In addition to being potent immunosuppressants, anti-CD3 monoclonal antibodies (mAb) are powerful mitogens in both humans and mice. The first antibody injection consistently induced an initial monocyte-dependent T cell activation with subsequent release of both monocyte- and T cell-derived cytokines [mainly tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma), interleukin (IL) 2, IL 3 and IL 6) into the circulation. This cytokine release is associated with a self-limiting, often severe, acute physical reaction in both patients and mice. We report here that a single injection of anti-TNF mAb prior to anti-CD3 administration not only neutralizes the biological activity of TNF but also strongly affects the release of other cytokines, with notably an up-regulation of IFN-gamma release and a down-regulation of IL 3 and IL 6 release. Conversely, pretreatment with anti-IFN-gamma mAb increases IL 3 and IL 6 production but does not affect TNF levels. Taken together, these data point to a pivotal role of IFN-gamma in the anti-CD3-induced cytokine cascade and reveal new regulatory pathways between TNF and IFN-gamma. With regard to the clinical implications of these findings, as anti-TNF mAb prevents anti-CD3-induced sickness in mice, whereas anti-IFN-gamma does not, such a therapeutic approach might be of value in OKT3-treated patients.