Effects of cadmium in vitro on microsomal steroid metabolism in the inner and outer zones of the guinea pig adrenal cortex

Studies were carried out to evaluate the effects of cadmium in vitro on microsomal steroid metabolism in the inner (zona reticularis) and outer (zona fasciculata and zona glomerulosa) zones of the guinea pig adrenal cortex. Microsomes from the inner zone have greater 21-hydroxylase than 17α-hydroxylase activity, resulting in the conversion of progesterone primarily to 11-deoxycorticosterone and of 17α-hydroxy progesterone principally to its 21-hydroxylated metabolite, 11-deoxycortisol. Microsomes from the outer zones, by contrast, have far greater 17α-hydroxylase and C_17,20-lyase activities than 21-hydroxylase activity. As a result, progesterone is converted primarily to its 17-hydroxylated metabolite, 17α-hydroxyprogesterone; and 17α-hydroxyprogesterone is converted principally to δ⁴-androstenedione, with only small amounts of 21-hydroxylated metabolites being produced. Addition of cadmium to incubations with inner zone microsomes causes concentration-dependent decreases in 21-hydroxylation and increases in 17α-hydroxylase and C_17,20-lyase activities, resulting in a pattern of steroid metabolism similar to that in normal outer zone microsomes. Cadmium similarly decreases 21-hydroxylation by outer zone microsomes but has no effect on the formation of 17-hydroxylated metabolites or on androgen (Δ⁴-androstenedione) production. In neither inner nor outer zone microsomes did cadmium affect cytochrome P-450 concentrations, steroid interactions with cytochrome(s) P-450, or NADPH–cytochrome P-450 reductase activities. The results indicate that cadmium produces both quantitative and qualitative changes in adrenal microsomal steroid metabolism and that the nature of the changes differs in the inner and outer adrenocortical zones. In inner zone microsomes, there appears to be a reciprocal relationship between 21-hydroxylase and 17α-hydroxylase/C_17,20-lyase activities which may influence the physiological function(s) of that zone.     

Unconjugated and sulfoconjugated steroids in plasma and zones of the adrenal cortex of the guinea pig

The following steroids were measured in their unconjugated and sulfoconjugated forms in plasma and in the outer and inner zones of the adrenal cortex of the guinea pig: pregnenolone, 17-hydroxypregnenolone, 21-hydroxypregnenolone, dehydroepiandrosterone and deoxycorticosterone. In plasma, pregnenolone and 21-hydroxypregnenolone were the predominant unconjugated steroids with concentrations 10-30 times higher than the other three steroids. Among the sulfoconjugated steroids, pregnenolone sulfate had a concentration 25-50 times higher than the other sulfoconjugates. For each steroid except 21-hydroxypregnenolone the sulfoconjugated form was present in a concentration 2-7 times higher than the unconjugated form. In the adrenal cortex, the content of 21-hydroxypregnenolone was significantly higher in the outer zone than in the inner zone and was present in amounts 3-100 times greater than the other unconjugated steroids in the outer zone. On the other hand, the content of pregnenolone was significantly greater in the inner zone than the outer zone, and was present in amounts 3-80 times greater than the other unconjugated steroids in the inner zone. With the exception of 21-hydroxypregnenolone and deoxycorticosterone, the steroid sulfoconjugates were significantly higher in the inner cortical zone. As in plasma, pregnenolone sulfate was the most abundant sulfoconjugated steroid. This report also describes preliminary studies concerning sulfurylated hydroxyl groups in different positions of 21-hydroxypregnenolone. The sulfoconjugate was prepared by using partially purified steroid sulfotransferase from the guinea pig adrenal. The results obtained indicated that of the total 21-hydroxypregnenolone conjugate formed, approximately 40% was the 21-sulfate and 20% the 3-sulfate, whereas 40% was non-hydrolyzable with the techniques used and was not further characterized.     

