Identification and functional characterization of the house finch interleukin-1β

Abstract

Interleukin-1β (IL-1β), an inflammatory cytokine of the IL-1 family, is primarily produced as a precursor protein by monocytes and macrophages, then matures and becomes activated through proteolytic catalysis. Although the biological characteristics of avian IL-1β are well known, little information is available about its biological role in songbird species such as house finches that are vulnerable to naturally-occurring inflammatory diseases. In this study, house finch IL-1β (HfIL-1β) was cloned, expressed, and its biological function examined. Both precursor and mature forms of HfIL-1β consisting of 269 and 162 amino acids, respectively, were amplified from total RNA of spleen and cloned into expression vectors. HfIL-1β showed high sequential and tertiary structural similarity to chicken homologue that allowed detection of the expressed mature recombinant HfIL-1β (rHfIL-1β) with anti-ChIL-1β antibody by immunoblot analysis. For further characterization, we used primary splenocytes and hepatocytes that are predominant sources of IL-1β upon stimulation, as well as suitable targets to stimulation by IL-1β. Isolated house finch splenocytes were stimulated with rHfIL-1β in the presence and absence of concanavalin A (Con A), RNA was extracted and transcript levels of Th1/Th2 cytokines and a chemokine were measured by qRT-PCR. The addition of rHfIL-1β induced significant enhancement of IL-2 transcript, a Th1 cytokine, while transcription of IL-1β and the Th2 cytokine IL-10 was slightly enhanced by rHfIL-1β treatment. rHfIL-1β also led to elevated levels of the chemokine CXCL1 and nitric oxide production regardless of co-stimulation with Con A. In addition, the production of the acute phase protein serum amyloid A and the antimicrobial peptide LEAP2 was observed in HfIL-1β-stimulated hepatocytes. Taken together, these observations revealed the basic functions of HfIL-1β including the stimulatory effect on cell proliferation, production of Th1/Th2 cytokines and acute phase proteins by immune cells, thus providing valuable insight into how HfIL-1β is involved in regulating inflammatory response.