Figure 4. DCex induce apoptosis in tumor cells via TNF superfamily ligands. A-B. DCex induce apoptosis in tumor cells. Fifty micrograms of wt mDCex were coincubated with 2,500 B16 melanoma cells in the presence of 2% DMSO or 200 μM Z-VAD-fmk for 5 h or 48 h and tested using the apoptosis specific Hoechst DNA hypo-staining (A) and MTT (B) assays, respectively. p value represents statistically significant differences between DCex cytotoxic activities in the presence of DMSO and Z-VAD-fmk C-D. DCex of TNF- (Tnf^−/−) and FasL- (Fasl^−/−) deficient mice have impaired antitumor cytotoxic activity. Mature Tnf^−/− DCex (C) and Fasl^−/− DCex (D) were compared with mature wtDCex for their abilities to kill B16 melanoma cells as measured using 48 h MTT assay. p values represent statistically significant differences between tumoricidal activities of wtDCex vs. Tnf^−/− DCex or Fasl^−/− DCex. E-F. Neutralization of TRAIL inhibits DCex-mediated tumoricidal activity. Fifty micrograms of wt mDCex were coincubated with 2,500 B16 melanoma cells in the presence of 20 μg of IgG control or anti-TRAIL antibodies for 5 h or 48 h and tested using the apoptosis specific DNA hypo-staining (E) and MTT (F) assays, respectively. p values represent statisticallly significant differences between DCex cytotoxic activities in the presence of IgG control and anti-TRAIL antibodies. Data are from representative experiments of 2 experiments performed and represent % cytotoxicity means ± SD of triplicates.