Journal: Frontiers in nutrition

Article Title: Hepatic PPARα Is Destabilized by SIRT1 Deacetylase in Undernourished Male Mice.

doi: 10.3389/fnut.2022.831879

Figure Lengend Snippet: FIGURE 3 | Regulation of PPARα protein level by SIRT1. (A) Transient transfection of human-derived HepG2 hepatocellular carcinoma cells with constitutively active SIRT1 decreased PPARα protein levels and acetylation, whereas transfection with dominant negative SIRT1 did not. (B) Incubating HepG2 cells with 100 µM of the natural SIRT1 activator resveratrol decreased PPARα protein and increased PPARα mRNA, similar to our mouse model of undernutrition. (C) Silencing SIRT1 in HepG2 cells increased PPARα protein and acetylation and (D) decreased ubiquitination of PPARα. Mean + SD; n = 3–4; ****P < 0.0001; ***P < 0.001; **P < 0.01; *adjusted P < 0.05. CA, constitutively active; DN, dominant negative; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; IP, immunoprecipitation; NS, not significant; PPARα, peroxisome proliferator-activated receptor-α; siRNA, small interfering RNA; SIRT1, NAD-dependent deacetylase sirtuin-1; Ub-PPARα, ubiquitinated PPARα.

Article Snippet: Alternatively, cells were transfected with 100 nM SIRT1 siRNA or control siRNA (#12241S and 6568S, Cell Signaling Technology, Danvers, MA) using Lipofectamine RNAiMAX Transfection Reagent (Invitrogen, Waltham, MA) according to the manufacturer’s instructions.

Techniques: Transfection, Derivative Assay, Dominant Negative Mutation, Ubiquitin Proteomics, Immunoprecipitation, Small Interfering RNA, Histone Deacetylase Assay

Journal: Frontiers in pharmacology

Article Title: Acacetin Protects Against High Glucose-Induced Endothelial Cells Injury by Preserving Mitochondrial Function via Activating Sirt1/Sirt3/AMPK Signals.

doi: 10.3389/fphar.2020.607796

Figure Lengend Snippet: FIGURE 5 | Acacetin rescued the impaired signal molecules induced by high glucose exposure in HUVECs. (A) Western blots of Sirt3 Sirt1, PGC-1α, pAMPK, t-AMPK in HUVECs cultured in 5.5 mM glucose or 33 mM glucose in the absence or presence of 0.3, 1, or 3 μM acacetin. (B) Acacetin reversed the decrease of Sirt3, Sirt1, PGC-1α and pAMPK in a concentration-dependent manner. n 5 individual experiments, **p < 0.01 vs. 5.5 mM glucose; ##p < 0.01 vs. 33 mM glucose alone.

Article Snippet: After 48 h transfection of control siRNA, Sirt1 siRNA or Sirt3 siRNA (Santa Cruz Biotechnology), the cells were incubated with 5.5 mM or 33 mM glucose culture medium in the absence or presence of 3 μM acacetin for 5 days, then collected for western blot analysis.

Techniques: Western Blot, Cell Culture, Concentration Assay

Journal: Frontiers in pharmacology

Article Title: Acacetin Protects Against High Glucose-Induced Endothelial Cells Injury by Preserving Mitochondrial Function via Activating Sirt1/Sirt3/AMPK Signals.

doi: 10.3389/fphar.2020.607796

Figure Lengend Snippet: FIGURE 6 | Sirt1 mediates the vascular protection of acacetin against hyperglycemia-induced injury by activating Sirt3/AMPK/PGC-1α. (A) Western blots of Sirt1, PGC-1α, Sirt3, pAMPK, t-AMPK in HUVECs cultured with 33 mM glucose and transfected with Sirt1 siRNA or Sirt3 siRNA or treated with the AMPK inhibitor dorsomorphin (Dorsom. 10 μM) in the absence or presence of 3 μM acacetin. (B) Silencing Sirt1, but not Sirt3 or inhibition of pAMPK decreased Sirt1 expression and abolished the Sirt1 activation by acacetin (n 5 individual experiments, **p < 0.01 vs. control siRNA 33 mM glucose alone; #p < 0.05, ##p < 0.01 vs. 33 mM glucose alone). (C) Acacetin reversal of the high glucose-induced reduction NAD+/NADH ratio was decreased by the NAMPT inhibitor GMX-1778 (10 nM) (n 5 individual experiments, **p < 0.01 vs. 5.5 mM glucose alone; ##p < 0.01 vs. 33 mM glucose alone). (D) Western blots of Sirt1 and Sirt3 in cells cultured with 33 mM glucose medium in the absence or presence of 3 μM acacetin and 33 mM glucose medium with 10 nM GMX-1778 in the absence or presence of 3 μM acacetin. (E) GMX-1778 decreased the expression of Sirt1 and Sirt3 proteins and abolished the upregulation of Sirt1 and Sirt3 by acacetin (n 5 individual experiments, **p < 0.01 vs. 33 mM glucose alone). (F) Schematic graph shows that acacetin protects against high glucose-induced vascular endothelial cell injury by increasing NAD+/NADH followed by Sirt1-mediated activation of Sirt3/AMPK/PGC-1α, thereby elevating mitochondrial antioxidation and inhibiting apoptosis.

