Subpial Demyelination Research Articles

Cross-talk between B cells, microglia and macrophages, and implications to central nervous system compartmentalized inflammation and progressive multiple sclerosis

B cells can be enriched within meningeal immune-cell aggregates of multiple sclerosis (MS) patients, adjacent to subpial cortical demyelinating lesions now recognized as important contributors to progressive disease. This subpial demyelination is notable for a 'surface-in' gradient of neuronal loss and microglial activation, potentially reflecting the effects of soluble factors secreted into the CSF. We previously demonstrated that MS B-cell secreted products are toxic to oligodendrocytes and neurons. The potential for B-cell-myeloid cell interactions to propagate progressive MS is of considerable interest. Secreted products of MS-implicated pro-inflammatory effector B cells or IL-10-expressing B cells with regulatory potential were applied to human brain-derived microglia or monocyte-derived macrophages, with subsequent assessment of myeloid phenotype and function through measurement of their expression of pro-inflammatory, anti-inflammatory and homeostatic/quiescent molecules, and phagocytosis (using flow cytometry, ELISA and fluorescently-labeled myelin). Effects of secreted products of differentially activated microglia on B-cell survival and activation were further studied. Secreted products of MS-implicated pro-inflammatory B cells (but not IL-10 expressing B cells) substantially induce pro-inflammatory cytokine (IL-12, IL-6, TNFα) expression by both human microglia and macrophage (in a GM-CSF dependent manner), while down-regulating their expression of IL-10 and of quiescence-associated molecules, and suppressing their myelin phagocytosis. In contrast, secreted products of IL-10 expressing B cells upregulate both human microglia and macrophage expression of quiescence-associated molecules and enhance their myelin phagocytosis. Secreted factors from pro-inflammatory microglia enhance B-cell activation. Potential cross-talk between disease-relevant human B-cell subsets and both resident CNS microglia and infiltrating macrophages may propagate CNS-compartmentalized inflammation and injury associated with MS disease progression. These interaction represents an attractive therapeutic target for agents such as Bruton's tyrosine kinase inhibitors (BTKi) that modulate responses of both B cells and myeloid cells. Stated in Acknowledgments section of manuscript.

An Animal Model for Chronic Meningeal Inflammation and Inflammatory Demyelination of the Cerebral Cortex.

Modeling chronic cortical demyelination allows the study of long-lasting pathological changes observed in multiple sclerosis such as failure of remyelination, chronically disturbed functions of oligodendrocytes, neurons and astrocytes, brain atrophy and cognitive impairments. We aimed at generating an animal model for studying the consequences of chronic cortical demyelination and meningeal inflammation. To induce long-lasting cortical demyelination and chronic meningeal inflammation, we immunized female Lewis rats against myelin oligodendrocyte glycoprotein (MOG) and injected lentiviruses for continuing overexpression of the cytokines TNFα and IFNγ in the cortical brain parenchyma. Immunization with MOG and overexpression of TNFα and IFNγ led to widespread subpial demyelination and meningeal inflammation that were stable for at least 10 weeks. We demonstrate here that immunization with MOG is necessary for acute as well as chronic cortical demyelination. In addition, long-lasting overexpression of TNFα and IFNγ in the brain parenchyma is sufficient to induce chronic meningeal inflammation. Our model simulates key features of chronic cortical demyelination and inflammation, reminiscent of human multiple sclerosis pathology. This will allow molecular, cellular and functional investigations for a better understanding of the adaptation mechanisms of the cerebral cortex in multiple sclerosis.

Lymphotoxin-alpha expression in the meninges causes lymphoid tissue formation and neurodegeneration.

