Vargas Vladimir E, Myers Dean A, Kaushal Kanchan M, Ducsay Charles A
1 Center for Perinatal Biology, Loma Linda University School of Medicine, Loma Linda, CA, USA.
2 Department of Obstetrics and Gynecology, University of Oklahoma Health Sciences Center, Oklahoma City, OK, USA.
Reprod Sci. 2018 Feb;25(2):230-238. doi: 10.1177/1933719117707056. Epub 2017 May 3.
We previously demonstrated decreased expression of key genes regulating cortisol biosynthesis in long-term hypoxic (LTH) sheep fetal adrenals compared to controls. We also showed that inhibition of the extracellular signal-regulated kinases (ERKs) with the mitogen-activated protein kinase (MEK)/ERK inhibitor UO126 limited adrenocorticotropic (ACTH)-induced cortisol production in ovine fetal adrenocortical cells (FACs), suggesting a role for ERKs in cortisol synthesis. This study was designed to determine whether the previously observed decrease in LTH cytochrome P45011A1/cytochrome P450c17 (CYP11A1/CYP17) in adrenal glands was maintained in vitro, and whether ACTH alone with or without UO126 treatment had altered the expression of CYP11A1, CYP17, and steroidogenic acute regulatory protein (StAR) in control versus LTH FACs. Ewes were maintained at high altitude (3820 m) from ∼40 days of gestation (dG). At 138 to 141 dG, fetal adrenal glands were collected from LTH (n = 5) and age-matched normoxic controls (n = 6). Fetal adrenocortical cells were challenged with ACTH (10 M) with or without UO126 (10 µM) for 18 hours. Media samples were collected for cortisol analysis and messenger RNA (mRNA) for CYP11A1, CYP17, and StAR was quantified by quantitative real-time polymerase chain reaction. Cortisol was higher in the LTH versus control ( P < .05). StAR mRNA was decreased in LTH versus control ( P < .05). U0126 alone had no effect on mRNA in either group. UO126 prevented the increase in CYP11A1 and CYP17 in control FACs. Basal CYP11A1 and CYP17 were not different in LTH versus control. ACTH increased CYP11A1 and CYP17 only in control FACs ( P < .05). U1026 attenuated the ACTH response indicative of a role for ERK in CYP11A1 and CYP17 expression. ACTH may require additional factors in FACs to fully regulate StAR expression.
我们之前证明，与对照组相比，长期缺氧（LTH）绵羊胎儿肾上腺中调节皮质醇生物合成的关键基因表达降低。我们还表明，用丝裂原活化蛋白激酶（MEK）/细胞外信号调节激酶（ERK）抑制剂UO126抑制ERK可限制促肾上腺皮质激素（ACTH）诱导的绵羊胎儿肾上腺皮质细胞（FACs）中皮质醇的产生，这表明ERK在皮质醇合成中发挥作用。本研究旨在确定之前观察到的LTH肾上腺中细胞色素P45011A1/细胞色素P450c17（CYP11A1/CYP17）的降低在体外是否持续存在，以及单独使用ACTH或联合UO126处理是否改变了对照组与LTH组FACs中CYP11A1、CYP17和类固醇生成急性调节蛋白（StAR）的表达。从妊娠约40天（dG）开始，将母羊饲养在高海拔（3820米）环境中。在138至141 dG时，从LTH组（n = 5）和年龄匹配的常氧对照组（n = 6）收集胎儿肾上腺。用ACTH（10 M）或联合UO126（10 µM）刺激胎儿肾上腺皮质细胞18小时。收集培养基样本进行皮质醇分析，并通过定量实时聚合酶链反应对CYP11A1、CYP17和StAR的信使核糖核酸（mRNA）进行定量。与对照组相比，LTH组的皮质醇水平更高（P <.05）。与对照组相比，LTH组的StAR mRNA降低（P <.05）。单独使用U0126对两组的mRNA均无影响。UO126可阻止对照组FACs中CYP11A1和CYP17的增加。LTH组与对照组的基础CYP11A1和CYP17无差异。ACTH仅在对照组FACs中增加CYP11A1和CYP17（P <.05）。U1026减弱了ACTH反应，表明ERK在CYP11A1和CYP17表达中发挥作用。ACTH可能需要FACs中的其他因素来完全调节StAR表达。
