HO-1在七氟烷预处理抑制大鼠氧糖剥夺海马神经元凋亡中的作用

Role of HO-1 in inhibition of oxygen-glucose deprivation-induced apoptosis in rat hippocampal neurons by sevoflurane preconditioning

目的 探讨血红素氧合酶-1（HO-1）在七氟烷预处理抑制大鼠氧糖剥夺海马神经元凋亡中的作用.方法 出生48 h内的Wistar大鼠,体外培养海马神经元,随机分为6组,每组108孔,正常对照组（C组）：不做任何处理;2%七氟烷预处理组（S1组）：2%七氟烷预处理60 min后正常培养24 h;OGD组：缺糖缺氧45 min后复糖复氧,再正常培养24 h以制备氧糖剥夺模型;2%七氟烷预处理＋OGD 组（S1＋OGD组）和4%七氟烷预处理＋OGD组（S2＋OGD组）：分别经2%、4%七氟烷预处理后制备氧糖剥夺模型;4%七氟烷预处理＋ZnPPⅨ＋OGD（Z组）：4%七氟烷预处理同时在培养液中加入ZnPPⅨ（终浓度10 μmol/L）,其余处理同S2＋OGD组.正常培养24 h时检测神经元存活率、凋亡率和HO-1蛋白及其mRNA的表达水平.结果 与C组比较,OGD组、S1＋OGD组、S2＋OGD组和Z组海马神经元存活率降低,凋亡率升高,海马神经元HO-1 mRNA及其蛋白表达上调,S1组海马神经元HO-1 mRNA 及其蛋白表达上调（P〈0.01）,神经元存活率、凋亡率差异无统计学意义（P〉0.05）;与OGD组比较,S1＋OGD组、S2＋OGD组海马神经元存活率升高,凋亡率降低,海马神经元HO-1 mRNA及其蛋白表达上调,Z组上述指标差异无统计学意义（P〉0.05）;与S1＋OGD组比较,S2＋OGD组海马神经元存活率升高,凋亡率降低,海马神经元HO-1 mRNA及其蛋白表达上调（P〈0.01）;与S2＋OGD组比较,Z组海马神经元存活率降低,凋亡率升高,海马神经元HO-1 mRNA及其蛋白表达下调（P〈0.01）.结论 HO-1可能参与了七氟烷预处理抑制大鼠氧糖剥夺海马神经元凋亡的机制.

Objective To investigate the role of HO-1 in inhibition of oxygen-glucose deprivation （OGD）-induced apoptosis in rat hippocampal neurons by sevoflurane preconditioning.Methods Hippoeanlpal neurons of newborn Wistar rats （〈48 h） were cultured in vitro.Tne neurons were randomly divided into 6 groups with 108 wells in each group：control group（group C）,2% sevoflurane preconditioning group （group S1）,OGD group,S1 ＋OGD group,4% sevoflurane preconditioning＋OGD group （group S2＋OGD）,and 4% sevoflurane preconditioning＋ZnPPⅨ＋OGD group（group Z）.Group C received no treatment.The neurons were cultured for 24 h after 2% sevoflurane preconditioning in group S1.For OGD experiments,the neurons were placed in deoxygenated glucose-free medium and sealed under 95% N2-5% CO2 in an anaerobic chamber equilibrated to 37℃ and 100%humidity for 45 min.then OGD was terminated by replacement of the stored medium and returning the cultures to a standard incubator maintained at 37℃ in 5% C02 and the neurons were cultured for 24 h as described by Ray et al. The OGD model was established after 2% and 4% sevoflurane preconditioning in group S1 ＋ OGD and S2 ＋ OGD respectively. In group Z, when the neurons were preconditioned with 4% sevoflurane, ZnPPⅨ was added to the culture medium at the same time, and the other procedures were the same as those in group S2 ＋ OGD. The neuron viability, apoptesis rate, and expression of HO-I protein and mRNA were detected at 24 h of culture. Results Compared with group C, neuron viability was significantly decreased,apoptosis rate was significantly increased, and expression of HO-1 protein and mRNA was up-regulated in group OGD, S1 ＋ OGD, S2 ＋ OGD and Z, expression of HO-1 protein and mRNA was up-regulated in group S1 （ P 〈 0.01 ）, but no significant change was found in neuron viability and apoptosis rate in group S1 （ P 〉 0.05）. Compared with group OGD, neuron viability was significantly increased, apoptosis rate was significantly decreased, and expression of HO-1 protein and mRNA was up-regulated in group S1 ＋ OGD and S2 ＋ OGD （ P 〈 0.01）, but no significant change was found in the indexes mentioned above in group Z （ P 〉 0.05 ）. Neuron viability was significantly higher, apoptosis rate lower and expression of HO-1 protein and mRNA higher in group S2 ＋ OGD than in group S1 ＋ OGD （ P 〈 0.01）. Neuron viability was significantly lower, apoptosis rate higher and expression of HO-1 protein and mRNA lower in group Z than in group S2＋OGD（P〈0.01）.Conclusion HO-1 is involved in the inhibition of OGD-indueed apoptosis in rat hippocampal neurons by sevoflurane preconditioning.