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1.
Vet Pathol ; 59(1): 132-137, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34490804

RESUMO

Canine multiple system degeneration (CMSD) is a progressive hereditary neurodegenerative disorder commonly characterized by neuronal degeneration and loss in the cerebellum, olivary nuclei, substantia nigra, and caudate nuclei. In this article, we describe 3 cases of CMSD in Ibizan hounds. All patients exhibited marked cerebellar ataxia and had cerebellar atrophy on magnetic resonance imaging. At necropsy, all cases showed varying degrees of cerebellar atrophy, and 2 cases had gross cavitation of the caudate nuclei. Histologic findings included severe degeneration and loss of all layers of the cerebellum and neuronal loss and degeneration within the olivary nuclei, substantia nigra, and caudate nuclei. Pedigree analysis indicated an autosomal recessive mode of inheritance, but the causative gene in this breed is yet to be identified. CMSD resembles human multiple system atrophy and warrants further investigation.


Assuntos
Doenças do Cão , Doenças Neurodegenerativas , Animais , Autopsia/veterinária , Cruzamento , Cerebelo/diagnóstico por imagem , Doenças do Cão/diagnóstico , Doenças do Cão/genética , Cães , Humanos , Doenças Neurodegenerativas/veterinária
2.
J Virol ; 86(9): 4793-800, 2012 May.
Artigo em Inglês | MEDLINE | ID: mdl-22379096

RESUMO

Cache Valley virus-induced malformations have been previously reproduced in ovine fetuses; however, no studies have established the course of infection of cells and tissues with Cache Valley virus. To address these questions, ovine fetuses at 35 days of gestation were inoculated in utero with Cache Valley virus and euthanized at 7, 10, 14, 21, and 28 days postinfection. On postmortem examination, arthrogryposis and oligohydramnios were observed in some infected fetuses. Morphological studies showed necrosis in the central nervous system and skeletal muscle of infected fetuses evaluated after 7 to 14 days postinfection, and hydrocephalus, micromyelia, and muscular loss were observed in infected fetuses after 21 to 28 days postinfection. Using immunohistochemistry and in situ hybridization, intense Cache Valley virus antigen and RNA staining was detected in the brain, spinal cord, skeletal muscle, and, to a lesser degree, in fetal membranes and other tissues of infected fetuses. Viral antigen and RNA staining decreased in targeted and infected tissues with the progression of the infection.


Assuntos
Vírus Bunyamwera , Infecções por Bunyaviridae/veterinária , Doenças dos Ovinos/virologia , Animais , Antígenos Virais/imunologia , Antígenos Virais/metabolismo , Vírus Bunyamwera/imunologia , Vírus Bunyamwera/isolamento & purificação , Infecções por Bunyaviridae/virologia , Sistema Nervoso Central/patologia , Córtex Cerebral/patologia , Doenças Fetais/veterinária , Doenças Fetais/virologia , Músculo Esquelético/patologia , Testes de Neutralização , RNA Viral/metabolismo , Ovinos , Medula Espinal/patologia
3.
Vet Immunol Immunopathol ; 211: 64-74, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31084897