Organized meningeal immune cell infiltrates are suggested to play an important role in cortical grey matter pathology in the multiple sclerosis brain, but the mechanisms involved are as yet unresolved. Lymphotoxin-alpha plays a key role in lymphoid organ development and cellular cytotoxicity in the immune system and its expression is increased in the CSF of naïve and progressive multiple sclerosis patients and post-mortem meningeal tissue. Here we show that persistently increased levels of lymphotoxin-alpha in the cerebral meninges can give rise to lymphoid-like structures and underlying multiple sclerosis-like cortical pathology. Stereotaxic injections of recombinant lymphotoxin-alpha into the rat meninges led to acute meningeal inflammation and subpial demyelination that resolved after 28 days, with demyelination being dependent on prior subclinical immunization with myelin oligodendrocyte glycoprotein. Injection of a lymphotoxin-alpha lentiviral vector into the cortical meningeal space, to produce chronic localized overexpression of the cytokine, induced extensive lymphoid-like immune cell aggregates, maintained over 3 months, including T-cell rich zones containing podoplanin + fibroblastic reticular stromal cells and B-cell rich zones with a network of follicular dendritic cells, together with expression of lymphoid chemokines and their receptors. Extensive microglial and astroglial activation, subpial demyelination and marked neuronal loss occurred in the underlying cortical parenchyma. Whereas subpial demyelination was partially dependent on previous myelin oligodendrocyte glycoprotein immunization, the neuronal loss was present irrespective of immunization. Conditioned medium from LTα treated microglia was able to induce a reactive phenotype in astrocytes. Our results show that chronic lymphotoxin-alpha overexpression alone is sufficient to induce formation of meningeal lymphoid-like structures and subsequent neurodegeneration, similar to that seen in the progressive multiple sclerosis brain.

Age-dependent gray matter demyelination is associated with leptomeningeal neutrophil accumulation.

People living with multiple sclerosis (MS) experience episodic CNS white matter lesions instigated by autoreactive T cells. With age, patients with MS show evidence of gray matter demyelination and experience devastating nonremitting symptomology. What drives progression is unclear and studying this has been hampered by the lack of suitable animal models. Here, we show that passive experimental autoimmune encephalomyelitis (EAE) induced by an adoptive transfer of young Th17 cells induced a nonremitting clinical phenotype that was associated with persistent leptomeningeal inflammation and cortical pathology in old, but not young, SJL/J mice. Although the quantity and quality of T cells did not differ in the brains of old versus young EAE mice, an increase in neutrophils and a decrease in B cells were observed in the brains of old mice. Neutrophils were also found in the leptomeninges of a subset of progressive MS patient brains that showed evidence of leptomeningeal inflammation and subpial cortical demyelination. Taken together, our data show that while Th17 cells initiate CNS inflammation, subsequent clinical symptoms and gray matter pathology are dictated by age and associated with other immune cells, such as neutrophils.

Age-related susceptibility to grey matter demyelination and neurodegeneration is associated with meningeal neutrophil accumulation in an animal model of MS

Abstract People living with multiple sclerosis (MS) experience episodic central nervous system (CNS) white matter lesions instigated by autoreactive T cells. With age, MS patients show evidence of grey matter demyelination and experience devastating non-remitting symptomology. What drives progression is unclear and has been hampered by the lack of suitable animal models. Here we show that passive experimental autoimmune encephalomyelitis (EAE) induced by an adoptive transfer of young Th17 cells induces a non-remitting clinical phenotype that is associated with persistent leptomeningeal inflammation and cortical pathology in old, but not young SJL/J mice. While the quantity and quality of T cells did not differ in the brains of old vs young EAE mice, an increase in neutrophils and a decrease in B cells was observed in the brains of old mice. Neutrophils were also found in the leptomeninges of a subset of progressive MS patient brains that showed evidence of leptomeningeal inflammation and subpial cortical demyelination. Taken together, our data show that while Th17 cells initiate CNS inflammation, subsequent clinical symptoms and grey matter pathology are dictated by age and associated with other immune cells such as neutrophils.

Accumulation of meningeal lymphocytes correlates with white matter lesion activity in progressive multiple sclerosis.

Subpial cortical demyelination is an important component of multiple sclerosis (MS) pathology contributing to disease progression, yet mechanism(s) underlying its development remain unclear. Compartmentalized inflammation involving the meninges may drive this type of injury. Given recent findings identifying substantial white matter (WM) lesion activity in patients with progressive MS, elucidating whether and how WM lesional activity relates to meningeal inflammation and subpial cortical injury is of interest. Using postmortem FFPE tissue blocks (range, 5–72 blocks; median, 30 blocks) for each of 27 patients with progressive MS, we assessed the relationship between meningeal inflammation, the extent of subpial cortical demyelination, and the state of subcortical WM lesional activity. Meningeal accumulations of T cells and B cells, but not myeloid cells, were spatially adjacent to subpial cortical lesions, and greater immune cell accumulation was associated with larger subpial lesion areas. Patients with a higher extent of meningeal inflammation harbored a greater proportion of active and mixed active/inactive WM lesions and an overall lower proportion of inactive and remyelinated WM lesions. Our findings support the involvement of meningeal lymphocytes in subpial cortical injury and point to a potential link between inflammatory subpial cortical demyelination and pathological mechanisms occurring in the subcortical WM.

