Alpha-human atrial natriuretic polypeptide inhibits 19-hydroxy-androstenedione secretion by human adrenal cells

We investigated the effect of ACTH, angiotensin II (AII), and alpha-human atrial natriuretic polypeptide (alpha-hANP) which plays an important role of water-electrolytes balance, on 19-hydroxyandrostenedione (19-hydroxyandrost-4-ene-3,17-dione, 19-OH-A-dione) secretion by cultured human adrenal cells. 19-OH-A-dione in culture media was measured using a specific RIA. Basal 19-OH-A-dione secretion by adrenal cells was 0.69 +/- 0.08 ng/3h/10(6) cells and significantly rose to 1.17 +/- 0.14 ng/3h/10(6) cells in the presence of ACTH, but not in the presence of A II. These results demonstrate that 196-OH-A-dione is directly secreted from adrenal cells. alpha-hANP significantly inhibited both basal and ACTH-stimulated 19-OH-A-dione secretions, as well as aldosterone. These results demonstrate that alpha-hANP inhibits aldosterone activity by means of the inhibition of both aldosterone and 19-OH-A-dione (an aldosterone amplifier) secretion by adrenal cells.     

19-hydroxyandrostenedione and 6 beta-hydroxyandrostenedione: new steroids regulated by the renin-angiotensin system in man

The changes of plasma 19-hydroxyandrostenedione (19-OH-A-dione) and 6 beta-hydroxyandrostenedione (6 beta-OH-A-dione) during the infusion of angiotensin II were evaluated and were compared with those of plasma aldosterone in man. Angiotensin II was infused into 5 normal subjects with an infusion pump at rates of 0.5, 1.0, 2.0 and 4.0 ng/kg per min. Each dose was infused for 20 min. Plasma 19-OH-A-dione rose significantly following the infusion of angiotensin II at a rate of 0.5 ng/kg per min and plasma 6 beta-OH-A-dione rose significantly following the infusion of angiotensin II at a rate of 1.0 ng/kg per min. In contrast, plasma aldosterone did not change significantly until the infusion rate reached 4.0 ng/kg per min. These results indicate that the secretion of 19-OH-A-dione and 6 beta-OH-A-dione is under the control of angiotensin II and the release of 19-OH-A-dione and 6 beta-OH-A-dione is induced earlier by the smaller doses of angiotensin II prior to the secretion of aldosterone. As 19-OH-A-dione and 6 beta-OH-A-dione amplify the action of aldosterone in bioassays using adrenalectomized rats and work as sodium-retaining and hypertensinogenic agents in intact rats, they are newly recognized biologically active steroids which are regulated by the renin-angiotensin system in man.     

Comparison of residual C-19 steroids in plasma and prostatic tissue of human, rat and guinea pig after castration: unique importance of extratesticular androgens in men

The concentrations of dehydroepiandrosterone (DHEA), its sulfate (DHEAS), androstenedione (A-dione), testosterone (T) and dihydrotestosterone (DHT) have been measured before and after castration in men and two animal models, namely the rat and the guinea pig. In adult men, the pre-castration levels of plasma DHEAS and DHEA were measured at 1839 +/- 320 and 2.4 +/- 0.5 ng/ml, respectively, while in both animal models, the concentrations of these two steroids were below 0.3 ng/ml. Orchiectomy in men reduced plasma T and DHT levels from 2.9 +/- 0.1 and 0.60 +/- 0.10 to 0.42 +/- 0.21 and 0.05 +/- 0.01 ng/ml (P less than 0.01), respectively, while there was no significant effect observed on DHEAS, DHEA and A-dione levels. By contrast, castration in the rat reduced the low levels of circulating DHEA and A-dione below the detection of the radioimmunoassay (RIA) used. In castrated guinea pig, a small quantity of plasma A-dione (0.07 +/- 0.02 ng/ml) was measured while DHEA was undetectable. Moreover, in the rat and guinea pig, plasma T and DHT levels became undetectable. Following administration of the antiandrogen Flutamide for two weeks in the castrated rat and guinea pig, prostate weight was not further reduced, thus indicating that there is no significant androgenic activity left following castration of these two species. In fact, castration in the rat and guinea pig caused a decrease in prostatic levels of DHT from 4.24 +/- 0.351 and 9.42 +/- 1.43 ng/g, respectively, to undetectable levels. In men, on the other hand, the prostatic DHT levels were only inhibited from 5.24 +/- 0.59 to 2.70 +/- 1.50 ng/g, respectively. As expected, when Flutamide was administered to the rat and the guinea pig, the levels of prostatic steroids remained undetectable while, in men, the DHT content in the prostate was further reduced to undetectable values. In summary, the plasma levels of DHEAS, DHEA, delta 4-dione are markedly different between men and both animal models used and furthermore, measurements of prostatic levels of androgens suggest that the high plasma levels of these steroids are likely responsible for the presence of important amounts of DHT in human prostate after castration.     

