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Optical coherence tomography angiography (OCTA) is an imaging modality that can be used for analyzing retinal vasculature. Quantitative assessment of en face OCTA images requires accurate segmentation of the capillaries. Using deep learning approaches for this task faces two major challenges. First, acquiring sufficient manual delineations for training can take hundreds of hours. Second, OCTA images suffer from numerous contrast-related artifacts that are currently inherent to the modality and vary dramatically across scanners. We propose to solve both problems by learning a disentanglement of an anatomy component and a local contrast component from paired OCTA scans. With the contrast removed from the anatomy component, a deep learning model that takes the anatomy component as input can learn to segment vessels with a limited portion of the training images being manually labeled. Our method demonstrates state-of-the-art performance for OCTA vessel segmentation.

Esclerosi múltiple; Cadena lleugera de neurofilaments sèrics Esclerosis múltiple; Cadena ligera de neurofilamentos séricos Multiple sclerosis; Serum neurofilament light chain Objective Evaluation of serum neurofilament light chain (sNfL), measured using high-throughput assays on widely accessible platforms in large, real-world MS populations, is a critical step for sNfL to be utilized in clinical practice. Methods Multiple Sclerosis Partners Advancing Technology and Health Solutions (MS PATHS) is a network of healthcare institutions in the United States and Europe collecting standardized clinical/imaging data and biospecimens during routine clinic visits. sNfL was measured in 6974 MS and 201 healthy control (HC) participants, using a high-throughput, scalable immunoassay. Results Elevated sNfL levels for age (sNfL-E) were found in 1238 MS participants (17.8%). Factors associated with sNfL-E included male sex, younger age, progressive disease subtype, diabetes mellitus, impaired renal function, and active smoking. Higher body mass index (BMI) was associated with lower odds of elevated sNfL. Active treatment with disease-modifying therapy was associated with lower odds of sNfL-E. MS participants with sNfL-E exhibited worse neurological function (patient-reported disability, walking speed, manual dexterity, and cognitive processing speed), lower brain parenchymal fraction, and higher T2 lesion volume. Longitudinal analyses revealed accelerated short-term rates of whole brain atrophy in sNfL-E participants and higher odds of new T2 lesion development, although both MS participants with or without sNfL-E exhibited faster rates of whole brain atrophy compared to HC. Findings were consistent in analyses examining age-normative sNfL Z-scores as a continuous variable. Interpretation Elevated sNfL is associated with clinical disability, inflammatory disease activity, and whole brain atrophy in MS, but interpretation needs to account for comorbidities including impaired renal function, diabetes, and smoking. Study funding was provided from the National Institutes of Health (K23NS117883 to E.S.S.; K01MH121582 to K.C.F.; U01NS111678 to P.A.C.), National Multiple Sclerosis Society (RG-1904-33834 to E.S.S.; RG-1904-33800 to P.A.C.), and Biogen.

Background: Overall Disability Response Score (ODRS) is a composite endpoint including Expanded Disability Status Scale, Timed 25-foot Walk, and 9-Hole Peg Test, designed to quantify both disability improvement and worsening in multiple sclerosis (MS). Objective: To assess the sensitivity and clinical meaningfulness of ODRS using natalizumab Phase 3 data sets (AFFIRM in relapsing-remitting MS and ASCEND in secondary progressive MS). Methods: Differences in ODRS over 96 weeks, ODRS at Week 96, and slope of ODRS change per year between natalizumab and placebo groups were analyzed. Correlation between ODRS and changes in patient-reported outcomes was also analyzed. Results: The difference (95% confidence interval (CI)) in the ODRS over 96 weeks between natalizumab and placebo groups was 0.34 (0.21–0.46) in AFFIRM ( p Conclusion: This analysis supports ODRS as a sensitive and potentially clinically meaningful disability outcome measure in MS.

A prodrome is an early set of signs or symptoms that indicate the onset of a disease before more typical symptoms develop. Prodromal stages are well recognized in some neurological and immune-mediated diseases such as Parkinson disease, schizophrenia, type 1 diabetes mellitus and rheumatoid arthritis. Emerging evidence indicates that a prodromal stage exists in multiple sclerosis (MS), raising the possibility of intervention at this stage to delay or prevent the development of classical MS. However, much remains unclear about the prodromal stage of MS and considerable research is needed to fully characterize the prodrome and develop standardized criteria to reliably identify individuals with prodromal MS who are at high risk of progressing to a diagnosis of MS. In this Roadmap, we draw on work in other diseases to propose a disease framework for MS that incorporates the prodromal stage, and set out key steps and considerations needed in future research to fully characterize the MS prodrome, identify early disease markers and develop standardized criteria that will enable reliable identification of individuals with prodromal MS, thereby facilitating trials of interventions to slow or stop progression beyond the prodrome.

