Abstract
Purpose :
Advanced glycation end products (AGEs) are toxic compounds resulting from the non-enzymatic modification of biomolecules by sugars or their metabolites. Ocular tissues are particularly vulnerable to glycative stress-derived toxicity due to their poor regenerative capacity. Strategies to diminish the levels of these toxic compounds could help fight the pathological dysfunction in ocular tissue caused by glycation-derived damaged.
This study tested the hypothesis that enhanced proteolytic capacity via mTOR pathway inhibition could accelerate the removal of glycated biomolecules, thereby minimizing toxicity derived from glycative stress.
Methods :
We exposed cultured human retinal pigment epithelial cells (RPE) and human lens epithelial cells (HLEC) to methylglyoxal (MGO), a highly reactive α-dicarbonyl that induces glycative stress in vitro, in presence of Rapamycin, a specific inhibitor of mTOR approved by the Food and Drug Administration. We evaluated the accumulation of AGEs by Western blotting and monitored cell viability using the CellTiter-Blue Cell Viability Assay. We also analyzed AGEs content in ocular tissues from wild type and p62-/- mice by immunohistochemistry.
Results :
MGO-treatment for 24 hours led to cytotoxicity in a dose-dependent manner in both cell types. At concentrations below 0.5 mM, MGO caused no significant loss of cell viability and no methylglyoxal-derived hydroimidazolone (MGH1), a marker of AGEs, was detected. Loss of cell viability is evident at concentrations above 1 mM with a concomitant accumulation of MGH1. Rapamycin treatment reduced glycation-derived toxicity in both cell types at highest concentrations of the glycating reagent. In vivo we found higher levels of MGH1 in retinal pigment epithelium from mice that lack the autophagic-receptor p62. Mouse embryonic fibroblasts derived from these p62-/- mice were more vulnerable to MGO-treatment compared to control and protection by Rapamycin was not observed in absence of p62, a selective autophagy adaptor protein for misfolded and ubiquitinated proteins.
Conclusions :
Our research indicates that mTOR inhibition is a new potential therapeutic target to diminish the accumulation of AGEs, which are drivers of different eye-related disorders. The shielding role requires, at least in part, the contribution of the autophagy receptor p62, suggesting a protective role of autophagy in the clearance of cytotoxic AGEs.
This is a 2020 ARVO Annual Meeting abstract.