Biochemical investigations have shown the presence of the enzyme phenylethanolamine N-methyltransferase (PNMT), which converts norepinephrine (NE) to epinephrine (E) in human pregnant tissues, e.g. myometrium and fetal membranes. The enzymatic activity of PNMT in myometrium is known to decrease during labor. In the present study, PNMT activity in chorio-decidua was measured and the effect of labor and prostaglandins (PG) on PNMT activity was determined. A cytosol fraction of chorio-decidua was incubated with [¹⁴C]-S-adenosyl-L-methionine and normetanephrine for 60min to measure PNMT activity. Fetal membranes were obtained at elective Cesarean section and normal vaginal delivery from full term pregnant women, and the activity of PNMT in chorio-decidua was compared. Significantly lower activity in chorio-decidua obtained from normal vaginal delivery than that from elective Cesarean section was observed. The concentration of 13, 14-dihydro-15-keto-prostaglandin F2α (DHK-PGF2α) in amniotic fluid obtained at normal vaginal delivery were measured by RIA, and the relationship of the DHK-PGF2α level to PNMT activity was studied. There was a significantly negative correlation between the concentration of DHK-PGF2α in amniotic fluid and PNMT activity in chorio-decidua. When various concentrations of PGF2α were pre-incubated with minced chorio-decidua for 2h, PNMT activity was decreased in a dose dependent manner. These results indicated that PGF2α might affect the bioconversion of NE to E in fetal membranes. The presence of this enzyme in fetal membranes may play an important role in regulating the concentration of catecholamines during pregnancy and parturition.

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To analyze the utility of insulin-like growth factor-I (IGF-I) and insulin-like growth factor binding protein-3 (IGFBP-3) radioimmunoassay in the diagnosis of growth hormone deficiency (GHD) we measured IGF-I and IGFBP-3 in sera from normal children (n=309), short children (n=99) and patients with GHD (n=73). In 80% and 93% of classical GHD (cGHD), IGF-I and IGFBP-3 levels, respectively, were below the age-related cutoff levels (lower limit). In 81% and 88% of normal short children (NS), IGF-I and IGFBP-3 levels, respectively, were above the age-related cutoff levels. Thus, both IGF-I and IGFBP-3 were good parameters for screening GHD. In contrast, in more than half of partial GHD (pGHD), either IGF-I or IGFBP-3 was above the age-related cutoff levels. The poor discrimination between patients with pGHD and NS by using these two parameters may be the result of their relatively similar GH levels, as compared to cGHD, or due to the limitations of GH stimulation tests. In about 80-90% of NS, IGF-I and IGFBP-3 were above the age-related cutoff levels at all ages. A hundred percent of cGHD under 10 years old had IGFBP-3 below the age-related cutoff levels, whereas 79% of cGHD under 10 years old had IGF-I below the age-related cutoff levels. Thus in the younger age groups, IGFBP-3 may be more sensitive than IGF-I. It may be because IGFBP-3 levels are relatively higher than those of IGF-I in younger subjects. IGFBP-3 may be less sensitive for diagnosing GHD in older children than in younger children because IGFBP-3 levels may also increase during puberty by mechanisms independent of the GH-IGF-I axis. There was a significant correlation between IGF-I and IGFBP-3 in all the subjects. However, IGF-I and IGFBP-3 classified subjects differently in 25% of patients with GHD and 19% of those with NS. This may reflect differences in daily coefficient of variation in IGF-I and IGFBP-3, in assay sensitivity and in non-GH dependent pubertal effect. The other explanation for the difference between these two parameters in terms of above and below the cutoff levels is that it may be due to the limitation of GH stimulation tests in the diagnosis of GHD.

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Three growth hormone (GH) deficient males with hypogonadotropic hypogonadism were treated with pulsatile luteinizing hormone-releasing hormone (LH-RH) administration. In two of them, the GH deficiency was idiopathic, but in the other it was secondary, caused by suprasellar germinoma. In response to LH-RH therapy, the serum testosterone(T), testicular volume, and body height increased in all three patients, and normal serum T levels and spermatogenesis were achieved in two patients. Gonadotropin responses to an LH-RH test preceding therapy did not seem to be an accurate predictor of the success of LH-RH therapy. We conclude that GH-deficient patients with hypogonadotropic hypogonadism can be expected to achieve normal pubertal development and spermatogenesis with pulsatile LH-RH administration.

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

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We describe a woman with acromegaly who had acanthosis nigricans and hirsutism. Serum growth hormone (GH) and testosterone levels were markedly elevated. Standard oral glucose tolerance test (OGTT) showed a diabetic curve and no suppressed GH levels. Fasting insulin levels were very high while plasma glucose levels were not hypoglycemic. Insulin tolerance test revealed blunted hypoglycemic response. Acanthosis nigricans was present in the right axilla and face. Ultrasonogram demonstrated bilateral polycystic changes in the ovaries. From the above findings this patient's condition is characteristic of a very rare syndrome consisting of acromegaly, polycystic ovaries (PCO), hyperandrogenism, hirsutism, insulin resistance and acanthosis nigricans.

