scholarly journals Enumeration of viral antigen-reactive helper T lymphocytes in human peripheral blood by limiting dilution for analysis of viral antigen-reactive T-cell pools in virus-seropositive and virus-seronegative individuals.

1989 ◽  
Vol 27 (10) ◽  
pp. 2316-2323 ◽  
Author(s):  
K A Clouse ◽  
P W Adams ◽  
C G Orosz
Keyword(s):  
T Cell ◽  
T Lymphocytes ◽  
Peripheral Blood ◽  
Viral Antigen ◽  
Human Peripheral Blood ◽  
Helper T Lymphocytes ◽  
Limiting Dilution

Differential chemokine expression profiles in human peripheral blood T lymphocytes: dependence on T-cell coreceptor and calcineurin signaling

Blood ◽  
2003 ◽  
Vol 101 (1) ◽  
pp. 216-225 ◽  
Author(s):  
Anthony D. Cristillo ◽  
Mirtha J. Macri ◽  
Barbara E. Bierer
Keyword(s):  
T Cell ◽  
T Lymphocytes ◽  
Signaling Pathways ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Expression Profiles ◽  
Lymphoid Tissues ◽  
Rnase Protection ◽  
Chemokine Expression ◽  
Calcineurin Activity

Abstract The chemokine superfamily consists of small (8-10 kDa) molecules that function to attract, selectively, different subsets of leukocytes. Binding of chemokines to their appropriate G-protein–coupled receptors is necessary for primary immune responses and for homing of leukocytes to lymphoid tissues. Here, we have characterized the signaling pathways in primary T lymphocytes that regulate chemokine gene induction using an RNase protection assay. Dependence on stimulation through the coreceptor CD28 and sensitivity to the calcineurin inhibitors cyclosporine and tacrolimus were studied using purified human peripheral blood lymphocytes. Lymphotactin (Ltn), macrophage inflammatory protein (MIP)–1α, and MIP-1β were all rapidly induced and sensitive to cyclosporine treatment. At later time points, the expression of MIP-1α and MIP-1β, but not of Ltn, was restored despite the inhibition of calcineurin activity. By contrast, the induction of interleukin-8 was delayed and was found to be cyclosporine insensitive. Calcineurin activity of IP-10 mRNA induction was contingent on the specific T-cell stimulation conditions, suggesting that IP-10 expression is modulated by calcineurin-dependent and -independent signaling pathways. Differential chemokine expression profiles result from the engagement of T-cell coreceptors and the requirement for, and the dependence on, calcineurin phosphatase activity.

scholarly journals Human natural killer cells isolated from peripheral blood do not rearrange T cell antigen receptor beta chain genes.

1986 ◽  
Vol 163 (1) ◽  
pp. 209-214 ◽  
Author(s):  
L L Lanier ◽  
S Cwirla ◽  
N Federspiel ◽  
J H Phillips
Keyword(s):  
T Cell ◽  
T Lymphocytes ◽  
Nk Cells ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Antigen Receptor ◽  
T Cell Antigen Receptor ◽  
Cell Antigen Receptor ◽  
Cell Antigen ◽  
Beta Chain Genes

The lineage of NK cells and their relationship to T lymphocytes have been controversial issues. Since rearrangement of the T cell antigen receptor beta chain genes occurs early in the ontogeny and differentiation of all T cells, this can be used as an unequivocal marker to discriminate T from non-T lymphocytes. Recent studies (16-18) examining T cell antigen receptor gene rearrangement and expression in certain IL-2-dependent NK cell lines and leukemias have revealed that some lines rearrange C beta genes, whereas others do not. However, it is important to establish whether these cell lines are representative of the major population of NK cells freshly derived from the host. Herein, we have purified granulocytes, CD16+ NK cells and T lymphocytes from human peripheral blood, prepared genomic DNA from each cell type, and then examined the organization of their T cell antigen receptor genes by restriction enzyme analysis using a C beta cDNA as probe. The C beta genes were in germline configuration in NK cells and granulocytes. In contrast, peripheral blood T lymphocytes showed rearrangement of the C beta gene. These data support the hypothesis that the majority of human peripheral blood NK cells are fundamentally distinct from T lymphocytes in lineage and nonself recognition.

