Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.4.3 (phospholipase C)
 18,461 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Apoptotic proteases cleave and inactivate survival signaling molecules such as Akt/PKB, phospholipase C (PLC)-gamma1, and Bcl-2. We have found that treatment of A431 cells with tumor necrosis factor-alpha in the presence of cycloheximide resulted in the cleavage of epidermal growth factor receptor (EGFR) as well as the activation of caspase-3. Among various caspases, caspase-1, caspase-3 and caspase-7 were most potent in the cleavage of EGFR in vitro. Proteolytic cleavage of EGFR was inhibited by both YVAD-cmk and DEVD-fmk in vitro. We also investigated the effect of caspase-dependent cleavage of EGFR upon the mediation of signals to downstream signaling molecules such as PLC-gamma1. Cleavage of EGFR by caspase-3 significantly impaired the tyrosine phosphorylation of PLC-gamma1 in vitro. Given these results, we suggest that apoptotic protease specifically cleaves and inactivates EGFR, which plays crucial roles in anti-apoptotic signaling, to abrogate the activation of EGFR-dependent downstream survival signaling molecules.
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PMID:Proteolytic cleavage of epidermal growth factor receptor by caspases. 1122 7

Phospholipase activities are thought to be involved in the activation of macrophages by lipopolysaccharide (LPS). Because our previous studies showed that the synthetic lipopeptide JBT3002 might activate macrophages via signaling pathways similar to those used by LPS, we investigated whether phospholipase activities are required for activation of macrophages by JBT3002. Treatment of RAW264.7 murine macrophage-like cells with JBT3002 stimulated expression of both inducible nitric oxide synthase (iNOS) and tumor necrosis factor-alpha (TNF-alpha) in a dose-dependent manner. The JBT3002-induced production of nitric oxide and TNF-alpha was significantly inhibited by tricyclodecan-9-yl xanthogenate (D609), a selective inhibitor of phosphatidylcholine (PC)-specific phospholipase C (PC-PLC). JBT3002-induced expression of steady-state mRNA for both iNOS and TNF-alpha was inhibited by D609. Cells treated with JBT3002 had greater production of diacylglycerol (DAG) in 2 min, which lasted for at least 30 min and could be blocked by D609. Activation of RAW264.7 cells was not affected by butanol, a PC-specific phospholipase D inhibitor, and treatment with JBT3002 did not affect phosphatidic acid formation. RAW264.7 cells treated with DAG analogue 1-oleoyl-2-acetyl-sn-glycerol, in the presence of interferon-gamma, produced TNF-alpha. These results suggested that activation of RAW264.7 cells by JBT3002 requires PC-PLC activity.
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PMID:Phosphatidylcholine-specific phospholipase C regulates activation of RAW264.7 macrophage-like cells by lipopeptide JBT3002. 1140 95

The purpose of these studies was to identify the role of phospholipases in the activation of macrophages by lipopolysaccharide (LPS). Tricyclodecan-9-yl-xanthogenate (D609), an inhibitor of phosphatidylcholine-specific phospholipase C (PC-PLC); butanol, an inhibitor of phosphatidylcholine phospholipase D (PC-PLD); and propranolol, an inhibitor of phosphatidate phosphohydrolase, were used in the study. Treatment of RAW264.7 murine macrophage-like cells with LPS resulted in expression of inducible nitric oxide synthase and tumor necrosis factor-alpha. The expression was partially inhibited by D609, butanol, or propranolol and was completely blocked by the combination of D609 and butanol. RAW264.7 cells constitutively produced low basal levels of diacylglycerol and phosphatidic acid; production of both was significantly increased after stimulation with LPS, reaching a peak in 2-3 min and remaining elevated after 30 min. In LPS-induced RAW264.7 cells, diacylglycerol was suppressed by each of the three inhibitors alone and almost abolished by D609 plus butanol or D609 plus propranolol. Phosphatidic acid was reduced to basal level by butanol after LPS stimulation for 2.5 min and by butanol plus D609 after LPS stimulation for 2.5 or 10 min. Taken together, these data indicate that activation of RAW264.7 cells by LPS can be mediated by the activities of both PC-PLC and PC-PLD.
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PMID:Phosphatidylcholine-specific phospholipase C and D in stimulation of RAW264.7 mouse macrophage-like cells by lipopolysaccharide. 1146 Mar 17

