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

1. The ability of acetylshikonin to inhibit the respiratory burst in rat neutrophils was characterized and the underlying mechanism of action was also assessed in the present study. 2. Acetylshikonin caused an irreversible and a concentration-dependent inhibition of formylmethionylleucyl-phenylalanine (fMLP) plus dihydrocytochalasin B (CB)- and phorbol 12-myristate 13-acetate (PMA)-induced superoxide anion (O2.-) generation with IC50 values of 0.48 +/- 0.03 and 0.39 +/- 0.03 microM, respectively. Acetylshikonin also inhibited the O2 consumption in neutrophils in response to fMLP/CB as well as to PMA. 3. Acetylshikonin did not scavenge the generated O2.- in the xanthine-xanthine oxidase system or during dihydroxyfumaric acid (DHF) autoxidation but, on the contrary, acetylshikonin enhanced the O2.- generation in these cell-free oxygen radical generating systems. 4. Acetylshikonin inhibited the formation of inositol trisphosphate (IP3) (39.0 +/- 7.8% inhibition at 10 microM, P < 0.05) in neutrophils in response to fMLP. 5. Both the neutrophil cytosolic protein kinase C (PKC) activity and the PMA-induced PKC associated with the membrane were unaffected by acetylshikonin. 6. Acetylshikonin did not affect the porcine heart protein kinase A (PKA) activity. Upon exposure to acetylshikonin, the cellular cyclic AMP level was decreased in neutrophils in response to fMLP. 7. The cellular formation of phosphatidic acid (PA) and, in the presence of ethanol, phosphatidylethanol (PEt) induced by fMLP/CB were inhibited by acetylshikonin (60.1 +/- 7.3 and 63.2 +/- 10.5% inhibition, respectively, at 10 microM, both P < 0.05). Moreover, acetylshikonin attenuated the fMLP/CB-induced protein tyrosine phosphorylation (about 90% inhibition at 1 microM). 8. In PMA-activated neutrophil particulate NADPH oxidase preparations, acetylshikonin did not inhibit, but enhanced, the O2.- generation in the presence of NADPH. However, acetylshikonin decreased the membrane associated p47phox in PMA-activated neutrophils (about 60% inhibition at 1 microM). 9. Collectively, these results suggest that the attenuation of protein tyrosine phosphorylation and a failure in the assembly of a functional NADPH oxidase complex probably contribute predominantly to the inhibition of respiratory burst in neutrophils by acetylshikonin. In contrast, the blockade of phospholipase C (PLC) and phospholipase D (PLD) pathways play only a minor role in this respect.
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PMID:Investigation of the inhibition by acetylshikonin of the respiratory burst in rat neutrophils. 917 81

The respiratory burst oxidase of phagocytes and B lymphocytes catalyzes the reduction of oxygen to superoxide anion (O-2) at the expense of NADPH. This multicomponent enzyme is dormant in resting cells but is activated on exposure to an appropriate stimulus. The phosphorylation-dependent mechanisms regulating the activation of the respiratory burst oxidase are unclear, particularly the phosphorylation status of the cytosolic component p67(phox). In this study, we found that activation of human neutrophils with formyl-methionyl-leucyl-phenylalanine (fMLP), a chemotactic peptide, or phorbol myristate acetate (PMA), a stimulator of protein kinase C (PKC), resulted in the phosphorylation of p67(phox). Using an anti-p67(phox) antibody or an anti-p47(phox) antibody, we showed that phosphorylated p67(phox) and p47(phox) form a complex. Phosphoamino acid analysis of the phosphorylated p67(phox) revealed only 32P-labeled serine residues. Two-dimensional tryptic peptide mapping analysis showed that p67(phox) is phosphorylated at the same peptide whether fMLP or PMA is used as a stimulus. In addition, PKC induced the phosphorylation of recombinant GST-p67(phox) in vitro, at the same peptide as that phosphorylated in intact cells. PMA-induced phosphorylation of p67(phox) was strongly inhibited by the PKC inhibitor GF109203X. In contrast, fMLP-induced phosphorylation was minimally affected by this PKC inhibitor. Taken together, these results show that p67(phox) is phosphorylated in human neutrophils by different pathways, one of which involves protein kinase C.
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PMID:Phosphorylation of the respiratory burst oxidase subunit p67(phox) during human neutrophil activation. Regulation by protein kinase C-dependent and independent pathways. 920 43

