Experimental Autoimmune Encephalomyelitis Lesions Research Articles

VEGF-A-mediated venous endothelial cell proliferation results in neoangiogenesis during neuroinflammation.

Newly formed leaky vessels and blood-brain barrier (BBB) damage are present in demyelinating acute and chronic lesions in multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE). However, the endothelial cell subtypes and signaling pathways contributing to these leaky neovessels are unclear. Here, using single-cell transcriptional profiling and in vivo validation studies, we show that venous endothelial cells express neoangiogenesis gene signatures and show increased proliferation resulting in enlarged veins and higher venous coverage in acute and chronic EAE lesions in female adult mice. These changes correlate with the upregulation of vascular endothelial growth factor A (VEGF-A) signaling. We also confirmed increased expression of neoangiogenic markers in acute and chronic human MS lesions. Treatment with a VEGF-A blocking antibody diminishes the neoangiogenic transcriptomic signatures and vascular proliferation in female adult mice with EAE, but it does not restore BBB function or ameliorate EAE pathology. Our data demonstrate that venous endothelial cells contribute to neoangiogenesis in demyelinating neuroinflammatory conditions.

Expression of sphingosine-1-phosphate receptor 1 in neuroinflammation of canine brains

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.

Involvement of NRON and TUG1 long noncoding RNAs in inflammation and the pathogenesis of EAE

There is growing evidence that the long noncoding RNAs (lncRNAs) contribute to the pathogenesis of various neurodegenerative diseases such as multiple sclerosis (MS). The role of lncRNAs nuclear repressor of NFAT (NRON) and Taurine up-regulated 1 (TUG1) in the inflammatory processes occurring in the experimental autoimmune encephalomyelitis (EAE) model of MS is yet to be investigated. Transcript levels of NRON and TUG1 in acute and chronic phases of EAE and cultured macrophages as well as the correlation between NRON and TUG1 expression with inflammatory cytokines, were evaluated in this study. EAE experimental model was induced in female C57BL/6 mice with subcutaneous injection of MOG35–55/CFA. Mice were scored for 28 days and then sacrificed. The expression of lncRNAs TUG1 and NRON in lumbar spinal cords, activated and controlled macrophages as well as the expression of IL-1, IL-6, and CDe-3 inflammatory cytokines, were assayed by real-time RT-PCR. The lncRNAs TUG1 and NRON were significantly down-regulated in lumbar spinal cords tissues in the acute phase of EAE compared to the control group. TUG1 and NRON were significantly down-regulated in macrophages treated with 10 ng lipopolysaccharide (LPS) compared to the control macrophages. A negative correlation was identified between NRON and TUG1 expression and IL-1, IL-6, and CDe-3 inflammatory cytokines. The present study demonstrates the dysregulation of lncRNAs TUG1 and NRON in spinal cord tissue lesions of EAE and activated macrophages, pointing to their potential role in the pathogenesis of EAE.

How myeloid cells shape experimental autoimmune encephalomyelitis: At the crossroads of outside-in immunity.

Experimental autoimmune encephalomyelitis (EAE) is an animal model of central nervous system (CNS) autoimmunity. It is most commonly used to mimic aspects of multiple sclerosis (MS), a demyelinating disorder of the human brain and spinal cord. The innate immune response displays one of the core pathophysiological features linked to both the acute and chronic stages of MS. Hence, understanding and targeting the innate immune response is essential. Microglia and other CNS resident MUs, as well as infiltrating myeloid cells, diverge substantially in terms of both their biology and their roles in EAE. Recent advances in the field show that antigen presentation, as well as disease-propagating and regulatory interactions with lymphocytes, can be attributed to specific myeloid cell types and cell states in EAE lesions, following a distinct temporal pattern during disease initiation, propagation and recovery. Furthermore, single-cell techniques enable the assessment of characteristic proinflammatory as well as beneficial cell states, and identification of potential treatment targets. Here, we discuss the principles of EAE induction and protocols for varying experimental paradigms, the composition of the myeloid compartment of the CNS during health and disease, and systematically review effects on myeloid cells for therapeutic approaches in EAE.

