Biomedical subjects

Wilco de Jager

Publications and source records attributed to Wilco de Jager.

10 recordsLinked to original sources

PubMed2006

Blood and synovial fluid cytokine signatures in patients with juvenile idiopathic arthritis: a cross-sectional study.

BACKGROUND: Juvenile idiopathic arthritis (JIA) consists of a heterogeneous group of disorders with, for the most part, an unknown immunopathogenesis. Although onset and disease course differ, the subtypes of JIA share the occurrence of chronic inflammation of the joints, with infiltrations of immunocompetent cells that secrete inflammatory mediators. OBJECTIVE: To identify a panel of cytokines specifically related to the inflammatory process in JIA. METHODS: Using a new technology, the multiplex immunoassay, 30 cytokines were measured in plasma of 65 patients with JIA, of which 34 were paired with synovial fluid. These data were compared with plasma of 20 healthy controls and 9 patients with type I diabetes, a chronic inflammatory disease. RESULTS: Patients with JIA had, irrespective of their subclassification, significantly higher levels of tumour necrosis factor alpha, macrophage inhibitory factor (MIF), CCL2, CCL3, CCL11, CCL22 and CXCL9 in plasma than controls. In paired plasma and synovial fluid samples of patients with JIA, significantly higher levels of interleukin (IL)6, IL15, CCL2, CCL3, CXCL8, CXCL9 and CXCL10 were present in synovial fluid. Cluster analysis in all patients with JIA revealed a predominant pro-inflammatory cytokine cluster during active disease and a regulatory/anti-inflammatory-related cytokine cluster during remission. Whether a discrimination profile of various cytokines could help in the determination of disease classification was tested. CONCLUSION: It is suggested that several cytokines (IL18, MIF, CCL2, CCL3, CCL11, CXCL9 and CXCL10) may correspond to the activation status during inflammation in JIA and could be instrumental in monitoring disease activity and outcomes of (new) immunotherapies.

Adolescent↗

PubMed2006

CSF protein profiling using Multiplex Immuno-assay: A potential new diagnostic tool for leptomeningeal metastases.

OBJECTIVE: The diagnosis of leptomeningeal metastases (LM) is based on clinical symptoms, magnetic resonance imaging (MRI) of brain and spine and cytological analysis of cerebrospinal fluid (CSF). The clinical picture of LM is highly variable and both cytological CSF analysis and contrast-enhanced MRI are limited in sensitivity. More sensitive tools are needed to diagnose LM. We measured a profile of proteins involved in adhesion and inflammation in the CSF of LM and control patients and determined their potential diagnostic value for LM. PATIENTS AND METHODS: Using Multiplex Immuno-Assay (MIA), the CSF concentrations of nine soluble adhesion molecules, cyto- and chemokines were measured in patients with cytologically proven LM (n = 57) and control patients with a systemic malignancy (n = 20), aseptic/viral meningitis (n = 11) or other (non-)neurological diseases (n = 19). RESULTS: We found high CSF levels of soluble Vascular Cell Adhesion Molecule-1 (sVCAM-1), soluble Intercellular Adhesion Molecule-1 (sICAM-1), Interleukin-8 (IL-8), Pulmonary and Activation Regulated Chemokine (PARC), Interleukin-18 (IL-18) and Interferon-gamma inducible protein (IP-10) in patients with LM. The CSF protein profile in LM patients differed significantly from the profile found in control patients. Multivariate logistic regression and ROC analysis showed that the MIA-measured CSF protein profile has an additive discriminating value for LM above standard CSF parameters. A combination of total protein, glucose, IL-8, PARC and IP-10 CSF levels proved to be most discriminative between LM and non-LM patients. CONCLUSION: Our results warrant a prospective study to determine whether a CSF protein profile, including IL-8, PARC and IP-10 has diagnostic value compared with CSF cytology, the golden standard for LM.

Adult↗

PubMed2006

Novel pan-DR-binding T cell epitopes of adenovirus induce pro-inflammatory cytokines and chemokines in healthy donors.

