Remitting Multiple Sclerosis Patients Research Articles

Prediction of disease activity and treatment failure in relapsing–remitting MS patients initiating daily oral DMTs

Background: Limited data exist regarding treatment response prediction to oral disease-modifying therapies (DMTs) in multiple sclerosis (MS). Objectives: We assessed the capacity of available scoring systems to anticipate disease activity parameters in naïve relapsing–remitting MS (RRMS) patients initiating daily oral DMTs, hypothesizing that they exhibit different predictive potentials. Methods: We conducted a retrospective study and applied the Rio Score (RS), modified Rio Score (mRS), and MAGNIMS Score 12 months after DMT initiation. At 36 months, we examined their ability to predict evidence of disease activity (EDA) components and treatment failure by logistic regression analysis. Results: Notably, 218 patients (62.4% females) initiating dimethyl fumarate, teriflunomide, and fingolimod were included. At 36 months, the RS high-risk group predicted evidence of clinical activity (odds ratio (OR) 10 [2.7–36.9]) and treatment failure (OR 10.6 [3.4–32.5]) but did not predict radiological activity (OR 1.9 [0.7–5]). The mRS non-responders group did not predict EDA and treatment failure. RS, mRS, and MAGNIMS 0 categories showed significantly lower EDA and treatment failure than the remainder. Conclusion: Scoring systems present different predictive abilities for disease activity parameters at 36 months in MS patients initiating daily oral therapies, warranting further adjustments (i.e. introduction of fluid biomarkers) to depict disease activity status fully.

MicroRNAs dysregulated in multiple sclerosis affect the differentiation of CG-4 cells, an oligodendrocyte progenitor cell line.

Demyelination is one of the hallmarks of multiple sclerosis (MS). While remyelination occurs during the disease, it is incomplete from the start and strongly decreases with its progression, mainly due to the harm to oligodendrocyte progenitor cells (OPCs), causing irreversible neurological deficits and contributing to neurodegeneration. Therapeutic strategies promoting remyelination are still very preliminary and lacking within the current treatment panel for MS. In a previous study, we identified 21 microRNAs dysregulated mostly in the CSF of relapsing and/or remitting MS patients. In this study we transfected the mimics/inhibitors of several of these microRNAs separately in an OPC cell line, called CG-4. We aimed (1) to phenotypically characterize their effect on OPC differentiation and (2) to identify corroborating potential mRNA targets via immunocytochemistry, RT-qPCR analysis, RNA sequencing, and Gene Ontology enrichment analysis. We observed that the majority of 13 transfected microRNA mimics decreased the differentiation of CG-4 cells. We demonstrate, by RNA sequencing and independent RT-qPCR analyses, that miR-33-3p, miR-34c-5p, and miR-124-5p arrest OPC differentiation at a late progenitor stage and miR-145-5p at a premyelinating stage as evidenced by the downregulation of premyelinating oligodendrocyte (OL) [Tcf7l2, Cnp (except for miR-145-5p)] and mature OL (Plp1, Mbp, and Mobp) markers, whereas only miR-214-3p promotes OPC differentiation. We further propose a comprehensive exploration of their change in cell fate through Gene Ontology enrichment analysis. We finally confirm by RT-qPCR analyses the downregulation of several predicted mRNA targets for each microRNA that possibly support their effect on OPC differentiation by very distinctive mechanisms, of which some are still unexplored in OPC/OL physiology. miR-33-3p, miR-34c-5p, and miR-124-5p arrest OPC differentiation at a late progenitor stage and miR-145-5p at a premyelinating stage, whereas miR-214-3p promotes the differentiation of CG-4 cells. We propose several potential mRNA targets and hypothetical mechanisms by which each microRNA exerts its effect. We hereby open new perspectives in the research on OPC differentiation and the pathophysiology of demyelination/remyelination, and possibly even in the search for new remyelinating therapeutic strategies in the scope of MS.