Nogo receptor-Fc delivered by haematopoietic cells enhances neurorepair in a multiple sclerosis model.

Nogo receptor 1 is the high affinity receptor for the potent myelin-associated inhibitory factors that make up part of the inflammatory extracellular milieu during experimental autoimmune encephalomyelitis. Signalling through the Nogo receptor 1 complex has been shown to be associated with axonal degeneration in an animal model of multiple sclerosis, and neuronal deletion of this receptor homologue, in a disease specific manner, is associated with preserving axons even in the context of neuroinflammation. The local delivery of Nogo receptor(1-310)-Fc, a therapeutic fusion protein, has been successfully applied as a treatment in animal models of spinal cord injury and glaucoma. As multiple sclerosis and experimental autoimmune encephalomyelitis exhibit large numbers of inflammatory cell infiltrates within the CNS lesions, we utilized transplantable haematopoietic stem cells as a cellular delivery method of the Nogo receptor(1-310)-Fc fusion protein. We identified CNS-infiltrating macrophages as the predominant immune-positive cell type that overexpressed myc-tagged Nogo receptor(1-310)-Fc fusion protein at the peak stage of experimental autoimmune encephalomyelitis. These differentiated phagocytes were predominant during the extensive demyelination and axonal damage, which are associated with the engulfment of the protein complex of Nogo receptor(1-310)-Fc binding to myelin ligands. Importantly, mice transplanted with haematopoietic stem cells transduced with the lentiviral vector carrying Nogo receptor(1-310)-Fc and recovered from the peak of neurological decline during experimental autoimmune encephalomyelitis, exhibiting axonal regeneration and eventual remyelination in the white matter tracts. There were no immunomodulatory effects of the transplanted, genetically modified haematopoietic stem cells on immune cell lineages of recipient female mice induced with experimental autoimmune encephalomyelitis. We propose that cellular delivery of Nogo receptor(1-310)-Fc fusion protein through genetically modified haematopoietic stem cells can modulate multifocal experimental autoimmune encephalomyelitis lesions and potentiate neurological recovery.

Expression of antioxidant enzymes in lesions of multiple sclerosis and its models

Oxidative stress promotes tissue injury in the central nervous system in neurological disorders such as multiple sclerosis (MS). To protect against this, antioxidant enzymes including superoxide dismutase-1 (SOD1), heme oxygenase-1 (HO-1), peroxiredoxin-5 (PRDX5) and glutathione peroxidase-4 (GPX4) may be upregulated. However, whether antioxidant enzyme elevation in mouse models of neurodegeneration corresponds to their expression in human diseases such as MS requires investigation. Here, we analyzed and compared the expression of SOD1, HO-1, PRDX5 and GPX4 in the murine spinal cord of three models of MS: focal lesions induced by (1) oxidized phosphatidylcholine or (2) lysophosphatidylcholine (lysolecithin), and (3) diffuse lesions of experimental autoimmune encephalomyelitis. Notably, CD68+ microglia/macrophages were the predominant cellular populations that expressed the highest levels of the detected antioxidant enzymes. Overall, the expression patterns of antioxidant enzymes across the models were similar. The increase of these antioxidant enzymes was corroborated in MS brain tissue using spatial RNA sequencing. Collectively, these results show that antioxidant capacity is relatively conserved between mouse models and MS lesions, and suggest a need to investigate whether the antioxidant elevation in microglia/macrophages is a protective response during oxidative injury, neurodegeneration, and MS.

