Selective expression of an arachidonate 12-lipoxygenase by pancreatic islet beta-cells.
The immunohistochemical distribution of arachidonate lipoxygenases in rat pancreas was characterized with specific polyclonal anti-5-lipoxygenase and anti-12-lipoxygenase antibodies. Immunohistochemical analysis of formaldehyde-fixed paraffin-embedded rat pancreas using anti-12-lipoxygenase antibody and biotin-avidin-peroxidase detection demonstrated specific staining of islets and no staining of pancreatic exocrine tissue. Less intense staining of pancreatic vascular myocytes and endothelial cells was also observed. Immunoblotting of isolated pancreatic islet extracts with the anti-12-lipoxygenase antibody demonstrated immunoperoxidase staining of a single protein band which comigrated with purified 12-lipoxygenase (relative molecular weight = 72,000) on sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. Dispersed cells prepared from isolated islets and then subjected to fluorescence-activated cell sorting and immunostaining exhibited 12-lipoxygenase antigen in beta-cell populations but not in non-beta-cell (predominantly alpha-cell) populations. Assays of enzymatic activity confirmed that the 12-lipoxygenase-catalyzed conversion of arachidonic acid to 12-hydroxyeicosatetraenoic acid methyl ester occurred only with purified beta-cells and not with islet non-beta-cells. No evidence of 5-lipoxygenase antigen or enzymatic activity was found in purified beta-cells or in islet non-beta-cells. We conclude that rat pancreatic islet beta-cells contain an arachidonate 12-lipoxygenase which shares antigenic epitopes with the homologous enzyme contained in tissues from other species. In addition, the selective localization of the 12-lipoxygenase to pancreatic beta-cells and its absence in pancreatic acinar cells and in islet non-beta-cells support observations suggesting that 12-lipoxygenase products may participate in glucose-induced insulin secretion from beta-cells.
Shannon
V R
VR
Department of Medicine, Washington University School of Medicine, St. Louis, Missouri 63110.
Ramanadham
S
S
Turk
J
J
Holtzman
M J
MJ
D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D000818:Animals; D001092:Arachidonate 12-Lipoxygenase / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D006893:Hydroxyeicosatetraenoic Acids / Q000378:metabolism; D015151:Immunoblotting; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D007515:Islets of Langerhans / Q000166:cytology / Q000201:enzymology*; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley
D006893:Hydroxyeicosatetraenoic Acids; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D001092:Arachidonate 12-Lipoxygenase