The ovulatory menstrual cycle is the result of the integrated action of the hypothalamus, pituitary, ovary, and endometrium. Like a metronome, the hypothalamus sets the beat for the menstrual cycle by the pulsatile release of gonadotropin-releasing hormone (GnRH). GnRH pulses occur every 1-1.5 h in the follicular phase of the cycle and every 2-<em>4</em> h in the luteal phase of the cycle. Pulsatile GnRH secretion stimulates the pituitary gland to secrete luteinizing hormone (LH) and follicle stimulating hormone (FSH). The pituitary gland translates the tempo set by the hypothalamus into a signal, LH and FSH secretion, that can be understood by the ovarian follicle. The ovarian follicle is composed of three key cells: theca cells, granulosa cells, and the oocyte. In the ovarian follicle, LH stimulates theca cells to produce <em>androstenedione</em>. In granulosa cells from small antral follicles, FSH stimulates the synthesis of aromatase (Cyp19) which catalyzes the conversion of theca-derived <em>androstenedione</em> to estradiol. A critical concentration of estradiol, produced from a large dominant antral follicle, causes positive feedback in the hypothalamus, likely through the kisspeptin system, resulting in an increase in GnRH secretion and an LH surge. The LH surge causes the initiation of the process of ovulation. After ovulation, the follicle is transformed into the corpus luteum, which is stimulated by LH or chorionic gonadotropin (hCG) should pregnancy occur to secrete progesterone. Progesterone prepares the endometrium for implantation of the conceptus. Estradiol stimulates the endometrium to proliferate. Estradiol and progesterone cause the endometrium to become differentiated to a secretory epithelium. During the mid-luteal phase of the cycle, when progesterone production is at its peak, the secretory endometrium is optimally prepared for the implantation of an embryo. A diagrammatic representation of the intricate interactions involved in coordinating the menstrual cycle is provided in Fig. 1.

A systematic review of the literature on sex steroid measurement in breast tissue identified only 19 articles meeting the following criteria: menopausal status given; steroids measured in tissue homogenates by conventional RIA with a purification step or by mass spectrometry; and values reported per g tissue or per g protein. Twelve articles were analyzed in detail for: ratios of sex steroid hormone levels in cancerous or benign tissues to blood levels, stratified by menopausal status; ratios between the different hormone levels within tissues or within blood; and difference in these ratios between tissue and blood compartments. Estrogen and androgen concentrations varied greatly in benign and cancerous tissues and in blood between individuals. Postmenopausal, but not premenopausal, estradiol concentrations were significantly higher in cancerous compared to benign breast tissue. The estradiol/estrone ratio was lowest in premenopausal benign tissue, and substantially higher in premenopausal cancerous tissue and postmenopausal benign and cancerous tissues. Estradiol and estrone levels were considerably higher in tissue than in plasma in both premenopausal and postmenopausal women. Androgen levels were generally higher in the benign than the cancerous tissue, and tissue androgen levels were higher than in plasma, suggesting in situ aromatization of androgens to estrogens in breast cancer tissue. Limited available data on levels of hydroxylated estrogens in breast tissue compared to corresponding levels in plasma or urine were reviewed, but due to the paucity of studies no conclusions can presently be drawn regarding the relationship of the 2-hydroxyestrone:16α-hydroxyestrone ratio to breast cancer risk and genotoxic effects of <em>4</em>-hydroxylated estrogens. Finally, data on hormone levels in breast adipose tissue were analyzed; high levels of <em>androstenedione</em> and testosterone and significant estrone and estradiol levels in breast adipocytes from postmenopausal breast cancer patients are consistent with an obesity-inflammation-aromatase axis occurring locally in breast tissue. The controversies regarding the source of intratumoral estrogens in the breast are summarized.

BACKGROUND

It has been postulated that intrauterine undernutrition may predispose to serious endocrine consequences, including precocious pubarche (PP), functional ovarian hyperandrogenism and insulin resistance syndrome.

OBJECTIVE

The aim of this study was to determine whether a history of PP was associated with the development of hyperandrogenism and/or metabolic consequences and to evaluate the effect of birth weight on this association.

METHODS

The study population comprised 27 Caucasian girls with a history of PP and 25 healthy girls of similar age (17.<em>4</em> +/- 1.3 vs. 17.7 +/- 0.9 years).

RESULTS

Gynecological age, irregular menses and oral contraceptive use were similar in the two groups. PP girls showed an increased Ferriman-Gallway Score [median (range) 8 (<em>4</em>-17) vs. 6 (2-10), P = 0.02] and tended to have more previous history of acne. No statistical differences were found between the groups for mean testosterone (1.7 +/- 0.7 vs. 1.<em>4</em> +/- 0.7 nmol/l, P = 0.<em>4</em>9) and dehydroepiandrosterone sulphate concentrations (7.2 +/- 3.7 vs. 5.8 +/- 2.0 micromol/l, P = 0.15), but mean Delta<em>4</em>-<em>androstenedione</em> concentrations (7.3 +/- 2.<em>4</em> vs. <em>4</em>.9 +/- 2.1 nmol/l, P = 0.007) and free androgen index (8.5 +/- 9.7 vs. 3.6 +/- 3.9 IU, P = 0.003) were significantly increased in the PP group. All girls showed normal glucose tolerance with an oral glucose tolerance test. Derived insulin resistance parameters were not statistically different between the two groups and fasting lipids were comparable in both groups. There was no significant effect of birth weight on androgen levels in the PP girls. Moreover, none of the PP girls demonstrated the above-described association.