1. The effects of bisindolylmaleimide GF 109 203X, reported to be a potent and highly selective inhibitor of protein kinase C (PKC), have been investigated on some human neutrophil functions. 2. GF 109 203X prevented O.2- production by NADPH-oxidase whatever the stimulus used for polymorphonuclear neutrophil (PMN) activation: directs PKC activators like phorbol myristate acetate (PMA) and dioctanoylglycerol, calcium ionophore (A23187), or receptor agonists like fMet-Leu-Phe (fMLP) and opsonized zymosan. 3. The effect of GF 109 203X was also examined on elastase exocytosis by neutrophils. PMA-mediated release was prevented by GF 109 203X. However, GF 109 203X had no effect on exocytosis induced by A23187 and the effect of this compound on the fMLP response changed according to its concentration. 4. These data suggest that PKC might be essential for stimulus-mediated O.2- production and also that PKC plays only a minor role in elastase secretion as compared to the role of the cytosolic calcium level.
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PMID:Effect of the protein kinase C inhibitor GF 109 203X on elastase release and respiratory burst of human neutrophils. 930 17

Malarial pigment (haemozoin; HZ) is generally considered to be a non-toxic, high-molecular-weight storage form of undigested, toxic, host-haemoglobin haem. The available information on HZ indicates that it is a very heterozygous material. Its exact structure, in terms of constituent proteins (remnants of host globin v. parasite proteins), the type of linkage between the haem moieties (mu-oxo haem dimers further aggregated by non-covalent hydrophobic bonds v. mutually independent haematin monomers), iron status in the haem (penta-co-ordinated, high-spin ferriprotoporphyrin IX v. esa-co-ordinated, low-spin ferriprotoporphyrin IX), and compositions (beta-haematin-like structure without functionally relevant proteins or other constituents v. a ferriprotoporphyrin-IX core with aggregated proteins and phospholipids of host and parasite origin) remains a subject of controversy. When investigated by macrophages, HZ is not inert but affects a number of functional parameters. Crude pigment, as present in infected erythrocytes and shed after schizont rupture, may be considered the 'natural diet' ingested by macrophages in infected blood. It is a powerful source of radicals that may generate lipoperoxides and derived, toxic hydroxyaldehydes such as 4-hydroxynonenal (HNE). High concentrations of HNE, which have been detected in HZ-fed macrophages, inhibit protein kinase C (PKC). Complexes between HNE and PKC have also been detected in immunoprecipitated PKC from HZ-fed macrophages. HNE-mediated inhibition of PKC (and of other, as yet unidentified enzymes and processes) may explain HZ-mediated effects. HZ-mediated inhibition of NADPH-oxidase, the enzyme responsible for oxidative bursts, may only be partially explained by PKC inhibition. As Hz-laden human and murine macrophages produce increased amounts of tumour necrosis factor-alpha, interleukins 1 and 6, and macrophage inflammatory proteins 1 alpha and 1 beta, HZ-macrophage interactions may contribute to the cytokine-mediated manifestations of malaria.
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PMID:Malarial pigment (haemozoin): a very active 'inert' substance. 932 87

Since selenite and other redox-active selenocompounds can modify protein kinase C (PKC) in the test tube, we have determined whether or not this redox regulation occurs inside the cell despite having high concentrations of GSH and the role of this regulation in the inhibition of tumor promotion. By using phorbol ester-promoted JB6 epidermal cell transformation assay, the concentrations of selenite, selenocystine, and selenodiglutathione which are optimal for chemopreventive activity were determined. At such concentrations (0.5 to 2 microM) in the cells treated with these agents, only a slight but transient decrease in PKC activity was observed when measured with a low (5 microM), but not with a high (100 microM) concentration of ATP. However, when the cells were serum starved or pretreated with 2-deoxyglucose, there was a pronounced but transient inactivation of PKC when assayed with both low and high concentrations of ATP. The inactivation was reversed in the cell by an endogenous mechanism or by treatment with thiol agents in the test tube. In spite of a substantial (90%) depletion of GSH in the cells by pretreatment with buthionine sulfoximine, there was no further increase in the redox modification of PKC by selenite as well as no change in the inhibitory effect of selenite on the phorbol ester-stimulated induction of ornithine decarboxylase, which is an intermediate marker related to cell transformation. While GSH is known to influence certain actions of selenium, it may not be required to mediate the effects of selenite tested in this study. The water-soluble cytosolic GSH did not interfere with the redox modification of PKC probably due to the shielding of the cysteine-rich region of the enzyme by a weak hydrophobic association with the membrane. Due to the presence of cofactors in the crude cell extracts, PKC was more sensitive to selenite than in the purified form and was inactivated by low concentrations of selenite (IC50 = 0.05 microM). This modification was reversed by thiol agents as well as by NADPH. A protein disulfide reductase, which can regenerate PKC, was present in the homogenate. Conceivably, selenite and other selenocompounds induce a redox modification of cellular PKC, compartmentally independent from the cytosolic GSH, but intimately connected to a NADPH-dependent reductase system, to mediate, at least in part, some of the cancer-preventive actions.
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PMID:Selenocompounds induce a redox modulation of protein kinase C in the cell, compartmentally independent from cytosolic glutathione: its role in inhibition of tumor promotion. 939 Jan 72

