Role of Lysyl Oxidase in the Mechanoregulation of Retinal Vascular Inflammation and Degeneration in diabetes

Abstract

Purpose : Vascular inflammation causes degeneration of retinal capillaries in early diabetic retinopathy (DR). Based on past reports that vascular inflammation in non-ocular conditions such as atherosclerosis can be mechanically regulated by lysyl oxidase (LOX), my lab is investigating whether LOX can mechanically regulate retinal vascular inflammation in early DR independent of the molecular and genetic DR stressors.

Methods : Retinal vessels were isolated from nondiabetic, diabetic (D), and D mice treated with LOX inhibitor β-aminopropionitrile (BAPN; 10 μg/kg in drinking water, 10 wks) (D+BAPN) for stiffness analysis using an atomic force microscope or RT-qPCR analysis of proinflammatory ICAM-1 and RAGE. Intravenous FITC-Con A injection and PAS staining were done to assess leukostasis and retinal capillary loss. Human retinal endothelial ECs (HRECs) grown on normal or stiff synthetic matrices were assessed for ICAM-1/RAGE mRNA expression, neutrophil-EC adhesion. Finally, neutrophils were treated with LOX (75 ng/ml; 6h) and assessed for F-actin dynamics (phalloidin staining), release of superoxide (DHE dye) and neutrophil elastase (EnzCheck Kit), and membrane assembly of NADPH-associated p47 and elastase-associated CD63 (immunolabeling).

Results : We show that retinal vessels become stiffer in diabetes, which is caused by LOX overexpression in retinal ECs. Treatment of ‘D’ mice with LOX inhibitor BAPN prevents the increase in retinal capillary stiffness, vascular ICAM-1/RAGE overexpression, leukostasis, and retinal capillary atrophy. Importantly, HRECs grown on synthetic matrices indicate that retinal capillary stiffening alone can promote retinal vascular inflammation and degeneration. Finally, LOX was found to activate neutrophils, as judged by increased superoxide and neutrophil elastase release and greater cytotoxicity towards retinal ECs, which are all blocked by preventing the LOX-induced F-actin depolymerization in neutrophils.

Conclusions : By uncovering a link between diabetes, LOX-dependent capillary stiffening and neutrophil actin regulation, and retinal vascular degeneration, these findings offer a new insight into DR pathogenesis that has important translational potential.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.