Background: Mitochondrial dysfunction plays an important role in multiple sclerosis (MS) disease progression. Plasma extracellular vesicles are a potential source of novel biomarkers in MS, and some of these are derived from mitochondria and contain functional mitochondrial components. Objective: To evaluate the relationship between levels of mitochondrial complex IV and V activity in neuronally enriched extracellular vesicles (NEVs) and brain and retinal atrophy as assessed using serial magnetic resonance imaging (MRI) and optical coherence tomography (OCT). Methods: Our cohort consisted of 48 people with MS. NEVs were immunocaptured from plasma and mitochondrial complex IV and V activity levels were measured. Subjects underwent OCT every 6 months and brain MRI annually. The associations between baseline mitochondrial complex IV and V activities and brain substructure and retinal thickness changes were estimated utilizing linear mixed-effects models. Results: We found that higher mitochondrial complex IV activity and lower mitochondrial complex V activity levels were significantly associated with faster whole-brain volume atrophy. Similar results were found with other brain substructures and retinal layer atrophy. Conclusion: Our results suggest that mitochondrial measures in circulating NEVs could serve as potential biomarkers of disease progression and provide the rationale for larger follow-up longitudinal studies.

Multiple sclerosis is associated with lesions not just in the white matter, but also involving the cortex. Cortical involvement has been linked to greater disease severity and hence understanding the factor underlying cortical pathology could help identify new therapeutic strategies for multiple sclerosis. The critical role of B cells in multiple sclerosis has been clarified by multiple pivotal trials of B-cell depletion in people with multiple sclerosis. The presence of B-cell rich areas of meningeal inflammation in multiple sclerosis has been identified at all stages of multiple sclerosis. Leptomeningeal inflammation is associated with greater extent of cortical demyelination and neuronal loss and with greater disease severity. Recent studies have identified several potential mechanisms by which B cells may mediate cortical injury including antibody production, extracellular vesicles containing neurotoxic substances and production of pro-inflammatory cytokines. Additionally, B cells may indirectly mediate cortical damage through effects on T cells, macrophages or microglia. Several animal models replicate the meningeal inflammation and cortical injury noted in people with multiple sclerosis. Studies in these models have identified Bruton’s tyrosine kinase inhibition and type II anti-CD20 antibodies as potential agents that can impact meningeal inflammation. Trials of anti-CD20 monoclonal antibodies in people with multiple sclerosis have unsuccessfully attempted to eliminate B cells in the leptomeninges. New strategies to target B cells in multiple sclerosis include Bruton’s tyrosine kinase inhibition and cell-based therapies aimed at B cells infected with Epstein–Barr virus. Future studies will clarify the mechanisms by which B cells mediate cortical injury and treatment strategies that can target B cells in the leptomeninges and CNS parenchyma.

Multiple sclerosis (MS) is a progressive inflammatory-demyelinating disease of the central nervous system. Increasing evidence suggests that vulnerable neurons in MS exhibit fatal metabolic exhaustion over time, a phenomenon hypothesized to be caused by chronic hyperexcitability. Axonal Kv7 (outward rectifying) and oligodendroglial Kir4.1 (inward rectifying) potassium channels have important roles in regulating neuronal excitability at and around nodes of Ranvier. Here, we studied the spatial and functional relationship between neuronal Kv7 and oligodendroglial Kir4.1 channels and assessed the transcriptional and functional signatures of cortical and retinal projection neurons under physiological and inflammatory-demyelinating conditions. We found that both channels became dysregulated in MS and experimental autoimmune encephalomyelitis (EAE) with Kir4.1 channels being chronically downregulated and Kv7 channel subunits being transiently upregulated during inflammatory demyelination. Further, we observed that pharmacological Kv7 channel opening with retigabine reduced neuronal hyperexcitability in human and EAE neurons, improved clinical EAE signs and rescued neuronal pathology in oligodendrocyte-Kir4.1-deficient mice. In summary, our findings indicate that neuron-oligodendrocyte compensatory interactions promote resilience through Kv7 and Kir4.1 channels and suggest pharmacological activation of nodal Kv7 channels as a neuroprotective strategy against inflammatory demyelination.

Retinal layer thickness is an important bio-marker for people with multiple sclerosis (PwMS). In clinical practice, retinal layer thickness changes in optical coherence tomography (OCT) are widely used for monitoring multiple sclerosis (MS) progression. Recent developments in automated retinal layer segmentation algorithms allow cohort-level retina thinning to be observed in a large study of PwMS. However, variability in these results make it difficult to identify patient-level trends; this prevents patient specific disease monitoring and treatment planning using OCT. Deep learning based retinal layer segmentation algorithms have achieved state-of-the-art accuracy, but the segmentation is performed on each individual scan without utilizing longitudinal information, which can be important in reducing segmentation error and reveal subtle changes in retinal layers. In this paper, we propose a longitudinal OCT segmentation network which achieves more accurate and consistent layer thickness measurements for PwMS.

Brain and flank tumors are of equivalent mass. Brain and flank tumor mass was not different on day 17 when tissues were harvested for analysis. Experiment conducted x 2 with {greater than or equal to} 5 mice/group.

TGF-β blockade does not add to RT + vaccination. TGF-β blockade with 1D11 (A) or LY2157299 (B) does not extend survival when combined with RT + rLM-OVA in mice with B16-OVA brain tumors. (N= 10 animals / group).

B16-OVA brain tumors stimulate secretion of TGF-β1 from microglia. Cytokine concentrations in the supernatants from OT-1 cells co-cultured with (A) CD11c+ APCs isolated from the spleen at a ratio of 1:5, (B) APC from brain tumor draining lymph nodes at a ratio of 1:1, (C) and CD11b+/CD45-mid microglia isolated from the brain at a ratio of 1:5.