Ageing promotes non-remitting experimental autoimmune encephalomyelitis with persistent meningeal inflammation and subpial demyelination in A/T SJL/J mice.

Abstract Current therapies for multiple sclerosis (MS) reduce the frequency of relapses but fail to prevent transition from relapse-remitting into progressive MS. Subpial demyelination, meningeal inflammation and neurodegeneration are hallmarks of progressive MS, however there is no single animal model that mimics all these features. Adoptive transfer of proteolipid protein (PLP)-primed Th17 cells into young (~12 weeks old) SJL/J recipient mice induces experimental autoimmune encephalomyelitis (EAE) characterized by subpial demyelination associated with meningeal tertiary lymphoid tissues (TLTs), however young mice develop remitting, non-progressive disease. Since age is a risk factor for transitioning to progressive MS, we hypothesized that adoptive transfer of PLP-primed Th17 cells into old (~12 months old) SJL/J recipient mice would promote a progressive phenotype. Indeed, transfer of cells into aged recipient mice resulted in a non-remitting EAE compared to young recipients despite a similar ability to produce T cell derived cytokines in the CNS at peak disease. However, old mice showed persistent meningeal inflammation as evidenced by more B/T cell rich TLTs in the meninges compared to young mice, and developed increased CNS pathology (subpial demyelination, microglia/macrophage activation, glial limitans disruption, axonal injury, loss of synapses), whereas young EAE mice had recovered most of the pathology seen at peak of disease by day 25 post-transfer. In summary, our model provides an opportunity to assess how subpial demyelination associated with meningeal inflammation impacts the transition from relapse-remitting to progressive MS in a single animal model.

Impact of B cell-depletion on plasma cells in a pre-clinical model of meningeal inflammation and subpial demyelination

Abstract Anti-CD20 B cell depletion therapy has provided evidence for the pathogenic roles of B cells in multiple sclerosis (MS). Terminally differentiated plasma cells (PCs) are spared by anti-CD20, and our lab has previously shown that during experimental autoimmune encephalomyelitis (EAE), IgA+ PCs mediate protection against neuroinflammation. One hypothesis is that in addition to depleting pro-inflammatory B cells, anti-CD20 also removes the major consumers of BAFF (i.e. B cells) thus promoting a favourable niche for regulatory PCs and may represent an additional mechanism by which anti-CD20 therapy confers protection. Adoptive transfer EAE of PLP-primed Th17 cells into SJL/J mice leads to ascending paralysis and meningeal inflammation which parallels brain pathology seen in human MS. Mice were prophylactically treated with anti-CD20 (10mg/kg) or isotype control antibodies which we re-administered throughout the course of the experiment and monitored daily for clinical symptoms. We find that anti-CD20 treatment effectively depleted B cells (but not T cells) in the periphery; within the central nervous system, meningeal immune clusters formed in both anti-CD20 and isotype control treated mice, with the former being devoid of B220+ B cells. Compared to control treated mice, anti-CD20 treated mice showed limited subpial demyelination and pathology adjacent to B cell-less meningeal immune clusters, and is accompanied by a reduction in clinical score. In the serum, anti-CD20 treatment elevated levels of BAFF and IgA, which inversely correlated with disease severity. We thus propose that anti-CD20 exerts its protective effects by providing a favourable niche for suppressive IgA+ PCs.

Selective vulnerability of inhibitory networks in multiple sclerosis

In multiple sclerosis (MS), a chronic demyelinating disease of the central nervous system, neurodegeneration is detected early in the disease course and is associated with the long-term disability of patients. Neurodegeneration is linked to both inflammation and demyelination, but its exact cause remains unknown. This gap in knowledge contributes to the current lack of treatments for the neurodegenerative phase of MS. Here we ask if neurodegeneration in MS affects specific neuronal components and if it is the result of demyelination. Neuropathological examination of secondary progressive MS motor cortices revealed a selective vulnerability of inhibitory interneurons in MS. The generation of a rodent model of focal subpial cortical demyelination reproduces this selective neurodegeneration providing a new preclinical model for the study of neuroprotective treatments.

