Resveratrol inhibits the progression of premature senescence partially by regulating v-rel avian reticuloendotheliosis viral oncogene homolog A (RELA) and sirtuin 1 (SIRT1)

Figures & data

Figure 1. Resveratrol promotes H₂O₂-induced proliferation activity and inhibits senescence and apoptosis of BMMSCs. Proliferation, senescence and apoptosis of BMMSCs treated with H₂O₂ were detected by (A) CCK-8 assay, (B) β-galactosidase staining assay and (C) flow cytometry assay, respectively. After treatment with different concentrations of resveratrol (1 µM, 5 µM, and 10 µM) for 24 h, the proliferation, senescence and apoptosis of H₂O₂-induced BMMSCs were detected by (D) CCK-8 assay, (E) β-galactosidase staining assay, and (F) flow cytometry assay. *p < 0.05, **p < 0.01, ***p < 0.001 versus the control group.

Figure 2. RELA is the target of resveratrol in premature senescence. (A) Swiss Target Prediction and Comparative Toxicogenomics Database were used to identify the potential targets of resveratrol; (B) the biological function of the selected target genes was analyzed by PANTHER and gene ontology; (C) Metascape and gene ontology were used to select genes with higher expression from the selected target genes with transcriptional activity.

Figure 3. Simulate and verify the combination of resveratrol and RELA. (A) Chemical structure of resveratrol; (B) the 3D structure model of RELA was conducted by SWISSMODEL and Autodock was used to calculate the possibility of binding between resveratrol molecules and RELA protein molecules; (C) the mRNA levels of RELA were detected by RT-qPCR; (D) the protein levels of RELA were detected by western blot. *p < 0.05, ***p < 0.001 versus the control group.

Figure 4. Overexpression of RELA weakens the effect of resveratrol in the treatment of premature senescence. (A) CCK-8 and (B) flow cytometry were used to detect BMMSCs proliferation activity and apoptosis rate induced by H₂O₂ in groups pc-NC, pc-NC + resveratrol and pc-RELA + resveratrol; (C) the expression of apoptosis-related proteins (Bax and Bcl-2) was detected by western blot; (D) β-galactosidase staining assay was performed to determine the senescence of BMMSCs; (E) the expression of senescence-related proteins (p53 p21 and p16) was detected by western blot; (F) the mRNA expression of RELA was detected by qRT-PCR; (G) the expression of RELA was detected by western blot. *p < 0.05, **p < 0.01, ***p < 0.001 versus another group.

Figure 5. SIRT1 is the target of RELA in premature senescence. (A) PRIDB was used to predict and screen the putative targets of RELA; (B) dual-luciferase reporter assay is conducted to verify the interaction between RELA and SIRT1; (C) after overexpression of RELA, the mRNA expression of SIRT1 were detected by qRT-PCR; (D) after overexpression of RELA, the protein expression of SIRT1 were detected by western blot; (E) the mRNA and protein expression of SIRT1 after treatment with different concentrations of resveratrol (1 µM, 5 µM, and 10 µM) for 24 h were evaluated by qRT-PCR and western blot. *p < 0.05, **p < 0.01, ***p < 0.001 versus another group.

Figure 6. Silencing SIRT1 weakens the effect of resveratrol in the treatment of premature senescence. (A) Efficiency of SIRT1 silence was tested. (B) CCK-8 and (C) flow cytometry were used to detect BMMSCs proliferation activity and apoptosis rate induced by H₂O₂ in groups si-NC, si-NC + resveratrol and si-RELA + resveratrol; (D) the expression of apoptosis-related proteins (Bax and Bcl-2) was detected by western blot; (E) β-galactosidase staining assay was performed to determine the senescence of BMMSCs; (F) the expression of senescence-related proteins (p53, p21 and p16) was detected by western blot. *p < 0.05, **p < 0.01, ***p < 0.001 versus another group.

Figure 7. Resveratrol inhibits premature senescence through targeted regulation RELA/SIRT1. (A) CCK-8 and (B) flow cytometry were used to detect BMMSCs proliferation activity and apoptosis rate induced by H₂O₂ in group pc-NC + pc-NC, pc-NC + pc-NC + resveratrol, pc-RELA + pc-NC + resveratrol and pc-RELA + pc-SIRT1+ resveratrol; (C) the expression of apoptosis-related proteins (Bax and Bcl-2) was detected by western blot; (D) β-galactosidase staining assay was performed to determine the senescence of BMMSCs; (E) the expression of senescence-related proteins (p53, p21 and p16) was detected by western blot. *p < 0.05, **p < 0.01, ***p < 0.001 versus another group.

Data availability statement

The raw data supporting the conclusions of this manuscript will be made available by the authors, without undue reservation, to any qualified researcher.