Search results for "Cytometry"
showing 10 items of 852 documents
Increasing functional avidity of TCR-redirected T cells by removing defined N-glycosylation sites in the TCR constant domain
2009
Adoptive transfer of T lymphocytes transduced with a T cell receptor (TCR) to impart tumor reactivity has been reported as a potential strategy to redirect immune responses to target cancer cells (Schumacher, T.N. 2002. Nat. Rev. Immunol. 2:512-519). However, the affinity of most TCRs specific for shared tumor antigens that can be isolated is usually low. Thus, strategies to increase the affinity of TCRs or the functional avidity of TCR-transduced T cells might be therapeutically beneficial. Because glycosylation affects the flexibility, movement, and interactions of surface molecules, we tested if selectively removing conserved N-glycoslyation sites in the constant regions of TCR alpha or …
PD-1 signalling in CD4+T cells restrains their clonal expansion to an immunogenic stimulus, but is not critically required for peptide-induced tolera…
2010
Summary The ultimate outcome of T-cell recognition of peptide–major histocompatibility complex (MHC) complexes is determined by the molecular context in which antigen presentation is provided. The paradigm is that, after exposure to peptides presented by steady-state dendritic cells (DCs), inhibitory signals dominate, leading to the deletion and/or functional inactivation of antigen-reactive T cells. This has been utilized in a variety of models providing peptide antigen in soluble form in the absence of adjuvant. A co-inhibitory molecule of considerable current interest is PD-1. Here we show that there is the opportunity for the PD-1/PD-L1 interaction to function in inhibiting the T-cell r…
A new model of chronic colitis in SCID mice induced by adoptive transfer of CD62L+ CD4+ T cells: insights into the regulatory role of interleukin-6 o…
2003
<i>Objective:</i> The proinflammatory cytokine interleukin (IL)-6 is involved in various chronic inflammatory processes. IL-6 is a predominant cytokine produced by lamina propria T cells in Crohn’s disease and experimental colitis. This study was designed to examine the effect of a neutralizing IL-6-receptor (IL-6R) antibody on the programmed cell death of mucosal T cells in the CD62L+ CD4+ SCID transfer model of chronic experimental colitis in mice and to gain more insight into the pathogenesis of this transfer colitis model. <i>Methods:</i> For adoptive transfer, we isolated CD62L+ CD4+ double-positive T cells from wild-type BALB/c mice followed by intraperitoneal …
Cooperation of Human Tumor-Reactive CD4+ and CD8+ T Cells after Redirection of Their Specificity by a High-Affinity p53A2.1-Specific TCR
2004
Abstract Efficient immune attack of malignant disease requires the concerted action of both CD8 + CTL and CD4 + Th cells. We used human leukocyte antigen (HLA)-A*0201 (A2.1) transgenic mice, in which the mouse CD8 molecule cannot efficiently interact with the α3 domain of A2.1, to generate a high-affinity, CD8-independent T cell receptor (TCR) specific for a commonly expressed, tumor-associated cytotoxic T lymphocyte (CTL) epitope derived from the human p53 tumor suppressor protein. Retroviral expression of this CD8-independent, p53-specific TCR into human T cells imparted the CD8 + T lymphocytes with broad tumor-specific CTL activity and turned CD4 + T cells into potent tumor-reactive, p53…
Induction of Interleukin 10–Producing, Nonproliferating Cd4+ T Cells with Regulatory Properties by Repetitive Stimulation with Allogeneic Immature Hu…
2000
The functional properties of dendritic cells (DCs) are strictly dependent on their maturational state. To analyze the influence of the maturational state of DCs on priming and differentiation of T cells, immature CD83− and mature CD83+ human DCs were used for stimulation of naive, allogeneic CD4+ T cells. Repetitive stimulation with mature DCs resulted in a strong expansion of alloreactive T cells and the exclusive development of T helper type 1 (Th1) cells. In contrast, after repetitive stimulation with immature DCs the alloreactive T cells showed an irreversibly inhibited proliferation that could not be restored by restimulation with mature DCs or peripheral blood mononuclear cells, or by…