Persistent elevation of intrathecal pro-inflammatory cytokines leads to multiple sclerosis-like cortical demyelination and neurodegeneration

Analysis of isolated meninges and cerebrospinal fluid (CSF) of post-mortem MS cases has shown increased gene and protein expression for the pro-inflammatory cytokines: tumour necrosis factor (TNF) and interferon-γ (IFNγ). Here we tested the hypothesis that persistent production of these cytokines in the meningeal compartment and diffusion into underlying GM can drive chronic MS-like GM pathology. Lentiviral transfer vectors were injected into the sagittal sulcus of DA rats to deliver continuous expression of TNF + IFNγ transgenes in the meninges and the resulting neuropathology analysed after 1 and 2 months. Injection of TNF + IFNγ viral vectors, with or without prior MOG immunisation, induced extensive immune cell infiltration (CD4+ and CD8+ T-cells, CD79a + B-cells and macrophages) in the meninges by 28 dpi, which remained at 2 months. Control GFP viral vector did not induce infiltration. Subpial demyelination was seen underlying these infiltrates, which was partly dependant on prior myelin oligodendrocyte glycoprotein (MOG) immunisation. A significant decrease in neuronal numbers was seen at 28 and 56 days in cortical layers II-V that was independent of MOG immunisation. RNA analysis at 28 dpi showed an increase in expression of necroptotic pathway genes, including RIP3, MLKL, cIAP2 and Nox2. PhosphoRIP3+ and phosphoMLKL+ neurons were present in TNF + IFNγ vector injected animals, indicating activation of necroptosis. Our results suggest that persistent expression of TNF in the presence of IFNγ is a potent inducer of meningeal inflammation and can activate TNF signalling pathways in cortical cells leading to neuronal death and subpial demyelination and thus may contribute to clinical progression in MS.

Extensive subpial cortical demyelination is specific to multiple sclerosis.

Cortical demyelinated lesions are frequent and widespread in chronic multiple sclerosis (MS) patients, and may contribute to disease progression. Inflammation and related oxidative stress have been proposed as central mediators of cortical damage, yet meningeal and cortical inflammation is not specific to MS, but also occurs in other diseases. The first aim of this study was to test whether cortical demyelination was specific for demyelinating CNS diseases compared to other CNS disorders with prominent meningeal and cortical inflammation. The second aim was to assess whether oxidative tissue damage was associated with the extent of neuroaxonal damage. We studied a large cohort of patients diagnosed with demyelinating CNS diseases and non‐demyelinating diseases of autoimmune, infectious, neoplastic or metabolic origin affecting the meninges and the cortex. Included were patients with MS, acute disseminated encephalomyelitis (ADEM), neuromyelitis optica (NMO), viral and bacterial meningoencephalitis, progressive multifocal leukoencephalopathy (PML), subacute sclerosing panencephalitis (SSPE), carcinomatous and lymphomatous meningitis and metabolic disorders such as extrapontine myelinolysis, thus encompassing a wide range of adaptive and innate cytokine signatures. Using myelin protein immunohistochemistry, we found cortical demyelination in MS, ADEM, PML and extrapontine myelinolysis, whereby each condition showed a disease‐specific histopathological pattern. Remarkably, extensive ribbon‐like subpial demyelination was only observed in MS, thus providing an important pathogenetic and diagnostic cue. Cortical oxidative injury was detected in both demyelinating and non‐demyelinating CNS disorders. Our data demonstrate that meningeal and cortical inflammation alone accompanied by oxidative stress are not sufficient to generate the extensive subpial cortical demyelination found in MS, but require other MS‐specific factors.

Siponimod therapy implicates Th17 cells in a preclinical model of subpial cortical injury.

Subpial demyelination is a specific hallmark of multiple sclerosis and a correlate of disease progression. Although the mechanism(s) that mediate pathogenesis in the subpial compartment remain unclear, it has been speculated that inflammation in the overlying meninges may be associated with subpial injury. Here we show that adoptive transfer of proteolipid protein-primed Th17 cells into SJL/J recipient mice induces subpial demyelination associated with microglial/macrophage activation, disruption of the glial limitans, and evidence of an oxidative stress response. This pathology was topologically associated with foci of immune cells in the meninges and occurred in the absence of measurable anti-myelin oligodendrocyte glycoprotein IgM or IgG antibodies. To test the role of brain-infiltrating leukocytes on subpial injury, we modulated sphingosine 1-phosphate (S1P) receptor1,5 activity with BAF312 (siponimod) treatment. Administration of BAF312, even after adoptively transferred T cells had entered the brain, significantly ameliorated clinical experimental autoimmune encephalomyelitis and diminished subpial pathology, concomitant with a selective reduction in the capacity of transferred T cells to make Th17 cytokines. We conclude that sustained subpial cortical injury is associated with the capacity for brain-resident T cells to produce Th17 cytokines, and this pathological process occurs in an S1P receptor1,5-dependent manner.

