Antibodies In Multiple Sclerosis Patients Research Articles

Serum Reactive Antibodies against the N-Methyl-D-Aspartate Receptor NR2 Subunit-Could They Act as Potential Biomarkers?

Synaptic dysfunction and disrupted communication between neuronal and glial cells play an essential role in the underlying mechanisms of multiple sclerosis (MS). Earlier studies have revealed the importance of glutamate receptors, particularly the N-methyl-D-aspartate (NMDA) receptor, in excitotoxicity, leading to abnormal synaptic transmission and damage of neurons. Our study aimed to determine whether antibodies to the NR2 subunit of NMDAR are detected in MS patients and evaluate the correlation between antibody presence and clinical outcome. Furthermore, our focus extended to examine a possible link between NR2 reactivity and anti-coagulant antibody levels as pro-inflammatory molecules associated with MS. A cross-sectional study was carried out, including 95 patients with MS and 61 age- and gender-matched healthy controls (HCs). The enzyme-linked immunosorbent assay was used to detect anti-NR2 antibodies in serum samples of participants along with IgG antibodies against factor (F)VIIa, thrombin, prothrombin, FXa, and plasmin. According to our results, significantly elevated levels of anti-NR2 antibodies were detected in MS patients compared to HCs (p < 0.05), and this holds true when we compared the Relapsing-Remitting MS course with HCs (p < 0.05). A monotonically increasing correlation was found between NR2 seropositivity and advanced disability (rs = 0.30; p < 0.01), anti-NR2 antibodies and disease worsening (rs = 0.24; p < 0.05), as well as between antibody activity against NR2 and thrombin (rs = 0.33; p < 0.01). The presence of anti-NR2 antibodies in MS patients was less associated with anti-plasmin IgG antibodies [OR:0.96 (95%CI: 0.92-0.99); p < 0.05]; however, such an association was not demonstrated when analyzing only RRMS patients. In view of our findings, NR2-reactive antibodies may play, paving the way for further research into their potential as biomarkers and therapeutic targets in MS.

Clinical and MRI Correlates of Antinuclear Antibodies in Multiple Sclerosis Patients

Clinical and MRI Correlates of Antinuclear Antibodies in Multiple Sclerosis Patients

Treatment- and population-specific genetic risk factors for anti-drug antibodies against interferon-beta: a GWAS

BackgroundUpon treatment with biopharmaceuticals, the immune system may produce anti-drug antibodies (ADA) that inhibit the therapy. Up to 40% of multiple sclerosis patients treated with interferon β (IFNβ) develop ADA, for which a genetic predisposition exists. Here, we present a genome-wide association study on ADA and predict the occurrence of antibodies in multiple sclerosis patients treated with different interferon β preparations.MethodsWe analyzed a large sample of 2757 genotyped and imputed patients from two cohorts (Sweden and Germany), split between a discovery and a replication dataset. Binding ADA (bADA) levels were measured by capture-ELISA, neutralizing ADA (nADA) titers using a bioassay. Genome-wide association analyses were conducted stratified by cohort and treatment preparation, followed by fixed-effects meta-analysis.ResultsBinding ADA levels and nADA titers were correlated and showed a significant heritability (47% and 50%, respectively). The risk factors differed strongly by treatment preparation: The top-associated and replicated variants for nADA presence were the HLA-associated variants rs77278603 in IFNβ-1a s.c.- (odds ratio (OR) = 3.55 (95% confidence interval = 2.81–4.48), p = 2.1 × 10−26) and rs28366299 in IFNβ-1b s.c.-treated patients (OR = 3.56 (2.69–4.72), p = 6.6 × 10−19). The rs77278603-correlated HLA haplotype DR15-DQ6 conferred risk specifically for IFNβ-1a s.c. (OR = 2.88 (2.29–3.61), p = 7.4 × 10−20) while DR3-DQ2 was protective (OR = 0.37 (0.27–0.52), p = 3.7 × 10−09). The haplotype DR4-DQ3 was the major risk haplotype for IFNβ-1b s.c. (OR = 7.35 (4.33–12.47), p = 1.5 × 10−13). These haplotypes exhibit large population-specific frequency differences. The best prediction models were achieved for ADA in IFNβ-1a s.c.-treated patients. Here, the prediction in the Swedish cohort showed AUC = 0.91 (0.85–0.95), sensitivity = 0.78, and specificity = 0.90; patients with the top 30% of genetic risk had, compared to patients in the bottom 30%, an OR = 73.9 (11.8–463.6, p = 4.4 × 10−6) of developing nADA. In the German cohort, the AUC of the same model was 0.83 (0.71–0.92), sensitivity = 0.80, specificity = 0.76, with an OR = 13.8 (3.0–63.3, p = 7.5 × 10−4).ConclusionsWe identified several HLA-associated genetic risk factors for ADA against interferon β, which were specific for treatment preparations and population backgrounds. Genetic prediction models could robustly identify patients at risk for developing ADA and might be used for personalized therapy recommendations and stratified ADA screening in clinical practice. These analyses serve as a roadmap for genetic characterizations of ADA against other biopharmaceutical compounds.

