Differential Activity of Systemic and Retinal 12/15-Lipoxygenases in a Mouse Model of Diabetes

Abstract

Purpose : The 12/15-lipoxygenase (12/15-LOX) has proinflammatory effects such as enhanced chemotaxis and vascular adhesion of leukocytes, which has been implicated in the pathogenesis of diabetic retinopathy (DR). Our previous studies have shown increased retinal 12/15-LOX expression and activity in the vitreous of diabetic patients with retinopathy, diabetic mouse retinas, and human retinal endothelial cells treated with high glucose. In the current study we aimed to evaluate the change in circulating 12/15-LOX activity in a type 1 diabetic mouse model and to characterize the contribution of endothelial 12/15-LOX versus circulating 12/15-LOX to leukocyte adhesion.

Methods : A lipidomic approach using liquid chromatography coupled with mass-spectrometry (LC-MS) was used to investigate the activity of circulating 12/15-LOX on linoleic acid (LA), arachidonic acid (AA), docosahexaenoic acid (DHA), and eicosapentaenoic acid (EPA) in the plasma of streptozotocin-induced diabetic mice (6-months of diabetes). Mouse retinal endothelial cells (mREC) as well as leukocytes isolated from 12/15-LOX knockout (KO) or wild type (WT) mouse were used to study leukocyte adhesion using the myeloperoxidase assay.

Results : Out of 29 bioactive lipids screened, only 3 metabolites showed significant changes in diabetic mouse plasma compared to normal controls. The increased metabolites included two products derived from the effect of 12/15-LOX on DHA (Resolvin D2 and 14-HDoHE) and one product derived from the lipoxygenation of LA (13-OxoODE). In contrast, there was no significant change in the plasma level of metabolites derived from 12/15-lipoxygenation of AA or EPA including 12- and 15-HETE or 12- and 15-HEPE, respectively. These data presumably reflect a differential role of circulating 12/15-LOX versus endothelial 12/15-LOX in mediating leukocyte adhesion. To this end, we performed in vitro leukocyte adhesion assay on LPS-activated mREC using leukocytes isolated from 12/15-LOX KO versus WT mice. Activated mREC significantly augmented the number of adherent leukocytes isolated from 12/15-LOX KO relatively to the same extent as those derived from WT mice.

Conclusions : Our current and previously studies suggest a differential role of endothelial 12/15-LOX versus the one in circulating blood cells in mediating the inflammatory responses during DR. This may facilitate the development of more precisely targeted treatment strategies.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.