Introduction: Multiple sclerosis (MS) is a chronic autoimmune disease of the central nervous system characterized by neuroinflammation leading to demyelination. The associated symptoms lead to a devastating decrease in quality of life. The cannabinoids and their derivatives have emerged as an encouraging alternative due to their management of symptom in MS. Objective: The aim of the study was to investigate the mechanism of action of cannabidiol (CBD), a nonpsychoactive cannabinoid, on molecular and cellular events associated with leukocyte recruitment induced by experimental autoimmune encephalomyelitis (EAE). Materials and Methods: C57BL/6 female mice were randomly assigned to the four experimental groups: C (control group), CBD (cannabidiol-treated group, 5 mg/kg i.p.; 14 days), EAE (experimental autoimmune encephalomyelitis-induced group), and EAE+CBD (experimental autoimmune encephalomyelitis-induced plus cannabidiol-treated group). Results: The results indicated that 5 mg/kg of CBD injected intraperitoneally between the 1st and 14th days of EAE could reduce the leukocyte rolling and adhesion into the spinal cord microvasculature as well cellular tissue infiltration. These results were supported by a decreased mRNA expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in the spinal cord. Conclusion: Purified CBD reduces in vivo VCAM and ICAM-mediated leukocyte recruitment to the spinal cord microvasculature at EAE peak disease.

Dysregulated microglia and astrocytes have been associated with progressive neurodegeneration in multiple sclerosis (MS), highlighting the need for strategies that additionally target intrinsic inflammation in the central nervous system (CNS). The objective of the present study was to investigate the glial response in experimental autoimmune encephalomyelitis (EAE)-induced mice treated with a combination of dimethyl fumarate (DMF) and pregabalin (PGB). For that, 28 C57BL/6J mice were randomly assigned to the five experimental groups: naïve, EAE, EAE-DMF, EAE-PGB, and EAE-DMF + PGB. Pharmacological treatments were initiated with the beginning of clinical signs, and all animals were euthanized at 28 dpi for the lumbar spinal cord evaluation. The results demonstrated a stronger attenuation of the clinical presentation by the combined approach. DMF alone promoted the downregulation of Iba-1 (microglia/macrophages marker) in the ventral horn compared with the non-treated EAE animals (P < 0.05). PGB treatment was associated with reduced Iba-1 immunofluorescence in both the dorsal (P < 0.05) and ventral horn (P < 0.05) compared to EAE vehicle-treated counterparts. However, the combined approach reduced the Iba-1 marker in the dorsal (P < 0.05) and ventral (P < 0.01) horns compared to non-treated EAE animals and further reduced Iba-1 in the ventral horn compared to each drug-alone approach (P < 0.05). In addition, the combination of DMF and PGB reduced activated astrocytes (GFAP) in both the dorsal and ventral horns of the spinal cord to a naïve-like level and upregulated Nrf-2 expression. Taken together, the data herein suggest robust attenuation of the glial response in EAE mice treated with DMF and PGB.

CNS lesions usually result in permanent loss of function and are an important problem in the medical field. In order to investigate neuroprotection/degeneration mechanisms and the synaptic plasticity of motoneurons, in addition to the potential for a variety of treatments, different experimental models of axonal injury have been proposed. Recent studies have tested the immunomodulatory drug dimethyl fumarate (DMF) for the treatment of neurodegenerative diseases and have shown promising outcomes. Therefore, in this work, we investigated the effects of DMF with regard to neuroprotection and its influence on the glial response in C57BL/6J animals subjected to crushing of the motor roots in the lumbar intumescence of the spinal cord. The animals were divided into a vehicle-treated injury group (0.08 % methylcellulose solution control group, n = 7) and injured groups treated with DMF at different doses (15, 30, 45, 90 and 180 mg/kg; n = 6-7 per dose). The 90 mg/kg dose showed the best neuroprotective results, so it was used for treatment over a period of eight weeks. Neuronal survival was assessed through Nissl staining, and functional recovery was evaluated with the CatWalk system (walking track test) and the von Frey test (mechanoreception). Immunohistochemistry was used to assess synaptic coverage and astroglial and microglial reactivity using the primary antibodies anti-synaptophysin (pre-synaptic terminal pan marker), GAD65 (GABAergic pre-synaptic terminations - inhibitory), and VGLUT1 (glutamatergic pre-synaptic terminations - excitatory). Glial reactions were evaluated with anti-IBA1 (microglia) and GFAP (astrocytes). Gene transcript levels of IL-3, IL-4, TNF-α, IL-6, TGF-β, iNOS-M1, and arginase-M2 were quantified by RT-qPCR. The results indicated that treatment with DMF, at a dose of 90 mg/kg, promoted neuroprotection and immunomodulation towards an anti-inflammatory response. It also resulted in greater preservation of inhibitory synapses and reduced astroglial reactivity, providing a more favorable environment for sensorimotor recovery.

