Showing papers on "Adrenal cortex published in 1994"

Disorders of steroid 11β-hydroxylase isozymes

Abstract: The most active corticosteroids are 11 beta-hydroxylated. Humans have two isozymes with 11 beta-hydroxylase activity that are respectively required for cortisol and aldosterone synthesis. CYP11B1 (11 beta-hydroxylase) is expressed at high levels and is regulated by ACTH, whereas CYP11B2 (aldosterone synthase) is normally expressed at low levels and is regulated by angiotensin II. In addition to 11 beta-hydroxylase activity, the latter enzyme has 18-hydroxylase and 18-oxidase activities and thus can synthesize aldosterone from deoxycorticosterone. Insights into the normal functioning of these enzymes are gained from studies of disorders involving them. Mutations in the CYP11B1 gene cause steroid 11 beta-hydroxylase deficiency, a form of congenital adrenal hyperplasia characterized by signs of androgen excess and by hypertension. Mutations in CYP11B2 result in aldosterone synthase (corticosterone methyloxidase) deficiency, an isolated defect in aldosterone biosynthesis that can cause hyponatremia, hyperkalemia, and hypovolemic shock in infancy and failure to thrive in childhood. These are both recessive disorders. Unequal crossing over between the CYP11B genes can generate a duplicated chimeric gene with the transcriptional regulatory region of CYP11B1 but sufficient coding sequences from CYP11B2 so that the encoded enzyme has aldosterone synthase (i.e. 18-oxidase) activity. This results in aldosterone biosynthesis being regulated by ACTH, a condition termed glucocorticoid-suppressible hyperaldosteronism. This form of genetic hypertension is inherited in an autosomal dominant manner.

Autoantibodies to cytochrome P450 enzymes P450scc, P450c17, and P450c21 in autoimmune polyglandular disease types I and II and in isolated Addison's disease

Abstract: Patients with idiopathic Addison's disease have autoantibodies reacting with adrenal cortex. If Addison's disease is associated with other endocrine immune diseases like autoimmune polyglandular diseases (APD) type I and type II, antibodies may recognize all steroid-producing cells. We showed previously that one antigen recognized by APD-I sera is the cytochrome P450c17 hydroxylase. We have now looked for antibodies to P450c17 and to two other key enzymes in the steroid biosynthetic pathway, the P450scc and P450c21, in a series of patients with isolated Addison's disease (8 patients) or with APD-I or APD-II (50 and 9 patients, respectively). The result of antienzyme antibodies were further correlated with the immunofluorescence pattern against adrenal gland, testis, ovary, and placenta, and with the clinical findings presented. In APD-I patients with Addison's disease and in APD-II patients, antibodies to at least one of the P450 enzymes were frequently found (positive findings in 81% and 78%, respectively). Such antibodies were less frequent in APD-I patients without Addison's disease (21%) and in the isolated Addison cases (25%). In APD-I, antibodies recognized as frequently P450c17 and P450scc, specific for all steroid-producing cells as the adrenal specific enzyme P450c21. In contrast, patients with APD-II or with the isolated Addison's disease reacted almost exclusively with P450c21. Immunofluorescence studies showed good correlation with the known fact that the zona glomerulosa of the adrenal cortex is devoid of the P450c17, that the Leydig cells of the testis and the theca interna cells of the ovary express P450c17 and P450scc, and that the placental trophoblasts express only P450scc. The presence of antibodies to P450scc or to at least one of the tested P450 enzymes correlated significantly to gonadal failure in the females but not in the males.

Hypothalamic-pituitary-adrenocortical axis changes in the rat after long-term treatment with the reversible monoamine oxidase-A inhibitor moclobemide

