Journal: Physiological reports
Article Title: β3 adrenergic agonism: A novel pathway which improves right ventricular-pulmonary arterial hemodynamics in pulmonary arterial hypertension.
doi: 10.14814/phy2.15549
Figure Lengend Snippet: FIGURE 8 Effects of treatments on expression and glutathionylation of endothelial nitric oxide synthase and soluble guanylyl cyclase subunits in the lungs of hypoxic mice. (a) Expression of endothelial nitric oxide synthase. Immunoblots of endothelial nitric oxide synthase (eNOS) in the lungs are shown. β-Actin was used as the loading control. Mean densitometries for eNOS normalized to β-actin are shown in the panel. Expression of eNOS was higher in the lungs of hypoxic mice compared with the normoxic control mice. N = 5–6 mice. *p = 0.008 compared with the normoxic controls on Kruskal–Wallis test. (b) Glutathionylation of endothelial nitric oxide synthase. Immunoblots (IB) of eNOS and GSH performed on eNOS immunoprecipitate (IP) from the lungs of mice are shown. IP of a nonspecific IgG with IB for GSH was used as a negative control. Mean densitometries for GSH IB normalized to eNOS IB are shown in the panel. Glutathionylation of eNOS (eNOS-GSH) was increased in the lungs of hypoxic mice treated with vehicle compared with the normoxic controls. Treatment with CL316243 reduced eNOS-GSH compared with the hypoxic control mice. N = 5–6 mice. *p = 0.009 compared with the normoxic controls and †p < 0.02 compared with the hypoxic controls on Kruskal–Wallis test. (c) Expression of soluble guanylyl cyclase (sGC) subunits. Immunoblots of sGC α1 and β1 subunits are shown. β-Actin was used as the loading control. Mean densitometries for sGC α1 and β1 subunits normalized to β-actin are shown in the panels. Expression of sGC α1, not β1 subunit, was reduced in the lungs of hypoxic mice compared with the normoxic controls. All treatments increased the expression of sGC α1 compared with the hypoxic controls. N = 5–7 mice. *p = 0.03 compared with the normoxic controls, †p < 0.03 compared with the hypoxic controls on univariate ANOVA.
Article Snippet: Briefly, the tissue homogenate (1 mg protein) was incubated with 2.5 μg anti- eNOS antibody (AF950, R&D) or with 2.5 μg IgG control (02- 6202, Sigma- Aldrich) for 1 h at 4°C and then with protein A/G- Plus agarose beads (SCZ- SC- 2003, Santa Cruz Biotechnology Inc.) for 1 h at 4°C.
Techniques: Expressing, Western Blot, Control, Negative Control
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