外源性p16基因对食管癌患者癌细胞系的生长抑制作用

目的 观察外源性p16基因质粒导入食管癌患者食管鳞癌细胞系Ec109后的表达及其对Ec109细胞增殖生长状况的影响。方法 根据转染质粒的不同或是否行p16 cDNA质粒转染分为三组，每组分别为5个样本。PcDNA3-P16转染组（P16转染组）：采用脂质体（Lipofectamine）为载体，将PcDNA3-P16表达质粒寻入Ec109细胞；空载体转染组：用PcDNA－3－neo空载体以上述相同方法转染，作空白对照；未转染组：未用PcDNA3－P16质粒转染，作阴性对照。应用Northern斑点杂交、免疫组织化学、免疫荧光检测转化细胞的P16蛋白变化；流式细胞仪分析细胞周期变化，DNA片段化电泳检测细胞凋亡。结果 P16转染组外源性p16基因在Ec109细胞中表达后，细胞生长速度减慢，克隆形成能力下降，瞬时表达时有细胞凋亡发生，稳定表达后细胞存在G1期阻滞；空载体转染组和未转染组均未观察到以上结果。结论 脂质体介导的外源性p16基因转染对Ec109细胞有生长抑制作用，p16基因瞬时表达可诱导细胞凋亡。

The Growth Suppress Effect of Exogenous p16 Gene on Human Esophageal Carcinoma Cell Lines

Objective To study the effect of transfection exogenous PcDNA3 P16 expression plasmid on proliferation of human esophageal squamous carcinoma cell lines Ec109. Methods According to the different transfected plasmid and whether the transfection were underwent, three group were divided and 5 sample included in each group.P16 transfection group: p16 cDNA was introducted into Ec109 cell line by the vector of Lipofectamine. Empty plasmid transfection: PcDNA3 neo plasmid was introducted with the same method as a blank control group.Non transfection group: the Ec109 cell that were not transfected as a negative control group. Immuno fluorensence microscope and Northern dot hybirdization were used to detect P16 protein status. Methyl tetrazo thialum (MTT) method and flow cytometry were used for analysis of cell cycle and growth inhibition. Cell apoptosis was detected by DNA fragment electrophoresis and flow cytometry. Results The cell vitality of transfected Ec109 cells decreased and the G1 phase arrest detected after PcDNA3 P16 expression plasmid was transfected and expressed in Ec109 cell lines. Cell apoptosis was observed while P16 protein transient expressed, but the other control groups had negative results. Conclusion Transfection exogenous p16 gene to Ec109 conducted by liposome would result in cell growth inhibition. Transient p16 gene expression could induce cell apoptosis.