ABSTRACT
The T-cell receptor (TCR) is a highly polymorphic surface receptor that allows T-cells to recognize antigenic peptides presented on the major histocompatibility complex (MHC). Changes in the TCR repertoire have been observed in several autoimmune conditions, and these changes are suggested to predispose autoimmunity. Multiple lines of evidence have implied an important role for T-cells in the pathogenesis of Systemic Sclerosis (SSc), a complex autoimmune disease. One of the major questions regarding the roles of T-cells is whether expansion and activation of T-cells observed in the diseases pathogenesis is antigen driven. To investigate the temporal TCR repertoire dynamics in SSc, we performed high-throughput sequencing of CD4+ and CD8+ TCRß chains on longitudinal samples obtained from four SSc patients collected over a minimum of two years. Repertoire overlap analysis revealed that samples taken from the same individual over time shared a high number of TCRß sequences, indicating a clear temporal persistence of the TCRß repertoire in CD4+ as well as CD8+ T-cells. Moreover, the TCRßs that were found with a high frequency at one time point were also found with a high frequency at the other time points (even after almost four years), showing that frequencies of dominant TCRßs are largely consistent over time. We also show that TCRß generation probability and observed TCR frequency are not related in SSc samples, showing that clonal expansion and persistence of TCRßs is caused by antigenic selection rather than convergent recombination. Moreover, we demonstrate that TCRß diversity is lower in CD4+ and CD8+ T-cells from SSc patients compared with memory T-cells from healthy individuals, as SSc TCRß repertoires are largely dominated by clonally expanded persistent TCRß sequences. Lastly, using "Grouping of Lymphocyte Interactions by Paratope Hotspots" (GLIPH2), we identify clusters of TCRß sequences with homologous sequences that potentially recognize the same antigens and contain TCRßs that are persist in SSc patients. In conclusion, our results show that CD4+ and CD8+ T-cells are highly persistent in SSc patients over time, and this persistence is likely a result from antigenic selection. Moreover, persistent TCRs form high similarity clusters with other (non-)persistent sequences that potentially recognize the same epitopes. These data provide evidence for an antigen driven expansion of CD4+/CD8+ T-cells in SSc.
Subject(s)
CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/metabolism , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/metabolism , Receptors, Antigen, T-Cell/genetics , Scleroderma, Systemic/etiology , Scleroderma, Systemic/metabolism , Adult , Antigens/immunology , Disease Susceptibility , Epitopes , Female , Gene Frequency , High-Throughput Nucleotide Sequencing , Humans , Immunologic Memory , Immunophenotyping , Longitudinal Studies , Lymphocyte Activation , Male , Middle Aged , Receptors, Antigen, T-Cell/metabolism , Receptors, Antigen, T-Cell, alpha-beta/genetics , Receptors, Antigen, T-Cell, alpha-beta/metabolism , Scleroderma, Systemic/pathologyABSTRACT
Systemic sclerosis (SSc) is a rare chronic disease of unknown pathogenesis characterized by fibrosis of the skin and internal organs, vascular alteration, and dysregulation of the immune system. In order to better understand the immune system and its perturbations leading to diseases, the study of the mechanisms regulating cellular metabolism has gained a widespread interest. Here, we have assessed the metabolic status of plasma and dendritic cells (DCs) in patients with SSc. We identified a dysregulated metabolomic signature in carnitine in circulation (plasma) and intracellularly in DCs of SSc patients. In addition, we confirmed carnitine alteration in the circulation of SSc patients in three independent plasma measurements from two different cohorts and identified dysregulation of fatty acids. We hypothesized that fatty acid and carnitine alterations contribute to potentiation of inflammation in SSc. Incubation of healthy and SSc dendritic cells with etoposide, a carnitine transporter inhibitor, inhibited the production of pro-inflammatory cytokines such as IL-6 through inhibition of fatty acid oxidation. These findings shed light on the altered metabolic status of the immune system in SSc patients and opens up for potential novel avenues to reduce inflammation.
