Differential synthesis, cellular localisation and secretion of interleukin-1α and interleukin-1β from ovine macrophages

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-1α and IL-1β from alveolar macrophages. Immunoprecipitation of IL-1α and IL-1β 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-1β than IL-1α was secreted by the macrophages. Flow cytometric analysis of IL-1α and IL-1β expression by fresh unstimulated macrophages and macrophages cultured for 5 h with LPS demonstrated that a proportion of the cell associated IL-1α, but not IL-1β, in stimulated macrophages was expressed at the cell surface. Analysis of IL-1 secretion by cultured alveolar macrophages, using IL-1α and IL-1β specific immunoassays, confirmed that IL-1β was the predominant secreted species of IL-1. While cell associated IL-1α and IL-1β were detected by immunoprecipitation and flow cytometric analysis of macrophages cultured in media alone or media supplemented with LPS, secreted IL-1β was detected only after stimulation of macrophages with LPS. This indicates a dissociation of IL-1β synthesis and secretion and is indicative of an IL-1β converting enzyme similar to that which has been described in the human and mouse models.