双歧杆菌通过Zonulin影响坏死性小肠结肠炎新生大鼠模型肠道通透性  

Effect of bifidobacterium on intestinal permeability and expression of zonulin in neonatal rats with necrotizing enterocolitis

作　　者：向玲[1] 杜维霞[1] 刘依枫 彭玲珑 韦红[1] 

机构地区：[1]重庆医科大学附属儿童医院新生儿科,儿童发育疾病研究省部共建教育部重点实验室,儿科学重庆市重点实验室,重庆市儿童发育重大疾病诊治与预防国际科技合作基地,重庆400014

出　　处：《第三军医大学学报》2016年第5期469-475,共7页Journal of Third Military Medical University

基　　金：国家自然科学基金(30200300);国家临床重点专科建设项目经费基金(卫办医政[2011]873)~~

摘　　要：目的探讨双歧杆菌对坏死性小肠结肠炎(necrotizing enterocolitis,NEC)新生大鼠模型肠道通透性及血清Zonulin表达的影响,分析双歧杆菌保护NEC可能的机制。方法将60只新生1 d的SD大鼠按随机数字表法分为3组(n=20):对照组、坏死性小肠结肠炎新生大鼠组(NEC组)及双歧杆菌干预坏死性小肠结肠炎新生大鼠组(NEC+BIF组)。3组于第1、2、3天固定时间点测量体质量;第4天处死大鼠,HE染色观察回盲部肠组织病理学变化并行病理学评分,异硫氰酸荧光素-葡聚糖(FITCDextran)示踪法检测肠黏膜通透性,ELISA检测血清Zonulin的水平,Western blot及免疫组化检测肠组织紧密连接蛋白的表达及分布,将3组大鼠血清Zonulin水平与肠道通透性及紧密连接蛋白表达量行相关性分析。结果与NEC组相比,双歧杆菌干预后大鼠肠上皮损伤明显改善;而与对照组相比,NEC组肠道通透性显著增高(P<0.05),血清Zonulin表达明显增加(P<0.05),肠上皮紧密连接蛋白Claudin-1、Occludin、ZO-1的表达明显减少(P<0.05),且分布不连续;与NEC组相比,NEC+BIF组肠道通透性降低(P<0.05),血清Zonulin表达降低(P<0.05),肠上皮紧密连接蛋白的表达增加(P<0.05)。相关性分析显示,3组大鼠血清Zonulin水平与肠道通透性呈正相关(r=0.923,P<0.01),而血清Zonulin水平与肠组织紧密连接Claudin-1、Occludin、ZO-1蛋白表达量均呈负相关(r=-0.779,P<0.01;r=-0.920,P<0.01;r=-0.619,P<0.01)。结论双歧杆菌可明显改善NEC大鼠的肠道通透性,其机制可能与抑制Zonulin的表达有关。Objective To determine the effect of Bifidobacterium on the intestinal permeability and serum expression of zonulin in neonatal rats with necrotizing enterocolitis （NEC）, and discuss the protective mechanism of Bifidobacterium from NEC. Methods According to random number table, 60 1-day-old Sprague-Dawley rats were divided into 3 groups： control group, NEC group, and Bifidobacterium intervention group （NEC＋BIF group） （n=20）. The weights of rats were measured in 3 consecutive days at same time. After day 4, all the rats were put to death, and the proximal intestines of ileocecal junction underwent pathological sectioning. HE staining was carried out to observe histological changes and score the tissue damage. The intestinal permeability was analyzed by fluorescein isothiocyanate dextran （FITC-Dextran） tracing. The serum zonulin was detected by ELISA. The expression and distribution of tight junction proteins （Claudin-1, Occludin, and ZO-1） were analyzed by Western blotting and immunohistochemical assay. Finally, the correlation analysis between the intestinal permeability, the expression of tight junction proteins and the serum level of zonulin was performed. Results Compared with the NEC group, the intestinal injury was obviously alleviated after Bifidobacterium intervention. Compared with the control group, the intestinal permeability and the level of zonulin in the NEC group were significantly increased （both P〈0.05）, and the expression of tight junction proteins （Claudin-1, Occludin, ZO-1） was significantly decreased （P〈0.05）, with discontinuous distribution. Compared with the NEC group, the intestinal permeability was decreased （P〈0.05）, the expression of zonulin was significantly decreased （P〈0.05）, and the expression of tight junction proteins was increased （P〈0.05） in the NEC＋BIF group. The correlation analysis showed that the level of zonulin was positively correlated with the intestinal permeability （r=0.923, P〈0.01）, and negatively correl

关 键 词：坏死性小肠结肠炎 双歧杆菌 ZONULIN 肠道通透性 紧密连接 

分 类 号：R363[医药卫生—病理学] R378.994[医药卫生—基础医学]