Subject(s)
Carnitine/blood , Fatty Acids/blood , Scleroderma, Systemic/blood , Adult , Aged , Cohort Studies , Cytokines/metabolism , Dendritic Cells/metabolism , Etoposide/pharmacology , Female , Fibrosis/genetics , Gene Expression/drug effects , Humans , Inflammation/genetics , Inflammation/immunology , Inflammation/metabolism , Male , Metabolome , Metabolomics/methods , Middle Aged , Organic Cation Transport Proteins/antagonists & inhibitors , Oxidation-Reduction/drug effects , Scleroderma, Systemic/immunology , Signal Transduction/drug effectsABSTRACT
Systemic sclerosis (SSc) is a severe autoimmune fibrotic disease characterized by fibrosis, vasculopathy, and immune dysregulation. Dendritic cells (DCs) are the most potent antigen-presenting cells, specialized in pathogen sensing, with high capacity to shape the immune responses. The most recent technological advances have allowed the discovery of new DC subsets with potential implications in inflammatory conditions. Alterations of DC distribution in circulation and affected tissue as well as impaired DC function have been described in SSc patients, pointing towards a crucial role of these cells in SSc pathogenesis. In particular, recent studies have shown the importance of plasmacytoid DCs either by their high capacity to produce type I interferon or other inflammatory mediators implicated in SSc pathology, such as chemokine C-X-C motif ligand 4 (CXCL4). In-vivo models of SSc have been vital to clarify the implications of DCs in this disease, especially DCs depletion and specific gene knock-down studies. This review provides these new insights into the contribution of the different DCs subsets in the pathogenesis of SSc, as well as to the novel developments on DCs in in-vivo models of SSc and the potential use of DCs and their mediators as therapeutic targets.
Subject(s)
Dendritic Cells/immunology , Scleroderma, Systemic/immunology , Animals , Dendritic Cells/pathology , Disease Models, Animal , Humans , Platelet Factor 4/genetics , Platelet Factor 4/immunology , Scleroderma, Systemic/genetics , Scleroderma, Systemic/pathologyABSTRACT
BACKGROUND: Localized Scleroderma (LoS) encompasses a group of idiopathic skin conditions characterized by (sub)cutaneous inflammation and subsequent development of fibrosis. Currently, lack of accurate tools enabling disease activity assessment leads to suboptimal treatment approaches. OBJECTIVE: To investigate serum concentrations of cytokines and chemokines implicated in inflammation and angiogenesis in LoS and explore their potential to be utilized as biomarker of disease activity. Additionally, to investigate the implication of potential biomarkers in disease pathogenesis. METHODS: A 39-plex Luminex immuno-assay was performed in serum samples of 74 LoS and 22 Healthy Controls. The relation between a validated clinical measure of disease activity (mLoSSI) and serum analytes was investigated. Additionally, gene and protein expression were investigated in circulating cells and skin biopsies. RESULTS: From the total of 39, 10 analytes (CCL18, CXCL9, CXCL10, CXCL13, TNFRII, Galectin-9, TIE-1, sVCAM, IL-18, CCL19) were elevated in LoS serum. Cluster analysis of serum samples revealed CCL18 as most important analyte to discriminate between active and inactive disease. At individual patient level, CCL18 serum levels correlated strongest with mLoSSI-scores (rsâ¯=â¯0.4604, Pâ¯<â¯0.0001) and in longitudinal measures CCL18 concentrations normalised with declining disease activity upon treatment initiation. Additionally, CCL18 was elevated in LoS serum, and not in (juvenile) dermatomyositis or spinal muscular atrophy. Importantly, CCL18 gene and protein expression was increased at the inflammatory border of cutaneous LoS lesions, with normal expression in unaffected skin and circulating immune cells. CONCLUSION: CCL18 is specific for disease activity in LoS thereby providing relevance as a biomarker for this debilitating disease.