PD-1 blockade does not add to RT + Vaccination. Adding PD-1 blockade to RT + rLM-OVA does not significantly improve survival in mice with B16-OVA brain tumors. (N = 10 animals / group)

Tumor Morphology and Volume Upon Death. A) Morphology of tumors calculated using ex vivo MRI as described. B) Calculated brain tumor volumes at the time of death as determined by ex vivo MRI (N = 3 mice/group).

Brain tumors are more tolerogenic than flank or lung tumors. (A) Representative FACS plots from tumor draining lymph nodes of mice with B16-OVA brain, flank, or lung tumors. (B) Summary graphs of the percentage of daughter cells producing IFN-γ recovered from the tumor draining lymph nodes of mice with B16-OVA brain, flank, or lung tumors (5 mice/group).

Purpose: Immune responses to antigens originating in the central nervous system (CNS) are generally attenuated, as collateral damage can have devastating consequences. The significance of this finding for the efficacy of tumor-targeted immunotherapies is largely unknown.Experimental Design: The B16 murine melanoma model was used to compare cytotoxic responses against established tumors in the CNS and in the periphery. Cytokine analysis of tissues from brain tumor–bearing mice detected elevated TGFβ secretion from microglia and in the serum and TGFβ signaling blockade reversed tolerance of tumor antigen-directed CD8 T cells. In addition, a treatment regimen using focal radiation therapy and recombinant Listeria monocytogenes was evaluated for immunologic activity and efficacy in this model.Results: CNS melanomas were more tolerogenic than equivalently progressed tumors outside the CNS as antigen-specific CD8 T cells were deleted and exhibited impaired cytotoxicity. Tumor-bearing mice had elevated serum levels of TGFβ; however, blocking TGFβ signaling with a small-molecule inhibitor or a monoclonal antibody did not improve survival. Conversely, tumor antigen–specific vaccination in combination with focal radiation therapy reversed tolerance and improved survival. This treatment regimen was associated with increased polyfunctionality of CD8 T cells, elevated T effector to T regulatory cell ratios, and decreased TGFβ secretion from microglia.Conclusions: These data suggest that CNS tumors may impair systemic antitumor immunity and consequently accelerate cancer progression locally as well as outside the CNS, whereas antitumor immunity may be restored by combining vaccination with radiation therapy. These findings are hypothesis-generating and warrant further study in contemporary melanoma models as well as human trials. Clin Cancer Res; 22(5); 1161–72. ©2015 AACR.

BACKGROUND A limited number of studies have investigated the presence of ongoing disease activity independent of clinical relapses in neuromyelitis optica spectrum disorder (NMOSD), and data are conflicting. The objective of our study was to examine whether patients with aquaporin-4 (AQP4)-IgG seropositive NMOSD exhibit progressive retinal neuroaxonal loss, independently of optic neuritis (ON) attacks. METHODS In this single-center, longitudinal study, 32 AQP4-IgG+ NMOSD patients and 48 healthy controls (HC) were followed with serial spectral-domain optical coherence tomography and visual acuity (VA) assessments. NMOSD patients with ON less than 6 months before baseline were excluded, whereas data from patients with ON during follow-up were censored at the last visit before ON. VA worsening was defined as a decrease in monocular letter acuity ≥5 letters for high-contrast VA and ≥7 letters for low-contrast VA. Analyses were performed with mixed-effects linear regression models adjusted for age, sex, and race. RESULTS The median follow-up duration was 4.2 years (interquartile range: 1.8-7.5). Relative to HC, NMOSD eyes had faster peripapillary retinal nerve fiber layer (pRNFL) (β = -0.25 µm/year faster, 95% confidence interval [CI]: -0.45 to -0.05, P = 0.014) and GCIPL thinning (β = -0.09 µm/year faster, 95% CI: -0.17 to 0, P = 0.05). This difference seemed to be driven by faster pRNFL and GCIPL thinning in NMOSD eyes without a history of ON compared with HC (GCIPL: β = -0.15 µm/year faster; P = 0.005; pRNFL: β = -0.43 µm/year faster, P < 0.001), whereas rates of pRNFL (β: -0.07 µm/year, P = 0.53) and GCIPL (β = -0.01 µm/year, P = 0.90) thinning did not differ between NMOSD-ON and HC eyes. Nine NMOSD eyes had VA worsening during follow-up. CONCLUSIONS In this longitudinal study, we observed progressive pRNFL and GCIPL atrophy in AQP4-IgG+ NMOSD eyes unaffected by ON. These results support that subclinical involvement of the anterior visual pathway may occur in AQP4-IgG+ NMOSD.