scholarly journals Diacylglycerol activates the influx of extracellular cations in T-lymphocytes independently of intracellular calcium-store depletion and possibly involving endogenous TRP6 gene products

2002 ◽  
Vol 364 (1) ◽  
pp. 245-254 ◽  
Author(s):  
Alessandra GAMBERUCCI ◽  
Emanuele GIURISATO ◽  
Paola PIZZO ◽  
Maristella TASSI ◽  
Roberta GIUNTI ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
T Lymphocytes ◽  
Peripheral Blood ◽  
Transient Receptor Potential ◽  
Human Peripheral Blood ◽  
Receptor Potential ◽  
Animal Cells ◽  
Bivalent Cation ◽  
Intracellular Calcium Store ◽  
Transient Receptor

In Jurkat and human peripheral blood T-lymphocytes, 1-oleoyl-2-acetyl-sn-glycerol (OAG), a membrane-permeant analogue of diacylglycerol, activated the influx of Ca2+, Ba2+ and Sr2+. OAG also caused plasma-membrane depolarization in Ca2+-free media that was recovered by the addition of bivalent cation, indicating the activation of Na+ influx. OAG-induced cation influx was (i) mimicked by the natural dacylglycerol 1-stearoyl-2-arachidonyl-sn-glycerol, (ii) not blocked by inhibiting protein kinase C or in the absence of phopholipase C activity and (iii) blocked by La3+ and Gd3+. Differently from OAG, both thapsigargin and phytohaemagglutinin activated a potent influx of Ca2+, but little influx of Ba2+ and Sr2+. Moreover, the influx of Ca2+ activated by thapsigargin and that activated by OAG were additive. Furthermore, several drugs (i.e. econazole, SKF96365, carbonyl cyanide p-trifluoromethoxyphenylhydrazone, 2-aminoethoxy diphenylborate and calyculin-A), while inhibiting the influx of Ca2+ induced by both thapsigargin and phytohaemagglutinin, did not affect OAG-stimulated cation influx. Transient receptor potential (TRP) 3 and TRP6 proteins have been shown previously to be activated by diacylglycerol when expressed heterologously in animal cells [Hofmann, Obukhov, Schaefer, Harteneck, Gudermann and Schultz (1999) Nature (London) 397, 259–263]. In both Jurkat and peripheral blood T-lymphocytes, mRNA encoding TRP proteins 1, 3, 4 and 6 was detected by reverse transcriptase PCR, and the TRP6 protein was detected by Western blotting in a purified plasma-membrane fraction. We conclude that T-cells express a diacylglycerol-activated cation channel, unrelated to the channel involved in capacitative Ca2+ entry, and associated with the expression of TRP6 protein.

scholarly journals Quantitative assessment of the pool size and subset distribution of cytolytic T lymphocytes within human resting or alloactivated peripheral blood T cell populations.

1983 ◽  
Vol 158 (2) ◽  
pp. 571-585 ◽  
Author(s):  
A Moretta ◽  
G Pantaleo ◽  
L Moretta ◽  
M C Mingari ◽  
J C Cerottini
Keyword(s):  
Monoclonal Antibody ◽  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Peripheral Blood ◽  
Great Majority ◽  
T Cell Subsets ◽  
Cytolytic T Lymphocytes ◽  
Subset Distribution

