Cytochrome p450-derived renal hetes: biological profiles and
differential stimulation by vasoactive peptides.
Carroll, Mairead A., Michael Balazy, Patricia Margiotta, Dong-Dong
Huang, J. R. Falck, and John C. McGiff.
Departments of Pharmacology, New York Medical College, (M.A.C.,
M.B., P.M., D-D.H., J.C.M.) and Department of Molecular Genetics,
University of Texas Health Science Center, Dallas, (J.R.F.)
APStracts 3:0144R, 1996.
The cytochrome P450 pathway is capable of metabolizing arachidonic
acid (AA) to w- and subterminal hydroxylase metabolites, 16-, 17-,
18-, 19- and 20-hydroxyeicosatetraenoic acids (P450-HETEs). We have
quantitated, by gas chromatography-mass spectrometry (GC/MS),
endogenous HETEs exiting the rabbit isolated perfused kidney elicited
by hormonal stimulation. Kidneys were perfused with Krebs'-Henseleit
solution, containing indomethacin (2.8 [mu]M), to prevent further
metabolism of HETEs by cyclooxygenase. Phenylephrine (2-3[mu]M) was
added to the perfusate to raise perfusion pressure to ca 80mmHg .
Angiotensin II (AII), arginine vasopressin (AVP) and bradykinin (BK)
were injected into the renal artery and perfusates collected
throughout the vasoactive response. After addition of an internal
standard, deuterated 19-HETE, perfusates were extracted, purified,
and P450-HETEs derivatized for GC/MS analysis. Under basal conditions
16-, 18-, 19- and 20-HETEs were released (range 50 to 270 pg/ml), 19
-HETE being the highest and five-fold greater than 16-HETE, the
lowest. Injection of 50 ng AII increased by 2- to 6-fold P450-HETE
release associated with an increase of 40+11 mmHg in perfusion
pressure. An equipressor dose of AVP (50 ng) did not release P450
-HETEs nor did a 5[mu]g dose of the vasodilator peptide BK which
decreased perfusion pressure by 22+6 mmHg. Authentic 19- and 20-HETE
isomers resulted in dose-dependent dilation, as did 18(R)- and 16(R)
-HETEs, whereas their enantiomers and 17-HETE isomers were without
effect on perfusion pressure. The vasodilator effects of 18(R)- and
16(R)-HETEs, like 20- and 19-HETEs, were inhibited by indomethacin.
Further, P450-HETEs exhibited both regio- and stereoselective
inhibition of proximal tubule ATPase activity. The (S) enantiomers of
16- and 17-HETE potently inhibited activity, whereas their (R)
isomers and other P450-HETEs had negligible effects on ATPase
activity. The quantity of HETEs released from the kidney either under
basal conditions or when stimulated by AII, and their biolgical
profile suggest that subterminal HETEs may participate in renal
mechanisms affecting vasomotion and tubular transport.
Received 15 February 1996; accepted in final form 25 March 1996.
APS Manuscript Number R97-6.
Article publication pending Am. J. Physiol. (Regulatory Integrative
Comp. Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 23 April 96