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Triadimefon is a broad-spectrum antifungal agent, which is widely used in agriculture to control mold and fungal infections. It is considered an endocrine disruptor. Whether triadimefon exposure can inhibit the development of fetal adrenal glands and the underlying mechanism remain unclear. Thirty-two pregnant female Sprague-Dawley rats were randomly divided into four groups. Dams were gavaged triadimefon (0, 25, 50, and 100 mg/kg/day) daily for 10 days from gestational day (GD) 12 to GD 21. Triadimefon significantly reduced the thickness of the zona fasciculata of male fetuses at 100 mg/kg, although it did not change the thickness of the zona glomerulosa. It significantly reduced the serum aldosterone levels of male fetuses at a dose of 100 mg/kg, and significantly reduced serum corticosterone and adrenocorticotropic hormone levels at doses of 50 and 100 mg/kg. Triadimefon significantly down-regulated the expression of Agtr1, Mc2r, Star, Cyp11b1, Cyp11b2, Igf1, Nr5a1, Sod2, Gpx1, and Cat, but did not affect the mRNA levels of Scarb1, Cyp11a1, Cyp21, Hsd3b1, and Hsd11b2. Triadimefon markedly reduced AT1R, CYP11B2, IGF1, NR5A1, and MC2R protein levels. Triadimefon significantly reduced the phosphorylation of AKT1 and ERK1/2 at 100 mg/kg without affecting the phosphorylation of AKT2. In contrast, it significantly increased AMPK phosphorylation at 100 mg/kg. In conclusion, exposure to triadimefon during gestation inhibits the development of fetal adrenal cortex in male fetuses. This inhibition is possibly due to the reduction of several proteins required for the synthesis of steroid hormones, and may be involved in changes in antioxidant contents and the phosphorylation of AKT1, ERK1/2, and AMPK., (Copyright © 2021. Published by Elsevier B.V.)

This review summarizes key knowledge regarding the development, growth, and growth disorders of the adrenal cortex from a molecular perspective. The adrenal gland consists of two distinct regions: the cortex and the medulla. During embryological development and transition to the adult adrenal gland, the adrenal cortex acquires three different structural and functional zones. Significant progress has been made in understanding the signaling and molecules involved during adrenal cortex zonation. Equally significant is the knowledge obtained regarding the action of peptide factors involved in the maintenance of zonation of the adrenal cortex, such as peptides derived from proopiomelanocortin processing, adrenocorticotropin and N-terminal proopiomelanocortin. Findings regarding the development, maintenance and growth of the adrenal cortex and the molecular factors involved has improved the scientific understanding of disorders that affect adrenal cortex growth. Hypoplasia, hyperplasia and adrenocortical tumors, including adult and pediatric adrenocortical adenomas and carcinomas, are described together with findings regarding molecular and pathway alterations. Comprehensive genomic analyses of adrenocortical tumors have shown gene expression profiles associated with malignancy as well as methylation alterations and the involvement of miRNAs. These findings provide a new perspective on the diagnosis, therapeutic possibilities and prognosis of adrenocortical disorders.

The adrenocortical gland undergoes structural and functional remodelling in the fetal and postnatal periods. After birth, the fetal zone of the gland undergoes rapid involution in favor of the definitive cortex, which reaches maturity with the emergence of the zona reticularis(zR) at the adrenarche. The mechanisms underlying the adrenarche, the process leading to pre-puberty elevation of plasma androgens in higher primates, remain unknown, largely due to lack of any experimental model. By following up fetal and definitive cortex cell lines in mice, we showed that activation of protein kinase A (PKA) signaling mainly impacts the adult cortex by stimulating centripetal regeneration, with differentiation and then conversion of the zona fasciculata into a functional zR. Animals developed Cushing syndrome associated with primary hyperaldosteronism, suggesting possible coexistence of these hypersecretions in certain patients. Remarkably, all of these traits were sex-dependent: testicular androgens promoted WNT signaling antagonism on PKA, slowing cortical renewal and delaying onset of Cushing syndrome and the establishment of the zR in male mice, this being corrected by orchidectomy. In conclusion, zR derives from centripetal conversion of the zona fasciculata under cellular renewal induced by PKA signaling, determining the size of the adult cortex. Finally, we demonstrated that this PKA-dependent mobilization of cortical progenitors is sexually dimorphic and could, if confirmed in humans, account for female preponderance in adrenocortical pathologies., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

