PMC:3537549 / 1929-8398 JSONTXT

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In order to adapt to the solitary way of life of ancestral felids, these pathogens have acquired elaborate means to persist within their host population over the course of time. The infection of new hosts upon the rare contact between individuals was evolutionarily assured by very efficient transmission strategies and the induction of latent, chronic and/or asymptomatic infections. Often kept in multi-cat households and placed in catteries or shelters, today’s domestic cat is consequently particularly susceptible to viral infections[1, 2]. The close proximity and high social contact rate of animals with different vaccination states in these stressful environments further increases the risk of infection. Moreover, the strong antigenic variability of several common feline viruses including the feline calicivirus (FCV), the feline coronavirus (FCoV) and the feline leukemia virus (FeLV) support escape from immune responses and lower the efficacy of currently available vaccines[3]. The availability of complementary prophylactic strategies could help protect pet cats in environments with high infectious pressure from long diseases and/or death caused by infections with fatal viruses.\nA promising addition to vaccination is the manipulation of innate immunity. Innate pathogen recognition relies on a set of sensory molecules, the Toll-like receptors (TLRs), which enable the immediate reaction of specific immune cells to pathogen “danger signals”, the so-called pathogen-associated molecular patterns (PAMPs)[4]. Due to their abundance in all bacterial as well as some viral genomes, oligodeoxynucleotides (ODN) containing unmethylated cytosine–phosphate–guanosine (CpG) motifs are effectively recognized as PAMPs by the vertebrate innate immune system[5]. Response to CpG ODN stimulation is conferred through TLR9, expressed mainly in the intracellular compartments of human B cells and plasmacytoid dendritic cells (pDCs)[6, 7]. Alarmed TLR9 is the initial instigator of gene expression profiles that strongly support antiviral mechanisms: upregulation of costimulatory molecules major histocompatibility complex (MHC) II, B7.1 and B7.2 on the surface of stimulated cells provides them with a stronger antigen presenting potential[8, 9] and production of cytokines such as type I interferon (IFN), interleukin (IL)-12, IFNγ, IL-6 and tumor necrosis factor (TNF)α, contribute to providing an optimal immune environment for the development of innate and adaptive responses against intracellular pathogens[10, 11]. Probably the most important antiviral property of CpG ODN resides in their potential to stimulate the production of high amounts of type I IFN by pDCs[12]. This family of cytokines, which includes IFNα, IFNω and IFNβ, has been shown to considerably enhance natural killer (NK) cell cytotoxicity[13], promote differentiation, maturation and immunostimulatory functions of monocytes and DCs[14], induce B cell production of immunoglobulin[15] and T-helper (Th)1 differentiation of T cells[16, 17]. Moreover, upon binding to their ubiquitously distributed receptor, type I IFNs effectively induce the synthesis of various intracellular proteins which interfere with the replication of a broad range of viruses[18]. The myxovirus-resistance protein (Mx) GTPase is a well-studied example of these intracellular antiviral effectors. This enzyme is known to be directly regulated by the type I IFN, and its detection is readily used as marker for upregulation and biological activity of this cytokine family[19].\nDistinct classes of ODN have been shown to induce differential immune responses[20]. Class A CpG ODN (CpG-A) consist of CpG motifs in a phosphodiester core, flanked on both ends by phosphorothioate poly (G) sequences. CpG ODN of this class are characterized by their potential to both induce massive type I IFN secretion by pDCs[12] and increase NK cytotoxicity[21], rendering them ideal candidates as prophylactic enhancers of innate immunity to viral infections. Conversely, class B CpG ODN (CpG-B), which encode multiple CpG motifs on a phosphorothioate backbone, promote monocyte maturation and B cell activation, thus substantially supporting the development of humoral immune responses[22–24]. Both CpG-A and CpG-B have indicated immunostimulatory properties in immune cells of mice, primates and many domestic species in vitro[25–31], while in vivo studies in outbred animals have mainly been carried out with CpG-B[24, 32, 33]. Concerning the cat, specifically synthesized CpG-A molecules were recently shown to induce IFNγ[34]; CpG-B molecules indicated the capability to induce proliferation of feline cells[35], and CpG-B-adjuvanted allergen indicated potential for immunotherapy in a feline asthma model[36]. Although class C[37, 38] and class P ODN[39] were developed in more recent years in an effort to combine the advantageous effects of both CpG-A and CpG-B and increase immunogenicity, CpG-A remain the strongest inducers of type I IFN described to date.\nThe potential of CpG ODN as prophylactic stand-alone inducers of innate defense mechanisms has been the subject of only few studies. Most works in this field initially described protection of mice against bacterial[40–44] and parasitic[45–47] infections. More recently, induction of resistance to viral infections was shown also in mouse models for Herpes Simplex Virus[48], neurotropic arenavirus[49], foot and mouth disease virus[50] and Vaccinia virus[51]. With exception of the latter, all these studies were carried out with CpG-B. In an outbred species, partial antiviral protection has only been described in two studies so far, in which reduced shedding of herpes and parainfluenza viruses was observed in lambs after administration also of a CpG-B[52, 53]. To our knowledge, prophylactic antiviral potential of CpG-A has not yet been described in outbred animals.\nWe carried out a series of experiments with the objective to characterize both the immunomodulatory and antiviral properties of CpG-A in the domestic cat. In vitro, the prototype of CpG-A, ODN 2216[12], strengthened the antiviral qualities of feline immune cells and stimulated their production of soluble molecules that inhibited the replication of five different families of viruses: Coronaviridae, Herpesviridae, Caliciviridae, Parvoviridae, and Retroviridae. In vivo, ODN 2216 induced a systemic antiviral state."}