Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We previously demonstrated that interleukin-2 induced murine lymphokine-activated killer cell activity is augmented by retinoic acid. The enhanced cytotoxicity is significantly correlated with the increase in PKC. In the present study, we have shown that retinoic acid increases the expression of perforin, a potent cytolytic mediator, at both protein and mRNA levels. This enhancement can be induced by a direct stimulation of PKC signaling pathway, as manipulated by a short term incubation of lymphokine-activated killer cells with 12-O-tetradecanoyl-phorbol-13-acetate and ionomycin, and suppressed by PKC inhibitors. These results suggest that PKC plays a regulatory role in the enhancement of the expression of cytolytic mediators in retinoic acid-augmented lymphokine-activated killer cells.
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PMID:Enhancement of perforin by retinoic acid is mediated by protein kinase C. 846 34

Simultaneous protein kinase C stimulation with phorbol 12,13-dibutyrate (PDBU) and calcium mobilization with ionomycin (Io) trigger cellular events leading to expression of proliferation-associated genes in human lymphocytes. The effect of a 16-hr exposure to PDBU and Io on the growth and cytotoxic activity of murine splenocytes and tumor-infiltrating lymphocytes (TIL) cocultured with interleukin-2 (IL-2) was studied. PDBU + Io increased the number of cytotoxic effector cells that could be generated in lymphokine-activated killer cells (LAK) cultures (40-fold) and to a lesser extent in TIL (10-fold). DNA synthesis of TIL increased significantly when exposed to PDBU + Io. Also, TIL stimulated with PDBU + Io demonstrated in vitro tumor-specific lytic activity significantly greater than that of control TIL (500 lytic units vs 50). The cell-surface phenotype of TIL treated with PDBU + Io was identical to that of control TIL (> 95% CD-3+, CD-4-, CD-8+). Results of adoptive immunotherapy using splenocytes stimulated by PDBU + Io and cultured in IL-2 were identical to those achieved when standard LAK cultures were used. However, treatment using TIL stimulated by PDBU + Io led to a significant reduction in the number of pulmonary nodules compared to standard TIL. In addition, PDBU + Io-stimulated TIL maintained significant in vivo activity without the need for systemic IL-2 administration. Pharmacologic manipulation of cytotoxic precursor cells is a useful strategy for improving the generation of murine cytotoxic effector cells. By using PDBU + Io, cytotoxic lymphocytes could be generated with improved in vitro and in vivo activity.
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PMID:Phorbol dibutyrate and ionomycin improve murine effector cell cytotoxicity. 847 68

For serial cultivation of normal human melanocytes media supplemented with the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) are largely employed. By using a culture medium that permits cultivation of melanocytes without TPA, the effects of TPA on melanocyte proliferation, phenotype, and susceptibility to lymphokine-activated killer cells were studied. Addition of 50 ng/ml TPA to the medium induced rapid dendrite formation and increased the cell proliferation rate by 16-63% in mitogen-rich media (four of seven cultures, p < 0.01), and by 237% in mitogen-reduced media (p < 0.001). Furthermore, several phenotypic changes indicating early stages of melanocyte transformation were induced by 50 ng/ml TPA. These included increased expression of melanoma progression-associated antigens such as A.1.43 and A.10.33, upregulation of nerve-growth factor receptor as well as of the melanocyte-activation marker HMB-45 and of histocompatibility class I antigens. In contrast, the expression of the differentiation marker K.1.2 and of intercellular adhesion molecule-1 was decreased in TPA-treated cultures. Most of these changes persisted even after removal of TPA from the culture medium (> or = 2 weeks). Staurosporine, a protein kinase C inhibitor, modulated melanocyte-antigen expression similar to TPA, suggesting that protein kinase C downmodulation rather than activation by TPA is involved. In addition to the antigenic alterations, the susceptibility of TPA-treated melanocytes to lymphokine-activated killer cell cytotoxicity decreased by 40% (p < 0.01), possibly due to their altered surface antigen expression. The presented data reveal that the tumor promoter TPA hitherto used as a supplement of melanocyte culture media induces profound phenotypic and functional changes of the cultured cells, indicating incipient transformation of normal human melanocytes in vitro.
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PMID:12-O-tetradecanoylphorbol-13-acetate not only modulates proliferation rates, but also alters antigen expression and LAK-cell susceptibility of normal human melanocytes in vitro. 849 88

