摘要
目的:Id蛋白即分化抑制因子(Id),在人类多种肿瘤中异常表达,并参与肿瘤细胞的增生、分化和凋亡等。文中旨在探讨人分化抑制因子3(Id3)基因体外转染在人肺腺癌细胞顺铂耐药株A549/DDP细胞中的表达及意义。方法:构建人Id3基因和增强型绿色荧光蛋白(EGFP)真核表达载体Id3/pEGFP,采用脂质体介导的转染技术将Id3融合基因(Id3-EGFP)导入A549/DDP细胞,激光共聚焦显微镜观察细胞内EGFP的表达情况;半定量RT-PCR检测Id3表达的改变;MTT法观察Id3对细胞的增殖抑制情况。结果:成功构建人Id3/pEGFP真核表达载体并转染A549/DDP细胞,倒置荧光显微镜下转染的A549/DDP细胞发出强绿色荧光,转Id3/pEGFP质粒的细胞其荧光主要表达于细胞核;RT-PCR结果显示Id3 mRNA在转染后的A549/DDP细胞能有效表达;MTT法结果表明,与对照组相比,转染Id3重组质粒的细胞增殖被抑制。结论:转染的Id3/pEGFP融合基因在A549/DDP细胞能有效表达,外源性Id3基因能抑制该细胞的增殖。
Objective： The Id proteins, also referred to as inhibitors of differentiation, express abnormally in a variety of human tumors and participate in the proliferation, differentiation and apoptosis of tumorous cells. We designed our study to investigate the expression of Id3 in Cisptatin resistant human lung adenocarcinoma A549/DDP cells in vitro and its significance. Methods： We constructed the recombinant eukaryotic expression vector Id3/pEGFP containing human Id3 cDNA and EGFP fusion genes, and transfected them into A549/DDP cells by the liposome-mediated method. After the transfection, we observed the EGFP expression by fluorescent microscopy, detected the alteration of the Id3 expression by semi-quantitative RT-PCR, and determined cell proliferation and inhibition by MTT assay. Results： The human eukaryotic expression vector Id3/pEGFP was successfully constructed and transfected into A549/DDP cells. RT-PCR showed that Id3 mRNA was effectively expressed in the Id3/ pEGFP-transfected A549/DDP cells, and the growth of these cells was significantly inhibited.Conclusion： The transfected Id3/EGFP fusion genes can be effectively expressed in A549/DDP cells, and the exogenous Id3 gene can suppress the proliferation of these ceils.
出处
《医学研究生学报》
CAS
2009年第9期939-943,I0004,共6页
Journal of Medical Postgraduates
基金
江苏省医学重点人才基金资助项目(批准号:RC2007117)
关键词
分化抑制因子3
肺腺癌
A549/DDP
表达
转染
Inhibitor of differentiation 3
Lung adenocarcinoma
A549/DDP
Expression
Transfection