Background Chronic active lesions (CAL) in multiple sclerosis (MS) have been observed even in patients taking high-efficacy disease-modifying therapy, including B-cell depletion. Given that CAL are a major determinant of clinical progression, including progression independent of relapse activity (PIRA), understanding the predicted activity and real-world effects of targeting specific lymphocyte populations is critical for designing next-generation treatments to mitigate chronic inflammation in MS. Methods We analyzed published lymphocyte single-cell transcriptomes from MS lesions and bioinformatically predicted the effects of depleting lymphocyte subpopulations (including CD20 B-cells) from CAL via gene-regulatory-network machine-learning analysis. Motivated by the results, we performed in vivo MRI assessment of PRL changes in 72 adults with MS, 46 treated with anti-CD20 antibodies and 26 untreated, over ~2 years. Findings Although only 4.3% of lymphocytes in CAL were CD20 B-cells, their depletion is predicted to affect microglial genes involved in iron/heme metabolism, hypoxia, and antigen presentation. In vivo, tracking 202 PRL (150 treated) and 175 non-PRL (124 treated), none of the treated paramagnetic rims disappeared at follow-up, nor was there a treatment effect on PRL for lesion volume, magnetic susceptibility, or T1 time. PIRA occurred in 20% of treated patients, more frequently in those with ≥4 PRL (p=0.027). Interpretation Despite predicted effects on microglia-mediated inflammatory networks in CAL and iron metabolism, anti-CD20 therapies do not fully resolve PRL after 2-year MRI follow up. Limited tissue turnover of Bcells, inefficient passage of anti-CD20 antibodies across the blood-brain-barrier, and a paucity of Bcells in CAL could explain our findings.

This scientific commentary refers to ‘Diverse injury responses of human oligodendrocytes to mediators implicated in multiple sclerosis’ by Pernin et al. (https://doi.org/10.1093/brain/awac075).

Objective To assess the independent contributions of clinical measures (relapses, Expanded Disability Status Scale [EDSS] scores, and neuroperformance measures) and nonclinical measures (new brain magnetic resonance imaging [MRI] activity and serum neurofilament light chain [sNfL] levels) for distinguishing natalizumab‐treated from placebo‐treated patients. Methods We conducted post hoc analyses using data from the AFFIRM trial of natalizumab for multiple sclerosis. We used multivariable regression analyses with predictors (EDSS progression, no relapse, new or enlarging MRI activity, brain atrophy, sNfL levels, and neuroperformance worsening) to identify measures that independently discriminated between treatment groups. Results The multivariable model that best distinguished natalizumab from placebo was no new or enlarging T2 or gadolinium‐enhancing activity on MRI (odds ratio; 95% confidence interval: 7.2; 4.7–10.9), year 2 sNfL levels

Vitamin D is a steroid hormone that has been widely studied as a potential therapy for multiple sclerosis and other inflammatory disorders. Pre-clinical studies have implicated vitamin D in the transcription of thousands of genes, but its influence may vary by cell type. A handful of clinical studies have failed to identify an in vivo gene expression signature when using bulk analysis of all peripheral immune cells. We hypothesized that vitamin D’s gene signature would vary by immune cell type, requiring the analysis of distinct cell types. Multiple sclerosis patients (n = 18) were given high-dose vitamin D (10,400 IU/day) for six months as part of a prospective clinical trial (NCT01024777). We collected peripheral blood mononuclear cells from participants at baseline and again after six months of treatment. We used flow cytometry to isolate three immune cell types (CD4+ T-cells, CD19+ B-cells, CD14+ monocytes) for RNA microarray analysis and compared the expression profiles between baseline and six months. We identified distinct sets of differentially expressed genes and enriched pathways between baseline and six months for each cell type. Vitamin D’s in vivo gene expression profile in the immune system likely differs by cell type. Future clinical studies should consider techniques that allow for a similar cell-type resolution.

Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system (CNS) characterized by varying degrees of secondary neurodegeneration. Retinal ganglion cells (RGC) are lost in MS in association with optic neuritis but the mechanisms of neuronal injury remain unclear. Complement component C3 has been implicated in retinal and cerebral synaptic pathology that may precede neurodegeneration. Herein, we examined post-mortem MS retinas, and then used a mouse model, experimental autoimmune encephalomyelitis (EAE), to examine the role of C3 in the pathogenesis of RGC loss associated with optic neuritis. First, we show extensive C3 expression in astrocytes (C3(+)/GFAP(+) cells) and significant RGC loss (RBPMS(+) cells) in post-mortem retinas from people with MS compared to retinas from non-MS individuals. A patient with progressive MS with a remote history of optic neuritis showed marked reactive astrogliosis with C3 expression in the inner retina extending into deeper layers in the affected eye more than the unaffected eye. To study whether C3 mediates retinal degeneration, we utilized global C3(−/−) EAE mice and found that they had less RGC loss and partially preserved neurites in the retina compared with C3(+/+) EAE mice. C3(−/−) EAE mice had fewer axonal swellings in the optic nerve, reflecting reduced axonal injury, but had no changes in demyelination or T cell infiltration into the CNS. Using a C3-tdTomato reporter mouse line, we show definitive evidence of C3 expression in astrocytes in the retina and optic nerves of EAE mice. Conditional deletion of C3 in astrocytes showed RGC protection replicating the effects seen in the global knockouts. These data implicate astrocyte C3 expression as a critical mediator of retinal neuronal pathology in EAE and MS, and are consistent with recent studies showing C3 gene variants are associated with faster rates of retinal neurodegeneration in human disease.