CONCLUSIONS

Precocious pubarche could be the first sign of future functional ovarian hyperandrogenism but a link between this condition and intrauterine undernutrition or insulin resistance could not be demonstrated in this study.

Four patients with androgen insensitivity had plasma LH and FSH measured at 20-min intervals for 2<em>4</em> h and at 15- to 30-min intervals for 3 h after the injection of LRH. Twenty-four-hour mean testosterone (T), estradiol, and <em>androstenedione</em> (delta <em>4</em>) levels were also measured. Patients with androgen insensitivity had significantly elevated LH levels (P less than 0.05) and an increase in the number of LH secretory episodes (P less than 0.001) compared to normal subjects. The amplitude of the LH secretory episodes, expressed as the absolute increment, was significantly higher than normal controls (P less than 0.005). The LH response to LRH (absolute increment) was twice that of normal, but was not significantly different from normal subjects. The 2<em>4</em>-h mean FSH levels were normal in three of the patients and elevated in one. This patient had the mildest degree of androgen insensitivity on clinical exam and the greatest degree of testicular atrophy. The 2<em>4</em>-h mean T, estradiol, and delta <em>4</em> levels were higher than normal, but only the delta <em>4</em> was significantly increased (P less than 0.05). To determine if the elevated LH levels were in response to a decrease in the free T level, we measured T-binding capacity (TBG), TBG was higher than normal controls but was not significantly different, suggesting that elevated LH levels were probably in response to a decrease in T action at the hypothalamic-pituitary level. This was further supported by the inability of prolonged dihydrotestosterone administration to affect LH secretion in one of the patients with the Reifenstein syndrome.

Congenital adrenal hyperplasia due to 3 beta-ol-hydroxysteroid dehydrogenase (3 beta-ol) deficiency is usually lethal. A partial deficiency in 3 beta-ol has been suggested in some women presenting with androgen excess. In this study, 2<em>4</em> women were investigated who had hirsutism and oligomenorrhea and high serum delta 5 androgens compared to delta <em>4</em> androgens. Of these women, 9 had significantly elevated 17-hydroxypregnenolone (17 Preg) to 17-hydroxyprogesterone (17 Prog) ratios when compared to controls under basal conditions. On further testing of 9 women with ACTH, <em>4</em> had significantly elevated 17 Preg to 17 Prog ratios. Eight women had elevated ratios of dehydroepiandrosterone sulfate to <em>androstenedione</em>, <em>4</em> had elevations of androstenediol (Adiol) to testosterone (T), and <em>4</em> had abnormal 17 Preg to cortisol ratios. Only 3 women out of the original 2<em>4</em> selected for study had elevated ratios for all <em>4</em> different steroid pairs measured. 17 Prog was normal in these women with 3 beta-ol deficiency in contradistinction to the high levels normally observed in women with congenital adrenal hyperplasia due to 21- or 11-hydroxylase deficiency. It is suggested that the cause of androgen excess in these women is high circulating levels of Adiol and, in part, the slightly elevated unbound T levels in these women. In conclusion, a subtle, incomplete form of 3 beta-ol deficiency may exist in adult women and is manifest by high delta 5 androgens such as dehydroepiandrosterone sulfate and Adiol and normal delta <em>4</em> androgens such as <em>androstenedione</em> and T.

Using dimethylbenz(a)anthracene (DMBA)-induced mammary tumors in the rat as model, comparison was made of the effect of treatment for 20 days with the aromatase inhibitor <em>4</em>-hydroxy<em>androstenedione</em> (<em>4</em>-OH-A) (7.5 mg, twice daily) or the antiandrogen flutamide (5 mg, twice daily) on tumor growth as well as on plasma and tumor content of estrogens, androgens, and their precursors and metabolites. Tumor number and size were markedly decreased following treatment with either drug, the effect of treatment being more important on size than number, and on new tumors which developed during treatment than on tumors already present at start of treatment. Treatment with the aromatase inhibitor <em>4</em>-OH-A caused a parallel decrease in plasma and tumor levels of pregnenolone (Preg), progesterone (P), and 17-OH P, while there was a marked increase in dehydroepiandrosterone (DHEA), androst-5-ene-3 beta,17 beta-diol (delta 5-diol), <em>androstenedione</em> (delta <em>4</em>-dione), testosterone (T), androstane-3 alpha, 17 beta-diol (3 alpha-diol), and androstane-3 beta,17 beta-diol (3 beta-diol), with no significant change in dihydrotestosterone (DHT) and 17 beta-estradiol levels. The marked increase in tissue T content coupled to a decrease in P levels could well contribute to the inhibition of tumor growth induced by <em>4</em>-OH-A. Flutamide, on the other hand, caused a marked fall in plasma and tissue levels of Preg, 17-OH Preg, P, and 17-OH P, with no significant change in the concentration of the other steroids, thus suggesting a possible role of the fall in tissue P levels in the inhibition of tumor growth. Since both drugs are potent inhibitors of DMBA-induced tumor growth in intact animals, better knowledge of their mechanism of action should add to our understanding of the multiple endocrine factors controlling the growth of these tumors.

