scholarly journals The effect of the immunophilin ligands rapamycin and FK506 on proliferation of mast cells and other hematopoietic cell lines.

1992 ◽  
Vol 3 (9) ◽  
pp. 981-987 ◽  
Author(s):  
T Hultsch ◽  
R Martin ◽  
R J Hohman
Keyword(s):  
Signal Transduction ◽  
Mast Cells ◽  
Cell Line ◽  
Cytotoxic T Lymphocyte ◽  
Cell Types ◽  
Cell Receptor ◽  
Immunosuppressive Drugs ◽  
Specific Cell ◽  
Fk506 Binding Protein ◽  
Rat Basophilic Leukemia Cells

The immunosuppressive drugs FK506 and cyclosporin A have an identical spectrum of activities with respect to IgE receptor (Fc epsilon RI)-mediated exocytosis from mast cells and T cell receptor-mediated transcription of IL-2. These findings suggest a common step in receptor-mediated signal transduction leading to exocytosis and transcription and imply that immunosuppressive drugs target specific signal transduction pathways, rather than specific cell types. This hypothesis is supported by studies on the effect of rapamycin on IL-3 dependent proliferation of the rodent mast cell line PT18. Rapamycin inhibits proliferation of PT18 cells, achieving a plateau of 80% inhibition at 1 nM. This inhibition is prevented in a competitive manner by FK506, a structural analogue of rapamycin. Proliferation of rat basophilic leukemia cells and WEHI-3 cells was also inhibited, at doses comparable to those shown previously to inhibit IL-2-dependent proliferation of cytotoxic T lymphocyte line (CTLL) cells. In contrast, proliferation of A-431 cells, a epidermoid cell line, was not affected by rapamycin. DNA histograms indicate that complexes formed between the rapamycin-FK506-binding protein (FKBP) and rapamycin arrest-proliferating PT18 cells in the G0/G1-phase. It is concluded that FKBP-rapamycin complexes may inhibit proliferative signals emanating from IL-3 receptors, resulting in growth arrest of cytokine-dependent, hematopoietic cells.

PKCθ-regulated signalling in health and disease

2014 ◽  
Vol 42 (6) ◽  
pp. 1484-1489 ◽  
Author(s):  
Pulak R. Nath ◽  
Noah Isakov
Keyword(s):  
Signal Transduction ◽  
T Cells ◽  
T Cell ◽  
Immunological Synapse ◽  
Cell Types ◽  
Cell Receptor ◽  
Nuclear Factor Κb ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Health And Disease

Protein kinase Cθ (PKCθ) is a key enzyme in T-lymphocytes where it plays an important role in signal transduction downstream of the activated T-cell receptor (TCR) and the CD28 co-stimulatory receptor. Antigenic stimulation of T-cells triggers PKCθ translocation to the centre of the immunological synapse (IS) at the contact site between antigen-specific T-cells and antigen-presenting cells (APCs). The IS-residing PKCθ phosphorylates and activates effector molecules that transduce signals into distinct subcellular compartments and activate the transcription factors, nuclear factor κB (NF-κB), nuclear factor of activated T-cells (NFAT) and activating protein 1 (AP-1), which are essential for the induction of T-cell-mediated responses. Besides its major biological role in T-cells, PKCθ is expressed in several additional cell types and is involved in a variety of distinct physiological and pathological phenomena. For example, PKCθ is expressed at high levels in platelets where it regulates signal transduction from distinct surface receptors, and is required for optimal platelet activation and aggregation, as well as haemostasis. In addition, PKCθ is involved in physiological processes regulating insulin resistance and susceptibility to obesity, and is expressed at high levels in gastrointestinal stromal tumours (GISTs), although the functional importance of PKCθ in these processes and cell types is not fully clear. The present article briefly reviews selected topics relevant to the biological roles of PKCθ in health and disease.

scholarly journals Immunomagnetic Isolation of Rat Bone Marrow-derived and Peritoneal Mast Cells

1997 ◽  
Vol 45 (12) ◽  
pp. 1715-1722 ◽  
Author(s):  
Maria Celia Jamur ◽  
Ana Cristina G. Grodzki ◽  
Andrea N. Moreno ◽  
William D. Swaim ◽  
Reuben P. Siraganian ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mast Cell ◽  
Mast Cells ◽  
Cell Types ◽  
Ultrastructural Level ◽  
Immunomagnetic Separation ◽  
Specific Cell ◽  
Morphological Examination ◽  
Immunomagnetic Isolation ◽  
Rat Bone

