Scleroderma-specific autoantibodies embedded in immune complexes mediate endothelial damage: an early event in the patho
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RESEARCH ARTICLE
Open Access
Scleroderma-specific autoantibodies embedded in immune complexes mediate endothelial damage: an early event in the pathogenesis of systemic sclerosis Elena Raschi1, Daniela Privitera1, Caterina Bodio1, Paola Adele Lonati1, Maria Orietta Borghi1,2, Francesca Ingegnoli2,3, Pier Luigi Meroni1,4 and Cecilia Beatrice Chighizola1,4*
Abstract Background: Consistently with their diagnostic and prognostic value, autoantibodies specific for systemic sclerosis (SSc) embedded in immune complexes (ICs) elicited a pro-inflammatory and pro-fibrotic cascade in healthy skin fibroblasts, engaging Toll-like receptors (TLRs) via their nucleic acid components. The objective of this study was to investigate the pathogenicity of SSc-ICs in endothelial cells. Methods: ICs were purified from the sera of SSc patients bearing different autoantibody specificities (antibodies against DNA topoisomerase I, centromeric proteins, RNA polymerase, and Th/To), patients with systemic lupus erythematosus (SLE) and primary anti-phospholipid syndrome (PAPS), or healthy controls (NHS) using polyethylene glycol precipitation. Human umbilical vein endothelial cells (HUVECs) were incubated with ICs, positive and negative controls. mRNA levels of endothelin-1 (et-1), collagenIα1 (colIα1), interferon (IFN)-α, and IFN-β were investigated by real-time PCR; et-1 and il-6 mRNA levels were assessed after pre-treatment with bafilomycin. ICAM-1 expression was evaluated by cell ELISA; secretion of IL-6, IL-8, and transforming growth factor (TGF)-β1 in culture supernatants was measured by ELISA. The expression of Fcγ receptors (CD64, CD32, and CD16) was assessed in endothelial cells at FACS analysis. Intracellular signaling pathways culminating with NFκB, p38MAPK, SAPK-JNK, and Akt were assessed by Western blotting. Healthy skin fibroblasts were stimulated with supernatants from HUVECs incubated with ICs, and TGF-β1 secretion and mRNA levels of colIα1 and matrix metalloproteinase (mmp)-1, protein expression of α smooth muscle actin (α-SMA), and IL-6 were evaluated by Western blotting; et-1 mRNA levels were assessed in fibroblasts pre-treated with IL-6 and TGF-β inhibitors and stimulated with ATA-ICs. (Continued on next page)
* Correspondence: [email protected] 1 Experimental Laboratory of Immunological and Rheumatologic Researches, IRCCS Istituto Auxologico Italiano, Via Zucchi 18, Cusano Milanino, 20095 Milan, Italy 4 Allergology, Clinical Immunology and Rheumatology Unit, IRCCS Istituto Auxologico Italiano, Piazzale Brescia 20, 20149 Milan, Italy Full list of author information is available at the end of the article © The Author(s). 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party mat
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