Regional distribution of microsomal drug and steroid metabolism in the guinea pig adrenal cortex

The regional distribution of steroid and drug metabolism was studied in intact cells and microsomal fractions obtained from the chromatically distinct inner (zona reticularis) and outer (zona fasciculata plus zona glomerulosa) zones of the guinea pig adrenal cortex. Cells isolated from the outer cortical zone produced far more cortisol than cells from the inner zone and cortisol production was stimulated by adrenocorticotropic hormone only in cells from the outer zone. Among the factors which may contribute to the greater cortisol production by the outer zone are a higher rate of 17 alpha-hydroxylation and ratio of 17 alpha- to 21-hydroxylase activities in that zone, both of which favor cortisol synthesis. In contrast, steroid 21-hydroxylase activity was far greater than 17 alpha-hydroxylase activity in microsomes obtained from the inner zone of the adrenal cortex. Microsomal metabolism of various xenobiotics such as benzo(a)pyrene and ethylmorphine proceeded far more rapidly in the inner than outer cortical zone. The zonal differences in metabolism appeared to result in part from differences in the ability of xenobiotics to interact with microsomal cytochromes P-450 in the two zones. The results indicate that the inner zone has a minor role in cortisol production by the adrenal cortex, but its involvement in the production of other steroids cannot be excluded. In contrast, the inner zone appears to have the major role in the metabolism of at least some xenobiotics which may account for its greater vulnerability to the toxic effects of chemicals requiring metabolic activation.     

Differential effects of adrenocorticotropic hormone on steroid hydroxylase activities in the inner and outer zones of the guinea pig adrenal cortex.

We have studied the effects of ACTH treatment on steroid hydroxylase activities in the inner (zona reticularis) and outer (zona fasciculata plus zona glomerulosa) zones of the guinea pig adrenal cortex. Animals received 5 or 10 U of ACTH daily for 6 days and enzyme activities were then assessed in isolated microsomal or mitochondrial preparations. In control animals, microsomal cytochrome P-450 concentrations were greater in the inner than outer zone, but mitochondrial P-450 levels were similar in the two zones. Microsomal 17 alpha-hydroxylase and mitochondrial 11 beta-hydroxylase activities were greater in the outer than inner zone, but microsomal 21-hydroxylase activity was greater in the inner zone. ACTH treatment decreased cytochrome P-450 concentrations in inner but not outer zone microsomes; mitochondrial P-450 levels were unaffected in both zones. ACTH caused a dose-dependent increase in inner zone 17 alpha-hydroxylase activity and decrease in 21-hydroxylase activity without affecting the activity of either enzyme in outer zone microsomes. ACTH also decreased 11 beta-hydroxylase activity in outer but not inner zone mitochondrial preparations. The net effect of ACTH treatment was to diminish the differences in steroid metabolism between the two zones. The results indicate that the effects of ACTH on steroid hydroxylase activities are both zone- and enzyme-dependent, suggesting the existence of multiple and independent regulatory mechanisms.     

Conversion of progesterone to corticosteroids by the midterm fetal adrenal and kidney

Midterm fetal adrenal and kidney tissue homogenates were incubated with 3H-progesterone (1 microM) and its conversion to te 3H-corticosteroids metabolites studied. Cortisol (36.3%) and corticosterone (4.7%) were isolated from the adrenal, and 11-deoxycortisol (32.5%) and deoxycorticosterone (21.1%) from the kidney. The results of these incubations confirmed the presence of 17- and 21-hydroxylase activities in both fetal tissues, and that of 11 beta-hydroxylase activity only in fetal adrenal tissue. We conclude that during pregnancy when progesterone levels are high, biosynthesis by the fetal kidney of 11-deoxycortisol, the most abundant corticosteroid formed by this tissue in this investigation, might provide to the fetal adrenal an important precursor for cortisol biosynthesis within the fetal compartment.     

