摘要
目的 观察热射病大鼠小肠上皮屏障通透性的改变,并通过观察肠上皮细胞紧密连接形态及肠上皮细胞紧密连接蛋白occludin蛋白表达的变化,初步探讨热射病对肠上皮屏障通透性的影响及机制.方法 将SD大鼠随机分为两组,每组10只（n=10）,热射病组及常温对照组,制备热射病大鼠模型,动物麻醉后放入42 ℃恒温箱热暴露50 min后取出,室温（26 ℃）放置2 h时,处死动物取材,观察热射病大鼠肠上皮细胞屏障通透性的改变（血浆中荧光剂FD4、内毒素）及炎性细胞因子浓度变化,并通过光镜（HE染色）及透射电镜观察小肠组织的大体及微观形态学改变.应用免疫组化及蛋白印迹杂交（Western bloting）技术检测各组肠上皮细胞紧密连接蛋白occludin蛋白表达变化情况.结果 热射病大鼠小肠上皮细胞屏障通透性（FD4、内毒素）及细胞因子水平显著高于正常对照组（P＜0.05）.形态学方面,光镜观察空肠HE染色,与正常大鼠比较,热射病组出现肠上皮细胞从微绒毛顶端滑塌,从多个镜下视野观察（≥3个视野/片）,此种表现不具有普遍性.透射电镜下,正常大鼠肠上皮细胞,紧密连接结构完整,呈致密的带状结构,热射病组紧密连接断裂,细胞间隙增宽,致密度减轻.免疫组织化学显示正常组大鼠肠上皮细胞,阳性信号沿细胞膜分布.热射病组荧光强度明显减弱.Western结果显示热射病组occludin蛋白表达明显低于对照组（P＜0.05）.结论 热射病引起紧密连接蛋白occludin的表达减少,紧密连接开放,从而破坏了肠上皮细胞间的紧密连接屏障,并因此引起肠上皮细胞屏障通透性的增高,导致内毒素血症及全身炎性介质反应.
Objective A stable rat model of heat stroke was established to investigate heat stroke-induced changes of the intestinal epithelial tight junction （TJ） barrier permeability, and to investigate the mechanism by observing the changes of TJ protein （occludin） expression and TJ morphology. Methods SD rats were randomly divided into two groups（n = 10）： heat stroke group and normal control group. Stable model of heat stroke was established. Anesthetized rats were exposed to 42 ℃ in a ventilated chamber for 50min, after heat exposure, the rats were placed in room temperature（26 ℃ ） for 2 h. Then the rats were sacrificed and samples were taken. The effect of heat stroke on intestinal epithelial barrier permeability was observed through changes of plasma FD4 and endotoxin concentration, cytokines concentration was detected as inflammatory indicators. The general and micro pathology was observed by light microscope （hematoxylin and eosin-staining, HE staining） and transmission electron microscope （TEM）. Occludin expression was investigated by Western bloting and immunochemistry. Results At 2 h after heat exposure, intestinal epithelial barrier permeability（ FD4 and endotoxin concentration）, and cytokines levels of heat stroke group were significantly higher than normal control group（P 〈 0. 05 ）. In morphology, by observing the light micrographs of HE jejunal tissue, the sloughing of epithelium off the basement membrane at the villus tips of the heat stroke group compared with the normal controlgroup. In many visual fields （VF）（ ≥6VF/slice）, this phenomenon was not universal. Under TEM, TJ of normal enterocytes was integrated with the compact zonal structure. At 2 h after heat exposure, TJ of heat stroke group was broken with widen intercellular space, and the density of TJ was decreased. In immunochemistry assay, the positive signal was distributed along the cell membrane in normal small intestinal tissue, while the signal was obviously decreased in heat stroke group.The results of Western bloting showed that occludin expresion of heat stroke group was significantly lower than normal control group（P 〈0.05）. Conclusion Heat stroke decreased occludin expression, opened the intestinal epithelial TJ, which destroyed intestinal epithelial tight junction barrier, induced high intestinal epithelial barrier permeability, produced endotoxemia and systemic inflammatory response syndrome （SIRS）.
出处
《国际外科学杂志》
2011年第1期7-11,F0003,共6页
International Journal of Surgery
