Journal: Molecules

Article Title: Paeonol Attenuates Atherosclerosis by Inhibiting Vascular Smooth Muscle Cells Senescence via SIRT1/P53/TRF2 Signaling Pathway

doi: 10.3390/molecules29010261

Figure Lengend Snippet: Paeonol inhibits the senescence of VSMCs and activates SIRT1 signaling in ApoE −/− mice. ( A ) Representative α-SMA and SA-β-gal immunofluorescence in artery intracuff lesions after treatment with paeonol in ApoE −/− mice. DAPI was used for nuclear staining. Scale bar, 100 μm. Paeonol decreased the percentage of ( B ) SA-β-gal and ( C ) SA-β-gal colocalized with α-SMA in artery intracuff lesions of ApoE −/− mice. ( D ) SIRT1, P53, and TRF2 protein levels were down-regulated after paeonol treatment by Western blotting analysis. The results are expressed as mean ± SD, ** p < 0.01 versus control; # p < 0.05, ## p < 0.01 versus model.

Article Snippet: The anti-SIRT1 antibody (Cat. No. R25721) and the goat anti-rabbit IgG secondary antibody (Cat. No. 511203) were obtained from Chengdu Zen Bioscience Co., Ltd. (Chengdu, China).

Techniques: Immunofluorescence, Staining, Western Blot

Journal: Molecules

Article Title: Paeonol Attenuates Atherosclerosis by Inhibiting Vascular Smooth Muscle Cells Senescence via SIRT1/P53/TRF2 Signaling Pathway

doi: 10.3390/molecules29010261

Figure Lengend Snippet: Paeonol increases the expression of SIRT1 and attenuates telomere damage in t-BHP-induced senescent VSMCs. VSMCs were exposed to t-BHP and then treated with paeonol, protein expression levels of SIRT1 ( A ) and TRF2 ( B ) were analyzed by Western blotting. ( C ) The level of TEL mRNA was increased after paeonol treatment by real-time PCR. ( D , E ) Paeonol decreased the levels of the SASP markers (IL-6 and TNF-α) in the cell supernatant using ELISA assay. ( F ) The cell proliferation capacity of VSMCs was determined by the EdU staining assay. The results are expressed as mean ± SD, ** p < 0.01 versus control; # p < 0.05, ## p < 0.01 versus t-BHP.

Article Snippet: The anti-SIRT1 antibody (Cat. No. R25721) and the goat anti-rabbit IgG secondary antibody (Cat. No. 511203) were obtained from Chengdu Zen Bioscience Co., Ltd. (Chengdu, China).

Techniques: Expressing, Western Blot, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Staining

Journal: Molecules

Article Title: Paeonol Attenuates Atherosclerosis by Inhibiting Vascular Smooth Muscle Cells Senescence via SIRT1/P53/TRF2 Signaling Pathway

doi: 10.3390/molecules29010261

Figure Lengend Snippet: SIRT1 is a potential cellular target of paeonol in VSMCs. ( A ) Paeonol promoted the resistance of SIRT1 to different temperature gradients, which was detected by CETSA in VSMCs. ( B ) Paeonol enhanced the resistance of SIRT1 to proteases, which was investigated by DARTS. ( C ) SIRT1 was successfully silenced in VSMCs by immunofluorescence staining and Western blot. Protein expression levels of senescence-associated proteins (P53, P16, P21) ( D ) and TRF2 ( E ) were analyzed by Western blotting in the control, t-BHP, siSIRT1, paeonol, and siSIRT1 + paeonol groups. ( F ) The level of TEL mRNA was assessed by real-time PCR. The results are expressed as mean ± SD, ** p < 0.01 versus control; # p < 0.05, ## p < 0.01 versus t-BHP; Δ p < 0.05, ΔΔ p < 0.01 versus paeonol.

Article Snippet: The anti-SIRT1 antibody (Cat. No. R25721) and the goat anti-rabbit IgG secondary antibody (Cat. No. 511203) were obtained from Chengdu Zen Bioscience Co., Ltd. (Chengdu, China).

