Adrenal Growth Research Articles

Agouti Related Peptide (AgRP) deficient mice show accelerated compensatory adrenal growth after unilateral adrenalectomy

Agouti related peptide (AgRP) is a well known regulator of food intake. This effect is exerted on the level of the hypothalamus by acting as an antagonist on several subtypes of the melanocortin receptors (MC-R). Aside from the central nervous system the only peripheral tissue with high expression of the AgRP gene is the adrenal gland.

Evidence against a role of human airway trypsin-like protease – the human analogue of the growth-promoting rat adrenal secretory protease – in adrenal tumourigenesis

A serine protease from rat adrenal cortex was recently characterized and named adrenal secretory protease (AsP). AsP is expressed in the adrenal cortex and is capable of cleaving pro-gamma-melanocyte-stimulating hormone (1-76 N-terminus of pro-opiomelanocortin) into fragments that act as adrenal mitogens. AsP may therefore play a crucial role in adrenal growth and tumourigenesis. The aim of this study was to further characterize the human homologue of AsP and its possible role in adrenal tumourigenesis. Starting with the rat cDNA sequence of AsP we detected high homology to the catalytic C-terminus of the human airway trypsin-like protease (HAT). Further analysis revealed that the HAT gene is the human homologue of a long splice variant of AsP, which we recently described as rat airway trypsin-like serine protease 1. In contrast to rodents, no short isoform of HAT was found in humans due to a stop codon in exon 6 which prevents the expression of a short isoform. While high expression of HAT mRNA was found in the trachea and in the gastrointestinal tract, expression in the adrenal was only very weak. RT-PCR and real-time PCR analysis revealed a complex tissue expression pattern of HAT, indicating a role for this protease in multiple tissues. We further investigated HAT expression in five normal adrenal glands, 15 adrenocortical adenomas (five hormonally inactive adenomas, five aldosterone-producing adenomas and five cortisol-producing adenomas), nine adrenocortical carcinomas, five phaeochromocytomas and two adrenal hyperplasias. Weak HAT expression was detectable in only two out of five normal adrenal glands, in one out of twenty-four adrenocortical tumours and four out of five phaeochromocytomas. However, the expression in the adrenal tissue was several orders of magnitude lower than in the trachea. In addition, we could not detect any HAT transcripts in a sample of fetal adrenal. Gene structure and tissue distribution of HAT, the human homologue of the rat adrenal secretory protease AsP, reveal major interspecies differences. The observation of very low expression levels in normal adrenal tissue and adrenocortical tumours casts doubt about a role for HAT in the physiological and pathological growth of adrenocortical cells.

Functional biology of the primate fetal adrenal gland: advances in technology provide new insight.

1. The main aim of the present review is to summarize recent experimental data from human and non-human primate models that have identified factors essential for adrenal development and other factors that may determine the regulation of the specific structural organization and function of the adrenal gland. 2. The fetal adrenal cortex has two morphologically distinct zones, with the outer definitive zone being comprised of tightly packed small cells, which appear to be steroidogenically quiescent until late gestation, and the inner fetal zone, which appears to be steroidogenically active throughout gestation. 3. In the primate fetus, growth of the adrenal gland involves hyperplasia, hypertrophy, migration and senescence. Cells appear to proliferate in the external portion of the definitive zone and then move centripetally and become non-proliferative in the fetal zone, where they acquire their steroidogenic capacities. 4. A variety of new technologies has been used to identify zonal-specific markers of the cortical zones within the developing human fetal adrenal gland. On microarray, 67 transcripts showed a minimum of a 2.5-fold difference between the fetal and adult adrenal gland. The vast majority of these genes had not been studied in relation to adrenal gland development or function. In combination with techniques such as laser capture microdissection, which has allowed the isolation of fairly pure zone-specific cell populations from the human fetal adrenal cortex, we can begin to unravel the complex interactions regulating adrenal growth and functional differentiation.

Relationship between Hormones of the Adrenal Cortex, Growth, and Puberty in Schoolchildren

Relationship between Hormones of the Adrenal Cortex, Growth, and Puberty in Schoolchildren

Differential actions of metyrapone on the fetal pituitary-adrenal axis in the sheep fetus in late gestation.

