EC:3.4.21.79 - FACTA Search

Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.79 (granzyme B)
3,301 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The action mechanism of FTY720, a novel immunosuppressant, is completely different from conventional immunosuppressants. The drug, which triggers apoptosis in murine and human lymphocytes, has a potent immunosuppressive activity to prevent allograft rejection without any severe side-effect. The present study was designed to determine whether FTY720 induces apoptotic cell death in activated lymphocytes infiltrated into liver grafts with ongoing rejection. FTY720 was orally administered at 5 mg/kg to the recipients on day 3 and day 4 after grafting, when the graft rejection was histologically confirmed. The intragraft patterns of IL-2, interferon-gamma (IFN-gamma), perforin, and granzyme B gene expression were detected by reverse transcriptase-polymerase chain reaction. The treatment reversed ongoing rejection and significantly prolonged recipient survival time compared with the control group. Light microscopic observation of the graft sections stained with the DNA nick-end labelling method showed that the apoptosis in the control allografts was mainly induced in hepatocytes, while that in the FTY720-treated allografts was in infiltrated lymphocytes. The rejection therapy with FTY720 did not alter the expression of IL-2, IFN-gamma, and perforin mRNAs, but slightly decreased granzyme B expression. Our results suggest that FTY720 does not alter the intrinsic lymphocyte function to produce the rejection-related cytokines, but strongly induces apoptotic cell death in the activated lymphocytes. Thus, FTY720 affords new insight into the mechanisms underlying improvements in immunosuppressive treatments.
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PMID:Induction of lymphocyte apoptosis in rat liver allograft with ongoing rejection by FTY720. 1120 66

To become competent killer cells, CD8(+) T cells require stimulation through signal transduction pathways associated with the T-cell receptor, costimulatory molecules such as CD28, and cytokine receptors such as the interleukin (IL)-2 receptor. We used wortmannin and LY294002, two inhibitors of phosphatidylinositol 3-kinase (PI3-K), to study the role of PI3-K in mouse cytotoxic T-lymphocyte (CTL) induction in response to mitogenic anti-CD3 antibody. Anti-CD3-induced CD8(+) T-cell proliferation and CTL development were inhibited dose dependently by both PI3-K inhibitors. IL-2 synthesis by anti-CD3-activated CD8(+) T cells was also diminished by PI3-K inhibition. PI3-K inhibition resulted in a modest decrease in anti-CD3-induced CD4(+) T-cell proliferation but failed to affect IL-2 expression by anti-CD3-activated CD4(+) T cells. PI3-K inhibition during CTL induction resulted in decreased levels of mRNAs coding for granzyme B, perforin, and Fas ligand. In addition, CTL induced in the presence of PI3-K inhibitors failed to conjugate normally with P815 target cells. Exogenous IL-2 did not reverse the effects of PI3-K inhibition on CD8(+) T-cell proliferation and CTL induction. These results support the conclusion that PI3-K activation is involved in T-cell receptor, CD28, and IL-2 receptor signaling of CD8(+) T cells. PI3-K is, therefore, an important component of multiple signal transduction pathways involved in CTL generation.
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PMID:Phosphatidylinositol 3-kinase inhibitors prevent mouse cytotoxic T-cell development in vitro. 1135 90

CD8(+) T cells use a number of effector mechanisms to protect the host against infection. We have studied human CD8(+) T cells specific for CMV pp65(495-503) epitope, or for staphylococcal enterotoxin B, for the expression patterns of five cytokines and cytolytic effector molecules before and after antigenic stimulation. Ex vivo, the cytolytic molecule granzyme B was detected in a majority of circulating CMV-specific CD8(+) T cells, whereas perforin was rarely expressed. Both were highly expressed after Ag-specific activation accompanied by CD45RO up-regulation. TNF-alpha, IFN gamma, and IL-2 were sequentially acquired on recognition of Ag, but surprisingly, only around half of the CMV-specific CD8(+) T cells responded to antigenic stimuli with production of any cytokine measured. A dominant population coexpressed TNF-alpha and IFN-gamma, and cells expressing TNF-alpha only, IFN-gamma only, or all three cytokines together also occurred at lower but clearly detectable frequencies. Interestingly, perforin expression and production of IFN-gamma and TNF-alpha in CD8(+) T cells responding to staphylococcal enterotoxin B appeared to be largely segregated, and no IL-2 was detected in perforin-positive cells. Together, these data indicate that human CD8(+) T cells can be functionally segregated in vivo and have implications for the understanding of human CD8(+) T cell differentiation and specialization and regulation of effector mechanisms.
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PMID:Functional heterogeneity of cytokines and cytolytic effector molecules in human CD8+ T lymphocytes. 1141 47