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS), characterized by demyelination, gliosis, and neurodegeneration. While the currently available disease-modifying therapies effectively suppress the immune attack on the CNS, there are no therapies to date that directly mitigate neurodegeneration. Glucagon-like peptide-1 (GLP-1) is a small peptide hormone that maintains glucose homeostasis. A novel GLP-1 receptor (GLP-1R) agonist, NLY01, was recently shown to have neuroprotective effects in the animal models of Parkinson’s disease and is now in a phase 2 clinical trial. In this study, we investigated the therapeutic potential of NLY01 in a mouse model of MS, experimental autoimmune encephalomyelitis (EAE). Our data show that NLY01 delays the onset and attenuates the severity of EAE in a prevention paradigm, when given before disease onset. NLY01 inhibits the activation of immune cells in the spleen and reduces their trafficking into the CNS. In addition, we show that NLY01 suppresses the production of chemokines that are involved in leukocyte recruitment to the site of inflammation. The anti-inflammatory effect of NLY01 at the early stage of EAE may block the expression of the genes associated with neurotoxic astrocytes in the optic nerves, thereby preventing retinal ganglion cell (RGC) loss in the progressive stage of EAE. In the therapeutic paradigm, NLY01 significantly decreases the clinical score and second attack in a model of relapsing–remitting EAE. GLP-1R agonists may have dual efficacy in MS by suppressing peripheral and CNS inflammation, thereby limiting neuronal loss. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13311-021-01088-5.

Oligodendrocytes and their progenitors upregulate MHC pathways in response to inflammation, but the frequency of this phenotypic change is unknown and the features of these immune oligodendroglia are poorly defined. We generated MHC class I and II transgenic reporter mice to define their dynamics in response to inflammatory demyelination, providing a means to monitor MHC activation in diverse cell types in living mice and define their roles in aging, injury and disease.

Background and Objectives:Rituximab is used widely for relapse prevention in neuromyelitis optica spectrum disorder (NMOSD) and myelin oligodendrocyte glycoprotein (MOG)-IgG associated disease (MOGAD), however data regarding the effectiveness and safety of long-term rituximab use in these conditions are limited. In this study we sought to evaluate long-term clinical outcomes in patients with aquaporin-4 IgG seropositive (AQP4-IgG+) NMOSD and MOGAD treated with rituximab.Methods:We performed a retrospective chart review of patients with AQP4-IgG+ NMOSD or MOGAD followed at the Johns Hopkins Neuromyelitis Optica Clinic and included patients who had received at least one dose of rituximab.Results:We identified 111 NMOSD and 23 MOGAD patients who fulfilled inclusion criteria. Median duration of rituximab treatment for the NMOSD patients was 3.7 years (range:0.5 to 13.2) and for the MOGAD patients was 2.1 years (range:0.5 to 7.0). The annualized relapse rate (ARR) decreased after rituximab initiation in both NMOSD (median ARR:pre-treatment 1.1, post-treatment 0; pConclusions:Rituximab treatment is associated with reduced annualized relapse rate in AQP4-IgG seropositive NMOSD, especially in the absence of gaps in treatment and/or B cell reconstitution. In MOGAD, while a reduction in relapses was observed after initiation of rituximab, this association appeared to be less robust than in AQP4-IgG seropositive NMOSD. Severe infections and hypogammaglobulinemia occurred in a significant proportion of patients, highlighting the need for close monitoring of infectious complications.Classification of Evidence:This study provides Class IV evidence that rituximab decreases the annualized relapse rate in AQP4-IgG seropositive NMOSD and MOGAD.

Anti-NMDA receptor (NMDAR) encephalitis patients have been reported to exhibit visual dysfunction without retinal thinning. The objective of our study was to examine the involvement of the visual pathway structure and function in anti-NMDAR encephalitis by assessing postrecovery visual function and retinal structure, and acute-phase occipital cortex function.In this cross-sectional study, patients diagnosed with anti-NMDAR encephalitis per consensus criteria underwent postrecovery visual acuity (VA) testing and optical coherence tomography (OCT) with automated retinal layer segmentation. Clinical data and acute-phase brain 18F-fluorodeoxyglucose (FDG) PET/CT (performed within 90 days of symptom onset, assessed qualitatively and semi-quantitatively) were retrospectively analyzed. VA and OCT measures were compared between anti-NMDAR and age, sex, and race-matched healthy controls (HC). When available, FDG-PET/CT metabolism patterns were analyzed for correlations with VA, and OCT measures.A total of 16 anti-NMDAR (32 eyes) and 32 HC (64 eyes) were included in the study. Anti-NMDAR exhibited lower low-contrast VA (2.5% contrast: -4.4 letters [95% CI; -8.5 to -0.3]; P = 0.04, 1.25% contrast: -6.8 letters [95%CI; -12 to -1.7]; P = 0.01) compared with HC, but no differences were found on OCT-derived retinal layer thicknesses. Acute-phase FDG-PET/CT medial occipital cortex metabolism did not correlate with follow-up low-contrast VA or ganglion cell/inner plexiform layer thickness (GCIPL) (n = 7, 2.5% contrast: r = -0.31; P = 0.50, 1.25% contrast: r = -0.34; P = 0.45, GCIPL: r = -0.04; P = 0.94).Although the visual system seems to be involved in anti-NMDAR encephalitis, no retinal structural or occipital cortex functional abnormalities seem to be responsible for the visual dysfunction. When detected acutely, occipital lobe hypometabolism in anti-NMDAR encephalitis does not seem to associate with subsequent retrograde trans-synaptic degenerative phenomena, potentially reflecting reversible neuronal/synaptic dysfunction in the acute phase of the illness rather than neuronal degeneration.

