锌指蛋白A20在炎症性肠病患儿肠黏膜中的表达及意义

Expression of zinc finger protein A20 in pediatric infiammatory bowel disease

【摘要】目的探讨炎症性肠病（IBD）患儿肠道炎症反应与锌指蛋白A20（A20）表达水平之间的关系。方法收集2008至2010年就诊于我院并行肠镜检查的患儿肠道黏膜标本共57份。将标本分为正常对照组（It=16）、IBD缓解期组（It=12）、IBD活动期组（n=13）和非IBD肠炎组（n=16）。内镜下取各组患儿末端回肠黏膜标本，采用荧光定量PCR和免疫组化法检测A20、NF—KB、IL-6、IL-8的表达水平。结果（1）NF—KB、A20在正常对照组肠黏膜中仅微量表达，IBD活动期组和非IBD肠炎组NF—KB、A20表达水平明显高于正常对照组（P均〈0．01）；（2）IBD缓解期组较正常对照组NF—KB[（9．35±4．84）％vs（0．57±0．44）％，P〈0．011、IL_6（t＇=1．34，P〉0．05）、IL-8（t=1．38，P〉0．05）表达水平高，而A20在mRNA水平（t=1．03，P〉0．05）和蛋白水平[（0．36±0．18）％vs（0．87±0．29）％，P〈0．01]上表达均偏低；（3）与非IBD肠炎组相比，IBD活动期组NF—KB[（24．17±11．27）％vs（55．29±21．84）％，P〈0．01]、IL-6（t=2．22，P〈0．05）、IL-8（t=2．97，P〈0．01）表达水平明显升高，而A20在mRNA（t=2．26，P〈0．05）和蛋白水平[（29．23±11．70）％VS（16．814-5．90）％，P〈0．01]上表达均较低；（4）IBD缓解期组与非IBD肠炎组相比，IL-6、IL-8表达水平差异无统计学意义（t＇值和t值分别为0．03和0．28，P均〉0．05），而A20在mRNA水平（t=4．42，P〈0．01）和蛋白水平[（29．23±11．70）％vs（0．47±0．25）％，P〈0．01]上表达均较低。结论IBD患儿存在肠道炎症反应过度而A20表达水平上调不足的现象；A20表达水平的异常可能参与了IBD的发生和发展。

Objective It is demonstrated that excessive activation of NF-KB is central to the pathogenesis of inflammatory bowel disease （IBD）. Zinc finger protein A20 （A20） is a key player in the negative feedback regulation of NF-KB signaling in response to multiple stimuli and has been described as central gatekeeper in inflammation and immunity. Mice genetically deficient in A20 develop severe intestinal inflammation and have increased susceptibility to dextran sodium sulfate（DSS）-induced colitis. Few studies have been done to explore the role of A20 in the pathogenesis of IBD. To clarify the relationship between intestinal inflammation and the expression level of A20 in IBD patients, the expression level of A20 and a series of inflammatory cytokines, such as NF-KB, IL-6, and IL-8, in children with IBD and controls were examined. Method Terminal ileal mucosal samples were obtained via endoscopy. Fifty-seven mueosal samples were divided into 4 groups： normal control group （n = 16 ）, IBD remission group （n -- 12 ） , IBD active group （n = 13） and non-IBD enteritis group （n = 16）. According to disease activity index scores, the IBD patients were divided into IBD remission group and IBD active group. Normal control group was consisted of patients with functional bowel disorders or intestinal polyps. Non-IBD enteritis was defined as changes in which endoscopy and histological examination showed inflammatory changes but could not be diagnosed as IBD. Real-time PCR was adopted for detecting the mRNA levels of A20, IL-6 and IL-8. Meanwhile immunohistoehemistry was performed to measure the expression of A20 and NF-KB. Result （ 1 ） The expression of A20 and NF-KB were very low in normal control group, but significantly up-regulated in IBD active group and non-IBD enteritis group （ P 〈 0. 01 for both） ; （2） Compared with normal control group, expression of NF-KB [（9.35±4. 84）% vs. （0.57±0.44）%, P〈0.01], IL-6 （t＇=1.34, P〉0. 05 ）, IL-8 （t = 1.38, P 〉 0. 05 ） increased in IBD remission group, while the expression of A20 in both mRNA （t=1.03, P〉0.05） and protein levels [（0.36±0.18）% vs. （0.87±0.29）%, P〈0.01] decreased; （3） Compared with non-IBD enteritis group, although the expression of NF-KB [ （24. 17±11.27）% vs. （55.29±21.84）%, P〈0.01], IL-6 （t=2.22, P〈0.05）, IL-8 （t =2.97, P〈0.01） were highly increased in IBD active group, the expression of A20 in both mRNA（ t = 2. 26, P 〈 0. 05 ） and protein levels [ （29.23 _＋ 11.70） % vs. （ 16.81 ＋ 5.90 ） % , P 〈 0. 01 ] significantly decreased ; （4） The expression of IL-6, IL-8 were similar in IBD remission group and non-IBD enteritis group （ both P 〉 0. 05 ）, but the expression of A20 was much lower in both mRNA （t = 4.42, P 〈 0. 01 ） and protein levels [ （29.23±11.70）% vs. （0.47±0.25）%, P 〈 0.01] in IBD remission group. Conclusion The results demonstrate that there is an excessive inflammatory response but insufficient up-regulation of A20 expression in IBD patients. Low levels expression of A20 may play an important role in the pathogenesis of IBD.