硫化氢预处理对大鼠全脑缺血/再灌注损伤的保护作用及机制

Protective Effect of H_2S Pretreatment on Cerebral Ischemia-reperfusion Injury and Its Mechanisms in Rats

目的探讨硫化氢对大鼠全脑缺血/再灌注损伤的保护作用及机制。方法采用双侧颈总动脉夹闭合并低血压方法建立全脑缺血/再灌注大鼠模型。将30只大鼠随机分为6组:(1)Sham组(n=5):大鼠接受假手术,仅暴露双侧颈总动脉和翼小孔,不作缺血处理;(2)脑缺血组(n=5):电凝针烧灼两侧翼小孔内的椎动脉,使椎动脉闭塞,术后24h闭塞双侧颈总动脉10min至脑缺血;(3)硫化氢预处理组(n=15):双侧颈总动脉闭塞30min前,进行腹腔注射硫氢化钠,分别为12、24、48μmol/kg剂量组,以上各组给药时均将药物按所需剂量溶解于0.5ml/100g生理盐水中;(4)NaCl预处理组(n=5):双侧颈总动脉闭塞30min前,腹腔注射等量生理盐水。观察硫化氢对海马组织超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量的影响,进行脑水肿测定,并检测脑内热休克蛋白70(HSP70)在海马组织中的表达水平。结果硫化氢低(P=0.042)、中(P=0.002)、高剂量组(P=0.000)海马组织中的SOD含量较脑缺血组明显增高,脑缺血组海马组织中的SOD活性较Sham组明显降低(P=0.003);硫化氢中(P=0.026)、高剂量组(P=0.015)海马组织中的MDA含量较脑缺血组明显降低;硫化氢中(P=0.018)、高剂量组(P=0.008)的脑组织含水量较脑缺血组明显降低,脑缺血组脑组织含水量较Sham组明显增高(P=0.009);硫化氢中剂量组的HSP70表达较脑缺血组明显减少(P=0.000),脑缺血组的HSP70表达较Sham组明显增加(P=0.000),NaCl预处理组与Sham组的HSP70表达差异有统计学意义(P=0.000)。结论硫化氢对全脑缺血/再灌损伤的保护作用机制可能与提高SOD活性,清除氧自由基,抑制脂质过氧化,下调HSP70蛋白在海马内的表达有关。

Objective To investigate the protective effect of H2S pretreatment after cerebral schemia/ reperfusion injury and its mechanisms in rats. Methods The rat model of global cerebral ischemia/reperfusion injury was established by bilateral common carotid arteries occlusion combined with hemorrhagic hypotension. 30 rats were randomly divided into four groups ： （ 1 ） sham group （ n = 5 ）, in which rats received sham surgery only, with their bilateral vertebral artery and bilateral common carotid artery exposed but without ischemia treat- ment; （2） global cerebral ischemia/reperfusion model group （ IR group, n = 5 ）, in which the global cerebral ischemia was induced by 10-min occlusion of bilateral common carotid arteries combined with hypotension; （3） H2S pretreatment group （n = 15）, in which H2S （12, 24, 48 ~mol/kg） was intraperitoneally injected before operation; （4） NaC1 pretreatment group （n = 5）, in which the rats were intraperitoneally injected with saline 30 minutes before operation. The activities of superoxide dismutase （SOD） and the levels of malondialdee- hyde （MDA） in brain were measured by spectrophotometry. Brain water content was detected. The expression of heat shock protein 70 （HSP70） in the hippocampus was determined by Western blotting. Results The SOD activities were significant increased in groups pretreated with 12μmol/kg H2S （ P = 0. 042）, 24p, mol/kg H2S （P =0. 002）, and 48μmol/kg H2S （P =0. 000）, and the SOD activity was significantly lower in the ischemia group than in the Sham group （P =0. 003）. The MDA activities in the 24p, mol/kg group （P =0. 026） and the 481μmol/kg group （P = 0. 015 ） groups were significantly lower than in the IR group. The brain water content was decreased in H2S pretreatment group （24μmol/kg and 48 μmol/kg） compared with IR group （P = 0. 018, P = 0. 008）, and it was also significantly higher in the IR group than in the sham group （ P = 0. 009 ）. The expression of HSP70 were decreased in H2S pretreatment group （24 μmol/kg） compared with the IR group （P = 0. 000）, and the expression of HSP70 were significantly higher in the IR group than in HSPT0 group （ P = 0. 000 ）. The expression of HSP70 also significantly differed between NaC1 group and HSP70 group （ P = 0. 000）. Conclusion H2S has protective effects on cerebral ischemia and reperfusion, which may be achieved by improving SOD activity, removing oxygen free radicals, inhibiting lipid peroxidation, and down-regnlating the expression of HSP70 in the hippocampus.