Mast cells are difficult to purify from heterogeneous cell populations and to preserve, especially for pre-embedding immunostaining at the ultrastructural level. We have developed a technique that permits the isolation of a pure population of mast cells suitable for immunocytochemical studies. A rat mast cell-specific monoclonal antibody (MAb AA4) conjugated to tosylactivated Dynabeads 450 was used to immunomagnetically separate mast cells from rat bone marrow and peritoneal cell suspensions. Approximately 85% of the mast cells were recovered in the positive population that comprised virtually pure mast cells. After microwave fixation, morphological examination showed that the cells were intact and retained their ultrastructural detail. Mast cells in all stages of maturation were immunolabeled with a panel of antibodies after immunomagnetic separation. The combination of immunomagnetic separation followed by immunostaining should prove useful for the study of mast cell maturation and for the characterization of other specific cell types that are present in tissues in only limited numbers.

scholarly journals Zika Virus Infects Human Placental Mast Cells and the HMC-1 Cell Line, and Triggers Degranulation, Cytokine Release and Ultrastructural Changes

Cells ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 975 ◽  
Author(s):  
Kíssila Rabelo ◽  
Antônio José da Silva Gonçalves ◽  
Luiz José de Souza ◽  
Anna Paula Sales ◽  
Sheila Maria Barbosa de Lima ◽  
...  
Keyword(s):  
Mast Cells ◽  
Cell Line ◽  
Zika Virus ◽  
Post Infection ◽  
Immune Cell ◽  
Cell Types ◽  
Ultrastructural Changes ◽  
Congenital Defects ◽  
Zikv Infection ◽  
Intracellular Changes

Zika virus (ZIKV) is an emergent arthropod-borne virus whose outbreak in Brazil has brought major public health problems. Infected individuals have different symptoms, including rash and pruritus, which can be relieved by the administration of antiallergics. In the case of pregnant women, ZIKV can cross the placenta and infect the fetus leading to congenital defects. We have identified that mast cells in the placentae of patients who had Zika during pregnancy can be infected. This led to our investigation on the possible role of mast cells during a ZIKV infection, using the HMC-1 cell line. We analyzed their permissiveness to infection, release of mediators and ultrastructural changes. Flow cytometry detection of ZIKV-NS1 expression 24 h post infection in 45.3% of cells showed that HMC-1 cells are permissive to ZIKV infection. Following infection, β-hexosaminidase was measured in the supernatant of the cells with a notable release at 30 min. In addition, an increase in TNF-α, IL-6, IL-10 and VEGF levels were measured at 6 h and 24 h post infection. Lastly, different intracellular changes were observed in an ultrastructural analysis of infected cells. Our findings suggest that mast cells may represent an important source of mediators that can activate other immune cell types during a ZIKV infection, which has the potential to be a major contributor in the spread of the virus in cases of vertical transmission.

scholarly journals T-cell calcium dynamics visualized in a ratiometric tdTomato-GCaMP6f transgenic reporter mouse

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Tobias X Dong ◽  
Shivashankar Othy ◽  
Amit Jairaman ◽  
Jonathan Skupsky ◽  
Angel Zavala ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Tracking ◽  
Cell Types ◽  
Cell Receptor ◽  
Receptor Activation ◽  
Specific Cell ◽  
Transgenic Expression ◽  
Reporter Mouse

Calcium is an essential cellular messenger that regulates numerous functions in living organisms. Here, we describe development and characterization of ‘Salsa6f’, a fusion of GCaMP6f and tdTomato optimized for cell tracking while monitoring cytosolic Ca2+, and a transgenic Ca2+ reporter mouse with Salsa6f targeted to the Rosa26 locus for Cre-dependent expression in specific cell types. The development and function of T cells was unaffected in Cd4-Salsa6f mice. We describe Ca2+ signals reported by Salsa6f during T cell receptor activation in naive T cells, helper Th17 T cells and regulatory T cells, and Ca2+ signals mediated in T cells by an activator of mechanosensitive Piezo1 channels. Transgenic expression of Salsa6f enables ratiometric imaging of Ca2+ signals in complex tissue environments found in vivo. Two-photon imaging of migrating T cells in the steady-state lymph node revealed both cell-wide and localized sub-cellular Ca2+ transients (‘sparkles’) as cells migrate.

