Submitted to: American Dairy Science Association Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: 5/5/2009
Publication Date: 7/12/2009
Citation: Nelson, C.D., Beitz, D.C., Reinhardt, T.A., Lippolis, J.D. 2009. Expression of Inducible Nitric Oxide Synthase is Up-Regulated by Production of 1,25-Dihydroxyvitamin D3 in Bovine Monocytes in Response to Toll-Like Receptor Signaling [abstract]. American Dairy Science Association. Paper No. 84. Interpretive Summary:
Technical Abstract: In the endocrine pathway of vitamin D signaling 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] is produced from 25-hydroxyvitamin D3 [25(OH)D3] by the enzyme CYP27B1 in the kidney. Production of 1,25(OH)2D3 in the kidney functions to regulate gene expression systemically. However, recent studies have shown that human monocytes express CYP27B1 and produce 1,25(OH)2D3 in response to toll-like receptor recognition of bacterial cell wall components. Activation of 25(OH)D3 by CYP27B1 in human monocytes improves antimicrobial activity via an autocrine pathway. The purpose of this study was to determine if 1,25(OH)2D3 is produced by CYP27B1 in bovine monocytes upon TLR signaling and if 1,25(OH)2D3 production in bovine monocytes increases expression of antimicrobial genes. Monocytes were isolated from peripheral blood of six lactating Holstein cows and cultured in media containing 10% FBS. Monocyte cultures were treated with 0-75 ng/ml 25(OH)D3 or 0-4 ng/ml 1,25(OH)2D3 and stimulated for 24 hours with lipopolysaccharide (LPS) a tripalmitoylated lipopeptide (Pam3CSK4) or peptidoglycan (PGN). Relative expression of CYP27B1, cathelicidin 4 (CATH4), CATH5, CATH6, and inducible nitric oxide synthase (iNOS) mRNA in monocytes was measured using quantitative real-time PCR. CYP27B1 expression in monocyte cultures was induced by stimulation with LPS, Pam3CSK4 or PGN (p < 0.05). Treatment of stimulated monocyte cultures with 25(OH)D3 or 1,25(OH)2D3 increased iNOS expression (p < 0.05). Treatment of un-stimulated monocyte cultures with 1,25(OH)2D3, but not 25(OH)D3, increased iNOS expression. Expression of CATH4, CATH5, and CATH6 genes in stimulated monocytes was not affected by 25(OH)D3 or 1,25(OH)2D3 treatment. In conclusion, bovine monocytes convert 25(OH)D3 to 1,25(OH)2D3 in response to TLR signaling and 1,25(OH)2D3 production up-regulates iNOS expression. This study is the first to show that an autocrine pathway of vitamin D signaling has an impact on the innate immune response in cattle.