The role of the phosphatidylinositol turnover in 12-hydroxyeicosatetraenoic acid generation from human platelets by Escherichia coli alpha-haemolysin, thrombin and fluoride.
Activation of human platelets with either the Escherichia coli alpha-haemolysin or thrombin, or with pharmacological agonists such as sodium fluoride (NaF), Ca-ionophore A23187 or phorbol myristate acetate (PMA), induced a similar pattern of serotonin release, unlike 12-hydroxyeicosatetraenoic acid (12-HETE) generation. In the presence of neomycin (0.1, 1, 10 mM), an inhibitor of the phosphatidylinositol-specific phospholipase C (PIP2-PLC), the E. coli alpha-haemolysin-induced 12-HETE generation was enhanced up to threefold in a dose-dependent manner. 12-HETE generation by NaF and thrombin was slightly inhibited at high neomycin concentrations (10 mM). Treatment of human platelets with E. coli alpha-haemolysin induced a different activation pattern of PIP2-PLC and phosphatidylinositol4-kinase (PI4-kinase), compared to NaF and thrombin. Haemolysin treatment resulted in a down-regulation of PIP2-PLC and PI4-kinase enzymatic activities by 20 +/- 9/40 +/- 8% compared to unstimulated cells; the decrease in enzymatic activities was observed within 2 min of stimulation and was still apparent after 60 min of stimulation. In NaF- and thrombin-stimulated platelets, dose- and time-dependent increases in PIP2-PLC (by up to 21 +/- 10%, 34 +/- 11%, respectively) and PI4-kinase (by up to 71 +/- 18, 54 +/- 14) activities were measured. Maximal enzymatic activities were reached after 5-20 min of stimulation (NaF, thrombin) followed by a decline to baseline levels (thrombin) or below baseline levels (NaF). Our results indicate that the phosphatidylinositolphosphate metabolism plays an important role in the regulation of 12-HETE release from human platelets by E. coli alpha-haemolysin.
König
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Lehrstuhl für Med. Mikrobiologie und Immunologie, AG Infektabwehrmechanismen, Ruhr-Universität Bochum, Germany.
König
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D019870:1-Phosphatidylinositol 4-Kinase; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D001426:Bacterial Proteins / Q000494:pharmacology*; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D029968:Escherichia coli Proteins; D006460:Hemolysin Proteins / Q000494:pharmacology*; D006801:Humans; D006893:Hydroxyeicosatetraenoic Acids / Q000097:blood*; D009355:Neomycin / Q000494:pharmacology; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D010716:Phosphatidylinositols / Q000097:blood*; D054801:Phosphoinositide Phospholipase C; D010727:Phosphoric Diester Hydrolases / Q000097:blood; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000097:blood; D012969:Sodium Fluoride / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D006460:Hemolysin Proteins; C036507:Hlya protein, E coli; D006893:Hydroxyeicosatetraenoic Acids; D010716:Phosphatidylinositols; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D012969:Sodium Fluoride; D017853:Phosphotransferases (Alcohol Group Acceptor); D019870:1-Phosphatidylinositol 4-Kinase; D010727:Phosphoric Diester Hydrolases; D054801:Phosphoinositide Phospholipase C; D013917:Thrombin; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D009355:Neomycin