Tanaka Saori, Hosogi Shigekuni, Sawabe Yukinori, Shimamoto Chikao, Matsumura Hitoshi, Inui Toshio, Marunaka Yoshinori, Nakahari Takashi
Laboratory of Pharmacotherapy, Osaka University of Pharmaceutical Sciences.
Biomed Res. 2016;37(3):167-78. doi: 10.2220/biomedres.37.167.
A PPARα (peroxisome proliferation activation receptor α) agonist (GW7647) activates nitric oxide synthase 1 (NOS1) to produce NO leading to cGMP accumulation in antral mucous cells. In this study, we examined how PPARα activates NOS1. The NO production stimulated by GW7647 was suppressed by inhibitors of PI3K (wortmannin) and Akt (AKT 1/2 Kinase Inhibitor, AKT-inh), although it was also suppressed by the inhibitors of PPARα (GW6471) and NOS1 (N-PLA). GW7647 enhanced the ACh (acetylcholine)-stimulated exocytosis (Ca(2+)-regulated exocytosis) mediated via NO, which was abolished by GW6471, N-PLA, wortmannin, and AKT-inh. The Western blotting revealed that GW7647 phosphorylates NOS1 via phosphorylation of PI3K/Akt in antral mucous cells. The immunofluorescence examinations demonstrated that PPARα existing with NOS1 co-localizes with PI3K and Akt in the cytoplasm of antral mucous cells. ACh alone and AACOCF3, an analogue of arachidonic acid (AA), induced the NOS1 phosphorylation via PI3K/Akt to produce NO, which was inhibited by GW6471. Since AA is a natural ligand for PPARα, ACh stimulates PPARα probably via AA. In conclusion, PPARα activates NOS1 via PI3K/Akt phosphorylation to produce NO in antral mucous cells during ACh stimulation.
过氧化物酶体增殖激活受体α（PPARα）激动剂（GW7647）激活一氧化氮合酶1（NOS1）以产生NO，导致胃窦黏液细胞中cGMP积累。在本研究中，我们研究了PPARα如何激活NOS1。GW7647刺激产生的NO受到PI3K抑制剂（渥曼青霉素）和Akt抑制剂（AKT 1/2激酶抑制剂，AKT-inh）的抑制，尽管它也受到PPARα抑制剂（GW6471）和NOS1抑制剂（N-PLA）的抑制。GW7647增强了通过NO介导的乙酰胆碱（ACh）刺激的胞吐作用（Ca(2+)调节的胞吐作用），而GW6471、N-PLA、渥曼青霉素和AKT-inh可消除这种作用。蛋白质免疫印迹显示，GW7647通过胃窦黏液细胞中PI3K/Akt的磷酸化使NOS1磷酸化。免疫荧光检查表明，与NOS1共存的PPARα在胃窦黏液细胞的细胞质中与PI3K和Akt共定位。单独的ACh和花生四烯酸（AA）类似物AACOCF3通过PI3K/Akt诱导NOS1磷酸化以产生NO，这被GW6471抑制。由于AA是PPARα的天然配体，ACh可能通过AA刺激PPARα。总之，在ACh刺激期间，PPARα通过PI3K/Akt磷酸化激活NOS1，在胃窦黏液细胞中产生NO。
