Recent studies have shown that thymosin fraction 5 (TF5) enhances production of interleukin-2 (IL-2) by phytohemagglutinin (PHA)-stimulated normal human peripheral blood mononuclear leukocytes (PBL). In this study we sought to determine whether this effect of TF5 might be mediated via the cyclooxygenase or lipoxygenase pathways. Our studies demonstrate that aspirin, an inhibitor of the cyclooxygenase pathway, given in vivo, or added to cultures in vitro, results in two-fold increased IL-2 production by PHA-stimulated PBL. This increase is comparable to that seen when PBL are cultured in vitro with TF5. When aspirin and TF5 are added simultaneously to PBL in the presence of PHA, an additive response is seen. An inhibitor of the lipoxygenase pathway, 15-hydroxyeicosatetraenoic acid, did not significantly change IL-2 production by PBL or influence the enhancement by TF5. Augmentation of IL-2 production by aspirin and/or TF5 was prevented by monocyte depletion of the PBL population. These results are interpreted as demonstrating (a) that TF5 and aspirin augment, by distinct mechanisms, IL-2 production by normal human PBL, (b) that the effects of both of these agents are mediated directly or indirectly via a monocyte population and (c) that aspirin, in addition to its analgesic and anti-inflammatory properties, may act as a modulator of immunological responsiveness, either alone or in combination with other biological response modifiers such as thymosin.