Rat Model Of Multiple Sclerosis Research Articles

Genetic variants of CC chemokine genes in experimental autoimmune encephalomyelitis, multiple sclerosis and rheumatoid arthritis

Multiple sclerosis (MS) is a complex disorder of the central nervous system, causing inflammation, demyelination and axonal damage. A limited number of genetic risk factors for MS have been identified, but the etiology of the disease remains largely unknown. For the identification of genes regulating neuroinflammation we used a rat model of MS, myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE), and carried out a linkage analysis in an advanced intercross line (AIL). We thereby redefine the Eae18b locus to a 0.88 Mb region, including a cluster of chemokine genes. Further, we show differential expression of Ccl2, Ccl11 and Ccl11 during EAE in rat strains with opposite susceptibility to EAE, regulated by genotype in Eae18b. The human homologous genes were tested for association to MS in 3841 cases and 4046 controls from four Nordic countries. A haplotype in CCL2 and rs3136682 in CCL1 show a protective association to MS, whereas a haplotype in CCL13 is disease predisposing. In the HLA-DRB1* 15 positive subgroup, we also identified an association to a risk haplotype in CCL2, suggesting an influence from the human leukocyte antigen (HLA) locus. We further identified association to rheumatoid arthritis in CCL2, CCL8 and CCL13, indicating common regulatory mechanisms for complex diseases.

The establishment of a multiple sclerosis model for clinical MRI study

Objective To establish a rat model of multiple sclerosis (MS) for clinical MRI study. Methods The Lewis (LEW) rats were immunized by myelin oligodendrocyte glycoprotein peptide 35-55 (MOG35-55) emulsified with complete Freunds adjuvant/incomplete Freunds adjuvant. Twenty LEW rats were selected. Group A rats were 5, group B rats were 10 and control group rats were 5. MRI and histopathology analysis were observed on group A and control group, and clinical course were observed on group B. The clinical course was observed and the brain and spinal cord of EAE rats were scanned on 3. 0 T MR system with quadrature wrist joint coil on the third day of the acute stage of disease. The T2/T1 weighted images and Gadolinium enhanced T1 weighted images in 3D volume were obtained respectively. The magnetization transfer ratio (MTR) images were calculated with special software in workstation. Rats were sacrificed within 24 h after MRI, and the histopathological changes of central nervous system were observed. Results Twice immunization of MOG35-55 was used and a rat model of MS was successfully induced with all the rats. The model had varied clinical symptoms. The lesions of central nervous system at acute stage on group A (5 rats) were depicted on MRI. The lesions located in the brain and the spinal cord with main manifestation of hyperintensity on T2 weighed images and hypointensity on T1 weighted images. Some lesions had Gd-DTPA enhancement, and the value of MTR decreased. The infiltration of mononuclear cells around blood vessels and the extensive distribution of lesions of demyelination in brain and spinal cord were verified by histological examination. Conclusions Twice immunization of MOG35-55 could produce a rat model which mimics MS.The model is stable with higher incidence, and the antigen is cheap and obtained easily. The intravital monitoring of brain and spinal cord lesions in MOG35-55 induced rat model is possible on 3.0 T clinical whole-body MR scanner with a quadrature wrist joint coil. MOG35-55 induced rat model is an ideal MS model for clinical MRI study. Key words: Multiple sclerosis; Models; animal; Rats; Magnetic resonance imaging

Imaging of Effector Memory T Cells during a Delayed-Type Hypersensitivity Reaction and Suppression by Kv1.3 Channel Block

Effector memory T (Tem) cells are essential mediators of autoimmune disease and delayed-type hypersensitivity (DTH), a convenient model for two-photon imaging of Tem cell participation in an inflammatory response. Shortly (3 hr) after entry into antigen-primed ear tissue, Tem cells stably attached to antigen-bearing antigen-presenting cells (APCs). After 24 hr, enlarged Tem cells were highly motile along collagen fibers and continued to migrate rapidly for 18 hr. Tem cells rely on voltage-gated Kv1.3 potassium channels to regulate calcium signaling. ShK-186, a specific Kv1.3 blocker, inhibited DTH and suppressed Tem cell enlargement and motility in inflamed tissue but had no effect on homing to or motility in lymph nodes of naive and central memory T (Tcm) cells. ShK-186 effectively treated disease in a rat model of multiple sclerosis. These results demonstrate a requirement for Kv1.3 channels in Tem cells during an inflammatory immune response in peripheral tissues. Targeting Kv1.3 allows for effector memory responses to be suppressed while central memory responses remain intact.

