Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.4.3 (phospholipase C)
18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The inducibility of glycosyl-phosphatidylinositol (GPI)-anchored proteins on affected paroxysmal nocturnal haemoglobinuria (PNH) neutrophils (PMN) after both in vitro and in vivo stimulation was investigated. Fc gamma R-III (CD16), decay-accelerating factor (DAF/CD55) and 20 kD homologous restriction factor (HRF20/CD59) were demonstrated to be concurrently deficient on unstimulated defective PNH PMN. Upon in vitro stimulation with either N-formyl-methionyl-leucyl-phenylalanine (fMLP), zymosan-activated serum (ZAS), or recombinant human granulocyte colony-stimulation factor (G-CSF), neither CD16 nor CD55 expression was induced on defective PNH PMN. G-CSF was administered to two patients with PNH when their conditions were complicated by bacterial infections, or to prevent infections associated with the extraction of teeth or cataract surgery. CD16 expression was induced on the defective PNH PMN in both cases during the administration of G-CSF, but the expression of CD55 and CD59 was not. CD16, induced on the defective PNH PMN during the administration of G-CSF, was phosphatidylinositol-specific phospholipase C (PIPLC)-sensitive, implying that it had GPI-linkage to the membranes. The patients treated with G-CSF recovered from infection or evaded infection. These observations suggest that a deficiency of GPI-anchored proteins is not always seen in defective PNH blood cells, at least under certain stimulation conditions.
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PMID:Induction of Fc gamma R-III (CD16) expression on neutrophils affected by paroxysmal nocturnal haemoglobinuria by administration of granulocyte colony-stimulating factor. 769 30

We report that engagement of a particular epitope near the C-terminal region of complement receptor type 3 (CR3, CD11b/CD18) alpha-chain with CD11b mAb VIM12 induces granulocyte activation with a rise in cytosolic-free Ca2+, actin polymerization, an up-regulation of CR3 cell surface expression and enhanced adhesiveness. Induction of enhanced adhesiveness and homotypic aggregation of human granulocytes represents an active process. It is temperature and energy dependent, requires divalent cations, and an intact cytoskeleton. The mAb VIM12-induced enhanced adhesiveness seems to be mediated, at least in part, by activated CR3 molecules. It can be significantly inhibited, although not completely abolished, with blocking mAbs against adhesiotopes of CR3. VIM12-induced adhesion could be blocked with the serine/threonine inhibitors okadaic acid and calyculin A and with dibuturyl-cAMP but not with the protein kinase inhibitors herbimycin A and staurosporine. We further present evidence that the particular molecular region of CR3 recognized by mAb VIM12 might be involved in the reported intramembrane sugar-lectin type interaction and complex formation between transmembrane CR3 and glycosylphosphatidylinositol (GPI)-anchored Fc gamma RIIIB (CD16) molecules on human granulocytes. Binding of mAb VIM12 to CR3 on granulocytes enhances the release of GPI-anchored Fc gamma RIIIB molecules from granulocytes upon phosphoinositol-phospholipase C treatment. The sugar preparation N-acetyl-D-glucosamine, previously shown to dissociate CR3-Fc gamma RIIIB complex formation, inhibits mAb VIM12 binding. Engagement of CR3 with mAb VIM12 may thus mimic a biologically relevant intramembrane cooperation between two distinct receptor molecules on human granulocytes.
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PMID:Granulocyte activation via a binding site near the C-terminal region of complement receptor type 3 alpha-chain (CD11b) potentially involved in intramembrane complex formation with glycosylphosphatidylinositol-anchored Fc gamma RIIIB (CD16) molecules. 773 Jun 47