Insulin resistance (IR) remains one of the major pathogenic mechanisms for non-insulin-dependent type 2 diabetes mellitus. We have previously modelled IR in H-411E liver cells in culture. In past experiments, we used both labeled glucose uptake, lipogenesis, and stimulation of calmodulin gene expression to quantify the ability of the antidiabetic drugs (pioglitazone and metformin) to reverse tumor necrosis factor-alpha (TNF-alpha)-induced IR in these insulin-treated cells. In these current experiments, H-411E liver cells were rendered IR by a combination of TNF-alpha and insulin. In other experiments, the ability of C2 ceramide (Cer) to inhibit insulin action and induce IR was assessed as well as the phospholipase C inhibitor D609 to reverse IR induced by these TNF-alpha-like agents. C2 Cer, like TNF-alpha, inhibited insulin action. D609 reversed TNF-alpha induced--and to a lesser extent, C2 Cer-induced--IR. At selected times, the cells were also treated with troglitazone (TRG) in 2 groups: (1) 1-time exposure and (2) chronic exposure followed by acute exposure. TRG concentrations ranged from 0.015 to 15.0 micromol/L. Our data demonstrate a powerful effect of TRG in reducing IR and restoring insulin sensitivity in TNF-alpha-treated H-411E cells. Furthermore, pretreatment with TRG, reflecting chronic exposure, as in human clinical use, was more potent than 1-time acute exposure. These data support the efficacy of using thiazolidinediones (TRG) in human type 2 diabetes, and support the use of this cell culture model to further study the effects of thiazolidinediones on TNF-alpha-induced insulin resistance.
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PMID:Mechanisms involved in tumor necrosis factor-alpha induction of insulin resistance and its reversal by thiazolidinedione(s). 1152 31

The Ca(2+) signaling system in an established immortalized rat parotid acinar cell line, Par-C5, was examined using the Ca(2+)-sensitive fluorescent indicator fura-2 and by measuring inositol 1,4,5-trisphosphate (IP(3)) formation. Agonist-induced increase in intracellular Ca(2+) ([Ca(2+)](i)) by mobilization of intracellular stores and influx across the cell membrane was stimulated by acetylcholine (ACh) and ATP, whereas noradrenaline-(NA)-induced a small [Ca(2+)](i) increase mediated primarily by release from intracellular Ca(2+) stores. [Ca(2+)](i) increase by ACh and ATP was mediated through the phosphoinositide signal pathway since both agonists significantly increased 1,4,5-IP(3) formation and Ca(2+) mobilization was abolished by the phospholipase C inhibitor U73122. In Ca(2+)-free medium, ACh or ATP discharged the IP(3)-sensitive Ca(2+) store and essentially abolished subsequent [Ca(2+)](i) response to thapsigargin (TG). Exposure to ionomycin and monensin after TG induced a further mobilization of Ca(2+), suggesting IP(3)-insensitive stores are present. Furthermore, depletion of IP(3)-sensitive Ca(2+) stores by TG, ACh and ATP enhanced plasmalemmal Ca(2+)-entry pathways. Exposure to tumor necrosis factor-alpha (TNF-alpha), a cytokine associated with lymphocyte invasion of salivary epithelial cells in autoimmune disorders, significantly reduced ACh-stimulated Ca(2+) mobilization. TNF-alpha inhibitory effect on Ca(2+) mobilization was not directly due to an interaction on muscarinic receptors since ACh-induced 1,4,5-IP(3) formation was not altered. These results in the Par-C5 cell line indicate 1) [Ca(2+)](i) is regulated by muscarinic and P2Y-nucleotide receptors and partly by alpha(1)-adrenergic receptors; 2) IP(3)-sensitive and -insensitive Ca(2+) stores exist; 3) Ca(2+) influx activated by ACh, ATP or TG is mediated by the store-operated Ca(2+) entry pathway; and 4) muscarinic agonist-stimulated Ca(2+) mobilization is altered by the cytokine TNF-alpha.
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PMID:Regulation of Ca(2+) signals in a parotid cell line Par-C5. 1168 33

