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There are several methods of extracting urine before assay for gonadotrophin and there has been some controversy about the best procedure. In an earlier investigation (Hipkin, 1967) two different kaolin—acetone procedures were compared. Both extracts contained gonadotrophin-inhibiting material which was probably responsible for the differences in potency when large doses of extract were assayed. Since the inhibitory material can interfere with the assay for gonadotrophin the tannic acid method of Johnsen (1958) has been investigated from this aspect.
Six 4 1. pools of urine were collected from patients known to have normal pituitary function. Half of each pool was extracted by Albert's method as modified by Borth, Lunenfeld & Menzi (1961) to give extract A, and the remainder by that of Johnsen (1958) to yield extract J. The extracts were tested for gonadotrophic and inhibitory activities by methods described previously (Hipkin, 1967).
The mean weight of solid extracted by procedure
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SUMMARY
Pooled urine from various groups of human subjects was extracted by the kaolin-acetone procedure of Albert (A procedure) and that of Loraine & Brown (L-B procedure). Two to three times as much solid was obtained with the L-B procedure. When the biological activity of the extracts was compared by the mouse uterine weight assay, they had similar potencies when the assay was conducted with small doses of extract. When larger doses of extract were used the A extract was 2·25 times as active as the L-B extract. Extracts obtained by both procedures contained gonadotrophin inhibiting material, but the L-B extract was 3·5 times more active in this respect. It was concluded that this was the reason for the apparent difference in response to the extracts at the higher dosage.
Gel filtration studies indicated that NaOH elution from kaolin and precipitation with 5 vol. acetone (as used in the L-B procedure), extracted more gonadotrophin-inhibiting material. Since this material was without gonadotrophic activity, NH4OH elution with 2 vol. acetone precipitation (as used in the A procedure) is preferable for urine extraction before gonadotrophin assays.
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Boiled urinary extracts inhibit the activity of 0·4 i.u. human chorionic gonadotrophin (HCG) in the mouse uterus assay (Hipkin, 1967) and a pineal source for this urinary gonadotrophin inhibitor has been suggested (Soffer, Fogel & Rudavsky, 1965). In intact rats, boiled urinary extracts augment the activity of small doses of HCG but this effect disappears when the animals are hypophysectomized (Hipkin 1970). Two pineal substances, melatonin and 5-methoxytryptophol, inhibit gonadal function (Mclsaac, Taborsky & Farrell, 1964). These materials have therefore been tested for their effects on HCG activity to determine whether they have the same properties as the urinary factor.
Animals were obtained from colonies bred at the Statens Seruminstitut. The doses of HCG and the experimental procedures were the same as those used previously (Hipkin, 1967, 1970), but instead of boiled urinary material, daily s.c. injections (0·2 ml each) of 5-methoxytryptophol or melatonin (20 μg to mice, 200 μg to rats)
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Sub-department ofEndocrine Pathology, Alder Hey Hospital, Eaton Road, Liverpool, LI2 2AP
(Received 12 September 1977)
Injections of ovine prolactin into normal subjects and certain patients increases the urinary excretion of calcium (McCalister & Welbourn, 1962; Beck, Gonda, Hamid, Morgen, Rubinstein & McGarry, 1964) and hypercalcaemia as well as hypercalcinuria is produced after administration of prolactin to experimental animals (Mahajan, Robinson & Horrobin, 1974). The levels of calcium in the serum and urine of patients with hyperprolactinaemia have therefore been studied.
Fourteen patients (one man, 13 women) were selected on the basis of raised levels of prolactin in the plasma. Clinical details are shown in Table 1. None of the patients had primary hypothyroidism and none was taking drugs except the man (S.B.), who was on cortisone and thyroxine replacement therapy. During the study, patients had their normal diet either at home or in hospital. Plasma prolactin was estimated by radioimmunoassay
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It is well established that primary and secondary deficiency of cortisol secretion by the adrenals causes a grossly impaired diuresis during the first 4 h after an oral water load, and that the levels of blood cortisol affect the osmotic threshold for secretion of antidiuretic hormone (ADH) by the neurohypophysis (Aubrey, Nankin, Moses & Streeten, 1965). The changes in blood cortisol concentration after a water load, however, have been less thoroughly studied, though Hatfield & Shuster (1959) found a fall in plasma cortisol levels, as measured by the Porter—Silber reaction. Since a variation in cortisol concentration might itself affect diuresis after a water load, such a response would be important in the understanding of the physiology of diuresis and was, therefore, re-investigated.
The experiments were performed on six healthy adult male subjects. No food or fluids were taken from 22.00 h the previous night. At 09.15 h a sample of
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Abstract
The effects of human recombinant basic fibroblastic growth factor (bFGF) on the secretion, viability, proliferation, attachment and morphology of ten dispersed human clinically non-functional (NF) adenomas were examined in vitro. Four clinically NF adenomas secreting FSH and/or LH in vitro were unaffected by 10 nm bFGF over a 4-h period. Over 4 days 10 nm bFGF stimulated LH secretion (66% and 72%, P<0·01) from two out of seven clinically NF adenomas secreting LH, whilst FSH (three tumours) and α-subunit secretion (three tumours) were unaffected. One adenoma co-secreting LH and α-subunit and one secreting LH alone were studied over 21 days; LH secretion fell progressively, but the decline was significantly less (P<0·05) with bFGF (10 nm) treatment after 14 and 21 days in both adenomas, whilst the fall in α-subunit secretion was unaffected by bFGF treatment. A 24-h GnRH test performed at the start and end of the 21-day period in one of these tumours showed an increase in both basal and stimulated LH secretion in the bFGF-treated group over control (124%, P<0·001). There was no effect of bFGF (10 nm) on viability, S-phase proliferation, attachment or morphology of adenoma cells over a 4-day period. These results suggest that bFGF has a role in tumorous LH secretion from these adenomas, but is not mitogenic (at least over 4 days) and is without effect on other parameters of in vitro differentiated function.
Journal of Endocrinology (1995) 144, 173–178