Abstract: The effects of the reversible monoamine oxidaseA (MAOA) inhibitor moclobemide on the rat hypothalamic-pituitary-adrenocortical (HPA) axis were studied. The time-course experiments showed that moclobemide, given via the drinking water (4.5 mg/kg/day), produces significant decreases (p < 0.05) in adrenal weight after 5 (-23%) and 7 weeks (-16%) of treatment. It was found that long-term moclobemide treatment had neuroanatomically distinct effects on corticosteroid receptor expression. Hippocampal mineralocorticoid receptor (MR) levels were upregulated at 2 (+65%), 5 (+76%) and 7 (+19%) weeks of treatment. Glucocorticoid receptor (GR) levels in this limbic brain structure were slightly up-regulated by 10% at 5 weeks, and indistinguishable from controls after 2 and 7 weeks of treatment. After 5 weeks of treatment, MR levels were unchanged in the hypothalamus, and increased by 44, 24 and 28% in the neocortex, amygdala and anterior pituitary, respectively. GR concentrations were elevated by 24 and 14% in the hypothalamus and anterior pituitary, respectively, whereas neocortical and amygdaloid receptor levels were not altered. After 5 weeks of moclobemide treatment, marked decreases in [125I]Tyr0-ovine corticotropin-releasing hormone ([125I])-oCRH binding capacity and proopiomelanocortin (POMC) mRNA content were observed in the anterior pituitary. Regarding the functional implications of long-term anti-depressant treatment, moclobemide treatment (5 weeks, 4.5 mg/kg/day) significantly attenuated stress (30-min novel environment)-induced plasma ACTH (-35%) and corticosterone (-29%) levels; no changes were observed in basal plasma ACTH and corticosterone levels. In conclusion, this study shows that moclobemide has a concerted influence on multiple elements of the HPA axis manifesting functionally as a reduced neuroendocrine responsiveness to stress. In previous experiments, it was found that the structurally and pharmacologically distinct antidepressant amitriptyline after long-term administration also attenuated HPA axis activity. We postulate that an adjustement of HPA axis activity may be regarded as a common denominator for clinically efficacious antidepressant drugs.

Ouabain is secreted by bovine adrenocortical cells.

Abstract: Ouabain is a specific inhibitor of the sodium pump. This steroid has been found in the mammalian circulation in significant amounts and may be of adrenal origin. Secretion of ouabain from adrenal cells has been little studied and the purpose of the present work was to determine the adrenal distribution of ouabain, aldosterone and cortisol, and to characterize the effects of ACTH and angiotensin II on the secretion of these steroids in primary cultures of bovine adrenocortical cells. In fresh bovine adrenals, the cortical to medullary ratios for aldosterone, cortisol and ouabain were 14, 4.25 and 2.5, respectively. All three steroids were detected in elevated amounts in the conditioned medium of primary cultures of adrenocortical cells. Reverse phase HPLC of the secreted ouabain immunoreactivity showed it was isopolar with commercial ouabain. In the presence of 10 nM ACTH or angiotensin II, the secretion of all three steroids increased significantly with similar time courses. The stimulated secretion of ouabain exceeded the intracellular content of this steroid in either control or activated cells by 3-5 fold. The amount of angiotensin II stimulated ouabain secretion was greater from cells incubated in larger volumes. These results show that ouabain is enriched in the bovine adrenal cortex, and is secreted by primary cultures of these cells. The secretion of ouabain is increased by ACTH and angiotensin II, is due to either de novo synthesis or transformation of an intracellular precursor that is not overtly immunoreactive, and is feedback regulated by either ouabain itself or a cosecreted factor. These cells may be useful to study stimulus-secretion coupling and the biosynthetic pathway of ouabain.

Functional difference between Ad4BP and ELP, and their distributions in steroidogenic tissues.

Abstract: Ad4BP, a zinc finger DNA-binding protein, was identified as a transcription factor regulating steroidogenic P-450 genes in a cAMP-dependent manner. Immunochemical and immunohistochemical studies with steroidogenic tissues, adrenal, ovary, and testis, were performed using the antiserum to Ad4BP. Ad4BP was expressed to the same extent in the three zones of the adrenal cortex. Immunohistochemical examination of ovarian follicle and corpus luteum showed the expression of Ad4BP. The granulosa and thecal cells, the two distinct types of the steroidogenic cells in the follicle, gave Ad4BP signals, which were stronger than in the latter cells than in the former. Immunoblot analyses of mature and regressed corpora lutea indicated a parallel expression of Ad4BP and side-chain cleavage P-450, and both proteins significantly decreased in the regressed tissues. Leydig cells surrounding seminiferous tubules gave clear immunostaining signals for Ad4BP. ELP, a mammalian counterpart of Drosophila FTZ-F1 detected in EC cel...

