Figure 9.

iNKT cells degranulate in response to low dose DAC-treated SK-MES-1 cells. SK-MES-1 cells were treated with PBS or DAC (50 nM, 100 nM and 1 μM) at 24, 48 and 72 hours. PBS and methanol (MeOH) were used as negative and vehicle controls. After a rest period of 3 days the cells were pulsed with vehicle, α-GalCer or 7DW8-5 for 24 hours, and then co-cultured for 4 h with iNKT cells and a mAb specific for CD107a. In some wells, 10 μg/ml of a blocking antibody specific for CD1d was included. PMA and ionomycin (P/I) treatment was used as a positive control. A, Flow cytometric dot plot showing cell-surface CD107a expression by iNKT cells after exposure to medium (left) or α-GalCer + DAC (right). B, Mean (± SEM) frequencies of iNKT cells that expressed CD107a after co-culture with control SK-MES-1 or DAC-treated SK-MES-1 cells pulsed with glycolipids. Percentages of iNKT cells that expressed CD107a in response to PBS- or DAC-treated SK-MES-1 cells were compared to those exposed to SK-MES-1 cells treated with vehicle alone using a two way ANOVA with Bonferroni's multiple comparison test. Results are representative of 3 experiments. C, % inhibition of cytolytic degranulation by iNKT cells in response to DAC-treated A549 cells pulsed with α-GalCer or 7DW8-5 or vehicle using an anti-CD1d mAb. Results are means of 4 experiments. (**p < 0.01, ***p < 0.001).