ALCAM沉默对新生大鼠坏死性小肠结肠炎的保护作用研究  

Protective effect of ALCAM silencing on necrotizing enterocolitis in neonatal rats

作　　者：吴臻斐 田由 章捷 林进汉[2] WU Zhenfei;TIAN You;ZHANG Jie;LIN Jinhan(Department of Surgery,Hangzhou Children's Hospital,Hangzhou 310005,China;不详)

机构地区：[1]杭州市儿童医院外科,310005 [2]温州医科大学附属育英儿童医院儿童外科

出　　处：《浙江医学》2023年第24期2595-2599,I0004,共6页Zhejiang Medical Journal

基　　金：浙江省医药卫生科技计划项目(2022KY1012)。

摘　　要：目的 研究活化白细胞黏附分子(ALCAM)沉默对坏死性小肠结肠炎(NEC)的影响及作用机制。方法 24只雄性SD新生大鼠按随机数字表法分为对照组、NEC组、沉默阴性对照组、ALCAM沉默组,每组6只。NEC组、沉默阴性对照组和ALCAM沉默组分别尾静脉注射0.9%氯化钠注射液、阴性对照腺相关病毒载体和ALCAM腺相关病毒载体;除对照组外,各组大鼠采用人工饲养与缺氧复氧和冷刺激诱导以建立NEC模型。采用ELISA法测定各组大鼠血清IL-1β、IL-6水平,采用HE染色法、Tunel染色法评价肠组织病理改变和细胞凋亡情况,采用免疫组化染色法测定CD166表达率,采用qRT-PCR法检测肠组织ALCAM的mRNA相对表达量,采用Western blot法检测肠组织ALCAM、E-钙黏蛋白(E-cadherin)、β-连环蛋白(β-catenin)相对表达量。结果 与对照组比较,NEC组大鼠血清IL-1β、IL-6水平升高,肠组织病理改变明显,细胞凋亡率增加,CD166表达率升高,肠组织ALCAM的mRNA及蛋白、β-catenin蛋白相对表达量升高,E-cadherin蛋白相对表达量降低,差异均有统计学意义(均P<0.01)。与沉默阴性对照组比较,ALCAM沉默组新生大鼠血清IL-1β、IL-6水平降低,肠组织病理改变减轻,细胞凋亡减少,CD166表达率降低,肠组织ALCAM的mRNA及E-cadherin蛋白相对表达量升高,ALCAM蛋白、β-catenin蛋白相对表达量降低,差异均有统计学意义(均P<0.05)。结论 沉默ALCAM表达对NEC新生大鼠的肠道损伤具有保护作用,其作用与炎性细胞因子水平下降、E-cadherin/β-catenin通路蛋白表达改变有关。Objective To investigate the effect of activated leucocyte cell adhesion molecule(ALCAM)silencing on necrotizing enterocolitis(NEC)in neonatal rats and its mechanism.Methods Twenty-four neonatal male SD rats were randomly divided into normal control group,NEC model group,ALCAM silencing control group and ALCAM silencing group with six rats in each group.The rat NEC model was induced by hypoxia-reoxygenation and cold stimulation method.Tail vein injection of normal saline(0.2 mL),negative adeno-associated virus vectors(0.2 mL)or ALCAM silencing adeno associated viral vector(0.2 mL)were given to the rats in NEC model group,ALCAM silencing control group,and ALCAM silencing group 1 h before modelling,respectively.qRT-PCR was used to analyze ALCAM gene expression in tissue,ELISA was used to detect levels of IL-1βand IL-6 in serum,hematoxylin-eosin staining and Tunel staining were used to evaluate pathological change and cell apoptosis,immunohistochemical staining was used to analyze CD166 content in tissue,and Western blot was used to analyze expression of ALCAM,E-cadherin,β-catenin proteins.Results Compared to normal control group,the NEC model group showed increased expression levels of ALCAM mRNA and CD166 in intestinal tissue,increased apoptosis rate,and increased serum IL-1βand IL-6 levels and the expression of ALCAM andβ-catenin proteins,while decreased E-cadherin protein expression(all P<0.01).Compared to ALCAM silencing negative control group,the mRNA and protein expression of ALCAM and the content of CD166 in intestinal tissue of ALCAM silencing group were decreased,the pathological changes were alleviated,apoptosis reduced,the levels of serum IL-1βand IL-6 and the protein expression of ALCAM andβ-catenin were decreased,while expression levels of E-cadherin protein were increased(all P<0.05).Conclusion ALCAM Silencing has a protective effect on the intestinal injury of neonatal NEC rats,which is related to the decline of inflammatory cytokines,changes in the expression of E-cadherin/β-catenin pathway p

关 键 词：坏死性小肠结肠炎 活化白细胞黏附分子 腺病毒载体 炎性细胞因子 E-钙黏蛋白/β-连环蛋白通路 

分 类 号：R722.1[医药卫生—儿科]