氢分子对H_2O_2体外诱导氧化损伤大鼠脊髓神经元的保护作用  

Protective effect of molecular hydrogen against H_2O_2-induced oxidative injury in cultured rat spinal cord neurons

作　　者：刘芳婷[1] 李建[1] 李向南[1] 徐盛眀 向正华[3] 袁红斌[1] 

机构地区：[1]第二军医大学长征医院麻醉科,上海200003 [2]第二军医大学长征医院骨科,上海200003 [3]第二军医大学基础部神经生物学教研室,上海200433

出　　处：《第二军医大学学报》2014年第3期233-239,共7页Academic Journal of Second Military Medical University

基　　金：国家自然科学基金(81200953);全军医学科技"十二五"科研项目(CWS11J121)~~

摘　　要：目的证明氢分子对过氧化氢(H2O2)诱导氧化损伤的大鼠脊髓神经元具有保护效应,并初步探讨其相关机制。方法将纯化培养7d后的大鼠脊髓神经元分为4组:(1)常规培养液(NM)对照组:以NM培养;(2)富氢培养液(HM)组:以HM培养;(3)NM+H2O2组:以NM培养2h后,加入100μmol/L H2O2和15μmol/L FeCl2继续培养;(4)HM+H2O2组:以HM培养2h后,加入100μmol/L H2O2和15μmol/L FeCl2继续培养。各组处理后每6h更换各自培养液及氧化剂,在12h后停止处理并检测神经元胞内活性氧(ROS)生成的水平、细胞凋亡情况,以及糖元合成激酶3β(GSK-3β)和p-GSK-3β的表达水平。结果与NM相比,HM可降低H2O2氧化损伤后神经元胞内ROS尤其是羟自由基(HO·)的生成(P<0.01),减少神经元凋亡的数量(P<0.01),下调caspase-3的表达(P<0.01),促进GSK-3β的磷酸化(P<0.01)。结论氢分子对H2O2氧化损伤的神经元具有保护效应,其机制与降低神经元胞内ROS尤其是HO·的生成、减少神经元凋亡的数量和抑制凋亡信号通路的激活以及促进GSK-3β的磷酸化利于神经元生长有关。Objective To investigate the protective effect of molecular hydrogen against hydrogen peroxide(H2O2)-induced oxidative injury in primary cultures of rat spinal cord neurons and the related mechanism.Methods Rat spinal cord neurons were cultured for 7days and were randomly assigned to four groups with different treatments-(1)Normal medium(NM)control:treated with NM;(2)Hydrogen-rich medium(HM):treated with HM;(3)NM+H2O2:treated with 100 μmol/L H2O2and 15μmol/L ferrous chloride(FeCl2)after pretreatment with NM for 2h;and(4)HM+H2O2:treated with 100μmol/L H2O2and 15μmol/L FeCl2after pretreatment with HM for 2h.The respective media were changed every 6hin each group,and 12hlater the cells were collected for assays of reactive oxygen species(ROS),hydroxyl radical(HO·), apoptosis,and the expression of glycogen synthase kinase-3β(GSK-3β)and p-GSK-3β.Results Compared with NM,HM significantly reduced the H2O2-induced intracellular production of ROS and HO·in purified neurons(P<0.01),significantly decreased the number of apoptotic neurons(P<0.01)and expression of caspase-3(P<0.01),and significantly promoted phosphorylation of GSK-3β(P<0.01).Conclusion Molecular hydrogen has protective effect against H2O2-induced oxidative injury in primary cultured neurons.The mechanisms might be related to reduction of intracellular production of ROS and HO·,inhibition of apoptosis in neurons and promotion of GSK-3βphosphorylation.

关 键 词：氧化损伤 过氧化氢 活性氧 氢 神经元 细胞凋亡 糖原合成激酶3β 

分 类 号：R744.9[医药卫生—神经病学与精神病学] R965.1[医药卫生—临床医学]