This study was performed to investigate the in vivo effects of staphylococcal alpha-toxin on phagocytosis and the secretion of proinflammatory cytokines at local sites of intraperitoneal toxin-challenged mice. A dosage of 45 hemolytic units (HU) of alpha-toxin induced a marked increase in the peritoneal neutrophil count. The toxin caused a 52% decrease in phagocytosis by peritoneal macrophages, compared with that of control mice receiving Staphylococcus aureus particles alone. However, no effect on phagocytosis in neutrophils was observed. A dosage of 45 HU toxin and the synergistic activity of S. aureus particles strongly induced interleukin (IL) 6 secretion but only mildly induced IL-1alpha secretion. The toxin did not induce the secretion of tumor necrosis factor-alpha (TNF-alpha). Interestingly, S. aureus culture supernatant induced the secretion of TNF-alpha in cultured macrophages. These results suggest that alpha-toxin damages the primary host defense system by inducing the oversecretion of IL-1alpha and IL-6, but not TNF-alpha, via a mechanism that requires the synergistic action of bacterial components.
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PMID:Staphylococcal alpha-toxin synergistically enhances inflammation caused by bacterial components. 1198 63

Previously, we histochemically examined the localization of tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) and its receptors in porcine ovarian follicles, and demonstrated a marked reduction in the expression of TRAIL-decoy receptor-1 (DcRI) in granulosa cells of atretic follicles. In the present study, to confirm the inhibitory activity of DcR1 in granulosa cells, granulosa cells prepared from healthy follicles were treated with phosphatidylinositol-specific phospholipase C (PI-PLC) to cleave glycophospholipid anchor of DcR1 and to remove DcR1 from the cell surface, and then incubated with TRAIL. PI-PLC treatment increased the number of apoptotic cells induced by TRAIL. The present finding indicated the possibility that TRAIL and its receptors were involved in induction of apoptosis in granulosa cells during atresia, and that DcR1 plays an inhibitory role in granulosa cell apoptosis.
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PMID:TRAIL-decoy receptor 1 plays inhibitory role in apoptosis of granulosa cells from pig ovarian follicles. 1206 77

Interleukin-1alpha (IL-1alpha) and tumor necrosis factor-alpha (TNF-alpha) markedly stimulate glucose utilization in primary cultures of mouse cortical astrocytes. The mechanism that gives rise to this effect, which takes place several hours after application of cytokine, has remained unclear. Experiments were conducted to identify the major signaling cascades involved in the metabolic action of cytokine. First, the selective IL-1 receptor antagonist (IL-1ra) prevents the effect of IL-1alpha on glucose utilization in a concentration-dependent manner, whereas it has no effect on the action of TNF-alpha. Then, using inhibitors of three classical signaling cascades known to be activated by cytokines, it appears that the PI3 kinase is essential for the effect of both IL-1alpha and TNF-alpha, whereas the action of IL-1alpha also requires activation of the MAP kinase pathway. Participation of a phospholipase C-dependent pathway does not appear critical for both IL-1alpha and TNF-alpha. Inhibition of NO synthase by L-NAME did not prevent the metabolic response to both IL-1alpha and TNF-alpha, indicating that nitric oxide is probably not involved. In contrast, the Na(+)/K(+) ATPase inhibitor ouabain prevents the IL-1alpha- and TNF-alpha-stimulated 2-deoxyglucose (2DG) uptake. When treatment of astrocytes with a cytokine was followed 24 h later by an acute application of glutamate, a synergistic enhancement in glucose utilization was observed. This effect was greatly reduced by ouabain. These data suggest that Na(+) pump activity is a common target for both the long-term metabolic action of cytokines promoted by the activation of distinct signaling pathways and the enhanced metabolic response to glutamate.
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PMID:Long-term modulation of glucose utilization by IL-1 alpha and TNF-alpha in astrocytes: Na+ pump activity as a potential target via distinct signaling mechanisms. 1211 71