We report the case of a patient with bilateral testicular tumors and congenital adrenal hyperplasia due to 21-hydroxylase deficiency. Catheterization of the left testicular and adrenal veins was performed. The presence of 11 beta-hydroxylated steroids in the spermatic veins confirmed the presence of testicular tumor secondary to adrenal rest cells. After adrenal suppression by dexamethasone combined with gonadal stimulation with hCG, a dramatic decrease in androgens and adrenal steroids was observed in the peripheral blood. Compared to the periphery, 21-deoxycortisol and 11 beta-hydroxy-delta <em>4</em>-<em>androstenedione</em> levels remained higher than that of 21-deoxycorticosterone in the gonadal vein, but not in the adrenal vein, which seems to indicate that the nature of this ectopic tissue is unusual and that its sensitivity to dexamethasone depends on the adrenocortical zones. No rise in estradiol or testosterone was obtained after hCG stimulation, suggesting that all of the testicular tissue was inactive or destroyed. This finding was confirmed by histological examination.

Adolescent females who have irregular menstrual periods may have the nonclassical form of congenital adrenal hyperplasia due to a mild deficiency of steroid 21-hydroxylase (NC 21-OHD). Hyperandrogenic signs such as acne, frontal hair loss, hirsutism, and irregular menstrual periods should alert the physician to the diagnosis of NC 21-OHD. An ACTH stimulation test in which serum hormone concentrations of 17-OHP, Delta(<em>4</em>)-<em>androstenedione</em>, and testosterone are determined will assist in the diagnosis of NC 21-OHD, but the definitive diagnostic test is an analysis of the mutations in the CYP21A2 gene. Typical mutations in the CYP21A2 gene in patients with NC 21-OHD are an exon 7 or an exon 1 mutation. Once the genotype establishes the diagnosis of NC 21-OHD, treatment should be initiated. Typical treatment is dexamethasone, 0.25 mg HS, which generally reverses the hyperandrogenic signs.

Measurements of the aromatase-inhibiting and antioxidative capacities of flavonoids in vitro showed that slight changes in flavonoid structure may result in marked changes in biological activity. Several flavonoids such as 7-hydroxyflavone and chrysin (5,7-dihydroxyflavone) were shown to inhibit the formation of 3H-17beta-estradiol from 3H-<em>androstenedione</em> (IC(50)<1.0 microM) in human choriocarcinoma JEG-3 cells and in human embryonic kidney cells HEK 293 transfected with human aromatase gene (Arom+HEK 293). Flavone and quercetin (3,3',<em>4</em>',5,7-pentahydroxyflavone) showed no inhibition (IC(50)>100 microM). None of the requirements for optimal antioxidative capacity (2,3-double bond with <em>4</em>'-hydroxy group, 3-hydroxyl group, 5,7-dihydroxy structure and the orthodihydroxy structure in the B-ring) is relevant for the maximum inhibition of aromatase by flavonoids. After oral administration to immature rats at a dose of 50 mg/kg body weight, which considerably exceeds amounts found in daily human diets, neither aromatase-inhibiting nonestrogenic flavonoids, such as chrysin, nor estrogenic flavonoids, such as naringenin and apigenin, induced uterine growth or reduced estrogen- or androgen-induced uterine growth. The inability of flavonoids to inhibit aromatase and, consequently, uterine growth in short-term tests may be due to their relatively poor absorption and/or bioavailability.

Abnormal steroid secretion may contribute to anovulation in insulin dependent diabetic patients with amenorrhoea. We have measured serum sex hormone-binding globulin (SHBG) and free and bound oestrogen and androgen levels in 17 such patients. As controls we included 17 patients with insulin dependent diabetes mellitus and normal menstrual cycles, 21 regularly menstruating normal women (both sampled during early follicular phase), and 23 non-diabetic patients with amenorrhoea. The diabetic patients with normal cycles had significantly higher serum concentrations of delta <em>4</em>-<em>androstenedione</em> and testosterone than the normal women (P less than 0.01). The amenorrhoeic diabetics in contrast had significantly lower serum concentrations of SHBG, 5 alpha-dihydrotestosterone and free and total oestradiol-17 beta than either group of menstruating women (P less than 0.05), and significantly lower concentrations of delta <em>4</em>-<em>androstenedione</em> (P less than 0.01), dehydroepiandrosterone sulphate (P less than 0.01), testosterone (P less than 0.01), and oestrone (P less than 0.05), than the cycling diabetics. The two amenorrhoeic groups had similar free and bound sex hormone concentrations except that delta <em>4</em>-<em>androstenedione</em> levels were significantly lower in the diabetics (P less than 0.01). We conclude that the low sex hormone levels in diabetic women with amenorrhea may be due to suppression of the hypothalamic-pituitary axis in view of the impaired LH secretion found in these patients and that excess androgen secretion seems not to be of aetiological importance in amenorrhea related to diabetes mellitus. The decreased steroid levels in amenorrheic diabetics is due to their suppressed ovarian function while the increased androgen levels in diabetics with regular cycles are probably of ovarian origin.

