The present study was undertaken to determine the differences in steroidogenic potential of isolated granulosa cells from normal and polycystic ovaries in vitro. Granulosa cells isolated from human ovarian follicles were cultured in a chemically defined medium containing gonadotropins and/or testosterone for 18 days.
The effect of human LH on progesterone (P) production of granulosa cells from normal ovaries in the mid-follicular phase was dose-dependent in the concentration ranging from 1 to 100ng/ml. The cultured granulosa cells from normal ovaries responded significantly to LH (100ng/ml) by showing an 8- to 20-fold increase in P production, whereas exogenous LH stimulated P production in the cultured granulosa cells from polycystic ovaries to a much lesser extent (2- to 4-fold increase).
The accumulation of estradiol (E₂) by the cultured granulosa cells, regardless of the follicular size, was very low, if any, in the absence of testosterone and/or FSH. In the absence of FSH, granulosa cells from large follicles in the preovulatory stage were more active (10-fold) in aromatization of exogenous testosterone (T: 100ng/ml) than those from medium-sized follicles in the mid-follicular phase. The addition of FSH alone failed to stimulate E2 biosynthesis in the absence of T, but exogenous FSH (200ng/ml) in the presence of T induced a marked increase (10-fold) in aromatase activity of the cultured granulosa cells from medium-sized follicles in the midfollicular phase. The cultured granulosa cells from polycystic ovaries, and also those from normal follicles under 6mm in diameter in the early and mid-follicular phases of the menstrual cycle, had the capacity to biosynthese E₂ in the presence of exogenous T and FSH.
These results strongly suggest that an intrinsic defect in the aromatase enzyme system may not exist in patients with polycystic ovaries, and that the persistent elevation of serum LH may lead to a disturbance of P production in response to exogenous LH in the cultured granulosa cells from polycystic ovaries.

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The effect of calcitonin (CT) on calcium content and enzyme activity in the hepatic mitochondria of intact rats was investigated. A single subcutaneous administration of CT (80 MRC mU/100g BW) produced a significant increase in the content of calcium, the activity of pyruvate carboxylase, succinate dehydrogenase and ATPase 15min after the hormone treatment. The significant increases in calcium content and pyruvate carboxylase activity were also observed 30min after CT administration, while succinate dehydrogenase and ATPase activity began to decrease. A physiological dose of CT (20 MRC mU/100g BW) caused a marked increase in calcium content and pyruvate carboxylase activity but not succinate dehydrogenase or ATPaseactivity. The removal of calcium by 10mM EGTA washing of the mitochondria produced a remarkable reduction in pyruvate carboxylase activity increased by CT administration. The addition of calcium ion of 2.5×10^-2-2.5×10¹nmoles Ca²⁺ per mg mitochondrial protein produced a marked increase in pyruvate carboxylase activity. The present results suggest that calcium taken up by the hepatic mitochondria after CT administration activates pyruvate carboxylase.

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The effect of gonadotropin and endogenous estrogen on the growing follicles (especially in relation to the presence or absence of theca cell layers) was studied morphometrically in intact immature female rats, using a chemical antiestrogen.
When pregnant mare serum gonadotropin (PMSG) was given in combination with clomiphene, the uterine weight decreased significantly as compared with that in the animals treated with PMSG alone (p<0.001). The administration of clomiphene and PMSG did not have a significant effect on the total number of follicles (growing plus atretic). The number of atretic follicles in those treated with PMSG alone tended to decrease as compared with sesame oil-treated animals. However, when PMSG was given in combination with clomiphene, the number of atretic follicles increased significantly as compared with the animals treated with PMSG alone (p<0.05). The number of growing follicles in the animals treated with PMSG alone tended to increase as compared with the animals treated with neither PMSG nor clomiphene. In contrast, when PMSG was given in combination with 5mg/day of clomiphene, the number of growing follicles tended to decrease. The administration of PMSG alone and PMSG combined with 1 and 5mg/day of clomiphene caused a significant decrease in the number of growing follicles without theca cell layers, as compared with sesame oil-treated animals (p<0.05, p<0.01 and p<0.001, respectively). On the other hand, the number of growing follicles with theca cell layers was increased markedly by the administration of PMSG alone (p<0.05).
It is suggested that the early stage of follicular growth in the absence of theca cell layers may be promoted by endogenous estrogen. In the presence of theca cell layers, in addition to being mediated by estrogen, gonadotropin-induced follicular growth may be promoted by the maturative effect of gonadotropin itself.

