V.H.T. James

Aromatase activity in normal breast and breast tumor tissues: In vivo and in vitro studies

Infusions of isotopically labeled [3H] androstenedione with measurement of [3H] estrone in normal breast and breast tumor tissue have been carried out in an attempt to determine the contribution that aromatization makes to the estrogen content of breast tissues. After infusion of [3H] androstenedione for 12h there was significant uptake of this steroid by normal breast and breast tumors. [3H] Estrone was detected in all samples of normal breast tissue examined so far but not in all tumors. Aromatase activity when measured in vitro was found to be higher in breast tumors than in fat next to the tumor or normal breast fat. Studies in which we have examined the effect of epidermal growth factor on aromatase activity in cultured breast adipose tissue suggests that the response may be influenced by a subjects menopausal status. Results from these preliminary studies suggest that the aromatization of androgens may make a significant contribution towards the estrogen content of some breast tumors and that growth factors may also be involved in regulating aromatase activity.

THE RELATIONSHIP BETWEEN 17β‐HYDROXYSTEROID DEHYDROGENASE ACTIVITY AND OESTROGEN CONCENTRATIONS IN HUMAN BREAST TUMOURS AND IN NORMAL BREAST TISSUE

The activity of 17β‐hydroxysteroid dehydrogenase (17βHSD) was measured in human breast tumours and in normal breast tissue from premenopausal, perimenopausal and postmenopausal women. Enzyme activity was higher in tumour tissue than in normal tissue from the same breast and under the conditions of the assay the oxidation of oestradiol was higher than the reduction of oestrone. The physiological status of the women in the study did not relate to the activity of the enzyme in either normal or tumour tissue although fibroadenomas had less activity than adenocarcinomas.

Diet-induced changes in sex hormone binding globulin and free testosterone in women with normal or polycystic ovaries: correlation with serum insulin and insulin-like growth factor-I.

The purpose of this study was to investigate the effect of calorie restriction on serum concentrations of sex hormone binding globulin (SHBG) in women with normal or polycystic ovaries (PCO) and to examine the possible role of insulin and insulin‐like growth factor‐I (IGF‐I) in mediating changes in SHBG levels. Six normal subjects with mean (SD) body mass index (BMI) 25.5 (2.2) and five subjects with PCO (BMI 36.1 (3.7)) were studied before and after 2 or (PCO only) 4 weeks of a very low calorie diet (330 kcal/day; Cambridge Diet). In both normal women and patients with PCO there was a twofold increase in SHBG concentrations after 2 weeks and this was sustained in the PCO subjects for a further 2 weeks. The rise in SHBG was accompanied by a fall in free testosterone concentrations. There were parallel changes in serum insulin and IGF‐I concentrations which decreased during the diet and there were significant negative correlations of SHBG with insulin in both normal subjects (r =−0.62) and women with PCO (r =−0.60). In addition, serum concentrations of an insulin‐dependent small molecular weight (34 kDa) binding protein for IGF‐I (IGF‐BPI) increased significantly during dieting in both groups and were negatively correlated with serum insulin (controls, r =−0.56; PCO, r =−0.68) and positively correlated with serum SHBG levels (controls, r = 0.69; PCO, r = 0.63).

Control of aromatase activity in breast cancer cells: The role of cytokines and growth factors

The aromatase complex has a key role in regulating oestrogen formation in normal and malignant breast tissues. Using dexamethasone-treated fibroblasts, derived from breast tumours, breast tumour cytosol and breast tumour-derived conditioned medium (CM) markedly stimulate aromatase activity. The cytokine, interleukin-6 (IL-6) has been identified as a factor present in CM which is capable of stimulating aromatase activity. To examine whether IL-6 may have a role in vivo in regulating breast tissue aromatase activity, IL-6 production and aromatase activity in breast tumour and adipose tissue from breast quadrants were examined. In 5/6 breasts examined so far, aromatase activity was highest in adipose tissue in the breast quadrant containing the tumour or on which the tumour impinged. There was a significant correlation (P < 0.05, Kendalls rank correlation) between IL-6 production and aromatase activity in these breast tissues. It is concluded that IL-6 may have an important role in regulating aromatase activity in breast tissues.

The plasma concentration of oestrone sulphate in postmenopausal women: Lack of diurnal variation, effect of ovariectomy, age and weight

The plasma concentration of oestrone sulphate has been determined in 47 normal postmanopausal women. Oestrone sulphate was measured by radioimmunoassay of the unconjugated oestrone released following enzymatic hydrolysis of the plasma sample. Procedural losses were corrected by the addition of tritiated oestrone sulphate as an internal recovery standard. The mean oestrone sulphate concentration was 295 ± 174 pg/ml with a range from 89–917 pg/ml. In four subjects studied we were unable to demonstrate any diurnal variation in the oestrone sulphate concentrations. We did not find any fall with age and present evidence that the ovary in the postmenopausal woman does not contribute to the circulating level of oestrone sulphate. The most significant observation was the correlation between the plasma oestrone sulphate concentration and the subjects weight (r = 0.59, P = < 0.001, n = 35). or the degree of obesity.