Article Snippet: After 48 h transfection of control siRNA, Sirt1 siRNA or Sirt3 siRNA (Santa Cruz Biotechnology), the cells were incubated with 5.5 mM or 33 mM glucose culture medium in the absence or presence of 3 μM acacetin for 5 days, then collected for western blot analysis.

Techniques: Western Blot, Cell Culture, Transfection, Inhibition, Expressing, Activation Assay, Control

Journal: Frontiers in pharmacology

Article Title: Acacetin Protects Against High Glucose-Induced Endothelial Cells Injury by Preserving Mitochondrial Function via Activating Sirt1/Sirt3/AMPK Signals.

doi: 10.3389/fphar.2020.607796

Figure Lengend Snippet: FIGURE 8 | Alterations of related proteins in aortic tissues of STZ-diabetic ApoE−/−mice. (A) Western blots of SOD1, SOD2, Sirt3, PGC-1α, Sirt1, Bcl-2, Bax, and β-actin in control aorta with and without acacetin and in STZ-diabetic ApoE−/−mice aorta with and without acacetin. (B) Analysis of SOD1, SOD2, Sirt3, PGC-1α, and Sirt1 (relative to β-actin). (C) Analysis of Bcl-2 and Bax proteins, relative to β-actin. Inset: ratio of Bcl-2/Bax. (D) Western blots of pAMPK and t-AMPK (total AMPK) and relative level of pAMPK/t-AMPK. n 6, *p < 0.05, **p < 0.01 vs. control or acacetin; #p < 0.05, ##p < 0.01 vs. STZ. (E) Sirt3 and CD31 immunohistochemical staining of aortic root section from mice of control, with acacetin, STZ, and STZ with acacetin treatment (×10 magnification).

Article Snippet: After 48 h transfection of control siRNA, Sirt1 siRNA or Sirt3 siRNA (Santa Cruz Biotechnology), the cells were incubated with 5.5 mM or 33 mM glucose culture medium in the absence or presence of 3 μM acacetin for 5 days, then collected for western blot analysis.

Techniques: Western Blot, Control, Immunohistochemical staining, Staining

Journal: Chinese medicine

Article Title: Taraxerone inhibits M1 polarization and alleviates sepsis-induced acute lung injury by activating SIRT1.

doi: 10.1186/s13020-024-01002-z

Figure Lengend Snippet: Fig. 7 Taraxerone plays a protective role in sepsis-induced ALI mice through the SIRT1 pathway. The EX527 (10 mg/kg) was administered intraperitoneally 2 h before the taraxerone (0, 30 mg/kg) treatment, and the lung tissues and BALFs were collected after the CLP model was constructed 12 h later. A With H&E staining, we examined the lung tissue for any pathological changes. The bar represented 100 μm. B The breathing frequency and dynamic compliance of mice were detected by BUXCO system. C Detection of IL-1β, IL-6, TNF-α, and IL-18 mRNA levels. D ROS level in BALFs were determined using flow cytometry and analyzed with FlowJo. E Changes in the proportion of M1 macrophages in BALFs were detected using flow cytometry, and the data were analyzed using FlowJo. F Effects of taraxerone (30 mg/kg) and EX527 (10 mg/kg) on NLRP3, Cleaved Caspase-1, Pro Caspase-1, ASC, Pho-P65, Ace-P65, P65, Pho-IκB, and IκB protein levels. G–I MDA content, GSH content, and SOD activity were determined to reflect the oxidative stress levels of lung tissues. Data are presented as mean ± SD (n ≥ 6 in each group). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001

Article Snippet: SIRT1 polyclonal antibody (Proteintech, China) was added to the supernatant overnight at 4 °C.

Techniques: Construct, Staining, Flow Cytometry, Activity Assay

Journal: Chinese medicine

Article Title: Taraxerone inhibits M1 polarization and alleviates sepsis-induced acute lung injury by activating SIRT1.

doi: 10.1186/s13020-024-01002-z

Figure Lengend Snippet: Fig. 8 Taraxerone plays a protective role in acute lung injury by activating SIRT1. Taraxerone activates SIRT1 to block the NF-κB-NLRP3 inflammasome axis and to suppress the excessive ROS, inhibiting the M1 polarization of macrophages, and exerting a protective effect on acute lung injury mice.

Article Snippet: SIRT1 polyclonal antibody (Proteintech, China) was added to the supernatant overnight at 4 °C.

Techniques: Blocking Assay