关键词: 自杀基因, 胞嘧啶脱氨酶基因, 基因治疗, 舌肿瘤, 腺病毒

Abstract:

Objective:The purpose of this investigation was to study the effects of replication-defective recombinant adenovirus-mediated cytosine deaminase (AdCMVCD) /5-fluorocytosine (5-FC) suicide gene system on tongue carcinoma cells.Methods:AdCMVCD was propagated in 293 tongue carcinoma cells, purified by cesium chloride gradient and titrated using tissue culture infectious dos- es-50 (TCID-50). Transmission electron microscope was used to observe the adenovirus transferred into tongue carcinoma cells (Tca8113 cell line), and reverse transcription polymerase chain reaction (RT-PCR) was performed to detect the expression of CD after infection of Tca8113 cells. The killing effects and bystander effects of AdCMVCD/5-FC system on tongue carcinoma cells were evaluated using MTTassay. The changes of cell cycles after treated with AdCMVCD/5-FC system were monitored with the flow cytometry (FCM).Results:The titration of adenovirus (AdCMVCD) was 613@109/ml plaque forming units (PFUs). Under the transmission electron microscope, the adenoviral particles were found in the nucleus of tongue carcinoma cells. The recombi- nant adenovirus expressed CD after infection of Tca8113 cells in vitro. AdCMVCD/5-FC had strong killing-effects on Tca8113 cells, and these effectswere 5-FC dose-dependent. The maximum killing efficacywas 97%. The bystander effectof this system in Tca8113 cells was good. 58% of cells were killed only when 1215% of cells were infected with AdCMVCD. Compared with the control groups,the cell treated by the AdCMVCD/5-FC accumulated in the S phase of the cell cycle (P<01001) and dropped to zero in the G2+Mphase (P<01001) and didnct change in the G1/G0phase (P>0105).Conclusion:The killing effects and the bystander effects ofAdCMVCD/5-FC system on tongue carcinoma cells are satisfying, and the AdCMVCD/5-FC system targeted on tongue carcinoma cells in the S phase.

Key words: suicidegene, cytosinedeaminase, genetherapy, tonguetumor, adenoviru