Journal: Biomedical research (Tokyo, Japan)

Article Title: PPARα induced NOS1 phosphorylation via PI3K/Akt in guinea pig antral mucous cells: NO-enhancement in Ca(2+)-regulated exocytosis.

doi: 10.2220/biomedres.37.167

Figure Lengend Snippet: Fig. 3 P-PI3K, P-Akt (Ser473), and P-NOS1 of antral mucosa stimulated by agonists. Agonists used were ACh (1 μM), GW7647 (50 nM), GW7647 plus ACh, AACOCF3 (1 μM), and AACOCF3 plus ACh. We used DMSO as the vehicle control. Panels A & B shows representative Western blotting for P-PI3K, P-Akt (Ser473), P-NOS1, and β-actin in antral mucosae. A: Control experiments. The addition of each agonist enhanced the bands of P-PI3K, P-Akt (Ser473), and P-NOS1. B: GW6471 (5 μM) experiments. The agonists did not enhance three bands in the presence of GW6471. C, D, E: The relative changes in the band densities of P-PI3K (C), P-Akt (Ser473) (D), and P-NOS1 (E) normalized by the corresponding β-actin bands. The stimulation with agonists significantly enhanced the band densities of P-PI3K, P-Akt (Ser473), and P-NOS1 (marked as ■). The enhancement was abolished by the prior treatment of GW6471 (marked as □). *Significantly different from the cor- responding values (P < 0.05).

Article Snippet: The antibodies used were an antiNOS1 antibody (LS-B3272; LifeSpan BioSciences, WA, USA), an anti-PI3K antibody (No. #4257; Cell Signaling Technology, MA, USA), an anti-Akt antibody (No. #9272; Cell Signaling), an anti-phosphoNOS1 antibody (ADI-KAP-NO032; Enzo Life Sciences, NY, USA), an anti-phospho-PI3K antibody (GTX50858; Gene Tex, CA, USA), and an antiphospho-Akt (Ser473) antibody (No. #9271; Cell Signaling Technology).

Techniques: Control, Western Blot

Journal: Biomedical research (Tokyo, Japan)

Article Title: PPARα induced NOS1 phosphorylation via PI3K/Akt in guinea pig antral mucous cells: NO-enhancement in Ca(2+)-regulated exocytosis.

doi: 10.2220/biomedres.37.167

Figure Lengend Snippet: Fig. 4 Effects of wortmannin and AKT-inh on P-PI3K, P-Akt (Ser473), and P-NOS1. A, B, C: Wortmannin (50 nM) experi- ments. (A) Representative Western blotting for P-Akt (Ser473) and P-NOS1. The stimulation with agonists did not enhance the band of P-Akt (Ser473) and P-NOS1. (B) The relative changes in the band densities of Akt (Ser473) normalized by the corresponding β-actin bands. (C) The relative changes in the band densities of NOS1 normalized by the corresponding β-actin bands. The agonists induced no significant changes in the band densities of P-Akt (Ser473) and P-NOS1 in the presence of wortmannin. D, E, F: AKT-inh (100 nM) experiments. (D) Representative Western blotting for P-PI3K and P- NOS1. The stimulation with agonists enhanced the band of P-PI3K, but not those of P-NOS1. (E) The relative changes in the band densities of P-PI3K normalized by the corresponding β-actin bands. The agonists significantly increased the band densities of P-PI3K. (F) The relative changes in the band densities of P-NOS1 normalized by the corresponding β-actin bands. The agonists did not change the band densities of P-NOS1. Thus, AKT-inh inhibited the increases in the P-NOS1 density, but did not those in P-PI3K. *Significantly different from the corresponding values (P < 0.05).

Article Snippet: The antibodies used were an antiNOS1 antibody (LS-B3272; LifeSpan BioSciences, WA, USA), an anti-PI3K antibody (No. #4257; Cell Signaling Technology, MA, USA), an anti-Akt antibody (No. #9272; Cell Signaling), an anti-phosphoNOS1 antibody (ADI-KAP-NO032; Enzo Life Sciences, NY, USA), an anti-phospho-PI3K antibody (GTX50858; Gene Tex, CA, USA), and an antiphospho-Akt (Ser473) antibody (No. #9271; Cell Signaling Technology).

Techniques: Western Blot