Deoxycorticosterone Concentrations Research Articles

SAT264 Effects Of Coenzymes In Gene Therapy For Congenital Adrenal Hyperplasia With AAV Vectors Into Model Mice

Abstract Disclosure: Y. Naiki: None. M. Miyado: None. R. Horikawa: None. N. Katsumata: None. S. Takada: None. M. Fukami: None. Congenital adrenal hyperplasia (CAH) is due to defects of steroid synthetic enzymes, which includes microsomal and mitochondrial P450s and they need coenzymes respectively. Microsomal P450s include 21-hydroxylase need P450 oxidoreductase which coded by POR and ferredoxin and ferredoxin reductase, which coded by FDX and FDXR, respectively, work for mitochondrial P450s include 11β-hydroxylase. 21-hydroxylase deficiency (21-OHD), in which CYP21A2 is mutated or deleted, and 11β-hydroxylase deficiency (11β-OHD), which caused by mutated CYP11B1 are the top two cause of CAH. CAH are treated with oral steroid supplementation, but optimal control remains difficult. Thus, new therapeutic approaches are still needed. We succeeded gene therapy for model mice of CAH and iPS cell from CAH patients (1). In this study, we examined the effects of induction of coenzyme genes with an AAV vector into a model mouse. A 21OHD mouse were made by breeding with H-2aw18 haplotype mice and treated with subcutaneous corticosteroid injection until 3 weeks of age. A 11β-OHD mouse were made by using the CRISPR/Cas9 method. A type 9 AAV (AAV9) containing Cyp21a cDNA (AAV9-Cyp21a1) was constructed with pAAV-CMV-shuttle. AAV9 containing Por cDNA (AAV9-Por), Cyp11b1 cDNA (AAV9-Cyp11b1), Fdx (AAV9-Fdx), Fdxr (AAV9-Fdxr) were also made. An AAV9-Cyp21a1 and AAV9-Por were mixed and injected into limbs muscles of a 21-OHD mouse and AAV9-Cyp11b1 mixed with AAV9-Fdx and AAV9-Fdxr injected into limbs muscles of 11β-OHD mouse. Serum progesterone (P4) and deoxycorticosterone (DOC) concentrations for 21-OHD mouse and DOC and corticosterone (B) concentrations for 11β-OHD mouse were measured before and after injection. P4 and DOC concentrations were measured by LC-MS/MS. An AAV9-Cyp21a1 and AAV9-Por injected mouse showed improved steroid synthesis up to 7 months after AAV9 induction. Comparing with a mouse injected only AAV9-Cyp21a1, significant reduction of P4/DOC was observed after 4 weeks of injection but there was no difference in the effectiveness of Cyp21a1 and Por combined treatment at the end of observation. Instead of a 21-OHD mouse, a 11β-OHD mouse did not show significant improvement by AAV mixture injected. These results indicate that extra-adrenal induction of Cyp21a1 and Por may ameliorate steroid metabolism in 21-OHD model mice. Although this coenzyme combined therapy showed some effects, but no long-term improvement of the efficacy exhibited. Instead of microsomal P450s, extra-adrenal induction with coenzyme genes did not work enough for mitochondrial P450s defects. Coenzymes of P450s are necessary for their activity, but our results suggest that the coenzyme combined gene therapeutic strategies are needed to be improved. Reference: (1) Naiki et al., Hum Gene Ther. 2022 Aug;33(15-16):801-809. Presentation: Saturday, June 17, 2023

High androgen concentrations in follicular fluid of polycystic ovary syndrome women

