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Protective effect of Metformin against H2O2-induced oxidative damage in human retinal pigment epithelial (RPE) cells through activation of AMPK/mTOR/Autophagy pathway
Zheng, W.
Conference Name第十二届广东省中西医结合学会神经科专委会年会
Source Publication第十二届广东省中西医结合学会神经科专委会年会
Conference Date2019-08
Conference Place广东

Objective: Age-related macular degeneration (AMD) is characterized by a progressive loss of their support cells, the retinal pigmented epithelium (RPE). Compared with normal RPE, AMD RPE exhibit increased susceptibility to oxidative stress, produce higher levels of reactive oxygen species (ROS) under stress conditions, and showed reduced mitochondrial activity. Therefore, it could be a great strategy to find some drugs that can effectively protect RPE cells which is desired to treat and slow the process of AMD. Metformin, a first line anti type 2 diabetes drug, has recently been shown to extend life-span in various species and therefore became the first anti-aging drug in clinical trial. However, the anti-oxidative effects of metformin and its underlying mechanisms in RPE cells are not well known. Hydrogen peroxide (H2O2) is an oxidizing agent, commonly used to cause irreversible oxidative damage and activation of the apoptotic cascade in various cell models. Therefore, H2O2-induced oxidative injury in RPE cell is a suitable model for investigating the effects of drug candidates for the treatment of AMD disorders. The AMP-activated protein kinase (AMPK) is a nutrient sensor playing an important role in metabolism and the regulation of whole body energy balance. The activation of AMPK is also activated by metformin and this activation is required for many metabolic effects of metformin including fatty acid oxidation. An increase in autophagosomes was observed in AMD RPE cultures. Metformin is also closely related with aging and autophagy. Based on these observations, the present study was designed to investigate the protective effects of metformin against H2O2-induced cytotoxicity in RPE cells and its underlying mechanisms. Content: In this experiment we used H2O2 to establish oxidative damage model. Hoechst staining and MTT to detect nuclear morphological changes and cell viability; flow cytometry, caspase3 activation kit to detect cell apoptosis; lactate dehydrogenase kit to detect cell membrane damage; mitochondrial membrane potential assay kit (JC-1) and CellROX Deep Red Reagent (ROS) to detect mitochondrial function changes; Immunofluorescence (IF) / Immunocytochemistry (ICC) / Western blot was used to detect changes in pathway protein levels. In vivo study, vitreous injection was used to establish animal model and intraperitoneal injection was used to give mice drugs. Results: Our result showed that metformin, in clinically relevant concentrations, protected the RPE cells from H2O2-induced cell death. Further study revealed that metformin significantly ameliorated cell death due to H2O2 insult by restoring abnormal changes in nuclear morphology, intracellular ROS, lactate dehydrogenase and mitochondrial membrane potential. Hoechst staining assay and flow cytometry analysis revealed that metformin significantly reduced the apoptosis of RPE cells exposed to H2O2. We also investigated the roles of the AMPK pathway in the protective effect of metformin. Western blotting analysis demonstrated that metformin time- and concentration- dependently stimulated the expression of AMPK, mTOR, Autophagy related markers in RPE cells while block AMPK or autophagy blocked the protective effect of metformin. Similar results were obtained in RPE cells. Conclusion: Taken together, these results indicated that metformin is able to protect the retinal pigmented epithelium cells from H2O2-induced cell death, at least in part, via the activation of AMPK/mTOR/Autophagy. As metformin is comparatively cheaper with very less side effects in clinic, our finding support its potential to be a drug for prevention and treatment of Age-related macular degeneration.

KeywordMetformin Age-related Macular Degeneration Rpe Cells Ampk
The Source to ArticlePB_Publication
Document TypeConference paper
Faculty of Health Sciences
Corresponding AuthorZheng, W.
Recommended Citation
GB/T 7714
Zheng, W.. Protective effect of Metformin against H2O2-induced oxidative damage in human retinal pigment epithelial (RPE) cells through activation of AMPK/mTOR/Autophagy pathway[C],2019.
APA Zheng, W..(2019).Protective effect of Metformin against H2O2-induced oxidative damage in human retinal pigment epithelial (RPE) cells through activation of AMPK/mTOR/Autophagy pathway.第十二届广东省中西医结合学会神经科专委会年会.
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