缺血预处理抗大鼠肠缺血再灌注肠黏膜损伤的蛋白质组学研究  

Proteomics study of intestinal mucosa after ischemic preconditioning against intestinal ischemic reperfusion injury in rats

作　　者：刘克玄[1] 李云胜[1] 王钟兴[1] 李偲[1] 刘家欣[1] 黄文起[1] 

机构地区：[1]中山大学附属第一医院麻醉科,广州510080

出　　处：《中华胃肠外科杂志》2009年第6期598-602,共5页Chinese Journal of Gastrointestinal Surgery

基　　金：国家自然科学基金（30672021）

摘　　要：目的采用蛋白质组研究技术分离、鉴定缺血预处理（IPC）抗大鼠肠缺血再灌注（Ⅱ／R）肠黏膜损伤相关蛋白，探讨其肠保护分子机制。方法将16只SD大鼠，随机分为Ⅱ／R组和IPC组。Ⅱ／R组阻断肠系膜上动脉60min后再开放60min;IPC组在阻断肠系膜上动脉前先阻断20min后再开放5min。余同Ⅱ／R组。再灌注结束即刻刮取肠黏膜，利用高分辨双向电泳对肠黏膜组织进行蛋白质分离．Image Master 2D Elite 5．0图像软件进行分析。应用基质辅助电离解析飞行时间质谱获取肽质量指纹图谱．检索数据库鉴定表达差异的蛋白质，明确其生物学功能。结果双向电泳发现，Ⅱ／R组及IPC组分别有蛋白质点（1404±20）个和（1338±20）个。10个点进行质谱分析，8个蛋白质点经过检索与已知蛋白质匹配．这些蛋白功能涉及到抗氧化、抑制凋亡及改善能量代谢。RT-PCR分析提示IPC上调醛糖还原酶的表达。Western blot分析提示IPC上调醛脱氢酶的表达。结论比较蛋白组学研究揭示IPC对肠缺血再灌注损伤的保护机制可能与其上调了一些具有抗氧化、抑制细胞凋亡及改善能量代谢作用的蛋白有关。Objective To identify associated proteins involved in the molecular response of ischemic preconditioning （IPC） against intestinal ischemia/reperfusion（Ⅱ/R） in the intestinal mucosa of rats. Methods Sixteen SD rats were randomly divided into Ⅱ/R and IPC groups. Ⅱ/R injury in rats was produced by clamping superior mesenteric artery for 60 min followed by 60 min reperfusion. IPC was elicited by 20 min ischemia and 5 min reperfusion before index ischemia. The intestinal mucosa was scratched immediately after 60 rain of reperfusion and total proteins were separated by immobilized pH gradient（IPG） based on two-dimensional gel electrophoresis（2-DE）. The differentially expressed proteins were analyzed using Image Master 2D Elite 5.0 image analysis software and identified by MALDI-TOFMS. The biological information of these proteins was searched in the database of these peptide mass finger-printing （PMF）. Western blotting and RT-PCR were used to validate the differentially expressed proteins. Results Image analysis revealed that averages of 1404±20 and 1338±20 were detected in Ⅱ/R and IPC groups. A total of 10 spots yielded good spectra, and 8 spots matched with known proteins after database searching. These proteins were mainly involved in anti-oxidation, inhibiting apoptosis and energy metabolism. Western blot confirmed up-regulation of aldehyde dehydrogenase and RT-PCR confirmed up-regulation of aldose reductase in IPC group. Conclusion The clues provided by comparative proteome strategy will shed light on molecular mechanisms of IPC against Ⅱ/R injury.

关 键 词：缺血预处理 小肠 再灌注损伤 蛋白质组学 

分 类 号：R57[医药卫生—消化系统]