Reproducibility of targeted lipidome analyses (Lipidyzer) in plasma and erythrocytes over a 6-week period

It is essential to measure lipid biomarkers with a high reproducibility to prevent biased results. We compared the lipid composition and inter-day reproducibility of lipid measurements in plasma and erythrocytes. Samples from 42 individuals (77% women, mean age 65 years, mean body mass index (BMI) 27 kg/m(2)), obtained non-fasted at baseline and after 6 weeks, were used for quantification of up to 1000 lipid species across 13 lipid classes with the Lipidyzer platform. Intraclass correlation coefficients (ICCs) were calculated to investigate the variability of lipid concentrations between timepoints. The ICC distribution of lipids in plasma and erythrocytes were compared using Wilcoxon tests. After data processing, the analyses included 630 lipids in plasma and 286 in erythrocytes. From these, 230 lipids overlapped between sample types. In plasma, 78% of lipid measurements were reproduced well to excellently, compared to 37% in erythrocytes. The ICC score distribution in plasma (median ICC 0.69) was significantly better than in erythrocytes (median ICC 0.51) (p-value < 0.001). At the class level, reproducibility in plasma was superior for triacylglycerols and cholesteryl esters while ceramides, diacylglycerols, (lyso)phosphatidylethanolamines, and sphingomyelins showed better reproducibility in erythrocytes. Although in plasma overall reproducibility was superior, differences at individual and class levels may favor the use of erythrocytes.Clinical epidemiolog

Acute phase inflammation is characterized by rapid changes in plasma/peritoneal fluid N-glycosylation in mice.

To access publisher's full text version of this article, please click on the hyperlink in Additional Links field or click on the hyperlink at the top of the page marked Files. This article is open access.Murine zymosan-induced peritonitis is a widely used model for studying the molecular and cellular events responsible for the initiation, persistence and/or resolution of inflammation. Among these events, it is becoming increasingly evident that changes in glycosylation of proteins, especially in the plasma and at the site of inflammation, play an important role in the inflammatory response. Using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS)-based glycosylation profiling, we investigated the qualitative and quantitative effect of zymosan-induced peritonitis on N-glycosylation in mouse plasma and peritoneal fluid. Our results show that both N-glycomes exhibit highly similar glycosylation patterns, consisting mainly of diantennary and triantennary complex type N-glycans with high levels (>95 %) of galactosylation and sialylation (mostly NeuGc) and a medium degree of core fucosylation (30 %). Moreover, MS/MS structural analysis, assisted by linkage-specific derivatization of sialic acids, revealed the presence of O-acetylated sialic acids as well as disialylated antennae ("branching sialylation") characterized by the presence of α2-6-linked NeuGc on the GlcNAc of the NeuGcα2-3-Galβ1-3-GlcNAc terminal motif. A significant decrease of (core) fucosylation together with an increase of both α2-3-linked NeuGc and "branching sialylation" were observed in N-glycomes of mice challenged with zymosan, but not in control mice injected with PBS. Importantly, substantial changes in glycosylation were already observed 12 h after induction of peritonitis, thereby demonstrating an unexpected velocity of the biological mechanisms involved.Dutch Arthritis Association (Reumafonds) LLP-24 Innovative Medicines Initiative Joint Undertaking (IMI JU)/ 115142-2 Netherlands Genomic Initiative/93511033 info:eu-repo/grantAgreement/EC/FP7/278535info:eu-repo/grantAgreement/EC/FP7/27853

Do high molecular weight adiponectin levels associate with radiographic progression in early rheumatiod arthritis and hand osteoarthritis?

Pathophysiology and treatment of rheumatic disease

Interaction between extracellular matrix molecules and microbial pathogens: evidence for the missing link in autoimmunity with rheumatoid arthritis as a disease model.

Rheumatoid arthritis (RA) is an autoimmune disease characterized by inflammation followed by tissue rebuilding or fibrosis. A failure by the body to regulate inflammation effectively is one of the hallmarks of RA. The interaction between the external environment and the human host plays an important role in the development of autoimmunity. In RA, the observation of anti-cyclic citrullinated peptide antibodies (ACPA) to autoantigens is well recognized. Citrullination is a post-translational modification mediated by peptidyl arginine deiminases, which exist in both mammalian and bacterial forms. Previous studies have shown how proteins expressed in the human extracellular matrix (ECM) acquire properties of damage-associated molecular patterns (DAMPs) in RA and include collagens, tenascin-C, and fibronectin (FN). ECM DAMPs can further potentiate tissue damage in RA. Recent work has shown that citrullination in RA occurs at mucosal sites, including the oral cavity and lung. Mucosal sites have been linked with bacterial infection, e.g., periodontal disease, where exogenous pathogens are implicated in the development of autoimmunity via an infectious trigger. Proteases produced at mucosal sites, both by bacteria and the human host, can induce the release of ECM DAMPs, thereby revealing neoepitopes which can be citrullinated and lead to an autoantibody response with further production of ACPA. In this perspectives article, the evidence for the interplay between the ECM and bacteria at human mucosal surfaces, which can become a focus for citrullination and the development of autoimmunity, is explored. Specific examples, with reference to collagen, fibrinogen, and FN, are discussed