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The effect of sustained stress on the plasma CRH level was studied in rats subjected to the stress of laparotomy conducted under ether anesthesia or water immersion-restraint. The role of AVP in ACTH secretion during such stress was also investigated. Concentrations of CRH and AVP in the hypothalamus, extrahypothalamic tissues and peripheral blood were measured by radioimmunoassays. Persistent secretion of ACTH was observed from 10 or 30min to 120min after the onset of each stress. Plasma CRH levels rose significantly 10min after the onset of ether-laparotomy stress and remained significantly elevated at 120min compared with controls. In the animals subjected to water immersion-restraint stress, plasma CRH tended to increase during the time course of the stress, reaching levels that were at least two times higher than the control. CRH concentrations in the median eminence (ME) during both types of stress decreased significantly at 120min. In the ether-laparotomy stressed rats, CRH in the neurointermediate lobe (NIL) decreased significantly at 120min, similar to the ME. Although a significant change in the adrenal CRH content was observed in the ether-laparotomy stressed rats, the involvement of adrenal CRH in ACTH secretion is unlikely as the absolute change in CRH was very small. These findings suggest that continuous CRH increase reflects a persistent secretion of CRH from the hypothalamic median eminence to the hypophysial portal vessels. It is possible that CRH secretion from the posterior pituitary gland is at least partly responsible for the persistent plasma ACTH increase in ether-laparotomy stress. Concentrations of AVP in the tissue and plasma showed no significant change during the sustained phase of each stress; the role of AVP in persistent ACTH secretion may therefore be small.

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We have investigated the effects of apoproteins on cholesterol esterase (CEase) in rat adrenal glands in order to clarify the mechanism of synthesis of free cholesterol which is the most important substrate for steroidogenesis. We prepared lipid mixtures containing cholesteryl oleate plus apoproteins with and without phosphatidylcholine as a substrate for CEase in order to investigate the effect of the substrate state on CEase. The substrate containing only cholesteryl oleate and apo-HDL increased both acid and alkaline CEase activities. Both acid and alkaline CEase activities were also increased by a substrate containing apo-HDL plus cholesteryl oleate and phosphatidylcholine more than by a substrate containing cholesteryl oleate plus apo-LDL with phosphatidylcholine or cholesteryl oleate with phosphatidylcholine. We have already reported that phosphatidylcholine is an important factor for the regulation of adrenal CEase. Therefore, the present studies show that apoproteins as well as phosphatidylcholine may be important factors for the regulation of adrenal CEase.

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We observed a patient manifesting spontaneous hypothyroidism with thyroid stimulation blocking antibody (TSBAb) long after treatment with antithyroid drugs (ATDs) for hyperthyroid Graves' disease. A 19-year-old female with Graves' disease was treated with ATDs for approximately 2 years; after cessation of ATDs, hyperthyroidism did not recur. Nine years later, she was again seen in our hospital because of symptoms indicative of hypothyroidism. Thyroid hormone replacement was commenced after laboratory confirmation of hypothyroidism. TSH-binding inhibitor immunoglobulin and TSBAb were both positive, while thyroid stimulating antibody (TSAb) was negative, at the time of diagnosis of hypothyroidism. These results indicated that the alterations in thyroid function in this patient appeared to relate to the presumed decline in the activity of TSAb and the appearance of TSBAb years after ATDs administration had been discontinued.

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The localization of galanin (GAL) was studied in the anterior pituitary of adult male macaque monkeys immunohistochemically. GAL-immunoreactive fiber bundles were noted in the posterior lobe of the pituitary. Cells immunoreactive for GAL were observed in the anterior lobe, and colocalization studies revealed that GAL-like immunoreactivity was present in gonadotrophs and thyrotrophs. The differences among the monkey, human and rat in GAL-immunoreactive localization may indicate that the regulation of GAL in the three species differs.

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The modulation of insulin release from the isolated pancreas using normal rats rendered hyperglycemic by in vivo glucose infusion was studied. In 24h and 48h hyperglycemic rats, the increase in the amount of insulin released was reduced in response to glucose challenge, but was unchanged in relation to arginine challenge. In 48h hyperglycemic rats insulin secretion after glucagon challenge was preserved. It was concluded that hyperglycemia itself makes the pancreatic B-cell selectively insensitive to glucose.

