scholarly journals Development of robust targeted proteomics assays for cerebrospinal fluid biomarkers in multiple sclerosis

2020 ◽  
Vol 17 (1) ◽  
Author(s):  
Astrid Guldbrandsen ◽  
Ragnhild Reehorst Lereim ◽  
Mari Jacobsen ◽  
Hilde Garberg ◽  
Ann Cathrine Kroksveen ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Cerebrospinal Fluid ◽  
Neurological Diseases ◽  
Trypsin Digestion ◽  
Csf Biomarkers ◽  
Biological Processes ◽  
High Quality ◽  
Significant Difference ◽  
Multiple Sclerosis Patients ◽  
Over Time

Abstract Background Verification of cerebrospinal fluid (CSF) biomarkers for multiple sclerosis and other neurological diseases is a major challenge due to a large number of candidates, limited sample material availability, disease and biological heterogeneity, and the lack of standardized assays. Furthermore, verification studies are often based on a low number of proteins from a single discovery experiment in medium-sized cohorts, where antibodies and surrogate peptides may differ, thus only providing an indication of proteins affected by the disease and not revealing the bigger picture or concluding on the validity of the markers. We here present a standard approach for locating promising biomarker candidates based on existing knowledge, resulting in high-quality assays covering the main biological processes affected by multiple sclerosis for comparable measurements over time. Methods Biomarker candidates were located in CSF-PR (proteomics.uib.no/csf-pr), and further filtered based on estimated concentration in CSF and biological function. Peptide surrogates for internal standards were selected according to relevant criteria, parallel reaction monitoring (PRM) assays created, and extensive assay quality testing performed, i.e. intra- and inter-day variation, trypsin digestion status over time, and whether the peptides were able to separate multiple sclerosis patients and controls. Results Assays were developed for 25 proteins, represented by 72 peptides selected according to relevant guidelines and available literature and tested for assay peptide suitability. Stability testing revealed 64 peptides with low intra- and inter-day variations, with 44 also being stably digested after 16 h of trypsin digestion, and 37 furthermore showing a significant difference between multiple sclerosis and controls, thereby confirming literature findings. Calibration curves and the linear area of measurement have, so far, been determined for 17 of these peptides. Conclusions We present 37 high-quality PRM assays across 21 CSF-proteins found to be affected by multiple sclerosis, along with a recommended workflow for future development of new assays. The assays can directly be used by others, thus enabling better comparison between studies. Finally, the assays can robustly and stably monitor biological processes in multiple sclerosis patients over time, thus potentially aiding in diagnosis and prognosis, and ultimately in treatment decisions.

scholarly journals Inflammation-related plasma and CSF biomarkers for multiple sclerosis

2020 ◽  
Vol 117 (23) ◽  
pp. 12952-12960 ◽  
Author(s):  
Jesse Huang ◽  
Mohsen Khademi ◽  
Lars Fugger ◽  
Örjan Lindhe ◽  
Lenka Novakova ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Cerebrospinal Fluid ◽  
Neurological Diseases ◽  
Area Under The Curve ◽  
Csf Biomarkers ◽  
Healthy Controls ◽  
Protein Biomarkers ◽  
Neurofilament Light Chain ◽  
Diagnostic Efficacy ◽  
Neurofilament Light

Effective biomarkers for multiple sclerosis diagnosis, assessment of prognosis, and treatment responses, in particular those measurable in blood, are largely lacking. We have investigated a broad set of protein biomarkers in cerebrospinal fluid (CSF) and plasma using a highly sensitive proteomic immunoassay. Cases from two independent cohorts were compared with healthy controls and patients with other neurological diseases. We identified and replicated 10 cerebrospinal fluid proteins including IL-12B, CD5, MIP-1a, and CXCL9 which had a combined diagnostic efficacy similar to immunoglobulin G (IgG) index and neurofilament light chain (area under the curve [AUC] = 0.95). Two plasma proteins, OSM and HGF, were also associated with multiple sclerosis in comparison to healthy controls. Sensitivity and specificity of combined CSF and plasma markers for multiple sclerosis were 85.7% and 73.5%, respectively. In the discovery cohort, eotaxin-1 (CCL11) was associated with disease duration particularly in patients who had secondary progressive disease (PCSF< 4 × 10−5,Pplasma< 4 × 10−5), and plasma CCL20 was associated with disease severity (P= 4 × 10−5), although both require further validation. Treatment with natalizumab and fingolimod showed different compartmental changes in protein levels of CSF and peripheral blood, respectively, including many disease-associated markers (e.g., IL12B, CD5) showing potential application for both diagnosing disease and monitoring treatment efficacy. We report a number of multiple sclerosis biomarkers in CSF and plasma for early disease detection and potential indicators for disease activity. Of particular importance is the set of markers discovered in blood, where validated biomarkers are lacking.

