The Functions And Mechanisms Of Connective Tissue Growth Factor In Bladder Cancer

Background and Objectives Urothelial bladder carcinoma(UBC),the most common and lethal genitourinary tract cancer in China,is characterized by complex biological behaviors and high risks of recurrence and progression.Chemotherapy is the essential approach for UBC treatment,however,chemoresistance considerably weakens the therapeutic effects.Connective tissue growth factor(CTGF),namely CCN2,a kind of matricellular protein secreted by normal or tumor cells,is demonstrated to promote cell proliferation,adhension,migration and invasion by binding integrin and activating downstream signal pathways in previous studies.CCN2 is aberrantly expressed in various cancers and exerts functions on tumor biobehaviors or chemosensitivity,while the functional mechanisms of CCN2 in UBC have not been illustrated until present.Hence,in levels of tissues,in vitro and in vivo,we invesitigated the detailed expression patterns of CCN2 in UBC tissues and analyzed the relevant mechanisms in modulation of UBC biobehaviors and chemosensitivity.The present study will help understanding the mechanisms of UBC development and provide evidence for novel stratergies of improving therapeutic effects in UBC.Materials and Methods 1.Surgical samples were collected from patients with nonmuscle invasive bladder cancer(NMIBC)or muscle invasive bladder cancer(MIBC)at our departement.Immunohistochemistry assay was employed to detect the expression of CCN2 in tumor tissues and adjacent normal urothelium and correlation analysis was performed to determine its association with clinicopathologic features and prognostic results.2.By using lipofectamine 2000,T24 and 5637 cells were transfected with CCN2 small interfering RNA(si RNA)or scramble oligonucleotides.Cell proliferation,migration and invasion were observed and cell cycle was analyzed after CCN2 down-regulation.3.T24 cells were transfected with CCN2 or scamble short hairpin RNA(sh RNA)vectors,stably transfected cells were screened and then subcutaneously inoculated into nude mice to induce tumor xenografts.By observation of tumor growth and pathologic analysis of xenografts,effect of CCN2 on UBC development was investigated in vivo.4.UBC cells with different levels of CCN2 expression were treated with mitomycin-C(MMC),and toxcities of MMC and cell apoptosis were detected.After collecting protein from above MMC-treated cells,Western blot assay was employed to analyze the effects of CCN2 knockdown on Akt and Erk pathways.Results 1.Compared with normal urothelium,we observed significant overexpression of CCN2 UBC tissues,especially in MIBC lesions.Higher expression of CCN2 was associated with advanced tumor stage and high pathologic grade as well as increased risks of recurrence and progression.2.CCN2 knockdown suppressed proliferation,migration and invasion of T24 and 5637 cells and blocked G1 to S cell cycle transition.3.In the xenograft model,targeting of CCN2 remarkably inhibited UBC tumor growth in nude mice.4.Down-regulation of CCN2 increased the toxicity of MMC in UBC cells and aggrevated MMC-induced cell apoptosis.When treated with MMC,UBC cells with CCN2 knockdown showed inhibition of phosphorylation of Akt and Erk.Conclusions 1.CCN2 is overexpressed in UBC,especially in MIBC tissues.High expression of CCN2 is associated with malignant pathologic features and unfavorable prognosis in UBC.2.Targeting of CCN2 inhibits proliferation,cell cycle,migration and invasion of UBC cells and suppresses tumor growth in nude mice.3.By inactivating Akt and Erk pathways,CCN2 knockdown can increase sensitivity of UBC cells to chemotherapeutic agent MMC.