Fig. 8

Resveratrol activates SIRT7 and inhibits metastasis. a Immunoblots showing SMAD4 acetylation level in an in vitro SIRT7 deacetylation assay: 2 mM NAD⁺, 1 µg GST-SIRT7, bead-bound HA-SMAD4, and Resveratrol (Res) were included as indicated. b Immunoblots showing SMAD4 acetylation level in wild-type or SIRT7 KO HEK293T cells treated with resveratrol. Note the SIRT7-dependent effect of SMAD4 deacetylation and SIRT7-independent effect of H3K9 deacetylation. c Resveratrol treatment accelerated the protein degradation of HA-SMAD4, and that was abolished in SIRT7 KO cells. d Representative immunoblots showing accelerated SMAD4 degradation upon resveratrol treatment in SIRT1 KO BT549 cells. e Quantification of d by Image J; f Phase contrast images of 4T1 cells showing epithelial or mesenchymal features. Noted that Resveratrol inhibited EMT. Scale bar, 100 µm. g Immunoblots showing decreased Smad4, Fn1, and Vim upon resveratrol treatment. Noted that p-Smad2 levels were not affected. h Wound healing assay for 4T1 cells treated with TGF-β1 (5 ng/ml) and/or resveratrol. Scale bar, 200 µm. i Quantification of migration ability in h, represented as relative migration area calculated by Image J. j Quantitative RT-PCR analysis of ANGPTL4 and CXCL8 mRNA levels in breast cancer cells treated with resveratrol or vehicle. k Representative images of H&E staining lung metastatic nodules in Balb/c mice injected with 4T1 cells and treated with resveratrol on day 7 (50 mg/kg). Scale bar, 1 cm. l Scatter blot showing lung metastatic nodules. m Kaplan–Meier survival of mice in k. *P < 0.05, calculated by log rank test. n Representative IHC staining of Smad4 in lung metastases from 4T1 mice k. Scale bar, 100 μm. *P < 0.05, **P < 0.01. P-values are calculated by one-way analysis of variance (ANOVA) e, non-parametric Mann–Whitney U-test l, Student’s t-test i, j or log rank test m. Data are shown as mean ± S.E.M