Silver staining of unconcentrated cerebrospinal fluid in agarose gel (Panagel) electrophoresis.

1984 ◽  
Vol 30 (5) ◽  
pp. 735-736 ◽  
Author(s):  
P D Mehta ◽  
S P Mehta ◽  
B A Patrick
Keyword(s):  
Multiple Sclerosis ◽  
Cerebrospinal Fluid ◽  
Silver Staining ◽  
Clinical Laboratory ◽  
Neurological Diseases ◽  
Agarose Gel ◽  
Oligoclonal Igg ◽  
Multiple Sclerosis Patients ◽  
Coomassie Brilliant ◽  
Oligoclonal Igg Bands

Abstract We subjected cerebrospinal fluid (CSF) from 20 patients with multiple sclerosis and 20 patients with other neurological diseases to agarose gel ( Panagel ) electrophoresis followed by staining with silver. Ten microliters of unconcentrated CSF from multiple sclerosis patients containing 0.4 to 0.8 microgram of immunoglobulin G was found to be optimum for detection of oligoclonal IgG bands, so identified by immunofixation. The band patterns for unconcentrated CSF stained with silver were almost identical to those for the same CSF concentrated 40-fold and stained with Coomassie Brilliant Blue. Silver staining thus enables the clinical laboratory to electrophorese unconcentrated CSF on commercially prepared ( Panagel ) plates.

T cells from multiple sclerosis patients recognize immunoglobulin G from cerebrospinal fluid

2003 ◽  
Vol 9 (3) ◽  
pp. 228-234 ◽  
Author(s):  
T Holmøy ◽  
B Vandvik ◽  
F Vartdal
Keyword(s):  
Multiple Sclerosis ◽  
T Cells ◽  
Cerebrospinal Fluid ◽  
T Cell ◽  
Mononuclear Cells ◽  
Neurological Diseases ◽  
Affinity Maturation ◽  
Peripheral Blood Mononuclear ◽  
Number Of Patients ◽  
Multiple Sclerosis Patients

Idiotopic sequences are created after V, D and J recombinations and by somatic mutations during affinity maturation of immuglobulin (Ig) molecules, and may therefore be potential immunogenic epitopes. Idiotope-specific T cells are able to activate and sustain the B cells producing such idiotopes. It is therefore possible that idiotope-specific intrathecal T cells could help maintain the persisting intrathecal synthesis of oligoclonal IgG observed in patients with multiple sclerosis (MS). This study was undertaken to examine T-cell responses to cerebrospinal fluid (CSF) IgG. Peripheral blood mononuclear cells (PBMC) from 14 of 21 MS patients and four of 17 control patients with other neurological diseases proliferated upon stimulation with autologous C SF IgG, while five and three, respectively, responded to serum IgG. By comparison, responses to myelin basic protein were recorded in only four MS and three control patients. Data from a limited number of patients indicate that the C SF IgG responsive cells were CD4+ and human leucocyte antigen DR restricted, that PBMC also respond to C SF IgG from other MS patients and that the C SF may contain T cells responding to autologous C SF IgG. This suggests that C SF IgG, or substances bound to this IgG, may represent T-cell immunogens, which could contribute to the intrathecal immune response in MS.

Correlation between DJ-1 levels in the cerebrospinal fluid and the progression of disabilities in multiple sclerosis patients

2008 ◽  
Vol 14 (8) ◽  
pp. 1056-1060 ◽  
Author(s):  
M Hirotani ◽  
C Maita ◽  
M Niino ◽  
SM Iguchi-Ariga ◽  
S Hamada ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Multiple Sclerosis ◽  
Cerebrospinal Fluid ◽  
Healthy Subjects ◽  
Significant Positive Correlation ◽  
Healthy Controls ◽  
Significant Difference ◽  
Relapsing Remitting ◽  
Multiple Sclerosis Patients

Objectives DJ-1 plays a key role in the anti-oxidative stress function. Increasing evidence supports the role of oxidative stress in the pathogenesis of multiple sclerosis (MS). The aim of this study was to investigate whether the DJ-1 levels were increased in patients with MS and to examine its association with the progression of MS. Methods Quantitative immunoblot assays were performed to evaluate the DJ-1 level in serum and cerebrospinal fluid (CSF) collected from relapsing–remitting patients with MS ( n = 29), disease controls subjects ( n = 14), and healthy subjects ( n = 44). Results No significant difference was observed in the serum DJ-1 level among the patients with MS, disease controls, and healthy controls. However, the CSF DJ-1 levels were significantly higher in the patients with MS than in the disease control subjects ( P < 0.0001). A significant positive correlation was also found between the CSF DJ-1 levels and the Multiple Sclerosis Severity Score ( P < 0.005, r = 0.501). Conclusions These results show that the CSF DJ-1 levels are significantly increased in the CSF of patients with MS and that the CSF DJ-1 levels may be associated with the disease progression of MS. Therefore, DJ-1 possibly plays an important role in the pathogenesis of MS.