Using Serum Metabolomics to Predict Development of Anti-drug Antibodies in Multiple Sclerosis Patients Treated With IFNβ.

Background: Neutralizing anti-drug antibodies (ADA) can greatly reduce the efficacy of biopharmaceuticals used to treat patients with multiple sclerosis (MS). However, the biological factors pre-disposing an individual to develop ADA are poorly characterized. Thus, there is an unmet clinical need for biomarkers to predict the development of immunogenicity, and subsequent treatment failure. Up to 35% of MS patients treated with beta interferons (IFNβ) develop ADA. Here we use machine learning to predict immunogenicity against IFNβ utilizing serum metabolomics data.Methods: Serum samples were collected from 89 MS patients as part of the ABIRISK consortium—a multi-center prospective study of ADA development. Metabolites and ADA were quantified prior to and after IFNβ treatment. Thirty patients became ADA positive during the first year of treatment (ADA+). We tested the efficacy of six binary classification models using 10-fold cross validation; k-nearest neighbors, decision tree, random forest, support vector machine and lasso (Least Absolute Shrinkage and Selection Operator) logistic regression with and without interactions.Results: We were able to predict future immunogenicity from baseline metabolomics data. Lasso logistic regression with/without interactions and support vector machines were the most successful at identifying ADA+ or ADA– cases, respectively. Furthermore, patients who become ADA+ had a distinct metabolic response to IFNβ in the first 3 months, with 29 differentially regulated metabolites. Machine learning algorithms could also predict ADA status based on metabolite concentrations at 3 months. Lasso logistic regressions had the greatest proportion of correct classifications [F1 score (accuracy measure) = 0.808, specificity = 0.913]. Finally, we hypothesized that serum lipids could contribute to ADA development by altering immune-cell lipid rafts. This was supported by experimental evidence demonstrating that, prior to IFNβ exposure, lipid raft-associated lipids were differentially expressed between MS patients who became ADA+ or remained ADA–.Conclusion: Serum metabolites are a promising biomarker for prediction of ADA development in MS patients treated with IFNβ, and could provide novel insight into mechanisms of immunogenicity.

Prevalence of Anti-JC Virus (JCV) Antibodies in the Multiple Sclerosis (MS) Population in Cyprus: A Retrospective Study.