Endocrine and neurogenic regulation of the Orphan nuclear receptors Nur77 and Nurr-1 in the adrenal glands

Abstract: nurr77 and nurr-1 are growth factor-inducible members of the steroid/thyroid hormone receptor gene superfamily. In order to gain insight into the potential roles of nur77 in the living organism, we used pharmacologic treatments to examine the expression of nur77 in the mouse adrenal gland. We found that nur77 and nurr-1 are induced in the adrenal gland upon treatment with pentylene tetrazole (Ptz; Metrazole). This induction is separable into distinct endocrine and neurogenic mechanisms. In situ hybridization analysis demonstrates that nur77 expression upon Ptz treatment in the adrenal cortex is localized primarily to the inner cortical region, the zona fasciculata-reticularis, with minimal induction in the zona glomerulosa. This induction is inhibitable by pretreatment with dexamethasone, indicating involvement of the hypothalamic-pituitary-adrenal axis in the activation of adrenal cortical expression. When mice were injected with adrenocorticotrophic hormone (ACTH), nur77 expression in the adrenal gland spanned all cortical layers including the zona glomerulosa, but medullary expression was not induced. Ptz also induces expression of both nur77 and nurr-1 in the adrenal medulla. Medullary induction is likely to have a neurogenic origin, as nur77 expression was not inhibitable by dexamethasone pretreatment and induction was seen after treatment with the cholinergic neurotransmitter nicotine. nur77 is also inducible by ACTH, forskolin, and the second messenger analog dibutyryl cyclic AMP in the ACTH-responsive adrenal cortical cell line Y-1. Significantly, Nur77 isolated from ACTH-stimulated Y-1 cells bound to its response element whereas Nur77 present in unstimulated cells did not. Moreover, Nur77 in ACTH-treated Y-1 cells was hypophosphorylated at serine 354 compared with that in untreated cells. These results, taken together with the previous observation that dephosphorylation of serine 354 affects DNA binding affinity in vitro, show for the first time that phosphorylation of Nur77 at serine 354 is under hormonal regulation, modulating its DNA binding affinity. Thus, ACTH regulates Nur77 in two ways: activation of its gene and posttranslational modification. A promoter analysis of nur77 induction in Y-1 cells indicates that the regulatory elements mediating ACTH induction differ from those required for induction in the adrenal medullary tumor cell line PC12 and in 3T3 fibroblasts.

Intimate contact of chromaffin and cortical cells within the human adrenal gland forms the cellular basis for important intraadrenal interactions.

Abstract: A new role for the adrenal medulla as a regulator of adrenocortical function has been postulated. However, there has been no idea as to how such a cellular interaction within the human adrenal gland could take place. In this study we were able to demonstrate with the help of specific immunostaining of cortical and chromaffin cells, respectively, that the two endocrine systems are interwoven with each other to an astonishing degree. Protrusions, clusters, islets, and single cortical cells were made visible by immunostaining with an antibody against 17 alpha-hydroxylase cytochrome P450 enzyme. They occurred diffusely within the entire adrenal medulla, providing ample contact zones for paracrine interactions. Specific immunostaining for the neuroendocrine protein chromogranin-A identified the occurrence of chromaffin cells within all three zones of the human adrenal cortex, including the zona glomerulosa. In an ultrastructural analysis, cortical and chromaffin cells were found in all zones in direct apposition, providing the possibility for direct intercellular exchange. The close morphological colocalization of cortical and chromaffin cells revealed in this study may constitute the basis for the growing evidence of relevant intraadrenal paracrine mechanisms within the human adrenal gland.

Regulation of corticotropin receptor number and messenger RNA in cultured human adrenocortical cells by corticotropin and angiotensin II.

Abstract: The regulation of ACTH receptor binding sites and mRNA by ACTH and angiotensin II (A-II) was studied using cultured human adrenal fasciculata reticularis cells (HAC). These cells expressed two major ACTH receptor transcripts of 1.8 and 3.4 kb and three minor ones of 4, 7, and 11 kb. ACTH increased the levels of all these transcripts in a time- and dose-dependent manner. At a maximal concentration of 10(-8) M, ACTH enhanced 21- and 4-fold the level of ACTH receptor mRNA and the number of receptors per cell, respectively. Pretreatment of HAC with A-II produced a dose-dependent enhancement of ACTH receptor mRNA that was associated with an increase of both ACTH receptor number and responsiveness to this hormone. The effects of A-II were completely blocked by an AT1 receptor subtype antagonist but not by an AT2 antagonist. The effects of ACTH together with A-II on ACTH receptor mRNA were greater than those induced by each hormone alone. These results show that ACTH receptor number and mRNA are positively regulated by the two main hormones (ACTH and A-II) which, in vivo, regulate adrenocortical functions. In addition, they also show that HAC are a target for A-II. Thus, regulation of ACTH receptors may be one mechanism by which ACTH and A-II regulate adrenocortical functions under both normal and pathological conditions.