RESUMO

S100A12 and S100A8/A9 (calprotectin) are released from activated mononuclear cells and belong to the group of damage associated molecular patterns. Fecal S100A12 and S100A8/A9 concentrations have been suggested as biomarkers of intestinal inflammation in dogs with chronic inflammatory enteropathies (CIE). However, the mucosal cellular infiltrate in dogs with CIE is primarily lymphocytic-plasmacytic. Whether fecal S100A12 and S100A8/A9 levels reflect the number and/or activity of intestinal mucosal mononuclear cells, or whether these proteins are also produced by other cells has not been investigated. Thus, the aim of this study was to evaluate intestinal mucosal S100A12 and S100A8/A9 positivity and a potential relationship with the respective protein concentrations in serum and fecal samples in dogs with CIE. Serum (single sample), fecal samples (from 3 consecutive days), and gastrointestinal tissue biopsies (i.e., stomach, duodenum, ileum, and colon) were evaluated from 21 dogs with CIE. Serum and fecal S100A12 and S100A8/A9 concentrations were measured by analytically validated in-house ELISAs. Tissue biopsies underwent routine histopathology and immunohistochemical evaluation for S100A12 and S100A8/A9 positivity (S100A12+ and S100A8/A9+, each recorded as positive cells/mm2). S100A12+ and S100A8/A9+ cells were identified in all segments of the gastrointestinal tract, but were predominantly localized in the lamina propria (LP). Duodenal LP S100A12 positivity correlated statistically significantly with that in the stomach and ileum (ρ = 0.66 and 0.69, both p < 0.01), but was inversely correlated with the severity of macrophage infiltration in the duodenum (ρ=-0.47, p = 0.042). Ileal LP S100A8/A9 positivity correlated positively with the extent of ileal neutrophil and macrophage infiltration (ρ=0.61, p = 0.047). Fecal S100A12 concentrations strongly correlated with the number of S100A12+ cells along the entire gastrointestinal tract (ρ = 0.76, p = 0.028), whereas serum S100A12 concentrations were inversely correlated to colonic S100A12+ cell counts (ρ=-0.50, p = 0.043). Mucosal S100A8/A9+ cell counts were not associated with the corresponding fecal or serum S100A8/A9 concentrations. These results suggest that the intestinal mucosa in dogs with CIE contains an increased number of activated (pro-inflammatory) phagocytes expressing and secreting the S100A12 protein, but the macrophage population seen on routine histopathology is predominantly mature (anti-inflammatory) with a reduced or absent expression of S100A12 and a normal or increased expression of S100A8/A9. However, the distribution of intestinal S100A8/A9 expression requires further study.


Assuntos
Doenças do Cão/imunologia , Doenças Inflamatórias Intestinais/veterinária , Mucosa Intestinal/metabolismo , Complexo Antígeno L1 Leucocitário/metabolismo , Proteína S100A12/metabolismo , Animais , Biomarcadores/análise , Biomarcadores/sangue , Doenças do Cão/sangue , Cães , Fezes/química , Feminino , Doenças Inflamatórias Intestinais/sangue , Doenças Inflamatórias Intestinais/imunologia , Complexo Antígeno L1 Leucocitário/análise , Complexo Antígeno L1 Leucocitário/sangue , Masculino , Neutrófilos/imunologia , Proteína S100A12/análise , Proteína S100A12/sangue
4.
J Histochem Cytochem ; 56(4): 359-70, 2008 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18071063

RESUMO

Arteries undergo marked structural and functional changes in human and experimental hypertension that generally involve smooth muscle cell (SMC) hypertrophy/hyperplasia as well as abnormal extracellular matrix turnover. In this study we examined time courses of changes in SMC activity and matrix protein content in a novel mini-pig aortic coarctation model. Cell proliferation was evaluated by immunostaining of Ki-67, apoptosis was assessed by TUNEL, and phenotypic changes were monitored by immunostaining three SMC contractile markers (caldesmon, calponin, and smoothelin). Changes in medial collagen and elastin were examined by picrosirius red and Verhoeff-van Gieson staining, respectively. LabVIEW-based image analysis routines were developed to objectively and efficiently quantify the (immuno)histochemical results. We found that significant cell proliferation and matrix production occurred in the early stages of this coarctation model and then declined gradually; the SMCs also tended to exhibit a less contractile phenotype following these cellular and extracellular changes. Specifically, different aspects of the phenotypic changes associated with hypertension occurred at different rates: cell proliferation and collagen production occurred early and peaked by 2 weeks, whereas changes in contractile protein expression continued to decrease over the entire 8-week study period. Temporal changes found in this study emphasize the importance of simultaneously tracing time courses of SMC growth and differentiation as well as matrix protein production and content. SMCs are multifunctional, and caution must be used to not overdefine phenotype. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials.


