含胸苷激酶的不同细胞对丙氧鸟苷敏感性的差别及死亡途径初探  

Different susceptibility of cell lines to thymidine kinase/ganciclovir mediated killing effect and the way of cell death

作　　者：张雷[1] 刘彤华[1] 崔全才[1] 高洁[1] 

机构地区：[1]中国医学科学院中国协和医科大学,北京协和医院病理科,100730

出　　处：《中华病理学杂志》2000年第1期34-38,共5页Chinese Journal of Pathology

基　　金：卫生部科学研究基金资助项目!(19981010)

摘　　要：目的　探讨含单纯疱疹病毒胸腺嘧啶核苷激酶 (HSV TK)基因细胞对丙氧鸟苷 (GCV)敏感度不均一的原因以及在GCV作用下细胞被杀伤的途径———坏死或凋亡。方法　将表达HSV TK基因的重组逆转录病毒载体 ,分别直接导入生长速度不同的 3个细胞系 ,胰腺癌细胞系 (PC 2 ,PC 7)和猪肾小管上皮细胞系 (LLC PK1) ,计算转染前后细胞倍增时间 ,MTT法检测 5 0 %细胞生长受抑制时的GCV浓度 (IC5 0 值 ) ;流式细胞术检测细胞周期 ;对GCV作用细胞和 (或 )移植瘤进行光镜和电镜的形态观察及原位凋亡检测。结果　倍增时间分别为 (31.2± 0 .1)、(48.3± 0 .1)、(5 3.9± 0 .1)h的细胞 ,其转染细胞IC5 0 值分别为 (0 .73± 0 .12 )、(0 .93± 0 .16 )、(1.2 2± 0 .0 6 ) μmol L ,呈正相关 ;细胞被阻滞在S期 ;经GCV作用的细胞肿胀崩解坏死 ,个别细胞表现凋亡 ;经GCV作用的移植瘤显示大片坏死。结论　HSV TK GCV系统对增殖旺盛的细胞作用更为显著 ,GCV作用的 3个转染细胞系细胞的死亡途径主要为坏死。Objective To investigate the mechanism of variability of herpes simplex virus (HSV) thymidine kinase (TK)/ganciclovir (GCV) mediated suicide effect obtained in three different cell lines and the ways in which this system mediated cell killing occurs. Methods Recombinant retroviral vector expressing HSV TK was transduced into three cell lines known with different growth rate (PC 2, PC 7 and LLC PK1). The doubling time of the transduced and parental cells was calculated. MTT method was used to detect the concentration of GCV at which cell growth was inhibited by 50% (IC 50 value). Cell cycle was analyzed by flow cytometry. Necrosis or apoptosis of cultured cells and/or xenografts was observed under light and electron microscope, and by in situ apoptosis detection. Results The doubling time of the three parental cell lines was (31.2±0.1) h, (48.3±0 1) h, and (53.9±0 1) h, separately. The IC 50 values of their HSV TK transduced cell lines to GCV were (0 73±0.12) μmol/L, (0.93±0.16) μmol/L and (1.22±0.06) μmol/L, respectively and the IC 50 value was correlated with the cell doubling time. Flow cytometry revealed S arrest. The majority of cells under treatment of GCV displayed swelling and collapse, but very few cells showed apoptosis. Large areas of necrosis were observed in the xenografts. Conclusions The cells with high growth rate are more susceptible to HSV TK/GCV mediated killing effect. Necrosis is the main way in which cells of the three HSV TK transduced cells lines die.

关 键 词：胰腺肿瘤 胸苷激酶 丙氧鸟苷 HSV-TK/GCV 

分 类 号：R363[医药卫生—病理学]