Mitochondrial steroid metabolism in the inner and outer zones of the guinea-pig adrenal cortex

Previous investigations have demonstrated that cells isolated from the outer zone (zona fasciculata + zona glomerulosa) of the guinea-pig adrenal cortex produce far more cortisol than those from the inner zone (zona reticularis). Studies were carried out to compare mitochondrial steroid metabolism in the two zones. Protein and cytochrome P-450 concentrations were similar in outer and inner zone mitochondria. However, the rate of 11 beta-hydroxylation was significantly greater in the outer zone despite the fact that substrates for 11 beta-hydroxylation (11-deoxycortisol, 11-deoxycorticosterone) produced larger type I spectral changes in inner zone mitochondria. The apparent affinities of 11-deoxycortisol and 11-deoxycorticosterone for mitochondrial cytochrome(s) P-450 were similar in the two zones. In both inner and outer zone mitochondria, 11 beta-hydroxylation was inhibited by metyrapone but unaffected by aminoglutethimide. Cholesterol sidechain cleavage activity, measured as the rate of conversion of endogenous cholesterol to pregnenolone, was far greater in outer than inner zone mitochondria. Addition of exogenous cholesterol or 25-hydroxycholesterol to the mitochondrial preparations did not affect pregnenolone production in either zone. Addition of pregnenolone to outer zone mitochondria produced a reverse type I spectral change (delta A 420-390 nm), suggesting displacement of endogenous cholesterol from cytochrome P-450. In inner zone mitochondria, pregnenolone induced a difference spectrum (delta A 425-410 nm) similar to the reduced vs oxidized cytochrome b5 spectrum. A b5-like cytochrome was found to be present in the mitochondrial preparations. Prior reduction of the cytochrome with NADH eliminated the pregnenolone-induced spectral change in inner zone mitochondria but had no effect in outer zone preparations. The results suggest that differences in mitochondrial steroid metabolism between the inner and outer adrenocortical zones account in part for the differences in cortisol production by cells in each zone.     

An adrenocortical tumor secreting weak mineralocorticoids

An adrenocortical carcinoma (15.5 g) secreting excessive amounts of steroids with weak mineralocorticoid activity in a 25-year-old woman was studied with particular reference to its in vivo and in vitro secretions of steroids. Severe hypertension, occasional low serum potassium and suppressed PRA were the major clinical findings, and were improved with removal of the tumor. In the preoperative stage, plasma levels of 11-deoxycorticosterone, 18-hydroxy-11-deoxycorticosterone, corticosterone and 18-hydroxycorticosterone were all increased. However, the plasma level of aldosterone was repeatedly normal. Although plasma levels of pregnenolone, 17-hydroxypregnenolone, progesterone and 17-hydroxyprogesterone were very high, those of other late step steroids, i.e. 11-deoxycortisol, cortisol, dehydroepiandrosterone, androstenedione and testosterone were almost normal. From these findings, a major etiological role of weak mineralocorticoids such as 11-deoxycorticosterone, 18-hydroxycorticosterone and corticosterone in her hypertension was suggested. Pregnenolone and 17-hydroxypregnenolone in tumor tissue were increased, but 11-deoxycorticosterone, corticosterone, aldosterone, cortisol and adrenal androgens such as dehydroepiandrosterone, androstenedione and testosterone were below normal or low normal. In vitro production of 11-deoxycorticosterone, aldosterone or cortisol by the tumor tissue slices was very low and scarcely responded to synthetic ACTH.     

Strain Differences in Adrenal Microsomal Steroid Metabolism in Guinea Pigs

We recently reported that CYP2D16, a xenobiotic-metabolizing P450 isozyme, was expressed at higher levels in adrenal microsomes from inbred Strain 13 guinea pigs than in those from outbred English Short Hair (ESH) animals. Studies were done to determine if there also were strain differences in adrenal microsomal steroid metabolism. In both inner (zona reticularis) and outer (zona fasciculata plus zona glomerulosa) zone preparations of the adrenal cortex, 21-hydroxylase activities were greater in microsomes from ESH than from Strain 13 guinea pigs. By contrast, 17-hydroxylase activities were similar in the two strains. In both strains, 21-hydroxylase activities were greater in inner than outer zone microsomes, but the opposite was found for 17-hydroxylase activities (outer>inner). Northern and Western analyses revealed higher levels of CYP21 mRNA and protein in adrenals from ESH than Strain 13 guinea pigs, but there were no strain differences in CYP17 mRNA or protein concentrations. Despite the zonal differences in adrenal 17-hydroxylase and 21-hydroxylase activities, CYP17 and CYP21 mRNA and protein levels were similar in the inner and outer zones within each strain of guinea pig. The results demonstrate strain differences in microsomal steroid metabolism that are explained by differences in CYP21 expression. By contrast, the zonal differences in steroid hydroxylase activities may be attributable to post-translational mechanisms.     

