Magnetic resonance elastography reveals altered brain viscoelasticity in experimental autoimmune encephalomyelitis

Cerebral magnetic resonance elastography (MRE) measures the viscoelastic properties of brain tissues in vivo. It was recently shown that brain viscoelasticity is reduced in patients with multiple sclerosis (MS), highlighting the potential of cerebral MRE to detect tissue pathology during neuroinflammation. To further investigate the relationship between inflammation and brain viscoelasticity, we applied MRE to a mouse model of MS, experimental autoimmune encephalomyelitis (EAE). EAE was induced and monitored by MRE in a 7-tesla animal MRI scanner over 4 weeks. At the peak of the disease (day 14 after immunization), we detected a significant decrease in both the storage modulus (G') and the loss modulus (G″), indicating that both the elasticity and the viscosity of the brain are reduced during acute inflammation. Interestingly, these parameters normalized at a later time point (day 28) corresponding to the clinical recovery phase. Consistent with this, we observed a clear correlation between viscoelastic tissue alteration and the magnitude of perivascular T cell infiltration at both day 14 and day 28. Hence, acute neuroinflammation is associated with reduced mechanical cohesion of brain tissues. Moreover, the reduction of brain viscoelasticity appears to be a reversible process, which is restored when inflammation resolves. For the first time, our study has demonstrated the applicability of cerebral MRE in EAE, and showed that this novel imaging technology is highly sensitive to early tissue alterations resulting from the inflammatory processes. Thus, MRE may serve to monitor early stages of perivascular immune infiltration during neuroinflammation

Gender effect on neurodegeneration and myelin markers in an animal model for multiple sclerosis

Multiple sclerosis (MS) varies considerably in its incidence and progression in females and males. In spite of clinical evidence, relatively few studies have explored molecular mechanisms possibly involved in gender-related differences. The present study describes possible cellular- and molecular-involved markers which are differentially regulated in male and female rats and result in gender-dependent EAE evolution and progression. Attention was focused on markers of myelination (MBP and PDGF\u3b1R) and neuronal distress and/or damage (GABA synthesis enzymes, GAD65 and GAD67, NGF, BDNF and related receptors), in two CNS areas, i.e. spinal cord and cerebellum, which are respectively severely and mildly affected by inflammation and demyelination. Tissues were sampled during acute, relapse/remission and chronic phases and results were analysed by two-way ANOVA

Purkinje cell loss in experimental autoimmune encephalomyelitis

Gray matter atrophy observed by brain MRI is an important correlate to clinical disability and disease duration in multiple sclerosis. The objective of this study was to link brain atrophy visualized by neuroimaging to its underlying neuropathology using the MS model, experimental autoimmune encephalomyelitis (EAE). Volumetric changes in brains of EAE mice, as well as matched healthy normal controls, were quantified by collecting post-mortem high-resolution T2-weighted magnetic resonance microscopy and actively stained magnetic resonance histology images. Anatomical delineations demonstrated a significant decrease in the volume of the whole cerebellum, cerebellar cortex, and molecular layer of the cerebellar cortex in EAE as compared to normal controls. The pro-apoptotic marker caspase-3 was detected in Purkinje cells and a significant decrease in Purkinje cell number was found in EAE. Cross modality and temporal correlations revealed a significant association between Purkinje cell loss on neuropathology and atrophy of the molecular layer of the cerebellar cortex by neuroimaging. These results demonstrate the power of using combined population atlasing and neuropathology approaches to discern novel insights underlying gray matter atrophy in animal models of neurodegenerative disease

BASHY Dye Platform Enables the Fluorescence Bioimaging of Myelin Debris Phagocytosis by Microglia during Demyelination

Multiple sclerosis (MS) is a demyelinating disease of the central nervous system that is characterized by the presence of demyelinated regions with accumulated myelin lipid debris. Importantly, to allow effective remyelination, such debris must be cleared by microglia. Therefore, the study of microglial activity with sensitive tools is of great interest to better monitor the MS clinical course. Using a boronic acid-based (BASHY) fluorophore, specific for nonpolar lipid aggregates, we aimed to address BASHY’s ability to label nonpolar myelin debris and image myelin clearance in the context of demyelination. Demyelinated ex vivo organotypic cultures (OCSCs) and primary microglia cells were immunostained to evaluate BASHY’s co-localization with myelin debris and also to evaluate BASHY’s specificity for phagocytosing cells. Additionally, mice induced with experimental autoimmune encephalomyelitis (EAE) were injected with BASHY and posteriorly analyzed to evaluate BASHY+ microglia within demyelinated lesions. Indeed, in our in vitro and ex vivo studies, we showed a significant increase in BASHY labeling in demyelinated OCSCs, mostly co-localized with Iba1-expressing amoeboid/phagocytic microglia. Most importantly, BASHY’s presence was also found within demyelinated areas of EAE mice, essentially co-localizing with lesion-associated Iba1+ cells, evidencing BASHY’s potential for the in vivo bioimaging of myelin clearance and myelin-carrying microglia in regions of active demyelination.This work was supported by Grant for Multiple Sclerosis Innovation-Merck Serono to AF, Young Investigator’s Projects for Collaborative Cross-disciplinary Studies from iMed.ULisboa to A.F. and F.S.; PTDC/QUI-QOR/29967/2017 and LISBOA-01-0145-FEDER-029967 to P.G.; the Spanish Ministry of Science, Innovation, and Universities (CTQ2017-89832-P for U.P.); the PhD grant 2021.04911.BD from Fundação para a Ciência e Tecnologia, Portugal (FCT) to M.V.P.; and in part by UIDB/04138/2020 and UIDP/04138/2020-from FCT to iMed.ULisboa