We have shown previously that basic fibroblast growth factor (bFGF) is a mitogen for human bone marrow (BM) stromal cells and that bFGF stimulates myelopoiesis in primary BM cultures. In this article, we demonstrate the presence of bFGF in two cell lineages in human BM and peripheral blood as well as the deposition of bFGF into the extracellular matrix of BM stromal cell cultures. In immunofluorescence experiments on BM and peripheral blood smears, megakaryocytes and platelets stained strongly for bFGF, whereas weaker staining was observed in immature and mature cells of the granulocyte series. The presence of bFGF in platelets was confirmed by enzyme-linked immunosorbent assay as well as by immunoprecipitation followed by immunoblotting. bFGF was synthesized by BM stromal cell cultures and was found either cell associated or localized in the nucleus and the nucleoli, and its location was dependent on the fixation procedure used. Addition of exogenous bFGF to stromal cells showed the presence of extracellular binding molecules for this cytokine. bFGF could be released from these sites by soluble heparin or phosphatidylinositol-specific phospholipase C. This study supports the role of bFGF as a stromal cell mitogen and stimulator of myelopoiesis. The data indicate that the stromal cells produce bFGF and that their extracellular matrix can serve as a reservoir for this growth factor. In addition, the results suggest a possible involvement of bFGF in platelet function as well as in megakaryocytopoiesis.
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PMID:Basic fibroblast growth factor expression in human bone marrow and peripheral blood cells. 842 58

The sphingolipids, sphingosylphosphorylcholine (SPPC) and sphingosine-1-phosphate (SPP), induce a rapid and transient rise in intracellular free calcium concentration ([Ca2+]i) in a variety of cell lines via activation of pertussis toxin-sensitive G protein-coupled receptors. We investigated whether these sphingolipids act on different receptors by testing the effect of varying concentrations of SPPC on [Ca2+]i in human leukemia HL-60 cells, which have been found to be nonresponsive to SPP. SPPC potently (EC50 = 1.5 microM) and rapidly increased [Ca2+]i in HL-60 cells in a pertussis toxin-sensitive manner. Differentiation of HL-60 cells through treatment with dibutyryl cAMP into granulocyte-like cells did not change the magnitude or the pertussis toxin sensitivity of the SPPC-induced [Ca2+]i rise, indicating that the receptor for SPPC is constitutively expressed in HL-60 cells. SPPC did not activate phospholipase C or D in HL-60 cells. However, SPPC, but not SPP, stimulated the generation of superoxide anions in dibutyryl cAMP-differentiated HL-60 cells as well as in human neutrophils, suggesting that the SPPC receptor may play a role in the inflammatory defense against invading microorganisms. On the basis of these results, we conclude that there apparently is a heterogeneity of G protein-coupled receptors for sphingolipids in mammalian cells.
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PMID:A distinct G(i) protein-coupled receptor for sphingosylphosphorylcholine in human leukemia HL-60 cells and human neutrophils. 864 55

Bacterial lipopolysaccharide (LPS)-induced exocytosis is one of the primary immune responses of the Limulus granulocyte (GR). Exocytosis can be mediated by guanine nucleotide-binding protein (G-protein)-linked surface receptors that activate phospholipase C (PLC) to produce inositol 1,4,5-triphosphate (IP3) and diacylglycerol (DAG). IP3 mobilizes intracellular Ca2+ ([Ca2+]i), which can lead to exocytosis. We used activators and inhibitors of known signal transduction pathways to investigate the signaling pathway responsible for LPS-induced exocytosis in the GR. These compounds have been shown to similarly effect pathways in vertebrate and invertebrate systems and this assumption is made here. Pretreatment of GRs with cholera and pertussis toxins, which modulate G-proteins, and U73122, which inhibits PLC, inhibited LPS-induced exocytosis, but pretreatment with the tyrosine kinase inhibitor herbimycin did not. In contrast, exocytosis was induced with fluoride (a G-protein activator) and thapsigargin with Mg2+ (an inhibitor of endomembranous Ca(2+)-ATPase). Exocytosis was not induced by phorbol ester, which mimics DAG to activate protein kinase C (PKC) and it was not effected by ethanol or chelerythrine, which inhibit phospholipase D and PKC, respectively. Microinjection of GRs with different concentrations of IP3, an IP3 analog (DL-2,3,6,trideoxy-myo-inositol 1,4,5-triphosphate), Mg2+, or Ca2+ induced different percentages of exocytosis in individual cells, while HEPES buffer did not. Microfluorometric analysis of intracellular Mg2+ ([Mg2+]i) and [Ca2+]i, using the dyes Mag Fura-2AM and Calcium Green 5N, respectively, revealed [Mg2+]i and [Ca2+]i fluxes during LPS-induced exocytosis. This study suggests that LPS induces exocytosis in the Limulus GR through activation of G-protein-coupled receptors, which stimulate the IP3 signaling pathway to induce both [Ca2+]i and [Mg2+]i fluxes to facilitate vesicular and plasma membrane fusion. This is the first demonstration of the signal transduction pathway responsible for the primary immune response of the GR.
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PMID:Signal transduction during exocytosis in Limulus polyphemus granulocytes. 901 85

