卡巴胆碱对肠缺血-再灌注大鼠外周血白细胞凋亡及细胞因子表达的影响

Effects of carbachol on apoptosis of peripheral white blood cells and expression of cytokines in rats suffering from gut ischemia/reperfusion injury

目的 观察拟胆碱药卡巴胆碱对肠缺血再灌注大鼠外周血淋巴细胞和中性粒细胞凋亡及细胞因子 m RNA表达的影响 ,探讨拟胆碱药的抗炎作用机制。方法 36只大鼠随机分为手术对照组、肠缺血 1h组、肠缺血 1h后再灌注 1h和 2 h组、卡巴胆碱 +肠缺血 1h组及卡巴胆碱 +肠缺血 1h后再灌注 2 h组。复制大鼠肠缺血再灌注模型 ;肠缺血前 10 m in经肠袋给予卡巴胆碱。检测不同时间点大鼠外周血白细胞计数及分类、淋巴细胞和中性粒细胞凋亡率及白细胞中细胞因子 m RNA表达水平。结果 肠缺血 1h,大鼠外周血白细胞计数减少 ,淋巴细胞比例增加 ,中性粒细胞比例减少 ,再灌注后上述指标均呈反向变化 ;卡巴胆碱可抑制肠缺血引起的外周血淋巴细胞凋亡率的增加 ,但增加中性粒细胞的凋亡率。外周血白细胞中致炎细胞因子肿瘤坏死因子 α(TNFα)和抗炎细胞因子白细胞介素 10 (IL 10 )表达在肠缺血 1h时均增加 ,但再灌注后 2h,IL 10、IL 4和 γ干扰素 (IFNγ)表达则明显减少 ,与 TNFα表达明显增加的趋势相反 ;卡巴胆碱可改善这一变化。结论 卡巴胆碱能减少肠缺血时淋巴细胞的凋亡 ,调节肠缺血再灌注时外周血白细胞中致炎和抗炎细胞因子表达失衡 ,在防治失控炎症反应、脓毒症和多器官功能障碍综合征中具有潜在临床应用价值。

Objective To evaluate the effect of cholinergic drug carbachol on apoptosis and expression of certain cytokines of peripheral blood lymphocytes and neutrophils in rats subjected to ischemia/reperfusion injury of the intestine. Methods Thirtysix Wistar male rats were randomly divided into groups as follows: sham operation, 1 and 2 hourgut ischemia, 1 hourgut ischemia followed by reperfusion for 1 hour and 2 hours , carbachol+1 hourgut ischemia , and carbachol+1 hourgut ischemia followed by reperfusion for 2 hours . Then the gut was subjected to ischemia/reperfusion. At different time points after the said injury, the total number and differential count of leukocytes, apoptosis rate of lymphocytes and neutrophils, and the mRNA expression levels of certain cytokines in peripheral blood leukocytes, were determined. Results At 1 hour after gut ischemia, the total number of leukocytes decreased (the ratio of lymphocytes increased and that of neutrophils decreased), but it increased after gut reperfusion. Carbachol could reduce the apoptosis rate of lymphocytes, but enhance that of neutrophils after gut ischemia. In leukocytes, mRNAs expression of inflammatory (TNFα) and antiinflammatory (IL10) cytokines were upregulated at 1 hour after gut ischemia , while that of IL10, IL4 and interferonγ were downregulated distinctly at 2 hours following reperfusion. All of these phenomena were ameliorated by giving carbachol. Conclusion Carbachol could reduce the peripheral lymphocytes apoptosis subsequent to gut ischemia, and regulate the balance between inflammatory and antiinflammatory cytokines expression in leukocytes during gut ischemia and reperfusion. The results suggest that carbachol might be a potential therapeutic agent in preventing uncontrolled inflammatory response as a result of ischemia/reperfusion injury of the intestine.