Synergism of Fas-Mediated Apoptosis and Tumor Necrosis Factor on Adriamycin Cytotoxicity to Transitional Cell Carcinoma

Abstract

Objective: To explore the accurate Fas antigenic expression in transitional cell carcinoma (TCC) cells and and compare its synergistic modulation on adriamycin cytotoxicity with α-tumor necrosis factor (TNF-α). Methods: The expression of Fas antigen in normal urothelium and TCC cell lines was measured by flow cytometry. The Fas/Fas ligand reaction-induced cytotoxic changes in tumor cells were evaluated by microculture MTT technique. The synergism efficacy of Fas-mediated apoptosis and TNF-α on adriamycin cytotoxicity of TCC cells was analyzed. The Fas/Fas ligand-induced apoptosis in tumor cells was studied by annexin-V apoptotic cell expression and DNA ladder fragmentation analysis. Results: 75% of TCC cell lines showed Fas antigen expression. The Fas expression was not influenced by in vitro growth density of tumor cells. Apoptosis of TCC cells was observed during 48 h of treatment after activation of the Fas/Fas ligand pathway. TNF-α had a higher cytotoxicity activity on TCC cells than Fas ligand (17 vs. 0.7%) while combination of both reagents had 30% increased synergistic cytotoxicity. The increasing rate of apoptotic body after 3 days of treatment was 74% in adriamycin, 32% in Fas ligand, 60% in TNF-α, 69% in Fas and adriamycin combination and Fas and TNF-α combination, 103% in combination of TNF-α and adriamycin, and 136% in combination of these three reagents individually. In the DNA ladder analysis, Fas ligand had a 1.5-fold increase of DNA fragmentation when compared with adriamycin while TNF-α had a 4.4-fold increase and triple combination had a 5.5-fold increase after 3 days of treatment. Conclusions: Although the Fas antigen expression is common in TCC cells and apoptosis can be activated by the Fas/Fas ligand pathway activation, the synergistic cytotoxicity of adriamycin by the Fas/Fas ligand pathway is lower than that of TNF-α.