活性氧在热打击诱导神经元细胞凋亡中的作用  

The effect of reactive oxygen species on heat stress-induced neuronal apoptosis

作　　者：刘云松[1] 邓旭斌[2] 苏磊[3] 

机构地区：[1]南方医科大学研究生院,广州军区广州总医院重症医学科,广州510010 [2]南方医科大学肿瘤研究所 [3]广州军区广州总医院重症医学科

出　　处：《中华急诊医学杂志》2014年第6期647-651,共5页Chinese Journal of Emergency Medicine

基　　金：全军十二·五重点课题（BWS12J018）

摘　　要：目的：通过观察热打击诱导活性氧（reactive oxygen species，ROS）爆发性增多对神经元细胞凋亡的影响，探讨重症中暑所致脑损害的发病机理。方法建立神经元细胞热打击模型，对照组将细胞置于标准37℃、5％CO2细胞培养箱，热打击组将细胞置于43℃细胞培养箱中热打击2 h，热打击后继续37℃、5％CO2细胞培养箱孵育，DCFH法检测热打击后0 h、0.5 h、1 h、2 h细胞内ROS含量；Annexin V-FITC/PI双染色方法和Westen blot检测热打击后0 h、3 h、6 h、12 h细胞凋亡率及caspase-3蛋白表达，同时检测ROS特异性清除剂MnTMPyP对热打击后12 h细胞凋亡的影响。结果与对照组比较，43℃热打击后0 h细胞内ROS增加，2 hROS呈爆发性增多（P＜0.05）；43℃热打击后3 h神经元细胞开始出现凋亡，12 h 诱导大量凋亡（43.2％，P＜0.05）、caspase-3蛋白表达明显增加（P＜0.05），其趋势与细胞凋亡趋势一致；MnTMPyP明显抑制了43℃热打击后12 h神经元细胞凋亡率（47.42％降至18.45％，P＜0.05）及caspase-3蛋白的表达。结论 ROS作为上游信号分子介导了热打击诱导的神经元细胞凋亡，但其介导神经元细胞凋亡的中间机制尚需进一步研究。Objective To observe the effect of heat stress-induced burst out of reactive oxygen on neuronal apoptosis and investigate pathogenesis of brain damage caused by severe heat stroke.Methods Neurons heat stress model is set up.Control group were incubated at 37 ℃,5%CO2 ,While heat stress group of cells were incubated at 43 ℃for 2 h,then all the cells were further incubated at 37 ℃for different time as indicated.The amounts of ROS were assayed by DCFH staining at 0 h,0.5 h,1 h,2 h after heat stress.Apoptosis was analyzed by flow cytometry using Annexin V-FITC/PI staining and expression of caspase-3 were determined by westen blotat 0 h、3 h、6 h、12 h after heat stress.In addition,MnTMPyP is the specificscavengers of ROS,which effect on apoptosis is also studied at 12 h after heat stress.Results Compared with control group,amounts of ROS was significant increased at 0 h after heat stress,the burst out of it was at 2 h after heat stress （P〈0.05 ）.Apoptosis was induced at 3h after heat stress ,it was significant increased at 12 h after heat stress （43.2%,P〈0.05 ）.The expression of caspase-3 was also significant increased at 12 h after heat stress （P〈0.05 ）,and its trend was consistent with apoptosis rate trend.In addition,the scavengers MnTMPyP significantly decreased the apoptosis （47.42% to 18.45%, P〈0.05 ）and expression of caspase-3 at 12 h after heat stress.Conclusions An upstream signaling molecules,ROS could mediate heat stress-induced neuronal apoptosis,but its intermediate mechanism needs for further studies.

关 键 词：重症中暑 脑损害 热打击 热打击模型 神经元细胞 凋亡 凋亡机制 活性氧 

分 类 号：R594.12[医药卫生—内科学]