Activation of CXCL5 induces intravesical chemoresistance of non-muscle invasive bladder cancer / 肿瘤

ABSTRACT

Objective: To explore the underlying mechanisms of drug-resistance of non-muscle invasive bladder cancer (NMIBC) to the intravesical chemotherapy. Methods: The chemoresistant bladder cancer cell line M-RT4 was established by continuous exposure of RT4 cells to mitomycin C (MMC). The chemoresistance and migration of M-RT4 cells were detected by CCK-8 assay and wound healing assay, respectively. The expressions of CXC chemokine ligand 5 (CXCL5) mRNA and protein in RT4 and M-RT4 cells as well as the recurrent NMIBC tissues were detected by real-time fluorescent quantitative PCR and Western blotting, respectively. The sensitivity of RT4 cells to MMC was observed after the treatment of recombinant human CXCL5 (rhCXCL5). The change of resistance of M-RT4 cells to MMC was detected by CCK-8 assay after siRNA transfection for CXCL 5 gene knockdown. The expressions of epithelial-mesenchymal transition (EMT)-associated factors [E-cadherin, cytokeratin-7 (Cyt-7), Claudin-1, N-cadherin, Vimentin and Snail] and nuclear factor-kappa B (NF-κB)-associated factors (NF-κB1, Rel A, NF-κB2 and Rel B) during the drug resistance of NMIBC cells were detected by Western blotting. Results: Compared to RT4 cells, the resistance of M-RT4 cells to MMC was significantly increased (P < 0.01), while the proliferation and migration of M-RT4 cells were enhanced (P < 0.05, P < 0.01). CXCL5 mRNA was over-expressed in the recurrent NMIBC tissues (P < 0.000 1), while the expression levels of CXCL5 mRNA and protein in M-RT4 cells were higher than those in RT4 cells (both P < 0.01). The sensitivity of RT4 cells to MMC was decreased after the treatment with rhCXCL5 (P < 0.01), whereas the sensitivity of M-RT4 cells to MMC was increased after CXCL 5 gene knockdown (P < 0.01). As compared with RT4 cells, the expressions of E-cadherin, Cyt-7 and Claudin-1 in M-RT4 cells were significantly decreased (all P < 0.01), but the expressions of N-cadherin, Vimentin, NF-κB1, Rel A, NF- κB2 and Rel B were increased (all P < 0.05). The treatment of rhCXCL up-regulated the expressions of Snail, NF-κB1, Rel A and Rel B in RT4 cells (all P < 0.05). Knockdown of CXCL 5 gene inhibited Snail activation (P < 0.05), up-regulated Cyt-7 expression (P < 0.01), and down-regulated the expressions of NF-κB1, Rel A and Rel B in M-RT4 cells (all P < 0.05). Conclusion: CXCL5 is over-expressed in NMIBC tissues and the chemoresistant bladder cancer cells. Knockdown of CXCL 5 gene expression may reduce the resistance of M-RT4 cells to MMC by reversing EMT, suggesting that CXCL5 is an important factor leading to the intravesical chemoresistance of NMIBC.