Maturational changes in steroidogenesis in the inner and outer zones of the guinea pig adrenal cortex

Studies were done to determine the effects of age on steroidogenesis in the inner (zona reticularis) and outer (zona fasciculta plus glomerulosa) zones of the guinea pig adrenal cortex. In 35-day-old animals, cortisol production by adrenal outer zone cells was approximately twice as great as that by inner zone cells. With aging, cortisol secretion by inner zone cells decreased to very low levels, but there was no detectable change in the capacity for cortisol production by the outer zone. However, the outer zone comprised a progressively decreasing fraction of the total adrenal mass in older animals. To determine the basis for the decline in cortisol production by inner zone cells with aging, the activities of several steroidogenic enzymes were determined. Microsomal 21-hydroxylase activity was greater in the inner than outer zone but was not significantly affected by age. By contrast, 17-hydroxylase activity was greater in the outer zone at all ages, and decreased with aging in the inner but not the outer zone. Mitochondrial cholesterol sidechain cleavage and 11β-hydroxylase activities were also higher in the outer than inner zone and declined in the zone only in older animals. The decrease in inner zone cholesterol sidechain cleavage activity with aging was proportionately greater than the age-dependent changes in other enzyme activities. The results indicate that the effects of aging on steroidogenesis are both zone- and enzyme-specific. The overall decline in cortisol secretion by the guinea pig adrenal cortex with aging is attributable to both a decrease in cortisol production by the cells of the zone reticularis and a disproportionate increase in the mass of the gland comprised by this zone. The decrease in cortisol secretion correlates closely with a decline in cholesterol sidechain cleavage activity in the zona reticularis, and may be causally related.     

Corticoid formation by the monkey fetal adrenal: Evaluation of 17-, 21- and 11-hydroxylase activities

There is indirect evidence that cortisol synthesis in the fetal rhesus monkey adrenal gland is limited at Day 135 of gestation but increases thereafter. This study was conducted to ascertain whether a reduced synthetic capacity is caused by a deficiency in 17-, 21- or 11-hydroxylase activity. For the sake of comparison 11- and 21-hydroxylases were also estimated in adult adrenals. 11-, 21-Hydroxylases were measured in the entire adrenal by the oxidation of NADPH by mitochondria and microsomes, respectively. 17-Hydroxylase was evaluated in outer and inner regions of the fetal gland by the formation of [³H]17-hydroxyprogesterone, -11-deoxycortisol, -cortisol and -androstenedione from [³H]progesterone. The maximum velocity of both the 11- and 21-hydroxylase was similar in fetal and adult glands indicating that corticoid formation in the fetus is not constrained by levels of these enzymes.[³H]Progesterone was extensively metabolized to -17-hydroxyprogesterone, -androstenedione, -11-deoxycortisol and -cortisol by homogenates from both regions of the fetal adrenal. The ratio of [³H]-cortisol to [³H]11-deoxycortisol was consistently higher in incubations of the inner glandular area. Together, these findings indicate that 17-hydroxylase is also active at Day 135 and that the 11-hydroxylase may be more concentrated in the fetal cortex. These data suggest in addition that the restriction in cortisol formation occurs at a step prior to the metabolism of progesterone to cortisol.     