Substantial subpial cortical demyelination in progressive multiple sclerosis: have we underestimated the extent of cortical pathology?

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Assessment of lesions on magnetic resonance imaging in multiple sclerosis: practical guidelines.

MRI has improved the diagnostic work-up of multiple sclerosis, but inappropriate image interpretation and application of MRI diagnostic criteria contribute to misdiagnosis. Some diseases, now recognized as conditions distinct from multiple sclerosis, may satisfy the MRI criteria for multiple sclerosis (e.g. neuromyelitis optica spectrum disorders, Susac syndrome), thus making the diagnosis of multiple sclerosis more challenging, especially if biomarker testing (such as serum anti-AQP4 antibodies) is not informative. Improvements in MRI technology contribute and promise to better define the typical features of multiple sclerosis lesions (e.g. juxtacortical and periventricular location, cortical involvement). Greater understanding of some key aspects of multiple sclerosis pathobiology has allowed the identification of characteristics more specific to multiple sclerosis (e.g. central vein sign, subpial demyelination and lesional rims), which are not included in the current multiple sclerosis diagnostic criteria. In this review, we provide the clinicians and researchers with a practical guide to enhance the proper recognition of multiple sclerosis lesions, including a thorough definition and illustration of typical MRI features, as well as a discussion of red flags suggestive of alternative diagnoses. We also discuss the possible place of emerging qualitative features of lesions which may become important in the near future.

A Mouse Model of Meningeal Inflammation and Subpial Demyelination Identifies an IL-17-mediated Mechanism of Cortical Injury that is Inhibited by Siponimod Therapy (S12.002)

A Mouse Model of Meningeal Inflammation and Subpial Demyelination Identifies an IL-17-mediated Mechanism of Cortical Injury that is Inhibited by Siponimod Therapy (S12.002)

Progressive multiple sclerosis: from pathogenic mechanisms to treatment.

During the past decades, better understanding of relapsing-remitting multiple sclerosis disease mechanisms have led to the development of several disease-modifying therapies, reducing relapse rates and severity, through immune system modulation or suppression. In contrast, current therapeutic options for progressive multiple sclerosis remain comparatively disappointing and challenging. One possible explanation is a lack of understanding of pathogenic mechanisms driving progressive multiple sclerosis. Furthermore, diagnosis is usually retrospective, based on history of gradual neurological worsening with or without occasional relapses, minor remissions or plateaus. In addition, imaging methods as well as biomarkers are not well established. Magnetic resonance imaging studies in progressive multiple sclerosis show decreased blood-brain barrier permeability, probably reflecting compartmentalization of inflammation behind a relatively intact blood-brain barrier. Interestingly, a spectrum of inflammatory cell types infiltrates the leptomeninges during subpial cortical demyelination. Indeed, recent magnetic resonance imaging studies show leptomeningeal contrast enhancement in subjects with progressive multiple sclerosis, possibly representing an in vivo marker of inflammation associated to subpial demyelination. Treatments for progressive disease depend on underlying mechanisms causing central nervous system damage. Immunity sheltered behind an intact blood-brain barrier, energy failure, and membrane channel dysfunction may be key processes in progressive disease. Interfering with these mechanisms may provide neuroprotection and prevent disability progression, while potentially restoring activity and conduction along damaged axons by repairing myelin. Although most previous clinical trials in progressive multiple sclerosis have yielded disappointing results, important lessons have been learnt, improving the design of novel ones. This review discusses mechanisms involved in progressive multiple sclerosis, correlations between histopathology and magnetic resonance imaging studies, along with possible new therapeutic approaches.

Spring cleaning: time to rethink imaging research lines in MS?