Assuntos
Aorta/patologia , Hipertensão/patologia , Miócitos de Músculo Liso/patologia , Actinas/metabolismo , Animais , Aorta/metabolismo , Apoptose , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ligação a Calmodulina/metabolismo , Proliferação de Células , Colágeno/metabolismo , Interpretação Estatística de Dados , Proteínas da Matriz Extracelular/metabolismo , Hipertensão/metabolismo , Processamento de Imagem Assistida por Computador , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Antígeno Ki-67/metabolismo , Proteínas dos Microfilamentos/metabolismo , Proteínas Musculares/metabolismo , Miócitos de Músculo Liso/metabolismo , Software , Suínos , Porco Miniatura , Fatores de Tempo , Calponinas
5.
Vet Dermatol ; 19(2): 52-8, 2008 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18336421

RESUMO

Dermatomyositis (DM) is a canine and human inflammatory disease of the skin and muscle that is thought to be autoimmune in nature. In dogs, DM occurs most often in the rough collie and Shetland sheepdog. Characteristic skin lesions typically develop on the face, ears, tail, and distal extremities. The severity of lesions varies and is thought to increase with stressful stimuli. Previous studies in the collie suggest that DM is inherited in an autosomal dominant fashion with incomplete penetrance. The work presented here concerns gene transcripts profiling and immunobiology of DM in the Shetland sheepdog. Gene transcript profiles were generated for affected and normal skin using a canine-specific oligonucleotide array having 49,929 probe sets. Two-hundred and eight-five gene transcripts, many of which are involved in immune function, were found to be differentially regulated in these tissues. Also reported are Western blot, immunohistochemistry, and immunofluorescence analyses which showed that staining patterns with sera from normal and affected dogs are quite similar. While our work suggests that canine DM is a disease that may be immune mediated, it did not detect the production of specific disease-associated autoantibodies.


Assuntos
Dermatomiosite/veterinária , Doenças do Cão/genética , Perfilação da Expressão Gênica/veterinária , Análise de Sequência com Séries de Oligonucleotídeos/veterinária , Animais , Western Blotting/veterinária , Dermatomiosite/genética , Dermatomiosite/imunologia , Dermatomiosite/patologia , Doenças do Cão/imunologia , Doenças do Cão/patologia , Cães , Feminino , Imuno-Histoquímica/veterinária , Masculino , Linhagem , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária
6.
PLoS One ; 12(1): e0170504, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28107431

RESUMO

Lack of safe and effective adjuvants is a major hindrance to the development of efficacious vaccines. Signaling via CD40 pathway leads to enhanced antigen processing and presentation, nitric oxide expression, pro-inflammatory cytokine expression by antigen presenting cells, and stimulation of B-cells to undergo somatic hypermutation, immunoglobulin class switching, and proliferation. Agonistic anti-CD40 antibodies have shown promising adjuvant qualities in human and mouse vaccine studies. An anti-CD40 monoclonal antibody (mAb), designated 2E4E4, was identified and shown to have strong agonistic effects on primary cells from multiple livestock species. The mAb recognize swine, bovine, caprine, and ovine CD40, and evoked 25-fold or greater proliferation of peripheral blood mononuclear cells (PBMCs) from these species relative to cells incubated with an isotype control (p<0.001). In addition, the mAb induced significant nitric oxide (p<0.0001) release by bovine macrophages. Furthermore, the mAb upregulated the expression of MHC-II by PBMCs, and stimulated significant (p<0.0001) IL-1α, IL6, IL-8, and TNF-α expression by PBMCs. These results suggest that the mAb 2E4E4 can target and stimulate cells from multiple livestock species and thus, it is a potential candidate for adjuvant development. This is the first study to report an anti-swine CD40 agonistic mAb that is also broadly reactive against multiple species.