Background and Purpose Progressive multifocal leukoencephalopathy (PML) is a debilitating disease of the central nervous system caused by the ubiquitous polyomavirus JC (JCV) in immunocompromised hosts. In recent years, a new subpopulation of patients at risk for PML has emerged, due to the growing use of immunomodulatory or immunosuppressive therapies in autoimmune diseases such as multiple sclerosis (MS). The anti-JCV antibody index is used as a stratification tool in assessing the risk of developing PML. The objective of this study was to retrospectively describe the prevalence of anti-JCV antibodies in the MS population in Cyprus. Methods We retrospectively collected the demographics of 214 MS patients in Cyprus who were screened for anti-JCV antibodies using the STRATIFY JCV™ assay between September 2011 and June 2018. Logistic regression analysis was used to examine the effect of demographic variables on seropositivity, and bivariate tests were used to assess the association between demographic characteristics and JCV AI index. Results A total of 214 MS patients in Cyprus were tested. Overall anti-JCV antibody prevalence was 45.8% (95% confidence interval 37.2%–55.8%). We could not establish a significant association between seropositivity and increasing age or sex. In the subgroup analysis of natalizumab-treated patients, the annual seroconversion rate was 4.5%. Conclusions Overall seroprevalence of anti-JCV antibodies in MS patients in Cyprus using the STRATIFY JCV assay was lower than the worldwide reported mean. Although previously reported, in our study, the anti-JCV antibody seropositivity was not associated with increasing age or sex.

Differential serostatus of Epstein-Barr virus in Iranian MS patients with various clinical patterns.

Background: Epidemiological evidence suggests a role of Epstein-Barr virus (EBV) in triggering the pathogenesis of Multiple Sclerosis (MS). The aim of this study was to assess the EBV-specific antibodies in MS patients with various clinical patterns and their association with the production of IFN-γ, IL-12, and IL-4 cytokines compared with healthy individuals. Methods: We measured EBNA-1 IgG, VCA IgG, and production of IFN-γ, IL-12 and IL-4 cytokines in patients with different clinical patterns and healthy controls using ELISA method. Results: There was a higher titer of anti-EBV antibodies in MS patients compared to healthy controls. SPMS patients generated higher EBNA-1 levels than those with RRMS and PPMS patients whereas; the level of VCA IgG was higher in the RRMS patients than PPMS. In PPMS patients, a significant increase was found in IFN-γ and IL-12 cytokines compared to other subtypes, whereas IL-4 cytokine had a decreased level compared to RRMS patients. Higher anti-EBV antibodies are associated with increased IL-12 cytokine in RRMS patients. However, no significant correlation was found between these antibodies and other secreted cytokines. Conclusion: EBV infection is one of the strong risk factors for MS. Acting on these factors could be useful to decrease the incidence and disease exacerbation of MS. Study of the antibody levels to EBV virus could be useful for evaluating MS risk score in each clinical subtypes.

Galectin-8 as an immunosuppressor in experimental autoimmune encephalomyelitis and a target of human early prognostic antibodies in multiple sclerosis.

Galectin-8 (Gal-8) is a member of a glycan-binding protein family that regulates the immune system, among other functions, and is a target of antibodies in autoimmune disorders. However, its role in multiple sclerosis (MS), an autoimmune inflammatory disease of the central nervous system (CNS), remains unknown. We study the consequences of Gal-8 silencing on lymphocyte subpopulations and the development of experimental autoimmune encephalitis (EAE), to then assess the presence and clinical meaning of anti-Gal-8 antibodies in MS patients. Lgals8/Lac-Z knock-in mice lacking Gal-8 expression have higher polarization toward Th17 cells accompanied with decreased CCR6+ and higher CXCR3+ regulatory T cells (Tregs) frequency. These conditions result in exacerbated MOG35-55 peptide-induced EAE. Gal-8 eliminates activated Th17 but not Th1 cells by apoptosis and ameliorates EAE in C57BL/6 wild-type mice. β-gal histochemistry reflecting the activity of the Gal-8 promoter revealed Gal-8 expression in a wide range of CNS regions, including high expression in the choroid-plexus. Accordingly, we detected Gal-8 in human cerebrospinal fluid, suggesting a role in the CNS immune-surveillance circuit. In addition, we show that MS patients generate function-blocking anti-Gal-8 antibodies with pathogenic potential. Such antibodies block cell adhesion and Gal-8-induced Th17 apoptosis. Furthermore, circulating anti-Gal-8 antibodies associate with relapsing-remitting MS (RRMS), and not with progressive MS phenotypes, predicting clinical disability at diagnosis within the first year of follow-up. Our results reveal that Gal-8 has an immunosuppressive protective role against autoimmune CNS inflammation, modulating the balance of Th17 and Th1 polarization and their respective Tregs. Such a role can be counteracted during RRMS by anti-Gal-8 antibodies, worsening disease prognosis. Even though anti-Gal-8 antibodies are not specific for MS, our results suggest that they could be a potential early severity biomarker in RRMS.