Previously, we identified peptides that stimulate phosphoinositide hydrolysis in several leukocyte cell lines from mixtures of random hexapeptide sequences. Moreover, the peptides activate phospholipase C via a pertussis toxin-sensitive G protein-coupled receptor. We now investigate the structure-activity relationship of the peptides with the goal of improving the activity of the peptides, as well as the biologic function of the peptides. Substitution of the L-methionine at the C terminus of peptides with D-methionine markedly increased the effectiveness of the peptides. The half-maximal effective concentrations of MKYMPm-NH2 and WKYMVm-NH2 for stimulation of phosphoinositide hydrolysis in U266 cells were 30 and 0.5 nM, respectively. By BIAcore analysis we confirmed the existence of a receptor for WKYMVm-NH2. Furthermore, the intracellular calcium concentration increase induced by WKYMVm-NH2 was not inhibited by several chemoattractants (FMLP, IL-8, platelet-activating factor, C5a, granulocyte-macrophage CSF, and granulocyte CSF) suggests that WKYMVm-NH2 has a unique cell surface receptor on leukocytes. WKYMVm-NH2 stimulated the phosphoinositide hydrolysis in U937, HL60, and U266 cells, as well as in human neutrophils. Moreover, WKYMVm-NH2 is more effective than FMLP in the production of superoxide in human neutrophils. The data suggest that WKYMVm-NH2 may have the ability to activate the microbicidal functions of human neutrophils.
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PMID:A peptide with unique receptor specificity: stimulation of phosphoinositide hydrolysis and induction of superoxide generation in human neutrophils. 902 31