Anti-inflammatory cytokine gene therapy decreases sensory and motor dysfunction in experimental Multiple Sclerosis: MOG-EAE behavioral and anatomical symptom treatment with cytokine gene therapy

Multiple Sclerosis (MS) is an autoimmune inflammatory disease that presents clinically with a range of symptoms including motor, sensory, and cognitive dysfunction as well as demyelination and lesion formation in brain and spinal cord. A variety of animal models of MS have been developed that share many of the pathological hallmarks of MS including motor deficits (ascending paralysis), demyelination and axonal damage of central nervous system (CNS) tissue. In recent years, neuropathic pain has been recognized as a prevalent symptom of MS in a majority of patients. To date, there have been very few investigations into sensory disturbances in animal models of MS. The current work contains the first assessment of hind paw mechanical allodynia (von Frey test) over the course of a relapsing-remitting myelin oligodendrocyte glycoprotein induced experimental autoimmune encephalomyelitis (MOG-EAE) rat model of MS and establishes the utility of this model in examining autoimmune induced sensory dysfunction. We demonstrate periods of both decreased responsiveness to touch that precedes the onset of hind limb paralysis, and increased responsiveness (allodynia) that occurs during the period of motor deficit amelioration traditionally referred to as symptom remission. Furthermore, we tested the ability of our recently characterized anti-inflammatory IL-10 gene therapy to treat the autoimmune inflammation induced behavioral symptoms and tissue histopathological changes. This therapy is shown here to reverse inflammation induced paralysis, to reduce disease associated reduction in sensitivity to touch, to prevent the onset of allodynia, to reverse disease associated loss of body weight, and to suppress CNS glial activation associated with disease progression in this model.

Tissue-Type Plasminogen Activator Is a Regulator of Monocyte Diapedesis through the Brain Endothelial Barrier

Inflammatory cell trafficking into the brain complicates several neurological disorders including multiple sclerosis. Normally, reliable brain functioning is maintained and controlled by the blood-brain barrier (BBB), which is essential to restrict the entry of potentially harmful molecules and cells from the blood into the brain. The BBB is a selective barrier formed by dedicated brain endothelial cells and dependent on the presence of intracellular tight junctions. In multiple sclerosis, a severe dysfunction of the BBB is observed, which is key to monocyte infiltration and inflammation in the brain. Proteolytic activity has been associated with these inflammatory processes in the brain. Our studies in plasma of rats indicated that the extracellular protease tissue-type plasminogen activator (tPA) correlates with the clinical signs of experimental allergic encephalomyelitis, a rat model of multiple sclerosis. In this study, we studied the function of the tPA during diapedesis of monocytes through a rat and human brain endothelial barrier. Monocyte-brain endothelial cell coculture experiments showed that monocytes induce the release of tPA by brain endothelial cells, which subsequently activates the signal transduction protein extracellular signal related kinase (ERK1/2), both involved in monocyte diapedesis. Importantly, live imaging and immunoblot analyses of rat brain endothelial cells revealed that tPA and ERK1/2 control the breakdown of the tight junction protein occludin. These studies identify tPA as a novel and relevant pathological mediator of neuroinflammation and provide a potential mechanism for this.

UCM707, an inhibitor of the anandamide uptake, behaves as a symptom control agent in models of Huntington's disease and multiple sclerosis, but fails to delay/arrest the progression of different motor-related disorders