Steroid biosynthesis by zona glomerulosa-fasciculata cells in primary culture of guinea-pig adrenals

Steroidogenesis was studied in guinea-pig glomerulosa-fasciculata cells maintained in primary culture for up to 7 days. The basal secretion which remained stable for the first 2 days in culture rapidly rose to reach a plateau on day 4 at levels 6-7-fold higher than those observed during the first 2 days of culture while the maximal response to ACTH in terms of cortisol and androstenedione secretion was fairly stable throughout the 7-day period. Exposure of glomerulosa-fasciculata cells to ACTH caused a stimulation of pregnenolone, 17-hydroxypregnenolone, progesterone, 17-hydroxyprogesterone, corticosterone, 11-deoxy-corticosterone, 11-deoxycortisol, cortisol, dehydroepiandrosterone, androstenedione, 11 beta-hydroxyandrostenedione and aldosterone while, after 48 h of incubation, a marked accumulation of end-products, namely cortisol and 11 beta-hydroxyandrostenedione, was observed. The half-maximal steroidogenic response to ACTH occurred at concentrations varying between 1.7 x 10(-11) and 1.1 x 10(-10) mol/l for the 12 steroids examined. Addition of 8-bromoadenosine 3', 5'-cyclic monophosphate stimulated steroid secretion in a dose-dependent manner. Maximal response to 8-bromoadenosine 3', 5'-cyclic monophosphate was obtained at 1 mmol/l, and no further rise of steroid secretion was observed after addition of ACTH. Incubation of glomerulosa-fasciculata cells with labeled corticosterone, cortisol and androstenedione indicates that only androstenedione can be converted into 11 beta-hydroxyandrostenedione, thus suggesting that this end-product is a good parameter of the C-19 steroid production by guinea-pig glomerulosa-fasciculata cells in primary culture. The present data confirm that guinea-pig glomerulosa-fasciculata cells in primary culture provide an interesting model for the study of the regulation of C-19 steroid formation by the adrenals.     

Fast and direct quantification of adrenal steroids by tandem mass spectrometry in serum and dried blood spots

We present a fast and reproducible method for steroid analysis (corticosterone, deoxycorticosterone, progesterone, 17alpha-hydroxyprogesterone, 11-deoxycortisol, 21-deoxycortisol, androstenedione, testosterone, dihydrotestosterone and cortisol) in small volumes of serum and in dried blood spot samples by LC-MS/MS. No derivatisation was needed. LC separation was achieved by using an Atlantis C18 column and water-methanol-formic acid gradient as a mobile phase and a flow rate of 250 microL/min over a run time of 6 min. Steroids were measured in MRM mode with electrospray interface (positive ion mode). Validation showed excellent precision, sensitivity, recovery and linearity with coefficients of determination r2>0.992.     

Chemical structure of corticosteroids and its relationship with their acute induction of lordosis in the female rat

The intravenous injection of several corticosteroids in the spayed estrogen-primed rat resulted, 5 min later, in a remarkable induction of lordosis response for deoxycorticosterone and 11-deoxycortisol. A lower but important effect was induced with corticosterone and 17 alpha-hydroxyprogesterone. A chemical structure-biological effect analysis showed that C-21 hydroxylation (e.g., deoxycorticosterone, 11-deoxycortisol, and corticosterone) induces the behavioral effect. However, C-11 or C-17 hydroxylation alone promotes the lordosis response to a lesser degree (corticosterone and 17 alpha-hydroxyprogesterone, respectively). However, the effect of these latter compounds is significant. From the results, it is concluded that deoxycorticosterone and 11-deoxycortisol along with other steroids such as 20 alpha-hydroxyprogesterone are suitable candidates for synergizing the well-known estrogen-progesterone induction of lordotic behavior.     

18-Hydroxy-11-deoxycortisol: a new steroid isolated from incubations of the adrenal with 11-deoxycortisol

