脂质体介导E1A-TK基因转染人肺腺癌细胞A549体外研究

Study on E1A-TK Gene Transfected into Human Pulmonary Adenocarcinoma Cell A549 by Lipofectamine in vitro

目的用脂质体转染真核表达质粒PcDNA3E1A-TK入人肺腺癌细胞A549,观察丙氧鸟苷(GCV)、X射线及二者联合对转染细胞的体外杀伤作用。方法用阳离子脂质体介导真核表达质粒PcDNA3E1A-TK转染人肺腺癌细胞A549,PCR、RT-PCR方法分别检测转染细胞中E1A-TK基因DNA整合、mRNA表达。MTT法测不同浓度GCV、不同剂量X射线及一定剂量X线照射后再给一定量GCV对转染细胞的生长抑制率。结果PCR、RT-PCR结果表明转染细胞有E1A-TK基因整合、mRNA表达。根据IC50,转染细胞对GCV、X射线的敏感性分别比亲代细胞提高111.3倍和2.9倍,在作用相同时间后,GCV+X线照射治疗对转染细胞的生长抑制作用比单纯GCV或X线治疗明显增强(P<0.01)。结论脂质体介导外源基因E1A-TK成功转入人肺腺癌A549细胞并获得表达;转染细胞获得了对GCV和X线的敏感性,GCV和X线对转染细胞有协同杀伤作用。

Objective To transfect human puhnonary adenocarcinoma cells A549 with the eucaryotic expression plasmids pcDNA3E1A-TK by lipofectamine, and to determine the sensitivity of the transfected cells to the cytotoxie actions of ganeiclovir（GCV） and X-ray. Methods Eucaryotic expression plasmids containing E1A-TK gene were transfected into human pulmonary adenocarcinoma cells A549 by lipofeetamine. PCR and RT-PCR were used to examine E1A-TK gene DNA integration and mRNA expression in the transfected cells, respectively. GCV, X-ray or GCV ＋ X-ray induced growth inhibition of the transfeeted cells were determined by MTr cytotoxicity assay. Results E1A-TK gene DNA integration and mRNA expression were found in the transfected cells. The IC50 of GCV and X-ray on the transfected cells was increased 113.3 and 2.9 folds, respectively. GCV and X-ray acted synergistically to increase the cell death of the transfeeted cells. Conclusion E1A-TK gene was successfully transfected into human pulmonary adenocarcinoma cells A549 by lipofectamine. The gene was successfully expressed and conferred sensitivity of the transfeeted cells to GCV and X-ray. GCV plus X-ray exhibited synergistic killing effect on the transfeeted ceils.