title: A novel inhibitory effect on prostacyclin synthesis of coupling factor 6 extracted from the heart of spontaneously hypertensive rats.
abstract: The possible presence of an unknown prostacyclin synthesis inhibitory substance has been reported in some strains of rats. We purified the inhibitory substance from the heart of spontaneously hypertensive rats by collecting active fractions after gel-filtration column chromatography and two steps of reverse-phase high performance liquid chromatography. The amino acid composition and automated gas-phase sequencing of the full-length substance and fragments cleaved by AspN indicated that the prostacyclin-inhibitory peptide was identical to coupling factor 6. Recombinant rat coupling factor 6, which was synthesized using a cleavable fusion protein strategy, attenuated base-line and bradykinin (10(-6) M)-induced prostacyclin synthesis and [3H]arachidonic acid (AA) release in human umbilical vein endothelial cells in a dose-dependent manner (10(-9)-10(-7) M). Exogenous AA- and prostaglandin H2-induced prostacyclin synthesis were unchanged even after treatment with 10(-7) M recombinant coupling factor 6. Base-line and bradykinin-induced [3H]AA release were suppressed by arachidonyltrifluoromethyl ketone, a relatively specific inhibitor of cytosolic phospholipase A2 at 40 microM, and simultaneous administration of coupling factor 6 showed no further effect. Neither oleyloxyethyl phosphorylcholine at 1 microM nor bromoenol lactone at 1 microM affected AA release. Preincubation (1 min) with 10(-7) M recombinant coupling factor 6 had no influence on adenosine diphosphate- and collagen-induced platelet aggregations. We conclude that coupling factor 6 possesses a novel function of prostacyclin synthesis inhibition in endothelial cells via suppression of Ca2+-dependent cytosolic phospholipase A2, although it is unclear whether coupling factor 6 functions in normal conditions or only in pathophysiological states.
authors.affiliation: Second Department of Internal Medicine, Hirosaki University School of Medicine, Hirosaki 036-8562, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Osanai
authors.forename: T
authors.initials: T
authors.lastname: Kamada
authors.forename: N
authors.initials: N
authors.lastname: Fujiwara
authors.forename: T
authors.initials: T
authors.lastname: Katoh
authors.forename: K
authors.initials: K
authors.lastname: Takahashi
authors.forename: M
authors.initials: M
authors.lastname: Kimura
authors.forename: K
authors.initials: K
authors.lastname: Satoh
authors.forename: K
authors.initials: K
authors.lastname: Magota
authors.forename: S
authors.initials: S
authors.lastname: Kodama
authors.forename: T
authors.initials: T
authors.lastname: Tanaka
authors.forename: K
authors.initials: K
authors.lastname: Okumura
mesh_terms: D000251:Adenosine Triphosphatases / Q000302:isolation & purification / Q000494:pharmacology / Q000502:physiology*; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D001920:Bradykinin / Q000494:pharmacology*; D002478:Cells, Cultured; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism*; D011464:Epoprostenol / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D006801:Humans; D006973:Hypertension / Q000378:metabolism*; D007700:Kinetics; D025261:Mitochondrial Proton-Translocating ATPases; D009206:Myocardium / Q000378:metabolism*; D009928:Organ Specificity; D010086:Oxidative Phosphorylation Coupling Factors / Q000494:pharmacology / Q000502:physiology; D051381:Rats; D011918:Rats, Inbred SHR; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D011994:Recombinant Proteins / Q000494:pharmacology; D014471:Umbilical Veins
chemical_list: D010086:Oxidative Phosphorylation Coupling Factors; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D016718:Arachidonic Acid; D011464:Epoprostenol; D000251:Adenosine Triphosphatases; C000628217:F(6) ATPase; D025261:Mitochondrial Proton-Translocating ATPases; D001920:Bradykinin