We intended to determine how the liver copes with the massive handling of lipids induced by OE (oleoyl-oestrone), as well as to characterize and differentiate the actual OE effects from those that may be only the consequence of decreased food intake. Thus we used male rats treated with oral OE (10 nmol/g per day) compared with a vehicle only PF (pair-fed) group and controls fed ad libitum (vehicle only). Plasma parameters, and total liver lipids, glycogen, DNA and total mRNA were measured. RNA was extracted and used for real-time PCR analysis of the gene expression of enzymes and regulatory factors of liver energy metabolism. Most hepatic proteins showed similar gene expressions in OE and controls, but the differences widened between OE and PF rats, showing that OE effects could not be merely attributed to a lower energy intake. The liver of OE-treated rats largely maintained its ability to mobilize glucose for the synthesis of fats; this was achieved in part by a peculiar combination of regulative modifications that facilitate both fatty acid disposal and restrained glucose utilization under conditions of limited food supply but ample availability of internal energy stores. In conclusion, the results presented suggest that the effect of OE on liver metabolism may be (at least in part) mediated through an insulin-sensitivity-dependent modulation of the expression of SREBP-1c (sterol-regulatory-element-binding protein-1c), resulting in the unique combined effect of mildly increased (or maintained) glucose disposal but also limited enhancement of lipogenesis.

Extracts of black cohosh (Actaea racemosa) and soy are used as 'natural' alternatives to conventional hormone replacement therapy (HRT) and there is some evidence that soy may protect against breast cancer by inhibiting the production of active oestrogens. This study compares the action of ethanolic extracts of black cohosh (BCE) and genistein on growth and enzyme activity in MCF-7 and MDA-MB-123 breast cancer cells. BCE inhibited growth at the two highest doses tested, i.e. 50 and 100 microg/ml, whilst genistein stimulated growth in the oestrogen receptor positive (ER(+)) MCF-7 cells, but at high doses it inhibited growth in both cell lines. BCE did not affect the conversion of androstenedione to oestradiol and only the highest doses (50 and 100 microg/ml) significantly inhibited the conversion of oestrone to oestradiol in MDA cells. In contrast, BCE induced a dose-dependent inhibition of the conversion of oestrone sulphate to oestradiol in both cell lines, whilst in human granulosa lutein (GL) cells enzyme activity was only inhibited at the highest dose of BCE. Genistein had no significant effect on enzyme activity in breast cancer cells and like BCE only the highest doses (10 and 50 microM) inhibited enzyme activity in human GL cells. In vivo genistein may have growth stimulatory effects on breast tissue but BCE not only inhibits growth but inhibits the conversion of oestrone sulphate to active oestradiol, considered by some, to be the preferred pathway of oestradiol synthesis in breast tissue.

The uptake, metabolism and subcellular distribution of oestradiol and oestriol in endometrial, myometrial and vaginal tissue of postmenopausal women under physiological conditions were studied by giving 3H-labelled oestradiol or oestriol in subphysiological doses by continuous infusion lasting 12 h before hysterectomy. The three tissues obtained from each woman were separated into three fractions: two cytosol fractions (free oestrogens and specifically bound) and one nuclear fraction. The results show an accumulation of both oestrogens in the target tissues, we found an approximately 33 times higher [3H]E2 concentration in endometrium (dpm per g) than in plasma (dpm/ml), 20 times in myometrium and 10 times in vaginal tissue. After the E3 infusions the tissue/plasma gradient was 37 for endometrium, 19 for myometrium and 11 for vagina. In plasma and tissues a metabolite of E3 could tentatively be identified as 16 alpha-hydroxyoestrone. The subcellular distribution showed that 60-80% of E2 and E3 is accumulated in the nuclear fraction of all tissues studied, no nuclear bound oestrone could be detected. From these results the conclusion was drawn that oestradiol still is the major tissue oestrogen in postmenopausal women and that it is mainly nuclear bound. Endometrium of postmenopausal women accumulates higher concentrations of E2 and E3 than vaginal tissue from the same individual, no preferential uptake of oestriol occurs under physiological conditions.

