TNF-alpha at 50-100 U/ml synergizes with IL-2 in enhancing LAK activity and IL-6 production in low-dose IL-2 (1-10 U/ml) culture of human PBL. High-dose TNF-alpha (> or = 200 U/ml) has less effect and even sometimes resulted in lowering of both LAK activity and IL-6 production below control levels. TNF-alpha-mediated regulation of low-dose IL-2 activation occurs even at late stages (effector phase) of LAK development. IL-6, as previously reported, acts at late stages of low-dose IL-2 culture to enhance LAK, but does not stimulate TNF-alpha production. The combined addition of TNF-alpha and IL-6 to late stages of IL-2 culture does not produce any additive or synergistic effect on LAK. We tested for the relative roles of TNF-alpha and IL-6 in late stage regulation of LAK development with antibodies (Abs) to these cytokines. Anti-IL-6 Ab abrogates late phase LAK enhancement by TNF-alpha, while anti-TNF-alpha Ab has no effect on IL-6 augmentation of LAK cytotoxicity. IL-2 added to PBL culture at doses greater than 10 U/ml induces production of both TNF-alpha and IL-6. Addition of anti-TNF-alpha Ab at late stages of high-dose IL-2 (> or = 20 U/ml) culture decreases both LAK cytotoxicity and IL-6 production, and the inhibition of LAK is reversed by the addition of IL-6. By contrast, anti-IL-6 Ab decreases LAK cytotoxicity, but does not alter TNF-alpha production, and the inhibition of LAK is not reversed by addition of TNF-alpha. These data indicate that TNF-alpha is important for both LAK development and IL-6 secretion in PBL, and that IL-6 is the proximate mediator in TNF-alpha regulation of these cytotoxic cell functions.