JC virus in cerebrospinal fluid samples of multiple sclerosis patients at the first demyelinating event

2007 ◽  
Vol 13 (5) ◽  
pp. 590-595 ◽  
Author(s):  
Roberto Alvarez-Lafuente ◽  
Marta García-Montojo ◽  
Virginia De Las Heras ◽  
Manuel Bartolomé ◽  
Rafael Arroyo
Keyword(s):  
Multiple Sclerosis ◽  
Cerebrospinal Fluid ◽  
Jc Virus ◽  
Neurological Diseases ◽  
Positive Sample ◽  
Viral Loads ◽  
Immunological Activation ◽  
Viral Particles ◽  
Multiple Sclerosis Patients

Objective To evaluate the possible involvement of JC virus (JCV) in the aetiology of multiple sclerosis (MS), through the comparison of DNA prevalences and viral loads of JCV in cerebrospinal fluid (CSF) of MS patients at the first demyelinating event and subjects suffering from other neurological diseases (OND). Methods Seventy-three CSF samples (43 from MS patients at the first demyelinating event, and 30 from patients with OND) were collected; all MS cases were followed up from 1 to 6.7 years after they were diagnosed with clinically definite MS. DNA was extracted and analysed by real-time PCR for the detection of JCV genomes. Results We found JCV DNA in the CSF of two MS patients (4.7%) with a mean viral load of 2.1 and 6.7 copies/mL of CSF. Among the patients of the OND group we did not find any positive sample. We did not find any difference in the course of the disease between MS patients with and without JCV genomes in their CSF along the follow up. Conclusion JCV seems to be only a bystander in the pathology of MS, and the presence of cell-free viral particles could be related to the immunological activation of the disease, mainly during relapses. Multiple Sclerosis 2007; 13: 590-595. http://msj.sagepub.com

Increase of CD4+TNFα+IL-2−T cells in cerebrospinal fluid of multiple sclerosis patients

2009 ◽  
Vol 15 (1) ◽  
pp. 120-123 ◽  
Author(s):  
N Shi ◽  
Y Kawano ◽  
T Matsuoka ◽  
FJ Mei ◽  
T Ishizu ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
T Cells ◽  
Cerebrospinal Fluid ◽  
Peripheral Blood ◽  
Neurological Diseases ◽  
Significant Change ◽  
Multiple Sclerosis Patients

Intracellular production of TNFα and IL-2 after stimulation with phorbol myristate/ionomycin was flowcytometrically measured in CD4+ T cells from peripheral blood (PB) and cerebrospinal fluid (CSF) of 29 patients with multiple sclerosis (MS), and 16 with other inflammatory and 41 with other non-inflammatory neurological diseases. In CSF, the percentages of CD4+TNFα+IL-2−T cells were significantly higher in patients with MS than either of the controls, whereas no difference was found in CD4+TNFα+IL-2+T or CD4+TNFα−IL-2+T cells. The increase was more pronounced at relapse than in remission. No significant change was detected in PB. These findings suggested that CD4+TNFα+IL-2−T cells are intrathecally upregulated in MS.

scholarly journals Investigation of IL-17A, IL-17F, IL-34 Levels of Patient with Multiple Sclerosis and Pseudotumor Cerebri

Acta Medica ◽  
2021 ◽  
pp. 1-8
Author(s):  
Hayriye Soytürk ◽  
Şeyda Karabörk ◽  
Şule Aydın Türkoğlu
Keyword(s):  
Multiple Sclerosis ◽  
Cerebrospinal Fluid ◽  
Demyelinating Disease ◽  
Pseudotumor Cerebri ◽  
Radiologically Isolated Syndrome ◽  
Significant Difference ◽  
Cytokine Levels ◽  
New Treatment ◽  
Multiple Sclerosis Patients ◽  
Relapsing Remitting Multiple Sclerosis