The human adult adrenal cortex is composed of the zona glomerulosa (zG), zona fasciculata (zF), and zona reticularis (zR), which are responsible for production of mineralocorticoids, glucocorticoids, and adrenal androgens, respectively. The final completion of cortical zonation in humans does not occur until puberty with the establishment of the zR and its production of adrenal androgens; a process called adrenarche. The maintenance of the adrenal cortex involves the centripetal displacement and differentiation of peripheral Sonic hedgehog-positive progenitors cells into zG cells that later transition to zF cells and subsequently zR cells., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The adrenal gland controls a plethora of crucial physiological functions, and dysfunction is associated with severe morbidity. Because of the vital importance of appropriate adrenal function, the development and function of the gland have been intensively studied, and these investigations have revealed fascinating developmental origins and a remarkable remodeling and regenerative capacity in the adult. This chapter, focusing on the adrenal cortex, will describe our current understanding of the development and maintenance of the adrenal gland, which has been advanced over recent years by the use of sophisticated genetic models in the study of both normal function and disease. This work has shed light on the transcriptional networks and signaling pathways involved in development and maintenance of the gland and in its pathology; these are discussed in the light of the wealth of physiological information gathered in studies of human and rodent adrenal development and function., (© 2013 Elsevier Inc. All rights reserved.)

During the early phases of development, adrenal glands share a common origin with kidneys and gonads. The action of diverse transcription factors, signalling pathways and endocrine signals is required for the individualization of the adrenal primordium and its subsequent differentiation into an adult adrenal gland, with massive remodelling taking place around the time of birth in humans. Here I summarize the most important steps by which the adrenal cortex is shaped and present an overview of the current understanding of the genes and molecular pathways implicated in adrenal development and involved in the pathogenesis of its congenital diseases. Evidence is accumulating that some pivotal factors acting during adrenocortical development also play an important role to regulate the growth of adrenocortical tumors, representing promising therapeutical targets for a biology-oriented therapy., (2010 Elsevier Ltd. All rights reserved.)

Adrenocortical dysplasia (acd) is a spontaneous autosomal recessive mouse mutation that exhibits a pleiotropic phenotype with perinatal lethality. Mutant acd embryos have caudal truncation, vertebral segmentation defects, hydronephrosis, and limb hypoplasia, resembling humans with Caudal Regression syndrome. Acd encodes Tpp1, a component of the shelterin complex that maintains telomere integrity, and consequently acd mutant mice have telomere dysfunction and genomic instability. While the association between genomic instability and cancer is well documented, the association between genomic instability and birth defects is unexplored. To determine the relationship between telomere dysfunction and embryonic malformations, we investigated mechanisms leading to the caudal dysgenesis phenotype of acd mutant embryos. We report that the caudal truncation is caused primarily by apoptosis, not altered cell proliferation. We show that the apoptosis and consequent skeletal malformations in acd mutants are dependent upon the p53 pathway by genetic rescue of the limb hypoplasia and vertebral anomalies with p53 null mice. Furthermore, rescue of the acd phenotype by p53 deficiency is a dosage-sensitive process, as acd/acd, p53(-/-) double mutants exhibit preaxial polydactyly. These findings demonstrate that caudal dysgenesis in acd embryos is secondary to p53-dependent apoptosis. Importantly, this study reinforces a significant link between genomic instability and birth defects.

The network regulating human adrenal development is complex. Studies of patients with adrenal insufficiency due to gene mutations established a central role for transcription factors GLI3, SF1 and DAX1 in the initial steps of adrenal formation. Adrenal differentiation seems to depend on adrenocorticotropic hormone (ACTH) stimulation and signalling, including biosynthesis and action of POMC, PC1, TPIT, MC2R, MRAP and ALADIN, all of which cause adrenocortical hypoplasia when mutated in humans. Studies of knockout mice revealed many more factors involved in adrenal development; however, in contrast to rodents, in humans several of those factors had no adrenal phenotype when mutated (e.g. WT1, WNT4) or, alternatively, human mutations have not (yet) been identified. Tissue profiling of fetal and adult adrenals suggested 69 genes involved in adrenal development. Among them were genes coding for steroidogenic enzymes, transcription and growth factors, signalling molecules, regulators of cell cycle and angiogenesis, and extracellular matrix proteins; however, the exact role of most of them remains to be elucidated.

