Theo J. Benraad

The effect of oral contraceptives on plasma free and salivary cortisol and cortisone

The effect of a low estrogen oral contraceptive (OC) on glucocorticoid levels in plasma and saliva as well as glucocorticoid binding was studied in 23 healthy women using 30 micrograms ethinyl estradiol (EE2) + 150 micrograms desogestrel (Marvelon) (II). Fifteen healthy females with normal menses served as controls (I). Blood and salivary samples were taken between 9.00 and 9.30 a.m. on the 18th day of menstrual or pill cycle. Assay accuracy had been optimised by applying extraction and chromatographic purification before radioimmunoassay (RIA) of cortisol and cortisone in both plasma and salivary samples. Free steroid assays were performed by applying the same procedure to equilibrium dialysates obtained after dialysing plasma against an equal volume of buffer, instead of measuring tracer distribution. Corticosteroid Binding Globulin (CBG) was measured by a commercial RIA. As expected, CBG as well as plasma total cortisol were elevated in the pill group. Interestingly both plasma free and salivary cortisol were higher than in controls (free cortisol I: 18.0 +/- 7.95 nmol/l; II: 32.3 +/- 9.03 nmol/l; salivary cortisol I: 9.2 +/- 3.88 nmol/l; II: 18.8 +/- 6.92 nmol/l. Salivary cortisol closely parallelled plasma free cortisol both within and between the groups, though at a much lower level (about 50%). Free cortisone was slightly lower in the pill group (I: 10.8 +/- 2.55 nmol/l; II 8.5 +/- 1.86 nmol/l) whereas salivary cortisone was 2.3 (I) and 4.4 (II) times higher than plasma free cortisone and tended to follow the plasma free and salivary cortisol pattern, both within and between the study groups.(ABSTRACT TRUNCATED AT 250 WORDS)

A sensitive radioimmunoassay of atrial natriuretic peptide in human plasma, using a tracer with an immobilized glycouril agent

A highly specific and sensitive radioimmunoassay (RIA) for alpha-human atrial natriuretic peptide (hANP[1-28]) in plasma was developed. The assay used a [125I]monoiodotyrosyl-hANP[1-28] tracer, prepared with an immobilized glycouril agent (Protag) and purified by high pressure liquid chromatography (HPLC), and a highly specific antiserum raised against hANP[1-28], coupled to keyhole limpet haemocyanin, in sheep. Plasma was extracted using C-18 Seppak cartridges. A good parallelism was found after dilution prior to extraction of plasma of patients with congestive heart failure (CHF) or of plasma of healthy subjects. Recovery of hANP[1-28] added to plasma was 96%. The limit of detection was 0.8 pg/tube, intra- and inter-assay variation were 9 and 12%, respectively. Mean plasma ANP values in 25 normal persons with a normal salt intake was 26.0 +/- 15.5 (+/- SD) pg/ml. Plasma levels of 18 subjects (7 normals, 11 CHF) were measured using four different antisera after the extraction step. High correlations were found between the values obtained with these four antisera.

Transition from Pituitary-Dependent to Adrenal-Dependent Cushing's Syndrome

PITUITARY-DEPENDENT bilateral adrenocortical hyperplasia (Cushings disease) is present in 70 to 80 percent of all patients with Cushings syndrome. In 20 to 40 percent of patients with Cushings d...

Atrial natriuretic peptide after myocardial infarction

Plasma concentrations of atrial natriuretic peptide (ANP) after acute myocardial infarction were measured at fixed times during 48 hours in 38 patients admitted to the hospital within 4.4 hours after the onset of symptoms. Three hours after admission, the mean concentration of ANP was significantly lower than that at the time of admission. Thereafter it rose steadily until 15 hours after admission. ANP concentrations measured in each patient at the time of admission and the individual mean ANP concentrations during the first 48 hours after admission correlated weakly but significantly with the size of the infarct and the left ventricular function. Neither the site of the infarct, the occurrence of reperfusion, nor the number of coronary vessels affected influenced the ANP concentration. In 24 patients in whom cardiac catheterization was performed, no relationship between ANP concentrations and left ventricular pressures was observed. Determination of ANP concentrations seems to be of little value in assessing cardiac function after acute myocardial infarction.