scholarly journals T Cell Calcium Dynamics Visualized in a Ratiometric tdTomato-GCaMP6f Transgenic Reporter Mouse

2017 ◽  
Author(s):  
Tobias X. Dong ◽  
Shivashankar Othy ◽  
Amit Jairaman ◽  
Jonathan Skupsky ◽  
Angel Zavala ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Tracking ◽  
Cell Types ◽  
Cell Receptor ◽  
Functional Expression ◽  
Specific Cell ◽  
Transgenic Expression ◽  
Reporter Mouse

AbstractCalcium is an essential cellular messenger that regulates numerous functions in living organisms. Here we describe development and characterization of “Salsa6f”, a fusion of GCaMP6f and tdTomato optimized for cell tracking while monitoring cytosolic Ca2+, and a transgenic Ca2+ reporter mouse with Salsa6f floxed and targeted to the Rosa26 locus for expression in specific cell types. Using CD4-Cre-Salsa6f mice, we report normal development and function of T cells expressing Salsa6f and demonstrate Ca2+ signaling dynamics during T cell receptor engagement in naïve T cells, helper Th17 T cells and regulatory T cells. Salsa6f expression also revealed functional expression of mechanosensitive Piezo1 channels in T cells. Transgenic expression of Salsa6f enables ratiometric imaging of Ca2+ signals in complex tissue environments found in vivo. Deep tissue two-photon imaging of T cells in the steady-state lymph node revealed a highly localized Ca2+ signaling behavior (“sparkles”) as cells migrate.

scholarly journals Distinct T cell receptor signaling requirements for perforin- or FasL-mediated cytotoxicity.

1996 ◽  
Vol 183 (4) ◽  
pp. 1697-1706 ◽  
Author(s):  
M T Esser ◽  
B Krishnamurthy ◽  
V L Braciale
Keyword(s):  
Signal Transduction ◽  
T Cell ◽  
T Cell Receptor ◽  
Fas Ligand ◽  
Cytotoxic T Lymphocyte ◽  
Interleukin 2 ◽  
Cell Receptor ◽  
Receptor Expression ◽  
Cell Surface Receptor ◽  
Tcr Signaling

A diverse array of signals are generated in a cytotoxic T lymphocyte (CTL) after the T cell receptor (TCR) engages the class I major histocompatibility complex (MHC) peptide complex. These signals result in a multitude of CTL effector functions, including cellular cytotoxicity, cell surface receptor expression, and cytokine secretion. We have examined signaling through the TCR in a wild type CD8+, MHC-restricted, antigen-specific CTL clone, 14-7, and its interleukin 2-dependent variant clone 14-7FD. We report here that 14-7FD is unable to kill via the perforin mechanism of killing, yet is able to kill via the Fas ligand/Fas mechanism and secrete interferon-gamma in an antigen-specific manner. 14-7FD has cytolytic granules that contain perforin and serine esterases, which are secreted after phorbol ester and Ca2+ ionophore treatment. Lastly, to investigate which TCR signaling requirements were operational in 14-7FD, we examined TCR-triggered intracellular Ca2+ mobilization in the two clones. After TCR engagement, 14-7FD failed to mobilize intracellular Ca2+, which may be the cause for its inability to trigger the perforin/granule exocytosis mechanism of killing. These results indicate that the signal transduction events that trigger perforin killing and the signaling requirements to induce FasL expression are distinct. We hypothesize that these two distinct TCR signal transduction requirements allow for separate activation of these two mechanisms of killing relating to their role in eradication of infected cells or regulation of immune responses.

scholarly journals T cell receptor-independent CD2 signal transduction in FcR+ cells.

