Journal: American Journal of Physiology - Lung Cellular and Molecular Physiology

Article Title: The inflammatory role of dysregulated IRS2 in pulmonary vascular remodeling under hypoxic conditions

doi: 10.1152/ajplung.00068.2020

Figure Lengend Snippet: Expression levels of Akt-FOXO1 pathway proteins are elevated in the lungs of IRS2+/− mice. Immunoblotting was used to analyze downstream signaling of IRS2 in the lungs of IRS2+/+ and IRS2+/- mice after 4 days of hypoxia stimulation. Expression levels of total (T) and phosphorylated (P) Akt, ERK, S6, FOXO1 (Thr24 and Ser256), and HIF-1α were assessed. Data are expressed as means ± SD. *P < 0.05, **P < 0.01, #P < 0.05, ##P < 0.01 versus indicated group (n = 3–4). Immunoblots shown are from one of three independent experiments conducted. IRS2, insulin receptor substrate 2; FOXO1, Forkhead box O1.

Article Snippet: Membranes were incubated with primary antibodies against Akt (No. 9272), phospho (p)-Akt (Ser473, No. 9271), p44/43 MAPK (ERK, No. 4695), p-ERK (No. 4370), ribosomal protein S6 (S6, No. 2217), p-S6 (No. 4858), FOXO1 (No. 2880), p-FOXO1 (Thr24 and Ser256, No. 9464 and No. 9461) (all from Cell Signaling Technology, Danvers, MA), mouse HIF-1α (No. NB100-105, Novus Biologicals, Littleton, CO), and β-actin (Santa Cruz Biotechnology, Dallas, TX).

Techniques: Expressing, Western Blot

Journal: American Journal of Physiology - Lung Cellular and Molecular Physiology

Article Title: The inflammatory role of dysregulated IRS2 in pulmonary vascular remodeling under hypoxic conditions

doi: 10.1152/ajplung.00068.2020

Figure Lengend Snippet: Expression levels of Akt-FOXO1 pathway proteins are elevated in the lungs of IRS2+/− mice. Immunoblotting was used to analyze downstream signaling of IRS2 in the lungs of IRS2+/+ and IRS2+/- mice after 4 days of hypoxia stimulation. Expression levels of total (T) and phosphorylated (P) Akt, ERK, S6, FOXO1 (Thr24 and Ser256), and HIF-1α were assessed. Data are expressed as means ± SD. *P < 0.05, **P < 0.01, #P < 0.05, ##P < 0.01 versus indicated group (n = 3–4). Immunoblots shown are from one of three independent experiments conducted. IRS2, insulin receptor substrate 2; FOXO1, Forkhead box O1.

Article Snippet: Membranes were incubated with primary antibodies against Akt (No. 9272), phospho (p)-Akt (Ser473, No. 9271), p44/43 MAPK (ERK, No. 4695), p-ERK (No. 4370), ribosomal protein S6 (S6, No. 2217), p-S6 (No. 4858), FOXO1 (No. 2880), p-FOXO1 (Thr24 and Ser256, No. 9464 and No. 9461) (all from Cell Signaling Technology, Danvers, MA), mouse HIF-1α (No. NB100-105, Novus Biologicals, Littleton, CO), and β-actin (Santa Cruz Biotechnology, Dallas, TX).

Techniques: Expressing, Western Blot

Journal: Redox Biology

Article Title: Dietary salt initiates redox signaling between endothelium and vascular smooth muscle through NADPH oxidase 4

doi: 10.1016/j.redox.2022.102296

Figure Lengend Snippet: Western analysis of aortic content of Runx2, osteopontin, and osteocalcin of endothelial Nox4 −/− and littermate mice with changes in dietary salt content. A , During ingestion of the 0.3% NaCl diet, Runx2 levels in the aorta did not differ between the strains. When fed a 4.0% NaCl diet, Runx2 increased in the aortas of littermate mice, when compared with the endothelial Nox4 −/− mice. Two-way ANOVA showed significant effects of Nox4 ( P < 0.05) and dietary salt ( P < 0.05) on catalase activity and Runx2 and an interaction effect ( P < 0.05) between Nox4 and dietary salt (n = 4 mice in each group; ns, not significant; * P < 0.05; Two-way ANOVA). B , When fed a 4.0% NaCl diet, osteopontin increased in the aortas of littermate mice, when compared with the endothelial Nox4 −/− mice. Two-way ANOVA showed significant effects of Nox4 ( P < 0.05) and dietary salt ( P < 0.05) on osteopontin and an interaction effect ( P < 0.05) between Nox4 and dietary salt (n = 4 mice in each group; ns, not significant; * P < 0.05; Two-way ANOVA). C , When fed a 4.0% NaCl diet, osteocalcin increased in the aortas of littermate mice, when compared with the endothelial Nox4 −/− mice. Two-way ANOVA showed significant effects of Nox4 ( P < 0.05) and dietary salt ( P < 0.05) on osteopontin, but no interaction effect ( P = 0.5943) between Nox4 and dietary salt (n = 4 mice in each group; * P < 0.05; Two-way ANOVA).

Article Snippet: Experiments also used commercial antibodies directed against FoxO1 (Cat# NB100-2312, Novus Biologicals, Centennial CO); p-FoxO1(ser256) (Cat# NB100-81927, Novus Biologicals), Runx2 (Cat# ab23981, Abcam Inc., Cambridge, MA), Runx2 (C-12) (Cat# sc-390715, Santa Cruz Biotechnology, Inc., Dallas, TX), PP2A (clone 1D6, Cat# 05–421, Millipore, Temecula, CA), osteopontin (Cat# ab283656, Abcam Inc., Cambridge, MA), osteocalcin (Cat# ab133612, Abcam Inc., Cambridge, MA), and GAPDH (Cat# Ab8245; Abcam Inc., Cambridge, MA), which served as a loading normalization control.

Techniques: Western Blot, Activity Assay