Fibrin-targeting molecular MRI in inflammatory CNS disorders

BackgroundFibrin deposition is a fundamental pathophysiological event in the inflammatory component of various CNS disorders, such as multiple sclerosis (MS) and Alzheimer’s disease. Beyond its traditional role in coagulation, fibrin elicits immunoinflammatory changes with oxidative stress response and activation of CNS-resident/peripheral immune cells contributing to CNS injury.PurposeTo investigate if CNS fibrin deposition can be determined using molecular MRI, and to assess its capacity as a non-invasive imaging biomarker that corresponds to inflammatory response and barrier impairment.Materials and methodsSpecificity and efficacy of a peptide-conjugated Gd-based molecular MRI probe (EP2104-R) to visualise and quantify CNS fibrin deposition were evaluated. Probe efficacy to specifically target CNS fibrin deposition in murine adoptive-transfer experimental autoimmune encephalomyelitis (EAE), a pre-clinical model for MS (n = 12), was assessed. Findings were validated using immunohistochemistry and laser ablation inductively coupled plasma mass spectrometry. Deposition of fibrin in neuroinflammatory conditions was investigated and its diagnostic capacity for disease staging and monitoring as well as quantification of immunoinflammatory response was determined. Results were compared using t-tests (two groups) or one-way ANOVA with multiple comparisons test. Linear regression was used to model the relationship between variables.ResultsFor the first time (to our knowledge), CNS fibrin deposition was visualised and quantified in vivo using molecular imaging. Signal enhancement was apparent in EAE lesions even 12-h after administration of EP2104-R due to targeted binding (M ± SD, 1.07 ± 0.10 (baseline) vs. 0.73 ± 0.09 (EP2104-R), p = .008), which could be inhibited with an MRI-silent analogue (M ± SD, 0.60 ± 0.14 (EP2104-R) vs. 0.96 ± 0.13 (EP2104-La), p = .006). CNS fibrin deposition corresponded to immunoinflammatory activity (R2 = 0.85, p < .001) and disability (R2 = 0.81, p < .001) in a model for MS, which suggests a clinical role for staging and monitoring. Additionally, EP2104-R showed substantially higher SNR (M ± SD, 6.6 ± 1 (EP2104-R) vs. 2.7 ± 0.4 (gadobutrol), p = .004) than clinically used contrast media, which increases sensitivity for lesion detection.ConclusionsMolecular imaging of CNS fibrin deposition provides an imaging biomarker for inflammatory CNS pathology, which corresponds to pathophysiological ECM remodelling and disease activity, and yields high signal-to-noise ratio, which can improve diagnostic neuroimaging across several neurological diseases with variable degrees of barrier impairment.

Emerging role of neuregulin-1beta1 in pathogenesis and progression of multiple sclerosis

Emerging role of neuregulin-1beta1 in pathogenesis and progression of multiple sclerosis