A novel cell layer without corticosteroid-synthesizing enzymes in rat adrenal cortex: Histochemical detection and possible physiological role

Abstract: A stratum of cells that did not contain both aldosterone synthase cytochrome P450 (cytochrome P450aldo) and cytochrome P45011 beta was found immunohistochemically between the zona glomerulosa and the zona fasciculata of the rat adrenal cortex. As cytochromes P450aldo and P45011 beta are the enzymes responsible for the biosynthesis of aldosterone and corticosterone, respectively, the cells there are considered to be incapable of synthesizing both aldosterone and corticosterone. Furthermore, the cells are regarded as inert in producing adrenal androgens, because rat adrenal cortex is known to lack steroid 17 alpha-hydroxylase. Thus, the stratum is composed of cells that do not synthesize any of the major corticosteroids in significant quantities. It was 5-10 cells thick under normal feeding conditions, but diminished to 4-5 cells thick when animals were maintained under Na restriction, which is known to stimulate the secretion of angiotensin-II. When the distribution of 5-bromo-2'-deoxyuridine-labeled nuclei in the adrenocortex from BrdU-administered rats was examined, the stained nuclei were concentrated in and around the cell stratum. The pulse-chase experiments showed that the labeled cells migrated out of this layer and into the zonae fasciculata-reticularis. On the basis of these findings, we suggest that the newly discovered cell layer is the progenitor cell zone of the rat adrenal cortex.

Clonal Composition of Human Adrenocortical Neoplasms

Abstract: The mechanisms of tumorigenesis of adrenocortical neoplasms are still not understood. Tumor formation may be the result of spontaneous transformation of adrenocortical cells by somatic mutations. Another factor stimulating adrenocortical cell growth and potentially associated with formation of adrenal adenomas and, less frequently, carcinomas is the chronic elevation of proopiomelanocortin-derived peptides in diseases like ACTH-dependent Cushing9s syndrome and congenital adrenal hyperplasia. To further investigate the pathogenesis of adrenocortical neoplasms, we studied the clonal composition of such tumors using X-chromosome inactivation analysis of the highly polymorphic region Xcen-Xp11.4 with the hybridization probe M27β, which maps to a variable number of tandem repeats on the X-chromsome. In addition, polymerase chain reaction amplification of a phosphoglycerokinase gene polymorphism was performed. After DNA extraction from tumorous adrenal tissue and normal leukocytes in parallel, the active X-chromosome of each sample was digested with the methylation-sensitive restriction enzyme HpaII. A second digestion with an appropriate restriction enzyme revealed the polymorphism of the region Xcen-Xp11.4 and the phosphoglycerokinase locus. Whereas in normal polyclonal tissue both the paternal and maternal alleles are detected, a monoclonal tumor shows only one of the parental alleles. A total of 21 female patients with adrenal lesions were analyzed; 17 turned out to be heterozygous for at least one of the loci. Our results were as follows: diffuse (n = 4) and nodular (n = 1) adrenal hyperplasia in patients with ACTH-dependent Cushing9s syndrome, polyclonal pattern; adrenocortical adenomas (n = 8), monoclonal (n = 7), as well as polyclonal (n = 1); adrenal carcinomas (n = 3), monoclonal pattern. One metastasis of an adrenocortical carcinoma showed a pattern most likely due to tumor-associated loss of methylation. In the special case of a patient with bilateral ACTH-independent macronodular hyperplasia, diffuse hyperplastic areas and a small nodule showed a polyclonal pattern, whereas a large nodule was monoclonal. We conclude that most adrenal adenomas and carcinomas are monoclonal, whereas diffuse and nodular adrenal hyperplasias are polyclonal. The clonal composition of ACTH-independent massive macronodular hyperplasia seems to be heterogeneous, consisting of polyclonal and monoclonal areas.

Distribution of catechol-O-methyltransferase enzyme in rat tissues.

Abstract: In the present study we show the distribution of catechol-O- methyltransferase (COMT) in various rat tissues with a highly specific antiserum prepared against recombinant rat COMT. Immunoprecipitation and immunocytochemical controls confirmed the COMT-specificity of the antibodies. The antiserum detected both the 24 KD soluble and the 28 KD membrane-bound forms of the enzyme. By immunohistochemical staining the COMT enzyme was found in most rat tissues. Staining was most intense in the liver and in the kidney, in agreement with previous studies and our immunoblotting results. In the gastrointestinal tract, epithelial cells of the stomach, duodenum, and ileum were immunoreactive for COMT. In pancreas, COMT immunoreactivity was found in insulin-producing β-cells and somatostatin-producing D-cells but not in glucagon-producing α-cells of the islets of Langerhans. In pituitary, COMT immunoreactivity was found in cleft cells, in pituicytes of the posterior lobe, and in the anterior lobe, partly in the same cells containing luteinizing hormone (LH). In other endocrine organs, COMT immunoreactivity was found in epithelial cells of the thyroid gland and in zona glomerulosa of the adrenal cortex. In the brain, brightest immunofluorescence was seen in ependymal cells of the cerebral ventricles and choroid plexus. Weak to moderate immunofluorescence was found in the neuropil of several brain areas, including striatum and cortex. Scattered small neurons in spinal sensory ganglia were also COMT immunoreactive. Previous immunocytochemical studies, enzyme activity determinations, and distribution of the COMT mRNA are in general agreement with the results presented here. The wide distribution of COMT in different tissues suggests an important role for this protein in inactivation of catechol compounds. (Less)