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The binding of R 1881 in cytosols from the normal, benign hypertrophic and cancerous human prostates was examined in the presence of molybdate and triamcinolone acetonide. The addition of 10mM Na₂MoO₄ resulted in an increase in the stability of the binder without any change in Kd. Triamcinolone acetonide added to the incubation of cytosol with R 1881 modified the inhibition pattern by other steroids, and the binding in the presence of triamcinolone acetonide exhibited the characteristics of androgen receptor. The complex of cytosol and R 1881 formed in the incubation in the presence of triamcinolone acetonide was sedimented at 8.5S.
Kd of the binding to R 1881 of the normal and pathological prostates was almost identical, but maximum binding sites of the androgen receptor were larger in benign hypertrophic and cancerous prostates than in normal tissues. For the assay of binding capacity in needle biopsy specimens, one point determinations were performed using 2.5nM R 1881 as the ligand. The number of binding sites obtained by this method were well correlated with those obtained by the Scatchard plot in various prostatic tissues.

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A variety of complications are encountered in primary aldosteronism. Among them, less well known is the association of functioning adrenocortical adenoma with neoplasia of other organs. Described here is a patient with primary aldosteronism who had undergone a surgical operation for thyroid cancer and died of menigoencephalitis due to Listeria monocytogenes. Moreover, the autopsy revealed that he had an adenocarcinoma of the rectum. To our knowledge, this is the first documented case of a patient who had both primary aldosteronism and a primary double cancer of the thyroid and rectum.

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Binding proteins for 1.25 (OH) ₂D₃ were investigated in thirty breast cancers. Human breast cancer was shown to contain specific, high affinity cytosol binding proteins for 1.25 (OH) ₂D₃ and 25 (OH) D₃. The binding protein for 1.25 (OH)₂D₃ sedimented at 3.7S and the binding protein for 25 (OH) D₃ at about 6.0S on sucrose density gradient analysis containing 0.3M KCI and 1mM dithiothreitol in buffer. Kd for 1.25 (OH)₂D₃ were from 0.1×10^-11M to 7.17times;10^-11M measured by Scatchard plots. Competition binding studies indicated that the relative specificity of the binding protein for 1.25 (OH) ₂D₃>25 (OH) D₃<1α(OH) D₃, 24, 25 (OH) ₂D₃>D₃> Estradiol-177beta;. 1.25 (OH)₂D₃ receptor-positive was detected in twenty-eight out of thirty breast cancers.

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The blood levels of progesterone (P) and estradiol (E₂) were artificially controlled in rats ovariectomized on days 19-20 of pregnancy (day 1: sperm in vaginal smears) by 48-168h intravenous infusions according to 15 different temporally variable programs. The predominance of P infused in various curves for longer than 72h, caused postmature deliveries on days 24-26 accompanying difficulties in labor and the lack of lactation, although changes in E₂ levels antagonistically accelerated the timing of delivery. The most successful delivery and subsequent lactation were induced by an infusion of total amount of 1.5mg P for 48h (days 19-21) at a linearly decreasing rate in combination with an infusion of a total amount of 875ng E₂ for 72h (days 19-22) at a slowly increasing rate, reaching a plateau in the morning of day 21 and rapidly decreasing to nil after that. Spontaneous delivery occurred on day 22 with a normal range of labor duration and no incidence of stillbirth. No further supply of sex steroids was required for the maintenance of lactation. Blood levels of prolactin shortly after delivery were much higher in these lactating ovariectomized rats than in non-lactating animals. The neuroendocrine dynamics at periparturient periods, which triggers the termination of pregnancy and then the induction of parturition and lactation, is discussed in relation to the results of these simulation experiments.

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The release of human chorionic gonadotropin (hCG) and its α-subunit (hCG-α) from the normal human placenta and the effect of some stimulatory agents on their release were studied in vitro using a perifusion system. Each perifusate was assayed for hCG and hCG-α in its own homologous radioimmunoassay systems.
Both hCG and hCG-α were released from the placenta at any stage of gestation in our perifusion system. Much more hCG than hCG-α was released from the placenta in early gestation. By comparison, however, hCG-α increased gradually with the gestational age. The amount of hCG-α released was almost equal to that of hCG in the placenta in the 17th gestational week. After the 22nd gestational week, hCG-α was released in larger quantities than hCG, and about 10times more hCG-α than hCG was released from the term placenta.
These results were also confirmed by gel filtration of perifusates on a Sephadex G-100 column. hCG-α, compared with hCG, was present in excess in gel filtrated perifusates in the last two trimesters.
By adding 1mM dibutyryl cyclic AMP to the perifusion medium, the release of both hCG and hCG-α was stimulated significantly. Synthetic luteinizing hormone releasing hormone (LH-RH) at concentrations of 10ng/ml and 100ng/ml had no effect, but at a high concentration (1μg/ml), LH-RH stimulated the release of them. Moreover, mouse epidermal growth factor (EGF) stimulated not only the release of hCG and hCG-α but also their production, because both hCG and hCG-α levels rose progressively with the time course in the presence of EGF.
The present studies demonstrate that the perifusion system of chorionic tissues is a useful method for investigating the release of hCG and its subunits in vitro.