1. The 17beta-hydroxy steroid dehydrogenase was solubilized during haemolysis of erythrocytes and was isolated from the membrane-free haemolysate. Membrane preparations isolated in different ways did not contain 17beta-hydroxy steroid dehydrogenase activity. The 17beta-hydroxy steroid dehydrogenase activity in the haemolysate was concentrated by repeated ammonium sulphate precipitation and gel filtration on Sephadex G-150. The 17beta-hydroxy steroid dehydrogenase activity of the purified preparation per unit weight of protein was 350-3000 times higher than the activity of the crude erythrocyte haemolysate. The 20alpha-hydroxy steroid dehydrogenase activity was lost during this purification procedure. 2. The 17beta-hydroxy steroid dehydrogenase was NADP-dependent and had a pH optimum for conversion of testosterone between 8.5 and 10. For the molecular weight of the enzyme a value of 64000 was calculated from Sephadex chromatography results. 3. p-Chloromercuribenzoate inhibited the enzymic activity. The oxidative activity of the enzyme for the 17beta-hydroxyl group was only partly inhibited when a large excess of 17-oxo steroids was added. The catalysing activity of the enzyme was influenced by the NADP(+)/NADPH ratio. The oxidation of the 17beta-hydroxyl group in the presence of NADP(+) proceeded faster than the reduction of the 17-oxo group with NADPH. When both reduced and oxidized cofactors were present the oxidation of the 17beta-hydroxyl group was inhibited to a considerable extent. 4. The enzyme had a broad substrate specificity and not only catalysed the conversion of androstanes with a 17beta-hydroxyl group, or 17-oxo group, but also the conversion oestradiolleft arrow over right arrowoestrone. In addition the steroid conjugates dehydroepiandrosterone sulphate and oestrone sulphate were also converted. There were no indications that more than one 17beta-hydroxy steroid dehydrogenase was present in the partially purified preparation.

The aim of the present study was to investigate the hypothesis that adrenal androgens are contributory to the development of endometrial cancer either by the oestrogenic action of 5-androstene-3 beta, 17 beta-diol (androstenediol) or through the inhibition of oestradiol metabolism. Concentrations of androstenediol, dehydroepiandrosterone (DHA), DHA sulphate (DHAS), oestrone and oestradiol were measured in plasma and endometrium from postmenopausal women with and without endometrial cancer. There was no difference between normal postmenopausal women and endometrial cancer patients with respect to either tissue or plasma adrenal androgens although there was a tendency for plasma DHAS levels to be increased in cancer patients (normal women: 640 +/- 156 ng/ml; cancer patients: 808 +/- 159 ng/ml). There was a positive correlation between endometrial tissue concentrations of androstenediol and DHA in both normal women (P less than 0.05) and cancer patients (P less than 0.01) but for DHAS the relationship was only significant for non-malignant tissue (androstenediol: DHAS, P less than 0.05; DHA: DHAS, P less than 0.02). A significant positive correlation was found between all three plasma adrenal androgens for both groups. In cancer patients there was a trend towards an inverse correlation between endometrial tissue concentrations of DHAS and the enzyme 17 beta-hydroxysteroid dehydrogenase (17OHSD) although the relationship was not significant (r = 0.49). In endometrium, oestradiol was present in significantly higher concentrations than oestrone whereas in plasma the reverse was the case. There was also a tendency for plasma oestradiol levels to be elevated in the cancer subjects. These data do not support a substantial role for adrenal androgens in endometrial cancer but suggest that a relationship may exist between DHAS and 17OHSD and that an imbalance between sulphatase and sulphotransferase activities may be involved.