SALIVARY TESTOSTERONE: RELATIONSHIP TO UNBOUND PLASMA TESTOSTERONE IN NORMAL AND HYPERANDROGENIC WOMEN

A sensitive radioimmunoassay (RIA) was used to measure salivary testosterone levels in normal women, in patients with polycystic ovaries (PCO), and in women with hirsutism. There was a highly significant correlation (r= 0·79, P<0·001) between the concentration of testosterone in saliva [12·3 ± 7·8 (SD) pg/ml] and the concentration of unbound testosterone in plasma (5·2 ± 3·1 pg/ml) in matched samples collected from 56 women including normals, patients with clinical signs of hyperandrogenism, and women treated with a combination of cyproterone acetate (CA) and ethinyl oestradiol (EE). The unbound plasma testosterone was measured in the dialysate directly using a sensitive RIA. Salivary and plasma testosterone levels in patients with PCO (20·6 ± 8·5 and 626 ± 187 pg/ml respectively, n= 14) and in those with hirsutism (13·9 ± 5·6 and 421 ± 170 pg/ml, n= 30) were significantly higher (P<0·001) than levels in normal women (7·7 ± 2·6 and 196 ± 68 pg/ml, n= 36). Treatment for 3 months with CA and EE resulted in a decrease (mean 68%) in salivary testosterone levels in all patients studied (n= 15), but the suppression of plasma testosterone (mean 34%) was not observed in all cases. It is concluded that measurement of salivary testosterone gives a useful indication of levels of biologically available androgen in hyperandrogenic women, before and during CA/EE therapy.

Aromatization of steroids in peripheral tissues

The action of glucocorticoids on aromatase activity in human adipose tissue has been investigated. Oestrogen production was increased in tissue maintained in the presence of dexamethasone and cortisol. Dexamethasone was effective at concentrations as low as 1 nmol/l. In contrast, over the 40 h incubation period, cortisol was only effective at concentrations in excess of 500 nmol/l, although after conditions of prolonged incubation, induction of aromatase activity was measurable using cortisol concentrations as low as 28 nmol/l. There were apparent variations in both basal levels of aromatic activity and in response to glucocorticoids in adipose tissue taken from various sites within the body.

RELATION BETWEEN PLASMA ŒSTRONE AND ŒSTRADIOL AND CLIMACTERIC SYMPTOMS

Plasma levels of oestrone and oestradiol-17beta were determined at 20 or 30 minute intervals for up to 24 hours in 26 postmenopausal or ovariectomised women of similar age, weight, and number of years since menopause or operation. Results in women with both superficial dyspareunia and flushes were compared with those in women with flushes only, and with those in symptomless women. Women with superficial dyspareunia had significantly lower mean concentrations of plasma-oestradiol, but not of oestrone, than symptomless women. Flushes were not related to plasma-oestrogen. The implications of these findings in relation to the optimum dose of oestrogen for treating climacteric symptoms are discussed.

Determination of deoxycorticosterone in plasma; Double isotope and immunoassay methods

Abstract Double isotope and radioimmunoassay (RIA) methods have been developed which are capable of determining deoxycorticosterone concentrations in human peripheral plasma. The levels found in control subjects were similar by both techniques. Using RIA, the mean level was 5 ng/100 ml plasma, with a range of 1 to 12 ng/100 ml.

The conversion of androstenedione to oestrone and production of oestrone in women with endometrial cancer

Abstract The conversion of androstenedione to oestrone and the production rate of oestrone has been measured in a group of women with endometrial cancer. In four postmenopausal subjects, conversion of androstenedione to oestrone ([ ρ ] BU AE 1 = 2.1 ± 0.7%, mean ± S.D.) and the production rate of oestrone (26.8 ± 5.7 μ g/24 h) were not excessive and similar to that reported for normal postmenopausal women. No significant change in either the extent of the conversion of androstenedione to oestrone or the production rate of oestrone was found when the postmenopausal women were re-investigated 2–3 months after ovariectomy and hysterectomy. For one premenopausal woman with endometrial cancer, conversion of androstenedione to oestrone ([ ρ ] BU AE 1 = 1.5%) and production of oestrone (63.4 μg/24h) were within the ranges reported for normal premenopausal women. A significant negative correlation ( r = −0.76, P