Histological and functional characteristics of the fetal human adrenals was studied in 119 normal fetuses aged 12 to 36 weeks development (WD). Immunocytochemical detection of steroidogenesis enzyme (3beta-HSD and P450 c21) and evaluation of cell proliferation using two nuclear markers (Ki-67 and PCNA) were performed in 70 of them. The human fetal adrenal cortex is composed of two morphologically distinct zones: the definitive peripheral zone and the fetal inner zone. From the 12th WD, we observed expression of an adherence protein (NCAM) and two steroidogenesis enzymes (3beta-HSD and P450 c21) in the definitive zone cells, attesting to the capacity of these cells to synthesize mineralocorticoids and/or cortisol. In the fetal zone, only P450 c21 immunoreactivity was detected. From the 14th WD, a transitional zone between the definitive zone and the fetal zone was identified by immunocytochemistry, with expression of 3b-HSD from the 21st WD. Only cells of the definitive zone proliferated from the 12th to 25th WD. The indexes of proliferation of PCNA and Ki-67, 40% and 25% respectively, decreased gradually and were lower than 1% at the 25th WD.

The human adrenal is an endocrine gland located at the superior part of the kidney. Composed of the adrenal cortex of mesoblastic origin and the adrenal medulla of neuroectoblastic origin, the human fetal adrenal grows considerably during the first three months of development. From 12 to 18 weeks of development (WD), the weight of the adrenals increases seven-fold. The gland's weight doubles from 18 to 28 WD and from 28 to 36 WD. At birth, the two adrenals weigh on average 10 g. At the 8th week, two zones are individualized in the adrenal cortex: the definitive zone and the fetal inner zone. At the second trimester, according to ultrastructural and biochemical studies, a third zone, called the transition zone, is individualized between the definitive zone and the fetal inner zone. The definitive zone persists, but the origin of the three zones (glomerular, fascicular and reticular) of adult adrenal cortex is not known. The fetal inner zone regresses from the 5th month of gestation and disappears totally one year after birth. At the 8th week, the immature neuroblasts migrate to the definitive zone, then to the fetal inner zone to compose the adrenal medulla, which develops essentially after birth and during the first year. Before the 10th week, the human fetal adrenal is able to produce steroid hormones, in particular dehydroepiandrosterone sulfate (DHEA-S); the secretion of cortisol remains discussed. The development of the human fetal adrenal is complex and is under the control of hormones (ACTH, LH and betaHCG), growth factors (ACTH essentially) and transcription factors (essentially SF1 and DAX-1). Knowledge of morphological and molecular phenomena of this development permits to understand the pathophisiology of congenital adrenal deficiencies.

The embryo of the domestic fowl (Gallus domesticus) tenders one distinctive advantage over general mammalian models for investigating the development of the hypothalamo-pituitary-adrenocortical (HPA) axis. This is the relative simplicity with which the embryonic endocrine environment can be influenced without confounding maternal influences. The ease of direct manipulation of the embryonic endocrine system has facilitated analysis of the development and function of the HPA axis in the chick embryo. As the chick embryo develops, functional activation of the adrenal gland is regulated at three different levels: the adrenal gland itself, the anterior pituitary, and the hypothalamus. The adrenal gland appears capable of independent secretion of glucocorticoids from day 8 until shortly after day 14 of embryonic development, at which point the pituitary influences adrenocortical activity. Around the same age, the hypothalamic level of control also begins. The information covered in this review will describe the major steps in the development of the HPA axis in the chicken embryo and show that the chicken has an emblematic HPA neuroendocrine axis.

