Fig. 6.

Beneficial effects of diosgenin (DG) mediated by the sirtuin 6/nuclear factor erythroid 2-related factor 2 (Sirt6/Nrf2) pathway. After 24 h of drug treatment, proteins were extracted for detection. (A) Western blot (WB) of phosphorylated-Sirt6 (p-Sirt6), kelch-like ECH-associated protein 1 (Keap1), solute carrier family 7 member 11 (SLC7A11), and glutathione peroxidase 4 (GPX4). (B) Quantitative analysis of Keap1, SLC7A11, and GPX4. (C) Quantitative analysis of p-Sirt6. (D) WB of nuclear-Nrf2 (n-Nrf2). (E) Quantitative analysis of n-Nrf2. (F) Cycloheximide (CHX) chase assay to assess Nrf2 protein stability: HUVECs were incubated with 30 μM cycloheximide with or without DG for the indicated times. (G) Co-immunoprecipitation and WB to detect the interaction between Nrf2 and Keap1: HUVECs were treated with Z-Leu-Leu-Leu-al (MG-132) (5 μM) with or without DG, followed by cell lysis. (H, I) Nrf2 immunofluorescence images (H) and quantitative analysis (I) in differentially treated HUVECs. Data are presented as the mean ± standard deviation (SD) from 5 independent experiments (n = 5). Statistical significance is denoted as ^∗∗∗P < 0.001.