RESUMO
Structural plasticity is critical for the functional diversity of neurons in the brain. Experimental autoimmune encephalomyelitis (EAE) is the most commonly used model for multiple sclerosis (MS), successfully mimicking its key pathological features (inflammation, demyelination, axonal loss, and gliosis) and clinical symptoms (motor and non-motor dysfunctions). Recent studies have demonstrated the importance of synaptic plasticity in EAE pathogenesis. In the present study, we investigated the features of behavioral alteration and hippocampal structural plasticity in EAE-affected mice in the early phase (11 days post-immunization, DPI) and chronic phase (28 DPI). EAE-affected mice exhibited hippocampus-related behavioral dysfunction in the open field test during both early and chronic phases. Dendritic complexity was largely affected in the cornu ammonis 1 (CA1) and CA3 apical and dentate gyrus (DG) subregions of the hippocampus during the chronic phase, while this effect was only noted in the CA1 apical subregion in the early phase. Moreover, dendritic spine density was reduced in the hippocampal CA1 and CA3 apical/basal and DG subregions in the early phase of EAE, but only reduced in the DG subregion during the chronic phase. Furthermore, mRNA levels of proinflammatory cytokines ( Il1ß, Tnfα, and Ifnγ) and glial cell markers ( Gfap and Cd68) were significantly increased, whereas the expression of activity-regulated cytoskeleton-associated protein (ARC) was reduced during the chronic phase. Similarly, exposure to the aforementioned cytokines in primary cultures of hippocampal neurons reduced dendritic complexity and ARC expression. Primary cultures of hippocampal neurons also showed significantly reduced extracellular signal-regulated kinase (ERK) phosphorylation upon treatment with proinflammatory cytokines. Collectively, these results suggest that autoimmune neuroinflammation alters structural plasticity in the hippocampus, possibly through the ERK-ARC pathway, indicating that this alteration may be associated with hippocampal dysfunctions in EAE.
Assuntos
Encefalomielite Autoimune Experimental , Esclerose Múltipla , Doenças dos Roedores , Camundongos , Animais , Esclerose Múltipla/metabolismo , Esclerose Múltipla/patologia , Esclerose Múltipla/veterinária , Hipocampo/metabolismo , Neurônios/patologia , Encefalomielite Autoimune Experimental/metabolismo , Encefalomielite Autoimune Experimental/patologia , Encefalomielite Autoimune Experimental/veterinária , Citocinas/metabolismo , Doenças dos Roedores/metabolismo , Doenças dos Roedores/patologiaRESUMO
Sphingosine-1-phosphate (S1P) is a signaling lipid mediator that is involved in multiple biological processes. The S1P/S1P receptor (S1PR) signaling pathway has an important role in the central nervous system. It contributes to physiologic cellular homeostasis and is also associated with neuroinflammation. Therefore, this study was performed to evaluate the expression of S1PR in dogs with meningoencephalitis of unknown etiology (MUE) and experimental autoimmune encephalomyelitis (EAE). The analysis used 12 brain samples from three neurologically normal dogs, seven dogs with MUE, and two canine EAE models. Anti-S1PR1 antibody was used for immunohistochemistry. In normal brain tissues, S1PR1s were expressed on neurons, astrocytes, oligodendrocytes, and endothelial cells. In MUE and EAE lesions, there was positive staining of S1PR1 on leukocytes. Furthermore, the expression of S1PR1 on neurons, astrocytes, oligodendrocytes, and endothelial cells was upregulated compared to normal brains. This study shows that S1PR1s are expressed in normal brain tissues and leukocytes in inflammatory lesions, and demonstrates the upregulation of S1PR1 expression on nervous system cells in inflammatory lesions of MUE and EAE. These findings indicate that S1P/S1PR signaling pathway might involve physiologic homeostasis and neuroinflammation and represent potential targets for S1PR modulators to treat MUE.