Differentiating multiple sclerosis from cerebral small vessel disease using diffusion tensor imaging and magnetic resonance spectroscopy on normally appearing thalami

BackgroundDiffusion tensor imaging (DTI) and magnetic resonance spectroscopy (1H-MRS) can detect the microstructural changes in normal-appearing conventional MRI. So, they may differentiate between multiple sclerosis (MS) cases and cerebral small vessel disease (CSVD). This work aimed to investigate if MRS and DTI are helpful in differentiating between MS and CSVD cases. MethodsThe study was conducted on 90 subjects divided into three groups: 30 relapsing–remitting MS patients, 30 patients with MRI showing CSVD, and 30 healthy controls. Diffusion tensor imaging measuring thalamic FA, ADC values, and 1H-MRS were conducted on patients and controls. ResultsThalamic FA values were significantly higher in the RRMS group than in the control and CSVD groups (P < 0.001, for each) but significantly lower in the CSVD group than the control group (P < 0.001). Moreover, thalamic ADC values were significantly higher in the CSVD group than in the control and MS groups (P < 0.001, for each). Also, thalamic NAA values were significantly lower in RRMS and CSVD groups than in controls (P < 0.001 for each). Still, they were significantly lower in the RRMS group than the CSVD group only on the left side (P = 0.004). The thalamic NAA/Cr values were significantly lower in RRMS (P < 0.001 for both sides) and CVSD than in controls (P = 0.044 and 0.036, for RT and LT sides, respectively). ConclusionsThalamic DTI and 1H-MRS can help detect the microstructural changes in normal-appearing thalami in RRMS and CSVD patients. Moreover, they can help differentiate MS from CSVD patients.

Multiple sclerosis susceptibility may be associated with the coding rs20541 (R130Q) IL-13 gene polymorphism in the Polish population

Some of the multiple autoimmune diseases have been already associated with IL-13 single-nucleotide polymorphisms (SNPs). However, there are only few studies regarding multiple sclerosis (MS) risk and IL-13 rs20541 (R130Q) polymorphism, and their results are conflicting. Therefore, the aim of our study was to investigate the frequency of the IL-13 gene rs20541 (R130Q) polymorphism in MS participants and its association with MS clinical subsets in the Polish population. We conducted a case‒control study including 94 relapsing remitting MS patients and 160 healthy volunteers. We genotyped the rs20541 polymorphism in the IL-13 gene and analysed the genotype frequency, age of MS onset and clinical condition (EDSS values) of the MS participants. Fisher’s exact test was used for statistical analysis, and the log-linear model was applied to test for associations. Allele A, as well as the AA and AG genotypes, was observed to be significantly more common in the MS subjects. The OR (odds ratio) for the A compared to the G allele was 1.71 (1.14–2.56), whereas OR 2.33 (0.86–6.26) and OR 1.92 (1.11–3.30) were obtained for the AA and AG genotypes, respectively. We did not identify any significant associations of the studied IL-13 SNP with the investigated clinical parameters of the MS participants. Our results suggest that the rs20541 polymorphism in the IL-13 gene may play an important role in MS predisposition but not in investigated clinical parameters in MS subjects of the Polish population.

Correlation between antibodies against the pathogenic pHERV-W envelope protein and the inflammatory phase of multiple sclerosis.

The role of retroviral envelope proteins belonging to the Human Endogenous Retroviral family 'W' (HERV-W), specifically syncytin-1 and pathogenic HERV-W (pHERV-W), as potential risk factors in multiple sclerosis (MS) has been established. This study aimed to investigate the humoral response to syncytin-1 and pHERV-W-derived peptides in a group of relapsing remitting MS patients categorized as having acute or stable disease. Furthermore, an inhibition assay was conducted to assess the extent of cross-reactivity between the two epitopes. The findings revealed that MS patients in the acute phase exhibited a higher specific antibody response to the pHERV-W env epitope compared to syncytin-1. This suggests a potential pathogenic role for pHERV-W env during the inflammatory stages of central nervous system involvement, and these antibody responses could serve as useful biomarkers for monitoring the progression of the disease.

Glutamate as a new path in discrimination between neuromyelitis optica spectrum disorder and multiple sclerosis

BackgroundNeuromyelitis optica spectrum disorder (NMOSD) used to be considered as a variant of multiple sclerosis (MS), however the recent detection of a highly specific serum biomarkers for NMOSD have made clear that NMOSD is a condition distinct from MS. The aim was to explore the role of serum glutamate level in the discrimination between NMOSD and relapsing remitting (RR) MS patients during and in between relapses. The study comprised two groups; first group, a total of 30 NMOSD patients, they were furtherly subdivided into NMOSD in remission, 15 patients without recent relapses in the last 3 months, NMOSD with relapse, 15 patients with recent relapses in the last 3 months, the second group, 30 definite, RRMS patients, they were further subdivided into RRMS in remission, 15 patients without recent relapses in the last 3 months RRMS with relapse, 15 patients with recent relapses in the last 3 months.ResultsWithout relapse, NMOSD patients have higher level of serum glutamate than RRMS patients with (P values = 0.005), a significant difference between EDSS in NMOSD patients and RRMS patients (P = 0.0001), The cut-off value of glutamate serum level between NMOSD in remission and RRMS in remission was > 10.3 μg/mL, yet its level for differentiation between group RRMS in remission and RRMS with relapse was > 12.6 μg/mL.ConclusionGlutamate cut-off value might be a reliable tool to discriminate between NMOSD and RRMS.