Activation of individual CD4+ T cells results in differential lymphokine expression: interleukin 2 (IL-2) is preferentially produced by T helper type 1 (TH1) cells, which are involved in cell-mediated immune responses, whereas IL-4 is synthesized by TH2 cells, which are essential for humoral immunity. The Ca(2+)-dependent factor NF-ATp plays a key role in the inducible transcription of both these lymphokine genes. However, while IL2 expression requires the contribution of Ca(2+)- and protein kinase C-dependent signals, we report that activation of human IL4 transcription through the Ca(2+)-dependent pathway is diminished by protein kinase C stimulation in Jurkat T cells. This phenomenon is due to mutually exclusive binding of NF-ATp and NF-kappa B to the P sequence, an element located 69 bp upstream of the IL4 transcription initiation site. Human IL4 promoter-mediated transcription is downregulated in Jurkat cells stimulated with the NF-kappa B-activating cytokine tumor necrosis factor alpha and suppressed in RelA-overexpressing cells. In contrast, protein kinase C stimulation or RelA overexpression does not affect the activity of a human IL4 promoter containing a mouse P sequence, which is a higher-affinity site for NF-ATp and a lower-affinity site for RelA. Thus, competition between two general transcriptional activators, RelA and NF-ATp, mediates the inhibitory effect of protein kinase C stimulation on IL4 expression and may contribute to differential gene expression in TH cells.
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PMID:Inhibition of NF-AT-dependent transcription by NF-kappa B: implications for differential gene expression in T helper cell subsets. 852 16

Contact between lymphokine-activated killer (LAK) cells and natural killer-resistant tumor targets SK-Mel-1 (human melanoma) or Raji (human lymphoma) stimulates phosphorylation of two M(r) 65,000 LAK proteins (pp65a and pp65b) with nearly identical isoelectric points. Phosphoamino acid analysis of pp65a and pp65b detected phosphorylation exclusively on serine residues. Phosphotyrosine could not be detected on either substrate after immunoblotting with an antiphosphotyrosine antibody, and herbimycin A treatment failed to inhibit p65 phosphorylation induced by target contact. However, phorbol myristate acetate treatment alone induced LAK pp65a and pp65b phosphorylation, suggesting phosphorylation may be mediated by protein kinase C or a protein kinase C-regulated kinase. The molecular weight and isoelectric points of pp65a and pp65b are similar to that reported for the human actin-bundling protein, L-plastin (L-fimbrin). On two-dimensional SDS-PAGE gel immunoblots, a peptide specific anti-L-plastin antiserum bound to pp65a and pp65b, suggesting that the phosphoproteins are similar or identical to L-plastin. In addition, two adjacent M(r) 65,000 LAK proteins were also detected by the antiserum and may correspond to unphosphorylated forms of L-plastin. On the basis of known properties of phosphorylated L-plastin, it is hypothesized that p65 phosphorylation in LAKs may regulate adhesion to tumor targets.
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PMID:Characterization of the M(r) 65,000 lymphokine-activated killer proteins phosphorylated after tumor target binding: evidence that pp65a and pp65b are phosphorylated forms of L-plastin. 854 53

Immune globulin for intravenous use (IVIG) has been used in many inflammatory conditions due to its immunomodulatory potential. The effector mechanisms are incompletely understood. This study dealt with the effects of IVIG on cytokine production in vitro. Cytokine synthesis was identified at the single-cell level using cytokine-specific MAb and indirect immunocytochemical techniques. Peripheral blood mononuclear cells (PBMC) were stimulated for 96 h by immobilized anti-CD3 MAb or by a combination of a protein kinase C activator (PMA) and a calcium ionophore (ionomycin). The addition of IVIG (6 mg/ml) caused a marked inhibition of proliferation and blast transformation despite unaffected cell survival. Anti-CD3-stimulated cultures containing IVIG exhibited a significant inhibition of production of T-cell derived lymphokines IL-2, IL-10, TNF-beta, IFN-gamma and TNF-alpha (made by both monocytes and T cells), while synthesis of the monokine IL-8 was significantly increased. The expression of IL-2 receptors was significantly suppressed. Similar but transient inhibition of most T-cell products (IL-2, IL-3, IL-4, IL-5, IL-10, TNF-beta and GM-CSF) was noted in the PMA/ionomycin-containing cultures. In contrast, no effects were found on IFN-gamma or TNF-alpha production. The superantigen streptococcal pyrogenic exotoxin-A (SPE-A) induced vigorous cell activation and extensive cytokine synthesis. IVIG was added either at the beginning or 24 h after the initiation of cultures in order to elucidate the importance of direct toxin-neutralization. Addition of IVIG from the beginning of cultures induced a strong reduction of blast transformation and an almost complete inhibition of lymphokine production, in particular of IFN-gamma and TNF-beta. Supplementation with IVIG 24 h after initiation of cultures also led to a significant decrease in lymphokine synthesis. Monokine production (IL-1 alpha, IL-1 beta, IL-1ra, IL-6 and IL-8) was either unaffected or even increased. These two facts argue against direct antigen-neutralization as being the only mechanism at work. However, in IVIG-exposed PBMC stimulated with LPS, IL-6 production was significantly reduced. A significant upregulation of IL-1ra was noticed in unstimulated PBMC cultured with IVIG. The results in all the experiments did not indicate a cytotoxic effect by IVIG on cell survival and the production of certain cytokines were unaffected. Instead, the authors believe that the results suggest a previously little examined functional link where the humoral immune response may have direct immunoregulatory effects on the cellular immune system.
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PMID:Intravenous immune globulin affects cytokine production in T lymphocytes and monocytes/macrophages. 862 37