Evaluation of Serum Auto Antibodies in Multiple Sclerosis Patients: A Case Control Study

Background and Purpose: Multiple sclerosis (MS) is believed to be an immune-mediated disorder that develops from an interaction of the individual's genetic and as yet unidentified environmental causes. The prevalence of auto-antibodies in multiple sclerosis patients and their clinical associations vary in various studies. The aim of this study was to determine serum auto antibodies in multiple sclerosis patients. Methods: This cross-sectional case-control study investigated anti-phospholipids antibody (APLA), antinuclear antibody (ANA), anti-cardiolipin antibody (ACLA), anti-neutrophilic antibodies (ANCA), anti-beta-2-glycoprotein I (anti β2GPI), and anti-double strand DNA (anti-ds-DNA) in 54 consecutive patients with relapsing remitting multiple sclerosis (RRMS) and clinically isolated syndrome (CIS) who were referred to Imam Reza outpatient clinic of Shiraz University of Medical Sciences. The results were compared with 25 healthy individuals as the control group. Results : Among 54 patients with relapsing-remitting MS or clinically isolated syndrome, at least one abnormal tests were found in 15 patients (27.9%), 6 (11.1%) had positive antinuclear antibodies, 3 (5.6%) had positive anti cardiolpin antibody (ACLA) and P-ANCA was positive in 2(3.7%) of patients and C-ANCA was positive in 1(1.9%) of patients. None of the patients had any clinical manifestations other than MS symptoms. In the patient group, anti-ds-DNA antibody was positive in 5.6% of cases; statistically it had no significant difference with the control group (0%) (P=0.7), but anti- phospholipids antibody (APLA) and B2GPI were negative in all patient and control groups. The females had more positive auto-antibodies in comparison to males, but statistically their difference was not significant. Conclusion: The results of this study showed that a significant number of patients with relapsing remitting multiple sclerosis and clinically isolated syndrome have positive serum auto-antibodies tests (including ANA, ANCA and ACLA) without clinical expression of any other autoimmune disease.

Auto‐Immunity Targeting the Enteric Nervous System May Cause Constipation in Multiple Sclerosis