Proliferative effect of androst-4-ene-3,17-dione and its metabolites in the androgen-sensitive LNCaP cell line

As a therapeutic approach for the treatment of androgen-sensitive diseases, it would be tempting to lower the level of the potent androgens testosterone (T) and dihydrotestosterone (DHT) by using inhibitors of type 3 and type 5 17beta-hydroxysteroid dehydrogenases (17beta-HSDs). However, the efficiency of such a strategy will be optimal only if androst-4-ene-3,17-dione (Delta4-dione), the precursor of T, does not possess per se agonist activity on the androgen receptor (AR). To determine if the proliferative effect previously observed on AR(+) cells for Delta4-dione originates from its direct (per se) action on AR or from its transformation into a metabolite, we started a series of experimentations using the human prostate cancer LNCaP cell line, which expresses a highly sensitive AR. By real-time RT-PCR analysis, we detected type 1 5alpha-reductase (5alpha-R), a small amount of type 5 17beta-HSD, but not type 2 5alpha-R nor type 3 17beta-HSD. We then studied the transformation of labeled Delta4-dione in LNCaP cells after 1-7 days and the most important metabolite detected was 5alpha-androstane-3,17-dione (A-dione), which is the product of 5alpha-R activity. We measured only low levels of androsterone (ADT) and epi-ADT. This result was next confirmed by using an inhibitor of 5alpha-R that completely inhibited the transformation of Delta4-dione into A-dione, and consequently into ADT and epi-ADT. The proliferative effect of Delta4-dione (carefully purified) on LNCaP (AR(+)) cells was next determined in presence or absence of the 5alpha-R inhibitor. Although the cells proliferate in the presence of Delta4-dione only, no cell proliferation was observed with a combination of Delta4-dione and 5alpha-R inhibitor, suggesting that Delta4-dione is not androgenic per se. We next determined that A-dione and epi-ADT stimulated cell growth with the same pattern and potency as Delta4-dione, whereas ADT had a 3.5-fold lower proliferative activity. In conclusion, Delta4-dione is not in itself an agonist steroid on LNCaP (AR(+)) cells, and its proliferative activity appears to be mediated by its transformation into A-dione and/or into epi-ADT.     

19-Norandrost-4-ene-3, 17-dione amplifies the action of aldosterone only in sodium-loaded conditions: Evidence for a new class of amplifiers of aldosterone

The amplification effect of 19-norandrost-4-ene-3, 17-dione (19-nor-A-dione) on aldosterone in normal and sodium-loaded conditions was evaluated using adrenalectomized rats fed a normal or high sodium diet. The administration of 19-nor-A-dione in normal or sodium-loaded conditions did not cause any significant change in urinary $\text{Na}\text{K}$ ratio. The simultaneous administration of subthreshold doses of aldosterone and 19-nor-A-dione in normal conditions also did not cause any significant change in urinary $\text{Na}\text{K}$ ratio. However, the simultaneous administration of subthreshold doses of aldosterone and 19-nor-A-dione in sodium-loaded conditions caused a significant decrease in urinary $\text{Na}\text{K}$ ratio. The decrease in urinary $\text{Na}\text{K}$ ratio was caused by a decrease in urinary Na excretion. These results demonstrate that 19-nor-A-dione, which did not amplify the action of aldosterone in normal conditions, amplified the action of subthreshold doses of aldosterone in sodium-loaded conditions. 19-Nor-A-dione is considered to be an amplifier of aldosterone which works only in sodium-loaded conditions.     

