人巨细胞病毒感染对人脐静脉内皮细胞周期和凋亡的影响 动脉粥样硬化发生的可能病因  

Effect of human cytomegalovirus infection on the cell cycle and apoptosis of human umbilical vein endothelial cell Possible etiological factors for the occurrence of atherosclerosis

作　　者：黄晨[1] 张荣怀[1] 王晓明[1] 郭海涛[2] 

机构地区：[1]解放军第四军医大学西京医院老年病二科 [2]解放军第四军医大学基础部生理学教研室,陕西省西安市710032

出　　处：《中国临床康复》2005年第27期56-58,共3页Chinese Journal of Clinical Rehabilitation

基　　金：陕西省科学技术研究发展计划项目(2001K10-G7)~~

摘　　要：目的:探讨人巨细胞病毒感染对人脐静脉内皮细胞细胞周期和细胞凋亡的影响。方法:实验于2003-06/2004/06在解放军第四军医大学老年病二科和解放军第四军医大学基础部生理学教研室完成。①细胞来源及分组:将液氮中冻存的人巨细胞病毒AD169株,用人胚肺成纤维细胞进行增殖培养6～8代;将人脐静脉内皮细胞消化成单细胞悬液,用DMEM培养基重新悬浮,细胞以一传三种于培养瓶继续培养至细胞贴壁并已伸展但未汇合,用台盼蓝拒染法检查活细胞数大于95%,对照组为空白对照,实验组接种人巨细胞病毒169病毒株。②组织学及实验室检查:凋亡的人脐静脉内皮细胞形态学特点在倒置相差显微镜下观察。两组人脐静脉内皮细胞增殖情况采用四甲基偶氮唑蓝比色法,以酶联免疫检测仪测得的A490值表示。两组人脐静脉内皮细胞的细胞周期变化采用流式细胞技术(在荧光激活细胞分选仪上进行细胞周期分析);检测两组人脐静脉内皮细胞的细胞凋亡情况以Annex-inⅤ染色法的以荧光强度判定。结果:①凋亡的人脐静脉内皮细胞形态学特点:正常的生长融合的人脐静脉内皮细胞呈铺路石状镶嵌排列,细胞边界清楚,胞浆丰富,胞核椭圆居中,核仁明显;凋亡的细胞有的变成大细胞。②人巨细胞病毒对人脐静脉内皮细胞增殖的影响:人巨细胞病毒感染人脐静脉内皮细胞后24h,对照组与人巨细胞病毒组A490值分别为1.09±0.08与2.08±0.09(t=7.96,P<0.05)。③人巨细胞病毒对人脐静脉内皮细胞细胞周期的影响:正常对照组G0/G1期的细胞为74.4%,加入人巨细胞病毒后为59.3%,较正常对照组降低20.3%(P<0.05)。④人巨细胞病毒对人脐静脉内皮凋亡的影响:正常对照组凋亡细胞为8.3%,当加入人巨细胞病毒后为5.8%,较对照组降低30.1%(P<0.01)。结论:人巨细胞病毒感染人脐静脉内皮细胞后,引起细胞生长周期和凋亡水平的改AIM： To investigate the influence of human cytomegalovirus infection on the cell cycle and apoptosis of human umbilical vein endothelial cell. METHODS： The experiment was carried out in the Second Department of Geriatrics, and the physiological staff room of the Department of Basic Medicine, the Fourth Military Medical University of Chinese PLA between from June 2003 to June 2004. ① Cell sources and grouping： The human cytomegalovirus AD169 strain, which was freezingly kept in liquid nitrogen, was proliferated and cultured with human embryonic lung fibroblasts for 6 to 8 passages; Human umbilical vein endothelial cells were digested to monocyte suspension, and suspended again with DMEM culture medium; The cells were inoculated to culture flask with one passed to three, and continued to cultured till adhesion and extension but no confluence, and the detection of trypan blue staining exclusion showed that more than 95% of the cells survived; The control group was blank control, human cytomegalovirus 169 strain was inoculated in the experimental group. ② Histological and laboratory examinations： The morphological characteristics of the apoptic human umbilical vein endothelial cells were observed under inverted phase contrast microscope; The proliferation of the human umbilical vein endothelial cells in both groups were detected with methyl-thiazoltetrazolium colorimetry, and expressed with the A490 value measured with enzyme-linked immune detection meter;, The changes of cell cycle of haman umbilical vein endothelial cells in both groups were detected with the flow cytometry, and cell cycle was analyzed with the fuorescent activated cell sorter; The apoptosis of human umbilical vein endothelial cells in both groups was judged with the fluorescent intensity detected by Annexin V staining. RESULTS： ① The morphological characteristics of the apoptic human umbilical vein endothelial cells： The normally grown and confluent human umbilical vein endothelial cells showed mosaic arrangement like cobblestone

关 键 词：巨细胞病毒感染 脐静脉 细胞学 细胞周期 细胞凋亡 

分 类 号：R543.3[医药卫生—心血管疾病]