It is not clear if an increase in intra-adrenal cortisol is required to mediate the actions of adrenocorticotropic hormone (ACTH) on adrenal growth and steroidogenesis during the prepartum stimulation of the fetal pituitary-adrenal axis. We infused metyrapone, a competitive inhibitor of cortisol biosynthesis, into fetal sheep between 125 and 140 days of gestation (term = 147 +/- 3 days) and measured fetal plasma cortisol, 11-desoxycortisol, and ACTH; pituitary pro-opiomelanocortin mRNA and adrenal expression of ACTH receptor (melanocortin type 2 receptor), steroidogenic acute regulatory protein (StAR), 11beta-hydroxysteroid dehydrogenase type 2 (11betaHSD2), cytochrome P450 cholesterol side-chain cleavage (CYP11A1), cytochrome P450 17-hydroxylase (CYP17), 3beta-hydroxysteroid dehydrogenase, and cytochrome P450 21-hydroxylase mRNA; and StAR protein in the fetal adrenal gland. Plasma ACTH and 11-desoxycortisol concentrations were higher (P < 0.05), whereas plasma cortisol concentrations were not significantly different in metyrapone- compared with vehicle-infused fetuses. The ratio of plasma cortisol to ACTH concentrations was higher (P < 0.0001) between 136 and 140 days than between 120 and 135 days of gestation in both metyrapone- and vehicle-infused fetuses. The combined adrenal weight and adrenocortical thickness were greater (P < 0.001), and cell density was lower (P < 0.01), in the zona fasciculata of adrenals from the metyrapone-infused group. Adrenal StAR mRNA expression was lower (P < 0.05), whereas the levels of mature StAR protein (30 kDa) were higher (P < 0.05), in the metyrapone-infused fetuses. In addition, adrenal mRNA expression of 11betaHSD2, CYP11A1, and CYP17 were higher (P < 0.05) in the metyrapone-infused fetuses. Thus, metyrapone administration may represent a unique model that allows the investigation of dissociation of the relative actions of ACTH and cortisol on fetal adrenal steroidogenesis and growth during late gestation.

Spatio-temporal expression of the ACTH receptor during mouse adrenal development: Implication for adrenal growth and differentiation

The ACTH receptor or melanocortin 2 receptor (MC2-R) is the key receptor regulating steroid biosynthesis in the adrenal cortex. Moreover, its expression has been implicated as a marker of adrenal tumors with higher differentiation and better clinical prognosis. Thus, ACTH as the ligand of the MC2-R might serve as a differentiation factor in the context of adrenal tumorigenesis. However, the role of ACTH during adrenal development is unknown and can not be determined without detailed knowledge about the expression of its receptor in the developing adrenal gland. Thus, timed pregnancies of 129Sv mice were performed and embryos harvested at day E9.5, E11.5, E13.5, E15.5, and E17.5 and 2 weeks post partum. MC2-R expression was assessed by means of RT-PCR and in situ hybridization (ISH) and correlated with the expression of a differentiation marker (side chain cleavage enzyme, SCC) as well as a proliferation marker (BrdU incorporation) at the various time points. Interestingly, RT-PCR of whole embryo cDNA demonstrated the expression of MC2-R mRNA as early as E9.5, at a time point when the urogenital ridge forms. In contrast, expression of steroidogenic factor 1 (SF-1) was detected at the earliest at E11.5, indicating a SF-1 independent expression of MC2-R in vivo. Comparable expression patterns were found between MC2-R mRNA and SCC mRNA. BrdU, which incorporates into cells undergoing mitosis was detected in a scattered distribution throughout the adrenal glands at each time point. Double staining experiments using BrdU immunohistochemistry and MC2-R ISH and SCC ISH, respectively, revealed the majority of cells expressing SCC or MC2-R to be BrdU negative. However, some BrdU/SCC positive cells could also be detected. These preliminary results suggest that the majority of adrenal cells with a differentiated phenotype resemble non-proliferating cells during adrenal development, whereas undifferentiated cells might represent a pool of proliferating stem cells that further differentiate in cells capable of steroidogenesis.

