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Patients with multiple sclerosis (MS) suffer with age an early immunosenescence process, which influence the treatment response and increase the risk of infections. We explored whether lipid-specific oligoclonal IgM bands (LS-OCMB) associated with highly inflammatory MS modify the immunological profile induced by age in MS. This cross-sectional study included 263 MS patients who were classified according to the presence (M+, n=72) and absence (M-, n=191) of LS-OCMB. CSF cellular subsets and molecules implicated in immunosenescence were explored. In M- patients, aging induced remarkable decreases in absolute CSF counts of CD4+ and CD8+ T lymphocytes, including Th1 and Th17 cells, and of B cells, including those secreting TNF-alpha. It also increased serum anti-CMV IgG antibody titers (indicative of immunosenescence) and CSF CHI3L1 levels (related to astrocyte activation). In contrast, M+ patients showed an age-associated increase of TIM-3 (a biomarker of T cell exhaustion) and increased values of CHI3L1, independently of age. Finally, in both groups, age induced an increase in CSF levels of PD-L1 (an inductor of T cell tolerance) and activin A (part of the senescence-associated secretome and related to inflammaging). These changes were independent of the disease duration. Finally, this resulted in augmented disability. In summary, all MS patients experience with age a modest induction of T-cell tolerance and an activation of the innate immunity, resulting in increased disability. Additionally, M- patients show clear decreases in CSF lymphocyte numbers, which could increase the risk of infections. Thus, age and immunological status are important for tailoring effective therapies in MS.

There are an increasing number of treatments available for multiple sclerosis (MS). The early identification of optimal responders to individual treatments is important to achieve individualized therapy. With this aim, we performed a multicenter retrospective longitudinal study including 186 MS patients treated with natalizumab who were followed for 2 years. We analyzed the following variables at recruitment: sex, current age, age at disease onset, disease duration, EDSS, number of T2 and Gd + lesions, IgG and IgM oligoclonal bands, HLA class II (DR, DRB, DQA, DQB, and DRB1*15:01), IgG and IgM antibody titers against human herpesvirus 6 (HHV-6) and the antibody response to Epstein–Barr virus (EBV) through the measurement of the anti-EBNA-1 and anti-VCA IgG titers, in relation to clinical response (no relapses or disability progression), and to NEDA-3 (no evidence of disease activity in terms of clinical response and no changes in MRI scans either) after 2-years follow-up. Baseline EDSS score, baseline EBNA-1 IgG titers and percentage change of HHV6 IgG titers between baseline and 6 month visits were significantly different in clinical responders and in NEDA-3 status (all of them remained significant in the multivariate analysis). We identified three variables for the early identification of natalizumab optimal responders in a rapid and cost-effective approach.

Primary progressive multiple sclerosis is a poorly understood disease entity with no specific prognostic biomarkers and scarce therapeutic options. We aimed to identify disease activity biomarkers in multiple sclerosis by performing an RNA sequencing approach in peripheral blood mononuclear cells from a discovery cohort of 44 untreated patients with multiple sclerosis belonging to different clinical forms and activity phases of the disease, and 12 healthy control subjects. A validation cohort of 58 patients with multiple sclerosis and 26 healthy control subjects was included in the study to replicate the RNA sequencing findings. The RNA sequencing revealed an interleukin 1 beta (IL1B) signature in patients with primary progressive multiple sclerosis. Subsequent immunophenotyping pointed to blood monocytes as responsible for the IL1B signature observed in this group of patients. Functional experiments at baseline measuring apoptosis-associated speck-like protein containing a CARD (ASC) speck formation showed that the NOD-leucine rich repeat and pyrin containing protein 3 (NLRP3) inflammasome was overactive in monocytes from patients with primary progressive multiple sclerosis, and canonical NLRP3 inflammasome activation with a combination of ATP plus lipopolysaccharide was associated with increased IL1B production in this group of patients. Primary progressive multiple sclerosis patients with high IL1B gene expression levels in peripheral blood mononuclear cells progressed significantly faster compared to patients with low IL1B levels based on the time to reach an EDSS of 6.0 and the Multiple Sclerosis Severity Score. In agreement with peripheral blood findings, both NLRP3 and IL1B expression in brain tissue from patients with primary progressive multiple sclerosis was mainly restricted to cells of myeloid lineage. Treatment of mice with a specific NLRP3 inflammasome inhibitor attenuated established experimental autoimmune encephalomyelitis disease severity and impro

