Orexin-A stimulates 3β-hydroxysteroid dehydrogenase expression and cortisol production in H295R human adrenocortical cells through the AKT pathway
Orexin-A stimulates 3β-hydroxysteroid dehydrogenase expression and cortisol production in H295R human adrenocortical cells through the AKT pathway
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DOI：
10.3892/ijmm.2014.1959
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发表时间：
2014-12-01
影响因子：
5.4
通讯作者：
Guo, Lei
中科院分区：
文献类型：
--
作者：
Chang, Xiaocen;Zhao, Yuyan;Guo, Lei
Orexin-A是一种调节肽，参与食物摄入、睡眠和觉醒的调节，具有多种内分泌和代谢功能。它通过刺激两种G蛋白偶联受体，食欲素受体1型（OX 1受体）和食欲素受体2型（OX 2受体），协调不同的中枢和外周过程。在本研究中，人肾上腺皮质细胞（NCI-H295 R细胞）与不同浓度的食欲素-A（10（-10）至10（-6）M）在体外孵育，并测定细胞中OX 1受体的mRNA和蛋白表达。此外，然后用或不用OX、受体特异性拮抗剂（SB 334867）、AKT拮抗剂（PF-04691502）或两者的组合处理用10-6 M食欲素-A处理的NCI-H295 R细胞。随后，分析细胞增殖、培养基中皮质醇含量以及3 β-羟基类固醇脱氢酶（3 β-HSD）的mRNA和蛋白表达。还在NCI-H295 R细胞中测定了AKT信号传导途径的活性。我们观察到OX 1受体mRNA和蛋白表达的增加是食欲素-A浓度依赖性的，10（-6）M食欲素-A发挥最强的作用。Orexin-A促进NCI-H295 R细胞增殖和皮质醇产生，并增加3 β-HSD的mRNA和蛋白表达;然而，这些作用被OX、受体拮抗剂、AKT拮抗剂以及两者的组合部分阻断。此外，食欲素-A显着增加AKT的磷酸化，与总AKT蛋白的水平保持不变。在PF-04691502（10（-6）M）、SB 334867（10（-6）M）和两者联合存在下，该效应被阻断。总之，我们的数据表明orexin-A对人肾上腺皮质细胞的存活和功能的影响是通过AKT信号通路介导的。
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 OX, receptor was determined in the cells. In addition, NCI-H295R cells treated with 10-6 M orexin-A were then treated with or without OX, 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 3 beta-hydroxysteroid dehydrogenase (3 beta-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 3 beta-HSD in the NCI-H295R cells; however, these effects were partly blocked by the OX, 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.