title: Targeting downstream transcription factors and epigenetic modifications following Toll-like receptor 7/8 ligation to forestall tissue injury in anti-Ro60 associated heart block.
abstract: Based on the consistent demonstration of fibrosis of the atrioventricular node surrounded by macrophages and multinucleated giant cells in anti-Ro antibody exposed fetuses dying with heart block, this study focuses on macrophage signaling stimulated by ssRNA associated with the Ro60 protein and the impact of antagonizing innate cell drivers such as TLR7/8. Transcriptome and epigenetic modifications which affect transcription factors, NF-κB and STAT1, were selected to evaluate the phenotype of macrophages in which TLR7/8 was ligated following treatment with either anti-Ro60/Ro60/hY3 RNA immune complexes or transfection with hY3. Based on microarray, TNF and IL6 were among the most highly upregulated genes in both stimulated conditions, each of which was significantly inhibited by preincubation with hydroxychloroquine (HCQ). In contrast, following stimulation of macrophages with either TNF-α or IFN-α, which do not signal through TLR, the resultant gene expression was refractory to HCQ. Ligation of TLR7/8 resulted in increased histone methylation as measured by increased H3K4me2, a requirement for binding of NF-κB at certain promoters, specifically the kB1 region in the TNF promoter (ChIP-qPCR), which was significantly decreased by HCQ. In summary, these results support that the HCQ-sensitive phenotype of hY3 stimulated macrophages reflects the bifurcation of TLR downstream signals involving NF-κB and STAT 1 pathways and for the former dimethylation of H3K4. Accordingly, HCQ may act more as a preventive measure in downregulating the initial production of IFN-α or TNF-α and not affect the resultant autocoid stimulation reflected in TNF-α and IFN-α responsive genes. The beneficial scope of antimalarials in the prevention of organ damage, inclusive of heart block in an anti-Ro offspring or more broadly SLE, may include in part, a mechanism targeting TLR-dependent epigenetic modification.
authors.affiliation: New York University School of Medicine, Department of Medicine, Division of Rheumatology, 550 First Avenue, New York, NY 10016, USA. Electronic address: bobdclancy@aol.com.
authors.forename: Robert M
authors.initials: RM
authors.lastname: Clancy
authors.affiliation: New York University School of Medicine, Department of Medicine, Division of Rheumatology, 550 First Avenue, New York, NY 10016, USA.
authors.forename: Androo J
authors.initials: AJ
authors.lastname: Markham
authors.affiliation: Garvan Institute of Medical Research, Darlinghurst, Sydney, NSW 2010, Australia.
authors.forename: Joanne H
authors.initials: JH
authors.lastname: Reed
authors.affiliation: New York University School of Medicine, Department of Medicine, Division of Rheumatology, 550 First Avenue, New York, NY 10016, USA.
authors.forename: Miroslav
authors.initials: M
authors.lastname: Blumenberg
authors.affiliation: Johns Hopkins University School of Medicine, Departments of Pathology and Oncology, 600 N. Wolfe Street, Baltimore, MD 21287, USA.
authors.forename: Marc K
authors.initials: MK
authors.lastname: Halushka
authors.affiliation: New York University School of Medicine, Department of Medicine, Division of Rheumatology, 550 First Avenue, New York, NY 10016, USA.
authors.forename: Jill P
authors.initials: JP
authors.lastname: Buyon
mesh_terms: D000974:Antibodies, Antinuclear / Q000276:immunology*; D000936:Antigen-Antibody Complex / Q000276:immunology / Q000378:metabolism; D002460:Cell Line; D004705:Endocytosis / Q000276:immunology; D044127:Epigenesis, Genetic; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D018390:Gene Targeting; D006327:Heart Block / Q000209:etiology* / Q000378:metabolism*; D006655:Histone Deacetylases / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008264:Macrophages / Q000276:immunology / Q000378:metabolism; D011485:Protein Binding; D012261:Ribonucleoproteins / Q000276:immunology; D051199:Toll-Like Receptor 7 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051200:Toll-Like Receptor 8 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics*; D000099207:SS-A Antigen; D000099186:Histone Deacetylase 3
chemical_list: D000974:Antibodies, Antinuclear; D000936:Antigen-Antibody Complex; D006655:Histone Deacetylases; D012261:Ribonucleoproteins; D051199:Toll-Like Receptor 7; D051200:Toll-Like Receptor 8; D014157:Transcription Factors; D000099207:SS-A Antigen; D000099186:Histone Deacetylase 3; C035356:SS-A antibodies
keywords: Anti-SSA/Ro autoantibodies; Epigenetics; Innate immunity; Macrophages; Toll-like receptor 7