Targeted Blood Brain Barrier Opening With Focused Ultrasound Induces Focal Macrophage/Microglial Activation in Experimental Autoimmune Encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) is a model of multiple sclerosis (MS). EAE reflects important histopathological hallmarks, dissemination, and diversity of the disease, but has only moderate reproducibility of clinical and histopathological features. Focal lesions are less frequently observed in EAE than in MS, and can neither be constrained to specific locations nor timed to occur at a pre-specified moment. This renders difficult any experimental assessment of the pathogenesis of lesion evolution, including its inflammatory, degenerative (demyelination and axonal degeneration), and reparatory (remyelination, axonal sprouting, gliosis) component processes. We sought to develop a controlled model of inflammatory, focal brain lesions in EAE using focused ultrasound (FUS). We hypothesized that FUS induced focal blood brain barrier disruption (BBBD) will increase the likelihood of transmigration of effector cells and subsequent lesion occurrence at the sonicated location. Lesion development was monitored with conventional magnetic resonance imaging (MRI) as well as with magnetic resonance elastography (MRE) and further analyzed by histopathological means. EAE was induced in 12 6–8 weeks old female C57BL/6 mice using myelin oligodendrocyte glycoprotein (MOG) peptide. FUS-induced BBBD was performed 6, 7, and 9 days after immunization in subgroups of four animals and in an additional control group. MRI and MRE were performed on a 7T horizontal bore small animal MRI scanner. Imaging was conducted longitudinally 2 and 3 weeks after disease induction and 1 week after sonication in control animals, respectively. The scan protocol comprised contrast-enhanced T1-weighted and T2-weighted sequences as well as MRE with a vibration frequency of 1 kHz. Animals were sacrificed for histopathology after the last imaging time point. The overall clinical course of EAE was mild. A total of seven EAE animals presented with focal T2w hyperintense signal alterations in the sonicated hemisphere. These were most frequent in the group of animals sonicated 9 days after immunization. Histopathology revealed foci of activated microglia/macrophages in the sonicated right hemisphere of seven EAE animals. Larger cellular infiltrates or apparent demyelination were not seen. Control animals showed no abnormalities on MRI and did not have clusters of activated microglia/macrophages at the sites targeted with FUS. None of the animals had hemorrhages or gross tissue damage as potential side effects of FUS. EAE-animals tended to have lower values of viscoelasticity and elasticity in the sonicated compared to the contralateral parenchyma. This trend was significant when comparing the right sonicated to the left normal hemisphere and specifically the right sonicated compared to the left normal cortex in animals that underwent FUS-BBBD 9 days after immunization (right vs. left hemisphere: mean viscoelasticity 6.1 vs. 7.2 kPa; p = 0.003 and mean elasticity 4.9 vs. 5.7 kPa, p = 0.024; right vs. left cortex: mean viscoelasticity 5.8 vs. 7.5 kPa; p = 0.004 and mean elasticity 5 vs. 6.5 kPa; p = 0.008). A direct comparison of the biomechanical properties of focal T2w hyperintensities with normal appearing brain tissue did not yield significant results. Control animals showed no differences in viscoelasticity between sonicated and contralateral brain parenchyma. We here provide first evidence for a controlled lesion induction model in EAE using FUS-induced BBBD. The observed lesions in EAE are consistent with foci of activated microglia that may be interpreted as targeted initial inflammatory activity and which have been described as pre-active lesions in MS. Such foci can be identified and monitored with MRI. Moreover, the increased inflammatory activity in the sonicated brain parenchyma seems to have an effect on overall tissue matrix structure as reflected by changes of biomechanical parameters.

Functional characteristics of Th1, Th17, and ex-Th17 cells in EAE revealed by intravital two-photon microscopy

BackgroundT helper (Th) 17 cells are a highly plastic subset of T cells, which in the context of neuroinflammation, are able to acquire pathogenic features originally attributed to Th1 cells (resulting in so called ex-Th17 cells). Thus, a strict separation between the two T cell subsets in the context of experimental autoimmune encephalomyelitis (EAE) is difficult. High variability in culture and EAE induction protocols contributed to previous conflicting results concerning the differential contribution of Th1 and Th17 cells in EAE. Here, we systematically evaluate the role of different T cell differentiation and transfer protocols for EAE disease development and investigate the functional dynamics of encephalitogenic T cells directly within the inflamed central nervous system (CNS) tissue.MethodsWe compiled the currently used EAE induction protocols reported in literature and investigated the influence of the different Th1 and Th17 differentiation protocols as well as EAE induction protocols on the EAE disease course. Moreover, we assessed the cytokine profile and functional dynamics of both encephalitogenic Th1 and Th17 cells in the inflamed CNS using flow cytometry and intravital two-photon laser scanning microscopy. Lastly, we used astrocyte culture and adoptive transfer EAE to evaluate the impact of Th1 and Th17 cells on astrocyte adhesion molecule expression in vitro and in vivo.ResultsWe show that EAE courses are highly dependent on in vitro differentiation and transfer protocols. Moreover, using genetically encoded reporter mice (B6.IL17A-EGFP.acRFP x 2d2/2d2.RFP), we show that the motility of interferon (IFN)γ-producing ex-Th17 cells more closely resembles Th1 cells than Th17 cells in transfer EAE. Mechanistically, IFNγ-producing Th1 cells selectively induce the expression of cellular adhesion molecules I-CAM1 while Th1 as well as ex-Th17 induce V-CAM1 on astrocytes.ConclusionsThe behavior of ex-Th17 cells in EAE lesions in vivo resembles Th1 rather than Th17 cells, underlining that their change in cytokine production is associated with functional phenotype alterations of these cells.