Together with recently advanced MRI technological capability, new needs and updated questions are emerging in imaging research in multiple sclerosis (MS), especially with respect to the identification of novel in vivo biomarkers of MS-relevant pathological processes. Expected benefits will involve approaches to diagnosis and clinical classification. In detail, three main points of discussion are addressed in this review: (1) new imaging biomarkers (centrifugal/centripetal lesion enhancement, central vein, paramagnetic rims at the lesion edge, subpial cortical demyelination); (2) thinking about high-resolution MR from a pathological perspective (from postmortem to in vivo staging); and (3) the clinical utility of quantitative MRI. In this context, research efforts should increasingly be focused on the direct in vivo visualization of "hidden" inflammation, beyond what can be detected with conventional gadolinium-based methods, as well as remyelination and repair, since these are likely to represent critical pathological processes and potential therapeutic targets. Concluding remarks concern the limitations, challenges, and ultimately clinical role of non-conventional MRI techniques.

Delineation of cortical pathology in multiple sclerosis using multi-surface magnetization transfer ratio imaging.

The purpose of our study was to evaluate the utility of measurements of cortical surface magnetization transfer ratio (csMTR) on the inner, mid and outer cortical boundaries as clinically accessible biomarkers of cortical gray matter pathology in multiple sclerosis (MS). Twenty-five MS patients and 12 matched controls were recruited from the MS Clinic of the Montreal Neurological Institute. Anatomical and magnetization transfer ratio (MTR) images were acquired using 3 Tesla MRI at baseline and two-year time-points. MTR maps were smoothed along meshes representing the inner, mid and outer neocortical boundaries. To evaluate csMTR reductions suggestive of sub-pial demyelination in MS patients, a mixed model analysis was carried out at both the individual vertex level and in anatomically parcellated brain regions. Our results demonstrate that focal areas of csMTR reduction are most prevalent along the outer cortical surface in the superior temporal and posterior cingulate cortices, as well as in the cuneus and precentral gyrus. Additionally, age regression analysis identified that reductions of csMTR in MS patients increase with age but appear to hit a plateau in the outer caudal anterior cingulate, as well as in the precentral and postcentral cortex. After correction for the naturally occurring gradient in cortical MTR, the difference in csMTR between the inner and outer cortex in focal areas in the brains of MS patients correlated with clinical disability. Overall, our findings support multi-surface analysis of csMTR as a sensitive marker of cortical sub-pial abnormality indicative of demyelination in MS patients.

Multiple sclerosis masquerading as Alzheimer-type dementia: Clinical, radiological and pathological findings

Background and objectives: We report a comprehensive clinical, radiological, neuropsychometric and pathological evaluation of a woman with a clinical diagnosis of AD dementia (ADem), but whose autopsy demonstrated widespread demyelination, without Alzheimer disease (AD) pathology. Methods and results: Initial neuropsychometric evaluation suggested amnestic mild cognitive impairment (aMCI). Serial magnetic resonance images (MRI) images demonstrated the rate of increase in her ventricular volume was comparable to that of 46 subjects with aMCI who progressed to ADem, without accumulating white matter disease. Myelin immunohistochemistry at autopsy demonstrated extensive cortical subpial demyelination. Subpial lesions involved the upper cortical layers, and often extended through the entire width of the cortex. Conclusions: Multiple sclerosis (MS) can cause severe cortical dysfunction and mimic ADem. Cortical demyelination is not well detected by standard imaging modalities and may not be detected on autopsy without myelin immunohistochemistry.

7 Tesla magnetic resonance imaging to detect cortical pathology in multiple sclerosis.

BackgroundNeocortical lesions (NLs) are an important pathological component of multiple sclerosis (MS), but their visualization by magnetic resonance imaging (MRI) remains challenging.ObjectivesWe aimed at assessing the sensitivity of multi echo gradient echo (ME-GRE) T2 *-weighted MRI at 7.0 Tesla in depicting NLs compared to myelin and iron staining.MethodsSamples from two MS patients were imaged post mortem using a whole body 7T MRI scanner with a 24-channel receive-only array. Isotropic 200 micron resolution images with varying T2 * weighting were reconstructed from the ME-GRE data and converted into R2 * maps. Immunohistochemical staining for myelin (proteolipid protein, PLP) and diaminobenzidine-enhanced Turnbull blue staining for iron were performed.ResultsProspective and retrospective sensitivities of MRI for the detection of NLs were 48% and 67% respectively. We observed MRI maps detecting only a small portion of 20 subpial NLs extending over large cortical areas on PLP stainings. No MRI signal changes suggestive of iron accumulation in NLs were observed. Conversely, R2 * maps indicated iron loss in NLs, which was confirmed by histological quantification.ConclusionsHigh-resolution post mortem imaging using R2 * and magnitude maps permits detection of focal NLs. However, disclosing extensive subpial demyelination with MRI remains challenging.