Background: Although it has been suggested that healthier lifestyle may optimize effects of the immunomodulation drugs for treating multiple sclerosis (MS), the knowledge regarding this kind of interactions is limited. Objective: The aim of the present study was to investigate the effects of treadmill exercise in combination with pharmacological treatment in an animal model for MS. Methods: C57BL/6J female mice were subjected to daily treadmill exercise for 4 weeks before immunization and 6 weeks before clinical presentation of disease. Dimethyl fumarate (DMF) or glatiramer acetate (GA) were administered after the first clinical relapse. Histopathological analyses were carried out in the lumbar spinal cord at peak disease and at 1 or 14 days post-treatment (dpt). Results: Exercised-GA treated animals demonstrated decreased astrocytic response in the spinal dorsal horn with an improvement in the paw print pressure. Exercised-DMF treated animals showed an increased microglial/macrophage response on both ventral and dorsal horn that were associated with clinical improvement and synaptic motoneuron inputs density. Conclusion: The present data suggest that prior regular exercise can modify the effects of pharmacological treatment administered after the first relapse in a murine model for MS.

Multiple sclerosis (MS) is a demyelinating disease with a wide range of symptoms including walking impairment and neuropathic pain mainly represented by mechanical allodynia. Noteworthy, exercise preconditioning may affect both walking impairment and mechanical allodynia. Most of MS symptoms can be reproduced in the animal model named experimental autoimmune encephalomyelitis (EAE). Usually, neurological deficits of EAE are recorded using a clinical scale based on the development of disease severity that characterizes tail and limb paralysis. Following paralysis recovery, subtle motor alterations and even mechanical allodynia investigation are difficult to record, representing sequels of peak disease. The aim of the present study was to investigate the walking dysfunction by the catwalk system (CT) in exercised and non-exercised C57BL/6 mice submitted to EAE with MOG35–55 up to 42 days post-induction (dpi).

Peripheral nerve injury results in retrograde cell body-related changes in the spinal motoneurons that will contribute to the regenerative response of their axons. Successful functional recovery also depends on molecular events mediated by innate immune response during Wallerian degeneration in the nerve microenvironment. A previous study in our lab demonstrated that TLR 2 and 4 develop opposite effects on synaptic stability in the spinal cord after peripheral nerve injury. Therefore, we suggested that the better preservation of spinal cord microenvironment would positively influence distal axonal regrowth. In this context, the present work aimed to investigate the influence of TLR2 and TLR4 on regeneration and functional recovery after peripheral nerve injury.

Experimental autoimmune encephalomyelitis (EAE) induces significant reduction of the body mass concomitant to sickness behavior and anorexia. We investigated whether regular physical exercise prevents metabolic alterations associated with body mass loss occurring during EAE inflammatory peak. Female C57BL/6 mice were assigned to the unexercised and exercised (trained) groups. In four weeks, EAE was induced in half of the animals in each group, and the exercise protocol was maintained until 10 days post-induction (dpi 10) completing 6 weeks of regular exercise (forced swimming). At dpi 14, the relative mass of metabolic tissues, serum levels of triglycerides, cholesterol, and glucose, glycogen contents in the muscle and liver, and muscle levels of cytokines were measured. A significantly decreased clinical score associated with attenuation of the body mass loss in exercised EAE animals, as compared to the non-exercised ones, was observed. The associated metabolic parameters were not modified by this approach, although negative correlations between some parameters and clinical score at dpi 14 were observed. Although the prior program of aerobic exercise is capable of decreasing clinical score and body mass loss, it is not sufficient to modify metabolic outcomes associated with inflammation at the EAE peak.