1991 ◽  
Vol 173 (4) ◽  
pp. 859-868 ◽  
Author(s):  
A R Arulanandam ◽  
S Koyasu ◽  
E L Reinherz
Keyword(s):  
Signal Transduction ◽  
T Cell ◽  
Mast Cells ◽  
Nk Cells ◽  
Signal Transduction Pathway ◽  
Cell Receptor ◽  
Surface Expression ◽  
Fc Receptor ◽  
Free Calcium ◽  
Murine Mast Cell

CD2 subserves both adhesion and signal transduction functions in T cells, thymocytes, and natural killer (NK) cells. In mature T lymphocytes, CD2-mediated signaling function apparently requires surface expression of T cell receptors (TCRs). In contrast, in CD2+ CD3- NK cells and thymocytes, signal transduction through CD2 is TCR independent. To resolve this paradox and characterize TCR-independent triggering mechanisms, we transfected a human CD2 cDNA into a murine mast cell line, C1.MC/57 (Fc epsilon RI+, Fc gamma RII+, Fc gamma RIII+), which is known to produce interleukin 6 (IL-6) as well as release histamine in response to crosslinking of Fc epsilon RI. In the CD2 transfectant, a combination of anti-T11(2) + anti-T11(3) monoclonal antibodies (mAbs) induced a rise in intracellular free calcium [( Ca2+]i), IL-6 production, and histamine release. As expected, no activation was mediated by the same mAbs in C1.MC/57. F(ab)'s fragments of the activatory combination of anti-T11(2) + anti-T11(3) mAbs induced IL-6 in the CD2-transfected mast cells, demonstrating an Fc gamma receptor ectodomain-independent triggering mechanism. In addition, either intact anti-T11(2) or anti-T11(3) IgG alone, which failed to induce [Ca2+]i mobilization in the transfectant, was able to induce IL-6 production. A mAb directed against both Fc gamma RII (previously denoted as Fc gamma RIIb) and Fc gamma RIII (previously denoted as Fc gamma RIIa) inhibits this induction. These results indicate that: (a) Ca2+ mobilization is not essential for IL-6 production; and (b) crosslinking of CD2 and Fc gamma receptors via intact anti-CD2 IgG stimulates IL-6 production. Thus, CD2-mediated IL-6 production occurs by both Fc receptor ectodomain-independent as well as Fc receptor ectodomain-dependent mechanisms in these nonlymphoid cells. Northern blot analysis demonstrates that although the mast cells do not express CD3 zeta or CD3 eta mRNA, they express Fc epsilon RI gamma mRNA. The latter is a known component of Fc gamma RIII as well as Fc epsilon RI, has significant homology to CD3 zeta/eta, and is thought to have a signal transduction function. In these mast cells, CD2 signaling machinery does not require CD3 zeta/eta and may be linked to the Fc epsilon RI gamma subunit. We predict that this subunit or a related structure may confer a TCR-independent signal transduction pathway upon CD2 in CD3- NK cells, thymocytes, and certain B lymphocytes.

scholarly journals The beta subunit of the Fc epsilon RI is associated with the Fc gamma RIII on mast cells.

1992 ◽  
Vol 175 (2) ◽  
pp. 447-451 ◽  
Author(s):  
T Kurosaki ◽  
I Gander ◽  
U Wirthmueller ◽  
J V Ravetch
Keyword(s):  
Signal Transduction ◽  
Mast Cells ◽  
Cell Receptor ◽  
Beta Subunit ◽  
Ligand Recognition ◽  
Gamma Chain ◽  
Effector Functions ◽  
Gamma Subunits ◽  
Alpha Beta ◽  
Common Signal

Fc epsilon RI is a tetrameric receptor, composed of a ligand recognition subunit, alpha, a beta chain, and dimeric gamma chains. Previous studies have indicated that the dimeric gamma chain is associated with Fc gamma RIIIA (CD16) on natural killer cells and macrophages as well as the clonotypic T cell receptor. Here we show that in mast cells, in addition to the dimeric gamma chains, the beta subunit is associated not only with Fc epsilon RI, but also with Fc gamma RIIIA. Functional reconstitution studies with a mastocytoma cell line indicate that Fc gamma RIIIA composed of alpha, beta, and gamma subunits has the capacity for signal transduction. These studies suggest that through the association of alternative ligand recognition subunits (alpha epsilon, alpha gamma), a common signal transduction complex (beta gamma 2) mediates similar biochemical and effector functions in response to immunoglobulin G (IgG) and IgE.

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