Subject(s)
Biomarkers , Chemokines, CC/metabolism , Scleroderma, Localized/metabolism , Biopsy , Chemokines/metabolism , Chemokines, CC/blood , Chemokines, CC/genetics , Cytokines/metabolism , Disease Susceptibility , Gene Expression , Gene Expression Profiling , Humans , Scleroderma, Localized/diagnosis , Scleroderma, Localized/etiology , Scleroderma, Localized/therapy , Severity of Illness Index , Skin TestsABSTRACT
Lung transplantation (LTx) is the last treatment for patients suffering from end-stage lung diseases. Survival post-LTx is hampered by the development of the bronchiolitis obliterans syndrome (BOS) and diagnosis is often late. Given the urgent clinical need to recognize BOS patients at an early stage, we analyzed circulating miRNAs to identify possible stratification markers for BOS development post-transplantation. Therefore, pro-fibrotic (miR-21, miR-155), anti-fibrotic (miR-29a) and fibrosis-unrelated (miR-103, miR-191) miRNAs were analyzed in serum of end-stage lung disease patients and during LTx follow-up. Significant elevated levels of serum miRNAs were observed for all investigated miRNAs in both chronic obstructive pulmonary disease and interstitial lung disease patients compared to healthy controls. The same miRNAs were also significantly increased in the serum of BOS+ vs. BOS- patients. Most importantly, miR-21, miR-29a, miR-103, and miR-191 levels were significantly higher in BOS+ patients prior to clinical BOS diagnosis. We demonstrated that a selected group of miRNAs investigated is elevated in end-stage lung disease and BOS+ patients, prior to clinical BOS diagnosis. Even if further research is expedient on the prognostic value of circulating miRNAs in BOS and lung conditions in general, these results strongly suggest that circulating miRNAs could be used as potential biomarkers for BOS development.
Subject(s)
Bronchiolitis Obliterans/blood , Lung Transplantation , MicroRNAs/blood , Adult , Biomarkers/blood , Female , Follow-Up Studies , Humans , Male , Middle Aged , Pulmonary Disease, Chronic Obstructive/blood , Retrospective StudiesABSTRACT
AIM AND BACKGROUND: Chronic inflammation associates with increased senescence, which is a strong predictor for cardiovascular disease. We hypothesised that inflammation accelerates senescence and thereby enhances the risk of cardiovascular disease in gout. METHODS: We assessed replicative senescence by quantifying telomere length (TL) in a discovery cohort of 145 Dutch patients with gout and 273 healthy individuals and validated our results in 474 patients with gout and 293 healthy participants from New Zealand. Subsequently, we investigated the effect of cardiovascular disease on TL of all participants. Also, we measured TL of CD4+ and CD8+ T lymphocytes, B lymphocytes, monocytes, natural killer cells and plasmacytoid dendritic cells. Additionally, we assessed the potential temporal difference in TL and telomerase activity. RESULTS: TL in PBMCs of healthy donors decreased over time, reflecting normal ageing. Patients with gout demonstrated shorter telomeres (p=0.001, R2=0.01873). In fact, the extent of telomere erosion in patients with gout was higher at any age compared with healthy counterparts at any age (p<0.0001, R2=0.02847). Patients with gout with cardiovascular disease had the shortest telomeres and TL was an independent risk factor for cardiovascular disease in patients with gout (p=0.001). TL was inversely associated with the number of gouty flares (p=0.005). CONCLUSIONS: Patients with gout have shorter telomeres than healthy participants, reflecting increased cellular senescence. Telomere shortening was associated with the number of flares and with cardiovascular disease in people with gout.
Subject(s)
Cardiovascular Diseases/metabolism , Gout/metabolism , Telomerase/genetics , Telomere/metabolism , Adult , Aged , Aged, 80 and over , B-Lymphocytes/metabolism , CD4-Positive T-Lymphocytes/metabolism , CD8-Positive T-Lymphocytes/metabolism , Cardiovascular Diseases/epidemiology , Case-Control Studies , Dendritic Cells/metabolism , Disease Progression , Female , Gout/epidemiology , Humans , Killer Cells, Natural/metabolism , Linear Models , Male , Middle Aged , Monocytes/metabolismABSTRACT
Splenic macrophages play a key role in immune thrombocytopenia (ITP) pathogenesis by clearing opsonized platelets. Fcγ receptors (FcγR) participate in this phenomenon, but their expression on splenic macrophages and their modulation by treatment have scarcely been studied in human ITP. We aimed to compare the phenotype and function of splenic macrophages between six controls and 24 ITP patients and between ITP patients according to the treatments they received prior to splenectomy. CD86, human leucocyte antigen D-related (HLA-DR) and FcγR expression were measured by flow cytometry on splenic macrophages. The major FcγR polymorphisms were determined and splenic macrophage function was assessed by a phagocytosis assay. The expression of the activation markers CD86 and HLA-DR was higher on splenic macrophages during ITP compared to controls. While the expression of FcγR was not different between ITP and controls, the phagocytic function of splenic macrophages was reduced in ITP patients treated with intravenous immunoglobulin (IVIg) within the 2 weeks prior to splenectomy. The FCGR3A (158V/F) polymorphism, known to increase the affinity of FcγRIII to IgG, was over-represented in ITP patients. Thus, these are the first results arguing for the fact that the therapeutic use of IVIg during human chronic ITP does not modulate FcγR expression on splenic macrophages but decreases their phagocytic capabilities.