Background and ObjectivesSerum neurofilament light chain (sNfL) and optical coherence tomography (OCT)–derived retinal measures (including peripapillary retinal nerve fiber layer [pRNFL] and macular ganglion cell layer/inner plexiform layer [GCIPL] thickness) have been proposed as biomarkers of neurodegeneration in multiple sclerosis (MS). However, studies evaluating the associations between sNfL and OCT-derived retinal measures in MS are limited.MethodsIn this retrospective analysis of a longitudinal, observational, single-center cohort study, sNfL levels were measured in people with MS and healthy controls (HCs) using single molecule array. Participants with MS were followed with serial OCT for a median follow-up of 4.5 years. Eyes with optic neuritis (ON) within 6 months of baseline OCT or ON during follow-up were excluded. Age-normative cutoffs of sNfL were derived using the HC data, and MS participants with sNfL greater than the 97.5th percentile for age were classified as having elevated sNfL (sNfL-E). Analyses were performed with mixed-effects linear regression models and adjusted for age, sex, race, and history of ON.ResultsA total of 130 HCs (age: 42.4 ± 14.2 years; 62% female) and 403 people with MS (age: 43.1 ± 12.0 years; 78% female) were included. Elevated sNfL levels were present at baseline in 80 participants with MS (19.9%). At baseline, sNfL-E participants had modestly lower pRNFL (−3.03 ± 1.50 μm; p = 0.044) and GCIPL thickness (−2.74 ± 1.02 μm; p = 0.007). As compared with those with sNfL within the reference range, eyes from NfL-E participants exhibited faster longitudinal thinning of the pRNFL (45% faster; −0.74 vs −0.51 μm/y; p = 0.015) and GCIPL (25% faster; −0.35 vs −0.28 μm/y; p = 0.021). Significant differences in rates of pRNFL and GCIPL thinning between sNfL groups were found only in those with relapsing-remitting MS but not progressive MS.DiscussionElevated baseline sNfL is associated with accelerated rates of retinal neuroaxonal loss in relapsing-remitting MS, independent of overt ON, but may be less reflective of retinal neurodegeneration in progressive MS.

Background and ObjectivesObservational studies suggest low levels of 25-hydroxyvitamin D (25[OH]D) may be associated with increased disease activity in people with multiple sclerosis (PwMS). Large-scale genome-wide association studies (GWAS) suggest 25(OH)D levels are partly genetically determined. The resultant polygenic scores (PGSs) could serve as a proxy for 25(OH)D levels, minimizing potential confounding and reverse causation in analyses with outcomes. Herein, we assess the association of genetically determined 25(OH)D and disease outcomes in MS.MethodsWe generated 25(OH)D PGS for 1,924 PwMS with available genotyping data pooled from 3 studies: the CombiRx trial (n = 575), Johns Hopkins MS Center (n = 1,152), and Immune-Mediated Inflammatory Diseases study (n = 197). 25(OH)D-PGS were derived using summary statistics (p < 5 × 10−8) from a large GWAS including 485,762 individuals with circulating 25(OH)D levels measured. We included clinical and imaging outcomes: Expanded disability status scale (EDSS), timed 25-foot walk (T25FW), nine-hole peg test (9HPT), radiologic activity, and optical coherence tomography-derived ganglion cell inner plexiform layer (GCIPL) thickness. A subset (n = 935) had measured circulating 25(OH)D levels. We fitted multivariable models based on the outcome of interest and pooled results across studies using random effects meta-analysis. Sensitivity analyses included a modifiedpvalue threshold for inclusion in the PGS (5 × 10−5) and applying Mendelian randomization (MR) rather than using PGS.ResultsInitial analyses demonstrated a positive association between generated 25(OH)D-PGS and circulating 25(OH)D levels (per 1SD increase in 25[OH]D PGS: 3.08%, 95% CI: 1.77%, 4.42%;p= 4.33e-06; R2= 2.24%). In analyses with outcomes, we did not observe an association between 25(OH)D-PGS and relapse rate (per 1SD increase in 25[OH]D-PGS: 0.98; 95% CI: 0.87–1.10), EDSS worsening (per 1SD: 1.05; 95% CI: 0.87–1.28), change in T25FW (per 1SD: 0.07%; 95% CI: −0.34 to 0.49), or change in 9HPT (per 1SD: 0.09%; 95% CI: −0.15 to 0.33). 25(OH)D-PGS was not associated with new lesion accrual, lesion volume or other imaging-based outcomes (whole brain, gray, white matter volume loss or GCIPL thinning). The results were similarly null in analyses using otherpvalue thresholds or those applying MR.DiscussionGenetically determined lower 25(OH)D levels were not associated with worse disease outcomes in PwMS and raises questions about the plausibility of a treatment effect of vitamin D in established MS.

