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QC 20180522

Immunological and inflammatory reactions have been suggested to have a role in the development of schizophrenia, a hypothesis that has recently been supported by genetic data. The aim of our study was to perform an unbiased search for autoantibodies in patients with a first psychotic episode, and to explore the association between any seroreactivity and the development of a Diagnostic and Statistical Manual of Mental Disorders, fourth edition (DSM-IV) disorder characterized by chronic or relapsing psychotic symptoms. We collected plasma samples from 53 patients when they were treated for their first-episode psychosis, and 41 non-psychotic controls, after which the patients were followed for a mean duration of 7 years. Thirty patients were diagnosed with schizophrenia, delusional disorder, schizoaffective disorder, bipolar disorder or a long-term unspecified nonorganic psychosis during follow-up, whereas 23 patients achieved complete remission. At the end of follow-up, plasma samples were analyzed for IgG reactivity to 2304 fragments of human proteins using a multiplexed affinity proteomic technique. Eight patient samples showed autoreactivity to the N-terminal fragment of the PAGE (P antigen) protein family (PAGE2B/PAGE2/PAGE5), whereas no such autoreactivity was seen among the controls. PAGE autoreactivity was associated with a significantly increased risk of being diagnosed with schizophrenia during follow-up (odds ratio 6.7, relative risk 4.6). An immunohistochemistry analysis using antisera raised against the N-terminal fragment stained an unknown extracellular target in human cortical brain tissue. Our findings suggest that autoreactivity to the N-terminal portion of the PAGE protein family is associated with schizophrenia in a subset of patients with first-episode psychosis., QC 20170824

Protein profiling through affinity proteomic approaches represents a powerful strategy for the analysis of human body fluids. Cerebrospinal fluid (CSF), being the fluid proximal to the central nervous system, is commonly analyzed in the context of neurological diseases, and can offer novel insights into the physiological state of the brain. Ultimately, and by analyzing the presence of brain-derived proteins in larger sets of samples that represent different phenotypes, profiling of CSF may serve as an important source to discover and verify disease-associated markers. Here, we describe a multiplexed and flexible protein profiling approach using antibody-based assays on suspension bead arrays. Through streamlined sample processing, protein biotinylation, and single-binder assay readout, this method enables high-throughput neuroproteomic analysis of up to 384 proteins in 384 samples., QC 20171019

The underlying molecular mechanisms of autoimmune diseases are poorly understood. To unravel the autoimmune processes across diseases, comprehensive and unbiased analyses of proteins targets recognized by the adaptive immune system are needed. Here we present an approach starting from high-density peptide arrays to characterize autoantibody repertoires and to identify new autoantigens. A set of ten plasma and serum samples from subjects with multiple sclerosis, narcolepsy, and without any disease diagnosis were profiled on a peptide array representing the whole proteome, hosting 2.2 million 12-mer peptides with a six amino acid lateral shift. On the basis of the IgG reactivities found on these whole-proteome peptide micro arrays, a set of 23 samples was then studied on a targeted array with 174 000 12-mer peptides of single amino acid lateral shift. Finally, verification of IgG reactivities was conducted with a larger sample set (n = 448) using the bead-based peptide microarrays. The presented workflow employed three different peptide microarray formats to discover and resolve the epitopes of human autoantibodies and revealed two potentially new autoantigens: MAP3K7 in multiple sclerosis and NRXN1 in narcolepsy. The presented strategy provides insights into antibody repertoire reactivity at a peptide level and may accelerate the discovery and validation of autoantigens in human diseases., QC 20170524

Purpose: This study is part of a larger effort aiming to expand the knowledge of brain-enriched proteins in human cerebrospinal fluid (CSF) and to provide novel insight into the relation between such proteins and different neurodegenerative diseases. Experimental design: Here 280 brain-enriched proteins in CSF from patients with Alzheimer's disease (AD), Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are profiled. In total, 441 human samples of ventricular CSF collected post mortem and lumbar CSF collected ante mortem are analyzed using 376 antibodies in a suspension bead array setup, utilizing a direct labelling approach. Results: Among several proteins displaying differentiated profiles between sample groups, we focus here on two synaptic proteins, neuromodulin (GAP43) and neurogranin (NRGN). They are both found at elevated levels in CSF from AD patients in two independent cohorts, providing disease-associated profiles in addition to verifying and strengthening previously observed patterns. Increased levels are also observed for patients for whom the AD diagnosis was not established at the time of sampling. Conclusions and clinical relevance: These findings indicate that analyzing the brain-enriched proteins in CSF is of particular interest to increase the understanding of the CSF proteome and its relation to neurodegenerative disorders. In addition, this study lends support to the notion that measurements of these synaptic proteins could potentially be of great relevance in future diagnostic tests for AD.

Background: Alzheimer's disease (AD) is a chronic neurodegenerative disorder accounting for more than 50% of all dementia cases. AD neuropathology is characterized by the formation of extracellular plaques and intracellular neurofibrillary tangles consisting of aggregated amyloid-beta and tau, respectively. The disease mechanism has only been partially elucidated and is believed to also involve many other proteins. Objective: This study intended to perform a proteomic profiling of post mortem AD brains and compare it with control brains as well as brains from other neurological diseases to gain insight into the disease pathology. Methods: Here we used label-free shotgun mass spectrometry to analyze temporal neocortex samples from AD, other neurological disorders, and non-demented controls, in order to identify additional proteins that are altered in AD. The mass spectrometry results were verified by antibody suspension bead arrays. Results: We found 50 proteins with altered levels between AD and control brains. The majority of these proteins were found at lower levels in AD. Pathway analyses revealed that several of the decreased proteins play a role in exocytic and endocytic pathways, whereas several of the increased proteins are related to extracellular vesicles. Using antibody-based analysis, we verified the mass spectrometry results for five representative proteins from this group of proteins (CD9, HSP72, PI42A, TALDO, and VAMP2) and GFAP, a marker for neuroinflammation. Conclusions: Several proteins involved in exo-endocytic pathways and extracellular vesicle functions display altered levels in the AD brain. We hypothesize that such changes may result in disturbed cellular clearance and a perturbed cell-to-cell communication that may contribute to neuronal dysfunction and cell death in AD., QC 20161207

Narcolepsy is a chronic sleep disorder with a yet unknown cause, but the specific loss of hypocretin-producing neurons together with a strong human leukocyte antigen (HLA) association has led to the hypothesis that autoimmune mechanisms might be involved. Here, we describe an extensive effort to profile autoimmunity repertoires in serum with the aim to find disease-related autoantigens. Initially, 57 serum samples from vaccine-associated and sporadic narcolepsy patients and controls were screened for IgG reactivity towards 10 846 fragments of human proteins using planar microarrays. The discovered differential reactivities were verified on suspension bead arrays in the same sample collection followed by further investigation of 14 antigens in 176 independent samples, including 57 narcolepsy patients. Among these 14 antigens, methyltransferase-like 22 (METTL22) and 5'-nucleotidase cytosolic IA (NT5C1A) were recognized at a higher frequency in narcolepsy patients of both sample sets. Upon sequence analysis of the 14 proteins, polymerase family, member 3 (PARP3), acyl-CoA-binding domain containing 7 (ARID4B), glutaminase 2 (GLS2) and cyclin-dependent kinase-like 1 (CDKL1) were found to contain amino acid sequences with homology to proteins found in the H1N1 vaccine. These findings could become useful elements of further clinical assays that aim towards a better phenotypic understanding of narcolepsy and its triggers., QC 20170109