Adenovirus can cause fatal infections in the immunocompromised host. To date, no effective anti-viral therapy is available. Adoptive therapy with adenovirus-specific T cells could be a promising treatment, but requires the identification of such T cells. Aim of this study was to identify conserved adenoviral T cell epitopes recognized in a majority of healthy individuals. By using a computer algorithm designed to predict pan-HLA-DR-binding T cell epitopes, we selected 19 peptides of adenovirus serotype 5. PBMCs from 26 healthy subjects were isolated and incubated with these peptides to test epitope-specific T cell proliferation. Six epitopes derived from E1B protein, hexon protein (two epitopes), DNA polymerase, E3A glycoprotein and fiber protein induced a proliferative T cell response in the majority of healthy controls. In vitro MHC binding assays confirmed the potential capacity of the adenovirus epitopes to bind multiple MHC alleles. The cytokine and chemokine profile induced by these epitopes was determined with a multiplex immunoassay and revealed a predominant pro-inflammatory pattern. Based on the broad recognition and the induced cytokine and chemokine profile, the detected epitopes can be regarded as potential candidates to select adenovirus-specific T cells for immune intervention in the immunocompromised host.

Adenoviridae↗

PubMed2006

Expression of the inflammatory chemokines CCL5, CCL3 and CXCL10 in juvenile idiopathic arthritis, and demonstration of CCL5 production by an atypical subset of CD8+ T cells.

This study focuses upon three chemokines, namely CCL5, CXCL10 and CCL3, which are potential novel therapeutic targets in arthritis. The aim of the study was to analyse the expression and production of these three chemokines within the joints of children with juvenile idiopathic arthritis (JIA) of the oligoarticular and polyarticular subtypes. All three of these chemokines are highly expressed at the level of mRNA, with the most significant increase in mRNA levels being demonstrated for CCL5 when compared with matched peripheral blood samples and controls. We show that high levels of all three chemokines are present in synovial fluid of children with JIA. We investigate the major source of CCL5 from inflammatory synovial cells, which we show to be CD8+ T cells. This CD8+ synovial T cell population has an unexpected phenotype that has not been described previously, being CCR7- yet predominantly CD28+ and CD45RA-. These cells contain high levels of stored intracellular CCL5, and rapid release of CCL5 takes place on T cell stimulation, without requiring new protein synthesis. In addition, we demonstrate that CCL5 is present in synovial biopsies from these patients, in particular on the endothelium of small and medium sized vessels. We believe this to be the first in depth analysis of these mediators of inflammation in JIA.

Adult↗

PubMed2006

Solid-phase and bead-based cytokine immunoassay: a comparison.

Cytokines and chemoattractive cytokines (chemokines) are present in a wide variety of body fluids such as plasma, cerebrospinal fluid, bronchoaveolar fluid, amniotic fluid, synovial fluid, middle ear effusion fluid, and urine. Cytokines can be detected using classical solid-phase sandwich immunoassays such as enzyme-linked immunosorbent assay (ELISA) or with a bead based multiplex immunoassay (MIA). The physical chemical properties of the different body fluids (such as pH and total protein content) differ, which may have an impact on the outcome of the cytokine assay. Both ELISA as well as MIA cytokine detection systems are constructed by sandwiching the protein of interest between a capture and reporter antibody. When the biological sample contains heterophilic antibodies (such as in patients with auto-immune diseases), these non-specific antibodies can cause false positive results. During pathological conditions, cytokines may be found over a wide concentration range; likewise have to cover this dynamic range in a similar fashion. The correct (statistical) analysis of standard curves and (multiplexed) data are critical for proper interpretation. Classical ELISA based cytokine assays are robust, easy to use and very well suited for measurement of single cytokines. Due to an increased interest in the integral approach to understand biological processes (the omics era), multiplex immunoassays for detection of cytokines and the interpretation of these assays are gaining popularity.

Animals↗

PubMed2006

Novel self-epitopes derived from aggrecan, fibrillin, and matrix metalloproteinase-3 drive distinct autoreactive T-cell responses in juvenile idiopathic arthritis and in health.

