The use of the central vein sign in the diagnosis of multiple sclerosis: a systematic review and meta-analysis

Background: The central vein sign (CVS) is a radiological feature proposed as a multiple sclerosis (MS) imaging biomarker able to accurately differentiate MS from other white matter diseases of the central nervous system. In this work, we evaluated the pooled proportion of the CVS in brain MS lesions and to estimate the diagnostic performance of CVS to perform a diagnosis of MS and propose an optimal cut-off value. Methods: A systematic search was performed on publicly available databases (PUBMED/MEDLINE and Web of Science) up to 24 August 2020. Analysis of the proportion of white matter MS lesions with a central vein was performed using bivariate random-effect models. A meta-regression analysis was performed and the impact of using particular sequences (such as 3D echo-planar imaging) and post-processing techniques (such as FLAIR*) was investigated. Pooled sensibility and specificity were estimated using bivariate models and meta-regression was performed to address heterogeneity. Inclusion and publication bias were assessed using asymmetry tests and a funnel plot. A hierarchical summary receiver operating curve (HSROC) was used to estimate the summary accuracy in diagnostic performance. The Youden index was employed to estimate the optimal cut-off value using individual patient data. Results: The pooled proportion of lesions showing a CVS in the MS population was 73%. The use of the CVS showed a remarkable diagnostic performance in MS cases, providing a pooled specificity of 92% and a sensitivity of 95%. The optimal cut-off value obtained from the individual patient data pooled together was 40% with excellent accuracy calculated by the area under the ROC (0.946). The 3D-EPI sequences showed both a higher pooled proportion compared to other sequences and explained heterogeneity in the meta-regression analysis of diagnostic performances. The 1.5 Tesla (T) scanners showed a lower (58%) proportion of MS lesions with a CVS compared to both 3T (74%) and 7T (82%). Conclusions: The meta-analysis we have performed shows that the use of the CVS in differentiating MS from other mimicking diseases is encouraged; moreover, the use of dedicated sequences such as 3D-EPI and the high MRI field is beneficial

Cerebrospinal fluid IgM levels in association with inflammatory pathways in multiple sclerosis patients

Background Intrathecal immunoglobulin M (IgM) synthesis has been demonstrated in the early disease stages of multiple sclerosis (MS) as a predictor factor of a worsening disease course. Similarly, increased cerebrospinal fluid (CSF) molecules related to B-cell intrathecal activity have been associated with a more severe MS progression. However, whether CSF levels of IgM are linked to specific inflammatory and clinical profile in MS patients at the time of diagnosis remains to be elucidated. Methods Using customized Bio-Plex assay, the protein levels of IgG, IgA, IgM, and of 34 other inflammatory molecules, related to B-cell, T-cell, and monocyte/macrophage activity, were analyzed in the CSF of 103 newly diagnosed relapsing-remitting MS patients and 36 patients with other neurological disorders. CSF IgM levels were also correlated with clinical and neuroradiological measures [advanced 3-T magnetic resonance imaging (MRI) parameters], at diagnosis and after 2 years of follow-up. Results A 45.6% increase in CSF IgM levels was found in MS patients compared to controls (p = 0.013). CSF IgM levels correlated with higher CSF levels of CXCL13 (p = 0.039), CCL21 (p = 0.023), interleukin 10 (IL-10) (p = 0.025), IL-12p70 (p = 0.020), CX3CL1 (p = 0.036), and CHI3L1 (p = 0.048) and were associated with earlier age of patients at diagnosis (p = 0.008), white matter lesion (WML) number (p = 0.039) and disease activity (p = 0.033) after 2 years of follow-up. Conclusion IgMs are the immunoglobulins mostly expressed in the CSF of naive MS patients compared to other neurological conditions at the time of diagnosis. The association between increased CSF IgM levels and molecules related to both B-cell immunity (IL-10) and recruitment (CXCL13 and CCL21) and to macrophage/microglia activity (IL-12p70, CX3CL1, and CHI3L1) suggests possible correlation between humoral and innate intrathecal immunity in early disease stage. Furthermore, the association of IgM levels with WMLs and MS clinical and MRI activity after 2 years supports the idea of key role of IgM in the disease course

Intrathecal versus peripheral inflammatory protein profile in MS patients at diagnosis: a comprehensive investigation on serum and CSF