Metyrapone Infusion Stimulates Adrenal Growth Without Activating the Cell Cycle or the IGF System in the Late Gestation Fetal Sheep

Recently, we have demonstrated that administration of metyrapone, to suppress cortisol synthesis and decrease negative feedback at the pituitary, results in an increase in circulating ACTH and adrenal growth in the late gestation sheep fetus. In these studies, we demonstrated a 2‐fold increase in adrenocortical growth using morphometric techniques. To elucidate the potential molecular mechanisms leading to the increase in adrenal growth, we examined adrenal expression of the cell cycle regulatory proteins (cyclin D1) and cyclin‐kinase inhibitory proteins (p16ink, p21Cip), and insulin‐like growth factor‐II (IGF‐II), IGF‐binding protein 2 (IGFBP‐2) and IGF‐I type 1 Receptor (IGF1R) from fetuses infused with metyrapone or vehicle for 15 days. There was a significant decrease in adrenal expression of cyclin D1 in metyrapone‐ (472.0 ± 29.7) compared with vehicle‐infused (662.7 ± 29.2) fetuses. There was no significant difference, however, in the adrenal expression of the cyclin‐kinase inhibitory proteins (p16ink or p21Cip) or in the IGF system (IGF‐II, IGFBP‐2 or IGF1R) mRNA between metyrapone‐ and vehicle‐infused. In summary, in this model of metyrapone‐activated adrenal cortical hypertrophy, growth occurs with a suppression of the rate‐limiting cell cycle protein and without activation of the IGF system.

043. Role of TGFβ in adrenal steroidogenesis before birth

During mammalian development there are periods when the fetal adrenal is either relatively refractory or increasingly sensitive to trophic stimulation. This pattern of regulation of adrenal growth and function ensures that the fetal lungs, liver, brain and kidney are exposed in a programmed temporal sequence to the genomic actions of circulating glucocorticoids. A range of studies in the rat and sheep have also demonstrated that exposure to excess glucocorticoids at inappropriate times in fetal life inhibits fetal growth and permanently reprograms the development of the cardiovascular and metabolic systems resulting in postnatal hypertension, abnormal hepatic glucose production and poor glucose tolerance In most mammalian species, there are therefore a range of mechanisms that protect the fetus from exposure to glucocorticoids of either maternal or fetal origin at inappropriate times in gestation. The factors that act to maintain periods of adrenal quiescence are not known. There is evidence that intra-adrenal transforming growth factor beta 1 (TGFβ1) is an inhibitor of adrenocortical steroidogenesis in the adult. In recent studies, we have demonstrated that expression of TGFβ1 is high in the fetal sheep adrenal at around 100�days gestation and that adrenal TGFβ1 expression then falls with increasing gestational age and is lowest immediately after birth. Following the activation of adrenal cytochrome P450 C17 (CYP17), there is an inverse relationship between adrenal TGFβ1 and CYP17 expression and TGFβ1 may therefore play a novel inhibitory role in the regulation of adrenal steroidogenesis during mid and late gestation. Whilst functional activation of the fetal adrenal is dependent on the fetal hypothalamo–pituitary axis, adrenal TGFβ1 mRNA expression is not altered by disconnection of the fetal hypothalamus and pituitary in late gestation. It therefore appears unlikely that TGFβ1 mRNA expression is regulated directly by either bioactive ACTH or cortisol in late gestation. The mechanism by which TGFβ1 expression is upregulated in mid gestation remains to be determined.

Clinical Impact of Recent Advances in the Biology of Adrenocortical Cancer

Adrenocortical carcinoma (ACC) is a rare but highly malignant tumor. Despite some progress in diagnostic and therapeutic approaches, the overall survival rate of patients with ACC remains poor. Surgery is the treatment of choice for patients with resectable tumors and for recurrent disease. However, most ACCs in adults are diagnosed with advanced disease, limiting therapeutic options. Thus, reliable clinical and molecular markers for the diagnosis of early stages of ACC and effective alternative therapies are needed. A prerequisite for the development of new diagnostic tools and therapeutic options in the management of patients with ACC is the elucidation of the molecular pathogenesis of adrenal tumorigenesis. Our understanding of molecular pathways involved in adrenocortical growth and differentiation has increased substantially over the last decades. Contributions to the elucidation of some aspects of the disruption of growth regulation of the adrenal cortex come from rare familiar syndromes associated with ACC, mouse models of adrenal tumor growth, and in vitro studies on adrenocortical tumor cell lines. In this review, we summarize the important molecular aspects of adrenal tumorigenesis and highlight some prospects for future clinical applications.