BackgroundAccording to current definitions of Polycystic Ovary Syndrome (PCOS), hyperandrogenism is considered as a key element in the pathogenesis of this common endocrinopathy. However, until now, studies about ovarian androgen profile in women are very rare. Our aim was then to characterise the expression profile of the androgens in follicular fluid of 30 PCOS patients, and compare it to those of 47 Control women and 29 women with only polycystic ovary morphology on ultrasounds (ECHO group).MethodsA retrospective, single-centre cohort study was performed. The intrafollicular concentrations of the key androgens were assessed and correlated with the intrafollicular levels of some adipokines of interest. Androgens were quantified by mass spectrophotometry combined with ultra-high-performance liquid chromatography, while adipokine concentrations were measured by ELISA assays.ResultsIn PCOS patients, the intrafollicular concentrations of the androgens synthesised by ovarian theca cells, i.e., 17OH-pregnenolone, dehydroepiandrosterone, Δ4-androstenedione and testosterone, were significantly higher than those of the androgens of adrenal origin, and positively correlated with the main PCOS clinical and biological features, as well as with the adipokines mostly expressed in the follicular fluid of PCOS patients, i.e. resistin, omentin, chemerin and apelin. Conversely, Control women showed the highest levels of 17OH-progesterone, deoxycorticosterone and 11-deoxycortisol. Confirming these results, apelin levels were negatively associated with pregnenolone and deoxycorticosterone concentrations, while visfatin levels, which were higher in the Control group, negatively correlated with the Δ4-androstenedione and testosterone ones.ConclusionsPCOS is characterised by a selective increase in the intrafollicular levels of the androgens synthesised by theca cells, strengthening the hypothesis that ovarian hyperandrogenism plays a central role in its pathogenesis. Further, the significant correlation between the intrafollicular concentrations of the androgens and most of the adipokines of interest, including apelin, chemerin, resistin and omentin, confirms the existence of a close relationship between these two hormonal systems, which appear deeply involved in ovarian physiology and PCOS physiopathology.

Simultaneous steroids measurement in dogs with hyperadrenocorticism using a column-switching liquid chromatography-tandem mass spectrometry method.

We developed an analytical method using an on-line column-switching liquid chromatography with triple quadrupole mass spectrometry (LC/MS/MS) for quantifying multiple steroids in serum. Using the developed method, we evaluated the serum concentration of nine steroids (cortisol, corticosterone, cortisone, 11-deoxycortisol, 21-deoxycortisol, deoxycorticosterone, progesterone, 17α-OH-progesterone and aldosterone) in dogs with hyperadrenocorticism (HAC). Serum was mixed with stable isotope internal standards and thereafter purified by the automated column-switching system. The limit of detection ranged 2–16 pg/ml for nine steroids. In the baseline samples, five steroids (cortisol, corticosterone, cortisone, 11-deoxycortisol, and 17α-OH-progesterone) were detected in all dogs. The concentrations of cortisone, 11-deoxycortisol, and 17α-OH-progesterone in dogs with HAC (n=19) were significantly higher those in dogs without HAC (n=15, P<0.02). After the adrenocorticotropic hormone stimulation test, six steroids (cortisol, corticosterone, cortisone, 11-deoxycortisol, 17α-OH-progesterone, and deoxycorticosterone) were above the limit of quantification in all dogs. Cortisol, corticosterone, cortisone, and deoxycorticosterone concentrations of dogs with HAC were significantly higher than those of dogs without HAC (P<0.02). In addition, 11-deoxycortisol and 17α-OH-progesterone concentration was higher in dogs with HAC than in dogs without HAC (P=0.044 and P=0.048, respectively). The on-line column-switching LC/MS/MS would be feasible for measuring multiple steroids in dog serum. The results suggest that cortisone, 11-deoxycortisol, and 17α-OH-progesterone would be related to HAC. Further studies are warranted to assess the clinical feasibility of steroid profile in dogs with HAC.

Maternal exercise alters rat fetoplacental stress response: Minimal effects of maternal growth restriction and high-fat feeding

IntroductionFetal growth restriction complicates 10% of pregnancies and increases offspring (F1) risk of metabolic disorders, including obesity and gestational diabetes mellitus (GDM). This disease predisposition can be passed onto the next generation (F2). Importantly, the risk of pregnancy complications in obese women can be exacerbated by a stressful pregnancy. Exercise can reduce adiposity and improve health outcomes in obese women and those with GDM. This study investigated the impacts of maternal growth restriction, obesity, exercise, and stress on fetal and placental endocrine function. MethodsUteroplacental insufficiency (Restricted) or sham (Control) surgery was induced on embryonic day (E) 18 in F0 Wistar-Kyoto rats. F1 offspring were fed a Chow or High-fat (HFD) diet from weaning and, at 16 weeks, were randomly allocated an exercise protocol; Sedentary, Exercised prior to and during pregnancy (Exercise), or Exercised only during pregnancy (PregEx). Females were mated and further randomly allocated to either undergo (Stress), or not undergo (Unstressed), physiological measurements during pregnancy. On E20, F2 fetal plasma (steroid hormones), tissues (brain, liver), and placentae (morphology, stress genes) were collected. ResultsMaternal growth restriction and high-fat feeding had minimal impact on fetoplacental endocrine function. PregEx and Exercise increased cross-sectional labyrinth and junctional zone areas. PregEx, but not Exercise, increased fetal deoxycorticosterone concentrations and reduced placental Hsd11b2 and Nr3c2 gene abundance. Maternal stress increased fetal corticosterone concentrations in Sedentary HFD dams and increased placental cross-sectional areas in PregEx mothers. DiscussionPregEx and Stress independently dysregulates the endocrine status of the developing fetus, which may program future disease.