The neuroendocrinology of the adrenal cortex

Abstract: Although until relatively recently assumed to be devoid of innervation, there is now ample proof that the adrenal cortex receives specific neurones of several types. A general interpretation of their roles in the regulation of adrenocortical function has not been forthcoming, probably because of the variety of the different experimental approaches which have been used, and the heterogeneous observations which have been made. We here summarize the evidence which is available, and offer the view that neural inputs may provide fine tuning of the responses to systemic factors such as ACTH, through direct actions on specific adrenocortical cells. However, neural regulation also provides an integrative function, through actions on the flow of blood through the gland, which itself exerts a powerful influence on adrenocortical function.

Purification and characterization of a substrate protein for mitochondrial ATP-dependent protease in bovine adrenal cortex.

Abstract: We have purified SP-22, a substrate protein for mitochondrial ATP-dependent protease in bovine adrenal cortex. Native SP-22 showed an M(r) of 350,000 +/- 20,000, and was composed of more than 10 molecules of an M(r) 21,600 subunit. Subcellular and submitochondrial fractionation of adrenocortical tissues revealed that SP-22 was localized in the mitochondrial matrix, suggesting that SP-22 is a natural substrate for ATP-dependent protease, a matrix enzyme. The concentration of SP-22 in adrenocortical mitochondrial fractions was 16 +/- 3 micrograms/mg proteins (mean +/- SD, n = 6) as determined by radioimmunoassay using specific anti-SP-22 antibody. Adrenal cortex showed the highest concentration among the 15 bovine tissues tested, followed by liver, renal cortex, adrenal medulla, heart, and renal medulla. We determined the amino acid sequence of SP-22, which is composed of 195 amino acids. Amino acid 47 was not identified by the sequencer. FAB-mass spectrometry of AA47-AA55 fragment revealed that AA47 was cysteine-sulfinic acid (Cys-SO2H). By a homology search in the NBRF-PIR data base, SP-22 was found to be 91% homologous to murine erythroleukemia cell MER-5 protein, which may have an important role in the induction of differentiation. SP-22 was also homologous to the C22 component of alkyl hydroperoxide reductase in Salmonella typhimurium, thiol-specific antioxidant in Saccharomyces cerevisiae, and some other proteins. Since a segment around AA47 was highly conserved, this residue may be important for the biochemical functions of SP-22.

Localization of Steroidogenic Enzymes in Adrenal Cortex and Its Disorders

Abstract: Immunolocalization and in situ hybridization analysis of steroidogenic enzymes demonstrated the localization of steroidogenesis in the adrenal cortex and its disorders. The findings obtained provided new insights into adrenocortical hormonal metabolism, especially through establishing endocrine-pathological correlation.

Increased cortisol response to exogenous adrenocorticotropic hormone in chronically stressed pigs: influence of housing conditions.

Abstract: In a longitudinal experiment, the influence of tethered housing (a condition of chronic stress) on the reactivity of the adrenal cortex to exogenous ACTH was investigated in gilts. To that end, the plasma cortisol response to synthetic ACTH (1-24; 10 micrograms/kg of BW; i.v. bolus injection via a permanent catheter) was determined before and after prolonged tethered housing. Two systems for tethered housing were used, one more restrictive than the other with regard to possibilities for visual and tactile contacts with conspecifics and visual control over the environment. The ACTH treatment induced a marked, transient plasma cortisol response in all gilts studied, irrespective of their housing conditions. Long-term tethered housing increased the ACTH-induced cortisol response. Possible effects of the experimental procedure or age-related effects could be excluded, because in control gilts, which were housed loose during the entire experimental period, the cortisol response to ACTH remained unaltered. The chronic stress-induced increase in the ACTH-induced cortisol response was considerably more pronounced and persistent in gilts that were deprived of possibilities for social contacts with conspecifics and visual control over the environment than in gilts with such possibilities. These data indicate that in tethered gilts adaptational changes occur at the level of the adrenal cortex that affect the ACTH-induced adrenocortical response. In addition, not only physical restraint but also restriction of social contact and visual control play an important role in the development of these changes.