Importance: Neurofilament light protein (NfL) is elevated in cerebrospinal fluid (CSF) of a number of neurological conditions compared with healthy controls (HC) and is a candidate biomarker for neuroaxonal damage. The influence of age and sex is largely unknown, and levels across neurological disorders have not been compared systematically to date. Objectives: To assess the associations of age, sex, and diagnosis with NfL in CSF (cNfL) and to evaluate its potential in discriminating clinically similar conditions. Data Sources: PubMed was searched for studies published between January 1, 2006, and January 1, 2016, reporting cNfL levels (using the search terms neurofilament light and cerebrospinal fluid) in neurological or psychiatric conditions and/or in HC. Study Selection: Studies reporting NfL levels measured in lumbar CSF using a commercially available immunoassay, as well as age and sex. Data Extraction and Synthesis: Individual-level data were requested from study authors. Generalized linear mixed-effects models were used to estimate the fixed effects of age, sex, and diagnosis on log-transformed NfL levels, with cohort of origin modeled as a random intercept. Main Outcome and Measure: The cNfL levels adjusted for age and sex across diagnoses. Results: Data were collected for 10059 individuals (mean [SD] age, 59.7 [18.8] years; 54.1% female). Thirty-five diagnoses were identified, including inflammatory diseases of the central nervous system (n = 2795), dementias and predementia stages (n = 4284), parkinsonian disorders (n = 984), and HC (n = 1332). The cNfL was elevated compared with HC in a majority of neurological conditions studied. Highest levels were observed in cognitively impaired HIV-positive individuals (iHIV), amyotrophic lateral sclerosis, frontotemporal dementia (FTD), and Huntington disease. In 33.3% of diagnoses, including HC, multiple sclerosis, Alzheimer disease (AD), and Parkinson disease (PD), cNfL was higher in men than women. The cNfL incr

Objective: to validate kappa free light chain (KFLC) and lambda free light chain (LFLC) indices as a diagnostic biomarker in multiple sclerosis (MS). Methods: we performed a multicenter study including 745 patients from 18 centers (219 controls and 526 clinically isolated syndrome (CIS)/MS patients) with a known oligoclonal IgG band (OCB) status. KFLC and LFLC were measured in paired cerebrospinal fluid (CSF) and serum samples. Gaussian mixture modeling was used to define a cut-off for KFLC and LFLC indexes. Results: the cut-off for the KFLC index was 6.6 (95% confidence interval (CI) = 5.2–138.1). The cut-off for the LFLC index was 6.9 (95% CI = 4.5–22.2). For CIS/MS patients, sensitivity of the KFLC index (0.88; 95% CI = 0.85–0.90) was higher than OCB (0.82; 95%CI = 0.79–0.85; p < 0.001), but specificity (0.83; 95% CI = 0.78–0.88) was lower (OCB = 0.92; 95% CI = 0.89–0.96; p < 0.001). Both sensitivity and specificity for the LFLC index were lower than OCB. Conclusion: compared with OCB, the KFLC index is more sensitive but less specific for diagnosing CIS/MS. Lacking an elevated KFLC index is more powerful for excluding MS compared with OCB but the latter is more important for ruling in a diagnosis of CIS/MS., NA

Recent studies have begun to point out the contribution of microbiota to multiple sclerosis (MS) pathogenesis. Theiler’s murine encephalomyelitis virus induced demyelinating disease (TMEV-IDD) is a model of progressive MS. Here, we first analyze the effect of intracerebral infection with TMEV on commensal microbiota and secondly, whether the early microbiota depletion influences the immune responses to TMEV on the acute phase (14 dpi) and its impact on the chronic phase (85 dpi). The intracranial inoculation of TMEV was associated with a moderate dysbiosis. The oral administration of antibiotics (ABX) of broad spectrum modified neuroimmune responses to TMEV dampening brain CD4+ and CD8+ T infiltration during the acute phase. The expression of cytokines, chemokines and VP2 capsid protein was enhanced and accompanied by clusters of activated microglia disseminated throughout the brain. Furthermore, ABX treated mice displayed lower levels of CD4+ and CD8+T cells in cervical and mesenteric lymph nodes. Increased mortality to TMEV was observed after ABX cessation at day 28pi. On the chronic phase, mice that survived after ABX withdrawal and recovered microbiota diversity showed subtle changes in brain cell infiltrates, microglia and gene expression of cytokines. Accordingly, the surviving mice of the group ABX-TMEV displayed similar disease severity than TMEV mice.