Intrathecal inflammation plays a key role in the pathogenesis of multiple sclerosis (MS). To better elucidate its relationship with peripheral inflammation, we investigated the correlation between cerebrospinal fluid (CSF) and serum levels of 61 inflammatory proteins. Paired CSF and serum samples were collected from 143 treatment-naïve MS patients at diagnosis. A customized panel of 61 inflammatory molecules was analyzed by a multiplex immunoassay. Correlations between serum and CSF expression levels for each molecule were performed by Spearman's method. The expression of sixteen CSF proteins correlated with their serum expression (p-value < 0.001): only five molecules (CXCL9, sTNFR2, IFNα2, Pentraxin-3, and TSLP) showed a Rho value >0.40, suggesting moderate CSF/serum correlation. No correlation between inflammatory serum patterns and Qalb was observed. Correlation analysis of serum expression levels of these sixteen proteins with clinical and MRI parameters pinpointed a subset of five molecules (CXCL9, sTNFR2, IFNα2, IFNβ, and TSLP) negatively correlating with spinal cord lesion volume. However, following FDR correction, only the correlation of CXCL9 remained significant. Our data support the hypothesis that the intrathecal inflammation in MS only partially associates with the peripheral one, except for the expression of some immunomodulators that might have a key role in the initial MS immune response

The prognostic value of white-matter selective double inversion recovery mri sequence in multiple sclerosis: an exploratory study

Using a white-matter selective double inversion recovery sequence (WM-DIR) that suppresses both grey matter (GM) and cerebrospinal fluid (CSF) signals, some white matter (WM) lesions appear surrounded by a dark rim. These dark rim lesions (DRLs) seem to be specific for multiple sclerosis (MS). They could be of great usefulness in clinical practice, proving to increase the MRI diagnostic criteria specificity. The aims of this study are the identification of DRLs on 1.5 T MRI, the exploration of the relationship between DRLs and disease course, the characterization of DRLs with respect to perilesional normal-appearing WM using magnetization transfer imaging, and the investigation of possible differences in the underlying tissue properties by assessing WM-DIR images obtained at 3.0 T MRI. DRLs are frequent in primary progressive MS (PPMS) patients. Amongst relapsing-remitting MS (RRMS) patients, DRLs are associated with a high risk of the disease worsening and secondary progressive MS (SPMS) conversion after 15 years. The mean magnetization transfer ratio (MTR) of DRLs is significantly different from the lesion without the dark rim, suggesting that DRLs correspond to more destructive lesions

Immune Response after COVID-19 mRNA Vaccination in Multiple Sclerosis Patients Treated with DMTs

The impact of disease-modifying therapies (DMTs) on the immune response to coronavirus disease-2019 (COVID-19) vaccines in persons with multiple sclerosis (pwMS) needs further elucidation. We investigated BNT162b2 mRNA COVID-19 vaccine effects concerning antibody seroconversion, inflammatory mediators' level and immunophenotype assessment in pwMS treated with cladribine (c-pwMS, n = 29), fingolimod (f-pwMS, n = 15) and ocrelizumab (o-pwMS, n = 54). Anti-spike immunoglobulin (Ig)-G detection was performed by an enzyme immunoassay; molecular mediators (GrB, IFN-gamma and TNF-alpha) were quantified using the ELLA platform, and immunophenotype was assessed by flow cytometry. ANCOVA, Student's t-test and Pearson correlation analyses were applied. Only one o-pwMS showed a mild COVID-19 infection despite most o-pwMS lacking seroconversion and showing lower anti-spike IgG titers than c-pwMS and f-pwMS. No significant difference in cytokine production and lymphocyte count was observed in c-pwMS and f-pwMS. In contrast, in o-pwMS, a significant increase in GrB levels was detected after vaccination. Considering non-seroconverted o-pwMS, a significant increase in GrB serum levels and CD4+ T lymphocyte count was found after vaccination, and a negative correlation was observed between anti-spike IgG production and CD4+ T cells count. Differences in inflammatory mediators' production after BNT162b2 vaccination in o-pwMS, specifically in those lacking anti-spike IgG, suggest a protective cellular immune response

CSF TNF and osteopontin levels correlate with the response to dimethyl fumarate in early multiple sclerosis

