| 1. |
- Albrecht, Inka, et al.
(författare)
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Development of autoantibodies against muscle-specific FHL1 in severe inflammatory myopathies
- 2015
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Ingår i: Journal of Clinical Investigation. - : AMER SOC CLINICAL INVESTIGATION INC. - 0021-9738 .- 1558-8238. ; 125:12, s. 4612-4624
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Tidskriftsartikel (refereegranskat)abstract
- Mutations of the gene encoding four-and-a-half LIM domain 1 (FHL1) are the causative factor of several X-linked hereditary myopathies that are collectively termed FHL1-related myopathies. These disorders are characterized by severe muscle dysfunction and damage. Here, we have shown that patients with idiopathic inflammatory myopathies (IIMs) develop autoimmunity to FHL1, which is a muscle-specific protein. Anti-FHL1 autoantibodies were detected in 25% of IIM patients, while patients with other autoimmune diseases or muscular dystrophies were largely anti-FHL1 negative. Anti-FHL1 reactivity was predictive for muscle atrophy, dysphagia, pronounced muscle fiber damage, and vasculitis. FHL1 showed an altered expression pattern, with focal accumulation in the muscle fibers of autoantibody-positive patients compared with a homogeneous expression in anti-FHL1-negative patients and healthy controls. We determined that FHL1 is a target of the cytotoxic protease granzyme B, indicating that the generation of FHL1 fragments may initiate FHL1 autoimmunity. Moreover, immunization of myositis-prone mice with FHL1 aggravated muscle weakness and increased mortality, suggesting a direct link between anti-FHL1 responses and muscle damage. Together, our findings provide evidence that FHL1 may be involved in the pathogenesis not only of genetic FHL1-related myopathies but also of autoimmune IIM. Importantly, these results indicate that anti-FHL1 autoantibodies in peripheral blood have promising potential as a biomarker to identify a subset of severe IIM.
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| 2. |
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| 3. |
- Carlberg, Konstantin, et al.
(författare)
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Integrated Single Cell and Spatial Transcriptomics Reveal Autoreactive Differentiated B Cells in Joints of Early Rheumatoid Arthritis
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Rheumatoid Arthritis (RA) is a prevalent autoimmune disease characterized by inflammation of peripheral joints. Patients can be subdivided by the presence or absence of Rheumatoid Factor and anti-citrullinated protein antibodies (ACPA) in their circulation. Inflammation of the joint tissue is associated with infiltration of leukocytes from the blood, which can result in generation of lymphoid structures composed of B and T cells. Previous studies have shown that both memory B cells and antibody-secreting plasma cells populate the rheumatic joint tissue when captured from established and often long-standing disease. However, it has remained unclear, whether these cells are autoreactive and whether the associated lymphoid structures are present at the site of inflammation already at the time of diagnosis. Here, we used an integrated single cell and spatial transcriptomic approach to study B and plasma cells in synovial tissue of ACPA- and ACPA+ RA patients at this early time point. We found evidence for T cell help to B cells and presence of memory B and plasma cell pools in ACPA- as well as in ACPA+ RA. Our results demonstrated common supportive microenvironments in both patient subgroups, clonal relationships between the memory B and plasma cell pools and autoreactivity within the plasma cell compartment. These findings challenge our understanding of the dynamics of local adaptive immune responses in the RA joint of ACPA- and ACPA+ patients at the time of diagnosis, with direct implications for B and T cell targeting therapies for both patient subgroups.
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| 4. |
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| 5. |
- Chemin, Karine, et al.
(författare)
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A Novel HLA-DRB1*10:01-Restricted T Cell Epitope From Citrullinated Type II Collagen Relevant to Rheumatoid Arthritis
- 2016
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Ingår i: Arthritis & Rheumatology. - : Wiley. - 2326-5191 .- 2326-5205. ; 68:5, s. 1124-1135
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Tidskriftsartikel (refereegranskat)abstract
- Objective. Antibodies against citrullinated type II collagen (Cit-CII) are common in the sera and synovial fluid of patients with rheumatoid arthritis (RA); however, the known T cell epitope of CII is not dependent on citrullination. The aim of this study was to identify and functionally characterize the Cit-CII-restricted T cell epitopes that are relevant to RA. Methods. Peripheral blood mononuclear cells (PBMCs) from HLA-DRB1*10:01-positive patients with RA and healthy donors were stimulated in vitro with candidate CII peptides. CD154 up-regulation was measured as a marker of antigen-specific activation, and anti-HLA-DR-blocking experiments confirmed HLA restriction. Cytokine production was measured using a Luminex technique. Direct peptide-binding assays using HLA-DRB1*10:01 and HLA-DRB1*04:01 monomeric proteins were performed. The T cell receptor (TCR) beta-chain of CD154-enriched antigen-specific T cells was analyzed using high-throughput sequencing. Results. A novel Cit-CII peptide was identified based on its ability to activate CD4+ T cells from HLA-DRB1*10:01-positive individuals. When stimulated in vitro, Cit-CII autoreactive T cells produced proinflammatory cytokines. Cit-CII311-325 bound (with low affinity) to HLA-DRB1*10:01 but not to HLA-DRB1*04:01, while the native form was unable to bind either protein. In addition, highly expanded clones were identified in the TCR beta repertoire of Cit-CII311-325-stimulated PBMCs. Conclusion. These results illustrate the ability of the citrullination process to create T cell epitopes from CII, a cartilage-restricted protein that is relevant to RA pathogenesis. The exclusive binding of Cit-CII311-325 to HLA-DRB1*10:01 suggests that recognition of citrullinated epitopes might vary between individuals carrying different RA-associated HLA-DR molecules.