Assuntos
Adjuvantes Imunológicos/uso terapêutico , Anticorpos Monoclonais/imunologia , Antígenos CD40/imunologia , Animais , Bovinos/imunologia , Reações Cruzadas/imunologia , Citocinas/metabolismo , Feminino , Citometria de Fluxo , Genes MHC da Classe II/imunologia , Cabras/imunologia , Células HEK293 , Humanos , Leucócitos Mononucleares/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Ovinos/imunologia , Suínos/imunologia
7.
Vet Clin Pathol ; 45(1): 135-47, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26765807

RESUMO

BACKGROUND: Canine S100 calcium-binding protein A12 (cS100A12) shows promise as biomarker of inflammation in dogs. A previously developed cS100A12-radioimmunoassay (RIA) requires radioactive tracers and is not sensitive enough for fecal cS100A12 concentrations in 79% of tested healthy dogs. An ELISA assay may be more sensitive than RIA and does not require radioactive tracers. OBJECTIVE: The purpose of the study was to establish a sandwich ELISA for serum and fecal cS100A12, and to establish reference intervals (RI) for normal healthy canine serum and feces. METHODS: Polyclonal rabbit anti-cS100A12 antibodies were generated and tested by Western blotting and immunohistochemistry. A sandwich ELISA was developed and validated, including accuracy and precision, and agreement with cS100A12-RIA. The RI, stability, and biologic variation in fecal cS100A12, and the effect of corticosteroids on serum cS100A12 were evaluated. RESULTS: Lower detection limits were 5 µg/L (serum) and 1 ng/g (fecal), respectively. Intra- and inter-assay coefficients of variation were ≤ 4.4% and ≤ 10.9%, respectively. Observed-to-expected ratios for linearity and spiking recovery were 98.2 ± 9.8% (mean ± SD) and 93.0 ± 6.1%, respectively. There was a significant bias between the ELISA and the RIA. The RI was 49-320 µg/L for serum and 2-484 ng/g for fecal cS100A12. Fecal cS100A12 was stable for 7 days at 23, 4, -20, and -80°C; biologic variation was negligible but variation within one fecal sample was significant. Corticosteroid treatment had no clinically significant effect on serum cS100A12 concentrations. CONCLUSIONS: The cS100A12-ELISA is a precise and accurate assay for serum and fecal cS100A12 in dogs.


Assuntos
Doenças do Cão/sangue , Ensaio de Imunoadsorção Enzimática/veterinária , Proteína S100A12/análise , Animais , Cães , Fezes/química , Feminino , Masculino , Coelhos , Valores de Referência , Reprodutibilidade dos Testes , Proteína S100A12/sangue
8.
Neurosci Lett ; 553: 153-8, 2013 Oct 11.
Artigo em Inglês | MEDLINE | ID: mdl-23973300

RESUMO

MicroRNAs (miRs) are endogenous small RNAs that regulate gene expression at the post-transcriptional level by mediating mRNA degradation or transcriptional inhibition. MiRs were implicated in the pathogenesis of numerous neurodegenerative diseases, including Parkinson's disease (PD). In this study we analyzed the possible role of miRs in the neurodegenerative process in a spontaneous autosomal recessive rat model for neurodegeneration developed in our laboratory. To investigate the role of miRs in the etiology of PD, we conducted miR expression profiling using microarrays. We found 20 miRs that are deregulated in affected rats and many of these are implicated in neurodegenerative disease, including PD. In this study we were particularly interested in the expression of miR-132, a miR that has been reported to be highly expressed in neurons, and to have a potential role in neurodegenerative diseases. We found a significant increase in miR-132 in affected rats by microarray and the result was confirmed by qPCR. Next we analyzed one of the known downstream targets of miR-132, nuclear receptor related 1 protein (Nurr1), which is essential in neurogenesis of midbrain dopaminergic neurons. Western blot analysis and immunohistochemistry revealed a significant decrease in Nurr1 protein expression in the mesencephalic neurons. Finally, we found a significant decrease in both serum and mesencephalon brain tissue of brain-derived neurotrophic factor (BDNF), which is known to be a direct target of Nurr1. Taken together, our findings suggest that miR-132 can regulate Nurr1 levels and might influence the development and function of midbrain dopaminergic neurons.