Objective: Maturation of adrenocortical function is important because a prepartum increase in fetal plasma glucocorticoids is required for survival after birth. Adrenal maturation may include alterations in the regulation of adrenocorticotropic hormone (ACTH) receptor expression. Therefore, we quantitated ACTH receptor expression in the ovine adrenal cortex during development and after manipulations to better understand the regulation of the adrenal receptor in vivo., Methods: For the ontogeny study, adrenals were obtained from fetuses at different stages of development, and the cortical tissue was stored at -80C until total RNA was extracted. The ACTH binding studies were done on adrenal membranes obtained from fetuses at two different ages using I125 (Phe-2, Nle-4) ACTH as the ligand. Plasma ACTH was measured by two-site immunoradiometric assay, and cortisol was measured by radioimmunoassay. ACTH receptor mRNA was quantitated by ribonuclease protection assay. The data were analyzed by analyses of variance., Results: ACTH receptor mRNA level progressively increased in fetal life; relative changes in receptor mRNA and binding were similar (3.0-fold and 2.4-fold, respectively). Physiologic increases in fetal plasma cortisol decreased adrenal ACTH receptor mRNA concentration, and there was a strong (r2=0.76, P<.002) linear relationship between fetal plasma ACTH concentration and receptor mRNA levels. Receptor mRNA stability increased in development, and message half-life was greater in adulthood than in fetal life., Conclusion: The data suggest that developmental increases in receptor expression are part of the maturation process in the fetal adrenal and that plasma ACTH concentration plays a major role in regulating ACTH receptor mRNA levels in vivo.

We examined fetal sheep adrenal glands from 99 to 130 days of gestational age (dGA) to see how connexin 43 (Cx43), the major if not the only adult adrenal gap junction protein, changes expression as the adrenal cortex emerges from the well-documented refractory period to participate in labor and delivery. Immunocytochemical technique and Western blot were used to examine changes in the quantity and quality of Cx43. In addition, adrenal glands of ACTH infused fetuses or of fetuses from dexamethasone injected ewes underwent image analysis quantification after Cx43 immunostaining. Finally, fetal adrenal glands, from fetuses splanchnic nerve sectioned (SPLX) at 125 dGA, were examined for the pattern of Cx43 immunostaining at 131 days of gestation. From 100 to 130 dGA, the amount of Cx43 in cells of the adrenal cortex increased steadily while the pattern of immunoreactivity changed from predominantly cytoplasmic to membrane bound. At 100-103 dGA, ACTH infusion increased the size of the cortex, but decreased the expression of Cx43 per unit area while dexamethasone had no effect on either parameter. Lastly, the expression of Cx43 as a membrane bound protein was not delayed or reversed by SPLX. We conclude that the described changes in Cx43 are most intriguing given their temporal relationship to the described preparturient increases in ACTH and cortisol in peripheral fetal plasma as term approaches and deserve further investigation.

Objective: Erythropoietin receptors (Epo-R) have been demonstrated on several nonhematopoietic cell types in animal models and in cell culture. Our objective was to determine the tissue distribution and cellular specificity of erythropoietin (Epo) and its receptor in the developing human fetus., Study Design: The expression of Epo and Epo-R mRNA was ascertained by RT-PCR for organs ranging in maturity from 5 to 24 weeks postconception. The cellular location of protein immunoreactivity was then determined using specific antiEpo and antiEpo-R antibodies. Antibody specificity was established by Western analysis., Results: mRNA for Epo and Epo-R was found in all organs in the first two trimesters. Immunolocalization of Epo was limited to the liver parenchymal cells, kidney interstitial cells and proximal tubules, neural retina of the eye, and adrenal cortex. As development progressed, immunoreactivity in the kidney became more prominent. In contrast, immunoreactivity for Epo-R was widespread throughout the body, in cell types including endothelial cells, myocardiocytes, macrophages, retinal cells, cells of the adrenal cortex and medulla, as well as in small bowel, spleen, liver, kidney, and lung., Conclusions: The distribution of Epo and its receptor is more widespread in the developing human than was initially postulated. Epo-R is expressed on many cell types during early fetal development, leading us to speculate that Epo acts in concert with somatic growth and development factors during this period. Further investigation is required to understand the nonhematopoietic role of Epo during human development.