The responses of plasma testosterone (T), sex hormone-binding globulin (SHBG) binding capacity, <em>androstenedione</em> (A), and luteinizing hormone (LH) to 21 km of marching exercise and to sleep deprivation stress were studied in army recruits. The effect of physical fitness on the exercise responses was evaluated, cross-sectionally, by comparing the stress responses in 11 fit and 11 less fit subjects, and, longitudinally, in 11 subjects after <em>4</em> months of physical training. The submaximal marching exercise did not significantly alter the plasma hormone levels compared to the control day levels. The fit subjects had a tendency toward smaller decreases in the mean plasma T and T/SHBG ratio during both control and exercise days than the less fit subjects. After <em>4</em> months of conditioning, the mean plasma T and T/SHBG ratio tended to decrease less during both control and exercise days, which was more evident in the well-conditioned subjects. The exercise caused a decrease in mean plasma LH, especially in the less fit subjects. After sleep deprivation stress, the morning levels of plasma T, A, and LH were significantly depressed, as a result of which the normal diurnal variation of the hormones disappeared.

Inhibition of estrogen production provides effective therapy for patients with hormone-dependent breast cancer. The source of estrogens in premenopausal women is predominantly the ovary, but after the menopause, estradiol is synthesized in peripheral tissues through the aromatization of androgens to estrogens. Uptake from plasma is the primary mechanism for maintenance of estradiol concentrations in breast cancer tissue in premenopausal women, whereas several steps may be operant in postmenopausal women. These include enzymatic synthesis of estradiol via sulfatase, aromatase, and 17 beta-hydroxysteroid dehydrogenase in the tumor itself. Aromatization of androgens secreted by the adrenal to estrogens in peripheral tissues and transport to the tumor via circulation in the plasma provides another means of maintaining breast tumor estradiol levels in postmenopausal women. These various sources contribute to the high tissue estrogen levels measured in breast tumor tissue. To effectively suppress tissue concentrations of estrogens and circulating estradiol in postmenopausal patients, various aromatase inhibitors have been developed recently. These include steroidal inhibitors such as <em>4</em>-hydroxy-<em>androstenedione</em> as well as non-steroidal compounds with imidazole and triazole structures. The most potent of these, CGS 20267, is reported to suppress levels of active estrogens (i.e., estrone, estrone sulfatase, and estradiol) by more than 95%. This compound can suppress both serum and 2<em>4</em>-hr urine estrogens to a greater extent than produced by the second generation inhibitor, CGS 169<em>4</em>9A. CGS 20267 is highly specific since it does not affect cortisol and aldosterone serum levels during ACTH stimulation tests nor sodium and potassium balance in 2<em>4</em>-hr urine samples.(ABSTRACT TRUNCATED AT 250 WORDS)

Serum sex hormones and endurance performance after a lacto-ovo vegetarian and a mixed diet. Med. Sci. Sports Exerc., Vol. 2<em>4</em>, No. 11, pp. 1290-1297, 1992. The effect of a lacto-ovo vegetarian (V) and a mixed, meat-rich (M) diet on the level of serum sex hormones, gonadotropins, and endurance performance of eight male endurance athletes was investigated in a 2 x 6 wk cross-over study. The energy contribution from carbohydrate, fat, and protein was 58%, 27%, and 15% on the V diet and 58%, 28%, and 1<em>4</em> E% on the M diet. For total fasting serum testosterone (T) there was a significant interaction between diet and time (P < 0.01). Thus, the V diet resulted in a lower total T level (13.7, 9.8-32.<em>4</em> nmol.l-1) (median and range) compared with the M diet (17.<em>4</em>, 11.8-33.5 nmol.l-1). During exercise after 6 wk on the diets total T was also significantly lower on the V than on the M diet (P < 0.05). Serum free testosterone, however, did not differ significantly during the 6 wk dietary intervention periods and neither did serum concentrations of sex hormone binding globulin, dihydrotestosterone, dehydroepiandrosterone sulphate, <em>4</em>-<em>androstenedione</em>, estrone, estradiol, estrone sulphate, or gonadotropins. Endurance performance time was higher for six and lower for two after the mixed diet compared with the vegetarian diet. This was not significant, however. In conclusion, 6 wk on a lacto-ovo vegetarian diet caused a minor decrease in total testosterone and no significant changes in physical performance in male endurance athletes compared with 6 wk on a mixed, meatrich diet.