Altered neural intrinsic oscillations in patients with multiple sclerosis: effects of cortical thickness.

To investigate the effects of cortical thickness on the identification accuracy of fractional amplitude of low-frequency fluctuation (fALFF) in patients with multiple sclerosis (MS). Resting-state functional magnetic resonance imaging data were collected from 31 remitting MS, 20 acute MS, and 42 healthy controls (HCs). After preprocessing, we first calculated two-dimensional fALFF (2d-fALFF) maps using the DPABISurf toolkit, and 2d-fALFF per unit thickness was obtained by dividing 2d-fALFF by cortical thickness. Then, between-group comparison, clinical correlation, and classification analyses were performed in 2d-fALFF and 2d-fALFF per unit thickness maps. Finally, we also examined whether the effect of cortical thickness on 2d-fALFF maps was affected by the subfrequency band. In contrast with 2d-fALFF, more changed regions in 2d-fALFF per unit thickness maps were detected in MS patients, such as increased region of the right inferior frontal cortex and faded regions of the right paracentral lobule, middle cingulate cortex, and right medial temporal cortex. There was a significant positive correlation between the disease duration and the 2d-fALFF values in the left early visual cortex in remitting MS patients (r = 0.517, Bonferroni-corrected, p = 0.008 × 4 < 0.05). In contrast with 2d-fALFF, we detected a positive correlation between the 2d-fALFF per unit thickness of the right ventral stream visual cortex and the modified Fatigue Impact Scale (MFIS) scores (r = 0.555, Bonferroni-corrected, p = 0.017 × 4 > 0.05). For detecting MS patients, 2d-fALFF and 2d- fALFF per unit thickness both performed remarkably well in support vector machine (SVM) analysis, especially in the remitting phase (AUC = 86, 83%). Compared with 2d-fALFF, the SVM model of 2d-fALFF per unit thickness had significantly higher classification performance in distinguishing between remitting and acute MS. More changed regions and more clinically relevant 2d-fALFF per unit thickness maps in the subfrequency band were also detected in MS patients. By dividing the functional value by the cortical thickness, the identification accuracy of fALFF in MS patients was detected to be potentially influenced by cortical thickness. Additionally, 2d-fALFF per unit thickness is a potential diagnostic marker that can be utilized to distinguish between acute and remitting MS patients. Notably, we observed similar variations in the subfrequency band.

A prospective observational longitudinal study with a two-year follow-up of multiple sclerosis patients on Cladribine

BackgroundCladribine was approved for the treatment of multiple sclerosis (MS). Real-world data is very limited. ObjectivesTo study the effectiveness and the safety of Cladribine treatment in only one group of MS patients after treatment with Cladribine for two years. MethodsThis observational, longitudinal prospective study. Eligible subjects were relapsing remitting MS patients who had at least two-year follow-up after Cladribine treatment. The primary endpoint was the proportion of relapse free patients. Secondary endpoints were ARR, change in EDSS scores, the proportion of patients with CDP, MRI activity, and NEDA-3 status, also the rate of occurrence of AEs. Patients were assessed for primary and secondary endpoints at the end of two years of follow-up. ResultsOf a total of seventy-two patients, 59 (81.9 %) were females, mean age of 36.32 + 10.06 years old, mean disease duration 7.21 + 6.19. Most patients (n = 32; 44.4 %) were naïve to any treatment. Forty patients (55.6 %) completed two courses of treatment. The primary endpoint showed that most of our cohort was relapse free (85 % versus 25 %; P < 0.001), Secondary endpoints showed that ARR was significantly reduced 0.15 + 0.36 versus 0.85 + 0.53; P < 0.01). Most of the cohort 90 % have no progression of disability. Few subjects had new T2 lesions (7.5 % versus 70.8 %; P < 0.001 and gadolinium enhancement 5 % versus 66.7 %; P < 0.001) in MRI compared to baseline. No evidence of disease activity 3 (NEDA-3) was achieved in 30 (75 %) patients. It was achieved in 87.5 % of naive patients versus 66.7 % in patients who received prior disease modification drugs before Cladribine initiation. Infections 6 (n = 6; 8.4 %) lymphocytopenia (n = 3; 4.2 %), and elevated liver enzymes (n = 1; 1.4 %) were reported. ConclusionCladribine treatment reduced significantly relapse rate and MRI activity. It was safe and tolerable. Early initiation of cladribine is associated with favorable outcomes.