A novel CNS-homing peptide for targeting neuroinflammatory lesions in experimental autoimmune encephalomyelitis

Using phage peptide library screening, we identified peptide-encoding phages that selectively home to the inflamed central nervous system (CNS) of mice with experimental autoimmune encephalomyelitis (EAE), a model of human multiple sclerosis (MS). A phage peptide display library encoding cyclic 9-amino-acid random peptides was first screened ex-vivo for binding to the CNS tissue of EAE mice, followed by in vivo screening in the diseased mice. Phage insert sequences that were present at a higher frequency in the CNS of EAE mice than in the normal (control) mice were identified by DNA sequencing. One of the phages selected in this manner, denoted as MS-1, was shown to selectively recognize CNS tissue in EAE mice. Individually cloned phages with this insert preferentially homed to EAE CNS after an intravenous injection. Similarly, systemically-administered fluorescence-labeled synthetic MS-1 peptide showed selective accumulation in the spinal cord of EAE mice. We suggest that peptide MS-1 might be useful for targeted drug delivery to CNS in EAE/MS.

Folate receptor-targeted positron emission tomography of experimental autoimmune encephalomyelitis in rats

BackgroundFolate receptor-β (FR-β) is a cell surface receptor that is significantly upregulated on activated macrophages during inflammation and provides a potential target for folate-based therapeutic and diagnostic agents. FR-β expression in central nervous system inflammation remains relatively unexplored. Therefore, we used focally induced acute and chronic phases of experimental autoimmune encephalomyelitis (EAE) to study patterns of FR-β expression and evaluated its potential as an in vivo imaging target.MethodsFocal EAE was induced in rats using heat-killed Bacillus Calmette-Guérin followed by activation with complete Freund’s adjuvant supplemented with Mycobacterium tuberculosis. The rats were assessed with magnetic resonance imaging and positron emission tomography/computed tomography (PET/CT) at acute (14 days) and chronic (90 days) phases of inflammation. The animals were finally sacrificed for ex vivo autoradiography of their brains. PET studies were performed using FR-β-targeting aluminum [18F]fluoride-labeled 1,4,7-triazacyclononane-1,4,7-triacetic acid conjugated folate ([18F]AlF-NOTA-folate, 18F-FOL) and 18 kDa translocator protein (TSPO)-targeting N-acetyl-N-(2-[11C]methoxybenzyl)-2-phenoxy-5-pyridinamine (11C-PBR28). Post-mortem immunohistochemistry was performed using anti-FR-β, anti-cluster of differentiation 68 (anti-CD68), anti-inducible nitric oxide synthase (anti-iNOS), and anti-mannose receptor C-type 1 (anti-MRC-1) antibodies. The specificity of 18F-FOL binding was verified using in vitro brain sections with folate glucosamine used as a blocking agent.ResultsImmunohistochemical evaluation of focal EAE lesions demonstrated anti-FR-β positive cells at the lesion border in both acute and chronic phases of inflammation. We found that anti-FR-β correlated with anti-CD68 and anti-MRC-1 immunohistochemistry; for MRC-1, the correlation was most prominent in the chronic phase of inflammation. Both 18F-FOL and 11C-PBR28 radiotracers bound to the EAE lesions. Autoradiography studies verified that this binding took place in areas of anti-FR-β positivity. A blocking assay using folate glucosamine further verified the tracer’s specificity. In the chronic phase of EAE, the lesion-to-background ratio of 18F-FOL was significantly higher than that of 11C-PBR28 (P = 0.016).ConclusionOur EAE results imply that FR-β may be a useful target for in vivo imaging of multiple sclerosis-related immunopathology. FR-β-targeted PET imaging with 18F-FOL may facilitate the monitoring of lesion development and complement the information obtained from TSPO imaging by bringing more specificity to the PET imaging armamentarium for neuroinflammation.