The objective of this study was to evaluate the influence of thyroxine (T<em>4</em>) and triiodothryonine (T3) on steroid production by bovine granulosa and thecal cells. Granulosa and thecal cells were obtained from small (1 to 5 mm) and large >> or = 8 mm) follicles of cattle, respectively, and cultured for <em>4</em> d. We conducted six experiments to evaluate the effect of 2 d of exposure to various doses of T3 or T<em>4</em>. In insulin- or insulin plus FSH-treated granulosa cells of experiment 1, 30 and 100 ng/ml of T<em>4</em> had no effect on aromatase activity or progesterone production. In experiment 2, in the presence of insulin and FSH, 1 and 3 ng/ml of T3 weakly (<1.<em>4</em>-fold) increased aromatase activity of granulosa cells but had no effect on progesterone production. Low doses of T<em>4</em> (3 to 30 ng/ml) tested in experiment 3 had no effect on aromatase activity but increased (to as much as 1.<em>4</em>-fold) progesterone production by granulosa cells. In experiment <em>4</em>, T<em>4</em> (30 ng/ml) increased (to 1.2-fold) progesterone production by granulosa cells only in the presence of FSH and had no effect on aromatase activity. In thecal cells of experiment 5, in the presence of insulin and LH, 30 and 100 mg/ml of T<em>4</em> increased <em>androstenedione</em> production to 2.3- and 2.8-fold, respectively; only 100 ng/ml of T<em>4</em> was effective at stimulating progesterone production by thecal cells. In experiment 6, 1 ng/ml of T3 increased thecal cell <em>androstenedione</em> production to 3.9-fold, whereas 3 ng/ml of T3 was without effect; progesterone production was not affected by T3. These results support the hypothesis that thyroid hormones may have direct stimulatory effects on ovarian function in cattle, acting at the level of granulosa and thecal cells.

BACKGROUND

Studies using purified enzyme preparations, placental microsomes or cell lines have shown that certain phytoestrogens can inhibit the enzymes that convert androgens to estrogens, namely aromatase and 17beta-hydroxysteroid dehydrogenase (HSD) type 1 and type 5. The study aim was to investigate the effects of selected phytoestrogens on aromatase and 17beta-HSD type 1 activity in primary cultures of human granulosa-luteal (GL) cells.

RESULTS

GL cells, cultured for <em>4</em>8 h in medium containing 5% fetal calf serum and for a further 2<em>4</em> h in serum-free medium with or without hFSH or hCG, were exposed to steroid substrates during the last 1-<em>4</em> h of the experiment. The production of progesterone in the presence of pregnenolone or estradiol synthesis from <em>androstenedione</em>, estrone or testosterone showed dose- and time-dependent increases. Whilst hCG priming had no effect on progesterone production, FSH priming induced mean 68 and 56% increases in the production of estradiol from <em>androstenedione</em> (A-dione) and estrone respectively, but had no significant effect on the metabolism of testosterone to estradiol. None of the phytoestrogens investigated had any acute effects on enzyme activity. In contrast, when GL cells were exposed to the compounds for 2<em>4</em> h prior to exposure to steroid substrates for <em>4</em> h, 10 micro mol/l apigenin and zearalenone significantly inhibited aromatase activity, whilst biochanin A and quercetin had no effect. None of the phytoestrogens inhibited FSH-induced 17beta-HSD type 1 activity, and only quercetin significantly inhibited progesterone production.

CONCLUSIONS

The inability of phytoestrogens to acutely inhibit steroidogenic enzymes in human GL cells (as has been shown in cell-free models) suggests that they are either rapidly metabolized to relatively inactive compounds or that the high enzyme activity in human GL cells masks any inhibitory effects of the compounds at the concentration tested.

Econazole, imazalil, and prochloraz, which have broad spectrum antimycotic activity, are shown to be potent inhibitors of steroid aromatase activity of human placental microsomes. The IC50 values for the inhibition of aromatase activity by econazole, imazalil, miconazole, prochloraz, clotrimazole, ketoconazole, and aminoglutethimide are 0.03, 0.15, 0.6, 0.7, 1.8, 60, and <em>4</em>5 microM, respectively. Econazole and <em>4</em>-hydroxy<em>androstenedione</em> also inhibit the steroid aromatase activity of human fetal liver, a finding which suggests that extraplacental aromatase may have many similarities to the placental enzyme. Econazole is a more effective inhibitor of placental aromatization of 19-hydroxy<em>androstenedione</em> than of <em>androstenedione</em>. This observation is consistent with the competitive nature of the inhibition of aromatase by imidazole antimycotic agents and the reduced affinity of the placental aromatase enzyme for 19-hydroxy<em>androstenedione</em> compared to <em>androstenedione</em>. The effectiveness of these imidazole antimycotic agents to inhibit the multiple hydroxylations of progesterone which are catalyzed by human fetal adrenal microsomes is also defined. While all of the imidazole antimycotic agents are potent inhibitors of the 16 alpha-, 17 alpha-, and 21-hydroxylations of progesterone, selective inhibitory profiles are apparent. Ketoconazole is a most potent inhibitor of human fetal adrenal progesterone 16 alpha- and 17 alpha-hydroxylases while clotrimazole and imazalil are the most potent inhibitors of progesterone 21-hydroxylase. These results are strongly supportive that imidazole drugs are selective inhibitors not only of steroid aromatase but also of other microsomal steroid hydroxylases.

