Figure 2. Autophagy is Inhibited by Rhein in Vitro.

(A) Chemical and molecular structures of emodin (A bioactive component of rhubarb). (B) NRK-52E cells were treated with rhein (C₁₅H₈O₆) 1 μg/ml and emodin (C₁₅H₁₀O₅) 10 μM with or without HBSS for 1 hour and subjected to western blot analysis of LC3 I/II and beclin1. (C) NRK-52E cells were exposed to rhein 1, 5, and 10 μg/ml with or without HBSS for 1 hour and subjected to western blot analysis of LC3 I/II and beclin1. (D) NRK-52E cells were treated with rhein 5 μg/ml with or without LiCl 10 mM for 1 hour and subjected to western blot analysis of LC3 I/II. (E) NRK-52E cells were transfected with mRFP-LC3 and treated with rhein 5 μg/ml with or without HBSS and bafilomycin A1 10 nM for 2 hours and subjected to fluorescence microscopy. Scale bar = 5 μm. (F) NRK-52E cells were exposed to HBSS and bafilomycin A1 10 nM with or without rhein 1 μg/ml for 0, 0.5, 1, 2 and 6 hours, and subjected to western blot analysis of LC3 I/II. Data are expressed as mean ± SD, *P < 0.05, ^**P < 0.01, ^§P < 0.05 vs. co-treatment of HBSS and bafilomycin A1 at 0.5 hour, ^#P < 0.05 vs. co-treatment of HBSS and bafilomycin A1 at 1 hour, ^##P < 0.01 vs. co-treatment of HBSS and bafilomycin A1 at 2 h, ^&P < 0.05 vs. co-treatment of HBSS and bafilomycin A1 at 6 hours, ^&&P < 0.01 vs. co-treatment of HBSS and bafilomycin A1 at 6 hours. NS, not statistically significant. Abbreviation: HBSS, Hank's balanced salt solution; LiCl, lithium chloride.