摘要
目的:探讨HIV-1 Tat蛋白对人外周血B淋巴细胞增殖、凋亡的影响及其机制。方法:采用流式细胞分选术分离HIV阳性患者外周血单核细胞的B淋巴细胞,分别转染pTat或pcDNA3.1各10μg(分别为pTat组与pcDNA3.1组),采用MTT实验检测细胞胞增殖情况,流式细胞术检测凋亡情况,DCHF-DA测定ROS水平,彗星试验检测细胞DNA损伤情况。结果:pTat组转染24h、48h的细胞增殖抑制率、细胞凋亡率及线粒体ROS水平均显著高于pcDNA3.1组(P<0.05)。pcDNA3.1组细胞的DNA大部分呈圆形荧光团,无拖尾现象;pTat组的细胞DNA拖尾现象,呈现典型彗星图像。与pcDNA3.1组相比,pTat组细胞DNA尾长、尾部DNA比例均显著增加(P <0.05)。结论:HIV-1 Tat蛋白可能通过增加线粒体ROS产生,诱导DNA损伤,进而抑制人外周血B淋巴细胞增殖并促进其凋亡。
Objective： To investigate the effect of HⅣ-1 Tat protein on the proliferation and apoptosis of human peripheral blood B lymphocytes and its mechanism. Methods： B lymphocytes from peripheral blood mononuclear cells of HⅣ-positive patients were isolated by flow cytometry and transfected with 10 μg of pTat or pcDNA3.1（pTat and pcDNA3.1, respectively）. Cell proliferation was detected by MTT assay, cell apoptosis was detected by flow cytometry, ROS levels was determined by DCHF-DA, and DNA damage was showed by comet assay. Results： The cell proliferation inhibition rate, apoptosis rate and mitochondrial ROS level in the pTat group transfected for 24 h and 48 h were significantly higher than those in the pcDNA3.1 group（P0.05）. Most of the DNA in the pcDNA3.1 group showed a circular fluorophore with no tailing; the DNA tailing phenomenon of pTat group showed a typical comet image.Compared with the pcDNA3.1 group, the DNA tail length and tail DNA ratio of pTat group were significantly increased（P〈0.05）.Conclusion： HⅣ-1 Tat protein may induce DNA damage by increasing mitochondrial ROS production, thereby inhibiting the proliferation of human peripheral blood B lymphocytes and promoting their apoptosis.
作者
索晓东
彭海
党英男
赵远
田登
SUO Xiao-dong;PENG Ha;DANG Ying-nan;ZHAO Yuan;TIAN Deng(Business Department,Qinghai blood center,Xining,Qinghai,810000,China;Clinical laboratory,Qinghai provincial people＇s hospital,Xining,Oinghai,810007,China;Clinical laboratory,Qinghai blood center,Xining,Qinghai 810000,China;Composition Department,Qinghai blood center,Xining,Qinghai,810000,China;Virus department,Inspection and Testing Center of Qinghai Center for Disease Prevention and Control,Xining,Qinghai,810007,China)
出处
《现代生物医学进展》
CAS
2018年第20期3845-3848,3944,共5页
Progress in Modern Biomedicine
基金
青海省重点科技攻关项目(9632009Y0062)