Item does not contain fulltext, We have previously identified eight novel autoantibody targets in the cerebrospinal fluid of multiple sclerosis (MS) patients, including sperm-associated Ag 16 (SPAG16). In the current study, we further investigated the autoantibody response against SPAG16-a protein with unknown function in the CNS-and its expression in MS pathology. Using isoelectric focusing, we detected SPAG16-specific oligoclonal bands in the cerebrospinal fluid of 5 of 23 MS patients (22%). Analysis of the anti-SPAG16 Ab reactivity in the plasma of a total of 531 donors using ELISA demonstrated significantly elevated anti-SPAG16 Ab levels (p = 0.002) in 32 of 153 MS patients (21%) compared with all other control groups with 95% specificity for the disease. To investigate the pathologic relevance of anti-SPAG16 Abs in vivo, anti-SPAG16 Abs were injected in mice with experimental autoimmune encephalomyelitis, resulting in a significant disease exacerbation. Finally, we demonstrated a consistent upregulation of SPAG16 in MS brain and experimental autoimmune encephalomyelitis spinal cord lesions, more specifically in reactive astrocytes. We conclude that SPAG16 is a novel autoantibody target in a subgroup of MS patients and in combination with other diagnostic criteria, elevated levels of anti-SPAG16 Abs could be used as a biomarker for diagnosis. Furthermore, the pathologic relevance of anti-SPAG16 Abs was shown in vivo.

[Background and Objectives] Resistant and susceptible mouse strains to experimental autoimmune encephalomyelitis (EAE), an inducible demyelinating experimental disease serving as animal model for multiple sclerosis, have been described. We aimed to explore MHC-independent mechanisms inducing resistance to EAE., [Methods] For EAE induction, female C57BL/6 (susceptible strain) and CD1 (resistant outbred strain showing heterogeneous MHC antigens) mice were immunized with the 35-55 peptide of myelin oligodendrocyte glycoprotein (MOG35-55). We studied T cell proliferation, regulatory and effector cell subpopulations, intracellular and serum cytokine patterns, and titers of anti-MOG serum antibodies., [Results] Upon immunization with MOG35-55, T lymphocytes from susceptible mice but not that of resistant strain were capable of proliferating when stimulated with MOG35-55. Accordingly, resistant mice experienced a rise in regulatory B cells (P = 0.001) and, to a lower extent, in regulatory T cells (P = 0.02) compared with C57BL/6 susceptible mice. As a consequence, MOG35-55-immunized C57BL/6 mice showed higher percentages of CD4+ T cells producing both IFN-gamma (P = 0.02) and IL-17 (P = 0.009) and higher serum levels of IL-17 (P = 0.04) than resistant mice., [Conclusions] Expansion of regulatory B and T cells contributes to the induction of resistance to EAE by an MHC-independent mechanism. © 2014 Nieves Marín et al.

Item does not contain fulltext, Limited information is available on the identity of antigens targeted by antibodies present in cerebrospinal fluid (CSF) of patients with clinically isolated syndrome (CIS). The aim of this study was to identify novel antigens for CIS and investigate their prognostic potential to predict conversion to multiple sclerosis (MS). We applied serological antigen selection (SAS) to identify antigens interacting with antibodies present in the pooled CSF from four CIS patients, who developed MS. Antibody reactivity towards CIS antigens identified by SAS was tested in CSF and serum from patients with CIS (n = 123/n = 108), MS (n = 65/n = 44), and other (inflammatory) neurological diseases (n = 75/n = 38) as well as in healthy control sera (n = 44). Using SAS, a panel of six novel CIS candidate antigens was identified. CSF antibody reactivity was detected in both CIS and relapsing-remitting (RR) MS. Serum reactivity was significantly increased in CIS and RR-MS as compared with controls (p = 0.03). For two antigens, the frequency of antibody-positive patients was higher in CIS patients who converted to MS as compared with CIS patients without conversion. We identified novel CIS antigens to which antibody reactivity was primarily detected in CIS and RR-MS as compared to controls. Possible prognostic potential could be demonstrated for two antigens.