Differentiation with dibutyryl cyclic AMP (dBcAMP) of the human, premonocytic U937 cell line toward a monocyte/granulocyte-like cell results in the cell acquiring an ability to release arachidonate upon stimulation. In contrast, the calcium ionophore ionomycin was able to stimulate phospholipase C, as measured by inositol 1,4,5-trisphosphate formation, to equal extents in both undifferentiated and dBcAMP-differentiated U937 cells. The role and regulation of cytosolic phospholipase A2 (cPLA2) in the production of arachidonate in these cells when either the chemotactic peptide fMLP or ionomycin are used as stimulus were investigated. The ionomycin- and fMLP-stimulated release of arachidonate were sensitive to the cPLA2 inhibitor arachidonyl trifluoromethylketone (IC50 values of 32 and 18 microM, respectively), but were not inhibited by E-6-(bromomethylene)-tetrahydro-3-(1-naphthalenyl)-2 H-pyran-2-one, a bromoenol lactone inhibitor of the calcium-independent phospholipase A2. These results, coupled with the inhibition of ionomycin-induced arachidonate production by electroporation of differentiated cells to introduce an anti-cPLA2, demonstrate that the cPLA2 is the enzyme responsible for arachidonate release in differentiated cells. SDS-PAGE and immunoblot analysis of differentiated cells showed the cells to contain both phosphorylated and unphosphorylated forms of cPLA2 (ratio of about 2: 3). Surprisingly, undifferentiated cells contain 30% more enzyme than differentiated cells and contain a higher percentage (approximately 75%) of the phosphorylated in the absence of stimulation. The inability of undifferentiated cells to produce arachidonate is not due to insufficient intracellular calcium concentrations since ionomycin induces large (820-940 nM) influxes of intracellular calcium in both differentiated and undifferentiated cells. This demonstrates that phosphorylation of cPLA2 andan influx of intracellular calcium are not sufficient to activate the enzyme to produce arachidonate. Instead, activation of a pertussis toxin-sensitive Gi alpha-type G-protein is required as evidenced by the production of arachidonate in undifferentiated cells stimulated with mastoparan, an activator of Gi alpha subunits, in combination with ionomycin. This activation of a Gi alpha-type G-protein is independent of modulations of adenylyl cyclase activity since cellular cAMP levels were not modulated upon treatment with mastoparan and ionomycin.
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PMID:Phosphorylation and calcium influx are not sufficient for the activation of cytosolic phospholipase A2 in U937 cells: requirement for a Gi alpha-type G-protein. 935 62

The numbers of patients in intensive care units, with immunosuppression, and of elderly people increase in parallel with antibiotic-resistant microorganisms. Therefore the demand for an effective antisepsis increases. Moreover, it became evident that the pathophysiology and the outcome of infection are dependent on the properties of the microorganisms, e.g. synthesis of endo- and exotoxins, and on the host defense, the immune system. In addition to the microbicidal action, we studied the effects of povidone-iodine (PVP-I, Betaisodona) on the generation, release and activity of exotoxins (alpha-hemolysin, phospholipase C, lipase), as well as on granulocyte-derived tissue-destructive enzymes (elastase, beta-glucuronidase) and microbial-induced cytokine generation from human neutrophils. Our results clearly show that PVP-I does not only kill a wide range of bacteria but also inhibits the generation and release of bacterial exotoxins; furthermore, it also inactivates bacterial exotoxins as well as granulocyte-derived tissue-destructive enzymes and cytokines. These data support the usefulness and efficacy of PVP-I as an effective therapeutic agent to combat infection.
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PMID:Effects of Betaisodona on parameters of host defense. 940 54

