Characterization and inhibition of 15-lipoxygenase in human monocytes: Comparison with soybean 15-lipoxygenase

M. M. Gleason, Camilo Rojas, K. S. Learn, M. H. Perrone, G. E. Bilder

Research output: Contribution to journalArticle

Abstract

These studies characterize a method to measure 15-lipoxygenase (15-LO) activity in human monocytes (HMC) exposed to interleukin-4 (IL-4) and compare the activity with that of soybean 15-LO. 15-LO activity was quantitated by measuring 15-[14C]hydroxyeicosa-5Z,8Z,11Z,13E-tetraenoic acid (15-HETE) production from the substrate [14C]arachidonic acid (AA) after high- performance liquid chromatographic or thin-layer chromatographic separation. 15-HETE production by HMC was significantly elevated after continuous exposure to a single dose (10 ng/ml) of IL-4 for 4 days, was maximal at 5 days, and remained elevated at 6 days. At 6 days 15-LO activity in IL-4- treated HMC was increased significantly (2.81 ± 0.70 fmol 15-HETE/cell) compared with background levels of 15-HETE in untreated HMC (0.098 ± 0.31 fmol 15-HETE/cell). 15-HETE production was linear in the range of 5 x 104 to 7 x 105 HMC/assay, from 2 to 160 μM AA, and during 5-10 min of incubation with AA. Ethyl 2-cyano-3-(3',4'-dihydroxyphenyl)propenoate (a caffeic acid analogue), N-methyl-4-benzyloxyphenyl acetohydroxamate (RG-6866), esculetin, and four novel lipoxygenase inhibitors, a phenylcyanomethylene analogue (RP- 27493) and three benzoxadiazine analogues (RP-64835, RP-65047, and RP- 65208), inhibited HMC 15-LO, with concentrations eliciting 50% of maximal inhibition (μM) of 0.21, 0.8, 6.3, 10, 22, 20, 6.3, 3, and 20 and 5.8, 5, 12, 1, 30, 0.8, 0.15, 0.05, and 2.8 for inhibition of soybean 15-LO, respectively. These data define conditions for optimal production of 15-HETE by IL-4-treated HMC. Known and novel lipoxygenase inhibitors inhibit IL-4- treated HMC 15-LO and soybean 15-LO, suggesting that IL-4-treated HMC provide an appropriate system to study human 15-LO activity.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Cell Physiology
Volume268
Issue number5 37-5
StatePublished - 1995
Externally publishedYes

Fingerprint

Arachidonate 15-Lipoxygenase
lipoxygenase
Soybeans
monocytes
Monocytes
soybeans
Interleukin-4
interleukin-4
Arachidonic Acid
Lipoxygenase Inhibitors
arachidonic acid
15-hydroxy-5,8,11,13-eicosatetraenoic acid
Human Activities
caffeic acid
Assays

Keywords

  • 15-HETE
  • 15-lipoxygenase
  • 5,8,11,14- eicosatetraynoic acid
  • human monocytes
  • interleukin-4
  • nordihydroguaiaretic acid

ASJC Scopus subject areas

  • Cell Biology
  • Clinical Biochemistry
  • Physiology
  • Agricultural and Biological Sciences(all)

Cite this

Characterization and inhibition of 15-lipoxygenase in human monocytes : Comparison with soybean 15-lipoxygenase. / Gleason, M. M.; Rojas, Camilo; Learn, K. S.; Perrone, M. H.; Bilder, G. E.

In: American Journal of Physiology - Cell Physiology, Vol. 268, No. 5 37-5, 1995.

Research output: Contribution to journalArticle

@article{20deee49efa24f82ac7a9b789ac0091c,
title = "Characterization and inhibition of 15-lipoxygenase in human monocytes: Comparison with soybean 15-lipoxygenase",
abstract = "These studies characterize a method to measure 15-lipoxygenase (15-LO) activity in human monocytes (HMC) exposed to interleukin-4 (IL-4) and compare the activity with that of soybean 15-LO. 15-LO activity was quantitated by measuring 15-[14C]hydroxyeicosa-5Z,8Z,11Z,13E-tetraenoic acid (15-HETE) production from the substrate [14C]arachidonic acid (AA) after high- performance liquid chromatographic or thin-layer chromatographic separation. 15-HETE production by HMC was significantly elevated after continuous exposure to a single dose (10 ng/ml) of IL-4 for 4 days, was maximal at 5 days, and remained elevated at 6 days. At 6 days 15-LO activity in IL-4- treated HMC was increased significantly (2.81 ± 0.70 fmol 15-HETE/cell) compared with background levels of 15-HETE in untreated HMC (0.098 ± 0.31 fmol 15-HETE/cell). 15-HETE production was linear in the range of 5 x 104 to 7 x 105 HMC/assay, from 2 to 160 μM AA, and during 5-10 min of incubation with AA. Ethyl 2-cyano-3-(3',4'-dihydroxyphenyl)propenoate (a caffeic acid analogue), N-methyl-4-benzyloxyphenyl acetohydroxamate (RG-6866), esculetin, and four novel lipoxygenase inhibitors, a phenylcyanomethylene analogue (RP- 27493) and three benzoxadiazine analogues (RP-64835, RP-65047, and RP- 65208), inhibited HMC 15-LO, with concentrations eliciting 50{\%} of maximal inhibition (μM) of 0.21, 0.8, 6.3, 10, 22, 20, 6.3, 3, and 20 and 5.8, 5, 12, 1, 30, 0.8, 0.15, 0.05, and 2.8 for inhibition of soybean 15-LO, respectively. These data define conditions for optimal production of 15-HETE by IL-4-treated HMC. Known and novel lipoxygenase inhibitors inhibit IL-4- treated HMC 15-LO and soybean 15-LO, suggesting that IL-4-treated HMC provide an appropriate system to study human 15-LO activity.",
keywords = "15-HETE, 15-lipoxygenase, 5,8,11,14- eicosatetraynoic acid, human monocytes, interleukin-4, nordihydroguaiaretic acid",
author = "Gleason, {M. M.} and Camilo Rojas and Learn, {K. S.} and Perrone, {M. H.} and Bilder, {G. E.}",
year = "1995",
language = "English (US)",
volume = "268",
journal = "American Journal of Physiology",
issn = "0363-6135",
publisher = "American Physiological Society",
number = "5 37-5",

}

TY - JOUR

T1 - Characterization and inhibition of 15-lipoxygenase in human monocytes

T2 - Comparison with soybean 15-lipoxygenase

AU - Gleason, M. M.