Upregulation of periostin in MOG-induced experimental autoimmune encephalomyelitis in mice

Periostin has dual roles as a multifunctional cytokine and an extracellular matrix protein that binds to several integrins on a variety of cells. This study evaluated whether the expression and cellular localization of periostin changes in the spinal cord of mice with experimental autoimmune encephalomyelitis (EAE), an animal model of human multiple sclerosis. Western blot analysis revealed that periostin was significantly upregulated in the spinal cord of EAE-induced mice in the effector stage, and did not significantly decrease in the recovery stage of EAE, compared with normal control. Periostin was constitutively detected in the neurons, ependymal cells, vascular endothelial cells, and astrocytes of normal mice. In the paralytic stage of EAE, periostin immunoreactivity was detected in some macrophages in the subarachnoid space (SAS), as well as in some microglial cells; in addition, periostin immunoreactivity was enhanced in astrocytes and vascular endothelial cells in EAE lesions. Collectively, these results suggest that periostin, either from monocyte-derived macrophages in SAS or glial cells in the parenchyma, partly facilitates the migration of inflammatory cells in the effector stage of EAE; moreover, periostin in the recovery stage of EAE appears to be involved in the plasticity of damaged central nervous system tissues.

Extrapituitary prolactin promotes generation of Eomes-positive helper T cells mediating neuroinflammation

Induction of eomesodermin-positive CD4+ T cells (Eomes+ T helper [Th] cells) has recently been correlated with the transition from an acute stage to a later stage of experimental autoimmune encephalomyelitis (EAE), an animal model for multiple sclerosis. Moreover, these cells' pathogenic role has been experimentally proven in EAE. While exploring how the pathogenic Eomes+ Th cells are generated during the course of EAE, we unexpectedly found that B cells and MHC class II+ myeloid cells isolated from the late EAE lesions strikingly up-regulated the expression of prolactin (PRL). We demonstrate that such PRL-producing cells have a unique potential to induce Eomes+ Th cells from naïve T cells ex vivo, and that anti-MHC class II antibody could block this process. Furthermore, PRL levels in the cerebrospinal fluid were significantly increased in the late phase of EAE, and blocking the production of PRL by bromocriptine or Zbtb20-specific siRNA significantly reduced the numbers of Eomes+ Th cells in the central nervous system (CNS) and ameliorated clinical signs in the later phase of EAE. The PRL dependency of Eomes+ Th cells was confirmed in a series of in vitro and ex vivo experiments. Collectively, these results indicate that extrapituitary PRL plays a crucial role in the CNS inflammation mediated by pathogenic Eomes+ Th cells. Cellular interactions involving PRL-producing immune cells could be considered as a therapeutic target for the prevention of chronic neuroinflammation.

Microglia pre-activation and neurodegeneration precipitate neuroinflammation without exacerbating tissue injury in experimental autoimmune encephalomyelitis

Human inflammatory or neurodegenerative diseases, such as progressive multiple sclerosis (MS), occur on a background of age-related microglia activation and iron accumulation as well as pre-existing neurodegeneration. Most experimental models for CNS diseases, however, are induced in rodents, which are naturally characterized by a homeostatic microglia phenotype, low cellular iron load and absence of neurodegeneration. Here, we show that naïve LEWzizi rats – Lewis rats with a zitter rat background – show a spontaneous phenotype partly mimicking the changes seen in human aging and particularly in the normal-appearing white and grey matter of patients with progressive MS. Using this model system, we further aimed to investigate (i) whether the acute monophasic MS model experimental autoimmune encephalomyelitis (EAE) transforms into chronic progressive disease and (ii) whether EAE-induced neuroinflammation and tissue damage aggravate on the LEWzizi background. We found that the pre-existing LEWzizi-specific pathology precipitated EAE-related neuroinflammation into forebrain areas, which are devoid of EAE lesions in normal Lewis rats. However, EAE-related tissue damage was neither modified by the LEWzizi-specific pathology nor did EAE-induced neuroinflammation modify the LEWzizi-related pathological process. Our data indicate that the interaction between pre-activated microglia and CD4+ autoreactive T cells during the induction and propagation of tissue damage in the CNS is limited.