To date, UCM707, (5Z,8Z,11Z,14Z)-N-(3-furylmethyl)eicosa-5,8,11,14-tetraenamide, has the highest potency and selectivity in vitro and in vivo as inhibitor of the endocannabinoid uptake. This may enable this compound to potentiate endocannabinoid transmission, with minimal side effects, in the treatment of several neurological disorders. In the present study, we examined whether the treatment with UCM707 produced beneficial effects, as other cannabinoid-related compounds have already shown, to alleviate motor deterioration or to delay/arrest neurodegeneration, in several models of neurological diseases such as Huntington's disease (HD), Parkinson's disease (PD) and multiple sclerosis (MS). UCM707 exhibited a notable anti-hyperkinetic activity in a rat model of HD generated by bilateral intrastriatal application of 3-nitropropionic acid. This effect was possibly associated with an amelioration of GABA and glutamate deficits induced by the toxin in the globus pallidus and the substantia nigra, respectively. However, UCM707 did not protect against the death of GABAergic neurons that occurs in rats with striatal atrophy generated by unilateral application of malonate, another animal model of HD, which is more useful to test neuroprotective strategies. In addition, UCM707 did not provide neuroprotection in rats with unilateral lesions of the nigrostriatal dopaminergic neurons caused by 6-hydroxydopamine, a rat model of PD. This was possibly due to the fact that UCM707 is devoid of anti-oxidant properties since another uptake inhibitor, AM404, that has these properties acted as a protective agent. Lastly, UCM707 was also unable to inhibit the development of the neurological impairment of rats with experimental autoimmune encephalomyelitis (EAE), an acute model of MS. However, UCM707, like other endocannabinoid uptake inhibitors reported previously, significantly reduced spasticity of the hindlimbs in a chronic relapsing EAE mice, a chronic model of MS. In summary, UCM707 might be a promising compound in HD to alleviate motor symptoms, which represents an important goal considering the current lack of efficient pharmacological treatments in this basal ganglia disorder. However, the compound was unable to delay neurodegeneration in this disorder and also in PD. In addition, UCM707 did not produce any neurological recovery from inflammatory attack in an EAE rat model of MS, although it retained the classic anti-spastic action shown by other uptake inhibitors in the EAE mouse model of this disease.

Brain-tethered T cells

On page 1805, Kawakami et al. show that antigen-specific CD4+ T cells that attack the brain become stationary within brain tissue, whereas nonspecific T cells cruise through without stopping. These intravital images—the first to capture T cells launching an autoimmune attack on the brain—suggest that antigen-specific T cells behave similarly in dense brain tissue as they do in lymph nodes. Activated CD4+ T cells specific for the endogenous brain protein myelin basic protein (MBP) trigger fatal encephalomyelitis in a rat model of multiple sclerosis. These cells migrate into the central nervous system (CNS) where they become reactivated. Figure A subset of MBP-specific T cells (green) become stationary in the brain during EAE, presumably after engaging their antigen on antigen-presenting cells (red). Previous studies by this group showed that both antigen-specific and nonspecific T cells gain access to the CNS, but only specific cells become reactivated once there. Kawakami et al. now show that a subset of MBP-specific T cells—likely those that have encountered antigen—stopped moving inside the brain, appearing tethered to a fixed point. T cells specific for a control antigen kept moving, suggesting that the presence of antigen was required for T cells to stop moving and to trigger disease. The authors were surprised to find that T cells move as rapidly through the dense brain tissue as in the more aqueous environment of the lymph nodes. How they do this is not known but the authors suggest that the T cells may produce proteases that help clear a path through the compact tissue.

Decreased endocannabinoid levels in the brain and beneficial effects of agents activating cannabinoid and/or vanilloid receptors in a rat model of multiple sclerosis

