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Effects of Anti-DNA Antibodies on Pleural Mesothelial Cells

In Vitro Studies to Explore the Pathogenetic Mechanism of Pulmonary Lupus

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Effects of Anti-DNA Antibodies on Pleural Mesothelial Cells by Hong Guo
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This dissertation, "Effects of Anti-DNA Antibodies on Pleural Mesothelial Cells: in Vitro Studies to Explore the Pathogenetic Mechanism of Pulmonary Lupus" by Hong, Guo, 郭紅, was obtained from The University of Hong Kong (Pokfulam, Hong Kong) and is being sold pursuant to Creative Commons: Attribution 3.0 Hong Kong License. The content of this dissertation has not been altered in any way. We have altered the formatting in order to facilitate the ease of printing and reading of the dissertation. All rights not granted by the above license are retained by the author. Abstract: Abstract of thesis entitled Effects of Anti-DNA Antibodies on Pleural Mesothelial Cells: in vitro Studies to Explore the Pathogenetic Mechanism of Pulmonary Lupus Submitted by Guo Hong For the degree of Master of Philosophy at the University of Hong Kong in August 2003 Pulmonary abnormalities, including pleuritis, are common in SLE. The pathogenic effects of autoantibodies in pulmonary lupus remain unclear. This study has investigated in vitro the pathophysiologic effect of anti-DNA containing IgG from lupus patients on pleural mesothelial cells and the related mechanisms. Our in vivo study has explored the pulmonary expression of inflammatory cytokines in lupus-prone mice and has evaluated the potential therapeutic effect of mycophenolate mofetil (MMF) on the release of cytokines. In in vitro experiments, IgG from 28 lupus patients and 13 healthy controls was purified by protein-G affinity chromatography. The amount of anti-dsDNA, anti-histone and anti-nucleohistone IgG were determined with ELISA. The lupus patients were divided into an active or an inactive group based on the SLE Disease Activity Index. The binding of IgG to a human pleural mesothelial cell-line (MeT-5A) under different conditions including pretreatment with DNase, pre-incubation with exogenous histone, DNA or nucleohistone was examined using flow cytometry and cellular ELISA. Gene expression and protein synthesis for IL-1β, MCP-1 and TGF-β1 in MeT-5A cells were determined using RT-PCR and ELISA. We demonstrated that the binding of IgG to MeT-5A cells was higher in the active lupus group than that in the inactive lupus group or controls. The binding decreased in the lupus group following the pretreatment of MeT-5A cells with DNase. The binding of IgG to MeT-5A cells was increased by 112% in the active lupus group following pre-incubation with histone, but not with DNA or nucleohistone. Gene expression and protein synthesis of MCP-1, TGF-β1, and IL-1β in MeT-5A cells were significantly enhanced following incubation with IgG from patients with active lupus when compared with the inactive lupus and control group. The concentration of anti-dsDNA antibodies correlated with the binding of IgG to MeT-5A cells as well as with cytokine synthesis by MeT-5A cells. In addition, the anti-histone antibody in the active lupus group was higher than that in the inactive lupus group and correlated with the cell binding or the synthesis of MCP-1. In our in vivo study, eight-week-old MRL/lpr mice were treated with MMF in vehicle for 8 or 12 weeks, while the control MRL/lpr and MRL mice received vehicle alone. The synthesis of MCP-1, TGF-β1, and IL-1β in the lungs were determined by RT-PCR and Western blot. We observed that gene expression of cytokines was significantly increased in MRL/lpr lupus mice as compared with the MRL controls. Gene expression of these cytokines and protein synthesis of TGF-β1 in the lung of MMF-treated MRL/lpr mice were significantly reduced following 8 or 12 weeks of treatment as compared with the untreated MRL/lpr mice. In summary, we have demonstrated that the in vitro binding of autoantibodies from active lupus patients to MeT-5A cells is closely associated with the disease activity. The binding of antibodies to MeT-5A cells is mediated via DNA and histone. Anti-dsDNA antibodies exert a direct pathogenic effect in lupus pleuritis by binding direct
Release date NZ
January 26th, 2017
Author
Contributor
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Country of Publication
United States
Illustrations
colour illustrations
Imprint
Open Dissertation Press
Dimensions
216x279x7
ISBN-13
9781361105498
Product ID
26645315

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