Prolactin (PRL), follicle stimulating hormone (FSH), luteinizing hormone (LH), oestrone (E1), and oestradiol (E2) levels were determined in 204 women who were receiving hormone replacement therapy for their climacteric symptoms. The changes in these hormone levels and the endometrial morphology were studied in order to determine the effects of the replacement therapy. The women were divided into two groups: the first group of 120 women was treated with conjugated oestrogens administered cyclically, plus norethisterone acetate. The second group of 84 women received oral oestriol succinate, also administered cyclically but without additional progestogens. The oestrogen-progestogen therapy resulted in a disappearance of the climacteric symptoms and a significant decrease of FSH and LH levels. Oestriol therapy was less effective than the conjugated oestrogens as a replacement therapy. Oestriol therapy also resulted in a less remarkable decrease of gonadotrophin levels. There were no significant changes in prolactin levels in either group of women. The endometrial histology did not change significantly after either of the two hormone replacement therapies.

Relationship between free oestrone and boar taint compounds in adipose tissue were studied in two groups of entire male pigs of different breeds. Group A consisted of 33 entire crossbred male pigs (dam Yorkshire and sire backcross Yorkshire x Wild Boar, generation seven). Group B consisted of 194 entire male pigs of crossbreeds between Swedish Hampshire (H) and Finnish Landrace (L), LH x H, H x LH, LH x LH (dam x sire). The measurements of free oestrone in adipose tissue were performed with a new method developed and validated in our laboratories. The standard curve was linear for concentrations of free oestrone ranging from 0.13 to 5.10 ng/g. The method exhibited parallelism of results between serial dilutions and a mean recovery of 97 +/- 13.7%. Intra-assay variations for samples with concentrations of free oestrone from 0.67 to 2.08 ng/g were from 9.23 to 11.94%. Inter-assay variations for the samples with concentrations of free oestrone from 0.89 to 2.96 ng/g were from 3.78 to 10.11%. The levels of free oestrone in fat from group A were well correlated with fat levels of androstenone (r = 0.66; p < 0.001) and levels of oestrone sulphate in peripheral plasma collected at the same time as the fat (r = 0.74, p < 0.001). The levels of free oestrone in fat from group B were significantly correlated to fat levels of androstenone (r = 0.68, p < 0.001) and skatole (r = 0.29, p < 0.001). In group B, age-related differences in fat levels of free oestrone, androstenone and skatole were studied. Free oestrone and skatole levels increased simultaneously at the age of 22 week (p < 0.05 for both), and androstenone levels increased at the age of 26 week (p < 0.05). It was suggested that the levels of free oestrone in adipose tissue might be used for the evaluation of hormonal status of male pigs as an alternative to plasma levels of testicular hormones. The levels of free oestrone might be involved in the regulation of skatole levels in fat as indicated by both the simultaneous increases in skatole and free oestrone levels in fat and positive correlation between skatole and free oestrone.

Spontaneous canine mammary inflammatory carcinoma (IMC) shares epidemiologic, histopathologic and clinical characteristics with the inflammatory breast carcinoma (IBC) disease in humans. We have analysed the steroids levels in serum and in tissue homogenates of IMC, the expression of two of their receptors (androgen and beta-estrogen) and of three enzymes included in the steroidogenesis pathway (aromatase (CYP19A1), steroid sulphatase (STS) and estrogen sulfotransferase (EST)) trying to explain the specific accumulation of steroids in IMC tissues generating deposits in the form of lipid droplets whose presence can be attributed to steroids secreted by IMC cells. According to our working hypothesis, oestrone sulphate would be the main component of these lipid droplets. The presence of these steroid deposits would contribute to the intense proliferation and invasive behaviour of IMC and IBC, although their involvement in angiogenesis is yet to be demonstrated.