Subject(s)
Autoimmune Diseases/immunology , Macrophages/immunology , Receptors, IgG/analysis , Receptors, IgG/genetics , Spleen/immunology , Thrombocytopenia/immunology , Adult , Aged , Autoimmune Diseases/surgery , Autoimmune Diseases/therapy , B7-2 Antigen/analysis , Female , Flow Cytometry , Humans , Immunoglobulin G/blood , Immunoglobulins, Intravenous/therapeutic use , Macrophages/physiology , Male , Middle Aged , Phagocytosis , Phenotype , Polymorphism, Genetic , Receptors, IgG/immunology , Spleen/cytology , Splenectomy , Thrombocytopenia/surgery , Thrombocytopenia/therapyABSTRACT
Retinal diseases generally are vision-threatening conditions that warrant appropriate clinical decision-making which currently solely dependents upon extensive clinical screening by specialized ophthalmologists. In the era where molecular assessment has improved dramatically, we aimed at the identification of biomarkers in 175 ocular fluids to classify four archetypical ocular conditions affecting the retina (age-related macular degeneration, idiopathic non-infectious uveitis, primary vitreoretinal lymphoma, and rhegmatogenous retinal detachment) with one single test. Unsupervised clustering of ocular proteins revealed a classification strikingly similar to the clinical phenotypes of each disease group studied. We developed and independently validated a parsimonious model based merely on three proteins; interleukin (IL)-10, IL-21, and angiotensin converting enzyme (ACE) that could correctly classify patients with an overall accuracy, sensitivity and specificity of respectively, 86.7%, 79.4% and 92.5%. Here, we provide proof-of-concept for molecular profiling as a diagnostic aid for ophthalmologists in the care for patients with retinal conditions.
Subject(s)
Eye Proteins/metabolism , Retinal Diseases/diagnosis , Retinal Diseases/metabolism , Adult , Aged , Aged, 80 and over , Algorithms , Aqueous Humor/metabolism , Biomarkers , Clinical Decision-Making , Cluster Analysis , Computational Biology/methods , Female , Humans , Male , Middle Aged , Proteome , Proteomics/methods , Reproducibility of Results , Sensitivity and SpecificitySubject(s)
Autoantibodies/immunology , Gene Expression Regulation , Receptors, Interleukin-7/genetics , Sjogren's Syndrome/genetics , Sjogren's Syndrome/pathology , Adult , Aged , Animals , Autoantibodies/blood , Biopsy, Needle , Case-Control Studies , Enzyme-Linked Immunosorbent Assay , Female , Humans , Immunohistochemistry , Male , Middle Aged , Receptors, Interleukin-7/metabolism , Reference Values , Retrospective Studies , Severity of Illness Index , Sjogren's Syndrome/immunology , Solubility , Statistics, Nonparametric , Young AdultABSTRACT
OBJECTIVES: Natural killer cell receptors (NKR) have been implicated in rheumatoid (RA) and psoriatic arthritis (PsA) pathogenesis. To gain more insight into their role, we characterised NKR (co-)expression patterns on NK and T cells and NK cell function in RA and PsA. METHODS: The frequency of NK and T cells expressing killer like immunoglobulin (KIR) and NKG2 receptors and natural cytotoxicity receptors was assessed by 10-colour flow cytometry in peripheral blood of 23 RA, 12 PsA patients and 18 healthy donors (HD). NK cell cytotoxicity and IFN-gamma production was assessed in 8 RA patients and 8 HD. RESULTS: In RA but not PsA, the frequency of NK cells (median; range) expressing NKG2A (42%; 14-81%) was elevated compared to HD (23%; 9-58%). NKG2A⺠NK cells predominantly lack KIR, but display normal cytotoxicity and IFN-γ production. In contrast, RA patients with normal NKG2A⺠NK cell frequency have less functional NK cells compared to HD. T cells expressing Fc-gamma receptor CD16 were elevated in RA (median 0.75%) versus HD (0.3%). Furthermore, T cells expressing the KIRs CD158ah in both RA (0.7%) and PsA (0.3%), and CD158e1e2 in RA (1.5%) were elevated compared to HD (0.2% and 0.4%, respectively). In RA, CD4⺠T cells expressing the KIRs CD158ah, CD158b1b2j and CD158e1e2 were low (<2%) but significantly elevated compared to HD. CONCLUSIONS: This study demonstrates the presence of an elevated, functionally active NKG2A⺠KIR- NK cell population in RA. Together with an elevated frequency of NKR-expressing T cells, these changes may reflect differential pathogenetic involvement.