Amplification of nocturnal oscillations in PRA and aldosterone during continuous heat exposure

To assess the effect of continuous heat exposure on the nocturnal patterns of renin, aldosterone, adrenocorticotropic hormone (ACTH), and cortisol, six young men were exposed to thermoneutral environment for 5 days, followed by a 5-day acclimation period in a hot dry environment (35 degrees C). Blood was collected at 10-min intervals during the second night at thermoneutrality (N0) and during the first (N1) and the last (N5) nights of heat exposure. Polygraphic recordings of sleep were scored according to established criteria. Continuous heat exposure led to progressive decreases in the 24-h urinary volume and in Na excretion, whereas urinary osmolality increased. After 5 days of uninterrupted heat, significant increases were found in plasma volume (P less than 0.05), osmolality (P less than 0.01), plasma Na (P less than 0.01), and protein levels (P less than 0.05). Sweat gland output increased during the first 3 days and then declined without any concomitant increases in body temperature. Compared with N0, there were no differences in plasma renin activity (PRA) and aldosterone (PA) profiles during N1 at 35 degrees C. However, during N5 the mean PRA and PA levels were significantly (P less than 0.05) enhanced, and their nocturnal oscillations were amplified (P less than 0.05). This amplification occurred mainly in the second part of the night when regular rapid-eye-movement and non-rapid-eye-movement sleep cycles were observed, leading to a general upward trend in the nocturnal profiles. The relationship between the nocturnal PRA oscillations and the sleep cycles was not modified. ACTH and cortisol patterns were not affected by continuous heat exposure.(ABSTRACT TRUNCATED AT 250 WORDS)     

Atrial natriuretic peptide in acute mountain sickness

To test the hypothesis that elevated atrial natriuretic peptide (ANP) may be involved in altered fluid homeostasis at high altitude, we examined 25 mountaineers at an altitude of 550 m and 6, 18, and 42 h after arrival at an altitude of 4,559 m, which was climbed in 24 h starting from 3,220 m. In 14 subjects, symptoms of acute mountain sickness (AMS) were absent or mild (group A), whereas 11 subjects had severe AMS (group B). Fluid intake was similar in both groups. In group B, urine flow decreased from 61 +/- 8 (base line) to 36 +/- 3 (SE) ml/h (maximal decrease) (P less than 0.05) and sodium excretion from 7.9 +/- 0.9 to 4.6 +/- 0.7) mmol.l-1.h-1 (P less than 0.05); ANP increased from 31 +/- 4 to 87 +/- 26 pmol/l (P less than 0.001), plasma aldosterone from 191 +/- 27 to 283 +/- 55 pmol/l (P less than 0.01 compared with group A), and antidiuretic hormone (ADH) from 1.0 +/- 0.1 to 2.9 +/- 1.2 pmol/l (P = 0.08 compared with group A). These variables did not change significantly in group A, with the exception of a decrease in plasma aldosterone from 189 +/- 19 to 111 +/- 17 pmol/l (P less than 0.01). There were no measurable effects of elevated ANP on natriuresis, cortisol, or blood pressure. The reduced diuresis in AMS may be explained by increased plasma aldosterone and ADH overriding the expected renal action of ANP. The significance of elevated ANP in AMS remains to be established.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hormonal regulation of potassium shifts during graded exhausting exercise