Patterns of inflammation, microstructural alterations, and sodium accumulation define multiple sclerosis subtypes after 15 years from onset.

Conventional MRI is routinely used for the characterization of pathological changes in multiple sclerosis (MS), but due to its lack of specificity is unable to provide accurate prognoses, explain disease heterogeneity and reconcile the gap between observed clinical symptoms and radiological evidence. Quantitative MRI provides measures of physiological abnormalities, otherwise invisible to conventional MRI, that correlate with MS severity. Analyzing quantitative MRI measures through machine learning techniques has been shown to improve the understanding of the underlying disease by better delineating its alteration patterns. In this retrospective study, a cohort of healthy controls (HC) and MS patients with different subtypes, followed up 15 years from clinically isolated syndrome (CIS), was analyzed to produce a multi-modal set of quantitative MRI features encompassing relaxometry, microstructure, sodium ion concentration, and tissue volumetry. Random forest classifiers were used to train a model able to discriminate between HC, CIS, relapsing remitting (RR) and secondary progressive (SP) MS patients based on these features and, for each classification task, to identify the relative contribution of each MRI-derived tissue property to the classification task itself. Average classification accuracy scores of 99 and 95% were obtained when discriminating HC and CIS vs. SP, respectively; 82 and 83% for HC and CIS vs. RR; 76% for RR vs. SP, and 79% for HC vs. CIS. Different patterns of alterations were observed for each classification task, offering key insights in the understanding of MS phenotypes pathophysiology: atrophy and relaxometry emerged particularly in the classification of HC and CIS vs. MS, relaxometry within lesions in RR vs. SP, sodium ion concentration in HC vs. CIS, and microstructural alterations were involved across all tasks.

Early remission in multiple sclerosis is linked to altered coherence of the Cerebellar Network

BackgroundThe development of permanent disability in multiple sclerosis (MS) is highly variable among patients, and the exact mechanisms that contribute to this disability remain unknown.MethodsFollowing the idea that the brain has intrinsic network organization, we investigated changes of functional networks in MS patients to identify possible links between network reorganization and remission from clinical episodes in MS. Eighteen relapsing–remitting MS patients (RRMS) in their first clinical manifestation underwent resting-state functional MRI and again during remission. We used ten template networks, identified from independent component analysis, to compare changes in network coherence for each patient compared to those of 44 healthy controls from the Human Connectome Project test–retest dataset (two-sample t-test of pre-post differences). Combining a binomial test with Monte Carlo procedures, we tested four models of how functional coherence might change between the first clinical episode and remission: a network can change its coherence (a) with itself (“one-with-self”), (b) with another network (“one-with-other”), or (c) with a set of other networks (“one-with-many”), or (d) multiple networks can change their coherence with respect to one common network (“many-with-one”).ResultsWe found evidence supporting two of these hypotheses: coherence decreased between the Executive Control Network and several other networks (“one-with-many” hypothesis), and a set of networks altered their coherence with the Cerebellar Network (“many-with-one” hypothesis).ConclusionGiven the unexpected commonality of the Cerebellar Network’s altered coherence with other networks (a finding present in more than 70% of the patients, despite their clinical heterogeneity), we conclude that remission in MS may result from learning processes mediated by the Cerebellar Network.