Two experiments in vivo and one experiment in vitro were conduced to examine the mechanisms involved, which lead to mammary secretion of oestrogens and its importance for milk production and udder health in cows. In experiment 1 in six cows of the White-Black breed on day 268 of pregnancy catheters were inserted into uterine vein of pregnant horn, the abdominal aorta and the caudal superficial epigastric (milk) vein. Blood samples for estimation of oestrone, oestrone sulphate, oestradiol-17alpha and -17beta by RIA were obtained daily from day 7 pre-partum until day 1 post-partum. Only the concentration of oestradiol-17beta was statistically higher (P< or =0.01) in mammary venous plasma than in aortal and uterine plasma. In experiment 2 forty late-pregnant cows were divided into two groups according to their milk production in the previous lactation: group 1 (n=20) high-yielding cows (>6500kg milk per lactation), and group 2 (n=20) low-yielding cows (<3700kg milk per lactation). Blood samples for measurement of oestradiol-17beta by RIA were collected from milk and tail veins every fourth day during a period from day 20 prior to parturition to day 4 post-partum. The concentration of oestradiol-17beta was significantly higher (P< or =0.01) in the milk vein than in the peripheral plasma from day 12 pre-partum to parturition. In high-yielding cows the level of oestradiol-17beta in mammary venous blood was significantly higher (P< or =0.01) than in low-yielding cows. In six cows with pathological udder oedema ante-partum the concentration of oestradiol-17beta in milk vein was significantly higher (P< or =0.05) than in control cows. There were no statistically significant differences in the levels of oestradiol-17beta in cows with clinical mastitis (n=10) during 2 weeks after parturition and without it (P> or =0.05). In an in vitro experiment, homogenates of mammary tissue collected on day 7 pre-partum from two cows were incubated with 3H-androstendione. After incubation the samples were extracted and 3H-oestradiol-17beta was separated by HPLC. 3H-oestradiol-17beta was formed in a total yield of 37%. These results indicate that oestrone, oestrone sulphate and oestradiol-17alpha are not secreted by bovine mammary gland. Furthermore, the secretion of oestradiol-17beta starts about day 12 pre-partum and is associated with milk yield and udder oedema. Preliminary in vitro study suggests the synthesis of oestradiol-17beta by mammary tissue.

The EURODIAB Complications Study, a clinic based epidemiological project including 3250 individuals with type 1 diabetes from 31 European centres analysed the natural dietary fibre intake and possible benefits for patients with diabetes. The mean intake of natural dietary fibre in the cohort of patients with type 1 diabetes was 17.3 g/day for all centres with a centre range of 13.9-21.9 g/day. The fibre consumption was lowest in patients from Eastern European centres compared to patients from centres in Southern and North-Western Europe. The fibre density was highest in patients from Southern Europe. Total fibre intake was significantly inversely related to HbA1c levels; severe ketoacidosis risk fell significantly with higher fibre intakes. Higher intakes of total fibre were independently associated with significantly higher levels of high density lipoprotein-cholesterol in male and in female patients. Fibre intakes in men with diabetes were also inversely related to ratios of total cholesterol to high density lipoprotein-cholesterol and to levels of low density lipoprotein-cholesterol. Higher fibre intakes are also associated with decreases in plasma oestradiol and oestrone levels. A protective effect of total fibre intake against cardiovascular disease was observed in females but not in males with diabetes.

Successful chronic cannulation of the foetal posterior vena cava and maternal utero-ovarian and jugular veins in five Jersey cows between days 240 and 260 of gestation enabled changes in plasma hormone levels preceding calving to be monitored. All cows delivered live calves within the expected range of gestation for the breed. Corticosteroids were assayed by competitive protein-binding and prostaglandin F, progesterone, oestrone and oestradiol-17beta by radioimmunoassay. Foetal corticosteroids rose slowly from 5.0 +/- 0.7 ng/ml at 20 days to 9.3 +/- 3.0 ng/ml at 10 days before term, then progressively increased to a mean of 74 ng/ml, though higher concentrations occurred following surgery. Foetal oestrone and oestradiol-17beta concentrations were both less than 50 pg/ml and showed little change toward term. The maternal utero-ovarian oestrogens increased slowly from 20 to 10 days pre-partum and then rose more rapidly reaching peak levels (2.9 +/- 0.6 ng/ml for oestrone and 1.4 +/- 0.3 ng/ml for oestradiol-17beta) 1 to 4 days before delivery. Maternal progesterone concentrations fell towards term, with a rapid decrease over the last 36-48 h before calving when they gradually increased until the last 24 h where was a dramatic rise, reaching peak levels (5.7 +/- 0.6 ng/ml) during labour.