This study was first designed to investigate systematically the kinetics of surface expression of scavenger receptors (SRs) and CD14 on alveolar macrophages in vivo and in vitro and their relation with local pro- and antiinflammatory responses in endotoxemia. The expression of SR and CD14 in lungs was down- and up-regulated, respectively, in the presence of endotoxemia, which might be due to decreased expression of SR and increased expression of CD14 on the surface of the resident macrophages. Down-regulation of SRs on alveolar macrophages not only induces decreased defensive function of the macrophages, it also enhances lipopolysaccharide (LPS)-induced activation of alveolar macrophages possibly through increasing LPS binding to CD14. Although CD14 is a key receptor responsible for LPS to activate macrophages, both phospholipase C and anti-CD14 antibody can completely inhibit activation of alveolar macrophages initiated by only LPS 1 ng/ml, as determined by tumor necrosis factor-alpha (TNFalpha) production, but it does not significantly change TNFalpha release upon cell stimulation by LPS 10 microg/ml. There was an intrinsic relation of enhanced intrapulmonary pro- and antiinflammatory responses with changes in SR and CD14 expression, which suggests that the down-regulation of SR and up-regulation of CD14 might be an important mechanism for the lung to change from a defense organ to an effector organ during sepsis.
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PMID:Intrapulmonary expression of scavenger receptor and CD14 and their relation to local inflammatory responses to endotoxemia in mice. 1255 30

Neutral sphingomyelinase (N-SMase) is one of the key enzymes involved in the generation of ceramide; however, the gene(s) encoding for the mammalian N-SMase is still not well defined. Previous studies on the cloned nSMase1 had shown that the protein acts primarily as lyso-platelet-activating factor-phospholipase C. Recently the cloning of another putative N-SMase, nSMase2, was reported. In this study, biochemical characterization of the mouse nSMase2 was carried out using the overexpressed protein in yeast cells in which the inositol phosphosphingolipid phospholipase C (Isc1p) was deleted. N-SMase activity was dependent on Mg(2+) and was activated by phosphatidylserine and inhibited by GW4869. The ability of nSMase2 to recognize endogenous sphingomyelin (SM) as substrate was investigated by overexpressing nSMase2 in MCF7 cells. Mass measurements showed a 40% decrease in the SM levels in the overexpressor cells, and labeling studies demonstrated that nSMase2 accelerated SM catabolism. Accordingly, ceramide measurement showed a 60 +/- 15% increase in nSMase2-overexpressing cells compared with the vector-transfected MCF7. The role of nSMase2 in cell growth was next investigated. Stable overexpression of nSMase2 resulted in a 30-40% decrease in the rate of growth at the late exponential phase. Moreover, tumor necrosis factor induced approximately 50% activation of nSMase2 in MCF7 cells overexpressing the enzyme, demonstrating that nSMase2 is a tumor necrosis factor-responsive enzyme. In conclusion, these results 1) show that nSMase2 is a structural gene for nSMase, 2) suggest that nSMase2 acts as a bona fide N-SMase in cells, and 3) implicate nSMase2 in the regulation of cell growth and cell signaling.
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PMID:Biochemical properties of mammalian neutral sphingomyelinase 2 and its role in sphingolipid metabolism. 1256 38


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