Serum potassium, aldosterone and insulin, and plasma adrenaline, noradrenaline and cyclic adenosine 3':5'-monophosphate (cAMP) concentrations were measured during graded exhausting exercise and during the following 30 min recovery period in six untrained young men. During exercise there was an increase in concentration of serum potassium (4.74 mmol.l-1, SEM 0.12 at the end of exercise vs 3.80 mmol.l-1, SEM 0.05 basal, P less than 0.001), plasma adrenaline (2.14 nmol.l-1, SEM 0.05 at the end of exercise vs 0.30 nmol.l-1, SEM 0.02 basal, P less than 0.001), plasma noradrenaline (1.10 nmol.l-1, SEM 0.64 at the end of exercise vs 1.50 nmol.l-1, SEM 0.05 basal, P less than 0.001), serum aldosterone (0.92 nmol.l-1, SEM 0.14 at the end of exercise vs 0.36 nmol.l-1, SEM 0.05 basal, P less than 0.01), and plasma cAMP (35.4 nmol.l-1, SEM 2.3 at the end of exercise vs 21.4 nmol.l-1, SEM 4.5 basal, P less than 0.05). While concentrations of serum potassium, plasma adrenaline and cAMP returned to their basal levels immediately after exercise, those of plasma noradrenaline and serum aldosterone remained elevated 30 min later (1.90 nmol.l-1, SEM 0.01, P less than 0.01; and 0.85 nmol.l-1, SEM 0.12, P less than 0.01, respectively). Serum insulin concentration did not change during exercise (6.47 mlU.l-1, SEM 0.58 at the end of exercise vs 5.47 mlU.l-1, SEM 0.41 basal, NS) but increased significantly (P less than 0.02) at the end of the recovery period (7.12 mlU.l-1, SEM 0.65).(ABSTRACT TRUNCATED AT 250 WORDS)     

Synthesis of 4,19-disubstituted derivatives of DOC. Radioreceptor assay of some corticosteroid derivatives in human mononuclear leukocytes

Several new 4,19-substituted steroids and previously synthesized corticosteroids were assayed for affinity to type 1 receptors in human mononuclear leukocytes. 11 beta,19-epoxy-4,21-dihydroxypregn-4-ene-3,20-dione (2) was hydrogenated with Pd-C to yield a mixture of all four dihydro derivatives 5, accompanied by 4,21-diacetoxy-11 beta,19-epoxy-3-hydroxypregnan-20-one (6) and 21-acetoxy-11 beta,19-epoxy-4-hydroxypregnane-3,20-dione (7). With hot acetic + p-toluenesulfonic acid 5 underwent rearrangement to 21-acetoxy-11 beta,19-epoxypregn-5-ene-4,20-dione (8) Pd-C hydrogenation of 3,21-diacetoxy-5 beta,19-cyclopregna-2,9(11)-diene-4,20-dione (10) gave 3,21-diacetoxy-5 beta,19-cyclopregn-5-ene-4,20-dione (11) and the 9,11-dihydro derivative of the latter. Treatment of 10 with warm HCl furnished 19-chloro-4,21-dihydroxypregna-4,9(11)-diene-3,20-dione (13). Pd-C hydrogenation of its diacetate 14 afforded the 4,5-dihydro derivative 18, 19-chloro-21-acetoxypregn-9(11)-en-20-one (15), its 4-acetoxy derivative 16 and the 3,4-diacetoxy derivative 17. When tested in a radioreceptor assay in human mononuclear leukocytes the synthesized compounds showed only low relative binding affinities (RBA) to type 1 receptor, the highest being 0.72% for 13 (aldosterone = 100%). For comparison, other RBA in this system were: 19-noraldosterone, 20%; 18-deoxyaldosterone, 5.8%; 18-deoxy-19-noraldosterone, 4.7%; 18,21-anhydroaldosterone, 0.37%; 17-isoaldosterone, 7.6% and apoaldosterone, 4.3%     

Investigation of the salivary 18-hydroxycorticosterone:aldosterone ratio in man using a direct assay