Tumor-infiltrating lymphocytes (TIL) are well known to be functionally impaired typified by the inability to lyse cognate tumor cells in vitro. We have investigated the basis for defective TIL lytic function in transplantable murine tumor models. CD8(+) TIL are nonlytic immediately on isolation even though they express surface activation markers, contain effector phase cytokine mRNAs, and contain perforin and granzyme B proteins which are packaged into lytic granules. Ag-specific lytic capability is rapidly recovered if purified TIL are briefly cultured in vitro and tumor lysis is perforin-, but not Fas ligand mediated. In response to TCR ligation of nonlytic TIL in vitro, proximal and distal signaling events are normal; calcium flux is rapid; mitogen-activated protein/extracellular signal-related kinase kinase, extracellular regulatory kinase 2, phosphoinositide-3 kinase, and protein kinase C are activated; and IL-2 and IFN-gamma is secreted. However, on conjugate formation between nonlytic TIL and cognate tumor cells in vitro, the microtubule-organizing center (MTOC) does not localize to the immunological synapse, thereby precluding exocytosis of preformed lytic granules and accounting for defective TIL lytic function. Recovery of TCR-mediated, activation-dependent MTOC mobilization and lytic activity requires proteasome function, implying the existence of an inhibitor of MTOC mobilization. Our findings show that the regulated release of TIL cytolytic granules is defective despite functional TCR-mediated signal transduction.
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PMID:CD8(+) tumor-infiltrating T cells are deficient in perforin-mediated cytolytic activity due to defective microtubule-organizing center mobilization and lytic granule exocytosis. 1167 13

Here we show that the genes for perforin, the three major T cell granzymes (A-C) and IFN-gamma are differentially expressed during primary activation of naive CD8(+) T cells, kinetically and at the single-cell level. When CD44(low)CD62L(high)CD8(+) lymph node T cells were activated with IL-2 and immobilized antibodies to CD3, CD8 and CD11a, expression of perforin, granzyme B and IFN-gamma mRNAs was induced by day 2, and increased in parallel with perforin-dependent cytolytic activity. Granzyme C and A transcripts were not detected until 1 and 3 days later respectively. Single-cell PCR showed that expression frequencies rose in parallel with total levels of each mRNA, but that individual cells expressed diverse combinations of perforin, granzyme A-C and IFN-gamma mRNAs. These expression patterns indicated that the delayed expression of granzymes A and C was not due to late activation of distinct cell subpopulations. Statistical analysis of the data suggested that each gene was differentially regulated at the single-cell level. Individual naive CD8(+) T cells gave rise over 7 days to clones that expressed all five products at the clonal level, but also expressed diverse combinations at the single-cell level. We conclude that, during primary activation, CD8(+) T cells progressively acquired the ability to express most or all of these genes, and that the variable expression patterns observed among single cells within clones and populations reflected transient rather than heritable differences in expression profile.
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PMID:The genes for perforin, granzymes A-C and IFN-gamma are differentially expressed in single CD8(+) T cells during primary activation. 1203 12