Gene expression, localization, and characterization of endothelin A and B receptors in the human adrenal cortex.

Abstract: Compelling evidence indicates that the endothelium-derived potent vasoconstrictor endothelin-1 (ET-1) stimulates aldosterone secretion by interacting with specific receptors. Although two different ET-1 receptors have been identified and cloned, the receptor subtype involved in mediating aldosterone secretion is still unknown. Accordingly, we wished to investigate whether the genes of ET-1 and of its receptors A and B are expressed in the normal human adrenal cortex. We designed specific primers for ET-1 and the ETA and ETB receptors genes and developed a reverse transcription polymerase chain reaction (RT-PCR) with chemiluminescent quantitation of the cDNA. In addition, we carried out 125I ET-1 displacement studies with cold ET-1, ET-3 and the specific ETA and ETB ligands BQ123 and sarafotoxin 6C. Localization of each receptor subtype was also investigated by autoradiography. Binding experiments were first individually analyzed by Scatchard and Hofstee plot and then coanalyzed by the nonlinear iterative curve fitting program Ligand. Histologically normal adrenal cortex tissue, obtained from kidney cancer patients (n = 7), and an aldosterone-producing adenoma (APA), which is histogenetically derived from the zona glomerulosa (ZG) cells, were studied. Results showed that the ET-1, ETA and ETB mRNA can be detected by RT-PCR in all adrenal cortices as well as in the APA. The best fitting of the 125I ET-1 displacement binding data was consistently provided by a two-site model both in the normal adrenal cortex (F = 22.1, P < 0.0001) and in the APA (F = 18.4, P < 0.0001). In the former the density (Bmax) of the ETA and ETB subtype was 2.6 +/- 0.5 pmol/mg protein (m +/- SEM) and 1.19 +/- 0.6, respectively. The dissociation constant (Kd) of ET-1, ET-3, S6C, and BQ-123 for each receptor subtype resulted to be within the range reported for human tissue for the ETA and ETB receptors. In the APA tissue the Bmax tended to be lower (1.33 and 0.8 pmol/mg protein, for the ETA and ETB, respectively) but the Kd were similar. Autoradiographic studies confirmed the presence of both receptor subtypes on the ZG as well as on APA cells. Thus, the genes of ET-1 and both its receptor subtypes ETA and ETB are actively transcribed in the human adrenal cortex. Furthermore, both receptor subtypes are translated into proteins in ZG and APA cells.

Origin of the angiotensin II secreted by cells.

Abstract: Circulating angiotensin II is unique in that it is formed in the blood by the interaction of circulating proteins. There are in addition many local renin-angiotensin systems in tissues in which angiotensin II is apparently secreted by various types of cells. This brief review considers the possible pathways for synthesis of locally produced angiotensin II in the brain, the anterior pituitary, the testes, the ovaries, the adrenal cortex, the kidneys, the heart, blood vessel walls, and brown and white fat. Synthesis by cells in culture is also reviewed. The possibility that certain cells contain a complete intracellular renin-angiotensin system is not ruled out, but there are problems with this hypothesis. Proteases other than renin may be involved, and there may be different pathways in different tissues. However, it appears that at least in some tissues, angiotensinogen is produced in one population of cells and transported in a paracrine fashion to other renin-containing cells, where it serves as the substrate for production of angiotensin II.

Role of the capacitative calcium influx in the activation of steroidogenesis by angiotensin-II in adrenal glomerulosa cells.

Abstract: Angiotensin-II (AngII)-induced Ca2+ influx in adrenal glomerulosa cells, a signal necessary for the stimulation of steroidogenesis by the hormone, is believed to involve two distinct mechanisms: 1) opening of voltage-operated Ca2+ channels, and 2) activation of a capacitative Ca2+ entry pathway that is dependent on calcium release from intracellular stores Nicardipine, a dihydropyridine calcium antagonist, has been used to investigate the role of these Ca2+ entry mechanisms in the steroidogenic response to AngII As demonstrated with the patch-clamp technique, micromolar concentrations of nicardipine completely blocked voltage-operated Ca2+ channel activity of both T- and L-types This agent similarly inhibited the rise of cytosolic free calcium concentration induced by potassium, but did not significantly affect the response to thapsigargin, an activator of the capacitative pathway Nicardipine reduced by only 22% the calcium influx stimulated by AngII, and the nicardipine-insensitive part of this response was abolished after exhausting the intracellular Ca2+ stores with thapsigargin Similarly, aldosterone secretion induced by AngII was only partially inhibited (40%) by nicardipine at concentrations that completely abolished the steroidogenic response to potassium Thapsigargin by itself was able to stimulate aldosterone production, an action highly potentiated by physiological concentrations of extracellular potassium These data strongly suggest that the major part of the calcium influx response to AngII, leading to aldosterone formation, involves a capacitative calcium entry pathway activated by the release of calcium from intracellular stores This mechanism of calcium influx could be responsible for some features of aldosterone response to the hormone, such as its poor sensitivity to dihydropyridines or its potentiation by potassium