Expression of genes encoding mineralocorticoid biosynthetic enzymes and the mineralocorticoid receptor, and levels of mineralocorticoids in the bovine follicle and corpus luteum.

Unlike sex steroids, mineralocorticoids have attracted limited attention in ovarian physiology. Recent studies on primates have indicated possible local synthesis and action of mineralocorticoids in the ovary. Here, we examined developmental changes in the levels of mineralocorticoids and expression of genes encoding their biosynthetic enzymes and receptor in the bovine ovary. The follicles and corpora lutea (CL) were collected from F1 heifers. Expression levels of 21α-hydroxylase (CYP21A2), 11β-hydroxylase-1 (CYP11B1), and the mineralocorticoid receptor (NR3C2) in granulosa cells (GC), thecal layers (TL), and CL tissues were quantified by real-time PCR, whereas mineralocorticoids in the follicular fluid were measured by enzyme immunoassay (EIA). TL and GC expressed CYP21A2 and NR3C2, whereas CYP11B1 was expressed at very low or undetectable levels. The expression levels of these genes were not significantly different among small/large and healthy/atretic follicles but were higher in TL than in GC. CYP21A2 and NR3C2 were expressed in all CL stages with higher expression observed in the mid-stage. CYP11B1 expression was only apparent in the mid-stage CL. Aldosterone was detected in all follicles, and its concentration was not significantly different among the follicular groups. In paired large-healthy/atretic follicles, the concentration of deoxycorticosterone, a precursor of aldosterone, was approximately ten-fold higher than that of aldosterone and not significantly different between healthy and atretic follicles. In conclusion, the presence of mineralocorticoids and expression of NR3C2 in the bovine follicle together with the developmental change in the expression of CYP21A2, CYP11B1, and NR3C2 in the CL suggest possible endocrine/paracrine/autocrine roles of mineralocorticoids in the bovine ovary.

Sex‐dependent changes in neuroactive steroid concentrations in the rat brain following acute swim stress

Sex differences in hypothalamic‐pituitary‐adrenal (HPA) axis activity are well established in rodents. In addition to glucocorticoids, stress also stimulates the secretion of progesterone and deoxycorticosterone (DOC) from the adrenal gland. Neuroactive steroid metabolites of these precursors can modulate HPA axis function; however, it is not known whether levels of these steroids differ between male and females following stress. In the present study, we aimed to establish whether neuroactive steroid concentrations in the brain display sex‐ and/or region‐specific differences under basal conditions and following exposure to acute stress. Brains were collected from male and female rats killed under nonstress conditions or following exposure to forced swimming. Liquid chromatography‐mass spectrometry was used to quantify eight steroids: corticosterone, DOC, dihydrodeoxycorticosterone (DHDOC), pregnenolone, progesterone, dihydroprogesterone (DHP), allopregnanolone and testosterone in plasma, and in five brain regions (frontal cortex, hypothalamus, hippocampus, amygdala and brainstem). Corticosterone, DOC and progesterone concentrations were significantly greater in the plasma and brain of both sexes following stress; however, the responses in plasma were greater in females compared to males. This sex difference was also observed in the majority of brain regions for DOC and progesterone but not for corticosterone. Despite observing no stress‐induced changes in circulating concentrations of pregnenolone, DHDOC or DHP, concentrations were significantly greater in the brain and this effect was more pronounced in females than males. Basal plasma and brain concentrations of allopregnanolone were significantly higher in females; moreover, stress had a greater impact on central allopregnanolone concentrations in females. Stress had no effect on circulating or brain concentrations of testosterone in males. These data indicate the existence of sex and regional differences in the generation of neuroactive steroids in the brain following acute stress, especially for the 5α‐reduced steroids, and further suggest a sex‐specific expression of steroidogenic enzymes in the brain. Thus, differential neurosteroidogenesis may contribute to sex differences in HPA axis responses to stress.

