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ABSTRACT
Samples of human and ruminant mammary secretions stimulated the proliferation of rat intestinal epithelial (RIE-1) cells in culture. The stimulation was dose-dependent, and samples taken prepartum had greater potency than those taken after parturition. When various hormones and growth factors known to be present in milk were tested, only epidermal growth factor (EGF), insulin and insulin-like growth factor I (IGF-I) stimulated the proliferation of RIE-1 cells. IGF-I was effective at lower concentrations than insulin, and the maximal stimulation induced by each of these two polypeptides was greater than that induced by EGF. The maximal stimulation induced by samples of mammary secretions was similar to that induced by insulin or IGF-I.
J. Endocr. (1987) 113, 285–290
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Plasma was obtained from adult Wistar rats maintained under 14 h of light and 10 h of darkness and shown by daily vaginal smears to have had two or more cycles of 4 days' duration. Oestradiol-17β concentrations were measured by a micro-modification (Exley, 1969) of the competitive protein binding technique of Corker & Exley (1970). In more than 75% of cases plasma from two rats was pooled (5–7 ml) and duplicate determinations carried out. At concentrations above 10 pg/ml the average variation between duplicates was ±15% of the mean. Below this level, although the mean detectable amount was 6 pg (corresponding to a concentration of about 2 pg/ml) precision was such (average agreement between duplicates ±35%) that values in this range must be regarded as estimates only. Plasma luteinizing hormone (LH) in samples from individual rats was determined by a modification (Naftolin & Corker, 1970) of the radioimmunoassay method of
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ABSTRACT
In prepubertal rats, removal of one testis (hemicastration) results in increased FSH secretion and a compensatory hypertrophy of the remaining testis. To determine whether these two events are related, testis weight was measured after inhibin-rich porcine follicular fluid (FF) was administered to hemicastrated rats twice daily for 14 days to block the compensatory rise in FSH. After hemicastration, serum immunoreactive FSH concentration approximately doubled and testis weight was increased by ∼60%. A slight but non-significant increase in serum bioactive FSH was also observed. Treatment of hemicastrated rats with FF completely prevented the increase in serum FSH and compensatory growth of the remaining testis, whereas concomitant treatment of these animals with ovine FSH reversed the inhibitory effects of FF. Serum inhibin concentrations were determined using two radioimmunoassay (RIA) systems; one assay utilized an antiserum generated against intact bovine inhibin and the other was directed against an α-inhibin fragment. Both assays showed a decline in inhibin levels following hemicastration. In addition, increases in serum inhibin concentrations were observed following FF administration in both RIAs; however, the relative increase in inhibin over levels in hemicastrated rats was greater using the intact inhibin assay.
In summary, these data suggest that the increase in FSH concentrations after hemicastration is related to a reduction in inhibin levels, and that this FSH rise is a primary signal for initiating compensatory testicular hypertrophy in prepubertal rats. Furthermore, exogenous FSH administration overcame the inhibitory effects of FF on the testes, suggesting that inhibin may not act directly at the gonadal level.
Journal of Endocrinology (1991) 130, 207–212
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SUMMARY
A granulosa cell multilocular cystadenoma was removed from a 7-yr.-old girl who showed breast growth and had a history of almost continuous vaginal bleeding for 9 days. Tissue, which microscopically was shown to consist almost exclusively of granulosa cells, was incubated with [4-14C]-DHA and [7-3H]pregnenolone in Krebs-Ringer bicarbonate solution. The ability of this tissue to synthesize oestrogens from these precursors was investigated. These results and those of pre- and post-operative urinary steroid estimations are discussed in relation to current theories of steroid biosynthesis.
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ABSTRACT
The concentration of epidermal growth factor (EGF) in human mammary secretions was about 50 nmol/l for several weeks prepartum. It then fell to about 13 nmol/l within 4 days after parturition, in parallel with the decrease in protein concentration which is associated with the onset of lactation. In contrast, the concentration of EGF in urine samples from the same donors remained constant throughout this period. All the immunoreactive EGF in mammary secretions competed at the EGF receptors on Swiss mouse 3T3 fibroblasts. The stimulation of these cells by samples of mammary secretions was, however, much greater than that induced by EGF alone, indicating the presence of other factors which synergize with EGF. Gel filtration of mammary secretions revealed two major peaks of mitogenic activity, corresponding to EGF and a factor of higher molecular weight. The latter could synergize with added EGF, insulin or bombesin, and thus falls into a different functional class from any of these factors.
J. Endocr. (1987) 112, 151–159
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ABSTRACT
125I-Labelled mouse epidermal growth factor (125I-EGF) was transferred intact and undegraded from circulating blood into milk in conscious lactating goats. Greater than 90% of the total radioactivity present in milk from the infused gland was in the aqueous phase and more than 72% was acidprecipitable. This radiolabelled material co-eluted with authentic EGF through gel filtration and was immunoprecipitable by a specific rabbit anti-mouse EGF immunoglobulin. Mammary uptake of125I-EGF infused into mammary arterial blood (close-arterial infusion) for 1 h varied from 20 to 83% at different stages of the reproductive cycle. Only 0·5–2·9% of the infused 125I-EGF was transferred into milk during the first 3 h after the start of the infusion, which represents 0·7–6·3% of mammary uptake of EGF.