1. We recently demonstrated the presence of phospholipase C-coupled bradykinin (BK) B2-receptors in human primary and SV40 virus-immortalized corneal epithelial (CEPI) cells. 2. The aims of the present studies were to demonstrate the specific binding of [3H]-BK to CEPI cell membranes and to study its pharmacological characteristics. In addition, we wished to study the functional coupling of the BK receptors to various physiological and pathological mechanisms in the CEPI cells, including phosphoinositide (PI) turnover, intracellular Ca2+-mobilization ([Ca2+]i), cell proliferation (via [3H]-thymidine incorporation), and the release of various cytokines, collagenase-1 (matrix metalloproteinase-1) and prostaglandin E2 (PGE2). 3. Specific [3H]-BK binding comprised 83 +/- 2% of the total binding, and was of high affinity (Kd = 1.66 +/- 0.52 nM, n = 5), saturable (Bmax = 640 +/- 154 fmol g(-1) wet weight) and reversible. Competition studies yielded the following affinity values for BK and a number of BK-related peptides: Hoe-140 (D-Arg-[Hyp3,Thi5,D-Tic7,Oic8]BK; icatibant): Ki = 0.17 +/- 0.07 nM; BK: Ki = 1.0 +/- 0.11 nM; [Tyr8]-BK: Ki = 12.9 +/- 2.3 nM; [des-Arg9]-BK: Ki > 9,200 nM (all n = 3-5)). 4. BK potently stimulated PI turnover (EC50 = 2.3 +/- 0.3 nM; n = 7) and [Ca2+]i mobilization (EC50 = 8-20 nM) in CEPI cells and both responses were inhibited in a concentration-dependent manner by 100 nM-10 microM Hoe-140, a selective B2-receptor antagonist, and also inhibited by the selective phospholipase C (PLC) inhibitor, U73122 (1-(6-((17beta-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1 H-pyrrole-2,5-dione) (IC50 = 3.0 +/- 1.6 microM). BK-induced [Ca2+]i mobilization was reduced by about 30% in the presence of 4 mM EGTA, but was not significantly affected by 100 nM nifedipine. 5. BK (0.1 nM-10 microM) significantly (P<0.05-0.001) stimulated [3H]-thymidine incorporation into CEPI cellular DNA. However, while interleukin-1alpha (IL-1alpha; 10 ng ml(-1)) potently stimulated the release of IL-6, IL-8 and granulocyte macrophage colony-stimulating factor from CEPI cells, BK (0.1 nM-10 microM) was without effect. 6. Whilst phorbol-12-myristate-13-acetate (PMA; 3 microg ml(-1)) and 10% foetal bovine serum (positive control agents) significantly stimulated the release of both MMP-1 and PGE2 from CEPI cells, BK (0.1 nM-10 microM) was without any significant effect under these conditions. 7. In conclusion, these data indicate that the CEPI cells express high-affinity [3H]-BK binding sites representing B2-subtype BK receptors coupled to PI turnover and [Ca2+]i mobilization which appear to stimulate [3H]-thymidine incorporation into cellular DNA. In contrast, BK failed to elicit the release of PGE2, various cytokines and MMP-1 from CEPI cells. These results suggest that BK may have a potential role in corneal epithelium wound healing by stimulating cell proliferation.
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PMID:Effects of bradykinin on signal transduction, cell proliferation, and cytokine, prostaglandin E2 and collagenase-1 release from human corneal epithelial cells. 955 96

In vivo, vascular walls are exposed to mechanical stretch, which may promote atherogenesis. This study was designed to investigate the effect of mechanical stretch on the production and gene expression of cytokines in endothelial cells (ECs) of human umbilical veins. ECs were cultured on flexible silicone membranes and exposed to cyclic mechanical stretch. Although the secretion levels of interleukin (IL)-1beta, tumor necrosis factor-alpha, IL-6, granulocyte (G) -colony stimulating factor (CSF), G and macrophage (M) -CSF, and M-CSF were not affected by cyclic stretch over 24 hours, the levels of IL-8 and monocyte chemotactic and activating factor (MCAF)/monocyte chemoattractant protein-1 (MCP-1) were significantly increased by cyclic stretch. Northern blot analysis indicated that the mRNA levels of IL-8 and MCAF/MCP-1 were upregulated by cyclic stretch as a function of its intensity. Cytochalasin D, which disrupts the actin cytoskeleton, abolished the stretch-induced gene expression of IL-8 and MCAF/MCP-1. In contrast, neither inhibition of stretch-activated ion channels nor disruption of microtubules affected the induction of these chemokines by cyclic stretch. Northern blot analysis using enzyme inhibitors showed that phospholipase C, protein kinase C, and tyrosine kinase were involved in the stretch-induced gene expression of IL-8 and MCAF/MCP-1, whereas cAMP- or cGMP-dependent protein kinase was not. In conclusion, cyclic stretch enhanced the secretion and gene expression of IL-8 and MCAF/MCP-1 in a stretch-dependent fashion, and the integrity of the actin cytoskeleton and activities of phospholipase C, protein kinase C, and tyrosine kinase may be essential in the process of stretch-induced gene induction of IL-8 and MCAF/MCP-1.
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PMID:Cyclic stretch upregulates production of interleukin-8 and monocyte chemotactic and activating factor/monocyte chemoattractant protein-1 in human endothelial cells. 963 28


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