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

Purpose To investigate the effect of the polyphenol resveratrol (3,5,4'-trihydroxy-trans-stilbene), rapamycin (RAP) and proteasome inhibitor MG-132 on cell death and autophagy 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.