Multiple sclerosis (MS) is an inflammatory disease of the central nervous system (CNS) that is primarily associated with somatic motor, sensory, and visual deficits. This disease can also cause gastrointestinal (GI) dysmotility, including constipation, but the etiology of this symptom in MS is poorly understood. Additionally, patients with MS typically have serum antibodies against CNS myelin and neuronal proteins that are also found in the enteric nervous system (ENS). Here, we test the hypothesis that constipation in MS is caused by auto‐antibodies that target the ENS. First, we investigated whether serum from MS patients contains antibodies that are immunoreactive against ENS targets of guinea pig myenteric plexus. We found that MS serum (n=20) targets ENS cells more intensely than age and gender matched healthy serum (n=20) (p&lt;0.05), with stained structures including enteric neurons and glia. Next, we tested whether mice with experimental autoimmune encephalomyelitis (EAE) exhibit signs of constipation by assessing whole GI transit, colonic propulsive motility, and fecal consistency. EAE was induced in C57BL/6 or SJL/J mice by immunization against mouse spinal cord homogenate. Whole GI transit was slower in EAE SJL mice than controls (p&lt;0.05) but was not different in EAE C57 mice. Both SJL and C57 mice with EAE had significantly slower colonic transit than control mice of the same background (C57 &amp; SJL p&lt;0.05). The percentage of fecal water content was significantly lower in SJL and C57 EAE mice compared to controls (C57 p&lt;0.01, SJL p&lt;0.0001). To test whether antibodies play a role in GI dysmotility in EAE, we assessed motility in B cell deficient EAE mice (muMt− mice). Colonic transit time was slower in EAE muMt− mice compared to muMt− control mice (p&lt;0.05), but there was no difference in fecal water content. In summary, these data demonstrate that serum antibodies in MS patients target the ENS and could cause constipation. Further, the EAE mouse model of MS has disrupted GI motility, and this effect is partially ameliorated in B cell deficient EAE mice. Together, these data suggest that GI dysmotility in MS involves an autoimmune mediated disruption of enteric neuro‐glial function.Support or Funding InformationSupported by a pilot grant from the NMSS, and a predoctoral fellowship to ETS from NIDDK/NIH.

Detection of potassium channel KIR4.1 antibodies in Multiple Sclerosis patients

The presence of KIR4.1 antibodies has been proposed to be a characteristic of Multiple Sclerosis (MS). This could have a significant impact on disease management. However, the validation of the initial findings has failed till date. Conflicting results have been attributed to difficulties in isolating the lower-glycosylated (LG) KIR4.1 expressed in oligodendrocytes, the putative target antigen of autoantibodies.The aim of this study is to verify the presence of KIR4.1 antibodies in MS patients, by independently replicating the originally-described procedure.Assay procedure consisted of KIR4.1 expression in HEK293 cells, 3-step elution to isolate LG-KIR4.1 in elution fraction 3, and ELISA. Sera of 48 MS patients and 46 HCs were studied in 21 working sessions.In a preliminary analysis, we observed different KIR4.1 antibody levels between MS patients and Healthy Controls (HCs). However, a high variability across working sessions was observed and the sensitivity of the assay was very low. Thus, stringent criteria were established in order to identify working sessions in which the pure LG-KIR4.1 was isolated. As per these criteria, we detected LG-KIR4.1 antibodies in 28% of MS patients and 5% of HCs.Unlike previous findings, this study is in agreement with the original report. We propose further efforts be made towards the development of a uniform method to establish the detection of KIR4.1 antibodies in MS patients.

Serum IgG against Simian Virus 40 antigens are hampered by high levels of sHLA-G in patients affected by inflammatory neurological diseases, as multiple sclerosis.

BackgroundMany investigators detected the simian polyomavirus SV40 footprints in human brain tumors and neurologic diseases and recently it has been indicated that SV40 seems to be associated with multiple sclerosis (MS) disease. Interestingly, SV40 interacts with human leukocyte antigen (HLA) class I molecules for cell entry. HLA class I antigens, in particular non-classical HLA-G molecules, characterized by an immune-regulatory function, are involved in MS disease, and the levels of these molecules are modified according with the disease status.ObjectiveWe investigated in serum samples, from Italian patients affected by MS, other inflammatory diseases (OIND), non-inflammatory neurological diseases (NIND) and healthy subjects (HS), SV40-antibody and soluble sHLA-G and the association between SV40-prevalence and sHLA-G levels.MethodsELISA tests were used for SV40-antibodies detection and sHLA-G quantitation in serum samples.ResultsThe presence of SV40 antibodies was observed in 6 % of patients affected by MS (N = 4/63), 10 % of OIND (N = 8/77) and 15 % of NIND (N = 9/59), which is suggestive of a lower prevalence in respect to HS (22 %, N = 18/83). MS patients are characterized by higher sHLA-G serum levels (13.9 ± 0.9 ng/ml; mean ± St. Error) in comparison with OIND (6.7 ± 0.8 ng/ml), NIND (2.9 ± 0.4 ng/ml) and HS (2.6 ± 0.7 ng/ml) subjects. Interestingly, we observed an inverse correlation between SV40 antibody prevalence and sHLA-G serum levels in MS patients.ConclusionThe data obtained showed a low prevalence of SV40 antibodies in MS patients. These results seems to be due to a generalized status of inability to counteract SV40 infection via antibody production. In particular, we hypothesize that SV40 immune-inhibitory direct effect and the presence of high levels of the immune-inhibitory HLA-G molecules could co-operate in impairing B lymphocyte activation towards SV40 specific peptides.