Multiple sclerosis (MS) is a chronic inflammatory, demyelinating, and neurodegenerative disease of the central nervous system (CNS). In people with MS, impaired remyelination and axonal loss lead to debilitating long-term neurologic deficits. Current MS disease-modifying drugs mainly target peripheral immune cells and have demonstrated little efficacy for neuroprotection or promoting repair. To elucidate the pathological mechanisms and test therapeutic interventions, multiple animal models have been developed to recapitulate specific aspects of MS pathology, particularly the acute inflammatory stage. However, there are few animal models that facilitate the study of remyelination in the presence of inflammation, and none fully replicate the biology of chronic demyelination in MS. In this review, we describe the animal models that have provided insight into the mechanisms underlying demyelination, myelin repair, and potential therapeutic targets for remyelination. We highlight the limitations of studying remyelination in toxin-based demyelination models and discuss the combinatorial models that recapitulate the inflammatory microenvironment, which is now recognized to be a major inhibitor of remyelination mechanisms. These models may be useful in identifying novel therapeutics that promote CNS remyelination in inflammatory diseases such as MS.

Leptomeningeal inflammation in multiple sclerosis is associated with worse clinical outcomes and greater cortical pathology. Despite progress in identification of this process in multiple sclerosis patients using post-contrast FLAIR imaging, early trials attempting to target meningeal inflammation have been unsuccessful. There is a lack of appropriate model systems to screen potential therapeutic agents targeting meningeal inflammation. We utilized ultra-high field (11.7 Tesla) MRI to perform post-contrast FLAIR imaging in SJL/J mice with experimental autoimmune encephalomyelitis induced using immunization with proteolipid protein peptide (PLP139-151) and complete Freund’s adjuvant. Imaging was performed in both a cross-sectional and longitudinal fashion at time-points ranging from 2 to 14 weeks post-immunization. Following imaging, we euthanized animals and collected tissue for pathological evaluation, which identified dense cellular infiltrates corresponding to areas of contrast-enhancement involving the leptomeninges. These areas of meningeal inflammation contained B cells (B220+), T cells (CD3+) and myeloid cells (Mac2+). We also noted features consistent with tertiary lymphoid tissue within these areas – presence of peripheral node addressin (PNAd) positive structures, CXCL13 producing cells and FDC-M1+ follicular dendritic cells. In the cortex adjacent to areas of meningeal inflammation we identified astrocytosis, microgliosis, demyelination and evidence of axonal stress/damage. Since areas of meningeal contrast enhancement persisted over several weeks in longitudinal experiments, we utilized this model to test the effects of a therapeutic intervention on established meningeal inflammation. We randomized mice with evidence of meningeal contrast enhancement on MRI scans performed at 6 weeks post-immunization, to treatment with either vehicle or evobrutinib (a Bruton’s tyrosine kinase inhibitor) for a period of 4 weeks. These mice underwent serial imaging and we examined the effect of treatment on the areas of meningeal contrast enhancement and noted a significant reduction in the evobrutinib group compared to vehicle (30% reduction versus 5% increase; P = 0.003). We utilized ultra-high field MRI imaging to identify areas of meningeal inflammation and to track them over time in SJL/J mice with experimental autoimmune encephalomyelitis and then utilized this model to identify Bruton’s tyrosine kinase inhibition as a novel therapeutic approach to target meningeal inflammation. The results of this study provide support for future studies in multiple sclerosis patients with imaging evidence of meningeal inflammation.

Patients with myelin oligodendrocyte glycoprotein antibody (MOG-IgG)-associated disease (MOGAD) suffer from severe optic neuritis (ON) leading to retinal neuro-axonal loss, which can be quantified by optical coherence tomography (OCT). We assessed whether ON-independent retinal atrophy can be detected in MOGAD.Eighty patients with MOGAD and 139 healthy controls (HCs) were included. OCT data was acquired with (1) Spectralis spectral domain OCT (MOGAD: N = 66 and HCs: N = 103) and (2) Cirrus high-definition OCT (MOGAD: N = 14 and HCs: N = 36). Macular combined ganglion cell and inner plexiform layer (GCIPL) and peripapillary retinal nerve fiber layer (pRNFL) were quantified.At baseline, GCIPL and pRNFL were lower in MOGAD eyes with a history of ON (MOGAD-ON) compared with MOGAD eyes without a history of ON (MOGAD-NON) and HCs (p 0.001). MOGAD-NON eyes had lower GCIPL volume compared to HCs (p 0.001) in the Spectralis, but not in the Cirrus cohort. Longitudinally (follow-up up to 3 years), MOGAD-ON with ON within the last 6-12 months before baseline exhibited greater pRNFL thinning than MOGAD-ON with an ON greater than 12 months ago (p 0.001). The overall MOGAD cohort did not exhibit faster GCIPL thinning compared with the HC cohort.Our study suggests the absence of attack-independent retinal damage in patients with MOGAD. Yet, ongoing neuroaxonal damage or edema resolution seems to occur for up to 12 months after ON, which is longer than what has been reported with other ON forms. These findings support that the pathomechanisms underlying optic nerve involvement and the evolution of OCT retinal changes after ON is distinct in patients with MOGAD. ANN NEUROL 2022;92:476-485.