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The amounts of immunoreactive proinsulin (IRP), immunoreactive insulin (IRI), and C-peptidelike immunoreactivity (CPR) in six insulinomas and one nesidioblastosis lesion were determined together with those in the surrounding pancreatic tissue. Four non-insulinoma and nondiabetic human pancreases were used as the control. The IRP in the seven tumors ranged from 5.85 μg/g to 65.45μg/g (mean±SEM, 28.70±8.01μg/g), while the IRP in the surrounding pancreatic tissue ranged from 2.08μg/g to 11.71μg/g (5.32±1.76μg/g). Control pancreases had an IRP content of 12.01±2.36μg/g. The IRI in the seven tumors ranged from 4.02U/g to 47.97U/g (14.40±6.35U/g), while that in the surrounding pancreatic tissue ranged from 0.28U/g to 3.64U/g (2.32±0.63U/g). Mean tumor CPR was 206.84±81.6μg/g and it was 29.16±9.15μg/g in the surrounding pancreatic tissue. The molar ratio of the IRP to IRI content was 6.83±1.95% for tumor tissue and 6.24±2.18% for the surrounding pancreatic tissue. These levels were similar to the ratio in the control pancreases (7.67±1.88%), in contrast to the higher serum IRP/IRI ratio in the tumor patients.

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To further elucidate the role of the preoptic-hypothalamic region in fluid and electrolyte balance we studied the effect of surgical preoptic-hypothalamic disconnection using either a large (preoptic-hypothalamic disconnection) or a small (medial preoptic-hypothalamic disconnection) microknife. Both the large and small cuts seemed to transect the posterior projection originating in the periventricular tissue surrounding the anteroventral third ventricle (AV3V) and extending to supraoptic nucleus, but the supraoptic-neurohypophysial pathway was severed only by the large cut. Seven-day metabolic studies showed a disruption in hydromineral balance only in large cut rats; they had increased water intake and urine volume on day 1, a near-recovery of function on days 2 and 3, and polydipsia and polyuria on days 4 to 7. There was no difference between small cut rats and sham-operated rats in metabolic measurements. The large cut rats also had sustained hypernatremia and hyperosmolality, which was enhanced after water restriction for 48h but was not accompanied by an increase in plasma arginine vasopressin. Our data therefore suggest that the efferent fibers running caudally from the AV3V are not involved in mediation of the hydromineral regulation of the AV3V.

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An elevated plasma atrial natriuretic peptide (ANP) concentration was observed in patients with chronic renal failure, and it was significantly (P<0.01) decreased by hemodialysis with or without fluid removal. The ANP concentration was decreased by dialysis without fluid removal and the decrease was significantly (P<0.01) correlated with the pre-dialysis value. The decrease in this peptide during fluid removal without diffusion was significantly (P<0.05) correlated with the decrease in the circulating plasma volume measured by dye dilution. The decrease in the ANP level was therefore considered to be related to a fall in right atrial pressure caused by the decline in the circulating plasma volume. The circulating plasma volume, osmotic pressure, and blood pressure were not changed by hemodialysis without fluid removal. Moreover, the filtrate concentration of ANP (mean: 5.5pg/ml) during fluid removal without dialysis was only about 8% of the plasma level, so filtration of ANP from the dialyzer was negligible. The decrease in ANP during hemodialysis without fluid removal may therefore have been caused by the removal of or a change in the level of substances, for example electrolytes, epinephrine, and uremic toxins. In addition to volume expansion, such a substance(s) might influence plasma ANP levels in patients with chronic renal failure.

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We have previously shown that methotrexate (MTX) and hydroxyurea (HU) stimulate expression of the human chorionic gonadotropin alpha and placental alkaline phosphatase genes and repress the expression of the c-myc oncogene in BeWo choriocarcinoma cells. In order to determine whether c-myc downregulation played a role in the induction of these placental genes, we treated BeWo choriocarcinoma cells with aphidicolin (APH), and 6-diazo-5-oxo-L-norleucine (DON), and compared the effects of these drugs with that of MTX. All of these drugs downregulate c-myc gene expression. At the doses used, both APH and DON repress c-myc expression to approximately the same extent, as well as inducing morphologic changes in BeWo similar to that of MTX, although neither stimulated hCG alpha expression. Both DON and APH stimulate placental alkaline phosphatase gene expression, but only MTX and DON stimulate cholesterol side chain cleavage enzyme gene expression. This indicates that the downregulation of c-myc gene expression is insufficient to stimulate the expression of all the placental genes stimulated by MTX.

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Remission of diabetes, i.e. significant amelioration from absolute insulin-dependency, has been sometimes observed in diabetic BB/Wor//Tky rats which were treated with insulin. In remission BB/Wor//Tky rats, plasma glucose levels improved to near normal level and insulin content was also preserved as much as that between diabetic and non-diabetic rats. In this process, we hypothesized that autoimmune insulitis was suppressed and remaining islet B-cells was restored from severe destruction by recovering in number and/or function. While, recently, a novel regenerating (reg) gene, identified in the regenerating pancreatic islets of surgical models, is reported to be related to the replication of pancreatic B-cells in vitro. Based on these findings, we histologically investigated whether the reg protein could be actually expressed or not in the islets from remission BB/Wor//Tky rats. As expected, reg protein was observed in the islets from remission BB/Wor//Tky rats mainly in accordance with pancreatic B-cells. Thus, the present findings suggested that the regeneration of pancreatic B-cells represented by the expression of reg protein might be, at least in part, relevent to remission induced by insulin therapy in spontaneously occurring Type 1 diabetes in BB/Wor//Tky rats.

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