Adaptations in Basal and Hypothalamic-Pituitary-Adrenal-Activated Deoxycorticosterone Responses Following Ethanol Self-administration in Cynomolgus Monkeys.

Acute ethanol activates the hypothalamic–pituitary–adrenal (HPA) axis, while long-term exposure results in a blunted neuroendocrine state, particularly with regards to the primary endpoint, cortisol, the primary glucocorticoid produced in the adrenal cortex. However, it is unknown if this dampened neuroendocrine status also influences other adrenocortical steroids. Plasma concentration of the mineralocorticoid and neuroactive steroid precursor deoxycorticosterone (DOC) is altered by pharmacological challenges of the HPA axis in cynomolgus monkeys. The present study investigated HPA axis regulation of circulating DOC concentration over the course of ethanol (4% w/v) induction and self-administration in non-human primates (Macaca fasciculata, n = 10). Plasma DOC, measured by radioimmunoassay, was compared at baseline (ethanol naïve), during schedule-induced polydipsia, and following 6-months of 22 h/day access to ethanol and water. The schedule induction of ethanol drinking did not alter basal DOC levels but selectively dampened the DOC response to pharmacological challenges aimed at the anterior pituitary (ovine corticotrophin-releasing hormone) and adrenal gland (post-dexamethasone adrenocorticotropin hormone), while pharmacological inhibition of central opioid receptors with naloxone greatly enhanced the DOC response during induction. Following 6 months of ethanol self-administration, basal DOC levels were increased more than twofold, while responses to each of the challenges normalized somewhat but remained significantly different than baseline. These data show that HPA axis modulation of the neuroactive steroid precursor DOC is markedly altered by the schedule induction of ethanol drinking and long-term voluntary ethanol self-administration. The consequences of chronic ethanol consumption on HPA axis regulation of DOC point toward allostatic modification of hypothalamic and adrenal function.

Changes in plasma concentrations of corticosterone and its precursors after ketoconazole administration in rats: An application of simultaneous measurement of multiple steroids using LC–MS/MS.

The adrenal gland is the most common toxicological target in the endocrine system, and inhibition of adrenal steroidogenesis by drugs can be fatal in humans. However, methods to evaluate the drug effect are limited. Recently, simultaneous measurement of multiple steroids, including precursors, has become possible. Here, we evaluated the usefulness of this simultaneous measurement for the evaluation of drug effects on adrenal steroidogenesis in vivo. For this purpose, we measured plasma concentrations of adrenal steroids in rats dosed with ketoconazole, a known inhibitor of adrenal steroidogenesis, and examined its relationship with the changes in histopathology and mRNA expression of steroidogenic enzymes in the adrenal gland. Ketoconazole (150mg/kg/day) was orally administered to male rats for 7 days. The adrenal weight was high, and the zona fasciculata/reticularis were hypertrophic with an accumulation of lipid droplets. mRNA expression of CYP11A1, a rate-limiting enzyme in adrenal steroidogenesis, was slightly high in the adrenal gland. Plasma concentration of deoxycorticosterone was markedly high, while there were no significant changes in that of corticosterone, progesterone, or pregnenolone. The changes in the adrenal gland and plasma concentration of steroids were thought to reflect inhibited metabolism of deoxycorticosterone to corticosterone through inhibition of CYP11B1, and compensatory reaction for the inhibition. The compensatory reaction was thought to have masked decrease of corticosterone. These results suggest that simultaneous measurement of multiple steroids can enable sensitive evaluation of drug effects on adrenal steroidogenesis in vivo, while providing insight into the underlying mechanism of the effect.