Subject(s)
Arthritis, Psoriatic , Arthritis, Rheumatoid , Killer Cells, Natural/immunology , NK Cell Lectin-Like Receptor Subfamily C/immunology , Adult , Aged , Arthritis, Psoriatic/immunology , Arthritis, Rheumatoid/immunology , Female , Humans , Immunity, Cellular , Lymphocyte Count , Male , Middle Aged , Receptors, KIR/immunology , Receptors, Natural Cytotoxicity Triggering/immunologyABSTRACT
Autoimmune muscle diseases (myositis) comprise a group of complex phenotypes influenced by genetic and environmental factors. To identify genetic risk factors in patients of European ancestry, we conducted a genome-wide association study (GWAS) of the major myositis phenotypes in a total of 1710 cases, which included 705 adult dermatomyositis, 473 juvenile dermatomyositis, 532 polymyositis and 202 adult dermatomyositis, juvenile dermatomyositis or polymyositis patients with anti-histidyl-tRNA synthetase (anti-Jo-1) autoantibodies, and compared them with 4724 controls. Single-nucleotide polymorphisms showing strong associations (P<5×10(-8)) in GWAS were identified in the major histocompatibility complex (MHC) region for all myositis phenotypes together, as well as for the four clinical and autoantibody phenotypes studied separately. Imputation and regression analyses found that alleles comprising the human leukocyte antigen (HLA) 8.1 ancestral haplotype (AH8.1) defined essentially all the genetic risk in the phenotypes studied. Although the HLA DRB1*03:01 allele showed slightly stronger associations with adult and juvenile dermatomyositis, and HLA B*08:01 with polymyositis and anti-Jo-1 autoantibody-positive myositis, multiple alleles of AH8.1 were required for the full risk effects. Our findings establish that alleles of the AH8.1 comprise the primary genetic risk factors associated with the major myositis phenotypes in geographically diverse Caucasian populations.
Subject(s)
Alleles , HLA Antigens/genetics , Myositis/genetics , Adolescent , Adult , Autoantibodies/immunology , Case-Control Studies , Dermatomyositis/genetics , Female , Genetic Association Studies , Genetic Predisposition to Disease , Genome-Wide Association Study , Haplotypes , Humans , Male , Middle Aged , Polymorphism, Single Nucleotide , Polymyositis/genetics , Risk Factors , White PeopleABSTRACT
BACKGROUND: Localized scleroderma (LoS) is characterized by a phase of disease activity followed by remission. However, disease recurrences occur. Knowledge concerning these recurrences can help prompt treatment, thereby preventing disease damage. OBJECTIVES: To investigate the frequency and characteristics of disease recurrences in paediatric- and adult-onset LoS, and to identify patient variables that are associated with a higher risk of disease recurrence. METHODS: Retrospective chart reviews were performed of patients with LoS. Data concerning the frequency and characteristics of the disease recurrences were collected. A multivariate analysis was performed to identify patient variables that were associated with a higher risk of disease recurrence. RESULTS: In total, 344 patients were included in the analysis, of whom 119 (35%) had paediatric-onset LoS and 225 (65%) had adult-onset LoS. Disease recurrence was present in 27% (n = 32) of the paediatric-onset group and 17% (n = 39) of the adult-onset group (P = 0·037). Multivariate analysis identified a statistically significant association between disease recurrence and the linear LoS of the limbs subtype, independent of age at disease onset. CONCLUSIONS: Recurrences in LoS occurred in almost one-quarter of the patients and were most frequent in the linear LoS of the limbs subtype, independent of age at disease onset. These disease recurrences can occur even after many years of quiescent disease. Awareness of the high recurrence rates may help treating physicians to recognize reactivation of the disease, leading to a decreased delay in treatment reinitiation.
