4195 23 archive 337 disk0/00/00/41/95 2008-04-07 14:27:00 2008-07-18 10:49:14 2008-07-16 17:05:50 article show Richard.Morrison@utas.edu.au Morrison RN Richard.Morrison@utas.edu.au Zou J Secombes CJ Scapigliati G Adams MB Mark.Adams@utas.edu.au Nowak BF B.Nowak@utas.edu.au pub 070401 300703 630303 restricted TNF-a; IL-1b; Atlantic salmon; Amoebic gill disease The definitive version is available at http://www.sciencedirect.com category => A1 categoryDesc => Refereed Article in a Scholarly Journal eprintID => 0 field1 => Fish and Shellfish Immunology field10 => field11 => field12 => field13 => field2 => United Kingdom field3 => 23 field4 => xx field5 => 1015-1031 field6 => Academic Press field7 => 1050-4648 field8 => field9 => funding => S grant => lastUpdate => 12/03/2008 rfcd => 300703 seo => 630303 themeArea => SPP title => Molecular cloning and expression analysis of tumour necrosis factor-alpha in amoebic gill disease (AGD)-affected Atlantic salmon (Salmo salar L.) tor => PB uid => 48522 update => no Tumour necrosis factor-alpha (TNF-a) is a key mediator of inflammation during amoebiasis of humans and mice. Atlantic salmon (Salmo salar L.) are also susceptible to infection by amoebae (Neoparamoeba spp.), inflicting a condition known as amoebic gill disease (AGD). Here, the role of TNF-a in AGD-pathogenesis was examined. Two Atlantic salmon TNF-a transcripts designated TNF-a1 and TNF-a2 together with their respective genes were cloned and sequenced. TNF-a1 is 1379 bp and consists of a 738 bp open reading frame (ORF) translating into a predicted protein of 246 amino acids. TNF-a2 is 1412 bp containing an ORF and translated protein the same lengths as TNF-a1. An anti-rainbow trout TNF-a polyclonal antibody that bound recombinant Atlantic salmon TNF-a1 and TNF-a2 was used to detect constitutive and inducible expression of TNF-a in various tissues. The anti-TNF-a antibody bound to a TNF-like protein z60 kDa that was constitutively expressed in a number of tissues in healthy Atlantic salmon. However, this protein was not detected in lysates from mitogen-stimulated head kidney leucocytes, despite up-regulation of TNF-a mRNAs under the same conditions. During the early onset of AGD in Atlantic salmon, there were no demonstrable differences in the gill tissue expression of TNF-a1, TNF-a2 nor the interleukin-1 beta (IL-1b), inducible nitric oxide synthase (iNOS) and interferon gamma (IFN-g) mRNAs compared to tissue from healthy fish. In Atlantic salmon with advanced AGD, IL-1b but not TNF-a1 or TNF-a2 mRNAs was up-regulated and was lesion-restricted. Given that Neoparamoeba spp. modulated both TNF-a2 and IL-1b in head kidney leucocytes in vitro, it appears that rather than being recalcitrant to Neoparamoeba spp.-mediated TNF-a expression, either the parasite can influence the cytokine response during infection, there is ineffective signalling for TNF-a expression, or there are too few cells at the site of infection with the capacity to produce TNF-a. These data support our previous observation that IL-1b mRNA expression is up-regulated in AGD-affected tissue and that TNF-a is not intrinsic in AGD-pathogenesis. 2007 published Fish and Shellfish Immunology 23 5 Academic Press 1015-1031 10.1016/j.fsi.2007.04.003 TRUE 1050-4648 http://dx.doi.org/10.1016/j.fsi.2007.04.003 4827 3 4195 1 application/pdf en staffonly cc_utas

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