An important factor in disruption of hepatic heme biosynthesis by porphyrinogenic drugs appears to be interaction with one or several cytochrome P<em>4</em>50 isozymes. Clarification of the nature of the interaction between porphyrinogenic drugs and cytochrome P<em>4</em>50 isozymes, as well as identification of the isozymes involved, will be helpful in extrapolating the results of animal experimentation to humans. Administration of griseofulvin to mice results in accumulation in the liver of two N-alkylated protoporphyrins (PPs). The major N-alkyl PP, N-griseofulvin PP, which is devoid of ferrochelatase-inhibitory activity, was shown to be the precursor of N-methyl PP, which is a potent ferrochelatase inhibitor. N-Griseofulvin PP was present predominantly as the Nc regioisomer rather than the anticipated NA regioisomer. Progesterone (PG) 6 beta-hydroxylase and <em>androstenedione</em> (AD) 6 beta-hydroxylase, diagnostic markers for cytochrome P<em>4</em>50 3A1/2 activity in rat liver, were identified in chick embryo liver. The in ovo administration of 3,5-diethoxycarbonyl-1,<em>4</em>-dihydro-2,6-dimethyl-<em>4</em>-ethylpyridine (<em>4</em>-ethyl DDC) and 3-[2-(2,<em>4</em>,6-trimethylphenyl)thioethyl]-<em>4</em>-methylsydnone (TTMS) caused inactivation of chick embryo hepatic PG and AD 6 beta-hydroxylases. Ascorbate was shown to inhibit uroporphyrin accumulation in cultures of chick embryos treated with porphyrinogenic drugs and in the livers of an ascorbate-requiring rat strain treated with a porphyrinogenic combination of chemicals. Ascorbate appears to act by inhibiting oxidation of uroporphyrinogen to uroporphyrin. Avian hepatocytes were cultured in <em>4</em>8-well plates and directly assayed within the wells for the activity of 7-ethoxyresorufin O-deethylation (EROD), and for porphyrin and protein concentration by a fluorescence plate reader. Uroporphyrin was the main porphyrin to accumulate in response to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in chick embryos, pheasants, ducks, and herring gulls; heptacarboxylated porphyrin predominated in turkey hepatocytes. The EROD-inducing potential of complex mixtures of polyhalogenated aromatic hydrocarbons from herring gull eggs was assessed and compared with that of TCDD. A nuclear heme pool appears to regulate the transcription of the cytochrome P<em>4</em>50 2B1/2B2 genes in rat liver, by modulating binding of a transcription factor(s) that appears to be involved in the transcriptional activation by phenobarbitone.

BACKGROUND

Physical activity is known to relieve the metabolic complications of polycystic ovary syndrome (PCOS), and exercise is also associated with telomere biology. We investigated the changes induced by progressive resistance training (PRT) in telomere content and metabolic disorder in women with PCOS and controls.

METHODS

Forty-five women with PCOS and 52 healthy women aged 18 to 37 years were submitted to PRT. A linear periodization of PRT was prepared based on a trend of decreasing volume and intensity throughout the training period. The volunteers performed PRT 3 times a week for <em>4</em> months. The participants' physical characteristics and hormonal concentrations were measured before and after PRT, as telomere content that was measured using quantitative real-time polymerase chain reaction.

RESULTS

Briefly, Progressive resistance training reduced waist circumference, body fat percentage, plasma testosterone and sex hormone-binding globulin concentrations, glycemia, and free androgen index. Fasting insulin and insulin resistance index were greater in women with PCOS. Androstenedione and homocysteine increased after PRT. There were no differences in telomere content between controls (0.96 ± 0.3 before vs 0.85 ± 0.21 after) and women with PCOS (0.9<em>4</em> ± 0.33 before vs 0.88 ± 0.39 after). Adjusted analysis showed telomere shortening after PRT in all women (0.95 ± 0.31 before vs 0.86 ± 0.31 after; P = .03). In women with PCOS, increased homocysteine levels were related to telomere reduction and increased androstenedione was positively correlated with telomere content after PRT.

CONCLUSIONS

Progressive resistance training had positive effects on the hormonal and physical characteristics of women with PCOS and controls, but telomere content was reduced and homocysteine level increased in all participants.

