ARACHIDONATE 15-LIPOXYGENASE IN HUMAN CORNEAL EPITHELIUM AND 12-LIPOXYGENASE AND 15-LIPOXYGENASE IN BOVINE CORNEAL EPITHELIUM - COMPARISON WITH OTHER BOVINE 12-LIPOXYGENASES
M. Liminga et al., ARACHIDONATE 15-LIPOXYGENASE IN HUMAN CORNEAL EPITHELIUM AND 12-LIPOXYGENASE AND 15-LIPOXYGENASE IN BOVINE CORNEAL EPITHELIUM - COMPARISON WITH OTHER BOVINE 12-LIPOXYGENASES, Biochimica et biophysica acta, L. Lipids and lipid metabolism, 1210(3), 1994, pp. 288-296
Lipoxygenases of bovine and human corneal epithelia were investigated.
The bovine epithelium contained an arachidonate 12-lipoxygenase and a
15-lipoxygenase. The 12-lipoxygenase was found in the microsomal frac
tion, while the 15-lipoxygenase was mainly present in the cytosol (100
000 x g supernatant). 12S-Hydroxyeicosatetraenoic acid (12S-HETE) and
15S-hydroxyeicosatetraenoic acid (15S-HETE) were identified by GC-MS a
nd chiral HPLC. BW A4C, an acetohydroxamic acid lipoxygenase inhibitor
, reduced the biosynthesis of 12S-HETE and 15S-HETE by over 90% at 10
mu M. IC50 for the 12-lipoxygenase was 0.3 mu M. The bovine corneal 12
-lipoxygenase was compared with the 12-lipoxygenases of bovine platele
ts and leukocytes. All three enzymes metabolized C-14-labelled linorei
c acid and alpha-linolenic acid poorly (5-16%) in comparison with [C-1
4]arachidonic acid. [C-14]Docosahexaenoic acid and [C-14]4,7,10,13,16-
docosapentaenoic acid appeared to be less efficiently converted by the
corneal enzyme than by the platelet and leukocyte enzymes. Immunohist
ochemical analysis of the bovine corneal epithelium using a polyconal
antibody against porcine leukocyte 12-lipoxygenase gave positive stain
ing. The cytosol of human corneal epithelium converted [C-14]arachidon
ic acid to one prominent metabolite. The product co-chromatographed wi
th 15S-HETE on reverse phase HPLC, straight phase HPLC and chiral HPLC
. Our results suggest that human corneal epithelium contains a 15-lipo
xygenase and that bovine corneal epithelium contains both a 15-lipoxyg
enase and a 12-lipoxygenase. The corneal 12-lipoxygenase appears to di
ffer catalytically from earlier described bovine 12-lipoxygenases.