^(60)Coγ射线照射对人脐静脉内皮细胞线粒体结构和功能的影响  

Effect of(60)^Coγ-ray irradiation on mitochondrial structure and function of human umbilical vein endothelial cells

作　　者：闫凯欣 赵亚伟 王溢豪 易静 段晗 陶宁 王华 胡舜英[1] YAN Kai-xin;ZHAO Ya-wei;WANG Yi-hao;YI Jing;DUAN Han;TAO Ning;WANG Hua;HU Shun-ying(Department of Cardiology,Chinese PLA General Hospital,Beijing 100853,China;Institute of Radiation Medicine,Academy of Military Medical Sciences,Beijing 100850,China;College of Life Sciences,Anhui Medical University,Hefei 230032,China)

机构地区：[1]解放军总医院心血管病医学部,北京100853 [2]军事科学院军事医学研究院辐射医学研究所,北京100850 [3]安徽医科大学生命科学学院,安徽合肥230032

出　　处：《中国药理学与毒理学杂志》2022年第9期657-664,共8页Chinese Journal of Pharmacology and Toxicology

基　　金：国家自然科学基金(82173450)。

摘　　要：目的探讨^(60)Coγ射线照射对血管内皮细胞线粒体结构和功能的影响。方法分别采用^(60)Coγ射线0(细胞对照组),5,10和20 Gy单次照射人脐静脉内皮细胞(HUVEC),照射后24和48 h,采用流式细胞术检测细胞凋亡;荧光探针H2DCF-DA标记,用激光共聚焦显微镜检测细胞产生活性氧(ROS)水平;JC-1探针标记,分别用流式细胞术和激光共聚焦显微镜检测细胞线粒体膜电位;通过钙黄绿素-AM探针标记,激光共聚焦显微镜定性检测线粒体膜通道孔(mPTP)开放状态;电镜观察线粒体结构变化;Western印迹法检测线粒体分裂相关标志物磷酸化动力相关蛋白1(p-Drp1)和磷酸化线粒体分裂因子(p-Mff)蛋白表达水平。结果与细胞对照组相比,照射后24和48 h,5,10和20 Gy组细胞凋亡率增加(P<0.01),细胞ROS水平增加(P<0.05,P<0.01),细胞mPTP开放增加(P<0.05,P<0.01)。流式细胞术和荧光显微镜检测结果显示,与细胞对照组相比,照射后24和48 h,5,10和20 Gy组线粒体膜电位下降(P<0.01)。电镜观察显示,与细胞对照组相比,照射组照射后48 h,单个线粒体面积增加(P<0.05,P<0.01),单个线粒体周长增加(P<0.05),线粒体密度下降(P<0.05,P<0.01),异常线粒体数量大幅增加(P<0.01)。与细胞对照组相比,各照射组照射后48 h,p-Drp1和p-Mff蛋白表达水平均显著增加(P<0.05,P<0.01)。结论^(60)Coγ射线照射加重血管内皮细胞线粒体损伤,促进线粒体分裂,诱发线粒体功能障碍。OBJECTIVE To investigate the effect of ^(60)Coγ-ray irradiation on mitochondrial structure and function of vascular endothelial cells.METHODS Human umbilical vein endothelial cells(HUVECs)were irradiated with ^(60)Coγ-ray at 0(cell control),5,10 and 20 Gy,respectively.The apoptosis of HUVEC cells was detected by apoptosis kit at 24 and 48 h after irradiation,the reactive oxygen species(ROS)levels by laser confocal microscopy after H2DCF-DA labeling,the mitochondrial membrane potential by flow cytometry and laser confocal microscopy after JC-1 probe labeling,and the open state of the mitochondrial permeability transition pore(mPTP)qualitatively by laser confocal microscopy through calcein-AM probe labeling.The structural changes of mitochondria were observed by electron microscopy.The mitochondrial division-related markers,phosphorylated power-related protein 1(p-Drp1)and p-mitochondrial fission factor(p-Mff)protein expressions were detected by Western blotting.RESULTS Compared with the cell control group(0 Gy),the cell apoptosis was increased(P<0.01),ROS levels were increased(P<0.05,P<0.01)and opening of mPTP was increased(P<0.05,P<0.01)in the 5,10and 20 Gy groups 24 and 48 h after irradiation.Flow cytometry and fluorescence microscopy results showed a decrease in mitochondrial membrane potential in the 5,10 and 20 Gy groups at 24 and 48 h after irradiation compared with the cell control group(P<0.01).Electron microscopic observations showed that the irradiated group showed an increase in individual mitochondrial areas(P<0.05,P<0.01)and in individual mitochondrial perimeters(P<0.05),a decrease in mitochondrial density(P<0.05,P<0.01),and a significant increase in the number of abnormal mitochondria(P<0.01)48 h after irradiation compared with the cell control group.Compared with the cell control group,the expression levels of p-Drp1 and p-Mff proteins were significantly increased in the irradiated groups 48 h after irradiation(P<0.05,P<0.01).CONCLUSION ^(60)Coγ-ray irradiation aggravates mitochondrial damage in

关 键 词：Γ射线 血管内皮细胞 线粒体功能 线粒体分裂 

分 类 号：R965[医药卫生—药理学] R818.03[医药卫生—药学]