PMNs are a major component of body defense against microbial invasion, involving reactive oxygen species in great quantity, which could benefit from antibiotic therapy. Recently, possible antibiotic effects on phagocyte functions (impairment or stimulation of reactive oxygen species production) were studied. In our study, an in vitro evaluation was made on macrolide activity on phagocyte respiratory burst functions, using assay of superoxide anion (O2.-) in response to four stimuli systems: N-formyl Met-Leu-Phe (fMLP), an analogue of bacterial chemotactic factors; 4 beta-phorbol 12-myristate 13-acetate (PMA), a direct activator of protein kinase C (PKC); calcium ionophore (A23187), which acts directly on calcium influx; and a bacterial strain, Staphylococcus aureus. We have shown that spiramycin, at therapeutic plasma concentrations, increased O2.- generation by bacteria and fMLP-stimulated PMNs, with rate of 26% for 1 microgram ml-1 and 34% for 5 micrograms ml-1, respectively. This pro-oxidant effect, however, weaker, was observed when PMNs were stimulated by PMA. A weak anti-oxidant effect was observed with A23187. For higher concentrations, spiramycin decreased strongly O2.- production, with IC50 values of 74 micrograms ml-1, 154 micrograms ml-1, 296 micrograms ml-1 and 400 micrograms ml-1 when PMNs were stimulated with bacteria, A23187, fMLP and PMA, respectively. The effect of spiramycin seemed to result from an intracellular mechanism by intervention of PMN oxidative metabolism (NADPH-oxidase activation), rather than a simple chemical interaction, because no effect has been observed in acellular models. For higher spiramycin concentrations, the decrease of O2.- production observed could not be taken into consideration because this concentration was not used in therapy. The enhanced of O2.- production observed could be used in therapy, so as to increase PMNs bactericidal activity.
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PMID:In vitro interaction between spiramycin and polymorphonuclear neutrophils oxidative metabolism. 960 67

Periodontal disease, a frequent complication of diabetes mellitus, is the major cause of tooth loss. However, studies on neutrophil function in patients with this condition have yielded contradictory findings. The NADPH oxidase activity of 40 diabetic patients with periodontosis who were on metabolic control was evaluated and compared with that in 40 healthy subjects. Superoxide anion production was measured by a photometric method, with NBT reduction at 490 nm in a microplate reader and by a microscopic method, with a percentage of positive PMNs with granules of formazan in the cytoplasm. When the PMN respiratory burst was activated by phorbol myristate acetate (PMA), a protein kinase C (PKC) soluble activator, superoxide production of diabetics (4.31 +/- 1.67 A x 10(-3)/min) and normal subjects (4.25 +/- 1.25 A x 10(-3)/min) was comparable by photometric method, whereas a significantly defective response to opsonized zymosan was observed when the microscopic method was used (58 +/- 17% in diabetics and 66 +/- 18% in controls; p = 0.05). Therefore in patients with diabetes the impact on PMN function is of multifactorial origin, and is probably correlated to the glucose level and to glycation of PMN protein, such as NADPH oxidase or myeloperoxidase. Alternatively, glucose in PMN may be reduced by aldose reductase to polyols, and this pathway requires NADPH, the coenzyme for the respiratory burst. Moreover, we found that superoxide production in response to opsonized zymosan was reduced in diabetic patients. The activation of protein tyrosine kinase (PTK) is an important mechanism underlying transmembrane signaling and, moreover, protein tyrosine phosphorylations, stimulated by zymosan receptor-mediated activation, might be caused by the activation of specific PTK, whereas activation by PMA is probably mediated through another PKC type.
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PMID:Respiratory burst of neutrophils in diabetic patients with periodontal disease. 970 64

The leukocyte NADPH oxidase of neutrophils is a membrane-bound enzyme that catalyzes the production of O-2 from oxygen using NADPH as the electron donor. Dormant in resting neutrophils, the enzyme acquires catalytic activity when the cells are exposed to appropriate stimuli. During activation, the cytosolic oxidase components p47phox and p67phox migrate to the plasma membrane, where they associate with cytochrome b558, a membrane-integrated flavohemoprotein, to assemble the active oxidase. Oxidase activation can be mimicked in a cell-free system using an anionic amphiphile, such as sodium dodecyl sulfate or arachidonic acid, as an activating agent. In whole cells and under certain circumstances in the cell-free system the phosphorylation of p47phox mediates the activation process. It has been proposed that conformational changes in the protein structure of cytosolic factor p47phox may be an important part of the activation mechanism. We show here that the total protein steady-state intrinsic fluorescence (an emission maximum of 338 nm) exhibited by the tryptophan residues of p47phox substantially decreased when p47phox was treated with anionic amphiphiles. A similar decrease in fluorescence was also observed when p47phox was phosphorylated with protein kinase C. Furthermore, a red shift of emission maximum and an increase of quenching by ionic quenchers and acrylamide were observed in the presence of activators. These results indicate the occurrence of a conformational change in the protein structure of p47phox. We propose that this alteration in conformation results in the appearance of a binding site through which p47phox interacts with cytochrome b558 during the activation process.
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PMID:Conformational changes of the leukocyte NADPH oxidase subunit p47(phox) during activation studied through its intrinsic fluorescence. 974 57