A method for the direct determination of 18-hydroxycorticosterone (18OHB) in human saliva has been developed and validated. Saliva was collected at 30 min and 1 h intervals between 0600 and 2200 h from healthy men and women for the determination of 18OHB (SHB), aldosterone (SA) and glucocorticoids (SGC = cortisol + cortisone). SHB was highly correlated with SA (r = 0.75; P less than 0.001) but even more highly with SGC (r = 0.89; P greater than 0.001). Multiple regression analysis confirmed that SGC was a more important determinant of SHB than was SA. Though the concentrations of 18OHB and aldosterone were highly correlated there was considerable variation in the 18OHB:aldosterone ratio during the period of saliva collection. This ratio tended to be highest in the morning and lowest in the evening and was weakly correlated with SGC level (r = 0.62; P less than 0.01). The 18OHB:aldosterone ratio in saliva approximates to, and is highly correlated with, that in plasma. We suggest that the fluctuations in SHB:SA ratio correspond to the relative rates of secretion of 18OHB and aldosterone and that this ratio is modulated either by ACTH or by cortisol. Whether this indicates that 18OHB is a by-product of glucocorticoid as well as aldosterone metabolism, or whether this implies a separate physiological role for the steroid remains to be clarified.     

Thermoregulatory responses of young and older men to cold exposure.

Nine young (20-25 years) and ten older (60-71 years) men, matched for body fatness and surface area:mass ratio, underwent cold tests in summer and winter. The cold tests consisted of a 60-min exposure, wearing only swimming trunks, to an air temperature of 17 degrees C (both seasons) and 12 degrees C (winter only). Rectal (Tre) and mean skin (Tsk) temperatures, metabolic heat production (M), systolic (BPs) and diastolic (BPd) blood pressures and heart rate (fc) were measured. During the equilibrium period (28 degrees C air temperature) there were no age-related differences in Tre, Tsk, BPs, BPd, or fc regardless of season, although M of the older men was significantly lower (P < 0.003). The decrease in Tre and Tsk (due to the marked decrease in six of the older men) and the increase in BPs and BPd were significantly greater (P < 0.004) for the older men during all the cold exposures. The rate of increase in M was significantly greater (P < 0.01) for the older group when exposed to 12 degrees C in winter and 17 degrees C in summer (due to the marked increase in four of the older men). This trend was not apparent during the 17 degrees C exposure in winter. There was no age-related difference in fc during the exposures. Significant decreases in Tre and Tsk and increases in M, BPs and BPd during the 12 degrees C exposure were observed for the older group (P < 0.003) compared to their responses during the 17 degrees C exposure in winter. In contrast, Tre, M, BPs in the young group were not affected as much by the colder environment. It was concluded that older men have more variable responses and some appear more or less responsive to mild and moderate cold air than young men.     

Reduced urinary aldosterone excretion rates with normal plasma concentrations of aldosterone in the very elderly

Although aldosterone production declines with age, so does the aldosterone metabolic clearance rate (MCR), and the net effect of age on the circulating level of aldosterone may be less than can be predicted from production rates alone. The effect of age on aldosterone production and plasma levels was studied in a group of elderly individuals at a very advanced age when susceptibility to the impacts of age might be particularly pronounced. Seventeen nursing home patients, ages 75-99 (mean age 86 years), had aldosterone production assessed from the urinary excretion rate of the acid hydrolyzable 18-glucuronide conjugate of aldosterone. Aldosterone excretion was low in the elderly when compared to a group of healthy, young to middle-aged subjects: 123 +/- 19 (SEM) vs. 234 +/- 18 ng/h (P less than 0.001). However, plasma aldosterone concentrations in the elderly were well within a range observed in much younger and fully ambulatory subjects: 14.1 +/- 1.3 in the elderly vs. 15.9 +/- 1.8 ng/dL in the young. The plasma aldosterone concentration was apparently maintained at a normal level by a coincident decrease in both the metabolic clearance rate and the aldosterone production rate. In conclusion, an aldosterone deficiency state resulting from an age-correlated reduction in aldosterone production is probably uncommon in the elderly.     

