慢病毒介导的CAV1过表达对HL-60细胞增殖与凋亡的影响  

Effect of Overexpression of CAV1Mediated by Lentivirus on Proliferation and Apoptosis of HL-60 Cells

作　　者：马微[1,2] 王迪迪[1] 王昭[1] 朱贵明[1] 张鹏霞[1] 

机构地区：[1]佳木斯大学基础医学院生化实验室,黑龙江佳木斯154007 [2]牡丹江医学院研究生处,黑龙江牡丹江157011

出　　处：《中国实验血液学杂志》2013年第4期905-910,共6页Journal of Experimental Hematology

基　　金：国家自然基金项目(编号81070428);黑龙江省自然科学基金项目(编号D201024);黑龙江省卫生厅科研课题项目(编号2010-484);佳木斯大学重点科研项目(编号2010-228);黑龙江省博士后科研启动项目(LBH-Q12005)

摘　　要：本研究旨在探讨慢病毒介导的CAV1过表达对HL-60细胞增殖与凋亡的影响。构建慢病毒重组表达载体pcDNA-EF1-CAV1,并与pPACK包装质粒混合物共转染至293TN细胞后,收集病毒液感染HL-60细胞,使CAV1基因在细胞中稳定转染并高表达。采用Western blot方法评价转染后HL-60 CAV1蛋白表达情况。采用CCK-8法、流式细胞术分别检测转染前后HL-60细胞的增殖活性和凋亡情况。结果表明:PCR阳性克隆筛选及核苷酸测序结果证实CAV1基因正确插入表达载体pcDNA-EF1-GFP中。重组慢病毒颗粒Lv-CAV1成功转染HL-60细胞,转染率达90%。Western blot检测结果显示,转染后48 h HL-60细胞中CAV1蛋白表达明显增高。CCK-8检测结果显示转染后48 h HL-60细胞增殖活性降低(P<0.05),流式细胞仪检测结果显示转染后HL-60细胞凋亡率明显增加(P<0.01)。结论:CAV1过表达对HL-60细胞具有抑制细胞的增殖活性、促进细胞凋亡的作用。This study was purposed to explore the effect of lentivirus-mediated CAV1 overexpression on proliferation and apoptosis in HL-60 cells.Recombinant lentiviral expression vector pcDNA-EF1-CAV1 was constructed,and cotransfected the 293TN cells with a mixture of pPACK packaging plasmids.Then collecting virus suspension infects the HL-60 cells,which make CAV1 gene stable transfection and high expression in the cells.The CAV1 protein expression status in HL-60 cells transfected was elvaluated through Western blot method.Proliferative activity and apoptosis of HL-60 cells before and after transfection were detected by CCK-8 method and flow cytometry,respectively.The results showed that the PCR-positive clone screening and results of nucleotide sequencing confirmed that the CAV1 gene inserted into the expression vector pcDNA-EF1-GFP correctly,recombinant lentiviral particles Lv-CAV1 transfected HL-60 cells successfully and with transfection rate up to 90%.The result of Western blot showed that CAV1 protein expression in HL-60 cells significantly increased at 48 hours after transfection.CCK-8 result indicated that cell proliferation activity increased at 48 h after transfection（P〈0.05）,flow cytometry testing results displayed that apoptosis rate of HL-60 cells obvirously decreased after transfection（P〈0.01）.It is concluded that the overexpression of CAV1 in HL-60 cells can inhibit cell proliferation activity and promote cell apoptosis.

关 键 词：CAV1 HL-60 慢病毒 细胞增殖 细胞凋亡 

分 类 号：R733.7[医药卫生—肿瘤] R979.1[医药卫生—临床医学]