Müller glial dysfunction during diabetic retinopathy in rats is reduced by the acrolein-scavenging drug, 2-hydrazino-4,6-dimethylpyrimidine

Diabetologia. 2018 Dec;61(12):2654-2667. doi: 10.1007/s00125-018-4707-y. Epub 2018 Aug 15.

Abstract

Aims/hypothesis: Recent 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.

Methods: An 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.

Results: 2-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/interpretation: These 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.

Keywords: Acrolein; Advanced lipoxidation end-products; Diabetic retina; Electroretinography; Hydrazino compounds; Inflammatory signalling; Müller glia; Oxidative stress; Retinopathy; Scavenging agents.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acrolein / toxicity*
  • Animals
  • Cell Survival / drug effects
  • Chromatography, Liquid
  • Diabetic Retinopathy / metabolism
  • Electroretinography
  • Enzyme-Linked Immunosorbent Assay
  • Ependymoglial Cells / drug effects*
  • Ependymoglial Cells / metabolism*
  • Free Radical Scavengers / therapeutic use*
  • Immunohistochemistry
  • Lysine / metabolism*
  • Male
  • Mass Spectrometry
  • Oxidative Stress / drug effects
  • Rats
  • Rats, Sprague-Dawley

Substances

  • Free Radical Scavengers
  • Acrolein
  • Lysine