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)

Previous studies have shown that thrombin-activated platelets interact through the P-selectin with neutrophils and monocytes. To identify other types of leukocytes capable of such an interaction, eosinophils, basophils, and lymphocytes were isolated from whole blood. Binding of these cells to activated platelets was examined in a double immunofluorescence assay and the results show that activated platelets not only bind to neutrophils and monocytes, but also to eosinophils, basophils, and subpopulations of T lymphocytes. Using monoclonal antibodies (MoAbs) specific for subsets of T cells, we could further demonstrate that the T cells which bind activated platelets are natural killer (NK) cells and an undefined subpopulation of CD4+ and CD8+ cells. All these interactions were dependent on divalent cations and were completely inhibited by an MoAb against P-selectin. Thus, P-selectin mediates the binding of activated platelets to many different types of leukocytes. Studies with leukocytes treated with proteases or neuraminidase have shown that the structures recognized by P-selectin are glycoproteins carrying sialic acid residues. Because the loss of binding of activated platelets to neuraminidase-treated neutrophils was almost complete, but only partial to treated eosinophils, basophils, and monocytes, the latter cell types may have different P-selectin ligands in addition to those present on neutrophils. We found that two previously identified ligands for P-selectin, the oligosaccharides Le(x) and sialyl-Le(x), had little or no inhibitory effect on adhesion of activated platelets to leukocytes and that binding was not inhibited by MoAbs against these oligosaccharides. In addition, there was no correlation between the expression of Le(x) on several cell types and their capacity to bind activated platelets. In contrast, the expression of sialyl-Le(x) on cells was almost perfectly correlated with their ability to bind activated platelets. Thus, while Le(x) cannot be a major ligand for P-selectin, a possible role for sialyl-Le(x) in P-selectin-mediated adhesion processes cannot be dismissed. Finally, activated platelets were found to bind normally to monocytes and neutrophils of patients with paroxysmal nocturnal hemoglobulinuria (PNH) and to neutrophils from which phosphatidyl inositol (PI)-linked proteins had been removed by glycosylphosphatidyl inositol-specific phospholipase C (GPI-PLC) digestion. This suggests that at least part of the P-selectin ligands on these cells are not GPI-anchored.
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PMID:P-selectin mediates Ca(2+)-dependent adhesion of activated platelets to many different types of leukocytes: detection by flow cytometry. 137 47

Adhesion of polymorphonuclear leukocytes (PMN) to endothelial cells is an essential step in inflammatory reactions. We characterized the effects of two important bacterial exotoxins, Escherichia coli hemolysin (HlyA) and Staphylococcus aureus alpha-toxin (S. alpha-toxin) on PMN adhesion to cultured HUVEC. Both toxins increased adherence of human PMN to HUVEC in a dose- and time-dependent manner, peaking after 30 min at 0.01 hemolytic units/ml HlyA or 0.5 microg/ml S. alpha-toxin. Pretreatment of HUVEC with anti-P-selectin mAbs or of PMN with anti-CD11b/CD18 mAb reduced HlyA- and S. alpha-toxin-related cell adhesion significantly. Increased P-selectin expression on toxin-treated endothelial cells was demonstrated by cell surface ELISA. Compared with endotoxin, HlyA and S. alpha-toxin did not induce the expression of E-selectin, ICAM-1, or VCAM-1. FACS analysis showed increased CD11b/CD18 expression on HlyA-, but not on S. alpha-toxin-stimulated PMN. Platelet-activating factor, an important costimulatory factor for PMN adhesion and activation, was also active in the exotoxin-stimulated adhesion system, as evidenced by studies using the platelet-activating factor receptor antagonist BN50727. HPLC analysis of endothelial cell extracts confirmed enhanced toxin-mediated PAF synthesis. The capacity of exotoxins to stimulate PMN adhesion to endothelial cells may be relevant in patients with severe local or systemic bacterial infections.
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PMID:Escherichia coli hemolysin and Staphylococcus aureas alpha-toxin potently induce neutrophil adhesion to cultured human endothelial cells. 889 49

P-selectin (CD62P) is a Ca2+-dependent endogenous lectin that can be expressed by vascular endothelium and platelets. The major ligand for P-selectin on leukocytes is P-selectin glycoprotein ligand-1 (PSGL-1). P-selectin can also bind to carcinoma cells, but the nature of the ligand(s) on these cells is unknown. Here we investigated the P-selectin binding to a breast and a small cell lung carcinoma cell line that are negative for PSGL-1. We report that CD24, a mucin-type glycosylphosphatidylinositol-linked cell surface molecule on human neutrophils, pre B lymphocytes, and many tumors can promote binding to P-selectin. Latex beads coated with purified CD24 from the two carcinoma cell lines but also neutrophils could bind specifically to P-selectin-IgG. The binding was dependent on divalent cations and was abolished by treatment with O-sialoglycoprotein endopeptidase but not endoglycosidase F or sialidase. The beads were stained with a monoclonal antibody (MoAb) to CD57 (HNK-1 carbohydrate epitope) but did not react with MoAbs against the sialylLe(x/a) epitope. The carcinoma cells and CD24-beads derived from these cells could bind to activated platelets or P-selectin transfected Chinese hamster ovary cells (P-CHO) in a P-selectin-dependent manner and this binding was blocked by soluble CD24. Transfection of human adenocarcinoma cells with CD24 enhanced the P-selectin-dependent binding to activated platelets. Treatment of the carcinoma cells or the CD24 transfectant with phosphatidylinositol-specific phospholipase C reduced CD24 expression and P-selectin-IgG binding concomitantly. These results establish a role of CD24 as a novel ligand for P-selectin on tumor cells. The CD24/P-selectin binding pathway could be important in the dissimination of tumor cells by facilitating the interaction with platelets or endothelial cells.
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PMID:CD24, a mucin-type glycoprotein, is a ligand for P-selectin on human tumor cells. 912 46