SNFL applicability as additional monitoring tool in natalizumab extended interval dosing regimen for RRMS patients

IntroductionExtended interval dosing (EID) of Natalizumab (NAT) has been proposed to reduce progressive multifocal leukoencephalopathy (PML) risk associated with standard interval dosing (SID) in people with multiple sclerosis (MS). Previous studies have suggested that NAT effectiveness is maintained in the great majority of patients who switch from SID to EID; monitoring of disease activity is currently based exclusively on clinical and MRI parameters. Frequent MRI are expensive and not always applicable, underlining the need for biological markers able to detect central nervous system lesions. Serum Neurofilament-light chain (sNFL) currently represents the most promising biomarker of disease activity, prognosis and treatment response in MS, and their clinical suitability is increasingly evident. The objective of the present study is to assess the applicability of sNFL as additional/alternative measure of treatment efficacy during EID regimen. MethodsWe measured sNFL by Simoa technology in longitudinal samples from 63 Relapsing Remitting (RR) MS patients switched from SID to EID. Inclusion criteria: diagnosis of RRMS, age 18-60 years; NAT SID for at least 12 months; NEDA-3 (no evidence of disease activity) for at least 12 months; availability of at least 2 serum samples collected 6 months apart. Patients’ follow-up time during EID was at least 12 months and 2 blood samples were collected after at least 6 and 12 months. Clinical examination was performed before each infusion, while MRI 6 and 12 months after NAT initiation and according to PML risk during the whole study. ResultsNo patients showed clinical or MRI activity during the whole follow-up. sNFL levels measured during SID and EID were comparable, without significant difference between groups. The effect of EID on NFL levels did not show significant effects (LMM, p> 0.05) and sNFL levels did not vary with time during SID or EID protocols (LMM, p> 0.05). Intra-individual sNFL levels demonstrated overall stability during SID and EID (median CV=11% between SID and EID samples). According to our previously published reference values, sNFL levels were in the normal range in all samples, both during SID and EID. ConclusionsOur results suggest that sNFL quantification can be used as an alternative/additional approach to MRI in managing individual patients. The present work provides a new clinical application of sNFL to monitor NAT efficacy.

Major depression favors the expansion of Th17-like cells and decrease the proportion of CD39+Treg cell subsets in response to myelin antigen in multiple sclerosis patients.

Mood disorders have been associated with risk of clinical relapses in multiple sclerosis (MS), a demyelinating disease mediated by myelin-specific T cells. We aimed to investigate the impact of major depressive disorder (MDD) and cytokine profile of T-cells in relapsing remitting MS patients. For our study, plasma and PBMC were obtained from 60 MS patients (30 with lifetime MDD) in remission phase. The PBMC cultures were stimulated with anti-CD3/anti-CD28 beads or myelin basic protein (MBP), and effector and regulatory T cell phenotypes were determined by flow cytometry. The cytokine levels, both in the plasma or in the supernatants collected from PBMC cultures, were quantified by Luminex. In some experiments, the effect of serotonin (5-HT) was investigated. Here, higher Th17-related cytokine levels in response to anti-CD3/anti-CD28 and MBP were quantified in the plasma and PBMC cultures of the MS/MDD group in comparison with MS patients. Further, elevated frequency of CD4+ and CD8+ T cells capable of producing IL-17, IL-22 and GM-CSF was observed in depressed patients. Interestingly, the percentage of myelin-specific IFN-γ+IL-17+ and IFN-γ+GM-CSF+ CD4+ T cells directly correlated with neurological disabilities. In contrast, the occurrence of MDD reduced the proportion of MBP-specific CD39+Tregs subsets. Notably, the severity of both neurological disorder and depressive symptoms inversely correlated with these Tregs. Finally, the addition of 5-HT downregulated the release of Th17-related cytokines in response to anti-CD3/anti-CD28 and myelin antigen. In summary, our findings suggested that recurrent major depression, by favoring imbalances of effector Th17 and Treg cell subsets, contributes to MS severity.

Flow Cytometry Analysis of Blood Large Extracellular Vesicles in Patients with Multiple Sclerosis Experiencing Relapse of the Disease.

The number of people living with multiple sclerosis (MS) in developed countries is increasing. The management of patients is hindered by the absence of reliable laboratory tests accurately reflecting the disease activity. Extracellular vesicles (EVs) of different cell origin were reportedly elevated in MS patients. We assessed the diagnostic potential, with flow cytometry analysis, of fresh large EVs (lEVs), which scattered more light than the 590 nm silica beads and were isolated from the blood plasma of relapsing remitting MS patients. Venous blood was collected from 15 patients and 16 healthy controls (HC). The lEVs were isolated from fresh platelet-free plasma by centrifugation, labelled with antibodies and the presence of platelet (CD41+, CD36+), endothelial (CD105+), erythrocyte (CD235a+), leukocyte (CD45+, CD19+, CD3+) and phosphatidylserine (Annexin V+) positive lEVs was analyzed using standard flow cytometry. Cryo-electron microscopy was used to verify the presence of EVs in the analyzed plasma fractions. MS patients experiencing acute relapse had slightly reduced relative levels (% of positive lEVs) of CD105+, CD45+, CD3+, CD45+CD3+ or CD19+ labelled lEVs in comparison to healthy controls. An analysis of other markers or a comparison of absolute lEV counts (count of lEVs/µL) did not yield any significant differences. Our data do not support the hypothesis that the exacerbation of the disease in RRMS patients leads to an increased numbers of circulating plasma lEVs which can be monitored by standard flow cytometry.

