title: The innate antiviral immune system of the cat: molecular tools for the measurement of its state of activation.
abstract: The innate immune system plays a central role in host defence against viruses. While many studies portray mechanisms in early antiviral immune responses of humans and mice, much remains to be discovered about these mechanisms in the cat. With the objective of shedding light on early host-virus interactions in felids, we have developed 12 real-time TaqMan(®) qPCR systems for feline genes relevant to innate responses to viral infection, including those encoding for various IFNα and IFNω subtypes, IFNβ, intracellular antiviral factor Mx, NK cell stimulator IL-15 and effectors perforin and granzyme B, as well as Toll-like receptors (TLRs) 3 and 8. Using these newly developed assays and others previously described, we measured the relative expression of selected markers at early time points after viral infection in vitro and in vivo. Feline embryonic fibroblasts (FEA) inoculated with feline leukemia virus (FeLV) indicated peak levels of IFNα, IFNβ and Mx expression already 6h after infection. In contrast, Crandell-Rees feline kidney (CrFK) cells inoculated with feline herpes virus (FHV) responded to infection with high levels of IFNα and IFNβ only after 24h, and no induction of Mx could be detected. In feline PBMCs challenged in vitro with feline immunodeficiency virus (FIV), maximal expression levels of IFNα, β and ω subtype genes as well as IL-15 and TLRs 3, 7 and 8 were measured between 12 and 24h after infection, whereas expression levels of proinflammatory cytokine gene IL-6 were consistently downregulated until 48h post inoculation. A marginal upregulation of granzyme B was also observed within 3h after infection. In an in vivo experiment, cats challenged with FIV exhibited a 2.4-fold increase in IFNα expression in blood 1 week post infection. We furthermore demonstrate the possibility of stimulating feline immune cells in vitro with various immune response modifiers (IRMs) already known for their immunostimulatory properties in mice and humans, namely Poly IC, Resiquimod (R-848) and dSLIM™, a synthetic oligonucleotide containing several unmethylated CpG motifs. Stimulation of feline PBMCs with dSLIM™ and R-848 effectively enhanced expression of IFNα within 12h by factors of 6 and 12, respectively, and Poly IC induced an increase in Mx mRNA expression of 28-fold. Altogether, we describe new molecular tools and their successful use for the characterization of innate immune responses against viruses in the cat and provide evidence that feline cells can be stimulated by synthetic molecules to enhance their antiviral defence mechanisms.
authors: Clinical Laboratory, Vetsuisse Faculty, University of Zurich, Winterthurerstr. 260, CH-8057 Zurich, Switzerland. crobert@vetclinics.uzh.ch
authors: Céline
authors: C
authors: Robert-Tissot
authors: Vera L
authors: VL
authors: Rüegger
authors: Valentino
authors: V
authors: Cattori
authors: Marina L
authors: ML
authors: Meli
authors: Barbara
authors: B
authors: Riond
authors: Maria Alice
authors: MA
authors: Gomes-Keller
authors: Andrea
authors: A
authors: Vögtlin
authors: Burghardt
authors: B
authors: Wittig
authors: Christiane
authors: C
authors: Juhls
authors: Regina
authors: R
authors: Hofmann-Lehmann
authors: Hans
authors: H
authors: Lutz
mesh_terms: D000818:Animals; D002371:Cat Diseases / Q000276:immunology / Q000821:virology*; D002415:Cats / Q000276:immunology* / Q000821:virology; D002460:Cell Line / Q000821:virology; D016181:Feline Acquired Immunodeficiency Syndrome / Q000276:immunology; D007113:Immunity, Innate / Q000235:genetics*; D016089:Immunodeficiency Virus, Feline / Q000276:immunology; D007372:Interferons / Q000235:genetics; D007963:Leukocytes, Mononuclear / Q000276:immunology / Q000821:virology; D008297:Male; D060888:Real-Time Polymerase Chain Reaction / Q000662:veterinary
chemical_list: D007372:Interferons