Non-invasive methods for monitoring reproductive status based on the measurement of urinary steroid conjugates were examined. Levels of urinary oestrone-3-glucuronide, oestrone-3-sulphate, oestradiol glucuronide, oestradiol sulphate and pregnanediol-3 alpha-glucuronide were determined during the ovarian cycle and pregnancy. Sequential hydrolysis showed oestradiol conjugates to be more abundant than oestrone conjugates. The levels of sulphates and glucuronides were similar in the follicular phase whereas sulphates predominated during the luteal phase and pregnancy. Although levels of oestrone-3-sulphate were two- to fourfold lower than those of oestradiol sulphate, measured after hydrolysis, the profiles throughout the cycle and pregnancy were similar. Levels of oestrone-3-sulphate, measured by direct assay, were below 1 mumol/mmol creatinine during the follicular phase, rising 3-4 days after ovulation to reach maximum values (2-8 mumol/mmol creatinine) in the mid-luteal phase. There was no consistent increase before ovulation. Levels during pregnancy rose gradually until days 70-90, after which there was no further increase (gestation length = 144 days). The pattern of pregnanediol-3 alpha-glucuronide was similar to that of oestrone-3-sulphate during the ovarian cycle but levels did not increase during pregnancy. The patterns of excretion of oestrogen and progesterone metabolites were similar to the pattern of the circulating hormones during the ovarian cycle. Circulating and urinary hormone patterns were similar for oestrogens throughout pregnancy but pregnanediol-3 alpha-glucuronide did not reflect progesterone secretion beyond day 70 of gestation.

Eleven unconjugated steroids were measured daily during a complete cycle in the peripheral plasma of 6 normally menstruating baboons (Papio hamadryas) by means of a radioimmunoassay procedure and the levels were compared with those found previously in 17 normally menstruating women. The patterns of progesterone and 20alpha-dihydroprogesterone were very similar to those found in women throughout the entire menstrual cycle. However, the ratio of these steroids differed markedly from that found in women. A great similarity of the follicular phase and the peri-ovulatory period profiles was observed in both species for 17-hydroxy-progesterone, pregnenolone, androstenedione, oestradiol and oestrone. The pattern of oestradiol:oestrone ratios was similar in both species. However, an elevation of the above five steroids, typical for the human luteal phase, was not found in baboons. The increase of testosterone values, seen in women at mid-cycle, was not detected in baboons. The plasma concentrations were lower in baboons than in humans for all the above steroids.

Estracyt, a compound of nitrogen-mustard linked to oestradiol phosphate, is used in the treatment of human prostatic cancer. The metabolism of this compound has been studied in different tissues of the rat both in vivo and in vitro. The phosphate group in position 17 of the oestradiol moiety is rapidly split off from the compound. An oestrone-cytostatic compound was extractable from the liver half an hour after the injection of Estracyt. In addition the in vitro results showed that only the liver was able to convert the oestradiol-cytostatic compound to an oestrone-cytostatic one. When animals were killed 24 h after a 3-day period of Estracyt treatment, the dominating metabolite in the ventral prostate was an oestronecytostatic compound, but traces of free oestrone could also be demonstrated. No such compound, however, was found in liver, diaphragm or blood at this time. It is concluded that in vivo an oestrone-cytostatic compound seems to be preferentially retained in the ventral prostate after Estracyt injection whilst the metabolic conversion of the oestradiol-cytostatic compound into an oestrone-cytostatic one possibly occurs in the liver.

Concentrations of oestradiol-17 beta and oestrone were measured in the uterine arterial and venous blood of anaesthetized sows on Days 11, 13 or 15 of the oestrous cycle or of pregnancy. Uterine arterial blood flow and the amounts of oestradiol-17 beta and oestrone in uterine flushings were determined in the same animals. In pregnant animals arterial and venous concentrations were significantly different (P less than 0.05) for oestradiol-17 beta on Days 11 and 13 and for oestrone on Day 15. Uterine content of both steroids was consistently greater in pregnant than in non-pregnant animals with a peak on Day 13. Uterine arterial blood increased from Day 11 to 13 of pregnancy then declined (P less than 0.08) by Day 15; no change in uterine blood flow occurred on the corresponding days of the oestrous cycle.

