摘要
目的 体外观察腺病毒载体介导的单纯疱疹病毒胸苷激酶基因及羟基无环鸟苷系统 (ADV -HSV -Tk/GCV)对人腺样囊性癌细胞 (ACC -2及ACC -M )的杀伤作用。方法 分别将人腺样囊性癌细胞株 (ACC -2及ACC -M )用腺病毒构建的绿色荧光蛋白按 1∶10 ,1∶2 0 ,1∶3 0转染 ,在 12～ 3 6h内用荧光显微镜观察细胞的转染率。MTT法观察ADV -HSV -TK在不同感染复数 (5～ 60 )以及不同浓度GCV处理时对ACC -2及ACC -M的细胞杀伤作用以及“旁观者效应”。结果 体外研究发现 ,ACC -2及ACC -M均能被腺病毒 -绿色荧光蛋白转染。最小感染复数为 10～ 2 0 ,转染率与感染复数呈正相关。在感染复数为 5 ,GCV浓度为 2 5 μg/ml时 ,能够发挥细胞杀伤效应且出现剂量依赖性。MTT法检测发现经ADV -HSV -TK处理后 ,当GCV浓度为 2 5 μg/ml或高于该浓度时ACC -2及ACC -M的细胞抑制率为 40 % -60 % ,而当GCV单独作用时则无此作用。转染ADV -HSV -TK的细胞与未转染细胞按不同比例混合提示“旁观者效应”的存在 ,且转染细胞的比例越高 ,被杀伤的细胞比例越大。结论 腺病毒载体介导的单纯疱疹病毒胸苷激酶基因及羟基无环鸟苷系统 (ADV -HSV -TK/GCV)在体外能够有效抑制腺样囊性癌细胞 ,然而 ,其细胞杀伤效应及“旁观者效应”
Objective To observe the killing effect of Adenovirus vectors encoding herpes simplex virus thymidine kinase gene/ganciclovir(ADV-HSV-TK/GCV) system on human Adenoid Cystic Carcinoma Cells(ACC-2 and ACC-M) in vitro.Methods Both the ACC-2 and ACC-M cell lines were transduced with the ADV-GFP at the proportion of 1∶10,1∶20,1∶30 for 12 to 36 hour. Next, the adenoid cystic carcinoma cell lines which exposure to ADV-HSV-TK at different M.O.I (5 to 60) were treated with ganciclovir (GCV) in different concentrations, while the two cell lines were transferred at an M.O.I of 100 and subsequently mixed with non- transferred cells at varying proportion, the cell killing and also the 'bystander effect' were detected with MTT assay.Results We have demonstrated that both ACC-2 and ACC-M can be transduced with AD-GFP in vitro. The minimal multiplicity of infection (MOI;for the purpose of this study the number of vector genomes per target cell) was 10 to 20,and the gene transduction efficiency was dependent on multiplicity of infection. The system could present the killing activity to both ACC-2 and ACC-M cells in dose-dependent manner at lower concentrations of MOI 5 and GCV(25 ug/ml.).Detection of MTT showed that when exposure to ADV-HSV-TK, the growth of Acc-2 and Acc-M cells were inhibited significantly by GCV when its concentration was equal to or over 25ug/ml, and the rate of inhibition was 40% to 60%, while the two cell lines were not or only slightly inhibited by GCV of any concentrations otherwise. The transferred ADV-HSV-TK cells mixed with un-transferred cells in different proportions revealed the existence of 'bystander effect', the more the transferred cells, the less the living cells existence. Conclusion The ADV-HSV-TK/GCV system is effective in killing ACC-2 and ACC-M in vitro. However, the system is still to be modified to enhance the killing effect and the 'bystander effect'.
出处
《临床口腔医学杂志》
2002年第6期415-418,共4页
Journal of Clinical Stomatology
基金
湖北省卫生厅 (0 1 0 4 - 0 30 3)