The association between IgG and IgM antibodies against cardiolipin, β2-glycoprotein I and Domain I of β2-glycoprotein I with disease profile in patients with multiple sclerosis.

Antiphospholipid antibodies (aPL) occur in patients with multiple sclerosis (MS) with a number of studies reporting elevated levels; their exact prevalence and pathogenic role remain unclear. Epidemiological studies associate MS with an increased risk of deep venous thromboembolism and stroke; overlapping clinical features with APS. Antibodies against the first domain - Domain I (DI) - of β2glycoprotein I (β2GPI), show the most clinical significance and evidence for pathogenicity in the antiphospholipid syndrome (APS), but have not yet been investigated in MS. Serum from a well-defined cohort of 127 MS patients and 92 healthy controls were tested for IgM and IgG antibodies against cardiolipin (CL), β2GPI and DI. Higher frequency of IgM and IgG anti-CL were found in MS patients (18.1% and 21.3%), compared to controls (1.1% in both cases, p<0.0001). We report that anti-DI antibodies were associated with MS patients, with 6.3% and 7.1% positive for IgM and IgG, respectively, compared to controls, 1.1% (p<0.05). IgM anti-CL antibodies were elevated in secondary progressive MS and primary progressive MS compared to relapse-remitting MS, (p<0.005). This study enrolled the largest number of patients with definite MS for studying the association with aPL. Although we confirmed IgM and IgG anti-CL antibodies occur in patients with MS, this is the first study that identified anti-DI antibodies in MS patients. This new finding may prove valuable and future studies are required to evaluate its role as a potential risk factor of thromboembolic phenomena in MS.

Expression of interferon type-I receptor isoforms, clinical response and development of neutralizing antibodies in multiple sclerosis patients – results of a prospective study

Abstract Background: One of the first line treatments for relapsing-remitting multiple sclerosis (RRMS) is interferon-β (IFNb), a cytokine with immune-modulatory effects. There is a high degree of variability in the response to the drug which is, among other factors, due to the presence of neutralizing antibodies (NABs) occurring late during therapy. Methods: The objective of this study was to determine whether the response to IFNb therapy and NAB development can be predicted based on the expression levels of the type-I interferon receptors IFNAR1, IFNAR2a, IFNAR2b, and IFNAR2c before start of treatment. The IFNAR expression levels in 163 samples of patients with relapsing-remitting MS were measured by real-time polymerase chain reaction (PCR). Results: Pre-treatment IFNAR2c expression levels were somewhat lower in patients who developed NAB during treatment compared to NAB-negative patients. No significant differences in the expression levels of other IFNAR subtypes and isotypes were found. Baseline IFNAR levels were not predictive of the clinical response after 2 years. Conclusions: Overall, there was a small, non-significant effect of IFNAR2c baseline levels on NAB development but no relation to clinical endpoints. Lower expression of IFNAR2c receptors could lead to higher IFNb levels inducing a higher rate of antibody response.

Herpetic infection and anti-varicella-zoster virus antibodies in multiple sclerosis patients treated with fingolimod

Herpetic infection and anti-varicella-zoster virus antibodies in multiple sclerosis patients treated with fingolimod

Significant low prevalence of antibodies reacting with simian virus 40 mimotopes in serum samples from patients affected by inflammatory neurologic diseases, including multiple sclerosis.