Relation of sensorimotor and cognitive cerebellum functional connectivity with brain structural damage in patients with multiple sclerosis and no disability.

Background and purposeTo investigate the relationship between the functional connectivity (FC) of the sensorimotor and cognitive cerebellum and measures of structural damage in patients with multiple sclerosis (MS) and no physical disability.MethodsWe selected 144 relapsing–remitting MS patients with an Expanded Disability Status Scale score of ≤1.5 and 98 healthy controls from the Italian Neuroimaging Network Initiative database. From multimodal 3T magnetic resonance imaging (MRI), including functional MRI at rest, we calculated lesion load, cortical thickness, and white matter, cortical gray matter, and caudate, putamen, thalamic, and cerebellar volumes. Voxel‐wise FC of the sensorimotor and cognitive cerebellum was assessed with seed‐based analysis, and multiple regression analysis was used to evaluate the relationship between FC and structural damage.ResultsWhole brain, white matter, caudate, putamen, and thalamic volumes were reduced in patients compared to controls, whereas cortical gray matter was not significantly different in patients versus controls. Both the sensorimotor and cognitive cerebellum showed a widespread pattern of increased and decreased FC that were negatively associated with structural measures, indicating that the lower the FC, the greater the tissue loss. Lastly, among multiple structural measures, cortical gray matter and white matter volumes were the best predictors of cerebellar FC alterations.ConclusionsIncreased and decreased cerebellar FC with several brain areas coexist in MS patients with no disability. Our data suggest that white matter loss hampers FC, whereas, in the absence of atrophy, cortical volume represents the framework for FC to increase.

MicroRNA-484 and Apoptotic Protease Activating Factor-1 Gene in Relapsing Remitting Multiple Sclerosis: the Possible Interplay

Background: Emerging evidence suggests that dysregulated apoptosis might be implicated in the pathogenesis of multiple sclerosis (MS). In this study we evaluated the expression of miR-484 and its potential target gene Apoptotic protease activating factor-1 (APAF-1) in relapsing remitting MS patients (RRMS), correlated their expression levels to patients’ clinical characteristics and investigated their role as potential disease biomarkers.

Perivascular space is associated with brain atrophy in patients with multiple sclerosis.

Perivascular space (PVS) is associated with neurodegenerative and neuroimmune diseases. Multiple sclerosis (MS) is traditionally a neuroimmune disease. However, studies show neurodegeneration also plays a vital role in MS. At present, most studies conclude severer PVS in MS is an imaging marker of neuroinflammation, while a 7T MRI study suggests that PVS in MS is associated with neurodegeneration. In this study, 82 MS patients (n=82) and 32 healthy controls (n=32) were enrolled. The following indexes were measured: the number, size and distribution of PVS, the PVS score, corpus callosum index (CCI), corpus callosum area (CCA), the ratio of the corpus callosum to the cranium (CCR), aligned third ventricle width (a3VW), and unaligned third ventricle width (u3VW). The PVS score (4 vs. 3, P=0.041), PVSs number (103.280±45.107 vs. 87.625±30.139, P=0.035), and enlarged perivascular spaces (EPVSs) number (9 vs. 1, P<0.001) of MS patients were significantly higher than in the healthy controls. PVSs number (23.5 vs. 13) and EPVSs number (1 vs. 0) in the basal ganglia (BG), and EPVSs number (3 vs. 0) in centrum semiovale (CSO) of MS patients were significantly higher than in the healthy controls, P<0.001. In MS patients, PVS was correlated with age and hypertension but not to the extended disability status scale (EDSS) score and other clinical data. In MS patients, PVS score was correlated with CCA (rs=0.272; P=0.013) and the CCR (rs=0.219; P=0.048), and PVSs number was correlated with CCA (rs=0.255; P=0.021), the correlation disappeared after adjusting hypertension and age. In MS patients in remission, PVSs number was correlated with CCA (rs=0.487; P=0.019), CCR (rs=0.479; P=0.021), and PVS score was correlated with CCA (rs=0.453; P=0.03). After adjustment of hypertension and age, the total number of PVSs was correlated with CCA (rs=0.419; P=0.049). The PVS load in MS patients was heavier than healthy people, especially in BG and CSO. PVS was not correlated with EDSS in MS patients. The PVS of MS patients was associated with CCA and CCR, and PVSs number was independently related with CCA in MS patients in remission.