Techniques: Immunofluorescence, Staining, Western Blot, Expressing, Real-time Polymerase Chain Reaction

Journal: Molecules

Article Title: Paeonol Attenuates Atherosclerosis by Inhibiting Vascular Smooth Muscle Cells Senescence via SIRT1/P53/TRF2 Signaling Pathway

doi: 10.3390/molecules29010261

Figure Lengend Snippet: Paeonol inhibits VSMC senescence by activating the SIRT1/P53/TRF2 signaling pathway. The results are expressed as mean ± SD, ** p < 0.01 versus control; # p < 0.05, ## p < 0.01 versus t-BHP; ΔΔ p < 0.01 versus t-BHP + paeonol.

Article Snippet: The anti-SIRT1 antibody (Cat. No. R25721) and the goat anti-rabbit IgG secondary antibody (Cat. No. 511203) were obtained from Chengdu Zen Bioscience Co., Ltd. (Chengdu, China).

Techniques:

Journal: Oncology letters

Article Title: p53 inhibits the upregulation of sirtuin 1 expression induced by c-Myc.

doi: 10.3892/ol.2017.6661

Figure Lengend Snippet: Figure 1. p53 inhibits the upregulation of Sirt1 protein expression level mediated by c‑Myc. (A) Left panel presents K562, H1299, U2OS and 293A cells trans fected with c‑Myc expression vectors. Right panel presents the quantitation of Sirt1 protein expression levels from K562, H1299, U2OS and 293A cells using GAPDH expression as the internal control (Student's t‑test; n=3; P<0.05). Error bars represent the mean ± standard error. (B) Left panel presents p53 wild type or knockout MEF/3T3 transfected with c‑Myc expression vectors. Right panel presents the quantitation of Sirt1 protein expression levels from p53 wild type or knockout MEF/3T3 using GAPDH expression level as the internal control (Student's t‑test; n=3; P<0.05). (C) Left panel presents p53+/+ or p53‑/‑ MEF/3T3 and HCT116 cells treated with 10058‑F4 with indicated concentrations for 24 h. Right panel presents the quantitation of Sirt1 protein expression levels from p53+/+ or p53‑/‑ MEF/3T3 and HCT116 cells using GAPDH expression level as the internal control (SNK‑q test; P<0.05). Cell lysates of A, B and C were analyzed by western blotting. Sirt1, sirtuin 1; p53, tumor protein p53. *P<0.05; **P<0.01.

Article Snippet: Anti-c-Myc (N262; dilution, 1:100; cat. no. sc-500771), anti-human Sirt1 (dilution, 1:200; cat. no. sc-15404), anti-p53 (dilution, 1:500; cat. no. sc-126) and anti-GAPDH (dilution, 1:500; cat. no. sc-47724) antibodies were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA).

Techniques: Expressing, Quantitation Assay, Control, Knock-Out, Transfection, Western Blot

Journal: Oncology letters

Article Title: p53 inhibits the upregulation of sirtuin 1 expression induced by c-Myc.

doi: 10.3892/ol.2017.6661

Figure Lengend Snippet: Figure 2. p53 inhibits the upregulation of Sirt1 mRNA level mediated by c‑Myc. (A) K562 cells were transfected with c‑Myc expression vectors or treated with 10058‑F4. (B) 293A cells were transfected with c‑Myc expression vectors or treated with 10058‑F4. (C) p53 wild type or knockout MEF/3T3 were transfected with c‑Myc expression vectors or treated with 10058‑F4. RNA was extracted from cells in A, B and C, and analyzed by quantitative polymerase chain reaction. Sirt1, sirtuin 1; p53, tumor protein p53.

Article Snippet: Anti-c-Myc (N262; dilution, 1:100; cat. no. sc-500771), anti-human Sirt1 (dilution, 1:200; cat. no. sc-15404), anti-p53 (dilution, 1:500; cat. no. sc-126) and anti-GAPDH (dilution, 1:500; cat. no. sc-47724) antibodies were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA).

