Synthesis of 12(R)- and 12(S)-hydroxyeicosatetraenoic acid by porcine ocular tissues.
Microsomal fractions from porcine ocular tissues synthesized 12(S)-5,8,10,14-hydroxyeicosatetraenoic acid [12(S)-HETE] from arachidonic acid by a membrane-bound lipoxygenase and 12(R)-HETE by the cytochrome P450-dependent monooxygenase system. Both activities were the highest in corneal microsomes. The 12(R)-HETE synthesizing activity of corneal microsomes was dependent on NADPH and inhibited by 0.1 mM SKF-525A, an inhibitor of P450 enzymes. The activity to form 12(R)-enantiomer was significantly enhanced by treatment of corneal epithelium with 3-methylcholanthrene or clofibrate. The induced activity was suppressed by cycloheximide, indicating that the induction of enzyme activities involved a translational process. The effect of these inducers on 12(R)-HETE synthesizing activity appeared to be additive. The activity to form 12(S)-enantiomer was markedly stimulated by 3 mM CaCl2. The 12-lipoxygenase of corneal microsomes was capable of oxygenating linoleic acid in addition to arachidonic acid, a characteristic of 12-lipoxygenases of the leukocyte type. 12(R)-HETE at 10(-6) M inhibited almost completely the Na,K-ATPase of corneal epithelium but had little or no effect on ciliary epithelial enzymic activity. 12(S)-HETE at 10(-6) M also inhibited corneal enzymic activity but to a lesser extent, and had no significant effect on ciliary epithelial Na,K-ATPase activity.
Department of Biochemistry, Jikei University School of Medicine, Tokyo, Japan.
T
T
Asakura
M
M
Matsuda
S
S
Matsuda
H
H
Shichi
D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D000818:Animals; D000869:Anterior Eye Segment / Q000378:metabolism; D001092:Arachidonate 12-Lipoxygenase / Q000378:metabolism; D016718:Arachidonic Acid / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D005123:Eye / Q000378:metabolism*; D006893:Hydroxyeicosatetraenoic Acids / Q000096:biosynthesis* / Q000494:pharmacology; D008861:Microsomes / Q000378:metabolism; D012160:Retina / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism; D013237:Stereoisomerism; D013552:Swine
D006893:Hydroxyeicosatetraenoic Acids; D016718:Arachidonic Acid; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D003577:Cytochrome P-450 Enzyme System; D001092:Arachidonate 12-Lipoxygenase; D000254:Sodium-Potassium-Exchanging ATPase