Recent studies have addressed the changes in endocannabinoid ligands and receptors that occur in multiple sclerosis, as a way to explain the efficacy of cannabinoid compounds to alleviate spasticity, pain, tremor, and other signs of this autoimmune disease. Using Lewis rats with experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, we recently found a decrease in cannabinoid CB1 receptors mainly circumscribed to the basal ganglia, which could be related to the motor disturbances characteristic of these rats. In the present study, using the same model, we explored the potential changes in several neurotransmitters in the basal ganglia that might be associated with the motor disturbances described in these rats, but we only found a small increase in glutamate contents in the globus pallidus. We also examined whether the motor disturbances and the changes of CB1 receptors found in the basal ganglia of EAE rats disappear after the treatment with rolipram, an inhibitor of type IV phosphodiesterase able to supress EAE in different species. Rolipram attenuated clinical decline, reduced motor inhibition, and normalized CB1 receptor gene expression in the basal ganglia. As a third objective, we examined whether EAE rats also exhibited changes in endocannabinoid levels as shown for CB1 receptors. Anandamide and 2-arachidonoylglycerol levels decreased in motor related regions (striatum, midbrain) but also in other brain regions, although the pattern of changes for each endocannabinoid was different. Finally, we hypothesized that the elevation of the endocannabinoid activity, following inhibition of endocannabinoid uptake, might be beneficial in EAE rats. AM404, arvanil, and OMDM2 were effective to reduce the magnitude of the neurological impairment in EAE rats, whereas VDM11 did not produce any effect. The beneficial effects of AM404 were reversed by blocking TRPV1 receptors with capsazepine, but not by blocking CB1 receptors with SR141716, thus indicating the involvement of endovanilloid mechanisms in these effects. However, a role for CB1 receptors is supported by additional data showing that CP55,940 delayed EAE progression. In summary, our data suggest that reduction of endocannabinoid signaling is associated with the development of EAE in rats. We have also proved that the reduction of CB1 receptors observed in these rats is corrected following treatment with a compound used in EAE such as rolipram. In addition, the direct or indirect activation of vanilloid or cannabinoid receptors may reduce the neurological impairment experienced by EAE rats, although the efficacy of the different compounds examined seems to be determined by their particular pharmacodynamic and pharmacokinetic characteristics.

Ciliary Neurotrophic Factor Protects Retinal Ganglion Cells from Secondary Cell Death During Acute Autoimmune Optic Neuritis in Rats

Multiple sclerosis (MS) is a chronic inflammatory disease of the CNS which leads to demyelination, axonal destruction and neuronal loss in the early stages. Available therapies mainly target the inflammatory component of the disease but fail to prevent neurodegeneration. To investigate the effect of ciliary neurotrophic factor (CNTF) on the survival of retinal ganglion cells (RGCs), the neurons that form the axons of the optic nerve, we used a rat model of myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis. Optic neuritis in this model was diagnosed by recording visual evoked potentials, and RGC function was monitored by measuring electroretinograms. This study demonstrates that CNTF has a neuroprotective effect on affected RGCs during acute optic neuritis. Furthermore, we demonstrate that CNTF exerts its neuroprotective effect through activation of the Janus kinase/signal transducer and activator of transcription pathway, mitogen activated protein kinases and a shift in the Bcl-2 family of proteins towards the anti-apoptotic side. In summary, our results demonstrate that CNTF can serve as an effective neuroprotective treatment in a rat model of MS that especially reflects the neurodegenerative aspects of this disease.

Neuroprotective effects and intracellular signaling pathways of erythropoietin in a rat model of multiple sclerosis

In multiple sclerosis (MS), long-term disability is primarily caused by axonal and neuronal damage. We demonstrated in a previous study that neuronal apoptosis occurs early during experimental autoimmune encephalomyelitis, a common animal model of MS. In the present study, we show that, in rats suffering from myelin oligodendrocyte glycoprotein (MOG)-induced optic neuritis, systemic application of erythropoietin (Epo) significantly increased survival and function of retinal ganglion cells (RGCs), the neurons that form the axons of the optic nerve. We identified three independent intracellular signaling pathways involved in Epo-induced neuroprotection in vivo: Protein levels of phospho-Akt, phospho-MAPK 1 and 2, and Bcl-2 were increased under Epo application. Using a combined treatment of Epo together with a selective inhibitor of phosphatidylinositol 3-kinase (PI3-K) prevented upregulation of phospho-Akt and consecutive RGC rescue. We conclude that in MOG-EAE the PI3-K/Akt pathway has an important influence on RGC survival under systemic treatment with Epo.

