Interleukin-8 (IL-8) plays a crucial role in the pathogenesis of inflammatory and hyperproliferative diseases in various organs. The purpose of the present investigation was to establish whether there is any naturally occurring inhibitor of IL-8. Here we demonstrate that an IL-8 inhibitor (IL-8INH) is present in the supernatant of polymorphonuclear (PMN) leukocytes. The release of IL-8INH could be increased by stimulating the PMN leukocytes by concanavalin A. IL-81NH blocks the IL-8-induced chemotaxis and Candida albicans killing activity of PMN leukocytes and epidermal cells in vitro, and IL-8-induced neutrophil infiltration in the mouse ear in vivo. The mechanism of action of IL-8INH involves blocking of 125I-IL-8 binding to the IL-8 receptor. Binding of 125I-IL-8 to neutrophils could not be displaced by the IL-8INH, however, preincubation of 125I-IL-8 with IL-8INH increased binding inhibition, suggesting an interaction between IL-8 and the inhibitor. Crosslinking of 125I-IL-8 to IL-8INH shows that IL-8INH binds specifically to 125I-IL-8, and the IL-8INH protein has an apparent molecular weight of 52 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The partial purification of the IL-8INH on DEAE-Sephadex anion-exchange chromatography column also suggests a 50-60-kDa inhibitor protein which blocks IL-8-induced effects on neutrophils by binding to IL-8.