title: Orexin-A stimulates 3β-hydroxysteroid dehydrogenase expression and cortisol production in H295R human adrenocortical cells through the AKT pathway.
abstract: Orexin-A is a regulatory peptide involved in the regulation of food intake, sleep-wakefulness, and it has various endocrine and metabolic functions. It orchestrates diverse central and peripheral processes through the stimulation of two G-protein coupled receptors, orexin receptor type 1 (OX1 receptor) and orexin receptor type 2 (OX2 receptor). In this study, human adrenocortical cells (NCI-H295R cells) were incubated with various concentrations of orexin-A (10-10 to 10-6 M) in vitro, and the mRNA and protein expression of OX1 receptor was determined in the cells. In addition, NCI-H295R cells treated with 10-6 M orexin-A were then treated with or without OX1 receptor specific antagonist (SB334867), AKT antagonist (PF-04691502), or a combination of both. Subsequently, cell proliferation, the cortisol content in the medium and the mRNA and protein expression expression of 3β-hydroxysteroid dehydrogenase (3β-HSD) were analyzed. The activity of the AKT signaling pathway was also determined in the NCI-H295R cells. We observed that the increase in the mRNA and protein expression of OX1 receptor was orexin-A concentration-dependent, with 10-6 M orexin-A exerting the most potent effect. Orexin-A enhanced cell proliferation and cortisol production, and increased the mRNA and protein expression of 3β-HSD in the NCI-H295R cells; however, these effects were partly blocked by the OX1 receptor antagonist, the AKT antagonist and the combination of both. Furthermore, orexin-A significantly increased the phosphorylation of AKT, with the levels of total AKT protein remaining unaltered. This effect was blocked in the presence of PF-04691502 (10-6 M), SB334867 (10-6 M) and the combination of both. On the whole, our data demonstrate that the effects of orexin-A on the survival and function of human adrenocortical cells are mediated through the AKT signaling pathway.
authors.affiliation: Departments of Endocrinology and Orthopedic Surgery, The First Affiliated Hospital, China Medical University, Shenyang, Liaoning 110001, P.R. China.
authors.forename: Xiaocen
authors.initials: X
authors.lastname: Chang
authors.affiliation: Departments of Endocrinology and Orthopedic Surgery, The First Affiliated Hospital, China Medical University, Shenyang, Liaoning 110001, P.R. China.
authors.forename: Yuyan
authors.initials: Y
authors.lastname: Zhao
authors.affiliation: Departments of Endocrinology and Orthopedic Surgery, The First Affiliated Hospital, China Medical University, Shenyang, Liaoning 110001, P.R. China.
authors.forename: Shujing
authors.initials: S
authors.lastname: Ju
authors.affiliation: Departments of Endocrinology and Orthopedic Surgery, The First Affiliated Hospital, China Medical University, Shenyang, Liaoning 110001, P.R. China.
authors.forename: Lei
authors.initials: L
authors.lastname: Guo
mesh_terms: D015096:3-Hydroxysteroid Dehydrogenases / Q000235:genetics / Q000378:metabolism*; D018268:Adrenocortical Carcinoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D001583:Benzoxazoles / Q000494:pharmacology; D015153:Blotting, Western; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D006854:Hydrocortisone / Q000096:biosynthesis / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins / Q000494:pharmacology*; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D009287:Naphthyridines; D009479:Neuropeptides / Q000494:pharmacology*; D000068796:Orexin Receptor Antagonists; D064446:Orexin Receptors / Q000235:genetics / Q000378:metabolism; D000068797:Orexins; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011728:Pyridones / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D014508:Urea / Q000031:analogs & derivatives / Q000494:pharmacology
chemical_list: C420062:1-(2-methylbenzoxazol-6-yl)-3-(1,5)naphthyridin-4-yl urea; C570662:2-amino-8-(4-(2-hydroxyethoxy)cyclohexyl)-6-(6-methoxypyridin-3-yl)-4-methylpyrido(2,3-d)pyrimidin-7(8H)-one; D001583:Benzoxazoles; C000596229:HCRT protein, human; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes; D009287:Naphthyridines; D009479:Neuropeptides; D000068796:Orexin Receptor Antagonists; D064446:Orexin Receptors; D000068797:Orexins; D011728:Pyridones; D011743:Pyrimidines; D014508:Urea; D015096:3-Hydroxysteroid Dehydrogenases; D051057:Proto-Oncogene Proteins c-akt; D006854:Hydrocortisone