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- Sun, M, et al.
(författare)
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Anticitrullinated protein antibodies facilitate migration of synovial tissue-derived fibroblasts
- 2019
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Ingår i: Annals of the rheumatic diseases. - : BMJ. - 1468-2060 .- 0003-4967. ; 78:12, s. 1621-1631
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Tidskriftsartikel (refereegranskat)abstract
- Rheumatoid arthritis (RA)-specific anti-citrullinated protein/peptide antibodies (ACPAs) might contribute to bone loss and arthralgia before the onset of joint inflammation. We aimed to dissect additional mechanisms by which ACPAs might contribute to development of joint pathology.MethodsFibroblast-like synoviocytes (FLS) were isolated from the synovial membrane of patients with RA. The FLS cultures were stimulated with polyclonal ACPAs (anti-CCP-2 antibodies) purified from the peripheral blood of patients with RA or with monoclonal ACPAs derived from single synovial fluid B cells. We analysed how ACPAs modulate FLS by measuring cell adhesion and mobility as well as cytokine production. Expression of protein arginine deiminase (PAD) enzymes and protein citrullination were analysed by immunofluorescence, and signal transduction was studied using immunoblotting.ResultsChallenge of FLS by starvation-induced stress or by exposure to the chemokine interleukin-8 was essential to sensitise the cells to ACPAs. These challenges led to an increased PAD expression and protein citrullination and an ACPA-mediated induction of FLS migration through a mechanism involving phosphoinositide 3-kinase activation. Inhibition of the PAD enzymes or competition with soluble citrullinated proteins or peptides completely abolished the ACPA-induced FLS migration. Different monoclonal ACPAs triggered distinct cellular effects in either fibroblasts or osteoclasts, suggesting unique roles for individual ACPA clones in disease pathogenesis.ConclusionWe propose that transient synovial insults in the presence of a certain pre-existing ACPA repertoire might result in an ACPA-mediated increase of FLS migration.
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- Sun, M, et al.
(författare)
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DIVERSITY OF ANTI-CITRULLINATED PROTEIN ANTIBODY COMPOSITIONS INFLUENCE SYNOVIAL FIBROBLAST REACTIVITY
- 2020
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Ingår i: ANNALS OF THE RHEUMATIC DISEASES. - : BMJ. - 0003-4967 .- 1468-2060. ; 79, s. 569-570
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Anti-citrullinated protein antibodies (ACPAs) play an important role in rheumatoid arthritis (RA) pathogenesis. We hypothesized that the effect of these antibodies is mediated by their binding to synovial fibroblasts and inducing an increased mobility of fibroblasts1.Objectives:In our study, we analyzed and compared fibroblast modulation by ACPA pools obtained from different patients or by a set of monoclonal ACPAs with different fine specificity that were obtained from different tissue sites.Methods:Synovial fibroblasts were isolated from RA patients synovial tissue biopsies. Individual polyclonal ACPA and control IgGs were purified from sera of four ACPA-positive RA patients by affinity purification on protein G and CCP-2 columns. Monoclonal antibodies were derived from memory B cell isolated from blood2, synovial fluid or bronchoalveolar lavage of RA patients. Whole antibodies and F(ab’)2 fragments were tested in fibroblast migration using IncuCyte live-cell analysis. Blocking experiments were performed with soluble citrullinated proteins in SF migration. Cross-reactivity of the antibodies to citrullinated and acetylated epitopes was tested using PAD inhibitors (Cl-amidine and GSK199), histone acetyltransferases (anacardic acid) and deacetylases (trichostatin A). Binding patterns of monoclonal ACPAs, both whole and F(ab’)2 fragments were analyzed in synovial biopsies obtained from both healthy donors and RA patients.Results:Three out of four tested individual ACPA were able to promote fibroblast migration. Five out of nine tested monoclonal ACPAs stimulated fibroblast migration. One of these antibodies, clone 1325:01B09 is characterized by cross-reactivity to citrullinated, homocitrullinated and acetylated targets. The effect of 1325:01B09 on fibroblast migration was completely abolished by Cl-amidine or by pre-incubating the antibody with citrullinated fibrinogen or histone but not citrullinated enolase or vimentin. Despite the cross-reactivity to acetylated epitopes, neither anacardic acid nor trichostatin A could modulate the 1325:01B09 effect on fibroblast migration. F(ab’)2 fragments of this antibody stimulated fibroblast migration and labelled podoplanin-positive fibroblasts in inflamed RA synovium similarly to the intact antibody, indicating an Fc-independent effect.Conclusion:The effect on fibroblast mobility was likely to be mediated by binding to citrullinated epitopes but not through Fc receptors. Detection of fibroblast modulating ACPAs in majority of RA patients indicated that fibroblasts might be key cellular targets in disease pathogenesis, although individual variability might exist in the composition of ACPA cellular targets.References:[1]Sun M, Rethi B, Krishnamurthy A, et al. Anticitrullinated protein antibodies facilitate migration of synovial tissue-derived fibroblasts. Ann Rheum Dis 2019;78(12):1621-31. doi: 10.1136/annrheumdis-2018-214967 [published Online First: 2019/09/05][2]Amara K, Lena Israelsson, Ragnhild Stålesen, et al. A Refined Protocol for Identifying Citrulline-specific Monoclonal Antibodies from Single Human B Cells from Rheumatoid Arthritis Patient Material. Bio-protocol 2019;9(16)Disclosure of Interests:Meng Sun: None declared, Bence Réthi: None declared, Akilan Krishnamurthy: None declared, Vijay Joshua: None declared, Alexandra Circiumaru: None declared, Marianne Engström: None declared, Caroline Grönwall: None declared, Vivianne Malmström Grant/research support from: VM has had research grants from Janssen Pharmaceutica, Khaled Amara: None declared, Lars Klareskog: None declared, Heidi Wähämaa: None declared, Anca Catrina: None declared
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