The inhibition of proliferation and invasion of bladder cancer BLS cells induced by RNA interference on NF-κBp65

Objective: To investigate the effects of RNA interference on NF-κB (nuclear factor-kappa B) p65 on the proliferation and invasion of bladder transitional cell carcinoma BLS cells, and to examine the expression of NF-κBp65 in human bladder transitional cell carcinoma tissues. Methods: The recombinant vector pRNAT-U6.1-NF-κBp65-shRNA/Neo in which the shRNA (short hairpin RNA) targeting NF-κBp65 was established, and then it was transfected into the BLS cells. The efficacy of gene silencing was confirmed by Western blotting. The proliferation, migration and invasion abilities of BLS cells after transfection with pRNAT-U6.1-NF-κBp65-shRNA/Neo were detected by MTT assay and colony-forming assay, respectively. The downstream targets of NF-κB, including c-myc, cyclin D1 and MMP9 (matrix metalloproteinase 9) were examined by Western blotting. The expressions of NF-κBp65 and MMP9 proteins in 60 cases of bladder transitional cell carcinoma tissues were detected by immunohistochemistry. Results: The recombinant vector pRNAT-U6.1-NF-κBp65-shRNA/Neo was successfully constructed, and the stably transfected cells were established. As compared with NC (negative control) NF-κBp65shRNA-NC-BLS cells which were tranfected with an empty vector, the abilities of proliferation, colony formation, migration and invasion of the NF-κBp65shRNA-BLS cells were obviously decreased (P < 0.05), as well as the expression levels of c-myc, cyclin D1 and MMP9 proteins were down-regulated (P < 0.05). The expression levels of NF-κBp65 and MMP9 in stages T_2-4 bladder transitional cell carcinoma and the cancer with lymphatic metastasis were higher than those in stages Tis-T₁ bladder transitional cell carcinoma and the cancer without lymphatic metastasis (P < 0.05). There was an obviously positive correlation between the NF-κBp65 and MMP9 expressions (r = 0.329, P < 0.05). Conclusion: NF-κB signal transduction way may be related to the proliferation and invasion of bladder cancer cells.