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Study on Killing Effects of HSV-tk/GCV System on Murine Liver Cancer Cells in vitro
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Abstract:
Objective: To investigate the killing effects of HSV-tk/GCV system on murine liver cancer cells in vitro. Methods: HSV-tk gene was cloned into retroviral vector (pLNCTK) by recombinant DNA technology. After packaging with PA317 cell line, murine liver cancer cell line MM45T.Li was infected by the recombinant retrovirus. The positive clones were obtained after G418 selection and it was termed MM45T. Li/TK. The integration and expression of tk gene in MM45T. Li/TK cells were identified by PCR and RT-PCR. The killing effects of GCV on TK cell alone or a mixture of TK and TK~(-) cells in different ratio were observed. Results: (1) PCR and RT-PCR analysis confirmed integration of HSV-tk gene in positive clone, and expression of HSV-tk gene at mRNA level. (2) There was no significant difference in cell proliferation or morphology between the MM45T. Li/TK and MM45T. Li. (3) The cytotoxicity of GCV in MM45T. Li/TK cells was 1 000 fold higher sensitive than that in parental MM45T.U cells. (4) When MM45T.Li/TK cells were mixed with parental MM45T. Li cells at a ratio of 25:75, significant bystander effect was observed in vitro after they were treated with nontoxic GCV. Conclusions: These results suggested that murine liver cancer cells were sensitive to HSV-tk/GCV system and had significant bystander effects.
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