Interferon-γ contributes to HLA-B27-associated unfolded protein response in spondyloarthropathies.
ABSTRACT HLA-B27 positivity strongly influences the susceptibility to and phenotype of spondyloarthropathies (SpA). This study was designed to screen factors that activate the promoter of HLA-B27 in U937 cells, and to assess whether these promoter-activating factors induce the unfolded protein response (UPR) in HLA-B27-expressing cells.
Cytometric Bead Array, flow cytometry, and real-time polymerase chain reaction were used to detect the expression of cytokines and UPR-associated proteins in peripheral blood and synovial fluid of patients with SpA. The HLA-B27 promotor transfectant was incubated separately with cytokines and Toll-like receptor ligands. After interferon-γ (IFN-γ) stimulation, expressions of GRP78, CHOP, and XBP-1 were tested in HLA-B27-expressing U937 cells and peripheral blood mononuclear cell (PBMC) of patients with ankylosing spondylitis (AS). (Clinical trial registration no. ChiCTR-OCC-11001565)
Expressions of GRP78, CHOP, and XBP-1 in monocytes/macrophages of SpA peripheral blood and synovial fluid were higher than those in healthy controls and patients with osteoarthritis (OA) (p < 0.05). Tumor necrosis factor-α (TNF-α) and IFN-α, IFN-ß, and IFN-γ were found to have activated the HLA-B27 promoter in the U937 cell line (p < 0.05). Following stimulation with IFN-γ, the expressions of GRP78, CHOP and XBP-1 in HLA-B27-transfected U937 cells and PBMC of HLA-B27-positive AS patients were more intense than those in A2-U937 cells, HLA-B27-negative AS patients, or healthy controls (p < 0.05).
Expressions of GRP78, CHOP, and XBP-1 were higher in monocytes/macrophages of patients with SpA than those in both OA patients and healthy controls, suggesting that UPR may participate in the pathogenesis of SpA. TNF-α and IFN-α, IFN-ß, and IFN-γ significantly activated HLA-B27 promoter in the U937 cell line, and IFN-γ, the strongest activating factor, may induce the UPR in HLA-B27-expressing cells.
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ABSTRACT: Objective To determine whether HLA–B27 expression alters the response of bone marrow monocytes from HLA–B27/human β2-microglobulin–transgenic (B27-Tg) rats to tumor necrosis factor α (TNFα) and, if so, whether this affects the cells involved in bone homeostasis. Methods Bone marrow monocytes were treated with RANKL or with TNFα to promote osteoclast formation. Osteoclasts were quantified by counting. Gene expression was measured using quantitative polymerase chain reaction analysis, and protein was detected by enzyme-linked immunosorbent assay, immunoblotting, or immunofluorescence. Effects of endogenously produced cytokines on osteoclast formation were determined with neutralizing antibodies. ResultsTNFα treatment enhanced osteoclast formation 2.5-fold in HLA–B27–expressing cells as compared to wild-type or to HLA–B7/human β2-microglobulin–expressing monocytes. TNFα induced ∼4-fold up-regulation of HLA–B27, which was associated with the accumulation of misfolded heavy chains, binding of the endoplasmic reticulum (ER) chaperone BiP, and activation of an ER stress response, which was not seen with HLA–B7. No differences were seen with RANKL-induced osteoclastogenesis. Enhanced interleukin-1α (IL-1α) production from ER-stressed bone marrow monocytes from B27-Tg rats was found to be necessary and sufficient for enhanced osteoclast formation. However, bone marrow monocytes from B27-Tg rats also produced more interferon-β (IFNβ), which attenuated the effect of IL-1α on osteoclast formation. ConclusionHLA–B27–induced ER stress alters the response of bone marrow monocytes from B27-Tg rats to TNFα, which is associated with enhanced production of IL-1α and IFNβ, cytokines that exhibit opposing effects on osteoclast formation. The altered response of cells expressing HLA–B27 to proinflammatory cytokines suggests that this class I major histocompatibility complex allele may contribute to the pathogenesis of spondyloarthritis and its unique phenotype through downstream effects involving alterations in bone homeostasis.Arthritis & Rheumatology 08/2013; 65(8). · 7.48 Impact Factor
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ABSTRACT: Mechanisms underlying the striking association of spondyloarthritis (SpA) with the MHC class I HLA-B27 remain elusive. SpA-like disease develops spontaneously in B*2705 transgenic rats in correlation with high HLA-B27 expression levels. The aim of this study was to examine the consequences of increased expression on oligomerization and intracellular redistribution of HLA-B27 alleles differently associated with SpA. HeLa cells were transfected with complementary DNA encoding for HLA-B proteins fused to yellow fluorescent protein and/or Renilla luciferase and harvested at an early and a later phase of expression. We monitored HLA-B intracellular trafficking and localisation by means of microscopy and live-cell imaging. Bioluminescence resonance energy transfer (BRET) and Western blot were used to monitor HLA-B oligomerization. At low expression levels, BRET signals were similarly elevated for SpA-associated HLA-B27 alleles but lower for the non-associated B*2706. Remarkably, at higher expression levels, HLA-B27 signals remained steady while those for HLA-B7 sharply decreased reaching B*2706 signals level, due, at least in part, to a decreased oligomer proportion without UPR outbreak. Such differential behavior was not abrogated by proteasome inhibition. With increased expression, all HLA-B proteins accumulated to a high density in cytoplasmic vesicles with labile form and size. The extent of this phenomenon was strictly correlated with level of predisposition to SpA. To our knowledge, this is the first time that a correlation is reported between the level of predisposition to SpA conferred by HLA-B27 alleles and their biochemical behavior. These findings open new perspectives in understanding the pathogenicity of HLA-B27. © 2014 American College of Rheumatology.Arthritis & rheumatology (Hoboken, N.J.). 04/2014;
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ABSTRACT: Inflammatory arthritis is a condition which is characterised by recurrent episodes of joint pain and swelling. It encompasses a spectrum of disorders ranging from rheumatoid arthritis to ankylosing spondylitis. In these conditions, for reasons that are poorly understood, the immune system raises an inflammatory response within the joint space. In some cases, autoantigens have been identified (e.g., anticitrullinated peptides in rheumatoid arthritis), but the absence of these, in the seronegative arthritides, for example, raises question as to the underlying pathogenesis. Interest has, therefore, turned to host-pathogen interactions and whether aberrant immune responses to these could explain the development of arthritis. This has been most widely studied in reactive arthritis (ReA), where an infectious episode precedes the development of the joint symptoms. In this review, we present the evidence for the role of host-bacterial interactions in the pathogenesis of joint inflammation with particular emphasis on ReA. We discuss a range of possible mechanisms including molecular mimicry, persistent low grade infections, and abnormal host responses to common bacterial causes of reactive arthritis as well as discussing some of the clinical challenges that we face in making the diagnosis and in treatment of persistent symptoms.International journal of inflammation. 01/2014; 2014:158793.