Human Papillomavirus Type 33 E7 Peptides Presented by HLA-DR*0402 to Tumor-Infiltrating T Cells in Cervical Cancer
2000
ABSTRACTSeveral characteristics make human papillomavirus (HPV) amenable to vaccination. Anti-HPV-directed vaccines are based on the observation that HPV E6 and E7 oncoproteins are constitutively expressed in HPV-positive cervical cancer and may serve as tumor rejection antigens. Five HPV types (16, 18, 31, 33, and 45) account for 80% of cervical cancer. Until now, the type of immune response capable of mediating an effective antitumor response has not been defined. In order to define the anticancer-directed immune response in situ, we characterized CD4+and CD8+sorted T cells from peripheral blood lymphocytes, freshly harvested tumor tissue, and tumor-infiltrating lymphocytes (TIL) from a p…
Optimized Protocol for the Detection of Multifunctional Epitope-Specific CD4+ T Cells Combining MHC-II Tetramer and Intracellular Cytokine Staining T…
2019
Analysis of multifunctional CD4+ T cells is fundamental for characterizing the immune responses to vaccination or infection. Major histocompatibility complex (MHC)/peptide tetramers represent a powerful technology for the detection of antigen-specific T cells by specific binding to their T-cell receptor, and their combination with functional assays is fundamental for characterizing the antigen-specific immune response. Here we optimized a protocol for the detection of multiple intracellular cytokines within epitope-specific CD4+ T cells identified by the MHC class II tetramer technology. The optimal procedure for assessing the functional activity of tetramer-binding CD4+ T cells was based o…
Monitoring of anti-vaccine CD4 T cell frequencies in melanoma patients vaccinated with a MAGE-3 protein.
2005
Abstract Quantitative evaluation of T cell responses of patients receiving antitumoral vaccination with a protein is difficult because of the large number of possible HLA-peptide combinations that could be targeted by the response. To evaluate the responses of patients vaccinated with protein MAGE-3, we have developed an approach that involves overnight stimulation of blood T cells with autologous dendritic cells loaded with the protein, sorting by flow cytometry of the T cells that produce IFN-γ, cloning of these cells, and evaluation of the number of T cell clones that secrete IFN-γ upon stimulation with the Ag. An important criterion is that T cell clones must recognize not only stimulat…
Nanoparticles and antigen-specific T-cell therapeutics: A comprehensive study on uptake and release
2015
Aim: T lymphocytes are used as cellular therapeutics in many disease entities including cancer. We investigated the uptake and retention of nanoparticles (NPs) by these nonphagocytic cells. Materials & methods: Uptake, release and toxicity of various polymeric NP preparations were analyzed by flow cytometry, confocal laser scanning microscopy and transmission electron microscopy. T-cell effector functions were measured using IFN-γ-ELISPOT and 51Chromium-release assays. Results: Amino-functionalized NPs were efficiently ingested by antigen-specific T cells without adversely influencing effector functions. NPs were stored in membrane-surrounded vesicles, with major proportions released e…
Microenvironment Regulation of IL23R/IL-23 Axis Drives Chronic Lymphocytic Leukemia (CLL) Progression
2015
Abstract Background : CLL displays a considerable degree of clinical heterogeneity, which is in part ascribable to clone-intrinsic biological features and that are also influenced by clone-extrinsic events related to the microenvironment. Among the dynamics-taking place within the CLL microenvironment, those finalized to the induction of an overly inflammatory milieu may significantly impact on the CLL natural history by hijacking the immunological microenvironment at the same time fostering clone fitness. IL-23 acts as a prototypical pro-inflammatory mediator representing a promising therapeutic target. We analyzed the ability of CLL cells to sense IL-23 through the IL-23R complex (consist…