新城疫病毒HN基因诱导人肺癌细胞SPC-A1凋亡的作用机制  

Mechanisms of Apoptosis in Human Lung Carcinoma Cells SPC-A1 Induced with Hemagglutinin-neuraminidase(HN) Gene from Newcastle Disease Virus

作　　者：连海[1] 金宁一[1] 李霄[1] 陈立刚[1] 张静敏[1] 管国芳[1] 孙丽丽[1] 李雪梅[1] 郑洪玲[1] 崔英姬[2] 

机构地区：[1]解放军军事医学科学院解放军基因工程重点实验室 [2]吉林大学农学部图书馆,长春130062

出　　处：《中国生物化学与分子生物学报》2006年第3期222-227,共6页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家重点基础研究发展规划(973计划)资助项目(No.G1999011902)~~

摘　　要：为了探索新城疫病毒血凝素神经氨酸酶(HN)基因对人肺癌细胞SPC-A1的作用及机制,将含有HN基因的重组质粒pVHN经脂质体介导转染人肺癌细胞SPC-A1,通过MTT方法检测细胞活力;采用吖啶橙/溴化乙锭(AO/EB)染色分析肿瘤细胞凋亡;罗丹明123和DCFA染色测定线粒体跨膜电位(ΔΨm)和活性氧水平变化;流式细胞仪分析MHC-Ⅰ分子表达;底物染色反应检测caspase-3活性.结果重组质粒pVHN转染人肺癌细胞SPC-A148h后,细胞活力明显降低;AO/EB染色可见明显的细胞凋亡形态学变化;与空质粒对照相比,线粒体ΔΨm下降(P<0·01),活性氧水平升高(P<0·05);细胞表面MHC-Ⅰ分子表达上调(P>0·05);caspase-3活性增强(P<0·01).以上结果提示,新城疫病毒HN基因能够上调SPC-A1细胞表面MHC-Ⅰ分子表达,并通过上调ROS水平,下调线粒体ΔΨm,进而激活caspase-3,最终诱导人肺癌细胞凋亡.To investigate the effect of hemagglutinin-neuraminidase（HN） gene from Newcastle disease virus on human lung carcinoma cells SPC-A1, the recombinant plasmid pVHN was introduced into human lung carcinoma cells SPC-A1 by liposome-mediated transfection. The viability of SPC-A1 cells was determined by MTT staining and apoptosis was detected by AO/EB staining. The alteration of mitochondrial transmembrane potential and ROS level of the cells was detected by flow cytometry （FCM） with rhodamine 123 and DCFA staining. The expression of MHC-Ⅰ was detected by FCM and the activation of caspase-3 was assayed by its substrate color reaction. Viability of SPC-A1 cells was decreased significantly and recombinant plasmid could lead to obviously morphological apoptotic changes of SPC-A1 cells by introduction of pVHN into SPC-A1 cells for 48 hours. Mitochondrial transmembrane potential was decreased（ P 〈 0.01 ） , ROS level of the cells was increased （ P 〈 0.05）, the expression of MHC-Ⅰ（ P 〉 0.05） and caspase-3 activity （ P 〈 0.01 ） was increased, compared with control cells. The above results showed increase of MHC-Ⅰ expression and significant apoptosis in SPC-A1 cells can be induced by recombinant plasmid pVHN. Apoptosis may be resulted from the increase of ROS level, the decrease of mitochondrial transmembrane potential and activation of Caspase-3.

关 键 词：新城疫病毒 HN基因 人肺癌细胞 凋亡 

分 类 号：Q255[生物学—细胞生物学] Q75