Chicken immunoregulatory Ig-like receptor families: an overview and expression details on ggTREM-A1.

Paired immunoregulatory receptors facilitate the coordination of the immune response at the cellular level. In recent years, our group characterized chicken homologues to mammalian immunoregulatory Ig-like receptor families. The first part of this review focuses on the current progress on chicken immunoregulatory Ig-like receptor families. One of these receptors is gallus gallus TREM-A1, which was described as the only member of the chicken TREM family with activating potential. The second part of this review presents a study initiated to further characterize ggTREM-A1 expression. For this purpose we established real-time RT-PCR and generated a specific mab to analyze the expression profile of ggTREM-A1 on mRNA and protein level, respectively. GgTREM-A1 mRNA was predominantly expressed in macrophages, but was also detected in brain, bone marrow, bursa, thymus, spleen and PBMC. Analyzing ggTREM-A1 surface expression by mab staining validated the expression on macrophages. Additionally, we showed high expression on blood monocytes, heterophils and NK cells and on monocytes isolated from bone marrow. Moreover, we detected ggTREM-A1 protein also on thrombocytes, B and T cell subsets, but antigen expression seemed to be lower and more variable in these cells. Immunohistochemistry of chicken brain tissue, combining ggTREM-A1 mab and various markers specific for various brain cell subsets showed expression of ggTREM-A1 on microglial cells, but also on neurons, astrocytes and oligodendrocytes. In conclusion, ggTREM-A1 is expressed on a variety of cells, relevant for the immune system, possibly combining physiological function of different mammalian TREM.

Hcp and VgrG1 are secreted components of the Helicobacter hepaticus type VI secretion system and VgrG1 increases the bacterial colitogenic potential.

The enterohepatic Epsilonproteobacterium Helicobacter hepaticus persistently colonizes the intestine of mice and causes chronic inflammatory symptoms in susceptible mouse strains. The bacterial factors causing intestinal inflammation are poorly characterized. A large genomic pathogenicity island, HHGI1, which encodes components of a type VI secretion system (T6SS), was previously shown to contribute to the colitogenic potential of H. hepaticus. We have now characterized the T6SS components Hcp, VgrG1, VgrG2 and VgrG3, encoded on HHGI1, including the potential impact of the T6SS on intestinal inflammation in a mouse T-cell transfer model. The H. hepaticus T6SS components were expressed during the infection and secreted in a T6SS-dependent manner, when the bacteria were cultured either in the presence or in the absence of mouse intestinal epithelial cells. Mutants deficient in VgrG1 displayed a significantly lower colitogenic potential in T-cell-transferred C57BL/6 Rag2(-/-) mice, despite an unaltered ability to colonize mice persistently. Intestinal microbiota analyses demonstrated only minor changes in mice infected with wild-typeH. hepaticus as compared with mice infected with VgrG1-deficient isogenic bacteria. In addition, competitive assays between both wild-type and T6SS-deficient H. hepaticus, and between wild-type H. hepaticus and Campylobacter jejuni or Enterobacteriaceae species did not show an effect of the T6SS on interbacterial competitiveness. Therefore, we suggest that microbiota alterations did not play a major role in the changes of pro-inflammatory potential mediated by the T6SS. Cellular innate pro-inflammatory responses were increased by the secreted T6SS proteins VgrG1 and VgrG2. We therefore concluded that the type VI secretion component VgrG1 can modulate and specifically exacerbate the innate pro-inflammatory effect of the chronic H. hepaticus infection.

TLR7 contributes to the rapid progression but not to the overall fatal outcome of secondary pneumococcal disease following influenza A virus infection.

Increased risk for bacterial superinfections substantially contributes to the mortality caused by influenza A virus (IAV) epidemics. While the mechanistic basis for this lethal synergism is still insufficiently understood, immune modulation through the viral infection has been shown to be involved. Since the pattern-recognition receptor (PRR) toll-like receptor 7 (TLR7) is a major sensor for the viral genome, we studied how IAV recognition by TLR7 influences the development of secondary pneumococcal infection. In a mouse model of IAV, TLR7-deficient hosts induced a potent antiviral response and showed unchanged survival. In secondary pneumococcal infection during acute influenza, TLR7ko mice showed a fatal outcome similar to wild-type (WT) hosts, despite significantly delayed disease progression. Also, when bacterial superinfection occurred after virus clearance, WT and TLR7-deficient hosts showed similar mortality, even though we found the phagocytic activity of alveolar macrophages isolated from IAV-pre-infected hosts to be enhanced in TLR7ko over WT mice. Thus, we show that a virus-sensing PRR modulates the progression of secondary pneumococcal infection following IAV. However, the fatal overall outcome in WT as well as TLR7ko hosts suggests that processes distinct from TLR7-triggering override the contribution of this single PRR.

CD8(+) T cells responding to alveolar self-antigen lack CD25 expression and fail to precipitate autoimmunity.

Although the contribution of CD8(+) T cells to the pathogenesis of noncommunicable lung diseases has become increasingly appreciated, our knowledge about the mechanisms controlling self-reactive CD8(+) T cells in the respiratory tract remains largely elusive. The outcome of the encounter between pulmonary self-antigen and naive CD8(+) T cells, in the presence or absence of inflammation, was traced after adoptive transfer of fluorescence-labeled CD8(+) T cells specific for the neo-self-antigen influenza A hemagglutinin into transgenic mice expressing hemagglutinin specifically in alveolar type II epithelial cells in order: to study the outcome of alveolar antigen encounter in the steady state and under inflammatory conditions; to define the phenotype and fate of CD8(+) T cells primed in the respiratory tract; and, finally, to correlate these findings with the onset of autoimmunity in the lung. We found that CD8(+) T cells remain ignorant in the steady state, whereas transient proliferation of self-reactive CD8(+) T cells is induced by forced maturation or licensing of dendritic cells, increases in the antigenic threshold, and targeted release of alveolar self-antigen by epithelial injury. However, these cells fail to acquire effector functions, lack the expression of the high-affinity IL-2 receptor CD25, and do not precipitate autoimmunity in the lung. We conclude that inadvertent activation of CD8(+) T cells in the lung is prevented in the absence of "danger signals," whereas tissue damage after infection or noninfectious inflammation creates an environment that allows the priming of previously ignorant T cells. Failure in effector cell differentiation after abortive priming, however, precludes the establishment of self-perpetuating autoimmunity in the lung.