L-DOPA, within 30 min after administration, induced a highly significant decrease of plasma prolactin levels (phase 1) in a number of groups of rats, differing in age and/or endocrine status, apparently by direct inhibition of prolactin release from the pituitary. Three hours after administration of L-DOPA these low plasma prolactin concentrations in treated animals had increased (phase 2) and did not differ significantly from levels in control animals, indicating that the effect of L-DOPA on plasma prolactin levels is only of short duration. During this process some interesting phenomena were observed, especially in the animals treated with oestrone. The elimination rate of prolactin from plasma was very high (t 1/2 = 2.8 min), as indicated by decreasing concentrations of the hormone during phase 1. Pituitary prolactin content did not change during phase 1, suggesting that prolactin synthesis was also stopped. Notwithstanding the high elimination rate, plasma prolactin regained initial concentrations in phase 2, suggesting release of a substantial part of the pituitary prolactin content. The latter,however, remained constant during the whole experiment (i.e. before L-DOPA administration and during phase 1 as well as phase 2). The results suggested another working mechanism of L-DOPA in decreasing plasma prolactin levels, namely by stimulating the uptake of this hormone in the periphery. After the effect of L-DOPA had ceased, most of the prolactin from the periphery returned into the bloodstream, causing a rapid restoration of plasma prolactin levels without substantial release from the pituitary. The nature of the processes responsible for the peripheral uptake of prolactin is discussed.

To evaluate the effects of ovarian surgery on the deranged episodic gonadotrophin release of women with the polycystic ovarian disease (PCOD), we studied 11 patients with the clinical and endocrinological features of PCOD before and after laparoscopic laser coagulations of ovarian surfaces and cysts. During both occasions, blood was collected at 15-min intervals for 8 h to determine LH and FSH secretory profiles and additionally for 3 h during GnRH injections (25 micrograms twice within 2 h) to assess pituitary responsiveness. Serum testosterone, androstendione and oestrogen (oestrone, oestradiol) levels were markedly reduced (P less than 0.05 or less) after surgery. Mean LH concentrations declined (P less than 0.001), while FSH levels increased (P less than 0.01) following laser treatments. The LH pulse frequencies (by Cluster analysis) did not change after ovarian surgery, but the LH pulse amplitudes were markedly reduced (P less than 0.01). Lower (P less than 0.05 or less) LH concentrations were attained in response to GnRH challenges, and the stimulated FSH release also tended to decrease after laser treatments. Thus, ovarian surgery in PCOD women resulted in reduced serum sex steroid concentrations and in divergent effects on serum LH and FSH levels. The attenuated pituitary LH responsiveness after ovarian surgery suggests action of sex steroids primarily at the pituitary site, while the increase in FSH concentrations may be attributed to other factors selectively modulating FSH release.

Sera from sheep immunized against oestrone (Group E1), oestradiol (Group E2), androstenedione (Group A) and testosterone (Group T) were given to ewes singly or as a mixture (Group M) of all 55 types as a single intravenous injection at the time of the start of mating. The number of lambs produced, the numbers of eggs shed and the display of oestrus were recorded. The ovulation rates were 1.8 in Group E1, 2.1 in Group E2, 1.6 in Group A, 1.8 in Group T and 2.1 in Group M compared with 1.3 for the controls (P, variation among groups, less than 0.001) in the first oestrous cycle. The effect persisted in those animals not conceiving to the first mating--1.3 in Group A, 1.8 in Group E1, 1.9 in Group E2 and 2.0 in Group M compared with 1.3 for the controls; all of the ewes in Group T conceived to mating at a single oestrus. The mean number of lambs born alive per ewe treated was 1.1 for Group A, 1.3 for Group E1, 1.3 for Group E2, 1.5 for Group T, 1.5 for Group M and 1.0 for the controls. The increase in the number of lambs born was due to a higher proportion giving birth to twins (P less than 0.01); no ewe gave birth to triplets. High conception rates were recorded for all treatments.