双歧杆菌分泌型粘附素对肠上皮细胞应激反应的影响  

The effects of bifidobacterial secretory adhesin on the stress response of intestinal epithelial cells

作　　者：钟世顺[1] 张振书[2] 李淑梅[3] 赖卓胜[2] 李东良[4] 

机构地区：[1]福建医科大学省立临床医学院福建省立医院消化内镜中心,福州350001 [2]南方医科大学南方医院消化科 [3]南京军区福州总医院476临床部内一科 [4]南京军区福州总医院肝胆内科

出　　处：《中华消化杂志》2011年第11期744-749,共6页Chinese Journal of Digestion

基　　金：基金项目：福建省青年人才资助项目（2006F3107）

摘　　要：目的观察双歧杆菌分泌型粘附素对肠上皮细胞应激反应后核因子（NF）-κB DNA结合活性、细胞胞质核因子抑制蛋白κB（IκB）、细胞胞核NF-κB p65的表达和肿瘤坏死因子（TNF）-α,白细胞介素（IL）-1β、IL-8等细胞因子表达的影响。方法经培养的肠上皮Lovo细胞分为5组，分别经LPS（100ng／ml）、H2O2（200μmol／L）直接刺激3h以及经双歧杆菌分泌型粘附素（30μg／ml）预孵30min后再予LPS（100ng／ml）、H2O2刺激（200μmol／L）刺激3h，正常对照组未作任何处理。采用凝胶电泳迁移率试验（EMSA）方法检测NF-κB DNA结合活性；Western印迹法分别检测细胞胞质IκB和细胞胞核NF-κBp65的表达；RT-PCR方法检测TNF-α、IL-1β、IL-8等细胞因子mRNA的表达情况。结果LPS和H2O2刺激后，细胞表现出较高的NF-κBDNA结合活性[分别为正常对照组的（6．20±0．35）倍和（4．16±0．52）倍]和细胞胞核NF-κBp65的表达[分别为（0．64±0．05）和（0．67±0．06）]，而细胞胞质IκB的表达则较弱[分别为（0.28±0．10）和（0．39±0．12）]；以双歧杆菌分泌型粘附素预孵后，前二者活性明显降低，而后者的表达明显增强；LPS和H2O2处理后，TNF-α、IL-1β、IL-8等细胞因子mRNA的表达水平均明显增高[LPS处理组分别为（0．92±0．10）、（0．38±0．03）、（1．44±0．25），H2O2处理组分别为（O．89±0．13）、（0．36±0．06）、（1．42±0．18）]，而粘附素预孵后则可显著降低它们的表达水平。NF-κB DNA结合活性与TNF-α、IL-1β、IL-8三种细胞因子的mRNA表达水平均呈显著正相关。结论双歧杆菌分泌型粘附素对LPS和H2O2诱导的肠上皮细胞NF-κB DNA结合活性有显著抑制作用，NF-κB的活化可能与TNF-α、IL-1β和IL-8等炎性细胞因子的表达调控有关。Objective To observe the effects of bifidobacterial sectory adhesin on NF-κB DNA binding activity, cytoplasm InB, nuclei NF-κBp65, TNF-α, IL-I±, and IL-8 expression in intestinal epithelial cell post stress response. Methods The cultured intestinal epithelial cell line Lovo cells were divided into 5 groups, which included directly stimulated by lypopolysaecharide （LPS） （100 ng/ml） or H2O2 （200 t±mol/L） for 3 hours groups, hifidobacterial sectory adhesin （30 ±g/ml） pre-incuhation for 30 minutes and then treated by LPS （100 ng/ml） or H2O2 （200 ±mol/L） for 3 hours groups and control group with no treatment. NF-κB DNA binding activity was evaluated by electrophoretic mobility shiftassay （EMSA）. The cytoplasm IκB and nuclei NF-κBp65 expression were determined by Western blot. The expressions of TNF-α, IL-1β and IL-8 at mRNA level were detected by semi-quantatitive assay of RT-PCR. Results The expressions of NF-κB DNA binding activity [-（6.20-t-0.35） times and （4.16＋0.52） times of that of non treated control group, respectively] and nuclei NF-κBp65 [,（0.64±0.05） and （0.67-t-0.06）] were higher in cells after LPS or H2O2 stimulated, however the cytoplasm IeB expression [-（0. 28 -I- 0. 10） and （0. 39 _＋ 0. 12） respectively＇] was weaker. After bifidobaeterial sectory adhesin pre-ineubation, the expressions of NF-nB DNA binding activity and nuclei NF-κBp65 decreased obviously, but cytoplasm IκB expression increased. After treated with LPS or H2O2, the mRNA expressions of TNF-α, IL-11] and IL-8 were significantly increased [LPS treated group： （0.92 ±0.10）, （0.38±0.03）, （1.44±0.25）, H2O2 treated group： （0.89±0.13）, （0.36±0.06）, （1.42 ±0. 18）]. After bifidobacterial sectory adhesin pre-incubation, their expressions decreased. The mRNA expressions of TNF-α, IL-1β and IL-8 have significantly positive correlation with NF-κB DNA binding activity. Conclusions There is a significant inhibition role of bifidobacterial sect

关 键 词：二裂菌属 细胞粘附 肿瘤细胞 培养的 肠 脂多糖类 过氧化氢 核转录因子 ΚB 细胞因子类 

分 类 号：R574[医药卫生—消化系统]