Assuntos
MicroRNAs/metabolismo , Degeneração Neural/metabolismo , Animais , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Mesencéfalo/metabolismo , Membro 2 do Grupo A da Subfamília 4 de Receptores Nucleares/metabolismo , Ratos
9.
Mol Biol Cell ; 24(12): 1918-32, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23615443

RESUMO

Desmin intermediate filaments (DIFs) form an intricate meshwork that organizes myofibers within striated muscle cells. The mechanisms that regulate the association of desmin to sarcomeres and their role in desminopathy are incompletely understood. Here we compare the effect nebulin binding has on the assembly kinetics of desmin and three desminopathy-causing mutant desmin variants carrying mutations in the head, rod, or tail domains of desmin (S46F, E245D, and T453I). These mutants were chosen because the mutated residues are located within the nebulin-binding regions of desmin. We discovered that, although nebulin M160-164 bound to both desmin tetrameric complexes and mature filaments, all three mutants exhibited significantly delayed filament assembly kinetics when bound to nebulin. Correspondingly, all three mutants displayed enhanced binding affinities and capacities for nebulin relative to wild-type desmin. Electron micrographs showed that nebulin associates with elongated normal and mutant DIFs assembled in vitro. Moreover, we measured significantly delayed dynamics for the mutant desmin E245D relative to wild-type desmin in fluorescence recovery after photobleaching in live-cell imaging experiments. We propose a mechanism by which mutant desmin slows desmin remodeling in myocytes by retaining nebulin near the Z-discs. On the basis of these data, we suggest that for some filament-forming desmin mutants, the molecular etiology of desminopathy results from subtle deficiencies in their association with nebulin, a major actin-binding filament protein of striated muscle.


Assuntos
Desmina/metabolismo , Filamentos Intermediários/metabolismo , Proteínas Musculares/metabolismo , Miofibrilas/metabolismo , Animais , Ligação Competitiva , Células Cultivadas , Embrião de Galinha , Citoesqueleto/metabolismo , Desmina/genética , Desmina/ultraestrutura , Cães , Eletroforese em Gel de Poliacrilamida , Recuperação de Fluorescência Após Fotodegradação , Humanos , Filamentos Intermediários/genética , Filamentos Intermediários/ultraestrutura , Cinética , Microscopia Confocal , Microscopia Eletrônica de Transmissão , Proteínas Musculares/genética , Proteínas Musculares/ultraestrutura , Mutação , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , Miofibrilas/ultraestrutura , Ligação Proteica , Sarcômeros/metabolismo
10.
Exp Dermatol ; 14(7): 481-90, 2005 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15946235

RESUMO

The purpose of this study is to reproduce in vitro a recessive keratinization defect of Norfolk terrier dogs characterized by a lack of keratin 10 (K10) production. Keratinocytes from skin biopsy samples of four normal dogs and two affected dogs were cultured organotypically with growth factor-supplemented media in order to stimulate cornification. The cultured epidermis from the normal dogs closely resembled the normal epidermis in vivo and cornified. The cultured epidermis from the affected dogs displayed many phenotypic alterations identified in skin biopsies from dogs with this heritable defect. Immunohistochemistry and immunoblotting showed a marked decrease in K10 from the cultures of the affected keratinocytes, compared to that in K10 from the cultures of the normal keratinocytes. Real-time reverse transcription polymerase chain reaction quantitation showed a 31-fold decrease in K10, a 1.75-fold increase in K1 and a 136-fold increase in K2e between the affected and the normal epidermis. Organotypic keratinocytes showed a 241-fold decrease in K10, a 31-fold decrease in K1 and a 1467-fold decrease in K2e between the affected and normal cultures. Although in vitro keratin expression did not precisely simulate in vivo, the morphology of the normal and the affected epidermis was largely preserved; thus, this culture system may provide an alternative to in vivo investigations for cutaneous research involving cornification.


Assuntos
Doenças do Cão/patologia , Queratinas/deficiência , Queratinas/genética , Dermatopatias Genéticas/veterinária , Animais , Sequência de Bases , Técnicas de Cultura de Células , Células Cultivadas , DNA/genética , Doenças do Cão/genética , Doenças do Cão/metabolismo , Cães , Queratina-10 , Queratinócitos/metabolismo , Queratinócitos/patologia , Queratinas/metabolismo , Fenótipo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Dermatopatias Genéticas/genética , Dermatopatias Genéticas/metabolismo , Dermatopatias Genéticas/patologia
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