Many investigations were carried out on the association between viruses and multiple sclerosis (MS). Indeed, early studies reported the detections of neurotropic virus footprints in the CNS of patients with MS. In this study, sera from patients affected by MS, other inflammatory (OIND) and non-inflammatory neurologic diseases (NIND) were analyzed for antibodies against the polyomavirus, Simian Virus 40 (SV40). An indirect enzyme-linked immunosorbent assay (ELISA), with two synthetic peptides, which mimic SV40 antigens, was employed to detect specific antibodies in sera from patients affected by MS, OIND, NIND and healthy subjects (HS). Immunologic data indicate that in sera from MS patients antibodies against SV40 mimotopes are detectable with a low prevalence, 6%, whereas in HS of the same mean age, 40 yrs, the prevalence was 22%. The difference is statistically significant (P = 0.001). Significant is also the difference between MS vs. NIND patients (6% vs. 17%; P = 0.0254), whereas no significant difference was detected between MS vs OIND (6% vs 10%; P>0.05). The prevalence of SV40 antibodies in MS patients is 70% lower than that revealed in HS.

A modified immune tolerant mouse model to study the immunogenicity of recombinant human interferon beta

Interferon beta may induce antibodies in multiple sclerosis patients and the incidence of immunogenicity depends on the type of product. These antibodies can reduce the efficacy of interferon beta. Two transgenic immune tolerant mouse models for human interferon beta (hIFNβ) (C57Bl/6, and C57Bl/6×FVB/N F1 hybrid mice) have been developed previously for studying immunogenicity. These models, however, may not be used for every interferon beta product because of the lack of immunogenicity in the wildtype genetic background. We therefore developed a modified transgenic mouse model by backcrossing the F1 hybrid C57Bl/6×FVB/N transgenic mice with wildtype FVB/N for 10 generations. These F10 offspring (referred to hear as FVB/N) have a genetic background consisting of mostly FVB/N (99.9%) and very little C57Bl/6 (0.1%), and are expected to have the more sensitive antibody producing phenotype of the parental FVB/N strain. The newly generated “FVB/N” strain was assessed for antibody formation against different rhIFNβ formulations compared to the C57Bl/6, and C57Bl/6×FVB/N transgenic mouse models.The new FVB/N transgenic mouse model was more sensitive for all tested rhIFNβ products, and the difference in antibody titers between the transgenic and non-transgenic mice of the FVB/N strain was much bigger compared to the antibody levels of the C57Bl/6, and C57Bl/6×FVB/N strains.

Lack of confirmation of anti-inward rectifying potassium channel 4.1 antibodies as reliable markers of multiple sclerosis

Background: auto-antibodies against the potassium channel inward rectifying potassium channel 4.1 (Kir4.1) have previously been identified in 46% of patients with multiple sclerosis (MS). Objectives: to confirm these findings. Methods: we evaluated the presence of anti-Kir4.1 antibodies by enzyme-linked immunosorbent assay (ELISA) and immunofluorescence in 268 MS patients, 46 patients with other neurological diseases (OND) and 45 healthy controls. Results: anti-Kir4.1 antibodies were found in 7.5% of MS patients, 4.3% of OND patients and 4.4% of healthy controls. Immunofluorescence analysis did not identify any specific staining. Conclusions: we confirmed the presence of anti-Kir4.1 antibodies in MS patients, but at a much lower prevalence than previously reported.

