FIGURE 6.

Caspase Inhibitor I rescues inhibition of TNF-α secretion in pretreated macrophages following incubation with actinomycin D-killed Jurkat T cells and LPS. A and B, induction of cell death in J774 macrophages co-cultured with Jurkat cells treated with etoposide (Etop.) or the indicated doses of actinomycin D (ActD). A, FACS analysis was gated on J774 cells only (by staining for the macrophage marker F4/80) and excluded all Jurkat cells. Representative untreated J774 cells (Live J774) and J774 cells induced to die by direct treatment with actinomycin D (Dead J774) are shown for comparison. AnnV, annexin V. B, combined data from three independent co-cultures of J774 and Jurkat cells. C, TNF-α production by J774 macrophages (mTNF-α) pretreated with Z-VAD-fmk or untreated, followed by co-culture with Jurkat T cells and LPS stimulation as described in the legend to Fig. 1. Data are presented as means ± S.E. from three independent experiments. *, statistically significant differences between the indicated values (p < 0.05; ANOVA). NS, not statistically significantly different. D, TNF-α production by J774 macrophages treated directly with actinomycin D ([ActD]) or co-cultured with Jurkat T cells previously treated with actinomycin D (Jurkat [ActD]), followed by overnight stimulation with LPS. Dashed lines represent response of untreated macrophages (100%) and suppression of macrophage response to 50 and 25% of untreated levels.