Alpha toxin from Clostridium perfringens type A, a phospholipase C, has been implicated in many of the localized and systemic features of gas gangrene. We demonstrated that human endothelial cells synthesize two vasoactive lipids, platelet-activating factor (PAF) and prostacyclin, in response to alpha toxin treatment. The stimulated synthesis of PAF required the enzymatic activity of the toxin and subsequent protein kinase C activation. Alpha toxin-treated endothelial cells accumulated the products of the phospholipase C reaction, diacylglycerol and ceramide, and exhibited a decrease in the enzymatic precursors phosphatidylcholine and sphingomyelin. Furthermore, the temporal accumulation of PAF depended on the concentration of the toxin in the overlying medium and was blocked in the presence of a neutralizing antibody. The cultured endothelial cells also exhibited enhanced neutrophil adhesion in response to alpha toxin which was mediated through the PAF receptor and P-selectin. P-selectin expression by endothelial cells and extravascular neutrophil accumulation were also observed in tissue sections from alpha toxin-injected Sprague-Dawley rats. These endothelial cell-mediated processes are important in maintaining vascular homeostasis and, when activated in a dysregulated manner by C. perfringens alpha toxin, may contribute to localized and systemic manifestations of gas gangrene including enhanced vascular permeability, localized neutrophil accumulation, and myocardial dysfunction.
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PMID:Alpha toxin from Clostridium perfringens induces proinflammatory changes in endothelial cells. 923 3

P-selectin mediates rolling of neutrophils and other leukocytes on activated endothelial cells and platelets through binding to P-selectin glycoprotein ligand-1 (PSGL-1). Certain PSGL-1 negative tumor cell lines can bind P-selectin under static conditions through the GPI-linked surface mucin, CD24, but the physiological significance of this interaction and whether it can occur under flow conditions is not known. Here, we show that CD24+ PSGL-1- KS breast carcinoma cells attach to and roll on recombinant P-selectin under a continuous wall shear stress, although at a lower density and higher velocity than CD24+ PSGL-1+ cells, such as HL-60. Adding excess soluble CD24 or removing CD24 from the cell surface with phosphatidylinositol-phospholipase C (PI-PLC) significantly reduced KS cell rolling on P-selectin. The ability of KS cells to roll on P-selectin was positively correlated with the CD24 expression level. Comparison with three other CD24+ cell lines established that expression of sialyl-Lewis(x) antigen was also necessary for CD24-mediated rolling on P-selectin. CD24 purified from KS cells supported rolling of P-selectin transfectants, but not L-selectin transfectants. Finally, KS cells rolled on vascular endothelium in vivo in a P-selectin-dependent manner. Together our data show that CD24 serves as a ligand for P-selectin under physiological flow conditions. Interaction of tumor cells with P-selectin via CD24 may be an important adhesion pathway in cancer metastasis.
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PMID:CD24 mediates rolling of breast carcinoma cells on P-selectin. 973 27

The surface expression of prion protein (PrP(C)) on human platelets, as detected by flow cytometry with the monoclonal antibody 3F4, increased more than two-fold (4300 v 1800 molecules/platelet) after full activation. Maximal surface expression of PrP(C) occurred within 3 min of platelet activation and declined to approximately half of maximal levels by 2 h at 37 degrees C. In comparison, PrP(C) on the surface of platelets, activated at 22 degrees C took 10 min to reach maximum but then remained constant for 2 h. In sonicated resting platelets, PrP(C) and P-selectin remained in intact granules after subcellular fractionation. Both glycoproteins were found in the ruptured membranes of activated platelets, suggesting that the PrP(C) was translocated from internal granules to the plasma membrane during activation, as is P-selectin. Platelet PrP(C) was not removed from the surface of platelets by phosphatidylinositol-specific phospholipase C (PIPLC) treatment but was degraded by proteinase K. Platelets may serve as a useful model for following the cellular processing of PrP(C).
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PMID:Increased expression of phosphatidylinositol-specific phospholipase C resistant prion proteins on the surface of activated platelets. 1099 93

