人核因子-κBp65 shRNA慢病毒载体构建及对肺癌细胞恶性生物学行为的影响

BACKGROUND AND OBJECTIVE: Nuclear factor-κB is an important transcription factor and is closely associated with a variety of malignant tumors. The biological behavior of lung tumor cells can be reversed by inhibiting the expression of NF-κBp65 directly or indirectly. Nuclear factor-κBp65 gene shRNA recombinant plasmids were constructed and then infected with A549 cells. New stable cell lines were selected, and the ability of migration and adhesion was identified. METHODS: Both scramble control sequence and interference sequence (shRNA) of human nuclear factor-κBp65 were designed and synthesized to build recombinant plasmids, with BamH I site at the 5′ end and Xho I and EcoR I sites at the 3′ end. A549 cells were infected, and stable transfection strains were selected by puromycin. Western blot and qRT-PCR methods were applied to assess the interference efficient of NF-κBp65 and the protein expression level of IκBα. Transwell and MTT assays were carried out to analyze the ability of migration and adhesion of A549 cells separately. RESULTS: Recombinant plasmids were successfully built, and A549/NF-κB p65 scramble and A549/NF-κB p65 shRNA stable transfection strains were also successfully screened. Both mRNA and protein expression levels of NF-κBp65 showed that A549/NF-κBp65 shRNA cells decreased compared with A549/NF-κB p65 scramble cells and A549 cells, whereas the protein level of IκBα significantly increased. Both migration and adhesion abilities were also reduced. CONCLUSION: In this study, both mRNA and protein expression levels of NF-κBp65 were effectively suppressed by RNA interference technique. NF-κBp65 inhibition can significantly reduce the migration and adhesion ability of A549 cells.