七氟醚对HT22小鼠海马神经元细胞DNA的损伤及其机制  

DNA damage of HT22 mouse hippocampal neuronal cells induced by sevoflurane and its mechanism

作　　者：王雪冬 王迎迎 刘索宁 王靖[4] 刘昊鹏 刘楠[1] 裴爱月[3] 秦晶[1] 冯春生[1] 朴美花[1] WANG Xuedong;WANG Yingying;LIU Suoning;WANG Jing;LIU Haopeng;LIU Nan;PEI Aiyue;QIN Jing;FENG Chunsheng;PIAO Meihua(Department of Anesthesiology,First Hospital,Jilin University,Changchun 130021,China;Department of Gastrointestinal Surgery,First Hospital,Jilin University,Changchun 130021,China;Clinical Medical College,Jilin University,Changchun 130021,China;Department of Endoscope Center,First Hospital,Jilin University,Changchun 130021,China)

机构地区：[1]吉林大学第一医院麻醉科,吉林长春130021 [2]吉林大学第一医院胃结直肠外科,吉林长春130021 [3]吉林大学第一医院内镜中心,吉林长春130021 [4]吉林大学

出　　处：《吉林大学学报（医学版）》2020年第2期240-247,共8页Journal of Jilin University：Medicine Edition

基　　金：国家自然科学基金资助课题(81901076,81771141)。

摘　　要：目的:观察吸入麻醉药七氟醚对HT22小鼠海马神经元细胞DNA的损伤,阐明七氟醚诱导脑神经毒性的可能机制。方法:HT22小鼠海马神经元细胞随机分为空白对照组、2%七氟醚组(2%Sevo 6 h组、2%Sevo 12 h组和2%Sevo 24 h组)、4%七氟醚组(4%Sevo 6 h组、4%Sevo 12 h组和4%Sevo 24 h组)和8%七氟醚组(8%Sevo 6 h组、8%Sevo 12 h组和8%Sevo 24 h组),应用四甲基偶氮唑盐比色(MTT)法和乳酸脱氢酶(LDH)法分别检测各组HT22小鼠海马神经元细胞存活率和死亡率。根据实验结果将细胞分为生理盐水组、生理盐水+4%Sevo 12 h组、生理盐水+8%Sevo 12 h组、N-乙酰半胱氨酸组(NAC组)、NAC+4%Sevo 12 h组和NAC+8%Sevo 12 h组。采用MTT法和LDH法分别检测NAC预处理后各组HT22小鼠海马神经元细胞存活率和死亡率;应用单细胞凝胶电泳检测各组HT22小鼠海马神经元细胞DNA双链断裂情况;Western blotting法检测各组HT22小鼠海马神经元细胞中DNA损伤相关蛋白8-OHdG、ATM、p-ATM和γ-H2AX表达量;DCFH-DA法检测各组HT22小鼠海马神经元细胞中活性氧(ROS)水平。结果:与空白对照组比较,2%Sevo组HT22小鼠海马神经元细胞存活率和死亡率差异无统计学意义(P>0.05);4%Sevo组和8%Sevo组HT22小鼠海马神经元细胞存活率降低(P<0.01),死亡率升高(P<0.01)。与4%Sevo组比较,同一时间8%Sevo组HT22小鼠海马神经元细胞存活率降低(P<0.05),死亡率升高(P<0.05)。与4%Sevo 6 h组比较,4%Sevo 12 h组和4%Sevo 24 h组HT22小鼠海马神经元细胞存活率降低(P<0.05),死亡率升高(P<0.05);与4%Sevo 12 h组比较,4%Sevo 24 h组HT22小鼠海马神经元细胞存活率降低(P<0.05),死亡率升高(P<0.05)。与8%Sevo 6 h组比较,8%Sevo 12 h组和8%Sevo 24 h组HT22小鼠海马神经元细胞存活率降低(P<0.05),死亡率升高(P<0.05);与8%Sevo 12 h组比较,8%Sevo 24 h组HT22小鼠海马神经元细胞存活率降低(P<0.05),死亡率升高(P<0.05)。与生理盐水组比较,生理盐水+4%Sevo 12 h组和生理�Objective:To observe the DNA damage of HT22 mouse hippocampal neuronal cells induced by inhaled anesthetic sevoflurane,and to clarify the possible mechanism of neurotoxicity induced by sevoflurane.Methods:The HT22 mouse hippocampal neuronal cells were randomly divided into blank control group,2%sevoflurane groups(2%Sevo 6 h group,2%Sevo 12 h group,2%Sevo 24 h group),4%sevoflurane groups(4%Sevo 6 h group,4%Sevo 12 h group,4%Sevo 24 h group)and 8%sevoflurane groups(8%Sevo 6 h group,8%Sevo 12 h group,8%Sevo 24 h group).The survival rates and the mortalities of HT22 mouse hippocampal neuronal cells in various groups were measured by MTT method and LDH methods,respectively.According to the experiment results,the cells were divided into normal saline group,normal saline+4%Sevo 12 h group,normal saline+8%Sevo 12 h group,NAC group,NAC+4%Sevo 12 h group,and NAC+8%Sevo 12 h group.MTT method and LDH method were used to detect the survival rates and the mortalities of HT22 mouse hippocampal neuronal cells in various groups after NAC pretreatment;single cell gel electrophoresis was used to detect the DNA double-strand breaks in the HT22 mouse hippocampal neuronal cells in various groups;Western blotting method was used to detect the expression amounts of 8-OHdG,ATM,p-ATM,andγ-H2AX in the HT22 mouse hippocampal neuronal cells in various groups.DCFH-DA method was used to detect the levels of reactive oxygen species(ROS)in the HT22 mouse hippocampal neuronal cells in various groups.Results:Compared with blank control group,there were no significant differences in the survival rates and the mortalities of HT22 mouse hippocampal neuronal cells in 2%Sevo groups(P>0.05);the survival rates of HT22 mouse hippocampal neuronal cells in 4%Sevo groups and 8%Sevo groups were significantly decreased(P<0.01),and the mortalities were significantly increased(P<0.01).Compared with 4%Sevo groups,the survival rates of HT22 mouse hippocampal neuronal cells in 8%Sevo groups at the same time were decreased(P<0.05),and the mortalities were increased

关 键 词：七氟醚 神经元细胞 DNA损伤 神经毒性 细胞死亡 

分 类 号：R614.21[医药卫生—麻醉学]