Interleukin-1α activation and localization in lipopolysaccharide-stimulated human monocytes and macrophages

Interleukin-1α (IL-1α) is a proinflammatory cytokine belonging to the IL-1 family. It is synthesized as a 33kDa precursor peptide that is cleaved by a calpain-like protease to a 16kDa propiece and a 17kDa mature IL-1α peptide. In contrast to its close relative, IL-1β, the role of IL-1α in inflammation is only partly understood. Human monocyte derived macrophages, stimulated with lipopolysaccharide (LPS) were analysed for production and localization of IL-1α by use of a monoclonal antibody (MAb) generated against recombinant precursor IL-1α. We found that the MAb detected IL-1α within the nuclei of the cells 2h (hours) after LPS stimulation and production continued for up to 20h. At no time could we demonstrate cleavage of the IL-1α precursor. The MAb was conjugated to fluorescein isothiocyanate (FITC) for use in flow cytometry. Based on the flow cytometric analysis CD68 positive cells were positive for IL-1α in agreement with CD68 being a marker for monocytes. Here, we demonstrate, for the first time, a method to visualize and measure the production of IL-1α in both human monocytes and macrophages. [Display omitted] •We made a monoclonal antibody, MAb1.3.2, against recombinant precursor IL-1α.•MAb1.3.2 was conjugated to fluorescein isothiocyanate (FITC) for use in flow cytometry.•We have identified the localization of IL-1α using MAb1.3.2 in human macrophages.•IL-1α producing PBMC's were both positive for CD68 and CD14.•Our study presents an assay suitable for comparison between donors or during disease.