Assuntos
Encéfalo , Doenças do Cão , Encefalomielite Autoimune Experimental , Receptores de Esfingosina-1-Fosfato , Animais , Cães , Doenças do Cão/metabolismo , Encefalomielite Autoimune Experimental/veterinária , Encefalomielite Autoimune Experimental/metabolismo , Encéfalo/metabolismo , Receptores de Esfingosina-1-Fosfato/metabolismo , Feminino , Masculino , Meningoencefalite/veterinária , Meningoencefalite/metabolismo , Doenças Neuroinflamatórias/veterinária , Doenças Neuroinflamatórias/metabolismo , Astrócitos/metabolismoRESUMO
Intravascular (IV) perfusion of tissue fixative is commonly used in the field of neuroscience as the central nervous system tissues are exquisitely sensitive to handling and fixation artifacts which can affect downstream microscopic analysis. Both 10% neutral-buffered formalin (NBF) and 4% paraformaldehyde (PFA) are used, although IV perfusion with PFA is most commonly referenced. The study objective was to compare the severity of handling and fixation artifacts, semiquantitative scores of inflammatory and neurodegenerative changes, and quantitative immunohistochemistry following terminal IV perfusion of mice with either 10% NBF or 4% PFA in a model of experimental autoimmune encephalitis (EAE). The study included 24 mice; 12 were control animals not immunized and an additional 12 were immunized with PLP139-151 subcutaneously, harvested at day 20, and fixed in the same fashion. Equal numbers (4 per group) were perfused with 10% NBF or 4% PFA, and 4 were immersion-fixed in 10% NBF. NBF-perfused mice had less severe dark neuron artifact than PFA-perfused mice (P < .001). Immersion-fixed animals had significantly higher scores for oligodendrocyte halos, dark neuron artifact, and perivascular clefts than perfusion-fixed animals. Histopathology scores in EAE mice for inflammation, demyelination, and necrosis did not differ among fixation methods. Also, no significant differences in quantitative immunohistochemistry for CD3 and Iba-1 were observed in immunized animals regardless of the method of fixation. These findings indicate that IV perfusion of mice with 10% NBF and 4% PFA are similar and adequate fixation techniques in this model.
Assuntos
Encefalomielite Autoimune Experimental , Doenças dos Roedores , Animais , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/veterinária , Fixadores , Formaldeído , Imuno-Histoquímica , Camundongos , Perfusão/veterinária , Polímeros , Fixação de Tecidos/métodos , Fixação de Tecidos/veterináriaRESUMO
Novel small molecules were synthesized and evaluated as retinoic acid receptor-related orphan receptor-gamma t (RORγt) inverse agonists for the treatment of inflammatory and autoimmune diseases. A hit compound, 1, was discovered by high-throughput screening of our compound library. The structure-activity relationship (SAR) study of compound 1 showed that the introduction of a chlorine group at the 3-position of 4-cyanophenyl moiety increased the potency and a 3-methylpentane-1,5-diamide linker is favorable for the activity. The carbazole moiety of 1 was also optimized; a quinazolinedione derivative 18i suppressed the increase of IL-17A mRNA level in the lymph node of a rat model of experimental autoimmune encephalomyelitis (EAE) upon oral administration. These results indicate that the novel quinazolinedione derivatives have great potential as orally available small-molecule RORγt inverse agonists for the treatment of Th17-driven autoimmune diseases. A U-shaped bioactive conformation of this chemotype with RORγt protein was also observed.
Assuntos
Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/agonistas , Quinazolinonas/química , Administração Oral , Animais , Sítios de Ligação , Agonismo Inverso de Drogas , Encefalomielite Autoimune Experimental/tratamento farmacológico , Encefalomielite Autoimune Experimental/patologia , Encefalomielite Autoimune Experimental/veterinária , Feminino , Humanos , Concentração Inibidora 50 , Interleucina-17/genética , Interleucina-17/metabolismo , Células Jurkat , Simulação de Acoplamento Molecular , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/genética , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Ligação Proteica/efeitos dos fármacos , Estrutura Terciária de Proteína , Quinazolinonas/administração & dosagem , Quinazolinonas/metabolismo , Quinazolinonas/farmacologia , Ratos , Ratos Endogâmicos Lew , Solubilidade , Relação Estrutura-Atividade , Células Th17/citologia , Células Th17/efeitos dos fármacos , Células Th17/metabolismoRESUMO
Human idiopathic inflammatory demyelinating diseases (IIDD) are a heterogeneous group of autoimmune inflammatory and demyelinating disorders of the central nervous system (CNS). These include multiple sclerosis (MS), the most common chronic IIDD, but also rarer disorders such as acute disseminated encephalomyelitis (ADEM) and neuromyelitis optica (NMO). Great efforts have been made to understand the pathophysiology of MS, leading to the development of a few effective treatments. Nonetheless, IIDD still require a better understanding of the causes and underlying mechanisms to implement more effective therapies and diagnostic methods. Experimental autoimmune encephalomyelitis (EAE) is a commonly used animal model to study the pathophysiology of IIDD. EAE is principally induced through immunization with myelin antigens combined with immune-activating adjuvants. Nonhuman primates (NHP), the phylogenetically closest relatives of humans, challenged by similar microorganisms as other primates may recapitulate comparable immune responses to that of humans. In this review, the authors describe EAE models in 3 NHP species: rhesus macaques ( Macaca mulatta), cynomolgus macaques ( Macaca fascicularis), and common marmosets ( Callithrix jacchus), evaluating their respective contribution to the understanding of human IIDD. EAE in NHP is a heterogeneous disease, including acute monophasic and chronic polyphasic forms. This diversity makes it a versatile model to use in translational research. This clinical variability also creates an opportunity to explore multiple facets of immune-mediated mechanisms of neuro-inflammation and demyelination as well as intrinsic protective mechanisms. Here, the authors review current insights into the pathogenesis and immunopathological mechanisms implicated in the development of EAE in NHP.
