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[This corrects the article DOI: 10.1371/journal.pgen.1005673.].
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Human ß-defensin 3 (hBD3) is a cationic antimicrobial peptide with potent bactericidal activity in vitro. HBD3 is produced in response to pathogen challenge and can modulate immune responses. The amplified recognition of self-DNA by human plasmacytoid dendritic cells has been previously reported, but we show here that hBD3 preferentially enhances the response to bacterial DNA in mouse Flt-3 induced dendritic cells (FLDCs) and in human peripheral blood mononuclear cells. We show the effect is mediated through TLR9 and although hBD3 significantly increases the cellular uptake of both E. coli and self-DNA in mouse FLDCs, only the response to bacterial DNA is enhanced. Liposome transfection also increases uptake of bacterial DNA and amplifies the TLR9-dependent response. In contrast to hBD3, lipofection of self-DNA enhances inflammatory signaling, but the response is predominantly TLR9-independent. Together, these data show that hBD3 has a role in the innate immune-mediated response to pathogen DNA, increasing inflammatory signaling and promoting activation of the adaptive immune system via antigen presenting cells including dendritic cells. Therefore, our data identify an additional immunomodulatory role for this copy-number variable defensin, of relevance to host defence against infection and indicate a potential for the inclusion of HBD3 in pathogen DNA-based vaccines.
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Células Dendríticas/inmunología , Escherichia coli/inmunología , Leucocitos Mononucleares/inmunología , Receptor Toll-Like 9/metabolismo , beta-Defensinas/metabolismo , Animales , Células Cultivadas , ADN Bacteriano/inmunología , Interacciones Huésped-Patógeno , Humanos , Inmunidad Innata , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptor Toll-Like 9/genéticaRESUMEN
Human ß-defensin 3 (hBD3) is a cationic host defence peptide and is part of the innate immune response. HBD3 is present on a highly copy number variable block of six ß-defensin genes, and increased copy number is associated with the autoimmune disease psoriasis. It is not known how this increase influences disease development, but psoriasis is a T cell-mediated disease and activation of the innate immune system is required for the initial trigger that leads to the amplification stage. We investigated the effect of hBD3 on the response of primary macrophages to various TLR agonists. HBD3 exacerbated the production of type I Interferon-ß in response to the viral ligand mimic polyinosinic:polycytidylic acid (polyI:C) in both human and mouse primary cells, although production of the chemokine CXCL10 was suppressed. Compared to polyI:C alone, mice injected with both hBD3 peptide and polyI:C also showed an enhanced increase in Interferon-ß. Mice expressing a transgene encoding hBD3 had elevated basal levels of Interferon-ß, and challenge with polyI:C further increased this response. HBD3 peptide increased uptake of polyI:C by macrophages, however the cellular response and localisation of polyI:C in cells treated contemporaneously with hBD3 or cationic liposome differed. Immunohistochemistry showed that hBD3 and polyI:C do not co-localise, but in the presence of hBD3 less polyI:C localises to the early endosome. Using bone marrow derived macrophages from knockout mice we demonstrate that hBD3 suppresses the polyI:C-induced TLR3 response mediated by TICAM1 (TRIF), while exacerbating the cytoplasmic response through MDA5 (IFIH1) and MAVS (IPS1/CARDIF). Thus, hBD3, a highly copy number variable gene in human, influences cellular responses to the viral mimic polyI:C implying that copy number may have a significant phenotypic effect on the response to viral infection and development of autoimmunity in humans.
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Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras del Transporte Vesicular/genética , ARN Helicasas DEAD-box/genética , Psoriasis/genética , Receptor Toll-Like 3/genética , beta-Defensinas/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras del Transporte Vesicular/metabolismo , Animales , Médula Ósea , Quimiocina CXCL10/genética , ARN Helicasas DEAD-box/metabolismo , Humanos , Inmunidad Innata/genética , Helicasa Inducida por Interferón IFIH1 , Liposomas/metabolismo , Macrófagos/metabolismo , Macrófagos/patología , Ratones , Ratones Noqueados , Poli I-C/administración & dosificación , Psoriasis/patología , Receptor Toll-Like 3/antagonistas & inhibidores , beta-Defensinas/metabolismoRESUMEN
Type I interferons (IFNs) are essential mediators of antiviral responses. These cytokines have been implicated in the pathogenesis of autoimmunity, most notably systemic lupus erythematosus (SLE), diabetes mellitus, and dermatomyositis, as well as monogenic type I interferonopathies. Despite a fundamental role in health and disease, the direct quantification of type I IFNs has been challenging. Using single-molecule array (Simoa) digital ELISA technology, we recorded attomolar concentrations of IFNα in healthy donors, viral infection, and complex and monogenic interferonopathies. IFNα protein correlated well with functional activity and IFN-stimulated gene expression. High circulating IFNα levels were associated with increased clinical severity in SLE patients, and a study of the cellular source of IFNα protein indicated disease-specific mechanisms. Measurement of IFNα attomolar concentrations by digital ELISA will enhance our understanding of IFN biology and potentially improve the diagnosis and stratification of pathologies associated with IFN dysregulation.