Lymphokines produced by non-transformed Th clones, Th1 and Th2, were classified into three groups based on their patterns of expression by different stimuli: Group I, GM-CSF and IL-2, characterized by a strict requirement of activation of both the PKC- and calcium-dependent pathways; Group II, IFN-gamma, IL-3, and IL-4, partially induced by calcium ionophore alone; and Group III, IL-5, IL-6, and IL-10, partially induced by either PMA or calcium ionophore alone. Transfection of constitutively active PKC or p21ras replaced the requirement for PMA in expression of these lymphokines, with the exception of GM-CSF. Production of Group II lymphokines was partially induced by constitutively active calcineurin. Production of Group I and II lymphokines was highly sensitive to cyclosporin A, while Group III lymphokines were relatively resistant. Addition of prostaglandin E2 (PGE2) and overexpression of catalytic subunit of protein kinase A inhibited lymphokine production in Th1 cells, but not in Th2 cells, with the exception of GM-CSF. Production of Group III lymphokines induced by PMA alone was upregulated by PGE2, but that of Group II and III lymphokines induced by calcium ionophore alone was not affected. These results suggest that one of the targets of PGE2 is downstream of the PKC-dependent pathway.
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PMID:Signal transduction in Th clones: target of differential modulation by PGE2 may reside downstream of the PKC-dependent pathway. 872 62

Thymocytes of T cell receptor transgenic mice with nonselecting and RAG-2 -/- backgrounds were developmentally arrested at the CD4+CD8+ stage before positive selection. These thymocytes underwent lineage commitment upon transient stimulation with a combination of ionomycin, a calcium ionophore, and phorbol 12-myristate 13-acetate (PMA), a protein kinase C activator, in suspension culture. The effective drug doses were limited within narrow ranges and much lower than those which induce proliferation of mature T cells. The doses corresponded to those which inhibit glucocorticoid-induced apoptosis in these thymocytes. CD4 lineage commitment required longer duration, higher intensity of the stimulation, or both, than CD8 lineage commitment. Functional helper T cells (Th1 and Th2) were induced from the CD4 lineage-committed cells upon secondary stimulation with a combination of ionomycin and PMA followed by lymphokine treatment. Cytotoxic T cells were induced from the CD8 lineage-committed cells upon incubation with concanavalin A and irradiated splenic dendritic cells, but not with the combination of ionomycin and PMA. These results indicate that positive selection is mimicked by the pharmacological stimulation in the absence of other cell types, but that final maturation of CD8 T cells may require a different signal.
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PMID:Differential induction of helper and killer T cells from isolated CD4+CD8+ thymocytes in suspension culture. 881 50

Treatment of lymphokine-activated killer (LAK) cells with phorbol ester (PMA) caused the downmodulation of LAK activity concomitantly with the inhibition of serine esterase (SE) release, which has been shown as a marker for perforin-dependent cell-mediated cytotoxicity. The reduction of perforin-dependent LAK activity by PMA-treatment appeared to be due to the disappearance of PMA-sensitive protein kinase C (PKC) isoforms such as PKC alpha, gamma, epsilon, theta. In contrast, Fas-mediated LAK activity was refractory against PMA-induced downregulation. Treatment of LAK cells with PMA caused a disappearance of cytotoxicity against Fas L5178Y tumor cells, while cytotoxicity against Fas+ transfectants was not affected by PMA treatment. Moreover, Fas-mediated LAK activity of perforin-knockout mice was not inhibited by PMA treatment. These results clearly demonstrated that Fas-mediated cytotoxicity could be dissociated from perforin-mediated cytotoxicity by their different requirement of PMA-sensitive PKC isoforms.
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PMID:The role of phorbol ester-sensitive protein kinase C isoforms in lymphokine-activated killer cell-mediated cytotoxicity: dissociation between perforin-dependent and Fas-dependent cytotoxicity. 920 76

Membrane glycoproteins (gps) play an important role in cell-cell interactions during epidermal maturation, and we have previously shown an up-regulation of PNA-binding gps in cultured human keratinocytes treated with interferon gamma (IFN-gamma). The protein kinase C (PKC) pathway is known to play a key role in the regulation of proliferation and differentiation of keratinocytes and is also reported to be involved in some IFN-gamma-mediated effects. In order to evaluate the cellular mechanisms and whether PNA-binding gp expression is related to the differentiative activity of the lymphokine, we studied the effects of PKC agonists and antagonists and the role of retinoic acid (RA), in the induction of these gps in cultured human keratinocytes stimulated with IFN-gamma and processed for protein analysis. The expression of PNA-binding gps was revealed by incubation of SDS-polyacrylamide gels with 125I-PNA. The PKC antagonists (H7, sphingosine) as well as RA downregulated the IFN-gamma-induced PNA-reactive gps, whereas staurosporine and TPA upregulated their expression. These results provide evidence that PNA-reactive gps are late highly IFN-gamma-sensitive markers of keratinocyte differentiation, drastically modulated through selective isoforms of PKC.
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PMID:Interferon gamma-induced PNA-binding glycoproteins as markers of human keratinocyte differentiation: biological evidence using protein kinase C agonists, antagonists and retinoic acid. 944 84


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