Cortisol has been shown to be metabolized in the zona glomerulosa of the adrenal gland through the same pathway involving the cytochrome P-450, corticosterone methyl oxidase by which corticosterone is transformed to 18-hydroxycorticosterone and aldosterone. When cortisol is the precursor, 18-hydroxycortisol and 18-oxocortisol are formed. 18-Hydroxycortisol can also be made at a similar rate in the bovine zona fasciculata and reticularis as in the zona glomerulosa. We studied the possibility that the formation of 18-hydroxycortisol in the zona fasciculata and reticularis might be through a different pathway involving initial 18-hydroxylation of 11-deoxycortisol before 11 beta-hydroxylation. Rat adrenal capsules or cores were incubated with 10 micrograms of cortisol or 11-deoxycortisol and the formation of 18-hydroxycortisol was measured by radioimmunoassay. Both capsules and cores transformed 11-deoxycortisol to 18-hydroxycortisol, but cortisol was only transformed in the capsular portion. Sixty-two rat adrenals were incubated with 10 mg of 11-deoxycortisol and the putative steroid, 18-hydroxy-11-deoxycortisol, was purified by TLC and HPLC and subjected to gas chromatography mass spectrometry. The mass spectra indicated that the steroid isolated was indeed 18-hydroxy-11-deoxycortisol. The function of this steroid is still unknown.     

The topology of the mitochondrial 11β-hydroxylase system in bovine adrenal cortex

Binding of deoxycorticosterone to cytochrome P-450 of the 11β-hydroxylase system in adrenal cortex mitochondria was inhibited by the nonpenetrating protein reagent diazobenzenesulfonate in damaged but not in intact mitochondria. The slowly penetrating hydrophilic substrate deoxycorticosterone 21-sulfate showed a slow binding to cytochrome P-450 as compared to the hydrophobic nonesterified steroid. In contrast, the esterified and nonesterified steroids bound equally fast in sonicated, aged or lysolecithin-treated mitochondria. These data imply that the steroid substrates must penetrate the inner mitochondrial membrane to interact with the 11β-hydroxylase system.     

Calf adrenocortical fasciculata cells secrete aldosterone when placed in primary culture

Aldosterone production occurs in the outer area of the adrenal cortex, the zona glomerulosa. The glucocortocoids cortisol and corticosterone, depending upon the species, are synthesized in the inner cortex, the zona fasciculata. Calf zona glomerulosa cells rapidly lose the ability to synthesize aldosterone when placed in primary culture unless they are incubated in the presence of the antioxidants butylated hydroxyanisol and selenous acid, the radioprotectant DMSO, and the cytochrome P-450 inhibitor metyrapone. In the presence of these additives, calf zona fasciculata cells in primary culture synthesize aldosterone at rates which can approach those from cells isolated from the zona glomerulosa. Calf zona glomerulosa and fasciculata cells both responded well to ACTH and angiotensin II, but the zona fasciculata cells respond very poorly compared to glomerulosa cells to increased potassium in the media. Rat zona fasciculata cells in primary culture under similar conditions did not synthesize aldesterone, suggesting that the regulation of the expression of the enzymes responsible for the biosynthesis of aldosterone in the two species is different. Two distinct cytochrome P-450 cDNAs which hydroxylate deoxycorticosterone at the 11β position have been described in the rat, human and mouse. Both cytochrome P-450 cDNAs have been cloned and expressed in non-steroidogenic cells, but only one is expressed in the zona glomerulosa and only this glomerulosa cytochrome P450 can further hydroxylate deoxycorticosterone to generate aldosterone. Two bovine adrenal cDNAs have been described with 11β-hydroxylase activity and their expression products in transiently transfected COS cells can convert deoxycorticosterone into aldosterone. Both enzymes are expressed in all zones of the adrenal cortex. Zonal regulation of aldosterone synthesis in the bovine adrenal gland may be due to an 11β-hydroxylase with aldosterone synthesizing capacity which has not yet been isolated. Alternatively, a single enzyme might be responsible for the several hydroxylations in the pathway between deoxycorticosterone and aldosterone and zonal synthesis might be controlled by unknown factors regulating the expression of C-18 hydroxylation. The incubation of zona fasciculata with antioxidants and metyrapone results in atypical expression of this activity by an unclear mechanism.     

