摘要
目的:探讨重组质粒pGL3-hTERT-tk/GCV对胃癌细胞的促凋亡作用。方法:以基因工程方法构建重组质粒pGL3-hTERT-tk和相应的荧光报告质粒pGL3-hTERT-tk-Luc^+;脂质体Lipofectamine^(TM)2000瞬时转染胃癌细胞系SGC-7901并用GCV干预,荧光显微镜观察细胞形态变化和转染效率,TUNEL标记和流式细胞术观察转染后胃癌细胞的凋亡;以上实验均以正常肝细胞L-02为对照。结果:经鉴定,重组质粒pGL3-hTERT-tk中tk片段的长度为1100 bp。荧光素酶标记的阳性、阴性对照及治疗报告质粒pGL3-hTERT-tk-Luc^+均能有效转染高表达端粒酶活性的胃癌细胞SGC-7901,转染效率为(8.2±1.14)%。重组质粒转染胃癌细胞后与GCV共育4 d,细胞的凋亡率为(60.0±1.56)%;被pGL3-hTERT-tk转染的肿瘤细胞细胞周期发生了变化,处于细胞周期早期的细胞大量凋亡,早期凋亡率为(47.1±1.35)%。结论:pGIB-hTERT-tk/GCV对胃癌细胞有强烈的杀伤作用,但不影响正常细胞的生长,有潜在临床应用前景。
Objective： To investigate the pro-apoptosis effect of pGL3-hTERT-tk/GCV on human gastric cancer cells in vitro. Methods： Recombinant plasmid pGL3-hTERT-tk and the corresponding reporter plasmid pGL3-hTERT-tk-Luc ＋ were constructed by gene engineering. The recombinant plasmids were, then used to transiently transfect gastric cancer cells SGC-7901 via LipofectamineTM2000 and was intervened by GCV. Fluorescence microscope was used to observe the changes of cell morphology and the transfection efficiency. Cell apoptosis was examined by TUNEL labeling and the apoptosis rate was determined by flow cytomtry. Normal hepatic cells L02 were used as control in all experiments. Results： The length of tk of therapeutic plasmid pGL3-hTERT-tk was 1 100 bp. pGL3-control-tk-Luc＋, pGL3-basic-tk-Luc＋ and pGL3- hTERT-tk-Luc＋ all could effectively transfect SGC-7901 cells with high telomerase activity, with the transfection rate being （ 8.2 ± 1.14） %. After SGC-7901 was transfected with therapeutic pGL3-hTERT-tk and cultured with GCV for 4 d, the apoptosis rate was （60.0 ± 1.56） % and cell cycle also significantly changed ; more cells at the early stage of cell cycle became apoptotic, with an apoptosis rate of （47. 1± 1.35 ）%. Conclusion： pGL3-hTERT-tk has strong killing effect against gastric cancer cells and has no influence on the growth of normal cells, showing a potential in future clinical application.
出处
《中国肿瘤生物治疗杂志》
CAS
CSCD
2008年第3期233-237,共5页
Chinese Journal of Cancer Biotherapy
基金
国家自然科学基金(No．30672405)
山西省青年科技基金(No.20001028)~~