Brain steroid contents in the catfish Heteropneustes fossilis: sex and gonad stage-specific changes

Neurosteroids are those which are synthesized in the central nervous system independently of supply by peripheral endocrine glands. In the present study, brain contents of the steroid hormones, estradiol-17β (E(2)), testosterone (T), corticosteroids, and progestins were investigated in both male and female catfish Heteropneustes fossilis in prespawning (vitellogenic) and spawning (post-vitellogenic) phases using ELISA or HPLC. The data show that the measured steroid hormones showed both stage-specific and sex-related variations. Brain E(2) was significantly higher in males in the prespawning phase and in females in the spawning phase. Testosterone was significantly higher in males in comparison with females in the prespawning phase. Cortisol was significantly higher in the prespawning and spawning phases in males than in females. Corticosterone level was low in the brain. 21-deoxycortisol and deoxycorticosterone were significantly higher in the prespawning phase than in the spawning phase. Male brain recorded the highest concentration of deoxycorticosterone. Progesterone (P(4)) was high in the prespawning phase and low in the spawning phase in both sexes. Levels of 17-hydroxy-4-pregnene-3,20-dione and 17,20β-dihydroxy-4-pregnen-3-one (17,20β-DP) and the metabolites of P(4) were the highest in females in the prespawning phase. The stage-specific and sexual differences in the content of the steroids suggest their biosynthesis in the brain, which may have implications in brain functions, in addition to reproductive regulation.

Close examination of steroidogenesis disorders in a DOC- and progesterone-producing adrenocortical carcinoma

We report a case of hypertension, hypokalemia, and amenorrhea accompanying an adrenocortical carcinoma. A 27-year-old woman was admitted to our hospital because of a left adrenal incidentaloma. She presented with hypertension, hypokalemia, and amenorrhea; her plasma renin activity was low, but her plasma aldosterone concentration was normal, as were cortisol and androgens. By contrast, her serum concentrations of deoxycorticosterone (DOC), 18-hydroxydeoxycorticosterone, and progesterone were high, and her urinary steroid profile showed elevated secretion of 17-deoxysteroids and 11-deoxysteroids (progesterone, DOC, 11-dehydrocorticosterone, and 11-deoxycortisol), and 3beta-hydroxy 5-en steroids (pregnenolone, 17-hydroxypregnenolone, and DHEA). Decreased ratios of metabolites of (1) 17-OHpregnenolone to pregnenolone and 17-OHprogesterone to progesterone, (2) corticosterone to DOC and cortisol to 11-deoxycortisol, and (3) progesterone to pregnenolone, 17-OHprogesterone to 17-OHpregnenolone and androstenedione to DHEA suggested the impairment of 17alpha-hydroxylase, 11beta-hydroxylase, and 3beta-HSD activities, respectively. After the tumor was removed, levels of all adrenal steroids were normalized. Based on the Weiss criteria, the tumor was diagnosed as an adrenocortical carcinoma, and immunohistochemical analysis of steroidogenic enzymes revealed disorganized steroidogenesis in the tumor tissue. With adrenocortical carcinomas, heterogeneity of individual steroid producing enzymes within tumor cells can lead to hypersecretion of various steroid intermediates, even when steroid end products are within the normal range.

Hypokalaemic paresis, hypertension, alkalosis and adrenal‐dependent hyperadrenocorticism in a dog

Generalised paresis, severe hypokalaemia and kaliuresis, metabolic alkalosis and hypertension, characteristic of mineralocorticoid excess, were identified in a dog with hyperadrenocorticism due to a functional adrenocortical carcinoma. Aldosterone concentration was decreased and deoxycorticosterone concentration increased in the presence of hypokalaemia. These metabolic abnormalities resolved with resection of the carcinoma. Mineralocorticoid excess in dogs with hyperadrenocorticism is generally considered to be of little clinical significance but resulted in the acute presentation of this patient. The possible pathogenesis of mineralocorticoid excess in this case of canine hyperadrenocorticism is discussed.

Hypothalamic–pituitary–adrenal axis and ethanol modulation of deoxycorticosterone levels in cynomolgus monkeys

