anti arpc5  (Boster Bio)

Journal: Heliyon

Article Title: Dapagliflozin impedes endothelial cell senescence by activating the SIRT1 signaling pathway in type 2 diabetes

doi: 10.1016/j.heliyon.2023.e19152

Figure Lengend Snippet: Dapagliflozin restores endothelial nitric oxide synthase (eNOS) activity and mitigates oxidative stress in senescent endothelial cells. (a and b) Senescence-associated β-galactosidase (SA-β-gal) staining to detect senescence in endothelial cells stimulated with HG (30 mM). Mannitol was used as a control to rule out osmotic effects. n = 6–8, scale bar = 50 μm. Data are presented as means ± SD. *P < 0.05 versus normal glucose; # P < 0.05 versus high glucose. ( c, d, and e) Immunoblotting to measure p53 and p21 protein levels in HG-stimulated endothelial cells. β-Actin was used for normalization. n = 3. Data are presented as means ± SD. *P < 0.05, versus normal glucose; # P < 0.05 versus high glucose. ( f) Griess reaction to detect endothelial nitric oxide (NO) production in the plasma of db/db mice. NO levels decreased in the plasma of db/db mice, and this effect was reversed by the addition of dapagliflozin. n = 3. Data are presented as means ± SD. *P < 0.05, versus db/m; # P < 0.05 versus db/db. ( g and h) The fluorescent probe dihydroethidium (DHE) used to detect intracellular reactive oxygen species production. n = 3, scale bar = 50 μm. ( i and j) Immunoblotting to measure total eNOS expression and eNOS phosphorylation status. n = 3. Data are presented as mean s± SD. *P < 0.05 versus normal glucose; # P < 0.05 versus high glucose. NG, normal glucose; D, dapagliflozin; HG: high glucose; M, mannitol.

Article Snippet: The blots were blocked in 5% fat-free milk and probed with primary antibodies against p-eNOS, eNOS, and SIRT1 (1:1000; Cell Signaling Technologies, Inc., Danvers, MA, USA, #9570, #32027, and #9475) and p53, p21, p16, and β-actin (1:5000; Proteintech, #10442–1-AP, #10355–1-AP; and 1:1000; Proteintech, #16717-1-AP).

Techniques: Activity Assay, Staining, Control, Western Blot, Clinical Proteomics, Expressing, Phospho-proteomics

Journal: Heliyon

Article Title: Dapagliflozin impedes endothelial cell senescence by activating the SIRT1 signaling pathway in type 2 diabetes

doi: 10.1016/j.heliyon.2023.e19152

Figure Lengend Snippet: Dapagliflozin rescues decreased SIRT1 expression in senescent endothelial cells. (a and b) SIRT1 expression in HG-stimulated endothelial cells normalized to β-actin levels. Mannitol was used as a control to rule out osmotic effects. n = 3. (c) NAD + /NADH ratios to evaluate SIRT1 activity in HG-stimulated endothelial cells. n = 3. ( d and e) Nicotinamide (NAM), a specific and potent pharmacological inhibitor of SIRT1, was used to inhibit SIRT1 activity. Cells were then treated with dapagliflozin and SA-β-gal staining was used to identify senescent cells. n = 6–8, Scale bar = 50 μm. ( f, g, and h) p53 and p21 protein levels were assessed using immunoblotting, with β-actin as a loading control. n = 3. Data are presented as means ± SD. *P < 0.05, versus normal glucose; # P < 0.05 versus high glucose; + P < 0.05 versus high glucose + dapa. NG, normal glucose; D, dapagliflozin; HG, high glucose; M, mannitol; N, nicotinamide.

Article Snippet: The blots were blocked in 5% fat-free milk and probed with primary antibodies against p-eNOS, eNOS, and SIRT1 (1:1000; Cell Signaling Technologies, Inc., Danvers, MA, USA, #9570, #32027, and #9475) and p53, p21, p16, and β-actin (1:5000; Proteintech, #10442–1-AP, #10355–1-AP; and 1:1000; Proteintech, #16717-1-AP).

Techniques: Expressing, Control, Activity Assay, Staining, Western Blot