An alternative explanation of hypertension associated with 17α-hydroxylase deficiency syndrome

The syndrome of 17α-hydroxylase deficiency is due to the inability to synthesize cortisol and is associated with enhanced secretion of both corticosterone and 11-deoxy-corticosterone (DOC). In humans, corticosterone and its 5α-Ring A-reduced metabolites are excreted via the bile into the intestine and transformed by anaerobic bacteria to 21-dehydroxylated products: 11β-OH-progesterone or 11β-OH-(allo)-5α-preganolones (potent inhibitors of 11β-HSD2 and 11β-HSD1 dehydrogenase). Neomycin blocks the formation of these steroid metabolites and can blunt the hypertension in rats induced by either ACTH or corticosterone. 3α,5α-Tetrahydro-corticosterone, 11β-hydroxy-progesterone, and 3α,5α-tetrahydro-11β-hydroxy-progesterone strongly inhibit 11β-HSD2 and 11β-HSD1 dehydrogenase activity; all these compounds are hypertensinogenic when infused in adrenally intact rats.Urine obtained from a patient with 17α-hydroxylase deficiency demonstrated markedly elevated levels of endogenous glycyrrhetinic acid-like factors (GALFs) that inhibit 11β-HSD2 and 11β-HSD1 dehydrogenase activity (>300 times greater, and >400 times greater, respectively, than those in normotensive controls). Thus, in addition to DOC, corticosterone and its 5α-pathway products as well as the 11-oxygenated progesterone derivatives may play a previously unrecognized role in the increased Na⁺ retention and BP associated with patients with 17α-hydroxylase deficiency.     

ACTH sensitivity of isolated human pathological adrenocortical cells: variability of responses in aldosterone, corticosterone, deoxycorticosterone and cortisol production

In vitro aldosterone, deoxycorticosterone, corticosterone and cortisol production of human adrenocortical cells derived from adenomas (Conn's syndrome, Cushing's syndrome), from hyperplastic adrenals (Cushing's syndrome) and from adrenals surrounding aldosteronoma are described. Cells from adenomas causing either Cushing's syndrome or Conn's syndrome harboured the highest basal and ACTH-stimulated corticosteroid production. Adrenocortical cells derived from micronodular hyperplasia causing Cushing's syndrome and cells from cortisol producing adenoma displayed predominantly cortisol and corticosterone secretion both under basal conditions and following stimulation with ACTH. Aldosteronoma cells showed highly variable aldosterone, deoxycorticosterone, corticosterone and cortisol response to ACTH. However, in aldosteronoma cell suspensions, the basal and ACTH-stimulated ratios of aldosterone to cortisol were increased when compared to ratios of steroids produced by cells from other adrenal tissues. Chronic treatment with spironolactone of patients with Conn's syndrome before surgery was associated with a decreased ratio of aldosterone to corticosterone, revealing that 18-hydroxylase in aldosteronoma cells may be inhibited during long-term therapy. Non-tumorous cells isolated from adrenals surrounding aldosteronoma displayed less aldosterone prior to and after stimulation with ACTH than aldosteronoma cells.     

Effects of cyproterone acetate on adrenal steroidogenesis in vitro

The effects of cyproterone acetate (CA) on steroidogenesis in isolated guinea-pig adrenal cells have been investigated by measuring the production of cortisol, its immediate precursors (11-deoxycortisol and 17-hydroxyprogesterone), and adrenal androgens (delta 4-androstenedione and dehydroepiandrosterone). Used at a dose of 2 micrograms/ml, CA provoked a sharp drop in the production of cortisol, aldosterone and 11-deoxycortisol. By contrast, 17-hydroxyprogesterone, delta 4-androstenedione and dehydroepiandrosterone were increased, which suggests that 21-hydroxylase activity is inhibited. With concentrations above 2 micrograms/ml CA, it would seem to be the 3-beta-ol-dehydrogenase-delta 4,5-isomerase complex that is affected, since dehydroepiandrosterone exhibited a sudden increase, whereas 17-hydroxyprogesterone and delta 4-androstenedione showed a relative decrease. The enzymatic system or systems involved therefore appear to be linked to the concentration of CA used but, whatever the case, the drop in cortisol production is accompanied by a decrease in aldosterone and an increase in adrenal androgen levels.