Endotoxin-Stimulated Monocytes Release Multiple Forms of IL-1β, Including a ProIL-1β Form Whose Detection Is Affected by Export

Abstract

Abstract The processing and release of 31-kDa proIL-1β to the mature 17-kDa form of IL-1β are still poorly understood. To help elucidate the mechanisms involved in IL-1β processing and release, we measured IL-1β forms released from endotoxin-stimulated monocytes by immunoprecipitation of [35S]methionine-labeled protein, by Western blots, and by our recently developed ELISA specific for proIL-1β. Our studies demonstrate that in addition to the 17-kDa mature IL-1β, IL-1β is also released as 31-, 28-, and 3-kDa molecules. The 31-kDa-released form of proIL-1β represented 20–40% of the total released IL-1β, as measured by SDS-PAGE with densitometry. This released proIL-1β was susceptible to ICE processing; however, this proIL-1β was not detectable by either a mature or proIL-1β-specific ELISA, suggesting that release induces a conformational change. The ELISA inability to detect proIL-1β was not due to inadequate sensitivity or subsequent degradation in the ELISA. Furthermore, while immunoaffinity-purified cytosolic proIL-1β could complex the type II IL-1R, released proIL-1β did not. Finally, the absence of a band shift in nondenaturing gel electrophoresis excluded proIL-1β binding to another protein. These findings imply that IL-1β is exported from monocytes as 3-, 17-, 28-, and 31-kDa forms and that the released 31-kDa form differs from cytosolic proIL-1β.