Objective: Multiple Sclerosis is an inflammatory demyelinating disease characterized by lymphocyte infiltration and demyelination of brain tissue and central nervous system. Materials and Methods: This study aimed to evaluate the interleukin (IL-17A, IL-17F, and IL-34 cytokine) levels in the cerebrospinal fluid of Relapsing-Remitting Multiple Sclerosis (n=23), radiologically isolated syndrome (n=5) and pseudotumor cerebri (n=15) cases. In this study, lumbar puncture cerebrospinal fluid obtained from the patients who were diagnosed with Multiple Sclerosis aged between 21-55. The PTC group included patients with pseudotumor cerebri aged 28-60 years. The levels of IL-17A, IL-17F, IL-34 cytokines were determined by ELISA kit. Results: In this study, Among the studied cytokines in the cerebrospinal fluid samples of the patients, median (min-max) values of IL-17A for the Demyelinated group and pseudotumor cerebri group were 50 (7-257) pg/ml and 2 (1-6) pg/ml respectively, a statistically significant difference (p<0.01) has been observed in between the two groups. Median (min-max) values of IL-17F for the Demyelinated group and pseudotumor cerebri group were 32 (6-891) pg/ml and 2 (1-3) pg/ml respectively, a statistically significant difference (p<0.01) has been observed between the two groups. Median (min-max) values of IL-34 for Demyelinated group and pseudotumor cerebri group were 16 (4-197) pg/ml and 2 (1-11) pg/ml respectively, a statistically significant difference (p<0.01) has been observed in between the two groups (Lower limit for the cytokine values have been determined as IL-17A: 3,93 pg/ml, IL-17F: 2,23 pg/ml, IL-34: 3,12 pg/ml). IL-34, was found to be high in Multiple sclerosis patients. This is important for the cerebral endothelial reaction in Multiple sclerosis. Conclusion: The high levels of IL-34 in cerebrospinal fluid samples suggest that it may be a new treatment strategy and an adjunct cytokine in the diagnosis of neuroinflammatory and neurodegenerative diseases such as multiple sclerosis and demyelinating disease. More extensive studies are needed to determine whether IL-34 can be a marker in the return of the disease from radiologically isolated syndrome to clinical MS.

Herpesviruses and human endogenous retroviral sequences in the cerebrospinal fluid of multiple sclerosis patients

2008 ◽  
Vol 14 (5) ◽  
pp. 595-601 ◽  
Author(s):  
R Álvarez-Lafuente ◽  
M García-Montojo ◽  
V De Las Heras ◽  
MI Domínguez-Mozo ◽  
M Bartolome ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Cerebrospinal Fluid ◽  
Neurological Diseases ◽  
Human Herpesvirus ◽  
Endogenous Retroviruses ◽  
Varicella Zoster ◽  
Barr Virus ◽  
Dna And Rna ◽  
Simplex Virus ◽  
Multiple Sclerosis Patients

Objective To analyze the possible role of human herpesvirus (HHVs) and human endogenous retroviruses (HERVs) infection in multiple sclerosis (MS) pathogenesis. Methods A total of 92 cerebrospinal fluid (CSF) samples were collected: 48 from MS patients at the first clinically evident demyelinating event, 23 from patients with other inflammatory neurological diseases (OINDs) and 21 from patients with other non-inflammatory neurological diseases (ONINDs). Total DNA and RNA were isolated, and the prevalences and viral loads of herpes simplex virus (HSV), varicella-zoster virus (VZV), Epstein–Barr virus (EBV), cytomegalovirus (CMV), HHV-6, HERV-H and HERV-W in the CSF of MS patients and controls were evaluated using a quantitative real-time polymerase chain reaction assay. Results (i) For HSV, 1/48 (2.1%, 86 copies/ml of CSF) MS patients and 1/23 (4.3%, 115.2 copies/ml of CSF) OIND patients (a myelitis case) had HSV sequences in the CSF; (ii) for EBV, only 1/48 (2.1%, 72 copies/ml of CSF) MS patients was positive for EBV; (iii) for HHV-6, only 5/48 (10.4%) MS patients had HHV-6 genomes in their CSF (128.1 copies/ml of CSF); (iv) we did not find any positive cases for VZV, CMV, HERV-H and HERV-W among MS patients or controls; (v) no cases of co-infections were found; (vi) the whole prevalence of HHVs was 7/48 (14.6%) for MS patients and 1/44 (2.3%) for controls ( p = 0.038). Conclusion The findings described here show that HHV infection is more frequent in the CSF of MS patients than in patients with other neurological diseases; however, only HHV-6 seems to be involved in the pathogenesis of MS in a subset of patients.

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