CD4+CD25+Foxp3+ regulatory T cells are dispensable for controlling CD8+ T cell-mediated lung inflammation.

Every person harbors a population of potentially self-reactive lymphocytes controlled by tightly balanced tolerance mechanisms. Failures in this balance evoke immune activation and autoimmunity. In this study, we investigated the contribution of self-reactive CD8(+) T lymphocytes to chronic pulmonary inflammation and a possible role for naturally occurring CD4(+)CD25(+)Foxp3(+) regulatory T cells (nTregs) in counterbalancing this process. Using a transgenic murine model for autoimmune-mediated lung disease, we demonstrated that despite pulmonary inflammation, lung-specific CD8(+) T cells can reside quiescently in close proximity to self-antigen. Whereas self-reactive CD8(+) T cells in the inflamed lung and lung-draining lymph nodes downregulated the expression of effector molecules, those located in the spleen appeared to be partly Ag-experienced and displayed a memory-like phenotype. Because ex vivo-reisolated self-reactive CD8(+) T cells were very well capable of responding to the Ag in vitro, we investigated a possible contribution of nTregs to the immune control over autoaggressive CD8(+) T cells in the lung. Notably, CD8(+) T cell tolerance established in the lung depends only partially on the function of nTregs, because self-reactive CD8(+) T cells underwent only biased activation and did not acquire effector function after nTreg depletion. However, although transient ablation of nTregs did not expand the population of self-reactive CD8(+) T cells or exacerbate the disease, it provoked rapid accumulation of activated CD103(+)CD62L(lo) Tregs in bronchial lymph nodes, a finding suggesting an adaptive phenotypic switch in the nTreg population that acts in concert with other yet-undefined mechanisms to prevent the detrimental activation of self-reactive CD8(+) T cells.

Evaluation of lumbosacral nerve root conduction in chickens by electrophysiological testing including high-resolution spinal magnetic stimulation.

The value of avian models in peripheral nerve research recently became substantiated by the immunobiological similarity of avian inflammatory demyelinating polyradiculoneuropathy to human Guillain-Barré syndrome providing an alternative animal model for experimental autoimmune neuritis. As electrophysiologic evaluation of nerve roots is essential part of the diagnosis of polyradiculoneuropathies in humans, it would be favourable to have similar research methods available for juvenile chickens. Hence, this study was performed (1) to establish a tool-set that allows for reproducible evaluation of the tibial/sciatic nerve and its nerve roots, (2) to achieve age-matched reference values, and (3) to trace the kinetics of peripheral nerve maturation within chickens. Nine chickens underwent serial electrodiagnostic examinations between the age of 6 and 15 weeks. Several methods of sensory and motor nerve fiber stimulation of the tibial/sciatic nerve were tested and modified or established. Ultimately, scalp-recorded somatosensory evoked potentials, compound muscle action potentials elicited by tibial/sciatic nerve electrical as well as spinal magnetic stimulation and motor nerve conduction velocity were available for tibial/sciatic nerve and nerve root evaluation in chickens. Base values were obtained for all investigations and parameters. Results indicated that the maturation of the nerve fibers is incomplete up to the age of 15 weeks. The methods tested here provide an excellent tool-set for quantitative tibial/sciatic nerve and nerve root assessment in avian polyradiculoneuropathies, especially within the scope of longitudinal monitoring of the disease course.

Acute paretic syndrome in juvenile White Leghorn chickens resembles late stages of acute inflammatory demyelinating polyneuropathies in humans.

BACKGROUND: Sudden limb paresis is a common problem in White Leghorn flocks, affecting about 1% of the chicken population before achievement of sexual maturity. Previously, a similar clinical syndrome has been reported as being caused by inflammatory demyelination of peripheral nerve fibres. Here, we investigated in detail the immunopathology of this paretic syndrome and its possible resemblance to human neuropathies. METHODS: Neurologically affected chickens and control animals from one single flock underwent clinical and neuropathological examination. Peripheral nervous system (PNS) alterations were characterised using standard morphological techniques, including nerve fibre teasing and transmission electron microscopy. Infiltrating cells were phenotyped immunohistologically and quantified by flow cytometry. The cytokine expression pattern was assessed by quantitative real-time PCR (qRT-PCR). These investigations were accomplished by MHC genotyping and a PCR screen for Marek's disease virus (MDV). RESULTS: Spontaneous paresis of White Leghorns is caused by cell-mediated, inflammatory demyelination affecting multiple cranial and spinal nerves and nerve roots with a proximodistal tapering. Clinical manifestation coincides with the employment of humoral immune mechanisms, enrolling plasma cell recruitment, deposition of myelin-bound IgG and antibody-dependent macrophageal myelin-stripping. Disease development was significantly linked to a 539 bp microsatellite in MHC locus LEI0258. An aetiological role for MDV was excluded. CONCLUSIONS: The paretic phase of avian inflammatory demyelinating polyradiculoneuritis immunobiologically resembles the late-acute disease stages of human acute inflammatory demyelinating polyneuropathy, and is characterised by a Th1-to-Th2 shift.