Development of the human fetal adrenals is characterized by rapid growth and high levels of steroidogenic activity during the latter two-thirds of pregnancy. By midgestation, the human fetal adrenals are composed of two distinct cortical zones: the predominant fetal zone, which occupies 80-90% of the cortical volume and produces large amounts of the delta 5-steroid dehydroepiandrosterone sulfate, and the narrow definitive zone, which surrounds the fetal zone. Late in gestation, the peripheral portion of the fetal zone develops into a third, functionally distinct compartment, the transitional zone, which is the likely site of cortisol synthesis. Soon after birth, the adrenal cortex is remodeled and the fetal zone disappears. The adult cortical zones are thought to arise from the definitive zone, which persists postnatally. Development of the human fetal adrenals is regulated primarily by corticortropin (ACTH) secreted from the fetal pituitary. However, as ACTH is not a mitogen per se, its proliferative actions on human fetal adrenal cortical cells are thought to be mediated by autocrine/paracrine growth factors produced by adrenal cortical cells in response to ACTH. In addition, these growth factors appear to modulate the functional response of fetal adrenal cortical cells to ACTH. The roles of several growth factors, including the insulin like growth factors I and II (IGF-I and IGF-II), epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), activin, inhibin, and the transforming growth factors alpha and beta (TGF-alpha and TGF-beta) have been examined. In cultured human fetal adrenal cortical cells, EGF, bFGF, and IGF-I and -II are mitogenic, whereas activin and TGF-beta inhibit proliferation. IGF-II, activin, and TGF-beta also modulate ACTH-stimulated steroidogenesis. Human fetal adrenal cortical cells express IGF-II, bFGF and the activin/inhibin subunits, and the abundance of mRNAs for each of these factors is up-regulated by ACTH, suggesting that these growth factors are autocrine/paracrine mediators of ACTH action. Thus, although human adrenal development is primarily regulated by ACTH, its actions appear to be mediated/modulated by a cohort of locally expressed growth factors, the net effect of which results in the unique growth and steroidogenic activity of the human fetal adrenal cortex.

Specific receptors for glucocorticoids were identified in the fetal and adult sheep adrenal cortex by a whole-cell binding assay using [3H]triamcinolone acetonide ([3H]TA) as the radiolabelled ligand. [3H]TA binding sites were saturable and of high affinity, with dissociation constant (Kd) of 2-3nM. Scatchard analysis revealed a single class of binding sites with a binding capacity (Bmax) of 207 and 5 fmol/10(6) cells for d100 fetuses and adults, respectively. By single point analyses at saturating [3H]TA concentration, we found that glucocorticoid receptors (GR) were present in the fetal adrenal cortex as early as d60. Highest concentrations were found at d100-110. GR decreased to d125, then increased to term (approx. d145) before declining again in newborn and adult animals. This demonstration of glucocorticoid receptors in ovine fetal adrenal cortical cells provides a mechanistic basis for the concept that glucocorticoids may act, perhaps in a paracrine or autocrine fashion, to influence adrenocorticotropin (ACTH)-induced activation of fetal adrenal function and the events leading to parturition.

In mammals, the fetal adrenal gland plays a key role during late gestation since the maturation of the fetus and the adaptation of the neonate to extra uterine life mainly depend on glucocorticoids. We review herein the present knowledge concerning the modalities and the endocrine control of the development of the pituitary-adrenal axis in the two most studied models: the ovine and the human fetus. Although the anatomy of the adrenal gland is different in these 2 species, the mechanisms involved in the maturation of adrenocortical cells appear rather similar. Moreover, in both species, the prepartum rise of corticosteroid plasma levels which conveys growth and differentiation of adrenocortical cells, is under pituitary control; this likely involves an increase in the "corticotropic activity" which results from changes in the cleavage of POMC and possibly from a synergistic of inhibitory effect of other hormones with ACTH.

