TY - JOUR
T1 - Metabolism of 12(R)-hydroxy-5,8,10,14-eicosatetraenoic acid (12(R)-HETE) in corneal tissues
T2 - Formation of novel metabolites
AU - Nishimura, Motonobu
AU - Schwartzman, Michal Laniado
AU - Falck, J R
AU - Lumin, Sun
AU - Zirrolli, Joseph A.
AU - Murphy, Robert C.
N1 - Funding Information:
i The present study was supported in part by NIH Grants EY 06513, HL34303, GM41206, and GM3178; the Mathers Foundation; and a postdoctoral research fellowship from the Fight for Sight Research Division of the National Society to Prevent Blindness. * To whom correspondence should be addressed. 3 Abbreviations used: PBS, phosphate-buffered saline; DMEM, becco’s modified Eagle’s medium; PFB, pentafluorobenzyh GC-MS, chromatography-mass spectrometry; LC-MS, liquid chromatography-mass spectrometry; CF-FAB, continuous flow fast atom bombardmenti TMS, trimethylsilane; ODS, octadecasilane; EI, electron impact; ECL, equivalent carbon length; BSTFA, N,O-his(trimethylsilyl)trifluoroacetamide; RP, reverse phase. The abbreviations used for the various eicosanoids are as proposed by the ad hoc Committee on Eicosanoid Nomenclature (Methods in Enzymology (Murphy, R. C., and Fitzpatrick, F. A., Eds.), Vol. 187, pp. l-9, Academic Press, San Diego) and are as follows: 12.HETE, 12.hydroxy-5,8,10,14-eicosatetraenoic acid; 12-HETrE, 12-hydroxy-5,8,14-eicosatrienoic acid; 12.oxo-ETrE, 12-0x0-5,8,14-eicosatrienoic acid; 12,20diHETE, 12,20-dihydroxy-5,8,10,14-eicosatetraenoic acid; 8-HHxTrE, 8-hydroxy-4,6,10-(Z,E,Z)-hexadecatrienoic acid, LTBl , leukotriene B,
PY - 1991/11/1
Y1 - 1991/11/1
N2 - 12(R)-Hydroxy-5,8,10,14-eicosatetraenoic acid [12(R)-HETE], a cytochrome P450 arachidonate metabolite, is metabolized by corneal tissues via three distinct metabolic pathways: β-oxidation, ω-hydroxylation, and keto-reduction. The major metabolite released from the intact rabbit corneal epithelium or cultured cells was identified by mass spectrometric analysis as 8-hydroxy-4,6,10-hexadecatrienoic acid, the tetranor metabolite derived following two steps of β-oxidation from the carboxy terminus. The β-oxidation pathway was expressed in both microsomes and mitochondria isolated from bovine corneal epithelium and was dependent on the addition of oxidizing equivalents. The major metabolite of 12(R)-HETE in subcellular fractions of bovine corneal epithelial cells was a dihydro compound, 12-hydroxy-5,8,14-eicosatrienoic acid (12-HETrE). This derivative is presumably formed by an oxidation of the hydroxyl group followed by two keto-reduction steps, since its formation was accompanied by the appearance of a keto metabolite identified as 12-oxo-5,8,14-eicosatrienoic acid. The ω-hydroxylation, in contrast to other cell types, was a minor route for 12(R)-HETE metabolism in these tissues. Since 12(R)-HETE has been implicated as a modulator of Na+-K+-ATPase activity and its related functions in ocular tissues, these findings raise the possibility that the newly described metabolites may be involved in regulating corneal functions. In addition, the presence of a keto reductase in the cornea may be of great importance following injury since 12(R)-HETrE resulting from 12(R)-HETE by this activity is a potent ocular proinflammatory compound.
AB - 12(R)-Hydroxy-5,8,10,14-eicosatetraenoic acid [12(R)-HETE], a cytochrome P450 arachidonate metabolite, is metabolized by corneal tissues via three distinct metabolic pathways: β-oxidation, ω-hydroxylation, and keto-reduction. The major metabolite released from the intact rabbit corneal epithelium or cultured cells was identified by mass spectrometric analysis as 8-hydroxy-4,6,10-hexadecatrienoic acid, the tetranor metabolite derived following two steps of β-oxidation from the carboxy terminus. The β-oxidation pathway was expressed in both microsomes and mitochondria isolated from bovine corneal epithelium and was dependent on the addition of oxidizing equivalents. The major metabolite of 12(R)-HETE in subcellular fractions of bovine corneal epithelial cells was a dihydro compound, 12-hydroxy-5,8,14-eicosatrienoic acid (12-HETrE). This derivative is presumably formed by an oxidation of the hydroxyl group followed by two keto-reduction steps, since its formation was accompanied by the appearance of a keto metabolite identified as 12-oxo-5,8,14-eicosatrienoic acid. The ω-hydroxylation, in contrast to other cell types, was a minor route for 12(R)-HETE metabolism in these tissues. Since 12(R)-HETE has been implicated as a modulator of Na+-K+-ATPase activity and its related functions in ocular tissues, these findings raise the possibility that the newly described metabolites may be involved in regulating corneal functions. In addition, the presence of a keto reductase in the cornea may be of great importance following injury since 12(R)-HETrE resulting from 12(R)-HETE by this activity is a potent ocular proinflammatory compound.
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U2 - 10.1016/0003-9861(91)90548-W
DO - 10.1016/0003-9861(91)90548-W
M3 - Article
C2 - 1929401
AN - SCOPUS:0025946705
SN - 0003-9861
VL - 290
SP - 326
EP - 335
JO - Archives of Biochemistry and Biophysics
JF - Archives of Biochemistry and Biophysics
IS - 2
ER -