Aggregation studies have become a useful criterion for analyzing leukocyte motility and activation in vitro. The T-cell-derived lymphokine human leukocyte inhibitory factor (LIF) is a modulator of many important polymorphonuclear (PMN) functions in addition to aggregation such as chemotaxis, lysosomal degranulation, phagocytosis, bactericidal killing, augmented antibody-dependent cellular cytotoxicity (ADCC), induction of neutrophil Fc-gamma, complement type-1 and FMLP receptors, and production of superoxide and H2O2. Our investigations focused on the ability of LIF to modulate the aggregation of macrophages (MO) induced by calcium ionophore A23187. The ionophore A23187 directly induced potent aggregation of macrophages, which was markedly enhanced when the cells were pretreated with LIF. However, the addition of LIF in the absence of other costimuli did not directly induce MO aggregation. LIF was shown to enhance PMN aggregation induced by N-formyl-Met-Leu-Phe (FMLP), but did not augment the aggregation of FMLP-stimulated macrophages, indicating a cellular specificity of aggregation-inducing costimuli following LIF priming. Additional cytokines examined for possibly inducing MO aggregation were interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF), and interleukin-6 (IL-6); all proved to be incapable of inducing aggregation directly, nor did they enhance the effect of A23187 ionophore on macrophage aggregation. Additionally, we found that LIF can directly stimulate MO to activate specific pathways of the arachidonic acid cascade, inducing the synthesis and release of thromboxanes and leukotriene B4. LIF did not augment the potent ability of A23187 to induce increased production of LTB4 or TxA2 by human MO. These new results coupled with our previously published data indicate that LIF can enhance the activation of both MO and PMN leukocytes when exposed to either A23187 or FMLP, respectively. Moreover, these data suggest that LIF can contribute directly to monocyte-macrophage leukocyte activation, in addition to PMN activation, during inflammatory responses, resulting in greater cell aggregation, activation, and specific proinflammatory arachidonic acid product release.