A microscopia intravital (MI) é uma poderosa ferramenta utilizada na área biomédica para a observação e imageamento in vivo de sistemas biológicos, tendo importante aplicação no estudo das interações leucócito-endotélio que ocorrem na microcirculação de diversos tecidos animais sob condições normais e patológicas. Dado que o controle das condições de aquisição das imagens na análise in vivo é bastante limitado, a presença de borramento e de artefatos de movimento nas sequências de quadros de vídeo adquiridas, resultantes da respiração e dos batimentos cardíacos inevitáveis do animal, constitui um dos principais obstáculos da MI. Isso compromete significativamente a detecção e o rastreamento dos leucócitos ao longo dos quadros, tanto em análises visuais como nas realizadas por métodos automáticos (computacionais). Assim, este trabalho realizou o estudo e aplicação de duas técnicas de deconvolução cega (algoritmo deconvblind e método de Shan et al.) para a correção de artefatos de movimento em imagens de vídeo de MI, fazendo uma avaliação quantitativa de como tal restauração afeta o desempenho de dois métodos de detecção automática de leucócitos (casamento de padrões e simetria de fase local). Análises de precisão versus revocação foram conduzidas a fim de se avaliar a eficácia da detecção automática em imagens de boa qualidade visual, imagens degradadas por movimento e imagens restauradas com cada uma das técnicas. A partir desse estudo, concluiu-se que o método de Shan et al. possui o maior potencial de restauração dos leucócitos e que as técnicas de deconvolução cega de fato têm utilidade como etapa de pré-processamento dos vídeos de MI para análises in vivo, especialmente quando o método de detecção automática de leucócitos é mais sensível a erros.

Intravital microscopy is an important experimental tool for the study of cellular and molecular mechanisms of the leukocyte–endothelial interactions in the microcirculation of various tissues and in different inflammatory conditions of in vivo specimens. However, due to the limited control over the conditions of the image acquisition, motion blur and artifacts, resulting mainly from the heartbeat and respiratory movements of the in vivo specimen, will very often be present. This problem can significantly undermine the results of either visual or computerized analysis of the acquired video images. Since only a fraction of the total number of images are usually corrupted by severe motion blur, it is necessary to have a procedure to automatically identify such images in the video for either further restoration or removal. This paper proposes a new technique for the detection of motion blur in intravital video microscopy based on directional statistics of local energy maps computed using a bank of 2D log-Gabor filters. Quantitative assessment using both artificially corrupted images and real microscopy data were conducted to test the effectiveness of the proposed method. Results showed an area under the receiver operating characteristic curve (AUC) of 0.95 (\(\hbox {AUC}=0.95;\) 95 % CI 0.93–0.97) when tested on 329 video images visually ranked by four observers.

Although previous studies have shown that forced exercise modulates inflammation and is therapeutic acutely for experimental autoimmune encephalomyelitis (EAE), the long-term benefits have not been evaluated. In this study, we investigated the effects of preconditioning exercise on the clinical and pathological progression of EAE. Female C57BL/6 mice were randomly assigned to either an exercised (Ex) or unexercised (UEx) group and all of them were induced for EAE. Mice in the Ex group had an attenuated clinical score relative to UEx mice throughout the study. At 42 dpi, flow cytometry analysis showed a significant reduction in B cells, CD4(+) T cells, and CD8(+) T cells infiltrating into the spinal cord in the Ex group compared to UEx. Ex mice also had a significant reduction in myelin damage with a corresponding increase in proteolipid protein expression. Finally, Ex mice had a significant reduction in axonal damage. Collectively, our study demonstrates for the first time that a prolonged and forced preconditioning protocol of exercise improves clinical outcome and attenuates pathological hallmarks of EAE at chronic disease. In this study, we show that a program of 6 weeks of preconditioning exercise promoted a significant reduction of cells infiltrating into the spinal cord, a significant reduction in myelin damage and a significant reduction in axonal damage in experimental autoimmune encephalomyelitis (EAE) mice at 42 dpi. Collectively, our study demonstrates for the first time that a preconditioning protocol of exercise improves clinical outcome and attenuates pathological hallmarks of EAE at chronic disease.

Patients suffering from neuropathic pain have a higher incidence of mood disorders such as depression. Increased expression of TNF has been reported in neuropathic pain and depressive-like conditions and most of the pro-inflammatory effects of TNF are mediated by TNF receptor 1. Here we sought to investigate 1) the occurrence of anhedonia in chronic neuropathic pain and the associated forms of hippocampal plasticity and 2) the involvement of TNFR1-mediated TNF signaling as a possible regulator of such events.

Intravital microscopy (IM) is an important experimental tool for the study of cellular and molecular mechanisms of leukocyte-endothelial interactions which occur in the microcirculation of various tissues and in various inflammatory conditions of in vivo specimens. Due to the limited control over the image acquisition conditions, the acquired video images can often have motion blur resulting mainly from the heartbeats and respiratory movements of the specimen. This problem can significantly undermine the results of both visual and computerized analysis over the IM videos. But since only a fraction of the total number of images is usually corrupted by severe motion blur, it is very important and desirable to have a procedure to automatically identify such images for posterior restoration. This work presents a new technique for the detection of motion blur in IM video images, based on directional (circular) statistics of local energy maps computed using a bank of 2D log-Gabor filters. In order to test the effectiveness of the proposed method, a quantitative assessment was conducted by testing each circular measure on 329 video images visually ranked by four observers. Results have shown areas under the ROC curves (AUC) of 0.95, outperforming the PSNR, SSIM and BIQAA metrics.