Differences in cortisol, aldosterone and testosterone responses to ACTH 1-17 administered at two different times of day

The effects of 100 micrograms, i.m. of the analog ACTH 1-17 administered at 0800 and 1800 on the secretion of cortisol, aldosterone and testosterone have been studied in normal subjects: 8 male and 8 female. The group as a whole and the males had significantly greater absolute and percent increments in plasma cortisol after administration at 1800. In the females, there was only a greater percent increment in cortisol after the evening administration. The heptadecapeptide always significantly stimulated serum aldosterone, with no difference between the two times of administration. In the females, ACTH 1-17 significantly stimulated testosterone, with a more protracted secretion after the evening administration. In the males, there was always a significant testosterone decrease after the administration of the drug, with no difference between morning and evening. In conclusion, 100 micrograms i.m. of the analog ACTH 1-17 stimulates cortisol secretion more when given during the circadian nadir of plasma cortisol, but only in men. ACTH 1-17 increases testosterone in women and decreases it in men, whereas it seems to increase aldosterone secretion in both sexes.     

Plasma cortisol and corticosteroid-binding globulin in essential hypertension

Plasma concentration of cortisol, total CBG-binding capacity, and blood pressure were measured in control subjects (n = 171), patients with essential hypertension (EH; n = 210) and their first-degree normotensive (NR; n = 84) or hypertensive (HR; n = 66) relatives. Mean (+/- SD) plasma cortisol was significantly (p less than 0.001) decreased in EH (10.1 +/- 4.3 g/dl) patients and HR (11.7 +/- 4.1). Plasma cortisol in NR did not differ from control values (14.3 +/- 4.5) but the distribution of individual values covered the entire control-EH (14.6 +/- 5.5) range. Mean (+/- SD) CBG-binding capacity was significantly (p less than 0.001) lower in EH (14.4 +/- 3.0), NR (17.5 +/- 2), HR (17.6 +/- 2.2) as compared to controls (20.9 +/- 2.1), indicating that the decline in EH and in most relatives was mainly in plasma CBG-bound cortisol. The plasma CBG-binding capacity for cortisol was significantly negatively correlated with mean arterial pressure (MAP) in both controls (p less than 0.001) and NR (p less than 0.01) but not in either HR (r = 0.02) or never-treated EH patients. Total afternoon plasma aldosterone was higher (p less than 0.01 vs. controls) in 93 untreated EH patients (11.2 +/- 4.8 ng/dl) than in either 161 first-degree relatives (8.1 +/- 3.4 ng/dl) or 117 controls (7.6 +/- 3.5 ng/dl). The respective aldosterone-binding globulin (ABG) binding capacities for aldosterone were 21.2 +/- 6.7, 20.1 +/- 9.3 and 9.8 +/- 4.0%. In all these subjects taken together, there was a positive correlation between MAP and ABG-binding capacity (r = 51; p less than 0.001). The association of reduced plasma cortisol and decreased CBG binding capacity in EH may be closely related to altered steroid metabolism, which may be partly explained by an abnormality resembling a relative deficiency in adrenal 17 alpha- and 11 beta-hydroxylation. In some EH patients, hypertension may be the result of the ineffectiveness of plasma cortisol in preventing slightly elevated endogenous ACTH levels leading to an increase in ACTH-sensitive steroids.     

Comparison of saliva and plasma 17-hydroxyprogesterone time-course response to hCG administration in normal men

17-Hydroxyprogesterone (17-OHP) time-course response to hCG (5000 IU) was studied simultaneously in the saliva and the plasma of 12 adult healthy men. Baseline levels in plasma and saliva were: 1.0 +/- 0.1 ng/ml (mean +/- SEM) and 24 +/- 2 pg/ml respectively. After hCG, a biphasic pattern was observed in both fluids with a similar early response but the peak elicited at 33 h in plasma was not observed in saliva where the levels were lower than those recorded at 24 h. Since saliva steroids are believed to reflect the plasma non-protein bound fraction, this difference was assumed to be due to the decrease of the unbound fraction of plasma 17-OHP in the late afternoon as a consequence of the increase of CBG-bound fraction since at that time cortisol levels are low. The ratio of saliva to plasma 17-OHP levels was significantly correlated with plasma cortisol levels: r = 0.44 (P less than 0.01; n = 140). However the similar response in saliva at 24 and at 48 h after hCG allows the evaluation of the endocrine testicular function using saliva instead of plasma.     