Alterations of host-gut microbiome interactions in multiple sclerosis.

SummaryBackgroundMultiple sclerosis (MS) has a complex genetic, immune and metabolic pathophysiology. Recent studies implicated the gut microbiome in MS pathogenesis. However, interactions between the microbiome and host immune system, metabolism and diet have not been studied over time in this disorder.MethodsWe performed a six-month longitudinal multi-omics study of 49 participants (24 untreated relapse remitting MS patients and 25 age, sex, race matched healthy control individuals. Gut microbiome composition and function were characterized using 16S and metagenomic shotgun sequencing. Flow cytometry was used to characterize blood immune cell populations and cytokine profiles. Circulating metabolites were profiled by untargeted UPLC-MS. A four-day food diary was recorded to capture the habitual dietary pattern of study participants.FindingsTogether with changes in blood immune cells, metagenomic analysis identified a number of gut microbiota decreased in MS patients compared to healthy controls, and microbiota positively or negatively correlated with degree of disability in MS patients. MS patients demonstrated perturbations of their blood metabolome, such as linoleate metabolic pathway, fatty acid biosynthesis, chalcone, dihydrochalcone, 4-nitrocatechol and methionine. Global correlations between multi-omics demonstrated a disrupted immune-microbiome relationship and a positive blood metabolome-microbiome correlation in MS. Specific feature association analysis identified a potential correlation network linking meat servings with decreased gut microbe B. thetaiotaomicron, increased Th17 cell and greater abundance of meat-associated blood metabolites. The microbiome and metabolome profiles remained stable over six months in MS and control individuals.InterpretationOur study identified multi-system alterations in gut microbiota, immune and blood metabolome of MS patients at global and individual feature level. Multi-OMICS data integration deciphered a potential important biological network that links meat intakes with increased meat-associated blood metabolite, decreased polysaccharides digesting bacteria, and increased circulating proinflammatory marker.FundingThis work was supported by the Washington University in St. Louis Institute of Clinical and Translational Sciences, funded, in part, by Grant Number # UL1 TR000448 from the National Institutes of Health, National Center for Advancing Translational Sciences, Clinical and Translational Sciences Award (Zhou Y, Piccio, L, Lovett-Racke A and Tarr PI); R01 NS10263304 (Zhou Y, Piccio L); the Leon and Harriet Felman Fund for Human MS Research (Piccio L and Cross AH). Cantoni C. was supported by the National MS Society Career Transition Fellowship (TA-180531003) and by donations from Whitelaw Terry, Jr. / Valerie Terry Fund. Ghezzi L. was supported by the Italian Multiple Sclerosis Society research fellowship (FISM 2018/B/1) and the National Multiple Sclerosis Society Post-Doctoral Fellowship (FG-190734474). Anne Cross was supported by The Manny & Rosalyn Rosenthal-Dr. John L. Trotter MS Center Chair in Neuroimmunology of the Barnes-Jewish Hospital Foundation. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Apoptotic protease activating factor-1 gene and MicroRNA-484: A possible interplay in relapsing remitting multiple sclerosis

BackgroundEmerging evidence suggests that dysregulated apoptosis might be implicated in the pathogenesis of multiple sclerosis (MS). The aim of the current study was to evaluate the expression of Apoptotic protease activating factor-1 (APAF1) mRNA and its potential regulator miR-484 in relapsing remitting MS patients (RRMS) and to investigate their role as potential disease biomarkers. MethodsAfter Bioinformatic analysis was conducted and revealed miR-484 involvement in the regulation of APAF-1 gene expression. Reverse Transcription-quantitative Real-Time PCR (RT-qPCR) was performed to detect the expression levels of APAF-1 and miR-484 in the peripheral blood mononuclear cells (PBMCs) of 34 RRMS patients recruited from the MS clinic of kasr al ainy hospital- faculty of medicine-Egypt and 34 healthy controls. ResultsAPAF-1 mRNA was significantly downregulated in patients whereas miR-484 expression was upregulated compared to controls (p < 0.01). Sensitivity and specificity of APAF-1 and miR-484 to diagnose MS was (85.3%, 76.5%) and (88.2% and 86.7%) respectively. ConclusionAPAF-1 and miR-484 could play a role as potential MS diagnostic biomarkers. However, absence of a control group of patients with other inflammatory diseases in our study warrants further research to corroborate our findings.