Ghrelin, an endogenous ligand for the growth hormone-secretagogue receptor, is expressed in the human adrenal cortex

Ghrelin is an endogenous ligand of the growth hormone secretagogue receptor (GHS-R), which was originally isolated from rat stomach. Ghrelin and GHS-R are also expressed in several peripheral tissues, including adrenal glands, and this prompted us to study ghrelin expression and ghrelin-binding site localization in the human adrenal cortex, and the possible effect of this peptide on corticosteroid-hormone secretion. Reverse transcription-polymerase chain reaction (RT-PCR) and radioimmune assay (RIA) showed sizeable expression of ghrelin mRNA and protein in six human adrenal cortexes. Autoradiography evidenced abundant [125I]ghrelin binding sites in the adrenal zona glomerulosa and outer zona fasciculata. However, ghrelin (10(-6) M) did not significantly affect either basal or agonist (ACTH and angiotensin-II)-stimulated early and late steps of steroid-hormone synthesis from adrenocortical slices (as measured by quantitative high pressure liquid chromatography). Since zona glomerulosa is the cambium layer involved in the growth maintenance of adrenal cortex, the present coupled RT-PCR, RIA and autoradiographic findings could suggest the involvement of ghrelin in the autocrine-paracrine regulation of human adrenal growth.

Adrenal TGFβ1 mRNA levels fall during late gestation and are not regulated by cortisol in the sheep fetus

During mammalian development there are periods when the fetal adrenal is either relatively refractory or increasingly sensitive to trophic stimulation. This pattern of regulation of adrenal growth and function ensures that the fetal lungs, liver, brain and kidney are exposed in a programmed temporal sequence to the genomic actions of circulating glucocorticoids. The factors which act to maintain periods of adrenal quiescence are not known. In the present study we have measured the level of messenger RNA (mRNA) expression of a putative inhibitor of adrenal steroidogenesis, transforming growth factor beta 1 (TGFβ1), and a key steroidogenic enzyme, cytochrome P450 17α hydroxylase (CYP17), during periods of adrenal quiescence and activation in the sheep fetus. We have also investigated the relative roles of the fetal hypothalamic-pituitary axis and cortisol in the regulation of expression of adrenal TGFβ1 and CYP17 mRNA during late gestation. Adrenal expression of TGFβ1 was greatest at around 100 days gestation, at a time when the fetal sheep adrenal is relatively refractory to trophic stimulation and there was an inverse relationship between the expression of TGFβ1 and CYP17 mRNA in the adrenal gland during the peripartum period. Whilst disconnection of the fetal hypothalamic-pituitary disconnection (HPD) axis resulted in a decrease in adrenal CYP 17 mRNA expression, there was no effect of fetal HPD, with or without cortisol replacement, on adrenal TGFβ1 mRNA expression in late gestation. Thus TGFβ1 may play a role in inhibiting adrenal steroidogenesis and ensuring that the adrenal remains relatively refractory to trophic stimulation during mid gestation. The maintenance of low adrenal TGFβ1 expression during late gestation is not dependent, however, on stimulation by the fetal hypothalamic-pituitary axis.

Convergence of Wnt Signaling and Steroidogenic Factor-1 (SF-1) on Transcription of the Rat Inhibin α Gene

The action of a variety of peptide hormones is critical for proper growth and differentiation of the urogenital ridge, which ultimately gives rise to the kidney, adrenal cortex, and gonad. One such class of peptides is the Wnt family of secreted glycoproteins that is classically involved in development of cell polarity and cell fate determination. Notably, alterations in Wnt-4 expression in mice and humans result in profound defects in urogenital ridge development, including dysregulation of kidney, gonadal, and adrenal growth. The nuclear receptor steroidogenic factor-1 (SF-1) has been implicated as a downstream effector of peptide hormone signaling during urogenital ridge development as evidenced by both the activation of SF-1-dependent transcription in the adrenal cortex by signaling molecules such as protein kinase A and by the adrenal and gonadal agenesis in mice with null mutations in SF-1. We hypothesized that Wnt-dependent signaling cascades regulate SF-1-dependent transcription of genes required for adreno-gonadal development. Specifically, the data demonstrate that beta-catenin synergizes with SF-1 to activate the alpha-inhibin promoter through formation of a transcriptional complex. The activation requires an intact SF-1 RE and is independent of TCF/Lef. These data support the recent observation that beta-catenin can participate in nuclear receptor-mediated transcriptional activation and extend the findings to the monomer binding class of orphan nuclear receptors.