The metabolites of deoxycorticosterone (DOC) and progesterone, allotetrahydrodeoxycorticosterone and allopregnanolone, are potent endogenous neuroactive steroids that are increased in rodent brain and plasma after hypothalamic-pituitary-adrenal (HPA) axis activation by acute stress or ethanol administration. However, little data are available for male nonhuman primates. To determine DOC concentrations in plasma samples from 11 monkeys following challenge of the HPA axis with naloxone, corticotropin-releasing factor (CRF), dexamethasone, adrenocorticotropic hormone (ACTH) following dexamethasone pretreatment and ethanol. DOC levels were measured in monkey plasma by radioimmunoassay. DOC levels were increased after naloxone (125 microg/kg and 375 microg/kg, respectively) and CRF administration (1 microg/kg), and decreased following dexamethasone (130 microg/kg) administration. ACTH (10 ng/kg) challenge, 4-6 h after 0.5 mg/kg dexamethasone, and administration of ethanol (1.0 g/kg and 1.5 g/kg) had no effect on DOC concentrations. DOC levels were positively correlated with cortisol and ACTH levels after the naloxone (375 microg/kg), CRF, and ACTH challenges. Finally, the suppression of DOC levels measured after dexamethasone was negatively correlated with subsequent alcohol self-administration. These results suggest that DOC levels in monkeys are regulated by the HPA axis and may contribute to physiological responses following activation.

Preventing eclampsia

Fetal growth restriction complicates 10% of pregnancies and increases offspring (F1) risk of metabolic disorders, including obesity and gestational diabetes mellitus (GDM). This disease predisposition can be passed onto the next generation (F2). Importantly, the risk of pregnancy complications in obese women can be exacerbated by a stressful pregnancy. Exercise can reduce adiposity and improve health outcomes in obese women and those with GDM. This study investigated the impacts of maternal growth restriction, obesity, exercise, and stress on fetal and placental endocrine function.Uteroplacental insufficiency (Restricted) or sham (Control) surgery was induced on embryonic day (E) 18 in F0 Wistar-Kyoto rats. F1 offspring were fed a Chow or High-fat (HFD) diet from weaning and, at 16 weeks, were randomly allocated an exercise protocol; Sedentary, Exercised prior to and during pregnancy (Exercise), or Exercised only during pregnancy (PregEx). Females were mated and further randomly allocated to either undergo (Stress), or not undergo (Unstressed), physiological measurements during pregnancy. On E20, F2 fetal plasma (steroid hormones), tissues (brain, liver), and placentae (morphology, stress genes) were collected.Maternal growth restriction and high-fat feeding had minimal impact on fetoplacental endocrine function. PregEx and Exercise increased cross-sectional labyrinth and junctional zone areas. PregEx, but not Exercise, increased fetal deoxycorticosterone concentrations and reduced placental Hsd11b2 and Nr3c2 gene abundance. Maternal stress increased fetal corticosterone concentrations in Sedentary HFD dams and increased placental cross-sectional areas in PregEx mothers.PregEx and Stress independently dysregulates the endocrine status of the developing fetus, which may program future disease.

Regulation of steroid 21-hydroxylation by 17 beta-estradiol in rat liver: in vivo and in vitro study.

In various extraadrenal organs, progesterone (P4) is converted to deoxycorticosterone (DOC) by steroid 21-hydroxylation. To investigate the regulation of extraadrenal 21-hydroxylase activity by 17 beta-estradiol (E2), the following two experiments were performed. Exp. 1). Three-week-old male SD rats were testectomized (TX) and injected with E2 (1 mg) or sesame-oil s.c. Sham rats were treated with sesame-oil. In these groups the serum concentration of DOC and corticosterone (B), microsomal 21-Hydroxylase activity and the expression of P450c21 mRNA of the liver and the adrenals were analyzed. Exp. 2). Isolated rat hepatocytes were cultured and stimulated by E2, 10(-9) approximately 10(-5) M. 21-Hydroxylase activity of these cells was analysed by the rate of production of DOC in the medium containing P4. The results of experiment 1 showed that both serum DOC concentration and 21-hydroxylase activity in the liver microsomal fraction were increased by E2 injection, but the expression of P450c21 mRNA was not detected in the liver even after E2 injection. In experiment 2, the activity of steroid 21-hydroxylase in isolated rat hepatocytes was stimulated by E2 in a dose dependent manner. These data provided evidence that: in rats the liver was one sites of the extraadrenal steroid 21-hydroxylase activity which had been stimulated by E2. The results also suggested that this hepatic enzyme was a different enzyme from the steroid 21-hydroxylase P450c21 expressed in the adrenal gland.