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In the present study, the role of the medial preoptic area (MPO) in the neural control of the nocturnal prolactin (PRL) surge was investigated in ovariectomized rats. Cervical stimulation (CS) or bilateral MPO lesions caused a marked nocturnal PRL surge at 0400h on the fourth day after CS or the lesions in ovariectomized rats in which blood samples were obtained by decapitation. However, operation for indwelling a catheter and serial blood collection completely eliminated the MPO lesioninduced nocturnal surge while they did not affecthe CS-induced surge. On the other hand, MPO lesions could not induce the nocturnal PRL surge in neonatally androgenized female rats. These resultsuggest that the MPO not only tonically inhibits the initiation of the nocturnal PRL surge but also has a buffer action on the PRL surge-suppressing action of stress. Furthermore, it may be possible that the failure of CS to initiate and maintain the nocturnal PRL surge in neonatally androgenized rats is not due to the inability of CS to disinhibit the inhibitory action of the MPO, but rather due to the extinction of the circadian rhythm itself of the nocturnal PRL surge in these rats.

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The metabolism of 3, 5-[3'-125I] triiodothyronin (T₃) and 3-[3', 5'-125I] triiodothyronine (rT₃) was studied in cultured monkey hepatocarcinoma cells (NCLP-6E), and the deiodinations of these iodothyronines were also investigated in cultured cell homogenates and in rat liver homogenates. The metabolites were analyzed by ionexchange column chromatography.
For nonphenolic ring deiodination of 3, 5-[3'-125I] triiodothyronine, the order of the inhibitory effect of excess unlabeled iodothyronine or its analog was as follows: 3, 3', 5-tiiodothyronine>triiodothyroacetic acid>tetraiodothyroacetic acid>thyroxine. This order did not differ between in the intact cells (NCLP-6E) and their homogenates.
The order of effectiveness of the excess unlabeled compounds on phenolic ring deiodination of 3-[3', 5'-125I] triiodothyronine in the intact cells was as follows: tetraiodothyroacetic acid>triiodothyroacetic acid, 3, 3', 5-triiodothyronine>thyroxine. This order was the same among monkey hepatocarcinoma cell homogenates, rat hepatoma cell homogenates and rat liver homogenates, and triiodothyroacetic acid was obviously more effective than 3, 3', 5-triiodothyronine.
It was concluded that 3, 3', 5-triiodothyronine had the highest affinity for nonphenolic ring deiodinase among iodothyronines and their analogs used in the present study and that tetraiodothyroacetic acid had the highest affinity for phenolic ring deiodinase. It seems, therefore, that the metabolites derived from the thyroid hormones might contribute to deiodinations which involve activation and inactivation of the hormones.

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Serum thyroxine-binding globulin (TBG) in 169 patients with various cancers was determined by radioimmunoassay (RIA). Eleven patients showed a high serum TBG level (>35μg/ml). Two of them had been treated with estrogen for prostate cancer. One patient had high serum TBG with serum hepatitis. Another 8 cases had normal liver function and also normal levels serum estrogen. Thus, about 4.7%(8/169) of the cancer patients had high serum TBG and mild hyperthyroxinemia caused by unknown mechanisms.
The high TBG level in these patients continued until just before death, or in some cases decreased to nomal after removal of cancer tumors by operation.
Caner is occasionally associated with an increase in serum TBG. Although the mechanism is not clear, the increased TBG in the cancerous state is interesting and has significance as a tumor marker.

