Autocrine production of TNF-α contributes to cytokine-induced apoptosis of human NK cells via the p80 TNF-αR. In each condition described below, purified CD3⁻CD56^bright+ NK cells were isolated from fresh blood10 and cultured at 1 × 10⁵ cells/well for 3 days in medium containing 15 ng/mL IL-15 and 3 ng/mL IL-12, after which nuclei were stained with PI to obtain a quantitative measurement of apoptosis. (A) Culture in the presence of a nonreactive receptor fusion protein (IL-4R-F_c , 500 μg/mL) shows 54% of nuclei in the hypodiploid peak, characteristic of apoptosis, while (B) culture in the presence of a TNFR-F_c fusion protein (500 μg/mL) partially neutralizes TNF-α and shows only 21% of nuclei in the hypodiploid peak. (C) Cultures containing a nonreactive isotype control MoAb (50 μg/mL) show 55% of nuclei in the hypodiploid fraction of DNA while (D) incubation with a blocking MoAb to the p80 TNF-αR (M1, 50 μg/mL) shows 32% of nuclei in the hypodiploid DNA fraction. (E) Cultures containing a nonreactive isotype control MoAb (50 μg/mL) show 64% of NK cell nuclei in the hypodiploid fraction of DNA and (F ) incubation with a blocking MoAb to the p60 TNF-αR (M50, 50 μg/mL) shows no measurable effect compared with control. Results were obtained with NK cells from a single donor and are representative of three experiments using different donors.