Adrenocorticotropic Hormone and PI3K/Akt Inhibition Reduce eNOS Phosphorylation and Increase Cortisol Biosynthesis in Long-Term Hypoxic Ovine Fetal Adrenal Cortical Cells

Abstract

This study was designed to determine the role of the MEK/ERK1/2 and PI3K/Akt pathways in cortisol production and endothelial nitric oxide synthase (eNOS) phosphorylation (peNOS) in the ovine fetal adrenal in response to long-term hypoxia (LTH). Pregnant ewes were maintained at high altitude (3820 m) for the last 100 days of gestation (dGa). At 138 to 142 dGa, fetal adrenal cortical cells (FACs) were collected from LTH and age-matched normoxic fetuses. Cortisol production and peNOS were measured in response to pretreatment with the MEK/ERK1/2 pathway inhibitor UO126 (UO) and adrenocorticotropic hormone (ACTH) stimulation. UO126 reduced ACTH-stimulated cortisol in both normoxic and LTH FACs. UO126 alone or in combination with ACTH reduced peNOS in the normoxic group, while ACTH alone or ACTH + UO inhibited peNOS in LTH FACs. Additionally, cortisol was measured in response to pretreatment with UO and treatment with 22R-hydroxycholesterol (22R-OHC) or water-soluble cholesterol (WSC) with and without ACTH stimulation. UO126 had no effect on 22R-OHC-treated cells, but reduced cortisol in cells treated with WSC and/or ACTH. Cortisol and peNOS were also measured in response to pretreatment with PI3K/Akt pathway inhibitor Wortmannin (WT) and ACTH stimulation. Wortmannin further increased cortisol under ACTH-stimulated conditions and, like ACTH, reduced peNOS in LTH but not normoxic FACs. Together, these data suggest that in LTH FACs MEK/ERK1/2 does not regulate peNOS but that UO acts downstream from eNOS, possibly at cholesterol transport, to affect cortisol production in LTH FACs, while the PI3K/Akt pathway, along with ACTH, regulates peNOS and plays a role in the fetal adaptation to LTH in FACs.

Fetal endocrine and metabolic adaptations to hypoxia: the role of the hypothalamic-pituitary-adrenal axis

“…Subsequent studies by Monau et al (110) showed that NO reduced ACTH-mediated cortisol production in LTH fetal adrenocortical cells (FACs) in vitro, whereas inhibition of NOS activity increased cortisol production in LTH cells, with no effect on normoxic cells. Furthermore, ACTH reduced eNOS activation via phosphorylation in LTH FACs (123) and NO-dependent inhibition of ACTH-induced cortisol production, which further supports the role of NO in regulating cortisol production in the LTH fetal adrenal (171). This may be possible by NO competing with the oxygen-binding site of CYP11A1 and CYP17 (74,170), disrupting the heme-oxygen complex attack by the enzyme on the steroid substrate.…”

Identify and validate GPC4 and VCAN as hub genes in primary bilateral macronodular adrenal hyperplasiaby WGCNA and DEG

“…In addition, cortisol may increase Bovine endometrial epithelial cells(BEECs) proliferation by raising the production of certain growth factors and activating the Wnt/-catenin and PI3K/AKT signalling pathways (30). PI3K/Akt inhibition decreases eNOS phosphorylation and boosts cortisol production in chronically hypoxic foetal sheep adrenal cortical cells (31). Therefore, we speculate that cortisol may stimulate adrenal gland proliferation via the aforementioned pathways.PI3K/Akt pathway may promote the secretion of cortisol by adrenal cortical cells.…”

Influence of nitric oxide signaling mechanisms in cancer

“…Once phosphorylated, Akt, the p65 subunit of NF-κB, translocates to the cell nucleus where it stimulates COX-2 gene transcription. 64 However, under normal conditions it is regulated by members of the mitogen-activated protein kinase (MAPK) family. Experimental studies have shown that activation of COX-2 induces the synthesis alpha factor, IL-6, and TNF-alpha, which are molecules that promote NO synthesis in macrophages and fibroblasts.…”