Integración de datos de imagen molecular y expresión génica

As far as all the background information about atlases and gene expression databases has already been analysed, now we need to define further the project, its objectives and facts of interest. Gene expression databases, in most cases, do not provide any kind of integration with anatomical information of where those genes are expressed. The characterization of the whole transcriptome for structures like the brain is of limited utility if we have no anatomical information. Combining the databases with the anatomical information provided by an anatomical atlas, we can have lots of advantages. First of all, the most immediate advantage that such integration would introduce would be user-friendliness. Part of this problem is solved with the aGEM tool already developed, that integrates different databases into a single user interface.Visual representation of the gene locations would improve user experience if it is integrated with aGEM. As a second advantage, this integration could facilitate the connection between imaging and gene expression information when defining or analysing results from preclinical experiments. The definition of an imaging protocol in order to study the phenotype of a transgenic animal model could benefit from the results of this project, since the researcher could look for anatomical structures related to the genes that have been manipulated. The results of image quantification are usually an statistical parametric map, that presents the statistical significance of a certain analysis for every voxel. Significant areas from this image could be related to the underlying genes by means of the proposed integration tool. So, the main objective of the project is to connect all the information provided by aGEM and by the atlas. For the issue, it is needed to find how the information is stored and related in aGEM, in order to extract enough information of interest to program a first version of the tool. Also, it is required to study which atlases are available and which one is the most suitable for our purposes. Once all these steps have been done, the kind of program which is going to be developed needs to be analysed. There are several possibilities, like a program in Java, C++, Matlab or a plugin for its integration in ImageJ. Once all the necessary information is extracted, an integration step must be done for the program to be operative. Then, when the information mapping is ready, the interface of the program can be written. The first version of program should be able to perform certain query types: 1. Anatomy query: Given a list of anatomy structures, the user should be able to select any of them and the program would launch a query showing the genes and their information expressed in the structure, which would be shown in the atlas images. 2. Gene query: The user selects a gene and the program would detect in which structures is the gene expressed and show them in the atlas images. The purpose in a first stage is to integrate part of the information contained in aGEM in a beta trial version of the interface, in order to check its real utility.Ingeniería Biomédic

Sex differences in the neuronal transcriptome and synaptic mitochondrial function in the cerebral cortex of a multiple sclerosis model

IntroductionMultiple sclerosis (MS) affects the cerebral cortex, inducing cortical atrophy and neuronal and synaptic pathology. Despite the fact that women are more susceptible to getting MS, men with MS have worse disability progression. Here, sex differences in neurodegenerative mechanisms are determined in the cerebral cortex using the MS model, chronic experimental autoimmune encephalomyelitis (EAE).MethodsNeurons from cerebral cortex tissues of chronic EAE, as well as age-matched healthy control, male and female mice underwent RNA sequencing and gene expression analyses using RiboTag technology. The morphology of mitochondria in neurons of cerebral cortex was assessed using Thy1-CFP-MitoS mice. Oxygen consumption rates were determined using mitochondrial respirometry assays from intact as well as permeabilized synaptosomes.ResultsRNA sequencing of neurons in cerebral cortex during chronic EAE in C57BL/6 mice showed robust differential gene expression in male EAE compared to male healthy controls. In contrast, there were few differences in female EAE compared to female healthy controls. The most enriched differential gene expression pathways in male mice during EAE were mitochondrial dysfunction and oxidative phosphorylation. Mitochondrial morphology in neurons showed significant abnormalities in the cerebral cortex of EAE males, but not EAE females. Regarding function, synaptosomes isolated from cerebral cortex of male, but not female, EAE mice demonstrated significantly decreased oxygen consumption rates during respirometry assays.DiscussionCortical neuronal transcriptomics, mitochondrial morphology, and functional respirometry assays in synaptosomes revealed worse neurodegeneration in male EAE mice. This is consistent with worse neurodegeneration in MS men and reveals a model and a target to develop treatments to prevent cortical neurodegeneration and mitigate disability progression in MS men