Luteal regression is associated with the generation of reactive oxygen species (ROS). To determine the nature of the ROS generator, cells isolated from luteinized rat ovaries were examined for ROS production using luminol-amplified chemiluminescence (LCL). Cells cultured for 2-48 h exhibited minimal LCL, but there was a significant (30- to 50-fold), rapid (maximum at 3-5 min), and dose-dependent increase in LCL in response to phorbol ester (phorbol 12-myristate 13-acetate; TPA; ED50 = 0.03 microM) and diacylglycerol (1,2-dioctanoyl-glycerol; ED50 = 30 microM). The TPA-induced response was cell number dependent and was virtually abolished by superoxide dismutase, freezing, or heating (95 degrees C for 5 min). Zymosan, known to induce a phagocytic response in leukocytes, stimulated a superoxide (O2-.) response with a slow onset (maximum at 40 to 60 min) and a maximum about one third of that observed for TPA. The response to TPA and zymosan was inhibited by the NADPH/NADH-oxidase inhibitor, diphenylene iodonium (ID50 = 5 microM for TPA), but not by the mitochondrial inhibitors, potassium cyanide, rotenone, or sodium azide. Fractionation of cells by centrifugal elutriation showed that TPA-stimulated O2-. production coeluted with the nonsteroidogenic cells and that little, if any, O2-. generation coeluted with the steroidogenic cells. Cells isolated 1, 2, and 4 h after in vivo treatment with a luteolytic dose of prostaglandin F2alpha (PGF2alpha) showed a significant increase in TPA-stimulated O2-. production at 2 h, whereas luteal cells or corpora lutea incubated directly with 1 microM PGF2alpha did not show any increase in response. Corpora lutea isolated from naturally regressed ovaries (18 days after ovulation) showed a significantly elevated level of TPA-stimulated O2-. production. In conclusion, there is a superoxide generator in luteinized ovaries that is activated through a protein kinase C pathway, localized in nonsteroidogenic cells, transiently increased during PGF2alpha-induced luteolysis in vivo, and elevated during natural luteal regression.
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PMID:Prostaglandin f2alpha treatment in vivo, but not in vitro, stimulates protein kinase C-activated superoxide production by nonsteroidogenic cells of the rat corpus luteum. 978 Mar 11

Classical chemoattractants and chemokines trigger integrin-dependent adhesion of blood leukocytes to vascular endothelium and also direct subsequent extravasation and migration into tissues. In studies of human polymorphonuclear neutrophil responses to formyl peptides and to interleukin 8, we show evidence of involvement of the atypical zeta protein kinase C in the signaling pathway leading to chemoattractant-triggered actin assembly, integrin-dependent adhesion, and chemotaxis. Selective inhibitors of classical and novel protein kinase C isozymes do not prevent chemoattractant-induced neutrophil adhesion and chemotaxis. In contrast, chelerythrine chloride and synthetic myristoylated peptides with sequences based on the endogenous zeta protein kinase C pseudosubstrate region block agonist-induced adhesion to fibrinogen, chemotaxis and F-actin accumulation. Biochemical analysis shows that chemoattractants trigger rapid translocation of zeta protein kinase C to the plasma membrane accompanied by rapid but transient increase of the kinase activity. Moreover, pretreatment with C3 transferase, a specific inhibitor of Rho small GTPases, blocks zeta but not alpha protein kinase C plasma membrane translocation. Synthetic peptides from zeta protein kinase C also inhibit phorbol ester-induced integrin-dependent adhesion but not NADPH-oxidase activation, and C3 transferase pretreatment blocks phorbol ester-triggered translocation of zeta but not alpha protein kinase C. These data suggest the involvement of zeta protein kinase C in chemoattractant-induced leukocyte integrin-dependent adhesion and chemotaxis. Moreover, they highlight a potential link between atypical protein kinase C isozymes and Rho signaling pathways leading to integrin-activation.
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PMID:Evidence of zeta protein kinase C involvement in polymorphonuclear neutrophil integrin-dependent adhesion and chemotaxis. 980 92


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