Rift Valley fever virus detection in susceptible hosts with special emphasis in insects.

Rift Valley fever phlebovirus (RVFV, Phenuiviridae) is an emerging arbovirus that can cause potentially fatal disease in many host species including ruminants and humans. Thus, tools to detect this pathogen within tissue samples from routine diagnostic investigations or for research purposes are of major interest. This study compares the immunohistological usefulness of several mono- and polyclonal antibodies against RVFV epitopes in tissue samples derived from natural hosts of epidemiologic importance (sheep), potentially virus transmitting insect species (Culex quinquefasciatus, Aedes aegypti) as well as scientific infection models (mouse, Drosophila melanogaster, C6/36 cell pellet). While the nucleoprotein was the epitope most prominently detected in mammal and mosquito tissue samples, fruit fly tissues showed expression of glycoproteins only. Antibodies against non-structural proteins exhibited single cell reactions in salivary glands of mosquitoes and the C6/36 cell pellet. However, as single antibodies exhibited a cross reactivity of varying degree in non-infected specimens, a careful interpretation of positive reactions and consideration of adequate controls remains of critical importance. The results suggest that primary antibodies directed against viral nucleoproteins and glycoproteins can facilitate RVFV detection in mammals and insects, respectively, and therefore will allow RVFV detection for diagnostic and research purposes.

Canine Central Nervous System Neoplasm Phenotyping Using Tissue Microarray Technique.

Tissue microarrays (TMAs) represent a useful technique for the simultaneous phenotyping of large sample numbers and are particularly suitable for histopathologic tumor research. In this study, TMAs were used to evaluate semiquantitatively the expression of multiple antigens in various canine central nervous system (CNS) neoplasms and to identify markers with potential discriminative diagnostic relevance. Ninety-seven canine CNS neoplasms, previously diagnosed on hematoxylin and eosin sections according to the World Health Organization classification, were investigated on TMAs, with each tumor consisting of 2 cylindrical samples from the center and the periphery of the neoplasm. Tumor cells were phenotyped using a panel of 28 monoclonal and polyclonal antibodies, and hierarchical clustering analysis was applied to group neoplasms according to similarities in their expression profiles. Hierarchical clustering generally grouped cases with similar histologic diagnoses; however, gliomas especially exhibited a considerable heterogeneity in their positivity scores. Multiple tumor groups, such as astrocytomas and oligodendrogliomas, significantly differed in the proportion of positive immunoreaction for certain markers such as p75NTR, AQP4, GFAP, and S100 protein. The study highlights AQP4 and p75NTR as novel markers, helping to discriminate between canine astrocytoma and oligodendroglioma. Furthermore, the results suggest that p75NTR and proteolipid protein may represent useful markers, whose expression inversely correlates with malignant transformation in canine astrocytomas and oligodendrogliomas, respectively. Tissue microarray was demonstrated to be a useful and time-saving tool for the simultaneous immunohistochemical characterization of multiple canine CNS neoplasms. The present study provides a detailed overview of the expression patterns of different types of canine CNS neoplasms.

Associated occurrence of p75 neurotrophin receptor expressing aldynoglia and microglia/macrophages in long term organotypic murine brain slice cultures.

Growth-promoting aldynoglia, characterized by the expression of the prototype immature Schwann cell marker p75 neurotrophin receptor (NTR) have been shown to occur in some demyelinating diseases. However, the mechanisms determining the emergence and fate of such cells are largely unknown. This study aimed at the identification of such cells and potential triggering factors using an in vitro slice culture approach. Organotypic cerebrum and brain stem slices of adult mice were cultivated for up to 18 days in vitro. Immunohistochemistry for the detection of p75(NTR), CD107b, periaxin, growth associated protein (GAP)-43, and glial fibrillary acidic protein (GFAP) was performed. The results for p75(NTR) were substantiated by the use of in situ hybridization. Cultivation was associated with a progressively increasing spontaneous occurrence of bi- to multipolar p75(NTR)-positive, but periaxin-negative glia, indicative of aldynoglial Schwann cell like cells. Similar cells stained intensely positive for GAP-43, a marker for non-myelinating Schwann cells. The number of p75(NTR) positive glia did not correlate with GFAP expression, but showed a strong correlation with a remarkable spontaneous response of CD107b positive phagocytic microglia/macrophages. Moreover, aldynoglial p75(NTR) immunoreactivity negatively correlated to neuronal p75(NTR) expression, which was lost during culturing. The present results demonstrate that the cultivation of organotypic murine brain slices is accompanied by a spontaneous response of both microglia/macrophages and p75(NTR) positive cells, suggestive of Schwann cell like aldynoglia. The findings highlights the role of microglia/macrophages, which seem to be an important triggering factor, facilitating the occurrence of this unique type of macroglia.

