根皮素对神经酰胺诱导小鼠神经元parthanatos的影响  

Effect of phloretin on ceramide-induced parthanatos in mice neurons

作　　者：马耀华[1] 邓文静[1] 秦进 牛瑞娜[1] 师云波[3] 荆婧[1] 范成河[3] MA Yaohua;DENG Wenjing;QIN Jin;NIU Ruina;SHI Yunbo;JING Jing;FAN Chenghe(Department of Neurointensive Care Unit,the First Affiliated Hospital,Zhengzhou University,Zhengzhou 450052;Department of Digestive Endoscopy Center,the First Affiliated Hospital,Zhengzhou University,Zhengzhou 450052;Department of Neurology,the First Affiliated Hospital,Zhengzhou University,Zhengzhou 450052)

机构地区：[1]郑州大学第一附属医院神经重症科,郑州450052 [2]郑州大学第一附属医院消化内镜中心,郑州450052 [3]郑州大学第一附属医院神经内科,郑州450052

出　　处：《郑州大学学报（医学版）》2023年第4期459-463,共5页Journal of Zhengzhou University(Medical Sciences)

基　　金：国家自然科学基金项目(81801167);河南省医学科技攻关计划项目(2018020129)。

摘　　要：目的:探讨根皮素对神经酰胺诱导小鼠神经元parthanatos的影响。方法:分离培养C57BL/6J胎鼠原代皮层神经元,分为对照组、神经酰胺组、根皮素组和根皮素+神经酰胺组。神经酰胺组和根皮素组分别用终浓度为20μmol/L的神经酰胺和根皮素处理24 h;根皮素+神经酰胺组先用终浓度为20μmol/L的根皮素处理24 h后,再用终浓度为20μmol/L的神经酰胺处理24 h;对照组用等体积的DMSO处理。采用CCK-8检测神经元活力,Hoechst33342/PI双染检测神经元的死亡,DCFH-DA探针检测细胞内活性氧(ROS)水平,Western blot检测多腺苷二磷酸核糖(PAR)聚合物和DNA损伤标志物γH2AX表达水平,彗星实验检测DNA损伤,细胞免疫荧光法检测细胞凋亡诱导因子(AIF)核转位。结果:神经酰胺处理后,神经元活力降低,神经元死亡率和ROS水平升高;根皮素+神经酰胺处理后,神经元活力升高,神经元死亡率和ROS水平降低(P<0.05);神经酰胺处理后,γH2AX蛋白表达水平升高,神经元拖尾长度和拖尾中的DNA百分比升高;根皮素+神经酰胺处理后,γH2AX蛋白表达水平降低,神经元拖尾长度和拖尾中的DNA百分比降低(P<0.05)。神经酰胺处理后,PAR蛋白表达水平升高,AIF发生核转位;根皮素+神经酰胺处理后,PAR蛋白表达水平降低,AIF核转位受到抑制(P<0.05)。结论:根皮素可能通过抑制氧化应激,进而抑制神经酰胺诱导的神经元发生parthanatos。Aim:To investigate the effect of phloretin on parthanatos induced by ceramide in mice.Methods:Primary cortical neurons from C57BL/6J embryonic mice were isolated,cultured and divided into control group,ceramide group,phloretin group,and phloretin+ceramide group.The cells were treated by final concentration of 20μmol/L ceramide or phloretin for 24 hours in ceramide group or phloretin group,and the cells in ceramide+phloretin group were treated by 20μmol/L ceramide for 24 hours first and then phloretin for 24 hours.Neuronal viability was determined by CCK-8 assay,neuronal death was studied by Hoechst33342/PI double staining assay,PAR polymer andγH2AX(DNA damage marker)expression levels were determined by Western blot,comet assay was used for detecting DNA damage,cellular immunofluorescence was used for detection of nuclear translocation of cell apoptosis inducing factor(AIF),and DCFH-DA probe was used for detecting intracellular active oxygen(ROS).Results:After ceramide treatment,neuronal activity decreased,neuronal death increased,the expression levels ofγH2AX and PAR protein increased,neuronal tail length and DNA percentage in the tail increased,and AIF nuclear translocation occured(P<0.05);after ceramide+phloretin treatment,neuronal activity increased,neuronal death decreased,the expression levels ofγH2AX and PAR protein decreased,neuronal tail length and DNA percentage in the tail decreased,and AIF nuclear translocation was suppressed(P<0.05).Conclusion:Phloretin may inhibit ceramide-induced parthanatos by decreasing oxidative stress in primary mice neurons.

关 键 词：根皮素 神经酰胺 神经元 parthanatos 小鼠 

分 类 号：R741[医药卫生—神经病学与精神病学]