T1w/FLAIR ratio standardization as a myelin marker in MS patients

IntroductionCalculation of a T1w/T2w ratio was introduced as a proxy for myelin integrity in the brain of multiple sclerosis (MS) patients. Since nowadays 3D FLAIR is commonly used for lesion detection instead of T2w images, we introduce a T1w/FLAIR ratio as an alternative for the T1w/T2w ratio. ObjectivesBias and intensity variation are widely present between different scanners, between subjects and within subjects over time in T1w, T2w and FLAIR images. We present a standardized method for calculating a histogram calibrated T1w/FLAIR ratio to reduce bias and intensity variation in MR sequences from different scanners and at different time-points. Material and methods207 Relapsing Remitting MS patients were scanned on 4 different 3 T scanners with a protocol including 3D T1w, 2D T2w and 3D FLAIR images. After bias correction, T1w/FLAIR ratio maps and T1w/T2w ratio maps were calculated in 4 different ways: without calibration, with linear histogram calibration as described by Ganzetti et al. (2014), and by using 2 methods of non-linear histogram calibration. The first nonlinear calibration uses a template of extra-cerebral tissue and cerebrospinal fluid (CSF) brought from Montreal Neurological Institute (MNI) space to subject space; for the second nonlinear method we used an extra-cerebral tissue and CSF template of our own subjects. Additionally, we segmented several brain structures such as Normal Appearing White Matter (NAWM), Normal Appearing Grey Matter (NAGM), corpus callosum, thalami and MS lesions using Freesurfer and Samseg. ResultsThe coefficient of variation of T1w/FLAIR ratio in NAWM for the no calibrated, linear, and 2 nonlinear calibration methods were respectively 24, 19.1, 9.5, 13.8. The nonlinear methods of calibration showed the best results for calculating the T1w/FLAIR ratio with a smaller dispersion of the data and a smaller overlap of T1w/FLAIR ratio in the different segmented brain structures. T1w/T2w and T1w/FLAIR ratios showed a wider range of values compared to MTR values. ConclusionsCalibration of T1w/T2w and T1w/FLAIR ratio maps is imperative to account for the sources of variation described above. The nonlinear calibration methods showed the best reduction of between-subject and within-subject variability. The T1w/T2w and T1w/FLAIR ratio seem to be more sensitive to smaller changes in tissue integrity than MTR. Future work is needed to determine the exact substrate of T1w/FLAIR ratio and to obtain correlations with clinical outcome.

Preserved T-cell response in anti-CD20-treated multiple sclerosis patients following SARS-CoV-2 vaccination

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has tremendous implications for the management of patients with autoimmune conditions such as multiple sclerosis (MS) under immune therapies targeting CD20+ B cells (aCD20). Here, we investigated humoral and cellular immune responses, including anti-spike titers, neutralization against SARS-CoV-2 wild-type (WT), delta, and omicron variant and T cell responses of aCD20-treated relapsing-remitting MS patients following SARS-CoV-2 vaccination compared with healthy controls. Blood samples were collected within 4-8 weeks following the second vaccination against SARS-CoV-2. Sera were analyzed for anti-SARS-CoV-2 spike antibodies and neutralization capacity against pseudovirus for wild-type (WT), delta, and omicron variant. Peripheral blood mononuclear cells (PBMCs) were stimulated with a SARS-CoV-2 peptide pool and analyzed via flow cytometry. The aCD20-treated MS patients had lower anti-SARS-CoV-2-spike titers, which correlated with B cell repopulation. Sera of aCD20-treated patients had reduced capacity to neutralize WT, delta, and omicron pseudoviruses in vitro. On the contrary, PBMCs of aCD20-treated patients elicited higher frequencies of CD3+ T cells and CD4+ T cells and comparable response of cytotoxic T cells, while Th1 response was reduced following restimulation with SARS-CoV-2. In summary, aCD20-treated patients have a reduced humoral immune response, depending on B cell repopulation, in accordance with preserved cellular immune response, suggesting partial cellular protection against SARS-CoV-2.