Immune complexes containing scleroderma-specific autoantibodies induce a profibrotic and proinflammatory phenotype in skin fibroblasts

Abstract

BackgroundIn systemic sclerosis (SSc), autoantibodies provide the most accurate tool to predict the disease subset and pattern of organ involvement. Scleroderma autoantibodies target nucleic acids or DNA/RNA-binding proteins, thus SSc immune complexes (ICs) can embed nucleic acids. Our working hypothesis envisaged that ICs containing scleroderma-specific autoantibodies might elicit proinflammatory and profibrotic effects in skin fibroblasts.MethodsFibroblasts were isolated from skin biopsies obtained from healthy subjects and patients with diffuse cutaneous SSc (dcSSc). ICs were purified by polyethylene-glycol precipitation from sera of SSc patients bearing different autoantibodies. ICs from patients with systemic lupus erythematosus (SLE) and primary anti-phospholipid syndrome (PAPS) and from normal healthy subjects (NHS) were used as controls. After incubation with ICs, fibroblasts were evaluated for ICAM-1 expression, interleukin (IL)-6, IL-8, monocyte chemoattractant protein (MCP)-1, matrix metalloproteinase (MMP)-2, tumor growth factor (TGF)-β1 and Pro-CollagenIα1 secretion, collagen (col)Iα1, mmp-1, toll-like receptor (tlr)2, tlr3, tlr4, tlr7, tlr8, tlr9, interferon (ifn)-α, ifn-β and endothelin-1 mRNA, and NFκB, p38MAPK and SAPK-JNK activation rate. Experiments were also performed after pretreatment with DNase I/RNase and NFκB/p38MAPK inhibitors.ResultsThe antigenic reactivity for each SSc-IC mirrored the corresponding serum autoantibody specificity, while no positivity was observed in NHS-ICs or sera. SSc-ICs but not NHS-ICs increased ICAM-1 expression, stimulated IL-6, IL-8, MMP-2, MCP-1, TGF-β1 and Pro-CollagenIα1 secretion, upregulated et-1, ifn-α, ifn-β, tlr2, tlr3 and tlr4, and activated NFκB, p38MAPK and SAPK-JNK. tlr9 was significantly upregulated by ARA-ICs, mmp-1 was significantly induced by ACA-ICs whereas colIα1 was not modulated by any SSc-ICs. SLE-ICs and PAPS-ICs significantly upregulated MMP-2 and activated NFκB, p38MAPK and SAPK-JNK. SLE-ICs and PAPS-ICs did not affect colIα1, mmp-1 and Pro-CollagenIα1. DNase I and RNase treatment significantly reduced the upregulation of study mediators induced by SSc-ICs. Pretreatment with NFκB/p38MAPK inhibitors suggested that response to anti-Th/To-ICs was preferentially mediated by p38MAPK whereas ATA-ICs, ACA-ICs and ARA-ICs engaged both mediators. In dcSSc fibroblasts, stimulation with SSc-ICs and NHS-ICs upregulated IL-6 and IL-8.ConclusionsThese data provide the first demonstration of the proinflammatory and profibrotic effects of SSc-ICs on fibroblasts, suggesting the potential pathogenicity of SSc autoantibodies. These effects might be mediated by Toll-like receptors via the interaction with nucleic acid fragments embedded in SSc-ICs.

Efferocytosis capacities of blood monocyte‐derived macrophages in systemic sclerosis

“…Moreover, unengulfed apoptotic cells may secondary undergo necrosis with release of intracellular and nuclear components that could also activate the immune system to produce autoantibodies . It has also been recently highlighted that IC containing intranuclear components and SSc‐specific ANA could trigger fibroblast activation and fibrosis in SSc . Therefore, the observed defect in efferocytosis of MDM in SSc could participate in the pathogenesis of autoimmunity and fibrosis via an increase release of antigen with subsequent formation of IC.…”

“…20 It has also been recently highlighted that IC containing intranuclear components and SSc-specific ANA could trigger fibroblast activation and fibrosis in SSc. 2 Therefore, the observed defect in efferocytosis of MDM in SSc could participate in the pathogenesis of autoimmunity and fibrosis via an increase release of antigen with subsequent formation of IC. Although the role of IC has been largely studied in mice models of SLE, their relevance in mice models of SSc is still to be determined.…”

“…Specific SSc‐ANA especially target centromere proteins (anti‐centromere antibodies), topoisomerase I (anti‐Topo I antibodies) and RNA‐polymerase III (anti‐RNA pol‐III). Although the pathogenic links between autoimmunity and fibrosis in SSc are still unclear, it has been recently highlighted that immune complex (IC) containing intranuclear components and SSc‐specific ANA could trigger fibroblast activation and fibrosis in SSc …”

“…Although the pathogenic links between autoimmunity and fibrosis in SSc are still unclear, it has been recently highlighted that immune complex (IC) containing intranuclear components and SSc-specific ANA could trigger fibroblast activation and fibrosis in SSc. 2 The mechanisms leading to the release of intranuclear components with subsequent emergence of autoantibodies and IC are still to be determined in SSc. Apoptotic cells could be a source of circulating nuclear antigens because, when they are not rapidly and/or efficiently cleared, they can switch from apoptosis to necrosis with subsequent release of their intracellular materials and triggering of proinflammatory signals.…”

Scleroderma-specific autoantibodies embedded in immune complexes mediate endothelial damage: an early event in the pathogenesis of systemic sclerosis

“…Evidence of the pathogenic role of SSc-speci c antibodies was raised by our group, with the demonstration that scleroderma-speci c autoantibodies bound to their antigens to form immune complexes (ICs) elicit proin ammatory and pro-brotic effects on healthy skin broblasts. According to our previous work, the effects of SSc-ICs on broblasts might be mediated by Toll-Like Receptors (TLRs) interacting with the nucleic acid fragments embedded in scleroderma ICs [14].…”