Journal: Free radical biology & medicine

Article Title: Nitroglycerin drives endothelial nitric oxide synthase activation via the phosphatidylinositol 3-kinase/protein kinase B pathway

doi: 10.1016/j.freeradbiomed.2011.09.020

Figure Lengend Snippet: PI3K involvement in GTN-dependent vasodilation. (A) Effect of wortmannin (PI3K inhibitor) pretreatment upon the acetylcholine (Ach)- or GTN-induced dilation of rat aortic rings. (B) Nitroglycerin EC50 values measured in rat aortic rings in the presence of the indicated concentrations of Akt 1/2 inhibitor. (C) Vasodilation experiment with mesenteric tissue recovered from p110γ-knockout mice. p110γ is the catalytic subunit of endothelial PI3K enzyme. Both pharmacologic inhibition and genetic knockout of PI3K inhibited GTN-induced dilation of conducing (aorta) and resistant (mesenteric artery) vessels. *P<0.05, **P<0.01.

Article Snippet: Human and bovine aortic endothelial cells (HAEC or BAEC; Allcells, Emeryville, CA, USA), human microvascular endothelial cells (HMEC), and primary mouse endothelial cells (MEC) were cultured at 37 °C, 5% CO 2 in corresponding endothelial medium with supplements including growth factors, 10% FBS, and 1% antibiotics–antimycotics (Sigma).

Techniques: Knock-Out, Inhibition

Journal: Free radical biology & medicine

Article Title: Nitroglycerin drives endothelial nitric oxide synthase activation via the phosphatidylinositol 3-kinase/protein kinase B pathway

doi: 10.1016/j.freeradbiomed.2011.09.020

Figure Lengend Snippet: PI3K/Akt-dependent phosphorylation of eNOS elicited by nitroglycerin. BAEC were challenged with nitroglycerin (500 nM) for 3 min before harvest. PI3K inhibitor wortmannin (100 nM) or Akt inhibitor (20 µM) was dissolved in DMSO and added to the cell cultures 1 h before GTN treatment; DMSO was also added at the same concentration (v/v) to control groups. Final DMSO concentration did not exceed 0.1%.

Article Snippet: Human and bovine aortic endothelial cells (HAEC or BAEC; Allcells, Emeryville, CA, USA), human microvascular endothelial cells (HMEC), and primary mouse endothelial cells (MEC) were cultured at 37 °C, 5% CO 2 in corresponding endothelial medium with supplements including growth factors, 10% FBS, and 1% antibiotics–antimycotics (Sigma).

Techniques: Phospho-proteomics, Concentration Assay, Control

Journal: Free radical biology & medicine

Article Title: Nitroglycerin drives endothelial nitric oxide synthase activation via the phosphatidylinositol 3-kinase/protein kinase B pathway

doi: 10.1016/j.freeradbiomed.2011.09.020

Figure Lengend Snippet: Time-dependent activation of Akt and eNOS paralleling PTEN phosphorylation by GTN. Representative Western blots showing PTEN phosphorylation (Ser 380), eNOS phosphorylation (Ser 1177 or 1179), and Akt phosphorylation (Ser 473) in (A) BAEC and (B) HMEC treated with vehicle or 500 nM GTN for the indicated amounts of time. Vehicle was added for 10 min in (A) and 15 min in (B). Results show rapid and sustained eNOS phosphorylation at the activation site Ser 1177, Akt phosphorylation at the activation site Ser 473, and PTEN phosphorylation at the inhibitory site Ser 380 by 500 nM GTN. Band intensities for phosphorylated eNOS in BAEC and for the experiments performed with HMEC were quantified using ImageJ and the values are reported as density units relative to control; *P<0.05, **P<0.01.

Article Snippet: Human and bovine aortic endothelial cells (HAEC or BAEC; Allcells, Emeryville, CA, USA), human microvascular endothelial cells (HMEC), and primary mouse endothelial cells (MEC) were cultured at 37 °C, 5% CO 2 in corresponding endothelial medium with supplements including growth factors, 10% FBS, and 1% antibiotics–antimycotics (Sigma).

Techniques: Activation Assay, Phospho-proteomics, Western Blot, Control

Journal: Free radical biology & medicine

Article Title: Nitroglycerin drives endothelial nitric oxide synthase activation via the phosphatidylinositol 3-kinase/protein kinase B pathway

doi: 10.1016/j.freeradbiomed.2011.09.020

Figure Lengend Snippet: Concentration-dependent Akt activation by GTN. (A) BAEC cells were treated with vehicle control (30% ethanol, 30% propylene glycol, 40% water) or the indicated GTN concentrations for 3 min before harvest. (B) Band densities were measured using ImageJ software. Results were quantified as relative Akt activation compared with control; *P<0.05 by Student's t test.

Article Snippet: Human and bovine aortic endothelial cells (HAEC or BAEC; Allcells, Emeryville, CA, USA), human microvascular endothelial cells (HMEC), and primary mouse endothelial cells (MEC) were cultured at 37 °C, 5% CO 2 in corresponding endothelial medium with supplements including growth factors, 10% FBS, and 1% antibiotics–antimycotics (Sigma).

Techniques: Concentration Assay, Activation Assay, Control, Software