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| 6. |
- Chemin, Karine, et al.
(författare)
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EOMES-positive CD4+ T cells are increased in PTPN22 (1858T) risk allele carriers.
- 2018
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Ingår i: European Journal of Immunology. - : Wiley. - 0014-2980 .- 1521-4141. ; 48:4, s. 655-669
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Tidskriftsartikel (refereegranskat)abstract
- The presence of the PTPN22 risk allele (1858T) is associated with several autoimmune diseases including rheumatoid arthritis (RA). Despite a number of studies exploring the function of PTPN22 in T cells, the exact impact of the PTPN22 risk allele on T-cell function in humans is still unclear. In this study, using RNA sequencing, we show that, upon TCR-activation, naïve human CD4+ T cells homozygous for the PTPN22 risk allele overexpress a set of genes including CFLAR and 4-1BB, which are important for cytotoxic T-cell differentiation. Moreover, the protein expression of the T-box transcription factor Eomesodermin (EOMES) was increased in T cells from healthy donors homozygous for the PTPN22 risk allele and correlated with a decreased number of naïve CD4+ T cells. There was no difference in the frequency of other CD4+ T cell subsets (Th1, Th17, Tfh, Treg). Finally, an accumulation of EOMES+CD4+ T cells was observed in synovial fluid of RA patients with a more pronounced production of Perforin-1 in PTPN22 risk allele carriers. Altogether, we propose a novel mechanism of action of PTPN22 risk allele through the generation of cytotoxic CD4+ T cells and identify EOMES+CD4+ T cells as a relevant T-cell subset in RA pathogenesis.
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| 7. |
- Gerstner, Christina, et al.
(författare)
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Multi-HLA class II tetramer analyses of citrulline-reactive T cells and early treatment response in rheumatoid arthritis
- 2020
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Ingår i: BMC Immunology. - : Springer Science and Business Media LLC. - 1471-2172. ; 21:1
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Tidskriftsartikel (refereegranskat)abstract
- Background HLA class II tetramers can be used for ex vivo enumeration and phenotypic characterisation of antigen-specific CD4+ T cells. They are increasingly applied in settings like allergy, vaccination and autoimmune diseases. Rheumatoid arthritis (RA) is a chronic autoimmune disorder for which many autoantigens have been described. Results Using multi-parameter flow cytometry, we developed a multi-HLA class II tetramer approach to simultaneously study several antigen specificities in RA patient samples. We focused on previously described citrullinated HLA-DRB1*04:01-restricted T cell epitopes from alpha-enolase, fibrinogen-beta, vimentin as well as cartilage intermediate layer protein (CILP). First, we examined inter-assay variability and the sensitivity of the assay in peripheral blood from healthy donors (n = 7). Next, we confirmed the robustness and sensitivity in a cohort of RA patients with repeat blood draws (n = 14). We then applied our method in two different settings. We assessed lymphoid tissue from seropositive arthralgia (n = 5) and early RA patients (n = 5) and could demonstrate autoreactive T cells in individuals at risk of developing RA. Lastly, we studied peripheral blood from early RA patients (n = 10) and found that the group of patients achieving minimum disease activity (DAS28 < 2.6) at 6 months follow-up displayed a decrease in the frequency of citrulline-specific T cells. Conclusions Our study demonstrates the development of a sensitive tetramer panel allowing simultaneous characterisation of antigen-specific T cells in ex vivo patient samples including RA 'at risk' subjects. This multi-tetramer approach can be useful for longitudinal immune-monitoring in any disease with known HLA-restriction element and several candidate antigens.
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| 8. |
- Hardt, Uta, et al.