Assuntos
Doenças Desmielinizantes/veterinária , Doença Autoimune do Sistema Nervoso Experimental/veterinária , Animais , Doenças Desmielinizantes/imunologia , Doenças Desmielinizantes/patologia , Encefalomielite Autoimune Experimental/patologia , Encefalomielite Autoimune Experimental/veterinária , Doença Autoimune do Sistema Nervoso Experimental/patologia , PrimatasRESUMO
In the present study, the use of dogs with experimental autoimmune encephalomyelitis (EAE) as a disease model for necrotizing encephalitis (NE) was assessed. Twelve healthy dogs were included in this study. Canine forebrain tissues (8 g), including white and grey matter, were homogenized with 4 mL of phosphate-buffered saline for 5 min in an ice bath. The suspension was emulsified with the same volume of Freund's complete adjuvant containing 1 mg/mL of killed Mycobacterium tuberculosis H37Ra. Under sedation, each dog was injected subcutaneously with canine brain homogenate at four sites: two in the inguinal and two in the axillary regions. A second injection (booster) was administered to all the dogs using the same procedure 7 days after the first injection. Clinical assessment, magnetic resonance imaging, cerebrospinal fluid analyses, necropsies, and histopathological and immunohistochemical examinations were performed for the dogs with EAE. Out of the 12 animals, seven (58%) developed clinically manifest EAE at various times after immunization. Characteristics of canine EAE models were very similar to canine NE, suggesting that canine EAE can be a disease model for NE in dogs.
Assuntos
Encéfalo/patologia , Doenças do Cão/imunologia , Encefalite/veterinária , Encefalomielite Autoimune Experimental/veterinária , Necrose/veterinária , Animais , Modelos Animais de Doenças , Cães , Encefalite/imunologia , Encefalomielite Autoimune Experimental/imunologia , Feminino , Imunofluorescência/veterinária , Imunização/veterinária , Imuno-Histoquímica/veterinária , Imageamento por Ressonância Magnética/veterinária , Masculino , Necrose/imunologiaRESUMO
Seventy-four 9-week old female C57BL/6J mice housed in a conventional facility were manipulated to induce experimental autoimmune encephalomyelitis, among which 26 developed clinical signs including lethargy, absence of defecation, and abdominal distension. By gross necropsy examination, there was distension of the cecum and colon with fecal impaction. By histologic examination, there was severe ulcerative and proliferative typhlocolitis. Fecal ELISA confirmed the presence of toxins A and B of Clostridium difficile. Alteration in immune status of the immunocompetent mice, due to stress caused by experimental manipulation or autoimmune disease, may have led to intestinal dysbiosis, followed by opportunistic infections resulting in C. difficile-associated disease. This report brings to light the occurrence of the disease in immunocompetent laboratory mice during experimental manipulations associated with alteration in immune status, and it discusses potential hazards associated with conventional housing within a hospital-associated research institute.
Assuntos
Abdome/patologia , Clostridioides difficile/metabolismo , Colite/veterinária , Constipação Intestinal/veterinária , Surtos de Doenças/veterinária , Camundongos Endogâmicos C57BL , Doenças dos Roedores/microbiologia , Doenças dos Roedores/patologia , Animais , Proteínas de Bactérias/metabolismo , Toxinas Bacterianas/metabolismo , Colite/microbiologia , Colite/patologia , Constipação Intestinal/patologia , Encefalomielite Autoimune Experimental/veterinária , Enterotoxinas/metabolismo , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Abrigo para Animais , CamundongosAssuntos
Doenças do Sistema Nervoso Central/veterinária , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/veterinária , Camundongos , Esclerose Múltipla , Ratos , Doenças dos Roedores , Animais , Autoantígenos/imunologia , Encefalomielite Autoimune Experimental/patologia , Humanos , Esclerose Múltipla/diagnóstico , Esclerose Múltipla/prevenção & controle , Esclerose Múltipla/terapia , RoedoresRESUMO
Cerebrospinal fluid (CSF) was removed from guineapigs by puncture of the cisterna magna and the total sample volume of 200-360 microl divided into 40 microl aliquots. After determination of albumin and IgG in these CSF aliquots it was found that successive samples gave different results. In general, up to 100 microl CSF could be removed before the protein concentration began to increase. In animals with chronic relapsing experimental allergic encephalomyelitis (CR-EAE) the rise in albumin concentration was accompanied by a corresponding fall in the number of white cells in later samples.