Young adult born neurons enhance hippocampal dependent performance via influences on bilateral networks

Adult neurogenesis supports performance in many hippocampal dependent tasks. Considering the small number of adult-born neurons generated at any given time, it is surprising that this sparse population of cells can substantially influence behavior. Recent studies have demonstrated that heightened excitability and plasticity may be critical for the contribution of young adult-born cells for certain tasks. What is not well understood is how these unique biophysical and synaptic properties may translate to networks that support behavioral function. Here we employed a location discrimination task in mice while using optogenetics to transiently silence adult-born neurons at different ages. We discovered that adult-born neurons promote location discrimination during early stages of development but only if they undergo maturation during task acquisition. Silencing of young adult-born neurons also produced changes extending to the contralateral hippocampus, detectable by both electrophysiology and fMRI measurements, suggesting young neurons may modulate location discrimination through influences on bilateral hippocampal networks.United States. National Institutes of Health (1DP2NS082126)National Institute of Mental Health (U.S.) (5R00MH085944)United States. National Institutes of Health (R01-DA028299)United States. Defense Advanced Research Projects Agency (W911NF-10-0059)Pew Charitable TrustsAmerican Federation for Aging ResearchAlfred P. Sloan FoundationNational Institute of Mental Health (U.S.) (1R21MH109941

Estrogen treatment prevents gray matter atrophy in experimental autoimmune encephalomyelitis

Gray matter atrophy is an important correlate to clinical disability in multiple sclerosis (MS), and many treatment trials include atrophy as an outcome measure. Atrophy has been shown to occur in experimental autoimmune encephalomyelitis (EAE), the most commonly used animal model of MS. The clinical severity of EAE is reduced in estrogen-reated mice, but it remains unknown whether estrogen treatment can reduce gray matter atrophy in EAE. In this study, mice with EAE were treated with either estrogen receptor (ER)-α ligand or ER-β ligand, and diffusion tensor images (DTI) were collected and neuropathology was performed. DTI showed atrophy in the cerebellar gray matter of vehicle-treated EAE mice compared with healthy controls but not in ER-α or ER-β ligand-treated EAE mice. Neuropathology demonstrated that Purkinje cell numbers were decreased in vehicle-treated EAE mice, whereas neither ER ligand-treated EAE groups showed a decrease. This is the first report of a neuroprotective therapy in EAE that unambiguously prevents gray matter atrophy while sparing a major neuronal cell type. Fractional anisotropy (FA) in the cerebellar white matter was decreased in vehicle- and ER-β ligand-treated but not in ER-α ligand-treated EAE mice. Inflammatory cell infiltration was increased in vehicle- and ER-β ligand-treated but not in ER-α ligand-treated EAE mice. Myelin staining was decreased in vehicle-treated EAE mice and was spared in both ER ligand-treated groups. This is consistent with decreased FA as a potential biomarker for inflammation rather than myelination or axonal damage in the cerebellum in EAE

Sex differences in autoimmune diseases

Women are more susceptible to a variety of autoimmune diseases including systemic lupus erythematosus (SLE), multiple sclerosis (MS), primary biliary cirrhosis, rheumatoid arthritis and Hashimoto's thyroiditis. This increased susceptibility in females compared to males is also present in animal models of autoimmune diseases such as spontaneous SLE in (NZBxNZW)F1 and NZM.2328 mice, experimental autoimmune encephalomyelitis (EAE) in SJL mice, thyroiditis, Sjogren's syndrome in MRL/Mp-lpr/lpr mice and diabetes in non-obese diabetic mice. Indeed, being female confers a greater risk of developing these diseases than any single genetic or environmental risk factor discovered to date. Understanding how the state of being female so profoundly affects autoimmune disease susceptibility would accomplish two major goals. First, it would lead to an insight into the major pathways of disease pathogenesis and, secondly, it would likely lead to novel treatments which would disrupt such pathways

Current developments in MRI for assessing rodent models of multiple sclerosis

MRI is a key radiological imaging technique that plays an important role in the diagnosis and characterization of heterogeneous multiple sclerosis (MS) lesions. Various MRI methodologies such as conventional T 1/T 2 contrast, contrast agent enhancement, diffusion-weighted imaging, magnetization transfer imaging and susceptibility weighted imaging have been developed to determine the severity of MS pathology, including demyelination/remyelination and brain connectivity impairment from axonal loss. The broad spectrum of MS pathology manifests in diverse patient MRI presentations and affects the accuracy of patient diagnosis. To study specific pathological aspects of the disease, rodent models such as experimental autoimmune encephalomyelitis, virus-induced and toxin-induced demyelination have been developed. This review aims to present key developments in MRI methodology for better characterization of rodent models of MS