Progesterone receptor activity and relapse-free survival in patients with primary breast cancer: the role of adjuvant chemotherapy

The prognostic significance of progesterone receptor activity (PgR) with regard to the estimated relapse-free survival (RFS) was studied in 350 one-center patients with primary breast cancer. All receptor assays were performed in one laboratory; PgR levels >10 fmol/mg protein were considered positive. Univariate as well as multivariate statistical analyses were used to examine the prognostic significance of several variables. Eighty-nine of the 350 patients received adjuvant CMF chemotherapy (cyclophosphamide, methotrexate, and 5-fluorouracil). The median observation period was 69 months (range 12–125 months).In the group of 261 patients who did not receive adjuvant CMF, the PgR-status lacked prognostic significance; only the lymph node status significantly affected the RFS (p<0.00001). In contrast, in the CMF-treated group of patients, the PgR-status was the most powerful predictor of recurrence (p<0.0001). The menopausal and the lymph-node status increased the predictive value of PgR (p<0.001). Premenopausal CMF-treated patients with PgR+ tumors had a significantly longer RFS than those with PgR− tumors (p<0.02). The present data urge the need for a reappraisal of the prognostic significance of PgR and of the mechanism of action of adjuvant chemotherapy in primary breast cancer.

Atrial Natriuretic Peptide

SummaryThe atrial natriuretic peptide (ANP) is part of a new family of cardiac hormones regulating water and salt homeostasis. Besides acting as a blood pressure-lowering agent, it also exerts potent natriuretic and diuretic effects. ANP can be considered an endogenous antagonist of the renin-angiotensin-aldosterone system and the antidiuretic hormone. One of the roles of ANP is to protect the body against fluid overload: it decreases intravascular fluid volume, which in turn diminishes cardiac secretion of ANP. The pharmacokinetic parameters of ANP reported in the literature vary widely. In general, ANP rapidly disappears from plasma with a high total body clearance. This is in agreement with the short-lived effects of the hormone. The actions of ANP have led to efforts to use this peptide hormone in the treatment of various cardiovascular dis-orders such as hypertension and congestive heart failure. Intravenous ANP administration indeed resulted in beneficial effects in these disorders. However, the peptide nature of ANP and its rapid elimination from the circulation limit its suitability as a drug. More promising is the development of long-acting ANP analogues and inhibitors of ANP degradation. Proper understanding of ANP pharmacokinetics is essential for the clinical use of these pharmacological agents.

Concordance and discordance of estrogen and progesterone receptor content in sequential biopsies of patients with advanced breast cancer: Relation to survival

In 75 patients with advanced breast cancer, sequential biopsies were analyzed for estrogen receptor (ER). In 50 of these patients progesterone receptor (PgR) was also measured. All pairs of biopsies met the following criteria: (i) interval between the two biopsies: at least 6 weeks; (ii) biopsies performed at least 6 weeks after stopping endocrine therapy; and (iii) concordant histology. Discordance in ER was found in 14 of 75 patients (18.7%); PgR was discordant in 14 of 50 patients (28.0%). No significant differences were found between concordant and discordant groups of patients in age at first diagnosis, menopausal state, diameter of the primary tumor, time interval between the two biopsies and intervening therapy. The initial ER level in patients whose ER changed from positive to negative was significantly lower than in patients whose ER remained positive. PgR levels exhibited a rise only when ER rose at the same time. Sequential assays have increased the prognostic significance of ER and as a consequence the estimated survival time for patients whose tumors were ER-negative in both biopsies was significantly shorter than for patients whose tumors were ER-negative in only one of the two biopsies. We found no prognostic significance for PgR in either single measurements or repeated biopsies.

Intrapatient comparison of treatment with chlorthalidone, spironolactone and propranolol in normoreninemic essential hypertension

