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• W2903942609 abstract "Background Increased type I interferon (IFN) has been shown to affect survival and activation of B cells in SLE. This study investigated novel mechanisms of endogenous production and autocrine activity of IFNβ in SLE B cells at the single-cell level. Methods IFNβ in B cells from SLE patients was analyzed using t-SNE platform based high dimensional flow cytometry. Intracellular IFNβ expression was visualized and analyzed by super-resolution confocal imaging and ImageStream analysis. Single cell gene expression analysis was carried out using the Fluidigm/BioMark system for targeted expression of low abundance genes, and the 10x Chromium platform for unbiased transcriptome analysis of up to 4,000 B cells per subject. Functional production of type I IFNs by B cells was analyzed using a human type I IFNs SEAP reporter HEK293 cell line. Results High dimension flow cytometry analysis identified intracellular IFNβ expression in pDCs, B cells, and CD4 T cells. B-cell endogenous IFNβ was required for optimal in vitro BCR and TLR7-induced activation and survival of B cells. Using a Fluidigm targeted-gene approach, B cells could be divided into type I IFN expressing (IFN+) or type I IFN stimulated gene (ISG+) subpopulations, suggesting B cells not only respond to type I IFNs but also express type I IFNs including IFNB and different IFNA genes. TLR7 and TLR3 were mainly expressed by IFN +cells whereas TLR9 was mainly expressed by ISG +B cells. The production of functional IFNβ and IFNα protein by single B cells from SLE subjects and was verified using a novel alkaline phosphatase live staining of HEK-blue reporter cells. There was enhanced IFNAR signaling by reporter cells in direct contact with SLE B cells which was blocked by anti-IFNβ and anti-IFNα. Interesting, anti-Ig crosslinking was required for optimal B-cell endogenous type I IFNs to stimulate responder cells. Unbiased single cells transcriptome analysis of SLE B cells using the 5’ 10X Chromium platform and Loupe V(D)J Browser indicated that gene clusters in type I IFN expressing or responding SLE B cells exhibited unique heavy- and light-chain gene expression repertoires. Conclusion (i) B cells are an important source of type I IFNs in modulating TLR and BCR responses in SLE; (ii) well-orchestrated and distinct programs in type I expression and responses genes in subsets of B cells, and (iii) distinct pathways of B cell survival and activation based on combined signaling through BCR, TLR, and IFNAR with a distinct BCR heavy- and light-chain repertoire. Acknowledgements This work was supported by grants from R01-AI-071110, R01 AI134023, Lupus Research Alliance Distinguished Innovator Award, I01B × 004049, and 1I01B × 000600 to J.D.M, 2T32AI007051–39 Immunology T32 Training Grant and the LFA Finzi Summer Fellowship to J.A.H, and the LRA Novel Research Award to H.-C.H." @default.
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• W2903942609 date "2018-08-01" @default.
• W2903942609 modified "2023-09-27" @default.
• W2903942609 title "II-05 B cells from SLE patients have increased endogenous production of IFNβ which is stimulated by BCR signaling and is required for survival of autoreactive B cells" @default.
• W2903942609 doi "https://doi.org/10.1136/lupus-2018-lsm.104" @default.
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