Activation of PGE2-secretion from gastric mucosa by a type I phospholipase C is mediated by a direct release of arachidonic acid.
We investigated the effects of an exogenous Type I phospholipase C (PLC) from clostridium perfringens on arachidonic acid release and prostaglandin synthesis from gastric mucosa by determining PGE2 release from organ cultured rabbit mucosal biopsies as well as PGE2 synthesis and substrate-dependent inactivation of the prostaglandin cyclooxygenase from endogenously released arachidonic acid in mucosal homogenate. PLC dose dependently stimulated PGE2 secretion from organ cultured mucosa to 145% and 245% at 0.1 and 1.0 U/ml during a 60 minute culture period. This effect was not affected by the calmodulin antagonist N-(6-aminohexyl)-1-5-chloro-1-naphthalene-sulfonamide (W-7) or the intracellular calcium chelator 1,2-bis-(2-aminophenoxy)ethane-N,N,N',N',-tetraacetic acid-acetoxymethyl ester (BAPTA-AM). PLC could not be substituted by phorbol-12-myristate 13-acetate (PMA), an analogue of the diacylglycerol second messenger functions. During a 15 minute preincubation of mucosal homogenate at 37 degrees C, 1mM CaCl2 stimulated PGE2 synthesis from endogenous arachidonic acid about 5-fold compared to an EDTA-control. In contrast, the residual prostaglandin synthesizing capacity, determined by incubation with excess 14C-labelled arachidonic acid, was reduced by CaCl2 to 37% of the EDTA-value. Quinacrine, an inhibitor of arachidonic acid release from phosphatidylethanolamine, reduced both the stimulation of PGE2 synthesis and the inactivation of prostaglandin cyclooxygenase. Therefore we conclude, that this Ca(2+)-effect reflects activation of the Ca-dependent phospholipase A2 (PLA2) and, as a consequence, substrate-induced inactivation of the prostaglandin cyclooxygenase.(ABSTRACT TRUNCATED AT 250 WORDS)
Preclik
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University of Ulm, Department of Internal Medicine II, Germany.
Stange
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EF
Ditschuneit
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H
D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism*; D002118:Calcium / Q000494:pharmacology; D002147:Calmodulin / Q000037:antagonists & inhibitors; D015232:Dinoprostone / Q000096:biosynthesis / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D005753:Gastric Mucosa / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D009924:Organ Culture Techniques; D010740:Phospholipases / Q000037:antagonists & inhibitors; D011817:Rabbits; D011863:Radioimmunoassay; D013449:Sulfonamides / Q000494:pharmacology; D010738:Type C Phospholipases / Q000378:metabolism*
D002147:Calmodulin; D013449:Sulfonamides; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D016718:Arachidonic Acid; D004533:Egtazic Acid; C017967:W 7; D010740:Phospholipases; D010738:Type C Phospholipases; D015232:Dinoprostone; D002118:Calcium