Hormonal and electrolyte response to exposure to 17,500 ft.

Hormone, electrolyte, and body fluid compartment changes were studied in subjects who either spent time at 10,000 ft before flying to 17,500 ft or were premedicated with acetazolamide and flown directly to 17,500 ft. In the former group, at 10,000 ft, renin and aldosterone were not different from control. Cortisol increased significantly from 9.8 to 19.5 mug/100 ml on the third day. At 17,500 ft, renin, aldosterone and cortisol were significantly elevated on day 3 but had returned to control levels by day 5. Sodium and potassium excretion was significantly reduced at both altitudes. Total body water, extracellular and plasma volume were reduced (P less than 0.05) at 17,500 ft. Subjects pretreated with acetazolamide and flown directly to 17,500 ft had significant increases (P less than 0.001) in plasma renin, aldosterone, and cortisol levels during the first 4 days at altitude. On day 1 there was a decrease of 45% in sodium and 38% in potassium excretion. On day 4 there was a decrease of 63% and 51%, respectively. These changes are not associated with the premedication. The initial changes may reflect the immediate response to stress and alkalosis followed by a return to control levels as the body adapts to altitude.     

The chemistry of 9 alpha-hydroxysteroids. 1. Preparation of 9 alpha,17 beta-dihydroxy-17 alpha-ethynylandrost-4-en-3-one

9 alpha-Hydroxyandrost-4-ene-3,17-dione 1, when allowed to react with dipotassium acetylide in tetrahydrofuran, resulted, after chromatographic separation, in 4-methyl-19-norandrosta-4,9-diene-1,17-dione 2, 4 xi-methyl-19-norandrosta-5(10),9(11)-diene-1,17-dione 3, 4-methyl-17 alpha-ethynyl-17 beta-hydroxy-19-norandrosta-4,9-dien-1-one 4, 4 xi-methyl-17 alpha-ethynyl-17 beta-hydroxy-19-norandrosta-5(10),9(11)-dien- 1-one 5, and 17 alpha-ethynyl-17 beta-hydroxy-9,10-secoandrost-4-ene-3,9-dione 6. Selective protection of delta 4-3-ketone of 9 alpha-hydroxyandrost-4-ene-3,17-dione 1 as its dienol methyl ether 7, and subsequent reaction with lithium acetylide-ethylenediamine followed by acidic hydrolysis, afforded 9 alpha,17 beta-dihydroxy-17 alpha- ethynylandrost-4-en-3-one 8.     

Dissociation of 19-hydroxy- 19-oxo-, and aromatizing-activities in human placental microsomes through the use of suicide substrates to aromatase

Suicide substrates of aromatase were used as chemical probes to determine if free 19-hydroxyandrost-4-ene-3,17-dione (19-OHA) and 19-oxoandrost-4-ene-3,17-dione (19-oxoA) are obligatory intermediates in the aromatization of androst-4-ene-3,17-dione (androstenedione) to oestrone by human placental aromatase. A radiometric-HPLC assay was used to monitor 19-hydroxy, 19-oxo-, and aromatized products formed in incubations of [14C]androstenedione and human placental microsomes. When microsomes were preincubated with the suicide substrates 10 beta-mercapto-estr-4-ene-3,17-dione (10 beta-SHnorA), or 17 beta-hydroxy-10 beta-mercaptoestr-4-ene-3-one (10 beta-SHnorT), it was found that 19-hydroxy-, 19-oxo- and aromatase activities were inhibited in parallel. However, when the suicide substrates 4-hydroxyandrost-4-ene-3,17-dione (4-OHA) and 19-mercaptoandrost-4-ene-3,17-dione (19-SHA) were preincubated with placental microsomes, significantly greater inhibition of formation of oestrogens was observed in comparison to the inhibition of formation of 19-hydroxy- and 19-oxo-metabolites. Furthermore, significantly more time-dependent inhibition of 19-oxoA formation was observed in comparison to inhibition of 19-OHA formation with these same inhibitors. These results suggest that 19-hydroxy- and 19-oxo-androstenediones are not free, obligatory intermediates in the aromatization of androstenedione by human placental aromatase, but rather are products of their own autonomous cytochrome P-450-dependent, microsomal enzymatic activities.     