Subject(s)
Scleroderma, Localized/therapy , Adolescent , Adult , Aged , Aged, 80 and over , Child , Child, Preschool , Female , Humans , Infant , Male , Middle Aged , Recurrence , Remission Induction/methods , Retrospective Studies , Risk Factors , Time-to-Treatment , Young AdultABSTRACT
BACKGROUND: Systemic sclerosis (SSc) is an autoimmune disease characterised by fibrosis of the skin and the internal organs. Except for anticentromere, antitopoisomerase I and antipolymerase III antibodies, there are no reliable circulating markers predicting susceptibility and internal organ complications. This study has exploited a proteome-wide profiling method with the aim to identify new markers to identify SSc phenotype. METHOD: 40 SSc patients were included for proteomic identification. Patients were stratified as having diffuse cutaneous SSc (dcSSc) (n=19) or limited cutaneous SSc (lcSSc) (n=21) according to the extent of skin involvement. As controls 19 healthy donors were included. Blood was drawn and plasma was stored before analysing with the SELDI-TOF-MS. For replication in serum, the cohort was extended with 60 SSc patients. RESULTS: Proteomic analysis revealed a list of 25 masspeaks that were differentially expressed between SSc patients and healthy controls. One of the peaks was suggestive for S100A8, a masspeak we previously found in supernatant of plasmacytoid dendritic cells from SSc patients. Increased expression of S100A8/A9 in SSc patients was confirmed in replication cohort compared with controls. Intriguingly, S100A8/A9 was highest in patients with limited cutaneous SSc having lung fibrosis. CONCLUSIONS: S100A8/A9 was robustly found to be elevated in the circulation of SSc patients, suggesting its use as a biomarker for SSc lung disease and the need to further explore the role of TLR in SSc.
Subject(s)
Calgranulin A/metabolism , Calgranulin B/metabolism , Proteomics , Scleroderma, Systemic/metabolism , Toll-Like Receptors/metabolism , Adult , Aged , Biomarkers/metabolism , Calgranulin A/immunology , Calgranulin B/immunology , Female , Humans , Male , Middle Aged , Phenotype , Prospective Studies , Pulmonary Fibrosis/immunology , Pulmonary Fibrosis/metabolism , Scleroderma, Systemic/immunology , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Toll-Like Receptors/immunologyABSTRACT
There is increasing evidence that gene copy number (CN) variation influences clinical phenotype. The low-affinity Fc receptor 3B (FCGR3B) located in the FCGR gene cluster is a CN polymorphic gene involved in the recruitment of polymorphonuclear neutrophils to sites of inflammation and their activation. Given the genetic overlap between systemic lupus erythematosus and systemic sclerosis (SSc) and the strong evidence for FCGR3B CN in the pathology of SLE, we hypothesised that FCGR3B gene dosage influences susceptibility to SSc. We obtained FCGR3B deletion status in 777 European Caucasian cases and 1000 controls. There was an inverse relationship between FCGR3B CN and disease susceptibility. CN of ≤ 1 was a significant risk factor for SSc (OR=1.55 (1.13-2.14), P=0.007) relative to CN ≥ 2. Although requiring replication, these results suggest that impaired immune complex clearance arising from FCGR3B deficiency contributes to the pathology of SSc, and FCGR3B CN variation is a common risk factor for systemic autoimmunity.