The role of phospholipase A2 in multiple Sclerosis: A systematic review and meta-analysis

Phospholipases A2 (PLA2) are a diverse group of enzymes that cleave the fatty acids of membrane phospholipids. They play critical roles in pathogenesis of neurodegenerative diseases such as multiple sclerosis by enhancing oxidative stress and initiating inflammation. The levels of PLA2 activity in MS patients compared to controls and role of inhibiting PLA2 activity on severity scores in different experimental models are not comprehensively assessed in the light of varying evidence from published studies. The objective of this systematic review is to determine the association between PLA2 activity and multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE). We performed a systematic review of six studies that assessed PLA2 activity in MS patients compared to controls and nine studies that assessed PLA2 activity in EAE. sPLA2 nor Lp-PLA2 activity were not increased in MS compared to controls in five of those six studies. A difference in sPLA2 activity was only found in a study that measured the enzyme activity in urine. However, inhibiting cPLA2 or sPLA2 led to lower clinical severity or no signs of EAE in mice, and a lower incidence of EAE lesions compared to animals without cPLA2 inhibition. These findings indicate that PLA2 appears to play a role in the pathogenesis of EAE.

CNS-resident classical DCs play a critical role in CNS autoimmune disease.

Experimental autoimmune encephalomyelitis (EAE) is an inflammatory demyelinating disease of the central nervous system (CNS), induced by the adoptive transfer of myelin-reactive CD4+ T cells into naive syngeneic mice. It is widely used as a rodent model of multiple sclerosis (MS). The development of EAE lesions is initiated when transferred CD4+ T cells access the CNS and are reactivated by local antigen-presenting cells (APCs) bearing endogenous myelin peptide/MHC class II complexes. The identity of the CNS-resident, lesion-initiating APCs is widely debated. Here we demonstrate that classical dendritic cells (cDCs) normally reside in the meninges, brain, and spinal cord in the steady state. These cells are unique among candidate CNS APCs in their ability to stimulate naive, as well as effector, myelin-specific T cells to proliferate and produce proinflammatory cytokines directly ex vivo. cDCs expanded in the meninges and CNS parenchyma in association with disease progression. Selective depletion of cDCs led to a decrease in the number of myelin-primed donor T cells in the CNS and reduced the incidence of clinical EAE by half. Based on our findings, we propose that cDCs, and the factors that regulate them, be further investigated as potential therapeutic targets in MS.

Vascular adhesion protein-1 is actively involved in the development of inflammatory lesions in rat models of multiple sclerosis

BackgroundVascular adhesion protein-1 (VAP-1) is an inflammation-inducible endothelial cell molecule and primary amine oxidase that mediates leukocyte entry to sites of inflammation. However, there is limited knowledge of the inflammation-related expression of VAP-1 in the central nervous system (CNS). Therefore, we investigated the expression of VAP-1 within the CNS vasculature in two focal rat models of experimental autoimmune encephalomyelitis (EAE) mimicking multiple sclerosis (MS).MethodsEAE was induced either with Bacillus Calmette-Guérin, resulting in a delayed-type hypersensitivity-like pathogenesis (fDTH-EAE), or with myelin oligodendrocyte glycoprotein (fMOG-EAE). A subgroup of fMOG-EAE rats were treated daily with a selective VAP-1 inhibitor (LJP1586; 5 mg/kg). On 3 and 14 days after lesion activation, rat brains were assessed using magnetic resonance imaging (MRI), and ex vivo autoradiography was conducted to evaluate the binding of Gallium-68-labelled VAP-1 ligand. Histology and immunohistochemistry (OX-42, VAP-1, intercellular adhesion protein-1 [ICAM-1], P-selectin) supported the ex vivo autoradiography.ResultsEAE lesions showed MRI-detectable signal changes and binding of the VAP-1-targeting radiotracer in both rat models. Some of the VAP-1 positive vessels showed morphological features typical for high endothelial-like venules at sites of inflammation. Inhibition of VAP-1 activity with small molecule inhibitor, LJP1586, decreased lymphocyte density in the acute inflammatory phase of fMOG-EAE lesions (day 3, P = 0.026 vs. untreated), but not in the remission phase (day 14, P = 0.70 vs. untreated), and had no effect on the amount of OX-42-positive cells in either phase. LJP1586 treatment reduced VAP-1 and ICAM-1 expression in the acute inflammatory phase, whereas P-selectin remained not detectable at all studied stages of the disease.ConclusionsOur results revealed that VAP-1 is expressed and functionally active in vasculature within the induced focal EAE lesions during the acute phase of inflammation and remains expressed after the acute inflammation has subsided. The study indicates that VAP-1 is actively involved in the development of inflammatory CNS lesions. During this process, the endothelial cell lesion-related vasculature seem to undergo a structural transformation from regular flat-walled endothelium to HEV-like endothelium.