Effect of bromocriptine on the control of plasma aldosterone diurnal variation in normal supine man.

In order to investigate the role of prolactin in the control of the circadian rhythm of plasma aldosterone (PA), plasma renin activity (PRA), cortisol (PC), aldosterone and prolactin (PRL) levels were determined in samples at 4-hour intervals from 5 normal supine men over a period of 24 h under basal conditions and subsequently over a period of 24 h during suppression of prolactin release by bromocriptine (CB-154). After suppression of prolactin, statistically signific1nt circadian rhythms in PC and PA have been detected with a moderate decrease of PA concentration, while the PC level remained unalterated. PRA rhythmicity persisted with a significant shift of acrophase and remarkable reduction of plasma levels. Moreover, during CB administration a significant correlation was obtained between PA and PC, while no correlation was detected between PA and PRA. These data are consistent with the following concepts: (a) the prolactin does not play a significant role in the regulation of circadian rhythm and concentration of plasma aldosterone in normal supine men, and (b) bromocriptine induces a remarkable reduction of PRA and a variable decrease in plasma aldosterone, but it does not influence the secretion of cortisol in normal subjects.     

Effects of ACTH on plasma levels of adrenal steroids in essential hypertension.

Plasma concentrations of progesterone (P), deoxycorticosterone (DOC), 17-hydroxyprogesterone (17-OH P), corticosterone (B), deoxycortisol (S), cortisol (F) and aldosterone (A) in 8 control subjects (mean age: 40.5 years) and 10 patients with essential hypertension (EH) (mean age: 48.5 years) were determined before, 4 and 8 hours after an infusion of ACTH at a rate of 25 units per 8 hours. Secretion rates (SR) of 18-hydroxy-11-deoxycorticosterone (18-OH DOC) were measured 24 hours before and again on the day of ACTH infusion. All subjects were studied on the fourth day of a diet containing 135 mEq of sodium and 90 mEq of potassium. There was no statistically significant difference between 8 control subjects and 10 patients with EH in the 7 plasma steroid levels and the SR of 18-OH DOC before ACTH infusion. The mean plasma P response to ACTH was slightly lower in controls than in patients with EH, while that of 17-OH P (in male subjects) was slightly higher. The mean plasma B response was significantly lower after 4 hours of ACTH infusion (p less than 0.01), while that of DOC was significantly higher after 8 hours of ACTH infusion (p less than 0.05) in patients with EH. The mean plasma S rose significantly more in patients with EH (p less than 0.025) at 4 and 8 hours after ACTH infusion. The mean plasma F response to ACTH infusion was slightly lower in patients with EH than in controls. The mean response of 18-OH DOC SR to ACTH infusion was slightly higher in patients with EH than in controls. The mean plasma A response was significantly higher in patients with EH than in controls 4 (p less than 0.05) and 8 hour (p less than 0.001) after an ACTH infusion. These results could be explained in part by abnormalities in the 17- and 11-hydroxylase systems, and that the abnormality in 11-hydroxylation was more pronounced than that in the 17-position. Furthermore, we suspect that the sensitivity of adrenal aldosterone to ACTH might be increased or another accelerated pathway to aldosterone biosynthesis might exist in patients with EH.