Fibroblasts of the adult adrenal cortex are considered to be nonsteroidogenic connective-tissue cells. However, it has been reported that in response to regenerative stimuli, adrenocorticotropic hormone (ACTH), and transformation to malignancy, these cells acquire characteristics of parenchymal cells, which includes delta 5, 3 beta-hydroxysteroid-dehydrogenase (delta 5, 3 beta-HSD) activity. To determine whether such delta 5, 3 beta-HSD activity in adult adrenocortical fibroblasts was due to the activation or augmentation of gene expression normally occurring during embryogenesis, a histochemical study of adrenocortical development, with particular attention to the connective-tissue capsule, was undertaken. Cryostat sections of rat embryos, from 14-days postconception (PC) to birth, and of adrenal glands 1-8, 44 and 90 days after birth were tested histochemically for delta 5, 3 beta-HSD. The same or adjacent sections were stained for PAS-positive material and reticulin, and with hematoxylin and eosin. delta 5, 3 beta-HSD activity overlapped with fibroblast-like cells and with extracellular connective-tissue components in the periphery of the glands from day-17 PC onward. delta 5, 3 beta-HSD activity over the capsule diminished shortly after birth and was absent in the adult. Appropriate controls showed that the staining within the capsule was specific and not an artifact. 3 beta-HSD activity in the capsule was more intensive when dehydroepiandrosterone (DHEA) was replaced by etiocholan-3 beta-ol-17-one (ETIO) as the steroid substrate. Furthermore, the spatial distribution of 3 beta-HSD activity in the cortex differed depending on the substrate used, and the distribution patterns changed with developmental age. (ABSTRACT TRUNCATED AT 250 WORDS)

The ability of human adult adrenal to utilize progesterone (P4) for cortisol (F) synthesis in vitro has been compared with that of fetal adrenal tissue. Explant cultures were studied for 6 days using Ham's F10 medium supplemented with 10% fetal bovine serum (FBS), with or without added P4 as substrate. Short-term (4h) incubations of fresh tissue minces were carried out in Ham's F10, with or without P4, in the absence of FBS. In contrast with the fetal gland, F production by cultured adult tissue was unaffected by addition of P4. During short-term incubations, P4 increased F production 150-fold in the fetal tissue as compared to 2- to 4-fold in the adult. ACTH had no acute effect on the P4 to F conversion in either tissue. These results demonstrate that the fetal adrenal exhibits a greater ability to utilize P4 for F production than the adult adrenal.

The aim of the present study was to assess whether glucocorticoids could be directly involved in the maturation of adrenocortical cells from 120-138 days old ovine fetuses. The cAMP response to ACTH1-24 of cells cultured for 24 hours in the presence of ACTH1-24 was 2 fold higher than that of control cells. However, the response of cells cultured in the presence of ACTH1-24 plus metyrapone or aminoglutethimide was lower than that of cells cultured in the presence of ACTH1-24 alone. Cells cultured for 48 hours in the presence of dexamethasone or cortisol released more cAMP than control cells when stimulated by ACTH1-24, but not in response to forskolin. However corticosteroid production stimulated by ACTH1-24, forskolin or dibutyryl cAMP was enhanced by dexamethasone treatment. These results suggest that glucocorticoids can affect the maturation of ovine fetal adrenocortical cells by an auto and/or a paracrine process, and that this effect is exerted, at least, at two different levels in the cell.

Studies of human fetal adrenal function and its control have revolved mainly around the remarkable capacity of the unique fetal zone of this gland to elaborate dehydroepiandrosterone sulfate. Another important function of the fetal adrenal, however, is its production of cortisol. Because the human fetal adrenal is deficient in 3 beta-hydroxysteroid dehydrogenase activity, cortisol has been thought to be formed from circulating progesterone. To further investigate this hypothesis, cortisol production by separated fetal and definitive zones of the midgestation human fetal adrenal in organ culture has been examined in the absence and presence of varying concentrations of progesterone and adrenocorticotropic hormone. Cortisol was measured by radioimmunoassay. In the absence of progesterone, cortisol production by both zones increased gradually over time in culture in response to adrenocorticotropic hormone. In the presence of progesterone, cortisol production by the definitive zone was unchanged. In contrast, the response of the fetal zone to progesterone was immediate: cortisol production increased significantly and remained high throughout the culture period. These results suggest a greater capacity of the fetal zone to utilize progesterone for cortisol production and are consistent with morphologic evidence that the active zone of the midgestation human fetal adrenal is the fetal zone, possessing not only the enzyme activity necessary for dehydroepiandrosterone sulfate production but, except for 3 beta-hydroxysteroid dehydrogenase, that for cortisol production as well.