AU - Rojas, Camilo

AU - Learn, K. S.

AU - Perrone, M. H.

AU - Bilder, G. E.

PY - 1995

Y1 - 1995

N2 - These studies characterize a method to measure 15-lipoxygenase (15-LO) activity in human monocytes (HMC) exposed to interleukin-4 (IL-4) and compare the activity with that of soybean 15-LO. 15-LO activity was quantitated by measuring 15-[14C]hydroxyeicosa-5Z,8Z,11Z,13E-tetraenoic acid (15-HETE) production from the substrate [14C]arachidonic acid (AA) after high- performance liquid chromatographic or thin-layer chromatographic separation. 15-HETE production by HMC was significantly elevated after continuous exposure to a single dose (10 ng/ml) of IL-4 for 4 days, was maximal at 5 days, and remained elevated at 6 days. At 6 days 15-LO activity in IL-4- treated HMC was increased significantly (2.81 ± 0.70 fmol 15-HETE/cell) compared with background levels of 15-HETE in untreated HMC (0.098 ± 0.31 fmol 15-HETE/cell). 15-HETE production was linear in the range of 5 x 104 to 7 x 105 HMC/assay, from 2 to 160 μM AA, and during 5-10 min of incubation with AA. Ethyl 2-cyano-3-(3',4'-dihydroxyphenyl)propenoate (a caffeic acid analogue), N-methyl-4-benzyloxyphenyl acetohydroxamate (RG-6866), esculetin, and four novel lipoxygenase inhibitors, a phenylcyanomethylene analogue (RP- 27493) and three benzoxadiazine analogues (RP-64835, RP-65047, and RP- 65208), inhibited HMC 15-LO, with concentrations eliciting 50% of maximal inhibition (μM) of 0.21, 0.8, 6.3, 10, 22, 20, 6.3, 3, and 20 and 5.8, 5, 12, 1, 30, 0.8, 0.15, 0.05, and 2.8 for inhibition of soybean 15-LO, respectively. These data define conditions for optimal production of 15-HETE by IL-4-treated HMC. Known and novel lipoxygenase inhibitors inhibit IL-4- treated HMC 15-LO and soybean 15-LO, suggesting that IL-4-treated HMC provide an appropriate system to study human 15-LO activity.

AB - These studies characterize a method to measure 15-lipoxygenase (15-LO) activity in human monocytes (HMC) exposed to interleukin-4 (IL-4) and compare the activity with that of soybean 15-LO. 15-LO activity was quantitated by measuring 15-[14C]hydroxyeicosa-5Z,8Z,11Z,13E-tetraenoic acid (15-HETE) production from the substrate [14C]arachidonic acid (AA) after high- performance liquid chromatographic or thin-layer chromatographic separation. 15-HETE production by HMC was significantly elevated after continuous exposure to a single dose (10 ng/ml) of IL-4 for 4 days, was maximal at 5 days, and remained elevated at 6 days. At 6 days 15-LO activity in IL-4- treated HMC was increased significantly (2.81 ± 0.70 fmol 15-HETE/cell) compared with background levels of 15-HETE in untreated HMC (0.098 ± 0.31 fmol 15-HETE/cell). 15-HETE production was linear in the range of 5 x 104 to 7 x 105 HMC/assay, from 2 to 160 μM AA, and during 5-10 min of incubation with AA. Ethyl 2-cyano-3-(3',4'-dihydroxyphenyl)propenoate (a caffeic acid analogue), N-methyl-4-benzyloxyphenyl acetohydroxamate (RG-6866), esculetin, and four novel lipoxygenase inhibitors, a phenylcyanomethylene analogue (RP- 27493) and three benzoxadiazine analogues (RP-64835, RP-65047, and RP- 65208), inhibited HMC 15-LO, with concentrations eliciting 50% of maximal inhibition (μM) of 0.21, 0.8, 6.3, 10, 22, 20, 6.3, 3, and 20 and 5.8, 5, 12, 1, 30, 0.8, 0.15, 0.05, and 2.8 for inhibition of soybean 15-LO, respectively. These data define conditions for optimal production of 15-HETE by IL-4-treated HMC. Known and novel lipoxygenase inhibitors inhibit IL-4- treated HMC 15-LO and soybean 15-LO, suggesting that IL-4-treated HMC provide an appropriate system to study human 15-LO activity.

KW - 15-HETE

KW - 15-lipoxygenase

KW - 5,8,11,14- eicosatetraynoic acid

KW - human monocytes

KW - interleukin-4

KW - nordihydroguaiaretic acid

UR - http://www.scopus.com/inward/record.url?scp=0029000373&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0029000373&partnerID=8YFLogxK

M3 - Article

C2 - 7762624

AN - SCOPUS:0029000373

VL - 268

JO - American Journal of Physiology

JF - American Journal of Physiology

SN - 0363-6135

IS - 5 37-5

ER -