Association between weight or Body Mass Index and hand osteoarthritis: a systematic review

Objective: To investigate the association between weight or Body Mass Index (BMI) and the development of hand osteoarthritis (OA). Methods: Systematic review of observational studies. Medical databases were searched up to April 2008. Articles which presented data on the association between weight and hand OA were selected. The qualities of these studies were then assessed by two independent reviewers using a 19 criteria scoring syst

Fc gamma receptor binding profile of anti-citrullinated protein antibodies in immune complexes suggests a role for FcγRI in the pathogenesis of synovial inflammation

OBJECTIVES: Anti-citrullinated protein antibodies (ACPA) are highly specific for rheumatoid arthritis (RA). Here, we studied binding of ACPA-IgG immune complexes (IC) to individual Fc gamma receptors (FcγR) to identify potential effector mechanisms by which ACPA could contribute to RA pathogenesis. METHODS: ACPA-IgG1 and control IgG1(IgG1 depleted of ACPA-IgG1) were isolated from plasma and synovial fluid (SF) of RA patients by affinity chromatography using CCP2 peptides. Subsequently, IC were generated using fluorescently labelled F(ab’)2 fragments against the F(ab’)2 region of IgG, or by using citrullinated fibrinogen. IC were incubated with FcγR-transfected CHO cell lines or neutrophils from healthy donors. FcγR binding of IC was analysed by flow cytometry in the presence or absence of specific blocking antibodies. RESULTS: ACPA-IgG1 IC predominantly bound to FcγRI and FcγRIIIA on FcγR-transfected CHO cell lines, while much lower binding was observed to FcγRIIA and FcγRIIB. ACPA-IgG1 IC showed reduced binding to FcγRIIIA compared to control IgG1 IC, in line with enhanced ACPA-IgG1 Fc core-fucosylation. Neutrophils activated in vitro to induce de novo expression of FcγRI showed binding of ACPA-IgG IC, and blocking studies revealed that almost 30% of ACPA-IgG IC binding to activated neutrophils was mediated by FcγRI. CONCLUSIONS: Our studies show that ACPA-IgG1 IC bind predominately to activating FcγRI and FcγRIIIA, and highlight FcγRI expressed by activated neutrophils as relevant receptor for these IC. As neutrophils isolated from SF exhibit an activated state and express FcγRI in the synovial compartment, this IC-binding could contribute to driving disease pathogenesis in RA.Drug Delivery Technolog

Lipid metabolism of leukocytes in the unstimulated and activated states

Lipidomics has emerged as a powerful technique to study cellular lipid metabolism. As the lipidome contains numerous isomeric and isobaric species resulting in a significant overlap between different lipid classes, cutting-edge analytical technology is necessary for a comprehensive analysis of lipid metabolism. Just recently, differential mobility spectrometry (DMS) has evolved as such a technology, helping to overcome several analytical challenges. We here set out to apply DMS and the Lipidyzer (TM) platform to obtain a comprehensive overview of leukocyte-related lipid metabolism in the resting and activated states. First, we tested the linearity and repeatability of the platform by using HL60 cells. We obtained good linearities for most of the thirteen analyzed lipid classes (correlation coefficient > 0.95), and good repeatability (%CV < 15). By comparing the lipidome of neutrophils (PMNs), monocytes (CD14+), and lymphocytes (CD4+), we shed light on leukocyte-specific lipid patterns as well as lipidomic changes occurring through differential stimulation. For example, at the resting state, PMNs proved to contain higher amounts of triacylglycerides compared to CD4+ and CD14+ cells. On the other hand, CD4+ and CD14+ cells contained higher levels of phospholipids and ceramides. Upon stimulation, diacylglycerides, hexosylceramides, phosphatidylcholines, phosphoethanolamines, and lysophosphoethanolamines were upregulated in CD4+ cells and PMNs, whereas CD14+ cells did not show significant changes. By exploring the fatty acid content of the significantly upregulated lipid classes, we mainly found increased concentrations of very long and polyunsaturated fatty acids. Our results indicate the usefulness of the Lipidyzer (TM) platform for studying cellular lipid metabolism. Its application allowed us to explore the lipidome of leukocytes.Proteomic