热休克预处理对大鼠肠缺血-再灌注损伤的保护效应

Protective effect of previous heat shock treatment on intestinal injury following mesenteric ischemia/reperfusion in rats

目的:探讨全身热休克预处理对肠缺血-再灌注(ischemia/reperfusion,IR)损伤的保护作用.方法:将40只SD♂大鼠随机分为4组:正常体温+假手术对照组(CTRL,n=10),正常体温+肠IR组(IR,n=10),41.5℃-42℃热休克+假手术组(42C,n=10),41.5℃-42℃热休克+肠IR组(42IR,n=10).Western blot检测肠黏膜HSP72蛋白表达,原位末端缺口标记法(TUNEL)检测肠黏膜上皮细胞凋亡,比色法检测肠黏膜caspase-3活性,Annexin-V/PI法流式细胞仪检测外周血白细胞凋亡比例.结果:42C、42IR组HSP72蛋白表达水平比CTRL、IR组显著增高(1.59±0.32、2.71±0.64 vs 0.41±0.1、0_30±0.04.P<0.01).42IR组caspase-3活性.白细胞凋亡比例比IR组相比有显著性差异(1.16±0.31 vs 2.32±0.54;39.65%vs 16.94%;P<0.01),且42IR组肠黏膜上皮细胞凋亡指数比IR组明显减少.42IR组与42C、CTRL组之间,caspase-3活性,肠黏膜上皮细胞凋亡指数,白细胞凋亡比例均无明显差异.结论:全身热休克预处理对肠缺血-再灌注损伤有保护作用,其机制可能与增加热休克诱导HSP72的表达和抑制外周血白细胞激活有关.

AIM： To investigate the protective effect of previous heat shock treatment on intestinal injury following mesenteric ischemia/ reperfusion （I/R）. METHODS： Forty adult male Sprague-Dawley rats were divided into 4 groups （10 in each group）： control group （CTRL）, ischemia/reper- fusion group （IR）, previous heat shock treatment group （42C） and ischemia reperfusion group （42IR）. The expression of HSP72 was examined in the small intestinal mucosa using Western blotting, and pathological changes in the ileal mucosal tissue were evaluated by microscopy.The apoptosis of intestinal mucosal epithelial cells was examined by terminal deoxylnucleotidyl transferase mediated-dUTP nick end labeling （TUNEL）. The enzymatic activity of caspase-3 in mucosal cells was assayed colorimetrically. The percentage of peripheral blood leukocytes undergoing apoptosis was measured by flow cytometry using an annexin-V/PI double staining assay. RESULTS： The expression of heat-shock protein 72 was significantly increased in 42C and 42IR rats compared with CTRL and IR rats （1.59 ± 0.32, 2.71 ± 0.64 vs 0.41 ± 0.1, 0.30 ± 0.04, P 〈 0.01）. The enzymatic activity of caspase-3 decreased significantly in the 42IR group compared with the IR group （1.16 ± 0.31 vs 2.32 ± 0.54, P 〈 0.01）. The percentage of peripheral blood leukocytes undergoing apoptosis was significantly higher in the 42IR group than in the IR group （39.65% vs 16.94%, P 〈 0.01）. The number of mucosal epithelia cells undergoing apoptosis was decreased in the 42IR the corhpared with the IR group. There were no significant differences in the enzymatic activity of caspase-3, in the percentage of peripheral blood leukocytes undergoing apoptosis and in the number of mucosal epithelial cells undergoing apoptosis among 42IR, 42C and CTRL rats. CONCLUSION： Whole body hyperthermia pretreatment can protect against subsequent mesenteric ischemia reperfusion injury by increased induction of heat shock protein 72 and inhibition of peripheral blood leukocyte activation in rats.