Cloning and characterization of Swine interleukin-17, preferentially expressed in the intestines.

摘要

Interleukin-17 (IL-17), initially reported as CTLA-8, is a proinflammatory cytokine produced mainly by activated T cells. In the present study, the cDNA of a swine IL-17 (PoIL-17) gene was cloned from activated neonatal thymocytes, and the recombinant PoIL-17 (rPoIL-17) was biologically characterized. The complete open reading frame (ORF) of PoIL-17 contains 462-bp coding deduced 153 amino acid residues, with a calculated molecular weight of 17.3 kDa. The amino acid sequence showed 72.9%, 64.9%, 64.7%, 60.1%, and 47.4% similarities with that of human, rat, mouse, Herpesvirus saimiri ORF 13, and chicken, respectively. The six cysteine residues conserved over species including the virus were observed in PoIL-17. We successfully prepared the recombinant mature form of PoIL-17 and analyzed its biologic activities for swine splenocytes. RT-PCR analysis revealed a marked upregulation of expression of IL-1beta, IL-8, tumor necrosis factor-alpha (TNF-alpha), granulocyte colony-stimulating factor (G-CSF), and monocyte chemotactic protein-1 (MCP-1) mRNA expression in splenocytes treated with 100 ng/ml rPoIL-17 for 3h. Furthermore, a swine chemokine, alveolar macrophage-derived neutrophil chemotactic factor II (AMCF-II), which was classified into the CXC subfamily was also augmented in mRNA level. This evidence indicates that recombinat PoIL-17 expressed in Escherichia coli was biologically active and exerted similar effects to those of a human (HuIL-17) and murine IL-17 (MuIL-17). The PoIL-17 mRNA is strongly expressed in the adult heart, skin, and, interestingly, intestinal tissues, including mesenteric lymph nodes but is restricted in neonatal tissues by using real-time quantitative RT-PCR. The gene sequence and biologically active recombinat protein for PoIL-17 will be useful for elucidation of the role of IL-17 in the regulation of intestinal immune responses.