Aromatase is present in human breast tumors and in breast cancer cell lines suggesting the possibility of in-situ estrogen production via the <em>androstenedione</em> to estrone and estradiol pathway. However, proof of the biologic relevance of aromatase in breast cancer tissue requires the demonstration that this enzyme mediates biologic effects on cell proliferation. Accordingly, we studied the effects of the aromatase substrate, <em>androstenedione</em>, on the rate of proliferation of wild-type and aromatase-transfected MCF-7 breast cancer cells. <em>Androstenedione</em> did not increase cell growth in wild-type MCF-7 cells which contained relatively low aromatase activity and produced <em>4</em>-fold more estrone than estradiol. In contrast, aromatase-transfected cells contained higher amounts of aromatase, produced predominantly estradiol, and responded to <em>androstenedione</em> with enhanced growth. An aromatase inhibitor fadrozole hydrochloride, blocked the proliferative effects of <em>androstenedione</em> providing evidence for the role of aromatase in this process. As further evidence of the requirement for aromatase, cells transfected with the neomycin resistance expression plasmid but lacking the aromatase cDNA did not respond to <em>androstenedione</em>. These studies provide evidence that aromatase may have a biologic role for in-situ synthesis of estrogens in breast cancer tissue.

In nonclassical steroid 21-hydroxylase deficiency, the genotype may be represented as a homozygous mild (nonclassical) form of the 21-hydroxylase defect or as a compound heterozygote, with one severe (classical) and one mild (nonclassical) 21-hydroxylase deficiency allele. We examined hormone levels in patients with nonclassical 21-hydroxylase deficiency in whom pedigree analysis and/or HLA linkage disequilibrium allowed unequivocal identification of the respective haplotypes as either classical or nonclassical. The results indicated that compound heterozygotes (21-OH defsevere/21-OH defmild) have an ACTH-stimulated 17-hydroxyprogesterone (17-OHP) response significantly greater than that of mild homozygotes (21-OH defmild/21-OH defmild): at 60 min, 8,131 +/- <em>4</em>,205 (+/-SD) (n = 17) vs. <em>4</em>,<em>4</em>68 +/- 2,123 ng/dl (n = 31) for the respective groups (P less than or equal to 0.01); at 360 min, 11,067 +/- 5,582 (n = 17) vs. 57<em>4</em>6 +/- 1565 (n = 8, P less than or equal to 0.01). Since serum cortisol levels were the same in both groups, the ratio of 17-OHP to cortisol was higher in the former group. Sixty minute ACTH-stimulated serum delta <em>4</em>-<em>androstenedione</em> levels also were significantly higher in compound heterozygotes than in mild homozygotes. Serum dehydroepiandrosterone and its sulfate were not significantly different between the two groups. Notably, compound heterozygotes were no more likely to have signs of androgen excess than were homozygotes for the mild gene defect. Stimulated levels of serum 17-OHP, 17-OHP/cortisol, delta <em>4</em>-<em>androstenedione</em> and dehydroepiandrosterone and its sulfate did not differ significantly between heterozygotes for the classical (21-OH defsevere/21-OHnormal) and nonclassical (21-OH defmild/21-OHnormal) 21-hydroxylase deficiency alleles. Thus, the presence of a single normal 21-hydroxylase allele is sufficient to obscure the difference between a severe and a mild 21-hydroxylase deficiency allele on the opposite haplotype. We conclude that the compound heterozygous patients as a group have a significantly higher response of 21-hydroxylase precursors to ACTH stimulation than do patients with the homozygous mild 21-hydroxylase deficiency state.

Cryptic 21-hydroxylase deficiency has been previously described in asymptomatic family members of patients with classical congenital adrenal hyperplasia (CAH). These family members were detected by high baseline 17-hydroxyprogesterone levels found in the course of family studies. The hormonal responses to ACTH of the family members with cryptic 21-hydroxylase deficiency were determined and compared to the responses of patients with CAH, patients with acquired adrenal hyperplasia, family members predicted to be heterozygous for CAH, family members predicted to be unaffected, and the general population. The ACTH-stimulated levels of 17-hydroxyprogesterone and delta <em>4</em>-<em>androstenedione</em> in the cryptic family members were elevated above the level of the general population or family members heterozygous for classical CAH, but below that of patients with CAH. The hormonal profile of patients with cryptic 21-hydroxylase deficiency is similar to that of patients with acquired adrenal hyperplasia. The response of family members heterozygous for the cryptic gene (21-OH CRYPTIC/21-OH NORMAL) was indistinguishable from that of family members heterozygous for the classical CAH gene (21-OH CAH/21-OH NORMAL). These studies support our previous proposal that patients with cryptic 21-hydroxylase deficiency are genetic compounds, having one gene for a severe enzyme deficiency and one gene for a mild 21-hydroxylase deficiency. Thus, the 21-hydroxylase genotype in cryptic 21-hydroxylase deficiency is 21-OH CAH/21-OH CRYPTIC.

