Project description:Genomic aberrations are of predominant importance to the biology and clinical outcome of patients with chronic lymphocytic leukemia (CLL), and FISH-based genomic risk classifications are routinely used in clinical decision making in CLL. One of the known limitations of CLL FISH is the inability to comprehensively interrogate the CLL genome for genomic changes. In an effort at overcoming the existing limitations in CLL genome analysis, we have analyzed high-purity DNA isolated from FACS-sorted CD19+ cells and paired CD3+ or buccal cells from 255 CLL patients for acquired genomic copy number aberrations (aCNA) using ultra-high-density Affymetrix SNP 6.0 arrays. Overall, two or more subchromosomal aCNA were found in 39% (100/255) of all cases analyzed, while ≥3 subchromosomal aCNA were detected in 20% (50/255) of cases. Subsequently, we have correlated genomic lesion loads (genomic complexity) with the clinical outcome measures time to first therapy (TTFT) and overall survival (OS). Using multivariate analyses incorporating the most important prognostic factors in CLL together with SNP 6.0 array-based genomic lesion loads at various thresholds, we identify elevated CLL genomic complexity as an independent and powerful marker for the identification of CLL patients with aggressive disease and short survival. we have analyzed high-purity DNA isolated from FACS-sorted CD19+ cells and paired CD3+ or buccal cells from 255 CLL patients for acquired genomic copy number aberrations (aCNA) using ultra-high-density Affymetrix SNP 6.0 arrays
Project description:CD4+ T cells (T helper cells) are cytokine-producing adaptive immune cells that activate or regulate the responses of different immune cells. They are known to play crucial roles in antibody class switching in B cells, neutrophil recruitment and activation of macrophages and CD8+ cytotoxic T cells. The activation and functional status of CD4+ T cells is important for adequate responses to pathogen infections but has also been associated with auto-immune disorders and survival in several cancers. In the current study, we carried out proteomic profiling of resting and activated primary human CD4+ T cells from healthy donors. In addition to identifying known markers of CD4+ T cell activation, we also identified protein kinases, protein phosphatases, and cytokines to be differentially expressed.
Project description:Promiscuous gene expression (pGE) of tissue-restricted antigens in medullary thyme ephitelial cells (mTECs) is essential for self-tolerance induction and to prevent autoimmunity. We sequenced single mTEC transcriptomes to explore gene expression heterogeneity and to discover patterns of regulation of pGE.
Project description:Promiscuous gene expression (pGE) of tissue-restricted antigens in medullary thyme ephitelial cells (mTECs) is essential for self-tolerance induction and to prevent autoimmunity. We sequenced single mTEC transcriptomes to explore gene expression heterogeneity and to discover patterns of regulation of pGE. This data set complements what's submitted earlier, under ArrayExpress accession E-MTAB-3346 ( http://www.ebi.ac.uk/arrayexpress/experiments/E-MTAB-3346/ ).
Project description:We explored how aging impacts transcriptional dynamics using single-cell RNA-sequencing to profile hundreds of CD4+ T cells from young and old mice from two divergent species. In young animals, immunological challenge drives a conserved transcriptomic switch from highly variable to tightly regulated gene expression, characterized by a strong up-regulation of a core activation program, coupled with a decrease in cell-to-cell variability. Aging significantly perturbed the activation of this core program, and increased expression heterogeneity across the population of cells in both species.
Project description:The identification of gene mutation and structural genomic aberrations that are critically involved in CLL pathogenesis is still evolving. One may postulate that genomic driver lesions with effects on CLL proliferation, apoptosis thresholds, or chemotherapy resistance should increase in frequency over time when measured sequentially in a large CLL cohort. We sequentially sampled a large, well-characterized CLL cohort at a mean of 4 years between samplings. The paired analysis included 156 patients, of whom 114 remained untreated and 42 received intercurrent therapies. Results: we identify a strong effect of intercurrent therapies on the frequency of acquisition of aCNAs in CLL. Importantly, the spectrum of acquired genomic changes was largely similar in patients that did or did not receive intercurrent therapies; therefore, various genomic changes that become part of the dominant clones are often already present in CLL cell populations prior to therapy. Further, we provide evidence that therapy of CLL with preexisting TP53 mutations results in the outgrowth of genomically very complex clones which dominate at relapse. Using complementary technologies directed at the detection of genomic events that are present in substantial proportions of of the clinically relevant CLL disease bulk, we capture aspects of genomic evolution in CLL over time, including increases in the frequency of genomic complexity, specific recurrent aCNAs, and TP53 mutations. Data from 156 paired samples (enrollment and one longitudinal sample) are included in this data set. Of these, 27 patients were assayed at two (or, rarely, more than two) time points. Please note: normal DNA and enrollment date tumor DNA CEL files and SNP call files will be found in a separate GEO data submission: GSE30777
Project description:The purpose of the study was to examine the role of the IRE1a-XBP1 pathway during Th2 lymphocyte activation and differentiation. In vitro Th2 cells were treated with 4μ8c, a drug that specifically inhibits IRE1a endonuclease activity, and transcriptomes were compared.
Project description:IL-27 is a potent antagonist of TH1-mediated inflammation, but the basis for this effect is not fully understood. Recent studies identified a population of T-bet+ CXCR3+ Treg that limit TH1-mediated immune pathology. The studies presented here demonstrate that IL-27-mediated STAT1 activation promotes Treg expression of T-bet and CXCR3. Infection with Toxoplasma gondii induced a similar Treg population that limits T cell responses and this population at mucosal sites is IL-27-dependent. Furthermore, transfer of Treg ameliorated the infection-induced CD4+ T cell-mediated pathology observed in IL-27p28-/- mice. Although IFN-γ promoted a similar population of cells in the periphery, it did not compensate for the absence of IL-27 at mucosal sites and microarray analysis revealed that Treg exposed to either cytokine have distinct transcriptional profiles. These findings suggest that IFN-γ and IL-27 have different roles in Treg biology but define IL-27 as a key cytokine that promotes the development of Treg specialized to control TH1 immunity. Three conditions were analyzed across two timepoints. Inducible regulatory T cells (iTreg) were generated in vitro in the presence of IL-27, IFNg or under 'Neutral' conditions as a control. Samples were collected at 10 hours and 2 days during the culture period. Three biological replicates were used for each condition.
Project description:Activation of CD8+ T cells depends exquisitely on the affinity of the T cell receptor (TCR) for a peptide MHC (pMHC) ligand complex. Here, we activated OT-I transgenic CD8+ T cells with pure peptide and examined early activation responses by single-cell RNA-sequencing. T cells were activated with the high affinity OT-I cognate peptide (N4=SIINFEKL) for 1, 3 or 6 hours, or with reduced affinity peptides (T4=SIITFEKL and G4=SIIGFEKL) or the non-binding peptide (NP68=ASNENMDAM) for 6 hours. Cells were then sorted into 96-well plates by FACS and RNA was sequenced following an adapted Smart-Seq2 protocol.
Project description:In this study, we have investigated the effect of LMP1 on gene expression in normal human GC B cells using a non-viral vector based system Experiment Overall Design: Gene expression was compared between LMP1-transfected and control vector-transfected GC B cells from three patients. RNA from the FACS-sorted transfected GC B cells was amplified. 10ug of fragmented cRNA was hybridized to HG-U133 Plus 2.0 microarrays. Differentially expressed genes were identified using significance analysis of microarrays (SAM) with a 1.5 fold change threshold and the q-value threshold set to 5%.