Steroid characteristics of mineralocorticoid adrenocortical hypertension

Adrenocortical causes of hypertension are established by examining the mineralocorticoid hormones produced in the zona glomerulosa and zona fasciculata. In the zona glomerulosa, aldosterone excess leads to hypertension, hypokalemia, and suppressed plasma renin activity, with increased concentrations of urinary aldosterone (either as the 18-glucuronide or free aldosterone) as an index of its production. Identifying a tumor by computed tomography scan verifies the diagnosis of a correctable lesion. If no tumor is found, several maneuvers are used to identify primary adrenal hyperplasia, a disorder with autonomous aldosterone production, for which reduction of adrenal mass is curative. The zona fasciculata has two major pathways: the 17-deoxy pathway, where deoxycorticosterone (DOC) and corticosterone are the significant steroids, and the 17-hydroxy pathway, which leads to cortisol production. Tumors of the 17-deoxy pathway, DOC-producing adenomas, have increased concentrations of DOC and its precursor steroids, normal concentrations of cortisol, and suppression of aldosterone production secondary to suppression of the renin system. Two enzymatic defects in the zona fasciculata, 11 beta- and 17 alpha-hydroxylase deficiency, can be first readily identified by the virilization in the former, hypogonadal features in the latter. Steroid patterns are diagnostic. DOC is produced in excess in both deficiencies and is the cause of the hypertension. Deficient or impaired 11 beta-hydroxy steroid dehydrogenase in the apparent mineralocorticoid excess syndrome or after licorice ingestion retards the conversion of cortisol to inactive cortisone in the kidney, leading to mineralocorticoid hypertension; this leads to suppression of the renin system and subsequently of aldosterone.

Steroids during development of genetic hypertension in rats of Lyon strain.

The urinary excretion and the plasma concentration of deoxycorticosterone (DOC), corticosterone, 18-hydroxy-DOC (18-OH-DOC), aldosterone, and 19-nor-DOC were measured by specific radioimmunoassays in genetically hypertensive (LH), normotensive (LN), and low blood pressure (LL) male rats of the Lyon strains at two ages that characterize the development of their systolic blood pressure (SBP). When compared with both LN and LL controls, 5-wk-old LH rats exhibited an increased urinary DOC and decreased urinary corticosterone excretions, which were significantly related to the SBP level (r' = 0.618 and -0.520; n = 23; P less than 0.01 for DOC and corticosterone, respectively). In addition, the adrenal synthesis of LH rats was found to rely on an increased 18-hydroxylase activity as indicated by elevated urinary 18-OH-DOC/corticosterone and aldosterone/corticosterone associated with a lower 11-beta-hydroxylase activity shown by the decreased urinary corticosterone/DOC. Twenty-wk-old LH rats with fully developed hypertension exhibited normal urinary excretion of steroids and a decrease in plasma DOC concentration, which negatively correlated with the SBP level (r' = -0.574; n = 25; P less than 0.01). In conclusion, the present study demonstrates that in the Lyon model of genetically hypertensive rats, compared with two genetically different control strains and maintained under physiological unstressed conditions, the development of hypertension is associated with an increased urinary excretion of DOC. After the full development of their hypertension, the mineralocorticoid synthesis in LH rats returns to normal or low levels which could, however, remain inappropriately high for their sodium body content.

Types of adrenocorticoids and their effect on organophosphorus-induced delayed neuropathy in chickens

The present study examined the effects of a glucocorticoid and a mineralocorticoid on organophosphorus-induced delayed neuropathy (OPIDN) as previous investigations have indicated that an endogenous steroid with both properties could alter this syndrome in chickens. The glucocorticoid triamcinolone and the mineralocorticoid deoxycorticosterone were provided in the diet beginning 1 day before and continuing 10 days after triortho-tolyl phosphate (TOTP, 360 mg/kg po), phenyl saligenin phosphate (PSP, 2.5 mg/kg im), and diisopropyl phosphorofluoridate (DFP, 1 mg/kg sc). In a manner similar to that seen with corticosterone, a low concentration (0.1 ppm) of triamcinolone reduced and a high concentration (10 ppm) exacerbated clinical signs. Concentrations of deoxycorticosterone under 80 ppm also partially delayed or ameliorated ataxia induced by TOTP, PSP, and DFP, but a combination of 0.1 ppm triamcinolone and 80 ppm deoxycorticosterone was not more effective than triamcinolone alone. Peripheral nerve damage was noted in all chickens given organophosphorus compounds, whether or not they had been given corticoids. Both steroids induced hydroxylase activity, but effects on most other enzyme systems examined were unremarkable. High concentrations of triamcinolone (10 ppm) could, however, also reduce liver cytochrome P450 levels and liver cholinesterase activity. Exacerbation of OPIDN was most notable in chickens under highest stress, as indicated by elevated heterophil-to-lymphocyte ratios. The clinical, pathological, biochemical, and hematological indices of exposure to adrenocorticoids and agents inducing OPIDN in chickens were, therefore, similar for both a synthetic glucocorticoid and the endogenous steroid corticosterone.