BackgroundPeptidylarginine deiminase 2 (PAD2) mediates the post-translational conversion of arginine residues in proteins to citrullines and is highly expressed in the central nervous system (CNS). Dysregulated PAD2 activity has been implicated in the pathogenesis of several neurologic diseases, including multiple sclerosis (MS). In this study, we sought to define the cellular and regional expression of the gene encoding for PAD2 (i.e. PADI2) in the human CNS using publicly available datasets and evaluate whether anti-PAD2 antibodies were present in patients with various neurologic diseases.MethodsA total of 491 study participants were included in this study: 91 people with MS, 32 people with neuromyelitis optica (NMO), 281 people with post-treatment Lyme disease (PTLD), and 87 healthy controls. To measure PADI2 expression in the CNS from healthy individuals, publicly available tissue and single cell RNA sequencing data was analyzed. Anti-PAD2 antibodies were measured in the serum of study participants using anti-PAD2 ELISA. Clinical and demographic variables were compared according to anti-PAD2 antibody positivity for the MS and PTLD groups and correlations between anti-PAD2 levels and disease severity were examined.ResultsPADI2 expression was highest in oligodendrocytes (mean ± SD; 6.4 ± 2.2), followed closely by astrocytes (5.5 ± 2.6), microglia/macrophages (4.5 ± 3.5), and oligodendrocyte precursor cells (3.2 ± 3.3). There was an increased proportion of anti-PAD2 positivity in the MS (19.8%; p = 0.007) and PTLD groups (13.9%; p = 0.057) relative to the healthy controls (5.7%), and these antibodies were not detected in NMO patients. There was a modest inverse correlation between anti-PAD2 levels and disease severity in people with MS (τ = −0.145, p = 0.02), with levels being the highest in those with relapsing-remitting disease. Similarly, there was a modest inverse correlation between anti-PAD2 levels and neurocognitive score (τ = −0.10, p = 0.027) in people with PTLD, with difficulty focusing, memory changes, fatigue, and difficulty finding words contributing most strongly to the effect.ConclusionPADI2 expression was observed in diverse regions and cells of the CNS, and anti-PAD2 autoantibodies were associated with less severe symptoms in subsets of patients with MS and PTLD. These data suggest that anti-PAD2 antibodies may attenuate inflammation in diseases of different etiologies, which are united by high PADI2 expression in the target tissue.

BackgroundMultiple sclerosis (MS) is a neurodegenerative disease, wherein aberrant immune cells target myelin-ensheathed nerves. Conventional magnetic resonance imaging (MRI) can be performed to monitor damage to the central nervous system that results from previous inflammation; however, these imaging biomarkers are not necessarily indicative of active, progressive stages of the disease. The immune cells responsible for MS are first activated and sensitized to myelin in lymph nodes (LNs). Here, we present a new strategy for monitoring active disease activity in MS, chemical exchange saturation transfer (CEST) MRI of LNs.Methods and resultsWe studied the potential utility of conventional (T2-weighted) and CEST MRI to monitor changes in these LNs during disease progression in an experimental autoimmune encephalomyelitis (EAE) model. We found CEST signal changes corresponded temporally with disease activity. CEST signals at the 3.2 ppm frequency during the active stage of EAE correlated significantly with the cellular (flow cytometry) and metabolic (mass spectrometry imaging) composition of the LNs, as well as immune cell infiltration into brain and spinal cord tissue. Correlating primary metabolites as identified by matrix-assisted laser desorption/ionization (MALDI) imaging included alanine, lactate, leucine, malate, and phenylalanine.ConclusionsTaken together, we demonstrate the utility of CEST MRI signal changes in superficial cervical LNs as a complementary imaging biomarker for monitoring disease activity in MS. CEST MRI biomarkers corresponded to disease activity, correlated with immune activation (surface markers, antigen-stimulated proliferation), and correlated with LN metabolite levels.

BackgroundNeurofilament light chain (NfL) is an axonal cytoskeletal protein that is released into the extracellular space following neuronal or axonal injury associated with neurological conditions such as multiple sclerosis (MS), amyotrophic lateral sclerosis (ALS), and other diseases. NfL is detectable in the cerebrospinal fluid (CSF) and blood. Numerous studies on MS have demonstrated that NfL is correlated with disease activity, predicts disease progression, and is reduced by treatment with MS disease-modifying drugs, making NfL an attractive candidate to supplement existing clinical and imaging measures in MS. However, for NfL to achieve its potential as a clinically useful biomarker for clinical decision-making or drug development, a standardized, practical, and widely accessible assay is needed. Our objective was to develop a novel NfL assay on an automated, globally available immunoassay platform and validate its performance.MethodsA prototype NfL assay was first developed and evaluated on the ADVIA Centaur® XP immunoassay system from Siemens Healthineers. The lower limit of quantitation (LLoQ), within-lab precision, assay range, cross-reactivity with neurofilament medium and heavy chains, and effect of interfering substances were determined. NfL assay values in serum and CSF were compared with radiological and clinical disease activity measures in patients with MS and ALS, respectively. This assay was further optimized to utilize serum, plasma, and CSF sample types on the Atellica® IM system and transferred to Siemens' CLIA laboratory where it was analytically validated as a laboratory-developed test (LDT).ResultsIn this study, an LLoQ of 1.85 pg/mL, within-lab precision ConclusionThe analytical performance of the NfL assay fulfilled all acceptance criteria; therefore, we suggest that the assay is acceptable for use in both research and clinical practice settings to determine elevated NfL levels in patients.