The kinetics of transfer of 125I-EGF were followed in two lactating goats. Radioactivity reached peak levels in milk about 120 min after the start of a 1 h close-arterial infusion into the mammary gland, with an initial lag of about 30 min when little transfer occurred. Transfer was slower in two non-lactating goats with maximal levels of activity in milk being reached after about 180 min. The results are consistent with a transcellular transfer, whereby the factor is bound to receptors on the baso-lateral membrane, internalized by epithelial cells and subsequently secreted across the apical membrane into the alveolar lumen. The low level of degraded labelled EGF in milk (and mammary vein blood) suggests a modification of the normal pathway of EGF degradation such that the delivery of internalized factor to lysosomes is avoided.
J. Endocr. (1986) 109, 325–332
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ABSTRACT
The polypeptide mitogen, epidermal growth factor (EGF), was originally isolated from mouse submaxillary gland (SMG). In mice, these glands are sexually dimorphic; the SMG of male animals typically contains up to 400 pmol EGF/mg protein whereas EGF concentrations in the SMG of female mice are only 5–20 pmol/mg protein. When mice were castrated at 8 weeks of age, EGF mRNA levels in the SMG fell rapidly to the low levels observed in control female animals. Although the concentration of EGF in the SMG, measured by radioimmunoassay, also fell to very low levels (13·8 ± 0·7 (s.e.m.) pmol/mg protein) in the castrated mice, the rate of decrease was considerably slower than that of the mRNA. In contrast to the loss of EGF and EGF mRNA from mice following castration, ovariectomy led to a rise in EGF mRNA levels in SMG, with a maximal increase (approximately 100-fold) 4–6 weeks after ovariectomy. Concentrations of EGF in the SMG also rose markedly in the ovariectomized animals, from a control value of 5·4 ± 0·8 to 34·7 ± 7·9 pmol/mg protein at 6 weeks. In mice, the kidney displays the second highest level of EGF gene expression. However, in contrast to the effects on SMG, kidney EGF mRNA was not affected by either castration or ovariectomy. These results provide further evidence for the tissue-specific control of EGF gene expression in response to steroid hormones.
Journal of Endocrinology (1989) 121, 501–506
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The rise in plasma corticosteroid concentration after the administration of corticotrophin (ACTH) is an index of the functional reserve of the adrenal cortex (Eik-Nes, Sandberg, Nelson, Tyler, & Samuels, 1954). A synthetic polypeptide β1–24 (Synacthen, Ciba) containing the amino acids 1–24 of the 39 found in pituitary ACTH (Kappeler & Schwyzer, 1961) is as active in stimulating the adrenal as corticotrophin (Landon, James, Cryer, Wynn & Frankland, 1964; Ohlsen & Hökfelt, 1965; Wood, Frankland, James & Landon, 1965). Wood et al. (1965) have shown that the rise in plasma corticosteroids (Mattingly, 1962) 30 min. after the i.m. administration of 250 μg. Synacthen can be used as a measure of adrenocortical reserve. We report our experience with this test carried out between 8 a.m. and 10 a.m. in two groups of hospital patients.
Group I comprised 21 women and nine men, aged from 20 to 73 yr. (mean age 46
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ABSTRACT
A homologous radioimmunoassay for the measurement of epidermal growth factor (EGF) levels in pig tissues and body fluids has been developed using an antiserum to recombinant porcine EGF. The assay is highly specific, showing no cross-reactivity with a variety of other polypeptides including the structurally related protein, transforming growth factor-α. Furthermore, <1% cross-reactivity was observed with mouse EGF emphasizing the necessity for homologous assays for EGF measurement. Immunoreactive EGF was present in extracts of pig kidney and pancreas (3·44 ±0·43 and 0·76 ±0·13 (s.e.m.) pmol/g wet weight respectively), but was not detected in extracts of submaxillary gland or liver. Although immunoreactive EGF was not detectable in uterine, allantoic or ovarian follicular fluids, colostrum contained EGF at biologically active concentrations (0·84±0·15 nmol/l). Immunoreactive EGF was also present in pig urine, with similar concentrations in samples from male or female animals. In addition, pig urine inhibited the binding of 125I-labelled EGF to 3T3 fibroblasts and stimulated DNA synthesis in quiescent monolayers of these cells, indicating that the immunoreactive material in urine is biologically active. Quantitative comparisons of the data presented here with that published previously indicate considerable species variation in the EGF levels of various tissues and body fluids.
Journal of Endocrinology (1992) 135, 77–83
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ABSTRACT
Seven Merino–Border Leicester cross–bred ewes were immunized with a purified fusion protein, produced by recombinant DNA methods, of the a subunit of bovine inhibin. Four animals were immunized with the fusion protein alone and three with a conjugate made by coupling the fusion protein to keyhole limpet haemocyanin (KLH) using glutaraldehyde. Each animal received four injections of the fusion protein over 93 days. The animals were synchronized using progestagen sponges and subjected to laparoscopy for the determination of ovulation rates in two consecutive cycles (days 115 and 135). The immunized animals had overall mean ovulation rates for each cycle of 3.4 and 3.4 which was significantly (P < 0.001) above the rates of 1.1 and 1.4 determined for the controls, which had either received no treatment (n=5) or had been immunized with 300 μg KLH (n=4). Analysis of antisera taken on day 115 showed significant fusion protein antibodies and iodinated inhibin–binding capacity in the test but not control groups. Furthermore, antisera to the fusion protein in four out of seven ewes neutralized the inhibin bioactivity of ovine follicular fluid in an in–vitro bioassay. These data demonstrate that neutralization of inhibin can be effected by immunization with bovine inhibin a subunit and that such immunization results in increased ovulation rates as predicted from the biological role of inhibin as a suppressor of FSH.
