Expression Of CD154 On Bladder Transitional Cell Carcinomas And Its Effects On Tumorigenesis

Objectives: To investigate CD154 expression in transitional cell carcinoma(TCC) of bladder and its correlation with tumor stage and tumor grade,and further to explore its effects on tumorigenesis by in vitro and in vivo experiments.Methods: 1.CD154 expression was examined immunohistochemically in a series of 56 TCCs and 10 normal urotheliums of the bladder,and its correlation with tumor stage and tumor grade was analysed statistically. 2. CD154 mRNA was extracted from splenic lymphocytes after stimulated with Phytohemagglutinin (PHA) in vitro for 6 hours.CD154 cDNA was expanded by RT-PCR.The recombinant CD154 plasmid was generated by ligation of the PCR gene product to the unique HindⅢ and Xho Ⅰ restriction sites of pcDNA3.1/Zeo(+).The recombinant CD154 plasmids were transfected into BTT739 cells,a kind of TCC of poor differentiation from mouse,and cells were selected with Zeocin for stable expression(BTT739-CD154 and BTT739-pcDNA3.1/Zeo+). 3.In vitro cell proliferation and apoptosis was measured by MTT and terminal deoxynucleotidyl transferase nick-end labeling(TUNEL) respectively,and comparations were performed betweenBTT739.BTT739-CD154 and BTT739-pcDNA3.1/Zeo(+).In vivo tumor growth of the three kinds of tumor cells was compared in syngeneic T739 mice.Results: 1. The expression of CD154 in TCC of the bladder decreased compared with that in normal transitional epithelium(P<0.05),with a significant association with both lower stage and lower stage(P<0.01, PO.025 respectively). 2.The mouse recombinant CD154 plasmid was constructed successfully. After gene transfection, the positive clones of BTT739 cells with stable CD154 (pcDNA3.1/Zeo+) expression were formed in a selective medium of Zeocin. 3.The expression of CD154 in BTT739 cells,without CD40 expresion,had no significant effects on cell proliferation and apoptosis in vitro(P — 0.115,P = 0.797respectively). However, BTT739-CD154 tumor growth was significantly suppressed in syngeneic mice compared with BTT739 and BTT739-pcDNA3.1/Zeo(+) cells.Conclusions: The expression of CD 154 in TCC of the bladder decreases compared with that in normal transitional epithelium,with a significant association with both lower stage and lower stage. CD154 expression has no effects on cell proliferation and apoptosis of CD40 negative tumor,but can activate immune system to suppress the tumor growth in vivo.Its role in the pathogenesis of CD40 positive tumor and CD154/CD40-based immunotherapy needs futher research.