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Title: NITRIC OXIDE PRODUCTION AND iNOS mRNA EXPRESSION IN IFN-8-STIMULATED CHICKEN MACROPHAGES TRANSFECTED WITH iNOS siRNAs

Author
item CHEESEMAN, JENNIFER - ISU AMES,IA
item Lillehoj, Hyun
item LAMONT, SUSAN - ISU AMES,IA

Submitted to: Avian Immunology Group Meeting
Publication Type: Abstract Only
Publication Acceptance Date: 7/25/2008
Publication Date: 9/1/2008
Citation: Cheeseman, J.H., Lillehoj, H.S., Lamont, S.J. 2008. Nitric oxide production and inos mrna expression in ifn-8-stimulated chicken macrophages transfected with inos sirnas. Avian Immunology Group Meeting held June 234-27, 2008, Brisbane, Australia.

Interpretive Summary:

Technical Abstract: Utilizing RNA interference technology with siRNA in the HD-11 macrophage cell line, we determined how the knock-down of the iNOS (inducible nitric oxide synthase) gene affected IFN-' induced macrophage production of nitric oxide (NO) and mRNA expression of genes involved in this biological pathway in the chicken. Chicken macrophages produce NO when stimulated with recombinant chicken IFN-', however, when transfected with iNOS siRNAs, the production of NO was significantly decreased. We observed a 14-28% reduction in NO production by IFN-' stimulated HD-11 cells at 48 hours after initial siRNA transfection compared to non-transfected IFN-'-stimulated macrophages. Significant knock-down of iNOS mRNA expression (15 to 50- fold lower) was observed for each of four iNOS siRNAs, when compared to non-transfected IFN-'-stimulated macrophages and to those treated with a negative control siRNA. The IFN-'-stimulated chicken macrophages transfected with iNOS siRNAs did not show altered levels of mRNA expression for genes involved in IFN-' signaling and iNOS pathways (IL-1ß, IL-6, IFN-', TGF-ß4, or SOCS-3) suggesting that the observed decrease in NO production is a direct result of siRNA mediated knock-down of iNOS, rather than IFN-'-induced changes in the other genes tested.