In order to directly assess the distribution of cytolytic T lymphocytes (CTL) and their precursors (CTL-P) in the two major subsets of human T cells, we have used limiting dilution microculture systems to determine their frequencies. The two subsets were defined according to their reactivity (or lack thereof) with B9.4 monoclonal antibody (the specificity of which is similar, if not identical, to that of Leu 2b monoclonal antibody). Both B9+ and B9- cells obtained by sorting peripheral blood resting T cells using the fluorescence-activated cell sorter (FACS) were assayed for total CTL-P frequencies in a microculture system that allows clonal growth of every T cell. As assessed by a lectin-dependent assay, approximately 30% of peripheral blood T cells were CTP-P. In the B9+ subset (which represents 20-30% of all T cells), the CTL-P frequency was close to 100%, whereas the B9- subset had a 25-fold lower CTL-P frequency. It is thus evident that 90% and 10% of the total CTL-P in peripheral blood are confined to the B9+ or B9- T cell subsets, respectively. Analysis of the subset distribution of CTL-P directed against a given set of alloantigens confirmed these findings. CTL-P frequencies were also determined in B9+ and B9- subsets derived from T cells that had been activated in allogenic mixed leucocyte cultures (MLC). Approximately 10% of MLC T cells were CTL-P. This frequency was increased 3.5-fold in the B9+ subset, whereas the B9- subset contained only a small, although detectable number of CTL-P. Moreover, the great majority of the (operationally defined) CTL-P in MLC T cell population were found to be directed against the stimulating alloantigens, thus indicating a dramatic increase in specific CTL-P frequencies following in vitro stimulation in bulk cultures.

scholarly journals Expression of IκBα in the nucleus of human peripheral blood T lymphocytes

Oncogene ◽  
1999 ◽  
Vol 18 (8) ◽  
pp. 1581-1588 ◽  
Author(s):  
Teresa Laín de Lera ◽  
Lola Folgueira ◽  
Angel G Martín ◽  
Catherine Dargemont ◽  
María-Antonia Pedraza ◽  
...  
Keyword(s):  
T Lymphocytes ◽  
Peripheral Blood ◽  
Human Peripheral Blood

PBMC-07- Stimulation of human peripheral blood mononuclear cells with immobilized anti-CD3 and soluble anti-CD28 antibodies. v1

2021 ◽  
Author(s):  
Alessia Furgiuele ◽  
Massimilano Legnaro ◽  
Alessandra Luini ◽  
Marco Ferrari ◽  
Emanuela Rasini ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Activation ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Cell Activation ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Functional Responses ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

This protocol was designed to activate the lymphocytes T of a population of peripheral blood mononuclear cells (PBMCs), simulating their physiological response to antigen/MHC complex acting on T Cell Receptors-TCR , in order to test their functional responses including cell proliferation and cytokine production. The co-stimulation protocol include: i)anti-CD3 antibody a polyclonal activator specific for invariant framework epitopes on TCR complex (in particular, we use UCHT1 clone an anti-human CD3 antibody that recognizes the ε-chain of CD3 which is used for immobilized option of activation) (http://static.bdbiosciences.com/documents/BD_Tcell_Human_CD3_Activation_Protocol.pdf) ii) anti-CD28 antibody used to cooperate with TCR signals promoting activation of T cells The procedure has been reproduced following the indications contained in the protocol of "EBiooscience" (https://tools.thermofisher.com/content/sfs/manuals/t-cell-activation-in-vitro.pdf). Pilot experiments on PBMC were carried out to determine the best concentrations of anti-CD3 and anti-CD28 to induce optimal proliferation of PBMC and production of cytokines TNF-α and IFN-γ. We found a dose dependent correlation between immobilized anti-CD3 and cells functional responses. The selected amount was 2 µg/mL for both anti-CD3 and anti-CD28 that was the concentration below the maximum response which allows also to test possible modulations by therapeutic agents. References http://static.bdbiosciences.com/documents/BD_Tcell_Human_CD3_Activation_Protocol.pdf https://tools.thermofisher.com/content/sfs/manuals/t-cell-activation-in-vitro.pdf https://www.bdbiosciences.com/ds/pm/tds/555330.pdf https://www.bdbiosciences.com/ds/pm/tds/555726.pdf BEFORE STARTING with this procedure Moreover, work under laminar flow hood when you are processing samples from the beginning to the end of the culture. Make sure you are using,sterile culture mediumand sterile plastic disposable as well.

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