NO is a potent cellular mediator which has been shown to modulate several immune mechanisms. Using human T lymphocytes as responder cells in a primary mixed lymphocyte reaction, we demonstrated that, at the initiation of the culture, exogenously provided NO via sodium nitroprusside, in non-toxic concentrations, inhibited both allogeneic proliferative and primary cytotoxic responses in a dose-dependent manner. In contrast, it had no effect on the cytotoxic activity of established human TCR (alpha)beta and TCR (gamma)delta cytotoxic T lymphocyte (CTL) clones. The NO inhibitory effect on primary cytotoxic T cell response correlates with inhibition of T cell blastogenesis. Furthermore, under our stimulation conditions, NO induced an inhibition of IL-2 production, an alteration of IL-2R(alpha) expression, and a down-regulation of NF-AT translocation in CD4(+) and CD8(+)allostimulated T cells. Furthermore, we demonstrate that the inhibition of allospecific CTL activity by the NO donor was at least in part related to an inhibition of granzyme B and Fas ligand transcription as revealed respectively by RNase protection and RT-PCR analysis. These results suggest that NO may function to fine tune human CD3(+) T cell activation and subsequent CTL generation.
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PMID:Analysis of the mechanisms of human cytotoxic T lymphocyte response inhibition by NO. 1235 82

Drugs can induce IgE mediated or T cell dependent immunological reactions. T cell dependent reactions are poorly understood, although T lymphocytes have been proposed as a protagonist in a number of non-immediate immunological reactions (NIR). The objective was to study in vivo different regulatory and proinflammatory cytokines and cytotoxic markers in patients with NIR to drugs. Twenty patients with NIR after drug intake were classified into two groups: Group A (severe), Stevens-Johnson syndrome and toxic epidermal necrolysis; and Group B (mild), maculopapular exanthema and desquamative exanthema. Another 25 subjects taking the same drugs but without reactions formed a control group. Samples were obtained within 24 hours of the reaction and 30 days later. IL-2, IL-4, IFN, TNF, perforin, granzyme B (GrB), and FasL mRNA expression levels were determined in peripheral blood mononuclear cells by competitive RT-PCR. There were 9 patients in Group A and 11 in Group B. The drugs involved were betalactams (8), anticonvulsants (6), allopurinol (1), sulfamethoxazole (1), amiodarone (1) dypirone (2), and erythromicine+paracetamol (1). At the acute stage there was a high increase of IL-2, IFN, and TNF mRNA expression in both groups vs. controls, perforin and GrB varied in each group with patients in Group A having the highest values, and FasL was only expressed in Group A. Relationships between the cytokines were only significant in Group B (p < 0.05). Only the relation between IFN-gamma and TNF-alpha was significant in Group A. There was a significant correlation between cytotoxic markers in both groups (A: p < 0.001, B: p < 0.01). These data demonstrate the complexity of the Th1 phenotype in NIR after drug intake. In patients with mild NIR, cytokines appear to play a closely related role, whereas cytotoxic markers appear more relevant in severe reactions.
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PMID:Gene expression levels of cytokine profile and cytotoxic markers in non-immediate reactions to drugs. 1249 Feb 85

Previously we showed that ethanol (EtOH) consumption suppressed IL-2-induced cytolytic activity of murine splenic natural killer (NK) cells. Although IL-2 receptor signaling is involved in activation of NK cells, neither the mechanism for this activation nor the role of EtOH consumption in modulating activation is completely understood. In this study we show by electrophoretic mobility-shift assay (EMSA) that enriched splenic NK cells from EtOH-consuming C57BL/6 mice exhibit reduced NF-kappaB and AP-1 binding activity in response to IL-2 stimulation as compared to the water-drinking mice. Semiquantitative RT-PCR and real-time PCR analyses indicated that EtOH consumption inhibits the induction of perforin, granzyme A, and granzyme B in response to IL-2. Pyrrolidine dithiocarbamate (PDTC) and N-tosyl-L-phenylalanine chloromethyl ketone (TPCK) blocked NFkappaB and AP-1 binding activity in nuclear extracts of IL-2-stimulated NK cells in an EMSA and also inhibited the IL-2-induced expression of perforin, granzyme A, and granzyme B gene expression in enriched NK cells. These inhibitors dramatically suppressed IL-2-stimulated NK cytolytic activity against YAC-1 lymphoma target cells. Taken together, these results suggest that NFkappaB and AP-1 are important regulators of NK cell cytolytic function through regulation of perforin, granzyme A, and granzyme B gene expression. The findings further suggest that the decreased cytolytic activity of IL-2-stimulated NK cytolytic activity in EtOH-consuming mice is due at least in part to impaired transactivation of these and possibly other genes involved in control of NK-cell target lysis.
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PMID:Alcohol consumption decreases IL-2-induced NF-kappaB activity in enriched NK cells from C57BL/6 mice. 1270 Apr 14

