论文部分内容阅读
目的验证Thy-1(CD90)基因与肺癌细胞迁移潜能的关系,并探讨其作用机制。方法利用真核表达载体pcDNA3.0构建包含Thy-1基因全长序列的重组表达质粒载体,转染A549细胞,G418筛选获得稳定克隆,采用RT-PCR及免疫荧光技术检测重组质粒转染后细胞内Thy-1 mRNA及蛋白表达水平的改变;通过体外Transwell迁移实验研究外源性Thy-1对A549细胞迁移能力的影响。结果构建了pcDNA3.0-Thy-1真核细胞表达载体;成功建立了稳定表达外源性人Thy-1蛋白的肺癌A549细胞株。Transwell实验提示,表达外源性Thy-1蛋白的A549细胞较对照组细胞体外迁移能力下降(P<0.0001);基因芯片结果提示,Thy-1基因在肺癌细胞中表达后,影响了25个细胞迁移相关基因的表达,在检测基因表达差异时,RT-PCR与基因芯片能够得到基本一致的结果。结论Thy-1基因的过表达能降低肺癌细胞系A549细胞的迁移能力,并改变A549细胞多个细胞迁移相关基因的表达,提示Thy-1在肺癌组织的表达状态可能影响肺癌的转移性。
Objective To verify the relationship between Thy-1 (CD90) gene and the migration potential of lung cancer cells and to explore its mechanism. Methods The eukaryotic expression vector pcDNA3.0 was used to construct a recombinant plasmid containing the full-length Thy-1 gene. The recombinant plasmid was transfected into A549 cells. The stable clones were screened by G418. The transfected cells were detected by RT-PCR and immunofluorescence Thy-1 mRNA and protein expression levels in vitro; Transwell migration in vitro experiments exogenous Thy-1 on A549 cell migration ability. Results The eukaryotic expression vector pcDNA3.0-Thy-1 was constructed. A549 cell line stably expressing exogenous human Thy-1 protein was successfully established. The results of Transwell assay indicated that the ability of ex vivo Thy-1 protein-expressing A549 cells to migrate in vitro was lower than that of control cells (P <0.0001). The results of gene microarray indicated that the expression of Thy-1 gene in lung cancer cells affected 25 cells Migration related gene expression, in the detection of gene expression differences, RT-PCR and gene chips can get basically the same result. Conclusion Thy-1 gene overexpression can reduce the migration ability of lung cancer cell line A549 and change the expression of multiple cell migration related genes in A549 cells, suggesting that the expression of Thy-1 in lung cancer may affect the metastasis of lung cancer.