The presence of Penicilin, Streptomicine, Tetraciclines, Bacitracine, Neomicine and Kanamicine has been investigated in 1.346 samples of natural mil from the following provinces in the Southern region of Spain: Córdoba, Jaén, Granada, Málaga, Cádiz, Huelva and Sevilla. The microbiological tests performed were: a) detection of non specific inhibiting substances by the reduction of methylene blue test (Frank, 1965) and b) Jaquet and Stee"s (1952) cavities method for the detection of antibiotic substances indicated with sensitive germs. The germs used were: Sarcina Lutea (Núm. 9341 of American Collection of Culture Types, ATCC), for the Penicilin; Bacillus Subtilis (ATCC 6633), for the Streptomicine; Bacillus Cereus var. Mycoides (ATCC; 11778), for Tetraciclines; Sarcina Subflava (ATCC 7468), for the Bacitracine, and Staphilococcus Epidermis (ATCC, 12228), for the Neomicine, and Staphilococcus Aureus (ATCC, 6538-P for the Kanamicine, For each microbiological test of antibiotic detection studied, the fines of regression and the minimal detectable limits were obtained experimentally. According to the FAO/OMS mixed Comittee (1970), antibiotics should be used in such away that residues can not be detected in foods for human consumption; snd the Spanish Alimentary Code prohibits the sale and use of foods containing antibiotics. The levels of contarnination by no specific inhibiting substances is alarmingly high since it contained in 32"62 per 100 of the samples of milk analysed. Penicilin was detected in 1"19 per 100 of the samples, Streptomicine in 1.11 per 100, Tetraciclines in 1.04 per 100, Bacitracine in 9.8 per 100, Kanamicine in 0.30 per 100, and Neomicine in 1.04 per 100. Of the 1.346 milk samples analysed, 11.66 per 100 contained one or more antibiotic. In the province of Córdoba 25.87 per 100 of the milk samples contained antibiotics; in that of Jaén, 8.46 per 100; in Seville, 6.50 per 100; in Cádiz, 5 per 100; in Granada, 4 per 100, and in the province of Malaga, 2, Se ha investigado la presencia de penicilina, estreptomicina, tetraciclinas, ba¬citracina, neomicina y kanamicina en 1346 muestras, de leche natural, procedentes de diferentes zonas de las siguientes provincias de la región sur de España: Córdoba, Jaén, Granada, Málaga, Cádiz, Huelva y Sevilla. Las pruebas microbiológicas han sido: a) detección de sustancias inhibidoras inespecíficas por la prueba de reducción del azul de metileno (Frank, 1965) y b) método de las cavidades de Jacquet y Steeg (1952) para la detección de las surtan cias antibióticas indicadas con gérmenes sensiblesios gérmenes sensibles utilizados han sido: para la la penicilina, la Sarcina lutea (núm. 9341 de la Colección Americana de cultivos tipo, ATCC); para la estreptomicina, el Bacillus subtilis (ATCC 6633); para las tetraciclinas, el Bacillus cereus var mycoides (ATCC 11778); para la bacitracina, la Sarcina subflava (ATCC 7468); para la neomicina, el Staphilococcus epidermis (ATCC 12228); y para la kanamicina, el Staphylococcus aureus (ATCC 6538-P). Se han obtenido experimentalmente, para cada prueba microbiológica de detección específica de los antibióticos estudiados, las líneas de regresión y los límites mínimos detectables. Según el Comité mixto FAO/OMS (1970) los antibióticos deben emplearse de forma que no se detecten residuos en los alimentos para consumo humano; y el Código alimentario español prohíbe la venta y utilización de alimentos con antibióticos. El nivel de contaminación por sustancias inhibidoras inespecíficas es alarmantemente alto ya que el 32"62 p. 100 de las muestras de leche analizadas lo contenían. La penicilina se ha detectado en el 1"19 p. 100 de las muestras, la estreptomicina en el 1"11 p. 100, las tetraciclinas en el 1"04 p. 100, la bacitracina en el 9"8 p. 100, la kanamicina en el 0"30 p. 100 y la neomicina en el 1"04 p. 100. De las 1346 muestras de leche analizadas, el 11"66 p. 100 contenían uño o más antibióticos. En la provincia de Córdoba el 25"87 p. 100 dulas

Contains fulltext : 89601.pdf (publisher's version ) (Closed access), BACKGROUND: Neurofilament proteins (Nf) are highly specific biomarkers for neuronal death and axonal degeneration. As these markers become more widely used, an inter-laboratory validation study is required to identify assay criteria for high quality performance. METHODS: The UmanDiagnostics NF-light (R)enzyme-linked immunoabsorbent assays (ELISA) for the neurofilament light chain (NfL, 68kDa) was used to test the intra-assay and inter-laboratory coefficient of variation (CV) between 35 laboratories worldwide on 15 cerebrospinal fluid (CSF) samples. Critical factors, such as sample transport and storage, analytical delays, reaction temperature and time, the laboratories' accuracy and preparation of standards were documented and used for the statistical analyses. RESULTS: The intra-laboratory CV averaged 3.3% and the inter-laboratory CV 59%. The results from the test laboratories correlated with those from the reference laboratory (R=0.60, p<0.0001). Correcting for critical factors improved the strength of the correlation. Differences in the accuracy of standard preparation were identified as the most critical factor. Correcting for the error introduced by variation in the protein standards improved the correlation to R=0.98, p<0.0001 with an averaged inter-laboratory CV of 14%. The corrected overall inter-rater agreement was subtantial (0.6) according to Fleiss' multi-rater kappa and Gwet's AC1 statistics. CONCLUSION: This multi-center validation study identified the lack of preparation of accurate and consistent protein standards as the main reason for a poor inter-laboratory CV. This issue is also relevant to other protein biomarkers based on this type of assay and will need to be solved in order to achieve an acceptable level of analytical accuracy. The raw data of this study is available online.