Cross-linking of the GPI-anchored protein Thy-1 results in T cell proliferation and IL-2 synthesis. However, the exact function of Thy-1 in the process of T cell activation remains unknown, as does the effect of costimulation on Thy-1-driven T cell responses. In this study, we have investigated the ability of Thy-1 to substitute for traditional signal 1 in the context of costimulation provided by dendritic cells. Dendritic cells dramatically enhanced T cell proliferation and IL-2 synthesis in response to Thy-1 triggering by anti-Thy-1 mAb. This effect was not dependent on dendritic cell Fcgamma receptors, but was a result of B7-mediated costimulation (signal 2). T cells were also activated when microbeads coated with a combination of anti-Thy-1 and anti-CD28 mAbs were used to supply signals 1 and 2, respectively. Thy-1-stimulated T cells adhere to target cells and express perforin, granzyme B, and Fas ligand, but fail to kill target cells due to an inability to reorganize their secretion machinery. Moreover, in contrast to TCR signaling, Thy-1 triggering failed to induce cytotoxicity in redirected lysis assays. We conclude that Thy-1 triggering can partially substitute for signal 1, which, in combination with a strong signal 2, leads to robust T cell proliferation, IL-2 synthesis, and cytotoxic effector molecule expression, but does not induce cytolytic function. The block at the level of cytotoxic effector function that results when T cells are activated in the absence of a classical, Ag-specific signal 1 may constitute a mechanism to ensure the specificity of CTL responses and prevent potentially harmful promiscuous cytotoxicity.
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PMID:Thy-1 signaling in the context of costimulation provided by dendritic cells provides signal 1 for T cell proliferation and cytotoxic effector molecule expression, but fails to trigger delivery of the lethal hit. 1281 84

Optimal differentiation of cytotoxic NK cells is important to provide protective innate immunity to patients after bone marrow transplantation. In vitro differentiation of CD56(+)CD3(-) NK cells takes weeks and is supported by several cytokines, including IL-2, IL-7, and IL-15, and thus can be useful for immunotherapy. However, IL-2 therapy is problematic in vivo, and NK cells differentiated in vitro with only IL-7 lack cytotoxicity. We assessed whether human NK cells initially differentiated in vitro from CD34(+)Lin(-) bone marrow cells with IL-7 could acquire cytotoxicity after exposure to additional cytokines and what changes promoted cytotoxicity. The cells cultured with IL-7 already had granzyme B as well as perforin, as previously reported, the proteins of cytotoxic granules. The cells also lacked LFA-1. After 1 wk of secondary culture with either IL-2 or IL-15, but not with IL-12 or IL-18, the IL-7-cultured cells acquired cytotoxicity. IL-2 or IL-15 also induced LFA-1. Ab to the LFA-1 subunits CD11a and CD18 blocked lysis by the NK cells, indicating that the new LFA-1 correlated with, and was essential for, the cytotoxic function of the in vitro generated cells. The LFA-1 also participated in target cell binding by the in vitro differentiated cells. In this study, we demonstrated a new function for IL-15, the induction of LFA-1 in NK progenitor cells, and that IL-15 does more than merely support NK progenitor cell proliferation. The efficacy after only 1 wk of IL-15 administration is a positive practical feature that may apply to human therapy.
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PMID:IL-15-mediated induction of LFA-1 is a late step required for cytotoxic differentiation of human NK cells from CD34+Lin- bone marrow cells. 1284 34

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