Juvenile idiopathic arthritis (JIA) is a heterogeneous autoimmune disease characterized by chronic joint inflammation. Knowing which antigens drive the autoreactive T-cell response in JIA is crucial for the understanding of disease pathogenesis and additionally may provide targets for antigen-specific immune therapy. In this study, we tested 9 self-peptides derived from joint-related autoantigens for T-cell recognition (T-cell proliferative responses and cytokine production) in 36 JIA patients and 15 healthy controls. Positive T-cell proliferative responses (stimulation index > or =2) to one or more peptides were detected in peripheral blood mononuclear cells (PBMC) of 69% of JIA patients irrespective of major histocompatibility complex (MHC) genotype. The peptides derived from aggrecan, fibrillin, and matrix metalloproteinase (MMP)-3 yielded the highest frequency of T-cell proliferative responses in JIA patients. In both the oligoarticular and polyarticular subtypes of JIA, the aggrecan peptide induced T-cell proliferative responses that were inversely related with disease duration. The fibrillin peptide, to our knowledge, is the first identified autoantigen that is primarily recognized in polyarticular JIA patients. Finally, the epitope derived from MMP-3 elicited immune responses in both subtypes of JIA and in healthy controls. Cytokine production in short-term peptide-specific T-cell lines revealed production of interferon-gamma (aggrecan/MMP-3) and interleukin (IL)-17 (aggrecan) and inhibition of IL-10 production (aggrecan). Here, we have identified a triplet of self-epitopes, each with distinct patterns of T-cell recognition in JIA patients. Additional experiments need to be performed to explore their qualities and role in disease pathogenesis in further detail.

Age of Onset↗

PubMed2005

Improved multiplex immunoassay performance in human plasma and synovial fluid following removal of interfering heterophilic antibodies.

Cytokines, chemokines and soluble adhesion molecules interact in a complex network within the immune system. Fingerprinting of these proteins may allow the use of these proteins as biomarkers for identification of disease, disease subtyping and monitoring therapeutic interventions. We developed a multiplex immunoassay (MIA) for the detection of 30 proteins in a variety of human body fluids such as plasma and synovial fluid (SF). The measurement of these proteins is hampered by the presence of human (auto-) antibodies, which can cause non-specific binding. We have validated a novel approach for the removal of interfering immunoglobulins using pre-absorption with protein-L. Interfering (auto-) antibodies, such as rheumatoid factor (RF), were removed using three methods; polyethylene glycol (PEG) precipitation, pre-absorption with human gamma-globulin or pre-absorption with protein-L. A significant decrease of RF was observed after a 2 h incubation with protein-L. RF IgM levels were reduced by 89% whereas total IgM, IgG and IgA levels were reduced by 60%. Residual immunoglobulins were blocked with rodent serum and did not interfere with the multiplex immunoassay. Comparing the MIA with a conventional enzyme-linked immunosorbent assay (ELISA) using a panel of spiked plasma samples resulted in correlation coefficients for all mediators between R2 = 0.88 and R2 = 0.99. Intra-assay variance was less than 10% whereas inter-assay variance ranged between 6% and 16%. Pathological samples with heterophilic antibodies hamper immunoassays such as ELISA and MIA. We show that pre-absorption with protein-L is a powerful tool for removal of interfering immunoglobulins from human bodily fluids to be used in immunoassays for studying changes in protein patterns.

Antibodies, Heterophile↗

PubMed2004

CD4+CD25bright regulatory T cells actively regulate inflammation in the joints of patients with the remitting form of juvenile idiopathic arthritis.

This study investigates the role of CD4(+)CD25(+) regulatory T cells during the clinical course of juvenile idiopathic arthritis (JIA). Persistent oligoarticular JIA (pers-OA JIA) is a subtype of JIA with a relatively benign, self-remitting course while extended oligoarticular JIA (ext-OA JIA) is a subtype with a much less favorable prognosis. Our data show that patients with pers-OA JIA display a significantly higher frequency of CD4(+)CD25(bright) T cells with concomitant higher levels of mRNA FoxP3 in the peripheral blood than ext-OA JIA patients. Furthermore, while numbers of synovial fluid (SF) CD4(+)CD25(bright) T cells were equal in both patient groups, pers-OA JIA patients displayed a higher frequency of CD4(+)CD25(int) T cells and therefore of CD4(+)CD25(total) in the SF than ext-OA JIA patients. Analysis of FoxP3 mRNA levels revealed a high expression in SF CD4(+)CD25(bright) T cells of both patient groups and also significant expression of FoxP3 mRNA in the CD4(+)CD25(int) T cell population. The CD4(+)CD25(bright) cells of both patient groups and the CD4(+)CD25(int) cells of pers-OA JIA patients were able to suppress responses of CD25(neg) cells in vitro. A markedly higher expression of CTLA-4, glucocorticoid-induced TNFR, and HLA-DR on SF CD4(+)CD25(bright) T regulatory (Treg) cells compared with their peripheral counterparts suggests that the CD4(+)CD25(+) Treg cells may undergo maturation in the joint. In correlation with this mature phenotype, the SF CD4(+)CD25(bright) T cells showed an increased regulatory capacity in vitro compared with peripheral blood CD4(+)CD25(bright) T cells. These data suggest that CD4(+)CD25(bright) Treg cells play a role in determining the patient's fate toward either a favorable or unfavorable clinical course of disease.