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In order to clarify the role of the pituitary conversion of L-thyroxine (T₄) to 3, 5, 3'-L-triiodothyronine (T₃) in regulating thyrotropin (TSH) secretion, the effect of altered thyroid states and fasting on intrapituitary T₃ generation was investigated by a paperchromatographic procedure using the anterior pituitary homogenates. Hepatic T₃ generation was also studied for comparison.
The rate of pituitary and hepatic T₃ generation in normal rats averaged 25.2±12.4 (mean±SE) fmoles T₃/mg protein/min and 33.8±12.7 fmoles T₃/mg protein/min, respectively. T₄ treatment raised the hepatic T₃ generation from T₄ (46.7±3.1 fmoles T₃/mg protein/min) and lowered the intrapituitary T₃ generation (4.5±0.2 fmoles T₃/ mg protein/min). On the contrary, thyroidectomy slowed the hepatic T₄ 5'-deiodination (11.0±2.8 fmoles T₃/mg protein/min), and accelerated the pituitary T₄ 5'-deiodination (64.3±1.4 fmoles T₃/mg protein/min). In 48h fasted rats, serum T₄, T₃ and TSH concentrations were all lower than those in fed rats, and both pituitary and hepatic T₃ generations were also suppressed.
Thus, altered thyroid states cause an opposite effect on pituitary and liver 5'-monodeiodination, whereas fasting causes similar changes. The findings suggest the existence of an autoregulatory mechanism for thyroid hormone activation within the target tissues.

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The tubulin content and biochemical components were determined in the cerebrum, cerebellum and hypothalamus from intact and T₃-treated male and female rats during early life.
T₃-treatment between 0 and 9days of age increased soluble protein, RNA, DNA and tubulin content (mg per g tissue) in the 10-day-old male cerebellum but not in the cerebrum and hypothalamus except for soluble protein and tubulin (mg per g tissue), respectively. Intracellular tubulin content (mg per mg DNA) was increased by the T₃-treatment in the 10-day-old male hypothalamus but not the other regions. When T₃ was administered between 10 and 19days, there was little effect of the treatment; increased tubulin (mg per g tissue) in the cerebrum and decreased RNA (mg per g tissue) and a ratio of tubulin to protein in the cerebellum from 20-dayold males. Less response to T₃-treatment was observed in female cerebrum and hypothalamus but not in the cerebellum, compared with the male.
These results suggest that the effect of T₃-treatment on brain is modified by several factors such as tissue specificity, age-dependency and sexual differences. Modification by these factors might depend, at least in part, on changes in the number of T₃-receptors due to the hormone treatment.

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The role of the lower brain stem in controlling preovulatory gonadotropin surges was investigated in female rats under acute experimental conditions. Electrolytic lesions or diethyldithiocarbamate implantations in the ventrolateral part of the medulla oblongata (VLMO), which were carried out at 1100-1330h on the day of proestrus, resulted in a blockade of the preovulatory surges of LH, FSH and PRL as well as subsequent ovulation. Such treatments in the dorsomedial part of the medulla oblongata did not affect gonadotropin surges or ovulation. By means of electrolytic lesions in the VLMO, norepinephrine concentrations were significantly reduced in the preoptic- anterior hypothalamic area at 1700-1800h on proestrus, though they did not change in the mid-posterior hypothalamus. Electrochemical stimulations of the suprachiasmatic part of the preoptic area or norepinephrine injections into the third ventricle at 1400-1500h on proestrus in animals with VLMO lesions succeeded in induce gonadotropin surges and ovulation.
These results suggest that the lower brain stem is involved in the induction of preovulatory gonadotropin surges and that the process may be mediated by the ascending noradrenergic system which originates in the VLMO.

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AA 560, one of potent nonsteroidal antiandrogens, was implanted in the arcuate nucleus of the hypothalamus of intact male rats through a stainless double cannula. The prostate weight and serum LH level significantly increased on the 5th day after the implantation of AA 560 when compared with those in the sham-operated animals. There existed a definite correlation between the weight of the ventral prostate and the serum LH level in AA 560 implanted animals.

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The effect of corticosteroids on the inception of remission was investigated in 10 patients with methimazole induced agranulocytosis. While remission started 6.00±0.92 days (mean±SE) after discontinuation of methimazole in 6 patients to whom corticosteroids were administered immediately after the cessation of methimazole treatment, it started at a significantly later date (9.25±0.48days after discontinuation of methimazole) in 4 patients in whom the corticosteroid treatment was started 5.5±0.5 days after methimazole ceased to be used. The results suggested that corticosteroids may quicken the recovery from methimazole-induced agranulocytosis.

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An attempt was made to purify androgen receptor of male rat liver cytosol which binds specifically with testosterone and androstenedione. A 30% ammonium sulfate precipitation afforded more than 5-fold purification of the androgen receptor and removed half of the amount of the enzymes involved in androgen metabolism.

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