Interleukin-6 messenger ribonucleic acid expression in human adrenal gland in vivo: new clue to a paracrine or autocrine regulation of adrenal function

Abstract: Interleukin-6 (IL-6) is an important mediator in the interaction of the hypothalamo-pituitary-adrenal axis with the immune system. Recently, a direct influence of IL-6 on adrenal steroidogenesis has been demonstrated. Therefore, we designed a study to determine whether IL-6 is expressed within the normal human adrenal gland. The combination of in situ hybridization and specific immunostaining was eminently suited to identify the cell types producing IL-6. IL-6 messenger ribonucleic acid occurred in the inner zone of the adrenal cortex in anti-17 alpha-hydroxylase-positive steroid cells. Also, CD68-positive macrophages in the zona reticularis showed a positive signal. No reaction was seen in chromaffin cells. We conclude that under normal conditions, IL-6 is expressed in specialized adrenocortical cells. Therefore, IL-6 may play an important role as a paracrine or autocrine factor in a local immune-adrenal interaction.

Is ouabain an authentic endogenous mammalian substance derived from the adrenal

Abstract: Ouabain has recently been reported to be an endogenous mammalian substance released by the adrenal cortex and present in normal plasma. We have attempted to confirm and extend this observation. Using a ouabain radioimmunoassay developed in this laboratory, we fractionated by high-performance liquid chromatography (HPLC) normal human plasma from healthy volunteers to determine the presence of ouabain immunoreactivity and compare this immunoreactivity with authentic ouabain. In most subjects no ouabain immunoreactivity that coeluted with authentic ouabain was observed. Some subjects had ouabain-immunoreactive material present at low levels, but it was largely attributable to cross-reactivity with diverse substances found not to be ouabain. Similar results were obtained after analysis of plasma collected from 10 patients entering a medical intensive care unit. Studies of serum-free medium conditioned by bovine adrenocortical cells showed some ouabain immunoreactivity. To determine whether this material might be a steroid product of cholesterol side-chain cleavage, we performed chemical blockade of steroidogenesis, which effectively suppressed progesterone production by these cells but had no consistent effect on ouabain immunoreactivity in this medium. Stimulation of steroidogenesis with 22-R-OH-cholesterol in bovine adrenocortical cells did not produce any increase in the ouabain immunoreactivity present in conditioned medium. Subsequent HPLC studies of ouabain immunoreactivity in bovine adrenocortical cell-conditioned medium indicated that authentic ouabain did not account for most of the ouabain immunoreactivity in serum-free medium. Studies with bovine adrenocortical cells incubated in a minimal salt and glucose medium indicated a small peak of immunoreactivity that may correspond to authentic ouabain.(ABSTRACT TRUNCATED AT 250 WORDS)

Augmented 17α-hydroxyprogesterone response to ACTH stimulation as evidence of decreased 21-hydroxylase activity in patients with incidentally discovered adrenal tumours (incidentalomas)