In order to better understand granule release from platelets, we developed an alpha-toxin permeabilized platelet model to study alpha-granule secretion. Secretion of alpha-granules was analyzed by flow cytometry using P-selectin as a marker for alpha-granule release. P-selectin surface expression occurred when platelets were permeabilized in the presence of Ca2+. Responsiveness to Ca2+ was lost 30 min after permeabilization but could be reconstituted with MgATP. Alpha-toxin-permeabilized, MgATP-exposed platelets also degranulated within a pH range of 5.4-5.9 without exposure to and independent of Ca2+. ATP, GTP, CTP, UTP, and ITP supported Ca2+-induced alpha-granule secretion, while H+-induced alpha-granule secretion occurred only with ATP and GTP. Both Ca2+- and H+-induced alpha-granule secretion required ATP hydrolysis. Kinase inhibitors blocked both Ca2+- and H+-induced secretion. These data suggest that alpha-granule secretion in this permeabilized platelet system shares many characteristics with granule secretion studied in other permeabilized cell models. Furthermore, these results show that H+ can trigger alpha-granule release independent of Ca2+.
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PMID:Alpha-granule secretion from alpha-toxin permeabilized, MgATP-exposed platelets is induced independently by H+ and Ca2+. 1008 26

Mechanisms responsible for the rapid tissue destruction in gas gangrene are not well understood. To examine the early effects of Clostridium perfringens exotoxins on tissue perfusion, a rat model of muscle blood flow was developed. Intramuscular injection of a clostridial toxin preparation containing both phospholipase C (PLC) and theta-toxin caused a rapid (1-2 min) and irreversible decrease in blood flow that paralleled formation of activated platelet aggregates in venules and arterioles. Later (20-40 min), aggregates contained fibrin and leukocytes, and neutrophils accumulated along vascular walls. Flow cytometry confirmed that these clostridial toxins or recombinant PLC induced formation of P-selectin-positive platelet aggregates. Neutralization of PLC activity in the clostridial toxin preparation completely abrogated human platelet responses and reduced perfusion deficits. It is concluded that tissue destruction in gas gangrene is related to profound attenuation of blood flow initiated by activation of platelet responses by PLC.
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PMID:Clostridial gas gangrene. I. Cellular and molecular mechanisms of microvascular dysfunction induced by exotoxins of Clostridium perfringens. 1095 Jul 74

Carbohydrates on tumor cells have been shown to play an important role in tumor metastasis. We demonstrated before that CD24, a Mr 35,000-60,000 mucine-type glycosylphosphatidylinositol-linked cell surface molecule, can function as ligand for P-selectin and that the sialylLex carbohydrate is essential for CD24-mediated rolling of tumor cells on P-selectin. To investigate the role of both antigens more closely, we transfected human A125 adenocarcinoma cells with CD24 and/or fucosyltransferase VII (Fuc TVII) cDNAs. Stable transfectants expressed CD24 and/or sialylLex. Biochemical analysis confirmed that in A125-CD24/FucTVII double transfectants, CD24 was modified with sialylLex. Only double transfectants showed rolling on P-selectin in vivo. When injected into mice, double transfectants arrested in the lungs, and this step was P-selectin dependent because it was strongly enhanced in lipopolysaccharide (LPS) pretreated wild-type mice but not in P-selectin knockout mice. CD24 modified by sialylLex was required on the tumor cells because the LPS-induced lung arrest was abolished by removal of CD24 from the cell surface by phosphatidylinositol-specific phospholipase C. A125-FucTVII single transfectants expressing sialylLex but not CD24 did not show P-selectin-mediated lung arrest. The sialylLex epitope is abundantly expressed on human carcinomas, and significant correlations between sialylLex expression and clinical prognosis exist. Our data suggest an important role for sialylLex-modified CD24 in the lung colonization of human tumors.
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PMID:The CD24/P-selectin binding pathway initiates lung arrest of human A125 adenocarcinoma cells. 1111 57

Thrombin-induced activation of RhoA and its involvement in the regulation of myosin II light chain(20) phosphorylation (MLC-P) in alpha-toxin permeabilized platelets was investigated. Permeabilized platelets, expressing normal levels of P-selectin, displayed a Ca(2+)-dependent increase in shape change and MLC-P. Thrombin activated RhoA as measured by a rhotekin-binding assay within 30 s of stimulation under conditions of constant [Ca(2+)](i). Under the same conditions and timecourse, thrombin or GTPgammaS induced an increase in MLC-P and platelet shape change which was not dependent on an increase in [Ca(2+)](i). The thrombin- and GTPgammaS-induced MLC-P in constant [Ca(2+)](i) was inhibited by the addition of Y27632, a Rho-kinase inhibitor. This study directly demonstrates that thrombin can activate RhoA in platelets in a timecourse compatible with a role in increasing MLC-P and shape change (not involving an increase in [Ca(2+)](i)). This is also Rho-kinase-dependent.
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PMID:Thrombin-induced activation of RhoA in platelet shape change. 1154 55


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