Adrenocorticoid receptors in C6 glioma cells: Effects on cell growth

Rat C6 glioma cells contain two receptors for adrenocorticoids--the predominant glucocorticoid receptor and low densities of the Type I corticosteroid (mineralocorticoid) receptor. Nanomolar concentrations of deoxycorticosterone, corticosterone and aldosterone, which fully occupy Type I receptors, produced a slight stimulatory effect on C6 cell growth in serum-free media. However, spironolactone, a Type I receptor antagonist, and pregnenolone, which does not bind to Type I receptors, had similar effects. Therefore, the slight growth stimulation produced by low steroid concentrations is not mediated by Type I or glucocorticoid receptors, but may be due to an effect on cell membrane properties or other receptor-independent action. Occupation of glucocorticoid receptors by higher concentrations of corticosteroids inhibited C6 cell growth.

Effects of metyrapone and etomidate on adrenal function and growth rate in female rats.

Two inhibitors of adrenal steroidogenesis were examined to determine whether the growth rate of female rats could be improved by lowering circulating plasma corticosterone concentrations. The first inhibitor, etomidate, is a potent narcotic agent and was found to have no effect on plasma corticosterone and deoxycorticosterone (DOC) concentrations at sub-narcotic doses. Growth rate, food intake, food conversion efficiency and adrenal weight were also unaffected by the drug. The second inhibitor, metyrapone, was shown in acute studies to have two distinct actions. In 6-week-old female rats moderate doses of metyrapone (50 mg/kg) had a stimulatory action resulting in increased plasma DOC and corticosterone concentrations. Higher doses of metyrapone (150-300 mg/kg) were increasingly less selective, causing an increase in plasma concentration of DOC, but attenuating the increase in corticosterone concentration, presumably by inhibiting the 11 beta-hydroxylase enzyme which allows the conversion of DOC to corticosterone. In adult rats (greater than 12 weeks old) the classical response to metyrapone was observed. Plasma DOC concentrations were increased, while corticosterone levels were reduced in a dose-dependent manner. In 5-week-old rats treated chronically, metyrapone (300 mg/kg) had no effect on plasma corticosterone, but increased plasma DOC concentration, depressed food intake and reduced growth rate. In 8-week-old rats treated chronically, the high dose of metyrapone (300 mg/kg) required to lower peak plasma corticosterone concentrations produced toxic effects resulting in the death of three animals. The remaining animals recovered rapidly, and gained more weight than controls over the final 6 days of the experiment. However, this experiment was terminated before the treated animals were able to catch up with the controls.(ABSTRACT TRUNCATED AT 250 WORDS)

113 11β-HYDROXYLASE DEFICIENCY-CONGENITAL DEFECT OR TUMOR

In 3 children with low renin hypertension and virilism, the hormonal profile distinguished the patient with an adrenal tumor producing DOC and androgens from the other 2 patients who had a genetic deficiency of steroid 11β-hydroxylase (11β-OHD). Clinical presentations were remarkably similar as were the baseline serum concentrations of deoxycorticosterone (DOC). The major difference in the baseline hormones consisted of very high DHEA levels in the tumor patient in contrast to very high androstenedione (Δ4) levels in the patients with the genetic defect of 11β-hydroxylation. The stimulation and suppression tests also distinguished the tumor patient. DOC did not stimulate with ACTH nor suppress with dexamethasone (dex). The PRA rose with dex suppression only in the patients with the genetic defect. Serum aldosterone was undetectable in all 3 patients in the baseline period. Conclusion: Though patients with adrenal tumors and those with a genetic deficiency of 11β-OHD have similar clinical presentations, similar PRA, aldosterone and DOC, the androgens and their steroid responses to ACTH and dex are clear distinguising features. (nd=not detectable).