The role of pro- and anti-inflammatory cytokines in the pathogenesis of spontaneous canine CNS diseases.

Dogs are comparatively frequently affected by various spontaneously occurring inflammatory and degenerative central nervous system (CNS) conditions, and immunopathological processes are a hallmark of the associated neuropathology. Due to the low regenerative capacity of the CNS a sophisticated understanding of the underlying molecular basis for disease initiation, progression and remission in canine CNS diseases represents a prerequisite for the development of novel therapeutical approaches. In addition, as many spontaneous canine CNS diseases share striking similarities with their human counterpart, knowledge about the immune pathogenesis may in part be translated for a better understanding of certain human diseases. In addition to cytokine-driven differentiation of peripheral leukocytes including different subsets of T cells recent research suggests a pivotal role of these mediators also in phenotype polarization of resident glial cells. Cytokines thus represent the key mediators of the local and systemic immune response in CNS diseases and their orchestration significantly decides on either lesion progression or remission. The aim of the present review is to summarize the growing number of data focusing on the molecular basis of the immune response during spontaneous canine CNS diseases and to detail the effect of cytokines on the immune pathogenesis of selected idiopathic, infectious, and traumatic canine CNS diseases. Steroid-responsive meningitis arteritis (SRMA) represents a unique idiopathic disease of leptomeningeal blood vessels characterized by excessive IgA secretion into the cerebrospinal fluid. Recent reports have given sophisticated insights into the cytokine-driven, immune-mediated pathogenesis of SRMA that is characterized by a biased T helper 2 cell response. Canine distemper associated leukoencephalitis represents an important spontaneously occurring disease that allows investigations on the basic pathogenesis of immune-mediated myelin loss. It is characterized by an early virus-induced up-regulation of pro-inflammatory cytokines with chronic bystander immune-mediated demyelinating processes. Lastly, canine spinal cord injury (SCI) shares many similarities with the human counterpart and most commonly results from intervertebral disk disease. The knowledge of its pathogenesis is largely restricted to experimental studies in rodents, and the impact of immune processes that accompany secondary injury is discussed controversially. Recent investigations on canine SCI highlight the pivotal role of pro-inflammatory cytokine expression that is paralleled by a dominating reaction of microglia/macrophages potentially indicating a polarization of these immune cells into a neurotoxic and harmful phenotype. This report will review the role of cytokines in the immune processes of the mentioned representative canine CNS diseases and highlight the importance of cytokine/cytokine interaction as a useful therapeutic target in canine CNS diseases.

Immunohistochemical characterization of inflammatory and glial responses in a case of necrotizing leucoencephalitis in a French bulldog.

A 3-year-old male French bulldog was presented with blindness, staggering and ataxia and was humanely destroyed due to worsening of the neurological signs. At post-mortem examination a non-suppurative leucoencephalitis with extensive malacia within the forebrain was found. In addition, a bilateral necrotizing optic neuritis and focal retinitis was detected. Immunohistochemistry revealed a CD3(+) T-cell dominated inflammatory response with intralesional reactive astrocytes expressing glial fibrillary acidic protein. Astroglia-like cells expressing vimentin, which is characteristic of immature astrocytes, were found within the malacic lesions. The pathological findings are similar to those described in idiopathic necrotizing leucoencephalitis (NLE) of Yorkshire terriers and substantiate the hypothesis that NLE is not a breed-specific disorder that exclusively affects Yorkshire terriers, but also the French bulldog.