Cytochrome P450 arachidonic acid omega-hydroxylation in the proximal tubule of the rat kidney.
20-Hydroxy-5,8,11,14-eicosatetraenoic acid (20-HETE) is a major cytochrome P450-dependent arachidonate metabolite in the rat kidney. In the present study we characterized the formation of 20-HETE in the proximal tubule, the nephron segment with the highest concentration of cytochrome P450 activities, including P450 arachidonic acid metabolism. Freshly isolated tubules showed a basal formation of 20-HETE, implying that it is an endogenous constituent of the proximal tubule. Conversion of exogenous arachidonic acid to 20-HETE in proximal tubule homogenates was enzymatic and NADPH-dependent (i.e., 0 and 65.5 +/- 1.1 pmol/mg/min in the absence and presence of NADPH, respectively). That its formation was not affected by indomethacin but inhibited following preincubation with 17-ODYA (17-octadecynoic acid) and 7-ER (7-ethoxyresorufin) suggested that a P450 monooxygenase activity was involved in its synthesis. This was further strengthened by the demonstration that antibody raised against the rat cytochrome P450 4A1, a major fatty acid omega-hydroxylase isozyme, inhibited 20-HETE formation, suggesting the involvement of a P450 4A1 or P450 4A1-like activity in this reaction. Pretreatment of rats with clofibrate and dexamethasone, inducers of the P450 4A gene family, yielded a twofold increase in the proximal tubular synthesis of 20-HETE as well as an increase in P450 4A1 mRNA. These results, together with previous demonstrations that 20-HETE vasoconstricts isolated blood vessels, namely, renal microvessels, and affects tubular ion transport, suggest a role for 20-HETE in the regulation of renal vascular tone and transport functions and further stress the importance of understanding the regulation of 20-HETE synthesis in the kidney.
Lin
F
F
Department of Pharmacology, New York Medical College, Valhalla 10595.
Abraham
N G
NG
Schwartzman
M L
ML
D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism*; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006893:Hydroxyeicosatetraenoic Acids / Q000096:biosynthesis; D006900:Hydroxylation; D007687:Kidney Tubules, Proximal / Q000378:metabolism*; D008297:Male; D012313:RNA / Q000302:isolation & purification; D051381:Rats; D017207:Rats, Sprague-Dawley
D006893:Hydroxyeicosatetraenoic Acids; D016718:Arachidonic Acid; D012313:RNA; D003577:Cytochrome P-450 Enzyme System