Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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B cell signature during inactive systemic lupus

ABSTRACT: Systemic lupus erythematosous (SLE) is an autoimmune disease with an important clinical and biological heterogeneity. B lymphocytes appear central to the development of SLE which is characterized by the production of a large variety of autoantibodies and hypergammaglobulinemia. In mice, immature B cells from spontaneous lupus prone animals are able to produce autoantibodies when transferred into immunodeficient mice, strongly suggesting the existence of intrinsic B cell defects during lupus. In order to approach these defects in humans, we compared the peripheral B cell transcriptomes of quiescent lupus patients to normal B cell transcriptomes. 17 patients with quiescent lupus (patient1-17) versus 9 controls (Control1-6,Control8-10).

ORGANISM(S): Homo sapiens

SUBMITTER: Jean-Nicolas SCHICKEL

PROVIDER: E-GEOD-30153 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:T cells from many patients with systemic lupus erythematosus are hypoproliferative in response to TCR ligation. This defect is mediated, at least in part, through down regulation of the TCR zeta chain. Investigation of this phenomenon in lupus-prone mice revealed that the hypoproliferative T cell phenotype apparent in BXSB males is mediated in part by the Yaa locus. The BXSB model of lupus is a recombinant inbred strain that was originally derived from a cross between a male SB/LE mouse and a female C57BL/6 mouse. BXSB is unique among lupus-prone mouse strains, in that males carry an autoimmune accelerating factor on the Y chromosome, termed Yaa, which accelerates disease onset in male BXSB mice so that fatal glomerulonephritis occurs around 6 months of age. It was recently reported that the Yaa locus is a translocation of a short stretch of genes including that encoding Toll-like receptor 7 (Tlr-7) from the X to the Y chromosome, leading to a two-fold over expression of a subset of these genes, including Tlr-7, and hypersensitivity of Yaa+ cells to TLR-7 agonists. To more fully understand the functional significance of the Yaa gene and how it plays a part in the modulation of CD4+ T cell responses, we performed microarray analysis using Affymetrix 430 2.0 arrays on spleens from B6 male B6.Yaa male samples. The analysis revealed an enhanced signature of innate immunity, including increased expression not only of Tlr7 and Tlr8, in line with reports that the Yaa locus leads to a duplication of the murine Tlr7 and Tlr8 genes , but also of Tlr2, Tlr5, Cd14, Lbp, C2, C3 and genes for several heat-shock proteins. T cells from B6.Yaa mice also displayed decreased TCR? expression. Interestingly, cell surface levels of the inhibitory molecule B7-H1 were increased on B6.Yaa B cells and monocytes/macrophages. In line with increased TLR-7 expression reported in B6.Yaa mice, the TLR-7 agonist imiquimod increased B7-H1 expression on normal B cells. Restoration of proliferation in co-cultures of B6 T cells and B6.Yaa APC with B7-H1 blocking Ab demonstrated a mechanistic contribution of B7-H1 up-regulation to Yaa-mediated APC-dependent T cell hypoproliferation. In addition, T cell hypoproliferation was induced in normal B6 T cells in vitro by the application of imiquimod. However, this defect was not mediated by B7-H1 or secreted factors but was independent of APC and mediated through direct imiquimod stimulation of T cells themselves. We postulate that excessive TLR-7 stimulation in Yaa+ mice mediates T cell hypoproliferation in part through up-regulation of B7-H1 and possibly also through direct T cell-mediated effects. Keywords: differential gene expression, RNA, spleen, C57Bl/6J, B6.SB-Yaa/J, male, wild type, Yaa Control mice: four C57Bl/6J male spleens Experimental mice: three B6.SB-Yaa/J male spleens and one pool of two B6.Yaa male spleens for the third Yaa sample (GSM236622) All mice were obtained from the Jackson Laboratories between March and December 2005. Spleen cell suspensions were prepared by teasing spleen tissue through 70 micron nylon screens and lysing red cells with ammonium chloride. All mice were 8 weeks of age at the time of the experiment. RNA was separately extracted, labeled, and hybridized from each animal.

