TY - JOUR
T1 - Metabolism of 12(S)-Hydroxy-5,8,10,14-eicosatetraenoic Acid and Other Hydroxylated Fatty Acids by the Reductase Pathway in Porcine Polymorphonuclear Leukocytes
AU - Wainwright, Sandra
AU - Falck, J. R.
AU - Yadagiri, Pendri
AU - Powell, William S.
N1 - Copyright:
Copyright 2017 Elsevier B.V., All rights reserved.
PY - 1990/10/1
Y1 - 1990/10/1
N2 - We have previously shown that porcine polymorphonuclear leukocytes (PMNL) reduce leukotriene B4 (LTB4) to 10,11-dihydro-LTB4, 10,11-dihydro-12-epi-LTB4, and 10,11-dihydro-12-oxo-LTB4 [Wainwright et al. (1990) Biochemistry 29, 1180-1185], We have now demonstrated that12(S)-hydroxy-5,8.10.14-eicosatetraenoic acid [12(S)-HETE] is metabolized by a similar pathway in porcine PMNL. 12(S)-HETE was metabolized to two products that were identified by gas chromatography-mass spectrometry and nuclear magnetic resonance spectroscopy as 12-hydroxy-5,8,14-eicosatrienoic acid (10,11-dihydro-12-HETE) and 12-oxo-5,8,14-eicosatrienoic acid (10, 11-dihydro-12-oxo-ETE). Derivatization of 12-hydroxy-5, 8, 14-eicosatrienoic acid with (R)-(+)-α-methoxy-α-(trifluoromethyl)phenylacetic acid, followed by chromatography on a silicic acid column, enabled the resolution of 12R and 12S stereoisomers, which were identified by cochromatography with synthetic standards. Incubation of 12(S)-HETE with PMNL for various times revealed that the stereochemistry of the 12-hydroxyl group of 12-hydroxy-5,8,14-eicosatrienoic acid was initially the same as that of 12(S)-HETE. However, after 40 min, 30% of the 12-hydroxy-5,8,14-eicosatrienoic acid had the opposite configuration at C12. 13-Hydroxy-9, 11-octadecadienoic acid (13-HODE) was metabolized in a similar fashion by porcine PMNL to 13-hydroxy-9-octadecenoic acid (11,12-dihydro-13-HODE) and 13-oxooctadecenoic acid (11,12-dihydro-13-oxo-ODE). The apparent Km values for the reduction of 12-HETE, LTB4, and 13-HODE were 0.21, 0.28, and 2.22 μM, respectively. All three substrates had the same apparent Vmax [0.029 pmol min−1 (106 cells)−1]. Competition experiments between LTB4 and 12-HETE indicated that they were metabolized by the same pathway. Various structurally related compounds were metabolized by porcine PMNL in the order LTB4 = 6-trans-LTB4 > 12-epi-6-trans,8-cis-LTB4 > 12-epi-6-trans,8-cis-LTB4 > 12-HETE > LTB5 > 15-HETE = 13-HODE ≫ 5-HETE > 9-HODE > 20-hydroxy-LTB4 > 12-hydroxy-5,8,10-heptadecatrienoic acid. Prostaglandins E2 and F2α were not metabolized to any detectable products by porcine PMNL.
AB - We have previously shown that porcine polymorphonuclear leukocytes (PMNL) reduce leukotriene B4 (LTB4) to 10,11-dihydro-LTB4, 10,11-dihydro-12-epi-LTB4, and 10,11-dihydro-12-oxo-LTB4 [Wainwright et al. (1990) Biochemistry 29, 1180-1185], We have now demonstrated that12(S)-hydroxy-5,8.10.14-eicosatetraenoic acid [12(S)-HETE] is metabolized by a similar pathway in porcine PMNL. 12(S)-HETE was metabolized to two products that were identified by gas chromatography-mass spectrometry and nuclear magnetic resonance spectroscopy as 12-hydroxy-5,8,14-eicosatrienoic acid (10,11-dihydro-12-HETE) and 12-oxo-5,8,14-eicosatrienoic acid (10, 11-dihydro-12-oxo-ETE). Derivatization of 12-hydroxy-5, 8, 14-eicosatrienoic acid with (R)-(+)-α-methoxy-α-(trifluoromethyl)phenylacetic acid, followed by chromatography on a silicic acid column, enabled the resolution of 12R and 12S stereoisomers, which were identified by cochromatography with synthetic standards. Incubation of 12(S)-HETE with PMNL for various times revealed that the stereochemistry of the 12-hydroxyl group of 12-hydroxy-5,8,14-eicosatrienoic acid was initially the same as that of 12(S)-HETE. However, after 40 min, 30% of the 12-hydroxy-5,8,14-eicosatrienoic acid had the opposite configuration at C12. 13-Hydroxy-9, 11-octadecadienoic acid (13-HODE) was metabolized in a similar fashion by porcine PMNL to 13-hydroxy-9-octadecenoic acid (11,12-dihydro-13-HODE) and 13-oxooctadecenoic acid (11,12-dihydro-13-oxo-ODE). The apparent Km values for the reduction of 12-HETE, LTB4, and 13-HODE were 0.21, 0.28, and 2.22 μM, respectively. All three substrates had the same apparent Vmax [0.029 pmol min−1 (106 cells)−1]. Competition experiments between LTB4 and 12-HETE indicated that they were metabolized by the same pathway. Various structurally related compounds were metabolized by porcine PMNL in the order LTB4 = 6-trans-LTB4 > 12-epi-6-trans,8-cis-LTB4 > 12-epi-6-trans,8-cis-LTB4 > 12-HETE > LTB5 > 15-HETE = 13-HODE ≫ 5-HETE > 9-HODE > 20-hydroxy-LTB4 > 12-hydroxy-5,8,10-heptadecatrienoic acid. Prostaglandins E2 and F2α were not metabolized to any detectable products by porcine PMNL.
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U2 - 10.1021/bi00495a017
DO - 10.1021/bi00495a017
M3 - Article
C2 - 2176862
AN - SCOPUS:0025043481
SN - 0006-2960
VL - 29
SP - 10126
EP - 10135
JO - Biochemistry
JF - Biochemistry
IS - 43
ER -