Spatiotemporal distribution of fibrinogen in marmoset and human inflammatory demyelination

Multiple sclerosis is an inflammatory demyelinating disease of the central nervous system. Although it has been extensively studied, the proximate trigger of the immune response remains uncertain. Experimental autoimmune encephalomyelitis in the common marmoset recapitulates many radiological and pathological features of focal multiple sclerosis lesions in the cerebral white matter, unlike traditional experimental autoimmune encephalomyelitis in rodents. This provides an opportunity to investigate how lesions form as well as the relative timing of factors involved in lesion pathogenesis, especially during early stages of the disease. We used MRI to track experimental autoimmune encephalomyelitis lesions in vivo to determine their age, stage of development, and location, and we assessed the corresponding histopathology post-mortem. We focused on the plasma protein fibrinogen-a marker for blood-brain barrier leakage that has also been linked to a pathogenic role in inflammatory demyelinating lesion development. We show that fibrinogen has a specific spatiotemporal deposition pattern, apparently deriving from the central vein in early experimental autoimmune encephalomyelitis lesions <6 weeks old, and preceding both demyelination and visible gadolinium enhancement on MRI. Thus, fibrinogen leakage is one of the earliest detectable events in lesion pathogenesis. In slightly older lesions, fibrinogen is found inside microglia/macrophages, suggesting rapid phagocytosis. Quantification demonstrates positive correlation of fibrinogen deposition with accumulation of inflammatory cells, including microglia/macrophages and T cells. The peak of fibrinogen deposition coincides with the onset of demyelination and axonal loss. In samples from chronic multiple sclerosis cases, fibrinogen was found at the edge of chronic active lesions, which have ongoing demyelination and inflammation, but not in inactive lesions, suggesting that fibrinogen may play a role in sustained inflammation even in the chronic setting. In summary, our data support the notion that fibrinogen is a key player in the early pathogenesis, as well as sustained inflammation, of inflammatory demyelinating lesions.

An Update on the Role of Matrix Metalloproteinases in the Pathogenesis of Multiple Sclerosis.

Matrix metalloproteinases (MMPs) are zinc-containing endopeptidases, proteins essential to the degradation of various tissue extracellular matrix proteins. Under normal conditions, MMPs participate in several physiological processes, both in the developing organism and the adult. Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system (CNS), affecting primarily young adults. Inflammatory infiltrations of the CNS parenchyma by autoreactive immune cells, that mediate myelin degradation in the form of the demyelination "plaque", are the pathological hallmark of the disease. Due to their capacity to orchestrate tissue penetration from various cells, MMPs have been elucidated in MS as mediators of blood-brain barrier disruption and CNS inflammation, thus contributing to the disease pathogenesis. This review focuses on clinical and experimental evidence of MMPs' pathogenetic role in MS and its animal model, experimental autoimmune encephalomyelitis (EAE). A MEDLINE search was performed for using the following terms: "metalloproteinases", "multiple sclerosis", "experimental autoimmune encephalomyelitis", "central nervous system" and "autoimmunity". Expression patterns of MMPs and their specific inhibitor molecules at the sites of MS and EAE lesions, as well as by specific cell types of the immune system, provide evidence of MMPs' role in the pathogenesis of CNS autoimmunity. Clinical evidence suggests differential profile of MMPs' expression in serum and cerebrospinal fluid (CSF) between MS patients with various disease types and healthy adults, rendering MMPs potential biomarkers for disease incidence and activity. MMPs' role in the processes of CNS inflammation, de- and remyelination, confers implications as therapeutic targets, either alone, or in relation with widely-used disease modifying treatments in MS.