OBJECTIVE

The production of E2 is paramount for the growth of estrogen receptor-positive breast cancer. Various strategies have been used, including the use of enzyme inhibitors against either aromatase (AROM) or steroid sulfatase (STS), in an attempt to ablate E2 levels. Both these enzymes play a critical role in the formation of estrogenic steroids and their inhibitors are now showing success in the clinic.

METHODS

We show here, in a xenograft nude mouse model, that the inhibition of both enzymes using STX681, a dual AROM and STS inhibitor (DASI), is a potential new therapeutic strategy against HDBC. MCF-7 cells stably expressing either AROM cDNA (MCF-7(AROM)) or STS cDNA (MCF-7(STS)) were generated. Ovariectomized MF-1 female nude mice receiving s.c. injections of either <em>androstenedione</em> (A(<em>4</em>)) or E2 sulfate and bearing either MCF-7(AROM) or MCF-7(STS) tumors were orally treated with STX6<em>4</em>, letrozole, or STX681. Treatment was administered for 28 days. Mice were weighed and tumor measurements were taken weekly.

RESULTS

STX6<em>4</em>, a potent STS inhibitor, completely blocked MCF-7(STS) tumor growth but failed to attenuate MCF-7(AROM) tumor growth. In contrast, letrozole inhibited MCF-7(AROM) tumors but had no effect on MCF-7(STS) tumors. STX681 completely inhibited the growth of both tumors. AROM and STS activity was also completely inhibited by STX681, which was accompanied by a significant reduction in plasma E2 levels.

CONCLUSIONS

This study indicates that targeting both the AROM and the STS enzyme with a DASI inhibits HDBC growth and is therefore a potentially novel treatment for this malignancy.

BACKGROUND

Sex steroid hormones have been proposed to play a role in the development of non-epithelial ovarian cancers (NEOC) but so far no direct epidemiologic data are available.

METHODS

A case-control study was nested within the Finnish Maternity Cohort, the world's largest biorepository of serum specimens from pregnant women. Study subjects were selected among women who donated a blood sample during a singleton pregnancy that led to the birth of their last child preceding diagnosis of NEOC. Case subjects were 41 women with sex cord stromal tumors (SCST) and 21 with germ cell tumors (GCT). Three controls, matching the index case for age, parity at the index pregnancy, and date at blood donation were selected (n = 171). OR and 95% CI associated with concentrations of testosterone, androstenedione, 17-OH-progesterone, progesterone, estradiol, and sex hormone-binding globulin (SHBG) were estimated through conditional logistic regression.

RESULTS

For SCST, doubling of testosterone, androstenedione, and 17-OH-progesterone concentrations were associated with about 2-fold higher risk of SCST [ORs and 95% CI of 2.16 (1.25-3.74), 2.16 (1.20-3.87), and 2.62 (1.27-5.38), respectively]. These associations remained largely unchanged after excluding women within 2-, 4-, or 6-year lag time between blood donation and cancer diagnosis. Sex steroid hormones concentrations were not related to maternal risk of GCT.

CONCLUSIONS

This is the first prospective study providing initial evidence that elevated androgens play a role in the pathogenesis of SCST.

CONCLUSIONS

Our study may note a particular need for larger confirmatory investigations on sex steroids and NEOC.

Previous studies of adrenal androgens and estrogens in critical illness were limited by measuring only selected sex steroids and by including men (who have confounding simultaneous changes in gonadal steroids). We evaluated relationships between changes in serum levels of cortisol (F), androgens, estrogens, and gonadotropins in 20 postmenopausal women with acute critical illness to determine if changes in adrenal androgens and estrogens paralleled gonadal axis suppression or adrenal stimulation. Two patterns of changes in sex steroids were observed. Admission serum levels of <em>androstenedione</em> (delta <em>4</em>-A), estradiol, and estrone, like F, were increased compared to healthy controls (P < 0.0001). delta <em>4</em>-A and estrone then decreased toward normal by day 5 in parallel with cortisol (r = 0.56 and 0.60). In contrast, admission serum dehydroepiandrosterone (DHEA) and DHEA-sulfate (DHEA-S) were not elevated and testosterone (T) was decreased in our patients compared to controls (P < 0.0005) in parallel with serum gonadotropin levels. Serum levels of DHEA and T continued to decrease by day 5 in parallel with gonadotropins. We conclude that in agonadal patients with acute critical illness, serum levels of DHEA-S and T are selectively decreased in relation to F, delta <em>4</em>-A, and estrogens. The decreased serum T levels suggest inhibition of 17 beta-OH-dehydrogenase and/or increased aromatization to estradiol. The marked increase in serum estrogen levels also suggests increased aromatization. The absence of increases in DHEA and DHEA-S suggest enhanced activity of 3 beta-hydroxysteroid dehydrogenase and/or inhibition of C17,20-lyase activity of P-<em>4</em>50c17. The clinical significance of this marked increase in the ratio of estrogens to androgens in acute illness requires further investigation.