Epstein–Barr virus and Mycobacterium avium subsp. paratuberculosis peptides are cross recognized by anti-myelin basic protein antibodies in multiple sclerosis patients

Epstein–Barr virus and Mycobacterium avium subsp. paratuberculosis (MAP) have been associated to multiple sclerosis (MS). We searched for antibodies against the homologous peptides Epstein–Barr virus nuclear antigen 1 (EBNA1)400–413, MAP_0106c protein (MAP)121–132, and myelin basic protein (MBP)85–98 on a MS Sardinian cohort, showing that these antibodies are highly prevalent among MS patients compared to healthy controls. Competitive assay demonstrated that antibodies recognizing EBNA1400–413 and MAP121–132 cross-react with MBP85–98, possibly through a molecular mimicry mechanism. Indeed, the fact that peptides from different pathogens can be cross-recognized by antibodies targeting self-epitopes supports the hypothesis that EBV and MAP might trigger autoimmunity through a common target.

Deficient Phosphorylation of Stat1 in Leukocytes Identifies Neutralizing Antibodies in Multiple Sclerosis Patients Treated with Interferon-Beta

BackgroundAnti interferon-beta (IFN-β) neutralizing antibodies (NAb) affect efficacy of treatment of multiple sclerosis patients, but exactly when the detrimental effects of NAbs offset therapeutic efficacy is debated. Quantification of intracellular pathway-specific phosphorylation by phospho-specific flow cytometry (phosphoflow) is a promising tool for evaluation of these effects in primary immune cells from treated patients at the single-cell level.MethodSamples for phosphoflow and gene expression changes were collected before administration of IFN-β and at four, six, and eight hours thereafter. Patients were NAb negative (n = 3) or were NAb positive with low/medium (n = 1) or high (n = 2) NAb titers. Levels of phosphorylation of six Stat transcription factors (pStat) in seven cell subtypes and expression levels of 71 pathway-specific genes in whole blood were measured. The data was subjected to principal component analysis (PCA), fifty-fifty MANOVA, ANOVA, and partial least square regression (PLSR).ResultsPCA of pStat levels clustered patients according to NAb class independently of time. PCA of gene expression data clustered patients according to NAb class but was affected by time and treatment. In the fifty-fifty MANOVA, NAb class was significant for both pStat levels and gene expression data. The ANOVA identified pStat1 protein in several cell subtypes as significantly affected by NAb class. The best fitting model for NAb prediction based on PLSR included pStat1 in monocytes, T cells, or lymphocytes and pStat3 in monocytes (r = 0.97). Gene expression data were slightly less predictive of NAb titers.ConclusionBased on this proof of concept study, we hypothesize that NAb effects can be monitored by evaluation of a single biomarker, pStat1, in either monocytes or T cells by phosphoflow directly after IFN-β administration. The method will significantly reduce cost relative to labor intensive in vitro methods and offers a patient-specific approach to NAb evaluation.

JCV epidemiology in MS (JEMS)—Epidemiology of anti-JCV antibody prevalence in multiple sclerosis patients—Portuguese data

BackgroundProgressive multifocal leukoencephalopathy, caused by oligodendrocyte lytic infection by JCVirus, is a growing concern for patients undergoing immune modulatory therapies for treatment of autoimmune diseases, such as multiple sclerosis (MS). The objective of JEMS was to describe the prevalence of anti-JCV antibodies in MS patients and to assess the various factors associated to it. MethodsSerum samples were collected and tested for anti-JCV antibody using the STRATIFY JCV™ assay in 131 Portuguese MS patients. Factors potentially associated with prevalence were also evaluated, as well as the effect of established risk factors. ResultsIn the population of 131 Portuguese patients included in the JEMS, the overall anti-JCV antibody prevalence was 69.5% (95% CI, 61.6–77.4). The anti-JCV antibody prevalence did not seem to be influenced by demographic characteristics, although results demonstrate a non-significant trend for increased prevalence with age. Disease characteristics, treatment duration, treatment history, prior immunosuppressive therapy use and natalizumab exposure duration did not seem to be associated with anti-JCV prevalence. ConclusionThe results of Portuguese MS patients participating in the JEMS study present some differences when compared with the global population and literature results. An overall prevalence higher than expected raises awareness for data confirmation with greater sample size studies.