Background: Disease activity in the first years after a diagnosis of relapsing-remitting multiple sclerosis (RRMS) is a negative prognostic factor for long-term disability. Markers of both clinical and radiological responses to disease-modifying therapies (DMTs) are advocated. Objective: The objective of this study is to estimate the value of cerebrospinal fluid (CSF) inflammatory markers at the time of diagnosis in predicting the disease activity in treatment-naïve multiple sclerosis (MS) patients exposed to dimethyl fumarate (DMF). Methods: In total, 48 RRMS patients (31 females/17 males) treated with DMF after the diagnosis were included in this 2-year longitudinal study. All patients underwent a CSF examination, regular clinical and 3T magnetic resonance imaging (MRI) scans that included the assessment of white matter (WM) lesions, cortical lesions (CLs) and global cortical thickness. CSF levels of 10 pro-inflammatory markers - CXCL13 [chemokine (C-X-C motif) ligand 13 or B lymphocyte chemoattractant], CXCL12 (stromal cell-derived factor or C-X-C motif chemokine 12), tumour necrosis factor (TNF), APRIL (a proliferation-inducing ligand, or tumour necrosis factor ligand superfamily member 13), LIGHT (tumour necrosis factor ligand superfamily member 14 or tumour necrosis factor superfamily member 14), interferon (IFN) gamma, interleukin 12 (IL-12), osteopontin, sCD163 [soluble-CD163 (cluster of differentiation 163)] and Chitinase3-like1 - were assessed using immune-assay multiplex techniques. The combined three-domain status of 'no evidence of disease activity' (NEDA-3) was defined by no relapses, no disability worsening and no MRI activity, including CLs. Results: Twenty patients (42%) reached the NEDA-3 status; patients with disease activity showed higher CSF TNF (p = 0.009), osteopontin (p = 0.005), CXCL12 (p = 0.037), CXCL13 (p = 0.040) and IFN gamma levels (p = 0.019) compared with NEDA-3 patients. After applying a random forest approach, TNF and osteopontin revealed the most important variables associated with the NEDA-3 status. Six molecules that emerged at the random forest approach were added in a multivariate regression model with demographic, clinical and MRI measures of WM and grey matter damage as independent variables. TNF levels confirmed to be associated with the absence of disease activity: odds ratio (OR) = 0.25, CI% = 0.04-0.77. Conclusion: CSF inflammatory markers may provide prognostic information in predicting disease activity in the first years after DMF initiation. CSF TNF levels are a possible candidate in predicting treatment response, in addition to clinical, demographic and MRI variables

CSF parvalbumin levels reflect interneuron loss linked with cortical pathology in multiple sclerosis

Introduction and methods: In order to verify whether parvalbumin (PVALB), a protein specifically expressed by GABAergic interneurons, could be a MS-specific marker of grey matter neurodegeneration, we performed neuropathology/molecular analysis of PVALB expression in motor cortex of 40 post-mortem progressive MS cases, with/without meningeal inflammation, and 10 control cases, in combination with cerebrospinal fluid (CSF) assessment. Analysis of CSF PVALB and neurofilaments (Nf-L) levels combined with physical/cognitive/3TMRI assessment was performed in 110 na\uefve MS patients and in 32 controls at time of diagnosis. Results: PVALB gene expression was downregulated in MS (fold change = 3.7 \ub1 1.2, P < 0.001 compared to controls) reflecting the significant reduction of PVALB+ cell density in cortical lesions, to a greater extent in MS patients with high meningeal inflammation (51.8, P < 0.001). Likewise, post-mortem CSF-PVALB levels were higher in MS compared to controls (fold change = 196 \ub1 36, P < 0.001) and correlated with decreased PVALB+ cell density (r = -0.64, P < 0.001) and increased MHC-II+ microglia density (r = 0.74, P < 0.01), as well as with early age of onset (r = -0.69, P < 0.05), shorter time to wheelchair (r = -0.49, P < 0.05) and early age of death (r = -0.65, P < 0.01). Increased CSF-PVALB levels were detected in MS patients at diagnosis compared to controls (P = 0.002). Significant correlation was found between CSF-PVALB levels and cortical lesion number on MRI (R = 0.28, P = 0.006) and global cortical thickness (R = -0.46, P < 0.001), better than Nf-L levels. CSF-PVALB levels increased in MS patients with severe cognitive impairment (mean \ub1 SEM:25.2 \ub1 7.5 ng/mL) compared to both cognitively normal (10.9 \ub1 2.4, P = 0.049) and mild cognitive impaired (10.1 \ub1 2.9, P = 0.024) patients. Conclusions: CSF-PVALB levels reflect loss of cortical interneurons in MS patients with more severe disease course and might represent an early, new MS-specific biomarker of cortical neurodegeneration, atrophy, and cognitive decline

A combined MRI and histological evaluation of cortical pathology in multiple sclerosis

