Ⅰ型单纯疱疹病毒胸苷激酶基因转染人卵巢癌细胞株后联合抗肿瘤药物诱导该细胞株凋亡的研究  

Adenovirus - mediated Herpes Simplex Virus Thymidine Kinase Gene Transference and Combined Drug Therapy Leads to Opoptosis of Human Epithelial Ovarian Cancer Cells

作　　者：姜洁[1] 孔北华[1] 江森[1] 刘春生[2] 

机构地区：[1]山东医科大学附属医院妇产科,济南250012 [2]山东医科大学附属医院肿瘤中心,济南250012

出　　处：《中华妇产科杂志》2000年第12期736-739,I015,共5页Chinese Journal of Obstetrics and Gynecology

基　　金：山东省优秀中青年科学家奖励基金!资助 (基金编号 :96 35 42 19)

摘　　要：目的　用复制缺陷型腺病毒载体将Ⅰ型单纯疱疹病毒胸苷激酶基因 (HSV1 tk)转染入卵巢上皮性癌细胞株TYK中 ,观察抗肿瘤药物核苷类似物羟甲基无环鸟苷 (GCV)和无环鸟苷 (ACV)对带有HSV1 tk基因的TYK细胞的体外杀伤作用及是否能诱导该细胞凋亡。方法　常规方法扩增、纯化重组腺病毒 ,聚合酶链反应 (PCR)扩增检测HSV1 tk基因的表达 ,测定重组腺病毒的感染效率。观察腺病毒感染后 ,GCV、ACV对带有HSV1 tk基因的TYK细胞的杀伤作用 ,DNA片段分析及电子显微镜观察GCV作用后TYK细胞形态及细胞凋亡。结果　病毒大量扩增、纯化。PCR扩增可见长度为40 4bp的特异性条带。当感染复数为 10 0时 ,腺病毒的感染效率为 98 9%。GCV、ACV的浓度一定时 ,随着感染复数的增高 ,细胞生长抑制率升高 ;而感染复数一定时 ,随着GCV、ACV浓度的增加 ,细胞生长抑制率亦有明显的升高。电子显微镜观察及DNA片段分析可见 ,GCV作用后带有HSV1 tk基因的TYK细胞发生细胞凋亡。结论　复制缺陷型腺病毒载体可有效地将HSV1 tk基因导入TYK细胞中 ,GCV、ACV对带有HSV1 tk基因的TYK细胞有杀伤作用 ,杀伤的机理可能是细胞凋亡。Objective To evaluate the treatment and apoptosis effect of adenovirus-mediated herpes simplex virus thymidine kinase (HSV 1-tk) gene transference followed by administration of ganciclovir (GCV) and acyclovir (ACV) on ovarian epithelial cancer cells. Methods Recombinant adenovirus was amplificated and purified by routine method. The expression of HSV 1-tk gene was assayed by polymerase chain reaction (PCR). The efficiency of recombinant Advtk transference was evaluated. The cytotoxicity efficacy of TYK cells that carry HSV 1-tk gene was evaluated followed by GCV、ACV administration after the transference of Advtk. The changes and apoptosis of TYK cells that carry HSV 1-tk gene were observed by means of analysis of DNA fragmentation and electronic microscopy. Results Adenovirus was amplificated and purified in large amount. PCR assay showed 404 bp special band. When the multiplicities of infection (MOI) was 100, the transduction rate was 98 9%. The inhibition rates of TYK cells that carry HSV 1-tk gene increased with the increase of MOI when the same concentration of GCV、 ACV were given. When the MOI was same, the inhibition rates were also increased with the increase concentration of GCV、ACV. Apoptosis of TYK cells that carry HSV 1-tk gene after administration of GCV was observed by means of analysis of DNA fragmentation and electronic microscopy. Conclusions The HSV 1-tk gene can effectively transfered into TYK cells by recombinant replicated-deficient adenovirus vector, GCV、ACV can effectively kill TYK cells that carry HSV 1-tk gene in vitro. Apoptosis may be the mechanism of the killing effect.

关 键 词：胸苷激酶 卵巢肿瘤 I型单单纯疱疹病毒 细胞凋亡 

分 类 号：R737.31[医药卫生—肿瘤]