Adolescent↗

PubMed2003

Simultaneous detection of 15 human cytokines in a single sample of stimulated peripheral blood mononuclear cells.

Cytokines secreted by cells of the immune system can alter the behavior and properties of immune or other cells. At a site of inflammation, sets of cytokines interact with immune cells, and their combined effect is often more important than the function of one isolated component. Conventional techniques, such as enzyme-linked immunosorbent assays, generally require large quantities of cells to characterize a complete cytokine profile of activated lymphocytes. The Bio-Plex system from Bio-Rad Laboratories combines the principle of a sandwich immunoassay with the Luminex fluorescent-bead-based technology. We developed a multiplex cytokine assay to detect different cytokines simultaneously in culture supernatant of human peripheral blood mononuclear cells stimulated with antigen and with mitogen. Fifteen human cytokines (interleukin 1alpha [IL-1alpha], IL-1beta, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-13, IL-15, IL-17, IL-18, gamma interferon, and tumor necrosis factor alpha) were validated with a panel of healthy individuals, rheumatoid arthritis patients, and juvenile idiopathic arthritis patients. Comparing the multiplex assay with a regular enzyme-linked immunosorbent assay technique with this donor panel resulted in correlation coefficients for all cytokines ranging from 0.75 to 0.99. Intra-assay variance proved to be less then 10%, whereas interassay variability ranged between 10 and 22%. This multiplex system proved to be a powerful tool in the quantitation of cytokines. It will provide a more complete picture in differences between activated lymphocyte cytokine profiles from healthy individuals and those from patients with chronic inflammatory diseases.

Arthritis, Juvenile↗

PubMedDate not supplied

Tolerogenic immune responses to novel T-cell epitopes from heat-shock protein 60 in juvenile idiopathic arthritis.

BACKGROUND: Juvenile idiopathic arthritis is a heterogeneous autoimmune disease characterised by chronic inflammation of one or more joints. In patients with this disease, T-cell reactivity to autologous heat-shock protein 60 (HSP60) is associated with a favourable prognosis. We sought to identify HSP60 T-cell epitopes to find potential targets for HSP60 immunotherapy and to assess whether immune responses to these epitopes contribute to the distinct clinical outcome of this disease. METHODS: We identified eight potential epitopes using a computer algorithm from both self and microbial HSP60 binding to many HLA-DR molecules. We analysed the pattern of T-cell responses induced by these HSP60 peptides in peripheral-blood mononuclear cells (PBMC) of 57 patients with juvenile idiopathic arthritis, 27 healthy controls, and 20 disease controls. We undertook in-vitro MHC binding studies with the identified peptides, and HLA class II typing of a subset of patients with juvenile idiopathic arthritis. FINDINGS: Five of the eight peptides identified yielded proliferative T-cell responses in 50-70% of PBMC from patients with juvenile idiopathic arthritis irrespective of MHC genotype, but not in PBMC from healthy or disease controls. Although PBMC from both patients with juvenile idiopathic arthritis and healthy controls produced interferon gamma in response to these peptides, only PBMC from patients with the disease produced interleukin 10. INTERPRETATION: The recorded T-cell-induction in juvenile idiopathic arthritis is tolerogenic. In patients with oligoarticular disease, the immune responses to the HSP60 epitopes identified could contribute to disease remission. RELEVANCE TO PRACTICE: The broad recognition of these HSP60 epitopes in a population of patients with polymorphic MHC genotypes opens the way for HSP60-peptide immunotherapy, representing a novel treatment option to specifically modulate the immune system in patients with juvenile idiopathic arthritis.

Adolescent↗