The effects of chlorthalidone, spironolactone and propranolol in reducing blood pressure were compared in the same 11 normoreninemic hypertensive patients. All three drugs decreased the blood pressure significantly and no agent had a superior blood pressure-lowering effect. The blood pressure did not normalize. The data suggest that no one variable--volume factors, relative hyperactivity of the renin-aldosterone system or beta-adrenergic hyperactivity--is the prime mover in normoreninemic hypertension. Long-term treatment with chlorthalidone resulted in slight hyperreninism (26.3 +/- 4.9 ng-ml-1-3 hours-1) (mean +/- standard error) with concomitant changes in plasma aldosterone (23.0 +/- 3.2 ng-100 ml-1). The body weight decreased significantly (--1.8 kg, P less than 0.005). Plasma potassium concentrations were low (3.2 +/- 0.1 mEq-liter -1). Creatinine clearance was unimpaired (117 +/- 6 ml-min-1). Treatment with spironolactone resulted in more marked hyperreninism (47.0 +/- 14.3 ng-ml-1-3 hours-1) and hyperaldosteronism (61.9 +/-11.8 ng-100 ml-1). The body weight decreased significantly (--1.9 kg, P less than 0.004). Significant hyperkalemia occurred (4.4 +/- 0.1 mEq-liter-1). The glomerular filtration rate decreased significantly to 93 +/- 3 ml-min-1 (P less than 0.004). Treatment with propranolol resulted in marked suppression of the plasma renin activity (1.8 +/- 0.2 ng-ml-1-3 hours-1) and plasma aldosterone levels (8.9 +/- 1.3 ng-100 ml-1). A significant increase in body weight occurred (+2.3 kg, P less than 0.013). The plasma potassium concentration increased to a level not significantly different from the value found after treatment with spironolactone (4.2 +/- 0.1 mEq-liter-1). The creatinine clearance decreased significantly to 99 +/- 5 ml-min-1 (P less than 0.008). Hyperreninemia (by spironolactone and chlorthalidone), effective hyperaldosteronism (by chlorthalidone) and volume retention (by propranolol) are considered to represent expressions of mechanisms counteracting the depressor effects of these different pharmacologic maneuvers, leading to the maintenance of supranormal blood pressure.

The HMG-CoA reductase inhibitor simvastatin suppresses human testicular testosterone synthesis in vitro by a selective inhibitory effect on 17-ketosteroid-oxidoreductase enzyme activity

In concentrations probably exceeding those achieved in vivo, the cholesterol lowering compound simvastatin was found to suppress the synthesis of the androgens androstenediol and testosterone in vitro by human testicular homogenates. It was demonstrated that simvastatin in addition to its known inhibitory effect on HMG-CoA reductase activity, also affects the later steps of testicular steroidogenesis by selectively inhibiting the 17-ketosteroid-oxidoreductase catalyzed conversion of dehydroepiandrosterone and androstenedione to androstenediol and testosterone respectively. There was no effect of simvastatin on the Cytochrome P-450-dependent microsomal enzymes. Although in doses conventionally used in the treatment of hypercholesterolemia, simvastatin does not affect testicular steroidogenesis, at higher doses--especially when inadvertently administered during early pregnancy--adverse effects on normal testosterone biosynthesis and thereby fetal development should be considered.

Complexes between urokinase-type plasminogen activator and its receptor in blood as determined by enzyme-linked immunosorbent assay

Complexes between urokinase‐type plasminogen activator (uPA) and its receptor (uPAR) were assessed in plasma and serum from 39 breast cancer patients and from 20 healthy individuals, applying a recently developed enzyme‐linked immunosorbent assay (ELISA) for the analysis of these complexes in tumor tissue extracts. The assay is based on a combination of rabbit polyclonal anti‐uPA antibodies for catching and a mouse anti‐uPAR monoclonal antibody (MAb) for detection. The specificity of the assessment of uPA:uPAR complexes was verified by simultaneous analysis of the individual blood samples in corresponding non‐sense ELISA formats, in which either the anti‐uPA catching antibody or the anti‐uPAR detecting antibody was substituted with an irrelevant antibody. Assessment of native uPA:uPAR complexes was ascertained by demonstrating the absence of any de novo formation of uPA:uPAR complexes in plasma and serum during the sample incubation step in the ELISA, as verified by the use of a peptide antagonist for uPAR. Plasma and serum samples contained almost identical levels of uPA:uPAR complexes. The levels of uPA:uPAR complexes were found to be significantly lower in serum from breast cancer patients compared to the serum of healthy donors, while the levels of (total) uPAR in plasma from breast cancer patients were significantly higher than in plasma from the healthy controls. In addition, the free, uncomplexed uPAR levels, estimated by subtraction of uPA:uPAR complex levels from (total) uPAR levels, were significantly elevated in plasma as well as in serum from breast cancer patients compared to healthy individuals. The uPA:uPAR complex levels were highly comparable to the uPA levels analyzed in the same plasma and serum samples, indicating that most if not all of the uPA present in these samples is complexed with uPAR. Int. J. Cancer 77:236–242, 1998.© 1998 Wiley‐Liss, Inc.