Identification of a serine protease involved with the regulation of adrenal growth.

The adrenal cortex is a dynamic organ in which the cells of the outer cortex continually divide. It is well known that this cellular proliferation is dependent on constant stimulation from peptides derived from the ACTH precursor pro-opiomelanocortin (POMC) because disruption of pituitary corticotroph function results in rapid atrophy of the gland. Previous results from our laboratory have suggested that the adrenal mitogen is a fragment derived from the N-terminal of POMC not containing the gamma-MSH sequence. Because such a peptide is not generated during processing of POMC in the pituitary, we proposed that the mitogen is generated from circulating pro-gamma-MSH by an adrenal protease. Using degenerate oligonucleotides, we identified a secreted serine protease expressed by the adrenal gland that we named adrenal secretory protease (AsP). In the adrenal cortex, expression of AsP is limited to the outer zona glomerulosa/fasciculata, the region where cortical cells are believed to be derived, and is significantly up-regulated during compensatory growth. Y1 adrenocortical cells transfected with a vector expressing an antisense RNA (and thus having reduced levels of endogenous AsP) were found to grow slower than sense controls while also losing their ability to utilize exogenous pro-gamma-MSH in the media supporting a role for AsP in adrenal growth. Digestion of an N-POMC peptide substrate encompassing the residues around the dibasic cleavage site at positions 49/50 with affinity-purified AsP showed cleavage not to occur at the dibasic site but two residues downstream leading us to propose the identity of the adrenal mitogen to be N-POMC (1-52).

Adrenal neuropeptides: regulation and interaction with ACTH and other adrenal regulators.

It is now well accepted that both the cortex and medulla of the mammalian adrenal gland receive a rich innervation. Many different transmitter substances have been identified in nerves supplying both cortex and medulla and, as well as catecholamines, a wide range of neuropeptides has been found in the adrenal gland. There have been several studies on the affects of age, sodium intake, stress, ACTH, and splanchnic nerve activity on the regulation of adrenal neuropeptide content. There is evidence that the abundance of each of these peptides is actively regulated. Although there have been many studies addressing the individual actions of various neurotransmitters on steroid secretion, adrenal blood flow, and adrenal growth, few have attempted to determine the nature of any interaction between neurotransmitters and the classical adrenal stimulants. There are, however, some significant interactions, particularly in the regulation of zona glomerulosa function. This review necessarily focuses on vasoactive intestinal peptide (VIP) and neuropeptide Y (NPY), as these are the most abundant transmitter peptides in the adrenal gland and the majority of studies have investigated their regulation and actions. However, substance P, calcitonin gene-related peptide (CGRP), neurotensin, and the enkephalins are included where appropriate. Finally, it has been suggested that certain neurotransmitters, particularly VIP, may interact with classical hormone receptors in the adrenal, notably the ACTH receptor. This review attempts to evaluate our current state of knowledge in each of these areas.

N-terminal proopiomelanocortin acts as a mitogen in adrenocortical tumor cells and decreases adrenal steroidogenesis.