Project description:In lupus autoimmunity, pathogenic IgG autoantibodies that fix complement and bind FcGammaR on inflammatory cells, are produced with help from T helper (Th1 and Th17) cells specific for peptides from nucleosomes of apoptotic cells; and these Th cells also infiltrate vital organs (1-9). Macrophages (e.g. tingible body MΦ), and DCs are normally tolerant to apoptotic cell antigens (10), but they are activated to present such autoantigens after binding to IgG immune complexes (IC) containing apoptotic cell derived DNA/RNA, which then dually stimulate their TLR and FcGammaR (11-16). Hence, to generate the activating IC, IgG-switched autoantibodies have to be made first by T cell help. Moreover, B cells become efficient antigen presenting cells (APC) to Th cells pre-primed by other APC (17), or if the B cells have developed high affinity somatically mutated receptors by T cell help (18). Thus, conventional APCs participate as disease progresses, but it is unknown who initially primes autoimmune Th cells. We fractionated spleen cells of lupus prone SNF1 mice in search of such APC. To produce lupus-prone SNF1 mice, NZB and SWR mice were purchased from The Jackson Laboratory, ME to breed the lupus-prone SNF1 hybrids (19). Female SNF1 mice, like BWF1, have high serum levels of IgG class anti-DNA and other anti-nuclear autoantibodies by 2 mo, and spontaneously begin to develop severe lupus nephritis by 5 mo age (20). We depleted spleen cells of lupus-prone SNF1 mice of cells with mature lineage markers, including conventional APC, and then isolated pure CD117+ (c-Kit+ or K+) cells from the lineage– (Lin– or L–) cells. These Lin–c-Kit+pure cells, which had morphology of hematopoietic progenitor cells, were the main type of APC inducing nuclear autoantigen-specific T helper cell (Th17) response upon feeding them with nucleosomes. These Lin–c-Kit+pure cell isolate is called "LinminuscKitplus pure" in the Sample titles shown below. CD117 (c-Kit) is also a marker for macrophage/dendritic cell precursors (MDP), which also express CX3CR1 (21-23). Therefore, from T and B-cell depleted spleen cells, we sorted out CD117+CX3CR1–, CD117+CX3CR1+, CD117–CX3CR1+, and CD117–CX3CR1– cell subsets, and then tested their abilities to induce Th responses to nucleosomes. The CD117+CX3CR1– cells were Lin– (Lin–c-Kit+CX3CR1– or L–K+Cx–; named as "LinminuscKitplusCX3CR1minus" in the sample titles below) were very similar to L–K+pure cells (LinminuscKitplus pure), which are also CX3CR1–, although isolated in a different way. Therefore, we compared gene expression profiles of nucleosome-pulsed APC that were isolated in those two ways, namely purified Lin–c-Kit+ pure (LinminuscKitplus pure) cells or the Lin–c-Kit+CX3CR1– (LinminuscKitplusCX3CR1minus) cell subset, with other types of APCs, namely the Lin–c-Kit+CX3CR1+ subset (named as "LinminuscKitminusCX3CR1plus" in sample titles below), or CD11c+CD11blowc-Kit– cells (Dendritic cells in the sample titles below). REFERENCES 1. Adams, S., P. Leblanc, and S. K. Datta. 1991. Junctional region sequences of T-cell receptor b chain genes expressed by pathogenic anti-DNA autoantibody-inducing T helper cells from lupus mice: Possible selection by cationic autoantigens. Proc. Natl. Acad. Sci. USA. 88:11271-11275. 2. Mohan, C., S. Adams, V. Stanik, and S. K. Datta. 1993. Nucleosome: A major immunogen for the pathogenic autoantibody-inducing T cells of lupus. J. Exp. Med. 177:1367-1381. 3. Kaliyaperumal, A., C. Mohan, W. Wu, and S. K. Datta. 1996. 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Andrzejewski, J. Andre-Schwartz, and R. S. Schwartz. 1978. Genetic studies of autoimmunity and retrovirus expression in crosses of New Zealand Black mice. I. xenotropic virus. J. Exp. Med. 147:854-871. 21. Fogg, D. K., C. Sibon, C. Miled, S. Jung, P. Aucouturier, D. R. Littman, A. Cumano, and F. Geissmann. 2006. A clonogenic bone marrow progenitor specific for macrophages and dendritic cells. Science 311:83-87. 22. Onai, N., A. Obata-Onai, M. A. Schmid, T. Ohteki, D. Jarrossay, and M. G. Manz. 2007. Identification of clonogenic common Flt3+M-CSFR+ plasmacytoid and conventional dendritic cell progenitors in mouse bone marrow. Nat. Immunol. 8:1207-1216. 23. Waskow, C., K. Liu, G. Darrasse-Jeze, P. Guermonprez, F. Ginhoux, M. Merad, T. Shengelia, K. Yao, and M. Nussenzweig. 2008. The receptor tyrosine kinase Flt3 is required for dendritic cell development in peripheral lymphoid tissues. Nat. Immunol. 9:676-683. 24. Du, P., W. A. Kibbe, and S. M. Lin. 2008. lumi: a pipeline for processing Illumina microarray. Bioinformatics 24:1547-1548. 25. Lin, S. M., P. Du, W. Huber, and W. A. Kibbe. 2008. Model-based variance-stabilizing transformation for Illumina microarray data. Nucleic Acids Res 36:e11. 26. Du, P., W. A. Kibbe, and S. M. Lin. 2007. nuID: a universal naming scheme of oligonucleotides for illumina, affymetrix, and other microarrays. Biol Direct 2:16. 27. Wettenhall, J. M., and G. K. Smyth. 2004. limmaGUI: a graphical user interface for linear modeling of microarray data. Bioinformatics 20:3705-3706. Gene profiling microarray and data analysis. Spleen cells from 3 batches of three 5 mo SNF1 female mice each time were used to obtain three independent isolations of each type of APC population. Total RNA was purified from each APC after incubation with nucleosomes (20µg/ml) for 6h. RNA expression analysis of each type of APC isolate in triplicate was performed at our Genomics Core Facility using Illumina Mouse WG-6 v2.0 Expression Beadchips, which provides coverage of around 45,281 genes and expressed sequence tags. Raw signal intensities of each probe were obtained using data analysis software (Beadstudio; Illumina) and imported to the Lumi package of Bioconductor for data analysis. Before transformation and normalization (24-26), A/P call detection was performed based on detection p value. 20,890 out of 45,281 probes with less than 0.01 were considered as valid signals. For each pairs of four comparisons: Lin–c-Kit+CX3CR1– versus Lin–c-Kit+ CX3CR1+; Lin–c-Kit+pure vs. Lin–c-Kit+CX3CR1+; Lin–c-Kit+CX3CR1– vs. DC; and Lin–c-Kit+pure vs. DC; differentially expressed genes were identified using an Analysis of Variance (ANOVA) model with empirical Bayesian variance estimation (27). Thus, 1,619 genes were identified as being differentially expressed (up or down) on the basis of a statistically significant (raw P-value < 0.01 and false discovery rate adjusted P-value < 0.05), and 1.5-fold change (up or down) in expression level in at least one of the comparisons, and out of these 230 genes were significantly UP-regulated by the above criteria in all 4 pair comparisons.

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B cell signature during inactive systemic lupus

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