title: Differential synthesis, cellular localisation and secretion of interleukin-1 alpha interleukin-1 beta from ovine macrophages.
abstract: In order to characterise the regulatory processes involved in expression of ruminant interleukin 1 (IL-1) biological activity, we have used specific monoclonal antibodies to assess synthesis, cellular localisation and secretion of ovine IL-1 alpha and IL-1 beta from alveolar macrophages. Immunoprecipitation of IL-1 alpha and IL-1 beta from lysates of macrophages cultured in media alone or media supplemented with lipopolysaccharide (LPS) revealed that both forms of IL-1 were synthesised as precursor proteins of 31-33 kDa. In contrast, both IL-1 species were immunoprecipitated from culture supernatants as 17 kDa molecules. Comparison of the precipitated bands from culture supernatants suggested that significantly more IL-1 beta than IL-1 alpha was secreted by the macrophages. Flow cytometric analysis of IL-1 alpha and IL-1 beta expression by fresh unstimulated macrophages and macrophages cultured for 5 h with LPS demonstrated that a proportion of the cell associated IL-1 alpha, but not IL-1 beta, in stimulated macrophages was expressed at the cell surface. Analysis of IL-1 secretion by cultured alveolar macrophages, using IL-1 alpha and IL-1 beta specific immunoassays, confirmed that IL-1 beta was the predominant secreted species of IL-1. While cell associated IL-1 alpha and IL-1 beta were detected by immunoprecipitation and flow cytometric analysis of macrophages cultured in media alone or media supplemented with LPS, secreted IL-1 beta was detected only after stimulation of macrophages with LPS. This indicates a dissociation of IL-1 beta synthesis and secretion and is indicative of an IL-1 beta converting enzyme similar to that which has been described in the human and mouse models.
authors.affiliation: Department of Veterinary Science, University of Melbourne, Vic., Australia.
authors.forename: P J
authors.initials: PJ
authors.lastname: Egan
authors.forename: A D
authors.initials: AD
authors.lastname: Nash
mesh_terms: D000818:Animals; D002460:Cell Line; D002474:Cell-Free System / Q000276:immunology; D005434:Flow Cytometry; D007118:Immunoassay / Q000662:veterinary; D007375:Interleukin-1 / Q000096:biosynthesis* / Q000378:metabolism*; D016676:Macrophages, Alveolar / Q000378:metabolism*; D008565:Membrane Proteins / Q000096:biosynthesis; D011233:Precipitin Tests / Q000662:veterinary; D012756:Sheep
chemical_list: D007375:Interleukin-1; D008565:Membrane Proteins