There is evidence that proopiomelanocortin (POMC)-derived peptides other than ACTH are involved in pituitary-dependent adrenal growth. We have synthesized the human N-terminal POMC fragment 1-28-POMC with the disulfide bridges in the correct position between cysteine residues 2-24 and 8-20 and studied the activity of these peptides in adrenocortical tumor cells in vitro. 1-28-POMC stimulated cell proliferation in human NCI-h295 and mouse Y-1 adrenal cancer cell lines and also in primary cultures of bovine adrenocortical cells in a concentration-dependent manner. 1-28-POMC led to rapid activation of the MAPKs extracellular signal-regulated kinases-1 and -2, but not c-Jun N-terminal kinase and p38, pathways. Steroid hormone production (cortisol, 17-hydroxyprogesterone, and dehydroepiandrosterone sulfate) in NCI-h295 cells was decreased by 1-28-POMC in a concentration-dependent fashion. However, protein levels of important regulators of steroidogenesis [steroidogenic factor-1, DAX-1 (dosage-sensitive sex reversal-adrenal hypoplasia congenita critical region on the X-chromosome 1), steroidogenic acute regulatory protein, and cytochrome P450 side-chain cleavage enzyme] remained unaffected by 1-28-POMC treatment. Our results provide evidence that synthetic 1-28-POMC induces adrenal tumor cell proliferation, inhibits adrenal steroidogenesis, and mediates its action by signaling via the extracellular signal-regulated kinase pathway. The distinct roles of 1-28-POMC and ACTH in the regulation of adrenal growth and steroidogenesis suggest that the adrenal cortex is under the dual opposing control of fragments from the same mother peptide POMC.

Tissue growth and remodeling of the embryonic and adult adrenal gland.

The adrenal gland provides a model system for the study of tissue remodeling in endocrine physiology. For example, proper adrenal development requires proliferation of the adrenogonadal primordia, separation of adrenal and gonadal precursors, and cell migration that unites the adrenal cortex and adrenal medulla. In the adult, normal adrenal function is assured by the adrenal gland's unique capacity for growth in response to both tissue injury and physiological demand. Identification of the molecular and genetic programs underlying tissue remodeling in the adrenal is important for understanding basic aspects of development and regeneration, as well as adrenal disease. Here, we will highlight the roles that nuclear receptors and pituitary hormones play in regulating fetal adrenal development and adult adrenal growth. In addition, we will review the most current data on how extracellular signaling pathways are coupled to the function of these important regulators of adrenal development and function.

Differential expression of VEGF receptors in adrenal atrophy induced by dexamethasone: a protective role of ACTH.

Although ACTH is important to adrenal growth and steroidogenesis, its role in vascular development and function has not been established in vivo. In the present study, we demonstrate the expression of mRNA for all four VEGF isoforms (mVEGF(120,144,164,188)) and for Flk-1/KDR and Flt-1 receptors in the mouse adrenal in vivo. Suppression of the pituitary adrenocortical axis by dexamethasone (0.5 mg x 100 g body wt(-1) x day(-1) during 6 days) induced a decrease in corticosterone levels, adrenal weights by 50% (P < 0.001), VEGF(188) mRNA, and Flk-1/KDR mRNA, whereas Flt-1 remained consistent during steroid treatment. A daily injection of ACTH-(1-39) restored the transcript for Flk-1/KDR and both VEGF(188) and plasma corticosterone to control levels. To gain further insights into the effects of ACTH, cultured endothelial cells (ECs) were treated with forskolin, which increases cAMP, the second messenger in ACTH action. We demonstrate that Flk-1/KDR protein expression was markedly increased by forskolin within 24-48 h of treatment in a dose-dependent manner (0.1-10 microM). The biological effect of ACTH on ECs was then tested by use of coincubations of fasciculata cells and ECs in 3D-collagen assay. Within 5-7 days of culture, ECs organized into multicellular structures that resemble networks of microvasculature, which characterize angiogenesis in vitro.

Capsaicin-sensitive adrenal sensory fibers participate in compensatory adrenal growth in rats.

Compensatory adrenal growth, in which one gland undergoes hyperplasia after removal of the other, is mediated by a neural reflex. In the present studies, a method employing capsaicin to selectively remove adrenal sensory fibers was developed and applied to determine whether adrenal capsaicin-sensitive fibers participate in compensatory adrenal growth. The splanchnic nerves of anesthetized male rats were treated with capsaicin or vehicle. Capsaicin treatment selectively removed adrenal calcitonin gene-related peptide-positive fibers. One week after drug treatment, rats underwent left adrenalectomy or sham surgery and recovered for 5 days. Capsaicin treatment bilaterally or to the left splanchnic nerve alone (i.e., the afferent nerve in the reflex) impaired compensatory adrenal growth at 5 days compared with vehicle controls, whereas capsaicin treatment to the right splanchnic nerve alone did not affect growth. Moreover, left adrenalectomy induced c-Fos immunolabeling in ipsilateral dorsal spinal cord that was prevented by capsaicin treatment. These data suggest that adrenal capsaicin-sensitive afferent nerves participate in compensatory adrenal growth and that this effect is primarily on the afferent limb of the reflex.