Multiple sclerosis (MS) is a chronic, demyelinating disease affecting the central nervous system. White matter lesions are the typical hallmark of the disease, however, cortical pathology has been demonstrated to influence MS disease progression. While neuropathological assessments evidenced areas of cortical demyelination, inflammation and neurodegeneration, in-vivo detection of cortical pathology is still challenging. Quantitative magnetic resonance imaging (qMRI), measuring biophysical properties sensitive to the composition/microstructure of the underlying tissue, has the potential to reveal subtle pathological changes. Despite the potentiality of the qMRI approach, a detailed multimodal characterization of cortical pathology is still lacking. The main objective of this thesis was to design and optimize acquisition and processing of a multimodal MRI protocol comprehending the estimations of T1 and T2* relaxation times, Magnetization Transfer Ratio (MTR) and Magnetization Transfer Saturation (MTsat) for the characterization of cortical pathology carried out on brain samples consisting of bi-hemispheric coronal slices donated from human MS donors. More in detail the thesis aimed to i) evaluate the feasibility of a B1 corrected variable flip angle approach for the estimation of T1 relaxation times in formalin-fixed ex-vivo samples at 3T field strength; ii)  evaluate the effect of cortical demyelination, histologically-validated, on each MRI modality estimated at both 3 and 7T and iii) compare the magnitude of the lesion contrast for each MRI modality obtained at the lower with respect to the higher field strength; iv) evaluate the association between each MRI modality acquired at 3 and 7T and activated microglia cell density in lesions and normal appearing tissue.  The MRI protocol was acquired on all the brain samples (8) on a 3T MRI system while a subset of the samples (5) were acquired also at 7T. Five of the samples acquired at 3T were selected to investigate the feasibility of the B1-corrected variable flip angle T1 mapping approach for ex-vivo analysis. Neuropathological assessments of myelin and activated microglia were performed on five samples (three acquired only at 3T and two acquired at both field strengths). Cortical demyelination was delineated on immunohistological stainings and registered to the MRI space.  The analysis of the T1 mapping approaches for ex-vivo investigations at 3T revealed that B1-correction improves, as compared to the uncorrected approach, the reliability of the variable flip angle (VFA) method both in terms of root mean square error and correlation with the gold standard inversion recovery approach. The analyses of histologically validated cortical demyelination revealed a significant decrease in MTsat values as well as an increase in T2* in areas of demyelination as compared to layer-matched normal appearing tissue in all the subjects when examining the global cortical voxel distributions. The same MRI modalities also reported the highest percentage of lesions with marked local contrast with respect to adjacent normal appearing tissue. MTsat also showed the strongest association with microglia activation in the cortex. In conclusion the qMRI approaches undertaken in this combined MRI-histology study demonstrated sensitivity to pathological changes in the cortex of MS patients, hardly visible with conventional MRI, that seemed to be amplified at higher field strength. MTsat have been demonstrated to be sensitive to cortical pathology and might be a candidate for the indirect assessment of demyelination and/or microglia activation in the cortex

Making Visible the Invisible: Automatically Measured Global and Regional Brain Volume Is Associated with Cognitive Impairment and Fatigue in Multiple Sclerosis

In multiple sclerosis (MS), the transition from relapsing-remitting to the secondary-progressive phase is characterized by a progression independent of relapse activity (PIRA), resulting in physical disability accumulation and invisible symptoms, i.e., fatigue and cognitive impairment (CI). These symptoms are related to neurodegenerative processes and have been correlated with MRI measures of brain atrophy only at a group level; however, the application in clinical practice of atrophy-based measurements for single-patient evaluation is yet to be fully investigated. In the present study, we aimed to evaluate the association between brain atrophy, measured with easy-to-use automatic software, and the “invisible” MS symptoms of cognition and fatigue. A total of 69 MS patients were included in the study; cognitive impairment and fatigue (FSS) (in addition to neurological disability, EDSS) were assessed and correlated with brain volumes calculated using the automated software QyScore® which is validated for single-patient use in the clinical setting. Results showed that the cognitive status was accurately reflected by measures of atrophy, with a sensitivity of up to 90%. CI patients showed a lower volume compared to cognitively normal patients in the whole brain (p = 0.017), gray matter (p = 0.042), insula (p = 0.035), cerebellum (p = 0.008), and limbic lobe (p = 0.049). FSS was associated with temporal lobe (r = −0.37, p = 0.013) and insular (r = −0.36, p = 0.019) volumes. The volumes of the same regions were also associated with EDSS. The global/regional atrophy results, assessed with automatic and easy-to-use software, correlated with cognitive and fatigue symptoms, thus supporting the clinical application in routine patient management