Abstract: Summary OBJECTIVE Recent studies have indicated that the syndrome of congenital adrenal hyperplasia due to 21-hydroxylase deficiency is closely associated with the development of benign adrenocortical tumours Tumour formation is thought to be a consequence of ACTH hypersecretion which results from the lack of glucocorticoid synthesis The aim of this study was to evaluate 21-hydroxylase activity in patients with an incidentally discovered adrenal mass ('Incidentaloma') without a history of congenital adrenal hyperplasia DESIGN A prospective study of 52 patients admitted to a single hospital PATIENTS Fifty-two consecutive subjects (mean age 56·0 years, range 24–81 years) with an incidentally discovered adrenal tumour were studied MEASUREMENTS The 21-hydroxylase reserve was assessed by intravenous bolus administration of 1-24 ACTH (tetracosactrin) with measurement of basal and stimulated serum 17α-hydroxyprogesterone (17-OHP) concentrations Impaired 21-hydroxylase activity was defined as an exaggerated 17-OHP response, with a 17-OHP increment exceeding 7-9nmolll Basal and stimulated cortisol concentrations, and basal ACTH were also measured RESULTS Baseline levels of 17-OHP were normal in 44 and elevated in 8 subjects In 37 patients (71·2%), the 17-OHP increment following ACTH administration exceeded 7·9nmol/l, demonstrating mildly decreased 21-hydroxylase activity In these subjects, the peak serum 17-OHP correlated with the tumour diameter In the patients with apparently normal 21-hydroxylase activity, no significant correlation was found between 17-OHP concentrations and tumour size All patients had a stimulated serum cortisol above 550 nmol/l reflecting intact adrenal gluco-corticoid reserve There were no other differences between the group with exaggerated and the group with normal 17-OHP Increment The tumours were removed from two women with augmented 17-OHP responses and this was followed by normalization of 17-OHP dynamics CONCLUSIONS Biochemical evidence for partial 21-hydroxyiase defiency is a common finding in patients with an adrenal Incidentaloma, even in the absence of a congential adrenal hyperpiasia history Exaggerated 17-OHP increment is not accompanied by decreased adrenal glucocorticoid reserve Normalization of the 17-OHP response after surgical treatment suggests that the phenomenon results from reduced 21-hydroxyiase activity in the tumour, which retains ACTH responsiveness

Impairment of Adrenocortical Function Associated with Increased Plasma Tumor Necrosis Factor-Alpha and Interleukin-6 Concentrations in African Trypanosomiasis

Abstract: African sleeping sickness (SS) is a severe, potentially lethal parasitic disease. The treatments of choice are the antiparasitic agents suramin, which is adrenotoxic, and/or melarsoprol. We evaluated the functional integrity of the hypothalamic-pituitary-adrenal (HPA) axis of patient with SS before, during, and after therapy with suramin and/or melarsoprol, in two sequential stages. First, we employed the standard adrenocorticotropic hormone (ACTH) 1-24 stimulation test (250 µg i.v.) to assess the maximal adrenocortical responsiveness of 69 patients with SS and 38 normal controls. We demonstrated paradoxically subnormal Cortisol responses before suramin therapy [net Cortisol response 60 min after stimulation: 10.5 ± 2.9 (mean ± SE) vs. 17.5 ± 1.0 µg/dl for controls, p = 0.004], with 27% of the patients falling within the adrenal insufficiency range (stimulated cortisol concentration <20 µg/dl). These responses subsequently and unexpectedly improved with suramin and/or melarsoprol therapy. Second, we performed a human corticotropin-releasing hormone (hCRH) test (100 µg i.v.) in 68 additional patients with SS and 14 control subjects to examine whether the glucocorticoid deficiency observed was primary and/or secondary. Compared to controls, the ACTH and Cortisol responses to hCRH were blunted (ACTH after 60 min: 29 ± 7 vs. 58 ± 8 pg/ml in controls, p = 0.014; cortisol: 15.2 ± 1.5 vs. 19.6 ± 0.7 µg/dl, p = 0.018), suggesting the presence of secondary adrenal insufficiency. There was improvement of both ACTH and cortisol responsiveness to hCRH with therapy, with cortisol recovery occurring before ACTH, suggesting an additional primary component of adrenal dysfunction in these patients. Plasma concentrations of tumor necrosis factor (TNF)-Α (16.0 ± 4.1 vs. 2.9 ± 1.4 pg/ml in controls, p = 0.003) and interleukin (IL)-6 (19.2 ± 7.3 vs. 1.3 ± 0.2 pg/ml, p = 0.0001), but not IL-1Β (2.0 ± 0.2 vs. 0.9 ± 0.2, p = NS), were elevated when adrenocrotical function impairment and disease activity were at their maximum, but gradually decreased into the normal range with therapy. We found a negative correlation between baseline cytokine concentrations and maximal Cortisol concentrations during hCRH testing (TNF-Α: r = –0.31, p = 0.003; IL-6: r = –0.34, p = 0.002). We conclude that unmedicated SS is associated with significant impairment of adrenocortical function which is reversed with suramin and/or melarsoprol therapy in the majority of patients. This impairment may be due to the elevated plasma cytokine concentrations, and may represent a natural adaptation of the HPA axis in inflammatory states. A controlled therapeutic trial is necessary to demonstrate whether supplemental glucocorticoids could be beneficial in SS.