Resveratrol, rapamycin and MG‐132 as inducers of autophagy in ARPE‐19 cells

Abstract Purpose To investigate the effect of the polyphenol resveratrol (3,5,4'‐trihydroxy‐trans‐stilbene), mTOR(definition)-inhibiting drug studied for extending healthspan and lifespan." style="text-decoration:underline dotted; text-underline-offset:2px; cursor:help;">rapamycin(definition) (RAP) and proteasome inhibitor MG‐132 on cell death and autophagy(definition) in human retinal pigment epithelium‐derived ARPE‐19 cells. Methods ARPE‐19 cells were exposed to different treatment regimens: 10‐50microM resveratrol, 50‐100 nM RAP, 50 microM chloroquine (CQ) and 50‐100 nM MG‐132 over 48 hours. The levels of LC3‐II, mammalian target of rapamycin (mTOR), Hsp70, p62 were determined by Western blot analysis; autophagic vacuoles (AVs) formation was detected by acridine orange, pDendra2‐hLC3 expression and transmission electron microscopy; cell death was quantified using annexin‐V‐FITC/propidium iodide (PI) labeling on flow cytometry. Results Exposure to RAP and MG‐132 caused a time‐ and concentration dependent induction of autophagy that could be inhibited by 3‐methyladenine (3‐MA), while the induction of an active autophagic flux could be verified with CQ treatment, a blocker of the autophagosome‐lysosome fusion. Similarly, resveratrol alone could induce autophagy in ARPE‐19 cells, serving as a pro‐survival signal in ARPE‐19 cells. Inhibition with 3‐MA increased the death rate of resveratrol treated ARPE‐19 cell, further proving the autophagy‐related protective role of resveratrol. Conclusion Resveratrol at lower concentrations, RAP and MG‐132 can provide a pro‐survival stimulus to ARPE‐19 cells by inducing autophagy. This property can possibly be used for prolonging the lifespan of retinal pigment epithelium in diseases such as age‐related macular degeneration.