Techniques: Transfection, Expressing, Knock-Out, Real-time Polymerase Chain Reaction

Journal: Oncology letters

Article Title: p53 inhibits the upregulation of sirtuin 1 expression induced by c-Myc.

doi: 10.3892/ol.2017.6661

Figure Lengend Snippet: Figure 3. p53 blocks the binding of c‑Myc to the Sirt1 promoter. (A) Alignment of the proximal core region of human, mouse and cow Sirt1 promoter sequences. The underlined bases indicate the p53 binding site and capital bases indicate the putative c‑Myc binding sites. (B) Human Sirt1 promoter reporter plasmids containing 2852, 200 or 102 bp upstream of the transcription start site were separately co‑transfected with pcDNA 3.1 or c‑Myc expres sion vectors into H1299 cells for 36 h. (C) H1299 cells were co‑transfected with wild type (‑200) or mutated Sirt1 promoter reporter plasmids (E2/mut or E3/mut) with c‑Myc expression vector. (D) HeLa nuclear extracts were incubated with biotinylated oligonucleotides 222 containing E2 box of human Sirt1 promoter or 276 (50 bp upstream of 222) and streptavidin‑agarose beads. The precipitated complexes were analyzed by western blotting. (E) H1299 cells were co‑transfected with Sirt1 promoter reporter plasmids with c‑Myc and p53 expression vectors as indicated. The relative luciferase activities of B, C and E are presented as the mean ± standard deviation of triplicate samples and were representative of three independent experiments. (F) Chromatin immunoprecipitation assay was performed in K562 and 293A cells to detect the recruitment of c‑Myc on the Sirt1 promoter E2 and E1 sites. The hTERT promoter was used as positive control. Sirt1, sirtuin 1; bp, base pair; mut, mutant; hTERT, human telomerase reverse transcriptase; p53, tumor protein p53. *P<0.05; **P<0.01.

Article Snippet: Anti-c-Myc (N262; dilution, 1:100; cat. no. sc-500771), anti-human Sirt1 (dilution, 1:200; cat. no. sc-15404), anti-p53 (dilution, 1:500; cat. no. sc-126) and anti-GAPDH (dilution, 1:500; cat. no. sc-47724) antibodies were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA).

Techniques: Binding Assay, Expressing, Plasmid Preparation, Incubation, Western Blot, Luciferase, Standard Deviation, Chromatin Immunoprecipitation, Positive Control, Mutagenesis, Reverse Transcription

Journal: International Journal of Molecular Medicine

Article Title: Resveratrol alleviates the cytotoxicity induced by the radiocontrast agent, ioxitalamate, by reducing the production of reactive oxygen species in HK-2 human renal proximal tubule epithelial cells in vitro

doi: 10.3892/ijmm.2015.2404

Figure Lengend Snippet: Role of sirtuin (SIRT)1 and SIRT3 in ioxitalamate-induced cytotoxicity. (A) Representative western blots of SIRT1 and phospho-SIRT1 were examined following 48 h of treatment with ioxitalamate and/or resveratrol. (B) Compared to treatment with ioxitalamate alone, changes in cell viability were examined following 48 h of treatment with ioxitalamate plus the SIRT1 inhibitor, EX-527, or the SIRT1 activator, SRT-1720, by MTT assay. ** p<0.01 and *** p<0.005.

Article Snippet: Primary antibodies were purchased from Cell Signaling Technology, Inc. [anti-cleaved caspase-3 (#9661), anti-survivin (#2808) and anti-LC3B (#2775)], and GeneTex [Irvine, CA, USA; anti-B-cell lymphoma 2 (Bcl-2; GTX127958), anti-cellular inhibitor of apoptosis protein (cIAP)1 GTX110087), anti-cIAP2 (GTX113128), anti-B-cell lymphoma-extra large (Bcl-xL; GTX105661), anti-receptor-interacting protein kinase 3 (RIP3; GTX107574), anti-sirtuin (SIRT)1; GTX61042), anti-phospho-SIRT1 (GTX61962) and SIRT3 (GTX89507)].

Techniques: Western Blot, MTT Assay

Journal: Autophagy

Article Title: CERKL regulates autophagy via the NAD-dependent deacetylase SIRT1

doi: 10.1080/15548627.2018.1520548

Figure Lengend Snippet: Table List of primary antibodies used in this study.

Article Snippet: Anti-phospho-SIRT1 (Ser27) , Cell Signaling Technology 2327 , Human SIRT1 p-S27 , 1:500 for WB.

Techniques: