Systemic Administration of Pegylated Arginase-1 Attenuates Progression of Diabetic Retinopathy in the db/db Mouse

Abstract

Background Diabetic retinopathy (DR) is a serious diabetes complication that affects millions worldwide. Current therapies are invasive and only slow late-stage progression, thus the development of less invasive, novel therapies that attenuate progression are paramount. In the diabetic milieu, inducible nitric oxide synthase (iNOS) expression is upregulated in retinal macrophages/microglia, which increases production of inflammatory cytokines and reactive oxygen species, worsening ischemia, blood-retinal-barrier (BRB) compromise, and disease progression. Arginase 1 (Arg1) is a ureohydrolase that competes with iNOS for their common substrate, L-arginine. Recent studies revealed a neuroprotective role of Arg1 in murine models of retinal ischemia/reperfusion injury and ischemic stroke and demonstrated the beneficial effects of Arg1 administration in these models. As DR also involves ischemia and inflammation, we hypothesized that exogenous Arg1 would be similarly beneficial through iNOS substrate depletion, resulting in decreased retinal inflammation and oxidative stress. Methods Using the obese, type 2 diabetic db/db mouse model that well-approximates human DR pathophysiology, the visual function, BRB, retinal inflammation, and oxidative stress were characterized in these mice aged 16-24-weeks and compared to lean, littermate controls. Visual function was determined by electroretinography (ERG) responses, visual acuity, and contrast threshold measurements. Retinas were collected for immunofluorescence or western blot analyses for iNOS, inflammatory cytokines (IL-1β, TNF-α), oxidative stress (3-nitrotyrosine, 4-hydroxynonenal), and BRB assessment, determined by both retinal albumin extravasation and co-labeling for ZO-1 (tight junction protein) and CD31 (endothelial cell-specific protein). To assess Arg1 therapeutic potential, recombinant Arg1 conjugated to polyethylene glycol (PEG-Arg1), for improved half-life, or a PEG-5000 vehicle control was administered intraperitoneally (25mg/kg) to 16-24-week-old db/db mice thrice weekly for 2 weeks and the same experiments were performed. Results Compared to lean littermate controls, db/db mice exhibited significant visual dysfunction, BRB compromise, and elevated retinal levels of iNOS, IL-1β, TNF-α, 3-nitrotyrosine, and 4-hydroxynonenal. PEG-Arg1 treatment significantly ameliorated visual function impairment and BRB compromise in these mice and markedly reduced iNOS levels, retinal inflammation, and oxidative stress compared to vehicle-treated db/db mice. Conclusion The retinal pathology of db/db mice significantly improved with PEG-Arg1 therapy. Though further investigation is necessary, these results suggest that systemic PEG-Arg1 administration has potential as a novel DR therapeutic.