Apoptosis of neurons in rats with experimental allergic encephalomyelitis

Experimental allergic encephalomyelitis (EAE) is a rat model of multiple sclerosis (MS). In MS and EAE, calpain expression is increased in cells of the central nervous system including astrocytes, oligodendrocytes and microglia; as well as infiltrating macrophages and T cells. However, calpain expression in neurons has not previously been reported in the EAE model. Since degeneration of neurons has been demonstrated in progressive cases of MS, and calpain has been implicated in cell death in EAE, it is hypothesized that neurons expressing increased levels of calpain may undergo apoptosis. To test the hypothesis, spinal cords were collected from control and EAE rats from 4 to 12 days after challenge with guinea‐pig myelin and complete Freund's adjuvant. Apoptosis in neurons was determined by combined TUNEL and immunoflourescent staining using a neuron‐specific marker (NeuN). Calpain expression in neurons was determined using double‐immunoflourescent staining with antibodies specific for calpain and NeuN. At 8 and 9 days postchallenge, neither apoptosis nor increased calpain expression were detected in neurons in control or 9 day EAE spinal cord sections. In contrast, at 11 days postinduction neurons in acute EAE were TUNEL‐positive, indicating that cells underwent apoptosis. Further studies are in progress to determine if and when oligodendrocytes in EAE rats are apoptotic. By correlating cell‐specific increases in calpain expression with apoptosis in EAE, a better understanding of the role of calpain in EAE and MS might be achieved. This increased knowledge could suggest a potential benefit of calpain inhibitors in the treatment of EAE and MS.Acknowledgements: Supported in part by grants from NIH‐NINDS.

Meyer R, Weissert R, Diem R, et al. Acute neuronal apoptosis in a rat model of multiple sclerosis.

Meyer R, Weissert R, Diem R, et al. Acute neuronal apoptosis in a rat model of multiple sclerosis.

Acute Neuronal Apoptosis in a Rat Model of Multiple Sclerosis

Demyelination caused by inflammation of the CNS has been considered to be a major hallmark of multiple sclerosis (MS). Using experimental autoimmune encephalomyelitis, a model of MS, we demonstrate that an immune-mediated attack of the optic nerve is accompanied by an early degeneration of retinal ganglion cells (RGCs). The decrease of neuronal cell density was correlated with functional disabilities as assessed by visual evoked cortical potentials and electroretinogram. Visual acuity was significantly reduced. DNA degradation and activation of caspase-3 in RGCs indicate that cell death of RGCs is apoptotic. These findings show for the first time that an inflammatory attack against myelin components can lead to acute neuronal cell loss by apoptosis.

MATTERS ARISING: Cavanagh replies:

Cortical reorganization occurring in multiple sclerosis (MS) patients is thought to play a key role in limiting the effect of structural tissue damage. Conversely, its exhaustion may contribute to the irreversible disability that accumulates with disease progression. Several aspects of MS-related cortical reorganization, including the overall functional effect and likely modulation by therapies, still remain to be elucidated. The aim of this work was to assess the extent of functional cortical reorganization and its brain structural/pathological correlates in Dark Agouti rats with experimental autoimmune encephalomyelitis (EAE), a widely accepted preclinical model of chronic MS. Morphological and functional MRI (fMRI) were performed before disease induction and during the relapsing and chronic phases of EAE. During somatosensory stimulation of the right forepaw, fMRI demonstrated that cortical reorganization occurs in both relapsing and chronic phases of EAE with increased activated volume and decreased laterality index versus baseline values. Voxel-based morphometry demonstrated gray matter (GM) atrophy in the cerebral cortex, and both GM and white matter atrophy were assessed by <i>ex vivo</i> pathology of the sensorimotor cortex and corpus callosum. Neuroinflammation persisted in the relapsing and chronic phases, with dendritic spine density in the layer IV sensory neurons inversely correlating with the number of cluster of differentiation 45-positive inflammatory lesions. Our work provides an innovative experimental platform that may be pivotal for the comprehension of key mechanisms responsible for the accumulation of irreversible brain damage and for the development of innovative therapies to reduce disability in EAE/MS. <b>SIGNIFICANCE STATEMENT</b> Since the early 2000s, functional MRI (fMRI) has demonstrated profound modifications in the recruitment of cortical areas during motor, cognitive, and sensory tasks in multiple sclerosis (MS) patients. Experimental autoimmune encephalomyelitis (EAE) represents a reliable model of the chronic-progressive variant of MS. fMRI studies in EAE have not been performed extensively up to now. This paper reports fMRI studies in a rat model of MS with somatosensory stimulation of the forepaw. We demonstrated modifications in the recruitment of cortical areas consistent with data from MS patients. To the best of our knowledge, this is the first report of cortical remodeling in a preclinical <i>in vivo</i> model of MS.