Abstract
Aims/hypothesisRecent studies suggest that abnormal function in Müller glial cells plays an important role in the pathogenesis of diabetic retinopathy. This is associated with the selective accumulation of the acrolein-derived advanced lipoxidation end-product, Nε-(3-formyl-3,4-dehydropiperidino)lysine (FDP-lysine), on Müller cell proteins. The aim of the current study was to identify more efficacious acrolein-scavenging drugs and determine the effects of the most potent on Müller cell FDP-lysine accumulation and neuroretinal dysfunction during diabetes.MethodsAn ELISA-based in vitro assay was optimised to compare the acrolein-scavenging abilities of a range of drugs. This identified 2-hydrazino-4,6-dimethylpyrimidine (2-HDP) as a new and potent acrolein scavenger. The ability of this agent to modify the development of diabetic retinopathy was tested in vivo. Male Sprague Dawley rats were divided into three groups: (1) non-diabetic; (2) streptozotocin-induced diabetic; and (3) diabetic treated with 2-HDP in their drinking water for the duration of diabetes. Liquid chromatography high-resolution mass spectrometry was used to detect 2-HDP reaction products in the retina. Immunohistochemistry, real-time quantitative (q)RT-PCR and electroretinography were used to assess retinal changes 3 months after diabetes induction.Results2-HDP was the most potent of six acrolein-scavenging agents tested in vitro (p < 0.05). In vivo, administration of 2-HDP reduced Müller cell accumulation of FDP-lysine at 3 months in rats rendered diabetic with streptozotocin (p < 0.001). A 2-HDP adduct was identified in the retinas of diabetic animals treated with this compound. 2-HDP supplementation was associated with reduced Müller cell gliosis (p < 0.05), reduced expression of the oxidative stress marker haem oxygenase-1 (p < 0.001) and partial normalisation of inwardly rectifying K+ channel 4.1 (Kir4.1) expression (p < 0.001 for staining in perivascular regions and the innermost region of the ganglion cell layer). Diabetes-induced retinal expression of inflammatory markers, inflammatory signalling compounds and activation of retinal microglial cells were all reduced in 2-HDP-treated animals. Retinal neurophysiological defects in diabetic animals, as indicated by changes in the electroretinogram 7 weeks after induction of diabetes, were also reduced by 2-HDP (p < 0.05–0.01 for b-wave amplitudes at flash intensities from −10 to +10 dB; p < 0.01 for time to peak of summed oscillatory potentials at +10 dB).Conclusions/interpretationThese findings support the hypothesis that Müller cell accumulation of FDP-lysine plays an important role in the development of diabetic retinopathy. Our results also suggest that 2-HDP may have therapeutic potential for delaying or treating this sight-threatening complication.
Highlights
Diabetic retinopathy is a leading cause of newly diagnosed blindness in people of working age in industrialised nations [1]
Oxidative stress results in the formation of lipid aldehydes, which covalently bind to nucleophilic amino-acid residues to form adducts known as advanced lipoxidation endproducts (ALE) [26]
This process has been implicated in the pathogenesis of a range of ocular diseases, including diabetic retinopathy
Summary
Diabetic retinopathy is a leading cause of newly diagnosed blindness in people of working age in industrialised nations [1]. There is no question that microvascular pathology is central to the development of this condition, in recent years it has become increasingly clear that diabetes significantly impacts neuronal and glial cell function in the retina [2, 3]. Many of these important functions become disturbed and the cells assume a reactive phenotype characterised by the upregulation of glial fibrillary acidic protein (GFAP) and the production of inflammatory chemokines and cytokines [5]. Given the importance of Müller cells in maintaining neurovascular structure and function in the retina, their dysfunction in diabetes is believed to be a major factor that contributes to the evolution of the microvascular and neurodegenerative lesions of diabetic retinopathy [6, 7]
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