Is the Fetal Lung a Mineralocorticoid Receptor Target Organ? Induction of Cortisol-Regulated Genes in the Ovine Fetal Lung, Kidney and Small Intestine

Abstract

Background: Lung, kidney and small intestine are involved in fetal volume regulation and amniotic fluid secretion and play a pivotal role in the transition from intrauterine to extrauterine life. Objective: This study was performed to determine the ontogeny of mineralocorticoid receptors (MR) and glucocorticoid receptors (GR), and of MR- and GR-regulated genes and proteins, serum and glucocorticoid-induced kinase (Sgk-1), epithelial sodium channel (ENaCα), and Na,K-ATPase α1. Methods: Lung, renal cortex and medulla, and small intestine were collected from fetuses at 80, 100, 120, 130 and 145 days’ gestation and from day 1 and 7 neonatal lambs. Real-time PCR was performed to determine mRNA concentration for MR, GR, the 11β-hydroxysteroid dehydrogenases (11β-HSD1 and 2), Sgk-1, ENaCα, and Na,K-ATPase α1. Protein expression of ENaCα and Na,K-ATPase α1 in whole cell and membrane fractions was determined by immunoblotting. Results: Expression of corticosteroid-induced genes in renal cortex increases at term; in small intestine the induction occurs postnatally. In contrast, in lung expression of MR and GR mRNAs were greater at 100 days to term than postnatally and 11β-HSD1 peaked at 145 days; the corticosteroid-induced genes also increased prenatally: Sgk-1 and ENaCα increased by 120 days, peaking at 145 days, and Na,K-ATPase α1 was greatest at 130 days. Conclusions: The expression of high levels of MR and 11β-HSD1 in preterm fetal lung suggest low endogenous fetal cortisol may exert actions at the high affinity MR in vivo, leading to increases in expression of sodium channels important in the regulation of lung liquid secretion and reabsorption.

Expression of ENaC subunits, chloride channels, and aquaporins in ovine fetal lung: ontogeny of expression and effects of altered fetal cortisol concentrations

“…This range of fetal ages was chosen to include the period of development of the fetal lung from the end of the canalicular phase through the alveolar phase to the time of birth (1). Data regarding lung, small intestine, and renal medulla and cortex expression of MR, GR, 11␤ HSD1 and 2, Na ϩ ,K ϩ -ATPase␣1, ENaC␣, and sgk1 from this set of animals have also been previously reported (18). Four additional ewes and their fetuses were also killed while in labor (142-148 days gestation) (labor, n ϭ 5 fetuses), and newborn lambs (n ϭ 5) were also killed in the first 48 h after delivery to determine the effects of labor and the associated increase in cortisol, and the changes in the immediate postpartum period after the high levels of cortisol have subsided and lung MR expression is decreased.…”

“…CFTR expression decreases and AQP expression increases in late gestation fetal sheep (4,24). We have previously found distinctive ontogenetic patterns of expression of ENaC␣ and Na ϩ ,K ϩ -ATPase␣1 subunit mRNAs and protein in the ovine fetal lung compared with the ovine fetal kidney or small intestine (18); these previous results indicate earlier increases in expression of these sodium transporters in lung than in kidney or small intestine. In contrast, in the fetal rat, expression of ENaC subunits and AQP1 in both lung and kidney occurs in the last days of fetal life or postnatally (21,40,41,45,46).…”

“…The fetal lung has low expression and activity of the cortisol inactivating enzyme 11␤ hydroxysteroid dehydrogenase activity (11␤ HSD2) (18,50), allowing cortisol binding to both receptors in the fetal lung. MR expression is considerably greater in the distal lung of the fetus than in the newborn or adult (19), suggesting that MR may play a role in fluid and salt reabsorption in the lung at the time of birth.…”

Jaagsiekte sheep retrovirus pseudotyped lentiviral vector-mediated gene transfer to fetal ovine lung

“…52 Membranes were blocked with 5% fat-free milk in TBS (1 h, room temperature), incubated overnight at 4 1C with primary HYAL2 antibody (1:1000; Abcam, Cambridge, MA, USA), rinsed in TBS-T and incubated at room temperature with secondary donkey anti-rabbit horseradish peroxidaseconjugated antibody (1:10 000; GE Healthcare, Piscataway, NJ, USA) for 2 h. Immunoblots were developed using a chemiluminescent substrate kit (SuperSignal West Pico, Pierce, Rockford, IL, USA) and the relative densities of HYAL2 bands were calculated (Scion Image, Bethesda, MD, USA; http://www.scioncorp.com). To account for minor differences in total protein loading, membranes were stripped and re-probed for mouse anti-actin antibody (Abcam).…”

Increased lung prolyl hydroxylase and decreased glucocorticoid receptor are related to decreased surfactant protein in the growth-restricted sheep fetus

“…SPs do not have a GRE, and thus GCs indirectly increase SP expression through activation of the thyroid transcription factor (TTF)-1 and by other transcription factors and cofactors such as GATA-6 (28,54,58). GC action within the fetal lung is also mediated through the action of the mineralocorticoid receptor (MR), which is known to play a role in normal lung development (44). Thus an increase in plasma cortisol concentration would be expected to increase SP expression in the lung, but, despite higher plasma cortisol concentrations in late gestation, the IUGR fetus in the carunclectomy model has reduced SP expression (66).…”