The interactions between a prior program of regular exercise and the development of experimental autoimmune encephalomyelitis (EAE)-mediated responses were evaluated. In the exercised EAE mice, although there was no effect on infiltrated cells, the cytokine and derived neurotrophic factor (BDNF) levels were altered, and the clinical score was attenuated. Although, the cytokine levels were decreased in the brain and increased in the spinal cord, BDNF was elevated in both compartments with a tendency of lesser demyelization volume in the spinal cord of the exercised EAE group compared with the unexercised.

Infection of mice with Plasmodium berghei NK65 represents a well-recognized malaria model in which infection is accompanied by an intense hepatic inflammatory response. Enzyme-inducible nitric oxide synthase is an important regulator of inflammation and leukocyte recruitment in microvessels, but these functions have yet to be evaluated in experimental malaria. In this study, we assessed the involvement of inducible nitric oxide synthase in inflammatory responses to murine experimental malaria induced by P. berghei NK65. We observed that wild type (WT) and nitric oxide synthase (iNOS)-deficient mice (iNOS−/−) mice showed similar levels of parasitemia following P. berghei NK65 infection, although infected iNOS−/− mice presented early mortality. Inducible nitric oxide synthase deficiency led to increased leukocyte rolling and adhesion to the liver in iNOS−/− mice relative to the WT animals, as observed via intravital microscopy. Infected iNOS−/− mice also exhibited increased hepatic leukocyte migration and subsequent liver damage, which was associated with high serum levels of the cytokines TNF-α, IL-6 and IL-10. Our data suggest potential role for the iNOS enzyme as a regulator of hepatic inflammatory response induced by P. berghei NK65-infection, and its absence leads to exacerbated inflammation and sequential associated-hepatic damage in the animals.

Although it is well known that regular exercise may promote neuroprotection, the mechanisms underlying this effect are still not fully understood. We investigated if swim training promotes neuroprotection by potentiating antioxidant pathways, thereby decreasing the effects of oxidative stress on glutamate and nitric oxide release. Male Wistar rats (n=36) were evenly randomized into a trained group (TRA) (5 days/week, 8 weeks, 30 min) and a sedentary group (SED). Forty-eight hours after the last session of exercise, animals were killed and brain was collected for in vitro ischemia. Cortical slices were divided into two groups: a group in which oxidative stress was induced by oxygen and glucose deprivation (OGD), and a group of non-deprived controls (nOGD). Interestingly, exercise by itself increased superoxide dismutase activity (nOGD, SED vs. TRA animals) with no effect on pro-oxidative markers. In fact, TRA-OGD slices showed lowered levels of lactate dehydrogenase when compared with SED-OGD controls, reinforcing the idea that exercise affords a neuroprotective effect. We also demonstrated that exercise decreased glutamate and nitrite release as well as lipid membrane damage in the OGD cortical slices. Our data suggest that under conditions of metabolic stress, swim training prevents oxidative damage caused by glutamate and nitric oxide release.

O objetivo do presente estudo foi avaliar os efeitos interativo e respectivo do treinamento fisico moderado de natacao e da dieta hiperlipidica sobre o metabolismo de carboidratos e lipideos no periodo de recuperacao ao exercicio em ratos machos adultos. Os animais foram divididos em sedentarios (S) e treinados (T). O treinamento consistiu de natacao a 90 min/dia, cinco dias/semana. Cada grupo foi subdividido em dieta padrao normocalorica (N) e dieta hipercalorica/hiperlipidica (H). Depois de oito semanas, os animais dos quatro grupos (SN, TN, SH e TH) foram sacrificados em repouso ou uma e duas horas apos o exercicio. A dieta hipercalorica (grupo SH vs. SN) aumentou o peso dos tecidos adiposos brancos, a glicemia, a concentracao plasmatica de triglicerideos e de colesterol total e diminuiu o conteudo de glicogenio. Por outro lado, o treinamento moderado de natacao (grupo TN vs. SN) diminuiu o peso dos tecidos adiposos brancos e melhorou o metabolismo de carboidratos, aumentando o conteudo de glicogenio. Quando associados (grupo TH vs. SH e TN), o treinamento nao foi capaz de reverter os efeitos da dieta para o peso dos tecidos adiposos brancos, conteudo de glicogenio e glicemia. Com relacao ao periodo de recuperacao, concluimos que os efeitos beneficos do treinamento em promover uma rapida recuperacao dos substratos endogenos, principalmente reposicao de glicogenio, sao prejudicados pela dieta hiperlipidica associada ao treinamento