(författare)
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Integrated single cell and spatial transcriptomics reveal autoreactive differentiated B cells in joints of early rheumatoid arthritis
- 2022
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Ingår i: Scientific Reports. - : Springer Nature. - 2045-2322. ; 12:1
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Tidskriftsartikel (refereegranskat)abstract
- B cells play a significant role in established Rheumatoid Arthritis (RA). However, it is unclear to what extent differentiated B cells are present in joint tissue already at the onset of disease. Here, we studied synovial biopsies (n = 8) captured from untreated patients at time of diagnosis. 3414 index-sorted B cells underwent RNA sequencing and paired tissue pieces were subjected to spatial transcriptomics (n = 4). We performed extensive bioinformatics analyses to dissect the local B cell composition. Select plasma cell immunoglobulin sequences were expressed as monoclonal antibodies and tested by ELISA. Memory and plasma cells were found irrespective of autoantibody status of the patients. Double negative memory B cells were prominent, but did not display a distinct transcriptional profile. The tissue architecture implicate both local B cell maturation via T cell help and plasma cell survival niches with a strong CXCL12-CXCR4 axis. The immunoglobulin sequence analyses revealed clonality between the memory B and plasma cell pools further supporting local maturation. One of the plasma cell-derived antibodies displayed citrulline autoreactivity, demonstrating local autoreactive plasma cell differentiation in joint biopsies captured from untreated early RA. Hence, plasma cell niches are not a consequence of chronic inflammation, but are already present at the time of diagnosis.
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| 9. |
- Houtman, Miranda, et al.
(författare)
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T cells are influenced by a long non-coding RNA in the autoimmune associated PTPN2 locus
- 2018
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Ingår i: Journal of Autoimmunity. - : ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD. - 0896-8411 .- 1095-9157. ; 90, s. 28-38
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Tidskriftsartikel (refereegranskat)abstract
- Non-coding SNPs in the protein tyrosine phosphatase non-receptor type 2 (PTPN2) locus have been linked with several autoimmune diseases, including rheumatoid arthritis, type I diabetes, and inflammatory bowel disease. However, the functional consequences of these SNPs are poorly characterized. Herein, we show in blood cells that SNPs in the PTPN2 locus are highly correlated with DNA methylation levels at four CpG sites downstream of PTPN2 and expression levels of the long non-coding RNA (IncRNA) LINC01882 downstream of these CpG sites. We observed that LINC01882 is mainly expressed in T cells and that anti-CD3/CD28 activated naive CD4(+) T cells downregulate the expression of LINC01882. RNA sequencing analysis of LINC01882 knockdown in Jurkat T cells, using a combination of antisense oligo-nucleotides and RNA interference, revealed the upregulation of the transcription factor ZEB1 and kinase MAP2K4, both involved in IL-2 regulation. Overall, our data suggests the involvement of LINC01882 in T cell activation and hints towards an auxiliary role of these non-coding SNPs in autoimmunity associated with the PTPN2 locus.
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| 10. |
- Lloyd, Katy A., et al.
(författare)
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Differential ACPA Binding to Nuclear Antigens Reveals a PAD-Independent Pathway and a Distinct Subset of Acetylation Cross-Reactive Autoantibodies in Rheumatoid Arthritis
- 2019
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Ingår i: Frontiers in Immunology. - : Frontiers Media SA. - 1664-3224. ; 9, s. 1-22
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Tidskriftsartikel (refereegranskat)abstract
- Rheumatoid arthritis (RA) associated anti-citrullinated protein autoantibodies (ACPA) target a wide range of modified proteins. Citrullination occurs during physiological processes such as apoptosis, yet little is known about the interaction of ACPA with nuclear antigens or apoptotic cells. Since uncleared apoptotic cells and neutrophil extracellular trap (NET) products have been postulated to be central sources of autoantigen and immunostimulation in autoimmune disease, we sought to characterize the anti-nuclear and anti-neutrophil reactivities of ACFA. Serology showed that a subset of anti-CCP2 seropositive RA patients had high reactivity to full-length citrullinated histones. In contrast, seronegative RA patients displayed elevated IgG reactivity to native histone compared to controls, but no citrulline-specific reactivity. Screening of 10 single B-cell derived monoclonal AGFA from RA patients revealed that four ACPA exhibited strong binding to apoptotic cells and three of these had anti-nuclear (ANA) autoantibody reactivity. Modified histones were confirmed to be the primary targets of this anti-nuclear ACPA subset following immunoprecipitation from apoptotic cell lysates. Monoclonal ACPA were also screened for reactivities against stimulated murine and human neutrophils, and all the nuclear-reactive monoclonal ACPA bound to NETs. Intriguingly, one ACPA mAb displayed a contrasting cytoplasmic perinuclear neutrophil binding and may represent a different NET-reactive ACPA subset. Notably, studies of CRISPR-Cas9 PAD4 KO cells and cells from PAD KO mice showed that the cytoplasmic NET-binding was fully dependent on PAD4, whilst nuclear- and histone-mediated NEI reactivity was largely PAD-independent. Our further analysis revealed that the nuclear binding could be explained by consensus-motif driven ACPA cross-reactivity to acetylated histones. Specific acetylated histone peptides targeted by the monoclonal antibodies were identified and the anti-modified protein autoantibody (AMPA) profile of the ACPA was found to correlate with the functional activity of the antibodies. In conclusion, when investigating monoclonal ACPA, we could group ACPA into distinct subsets based on their nuclear binding-patterns and acetylation-mediated binding to apoptotic cells, neutrophils, and NETs. Differential anti-modified protein reactivities of RA-autoantibody subsets could have an important functional impact and provide insights in RA pathogenesis.
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