western blot analysis.Results Both HE staining and immunohistochemistry results showed the integrate intestinal villi with the continuous Claudin-1expression alone the villi in the sham group;the intestinal villi of the I/R group partially detached,thickened,crooked and fractured,with the obvious disconnection of Claudin-1staining alone the top of the villi;while the intestinal villi of the I/R＋GSNO group were neatly arranged and damage to intestinal mucosa was much alleviated,accompanied with the marked restoration of the continuity of claudin-1staining.Compared to the sham group,claudin-1protein level of for the I/R group and the I/R＋GSNO group decreased by 32.5%and 13.8%respectively（P0.05）;and compared to the I/R group,claudin-1protein level of the I/R＋GSNO group increased by 27.8%（P0.05）.Conclusion Protein level of claudin-1would decreases after I/R,and pretreatment with GSNO can effectively relieve the damage of intestinal mucosal structure as well as intestinal tight junction barrier through upregulating the expression of claudin-1protein.