Maternal undernutrition throughout pregnancy increases adrenocorticotrophin receptor and steroidogenic acute regulatory protein gene expression in the adrenal gland of twin fetal sheep during late gestation

We have previously demonstrated that maternal undernutrition during either the ‘periconceptional’ (i.e. from 60 days (d) before until 7 d after mating) or ‘gestational’ periods (i.e. from 8 d after mating until the end of pregnancy) have differential effects on the subsequent development of the hypothalamo-pituitary-adrenocortical (HPA) axis and on adrenal growth and steroidogenesis in the sheep fetus during late gestation (term=147±3 d gestation). The specific mechanisms by which periconceptional or gestational undernutrition result in activation of the fetal HPA axis in late gestation are unclear. We have therefore investigated the impact of maternal nutrient restriction imposed either during the periconceptional period, or between 8 and 147 d gestation on the expression of specific genes in the fetal pituitary and adrenal which regulate adrenal steroidogenesis in late gestation. Ewes were maintained on either a Control (C) or Restricted (R, 70% of C) diet from 60 d before until 7 d after mating (periconceptional period) and then maintained on either a Control or Restricted diet from 8 d after mating for the remainder of pregnancy (gestational period). Four nutritional treatment groups were therefore generated (C–C, C–R, R–R and R–C). Whilst periconceptional undernutrition (R–R and R–C groups) resulted in higher fetal plasma adrenocorticotrophic hormone (ACTH) at 135–146 d gestation, there was no change in the relative level of expression of the ACTH receptor (MC2R), steroidogenic acute regulatory protein (StAR) or steroidogenic enzyme mRNAs in the fetal adrenal in late gestation. Exposure to gestational undernutrition (R–R and C–R groups), however, resulted in a stimulation in the relative level of expression of MC2R mRNA ( P=0.001) and StAR mRNA ( P=0.007) in the fetal adrenal during late gestation. This study provides new insights into the potential mechanisms by which alterations of the nutrient environment of the fetus at different stages of gestation may result in differential activation of the fetal HPA axis.

Steroidogenic acute regulatory protein expression is decreased in the adrenal gland of the growth-restricted sheep fetus during late gestation.

Functional development of the adrenal cortex is critical for fetal maturation and postnatal survival. In the present study, we have determined the developmental profile of expression of the mRNA and protein of an essential cholesterol-transporting protein, steroidogenic acute regulatory protein (StAR), in the adrenal of the sheep fetus. We have also investigated the effect of placental restriction (PR) on the expression of StAR mRNA and protein in the growth-restricted fetus. Adrenal glands were collected from fetal sheep at 82-91 days (n = 10), 125-133 days (n = 10), and 140-144 days (n = 9) and from PR fetuses at 141-145 days gestation (n = 9) (term = 147 +/- 3 days gestation). The adrenal StAR mRNA:18S rRNA increased (P < 0.05) between 125 days (7.44 +/- 1.61) and 141-144 days gestation (13.76 +/- 1.88). There was also a 13-fold increase (P < 0.05) in the amount of adrenal StAR protein between 133 and 144 days gestation in these fetuses. However, the amount of StAR protein (6.9 +/- 1.7 arbitrary densitometric units [AU]/microg adrenal protein) in the adrenal of the growth-restricted fetal sheep was significantly reduced, when compared with the expression of StAR protein (17.1 +/- 1.9 AU/microg adrenal protein) in adrenals from the age-matched control group. In summary, there is a developmental increase in the expression of StAR mRNA and protein in the fetal sheep adrenal during the prepartum period when adrenal growth and steroidogenesis is dependent on ACTH stimulation. We have found that, while the level of expression of StAR protein is decreased in the adrenal gland of the growth-restricted fetus during late gestation, this does not impair adrenal steroidogenesis. Our data also suggest that the stimulation of adrenal growth and steroidogenesis in the growth-restricted fetus may not be ACTH dependent.