title: Regulation of toll-like receptors 3, 7 and 9 in porcine alveolar macrophages by different genotype 1 strains of porcine reproductive and respiratory syndrome virus.
abstract: The toll-like receptors (TLRs) play an important role in the innate host defense against pathogens. Endosomal TLRs, TLR3, TLR7/8, and TLR9 are involved in antiviral responses by promoting the production of antiviral cytokines such as type I interferons. Porcine reproductive and respiratory syndrome (PRRS) is an important disease causing economically high losses to the swine industry worldwide and caused by a single stranded positive sense RNA virus, known as PRRS virus (PRRSV). Studies focused on the interaction between PRRSV and TLRs are scarce. The aim of the present study was to evaluate the expression of TLR3, TLR7 and TLR9 in porcine alveolar macrophages (PAM) infected with different genotype 1 PRRSV strains previously sequenced and characterized by their ability to induce TNF-α: 3262 (TNF-α inducer), 3267 (TNF-α not inducer) and an attenuated vaccine strain (strain Deventer, PorcilisPRRS, Merck) that replicates scarcely in PAM. PAM were infected with the different PRRSV strains (at 0.1 multiplicity of infection) for 48 h or mock-stimulated with PAM supernatants. Cells were collected at different time-points (0 h, 6 h, 12 h, 24 h, 36 h, 48 h) to determine the kinetics of viral replication by quantitative RT-PCR (qRT-PCR) and the expression of TLR3, 7 and 9 by qRT-PCR, flow cytometry and indirect immunofluorescence assay. Although infection with PRRSV did not affect significantly relative levels of any TLR mRNA transcript (normalized to β-actin expression), this infection resulted in significant differences in the proportion of cells expressing TLR3. Thus, in PAM infected with PRRSV strain 3262 the proportion of TLR3+ cells significantly increased from 24h compared with the controls; in contrast strain 3267 resulted in a lower proportion of TLR3+ PAM. Interestingly, strain 3262 replicate to lower levels than 3267 at comparable post-inoculation times. For strain DV, the results indicated that this strain did not replicate substantially in PAM and did not stimulated TLR3 expression. These observations suggest that at least TLR3 is regulated differentially by different genotype 1 PRRSV strains and this seems to be related apparently to the replication levels of each strain, as well as, to the TNF-α inducing capability. The fact that mRNA transcripts were kept constant also suggests that this regulation occurs at a post-transcriptional level.
authors.affiliation: Centre de Recerca en Sanitat Animal (CReSA), UAB-IRTA, Campus de la Universitat Autònoma de Barcelona, 08193 Bellaterra (Cerdanyola del Vallès), Spain.
authors.forename: Liudmila
authors.initials: L
authors.lastname: Kuzemtseva
authors.affiliation: Centre de Recerca en Sanitat Animal (CReSA), UAB-IRTA, Campus de la Universitat Autònoma de Barcelona, 08193 Bellaterra (Cerdanyola del Vallès), Spain.
authors.forename: Eugenia
authors.initials: E
authors.lastname: de la Torre
authors.affiliation: Centre de Recerca en Sanitat Animal (CReSA), UAB-IRTA, Campus de la Universitat Autònoma de Barcelona, 08193 Bellaterra (Cerdanyola del Vallès), Spain.
authors.forename: Gerard
authors.initials: G
authors.lastname: Martín
authors.affiliation: Centre de Recerca en Sanitat Animal (CReSA), UAB-IRTA, Campus de la Universitat Autònoma de Barcelona, 08193 Bellaterra (Cerdanyola del Vallès), Spain.
authors.forename: Ferran
authors.initials: F
authors.lastname: Soldevila
authors.affiliation: Roslin Institute and Royal (Dick) School of Veterinary Studies, Easter Bush Campus, Midlothian EH25 9RG, United Kingdom.
authors.forename: Tahar
authors.initials: T
authors.lastname: Ait-Ali
authors.affiliation: Centre de Recerca en Sanitat Animal (CReSA), UAB-IRTA, Campus de la Universitat Autònoma de Barcelona, 08193 Bellaterra (Cerdanyola del Vallès), Spain; Department de Sanitat i Anatomia Animals, Universitat Autònoma de Barcelona, 08193 Bellatera, Barcelona, Spain.
authors.forename: Enric
authors.initials: E
authors.lastname: Mateu
authors.affiliation: Centre de Recerca en Sanitat Animal (CReSA), UAB-IRTA, Campus de la Universitat Autònoma de Barcelona, 08193 Bellaterra (Cerdanyola del Vallès), Spain; Department de Sanitat i Anatomia Animals, Universitat Autònoma de Barcelona, 08193 Bellatera, Barcelona, Spain. Electronic address: laila.darwich@uab.cat.
authors.forename: Laila
authors.initials: L
authors.lastname: Darwich
mesh_terms: D000818:Animals; D005786:Gene Expression Regulation; D005838:Genotype; D054884:Host-Pathogen Interactions / Q000235:genetics / Q000276:immunology; D007113:Immunity, Innate / Q000235:genetics; D016898:Interferon-alpha / Q000096:biosynthesis; D016676:Macrophages, Alveolar / Q000276:immunology* / Q000821:virology*; D019318:Porcine Reproductive and Respiratory Syndrome / Q000235:genetics / Q000276:immunology / Q000821:virology; D019316:Porcine respiratory and reproductive syndrome virus / Q000145:classification / Q000235:genetics* / Q000276:immunology*; D034421:Sus scrofa / Q000235:genetics* / Q000276:immunology* / Q000821:virology; D013552:Swine; D051193:Toll-Like Receptors / Q000235:genetics* / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis; D014779:Virus Replication / Q000235:genetics
chemical_list: D016898:Interferon-alpha; D051193:Toll-Like Receptors; D014409:Tumor Necrosis Factor-alpha
keywords: Genotype 1 PRRSV; Innate immunity; Porcine alveolar macrophages; Toll-like receptors