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Flow Cytometric Measurement of SLAM-Associated Protein and X-Linked Inhibitor of Apoptosis

Flow cytometry is a valuable tool for the detection and characterization of proteins expressed by individual cells. Flow cytometry can be used to measure cell expression of 2 intracellular proteins that are involved in the regulation of immune homeostasis, SLAM-associated protein (S ...

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Designs for Massively Parallel Sequencing Approaches to Identify Causal Mutations in Human Immune Disorders

Massively parallel sequencing technologies provide new opportunities to discover causal variants and �narrow down candidate genes responsible for human Mendelian disorders. Such information can in turn provide new insights into understanding the basic science behind, as we ...

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Isolation of RNA and the Synthesis and Amplification of cDNA from Antigen-Specific T Cells for Genome-Wide Expression Analysis

Genome-wide gene expression analysis has become a very powerful routine tool for the study of distinct differentiation states. However, the examination of total populations of cells that contain high levels of heterogeneity, such as the total CD8+ T cell population during an immune respo ...

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Molecular Measurement of T Cell Receptor Excision Circles

This chapter provides protocols necessary for quantifying human, mouse, and nonhuman primate signal joint T cell receptor excision circles (sjTRECs) produced during T cell receptor alpha (TCRA) gene rearrangement. These non-replicated episomal circles of DNA are generated by the r ...

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Real-Time Quantitative (RQ-)PCR Approach to Quantify the Contribution of Proliferation to B Lymphocyte Homeostasis

The cells of the adaptive immune system, B and T lymphocytes, each generate a unique antigen receptor through V(D)J recombination of their immunoglobulin (Ig) and T-cell receptor (TCR) loci, respectively. Such rearrangements join coding elements to form a coding joint and delete the interv ...

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Quantitating Lymphocyte Homeostasis In Vivo in Humans Using Stable Isotope Tracers

Humans have a remarkable ability to maintain relatively constant lymphocyte numbers across many decades, from puberty to old-age, despite a multitude of infectious and other challenges and a dramatic decline in thymic output. This phenomenon, lymphocyte homeostasis, is achieved by m ...

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Plasmacytoid Dendritic Cells in Tolerance

Dendritic cells (DC) are professional antigen-presenting cells (APCs) that modulate the outcome of the immune response toward immunity or tolerance. There are a large variety of DC subsets according to surface phenotype, function, and tissue distribution. Murine plasmacytoid DC (p ...

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DCs in Immune Tolerance in Steady-State Conditions

Dendritic cells (DCs) are antigen-presenting cells (APCs) characterized by a unique capacity to stimulate na�ve T cells and initiate primary immune responses. Recent studies suggest that DCs are also involved in the induction of immunological tolerance in peripheral tissues under s ...

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Identifying Regulatory B Cells (B10 Cells) that Produce IL-10 in Mice

Regulatory B cells that produce IL-10 are now recognized as an important component of the immune system. We have identified a rare antigen-specific regulatory B-cell subset with a unique CD1dhiCD5+CD19hi phenotype in the spleens of wild-type mice. We call these cells B10 cells because they are ...

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Regulatory CD4 CD8 Double Negative T Cells

Peripheral αβTCR+CD3+CD4–CD8– NK1.1/CD56– double-negative (DN) Treg cells are a relatively rare subset of regulatory cells found in both humans and mice, typically comprising less than 5% of the total peripheral T-cell pool. Numerous studies have shown that DN Tregs can inhibit CD4+ and CD8+ T-c ...

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Phenotypic and Functional Characterization of CD8+ T Regulatory Cells

Increasing evidence shows the presence and significance of CD8+ T regulatory cells (CD8+ Tregs) in both human and rodent transplant recipients, as well as in autoimmune disease models. We, hereafter, review all available data on the phenotypic and functional characterization of CD8+ Treg ...

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Ex Vivo Generation of Regulatory T Cells: Characterization and Therapeutic Evaluation in a Model of Chronic Colitis

Naturally occurring regulatory T cells (nTregs; CD4+CD25+Foxp3+) are capable of suppressing the chronic inflammation observed in a variety of different animal models of autoimmune and chronic inflammatory diseases such as inflammatory bowel diseases, diabetes, and arthritis. A ...

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Immunoregulatory Properties of Heme Oxygenase-1

Heme oxygenase-1 (HO-1) is one of the three isoforms of the heme oxygenase enzyme that catabolyzes the degradation of heme into biliverdin with the production of free iron and CO. HO-1 is induced by its substrate and by other stimuli, including agents involved in oxidative stress and proinflammat ...

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Induction of Tolerogenic Dendritic Cells by NF-B Blockade and Fc Receptor Modulation

Autoimmune diseases develop as a result of an unbalanced adaptive immunity that targets self-antigens and causes destruction of healthy host tissues. Maintenance of peripheral immune tolerance to self- antigens is mainly mediated by dendritic cells (DCs), professional antigen- ...

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Transforming Growth Factor-Beta: Recent Advances on Its Role in Immune Tolerance

Transforming growth factor (TGF- β1) is a pleiotropic cytokine, secreted by immune and nonhematopoietic cells. TGF-β is involved in many different critical processes, such as embryonal development, cellular maturation and differentiation, wound healing, and immune regulation. ...

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Regulatory T Cell Enrichment by IFN- Conditioning

IFN-γ was originally characterized as a proinflammatory cytokine with T helper type 1 inducing activity, but it is now clear that it also has important immunoregulatory functions. Regulatory T cells play an important role in models of autoimmunity, GVHD, and transplantation, and offer pot ...

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Indoleamine 2,3-Dioxygenase and Regulatory Function: Tryptophan Starvation and Beyond

Indoleamine 2,3-dioxygenase (IDO) is an ancestral enzyme that, initially confined to the regulation of tryptophan availability in local tissue microenvironments, is now considered to play a wider role that extends to homeostasis and plasticity of the immune system. Thus, IDO biology h ...

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Fine-Tuning Antitumor Responses Through the Control of GalectinGlycan Interactions: An Overview

In recent years, we have witnessed critical advances in genomics and proteomics which contributed to delineate the “tumor progression signature”. This includes the altered expression of genes and proteins not only in tumor cells, but also in tumor-associated stromal, endothelial, and ...

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Regulation of Lymphocytes by Nitric Oxide

Shortly after the identification of nitric oxide (NO) as a product of macrophages, it was discovered that NO generated by inducible NO synthase (iNOS) inhibits the proliferation of T lymphocytes. Since then, it has become clear that iNOS activity also regulates the development, differenti ...

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Organ Transplantation: Modulation of T-Cell Activation Pathways Initiated by Cell Surface Receptors to Suppress Graft Rejection

T-cell activation depends upon two types of signals: a T-cell-receptor-mediated antigen-specific signal and several non-antigen-specific ones provided by the engagement of costimulatory and/or inhibitory T-cell surface molecules. In clinical transplantation, T-cell co ...

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