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S. A. BARNETT and ELIZABETH M. COLEMAN

SUMMARY

Breeding stocks of mice of strain A2G have been studied at environmental temperatures of −3° and 21° C. The mean age of opening of the vagina was 33 days at −3° C, 26 days at 21° C. The mean body weight at opening was 13 g in both temperatures. The vaginal smear of typical oestrus appeared at a mean age of 61 days at −3° C, 38 days at 21° C; it was preceded by variable numbers of anomalous smears containing squamous cells. The mean length of the oestrous cycle was 8·5 days at −3° C, 4·8 days at 21° C. Females transferred from 21° to −3° C had longer cycles at first, but tended to return to normal after some months. The interval between parturitions had two modes, at about 3 and 6 weeks respectively: most intervals were around 6 weeks at −3° C, 3 weeks at 21° C. There was evidence of a negative correlation between the numbers weaned in successive parturitions, when the interval between parturitions was near the minimum. The slowing of the reproductive cycle at −3° C may be attributed to the prior demands of catabolism; but this does not account for the recovery of the mice transferred from warm to cold.

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M. E. Coleman and T. D. Etherton

ABSTRACT

The insulin-like growth factor-binding proteins (IGFBPs) in sera of growing pigs were partially characterized with respect to their size, immunological relationships to other known IGFBPs and their regulation by porcine (p) GH. Castrated male pigs (14–16 weeks of age) were treated with either vehicle or pGH (up to 100 μg/kg body weight per day) by daily i.m. injection for 7 days. Blood samples were collected by jugular venepuncture at the time of injection. Five IGFBPs of 43, 40, 34, 30 and 26 kDa were identified on ligand blots of porcine sera. A 30 kDa IGFBP, in addition to the 43 and 40 kDa IGFBPs, was immunoprecipitated by antiserum to pIGFBP-3 and found to contain N-linked carbohydrate suggesting that it is a fragment of pIGFBP-3 as has been noted for a 29 kDa N-glycosylated IGFBP in rat sera. The 34 kDa IGFBP in pig sera was precipitated by antisera to rat IGFBP-2 and contained no N-linked carbohydrate. Administration of pGH to normal growing pigs not only increased pIGFBP-3 levels but elicited a dose-dependent suppression of levels of the 34 kDa IGFBP as well. In summary, the M r pattern of IGFBPs in the sera of growing pigs is similar to that observed in fetal and maternal pig sera and in other species. Furthermore, we report that administration of pGH to normal pigs suppresses the expression of an IGFBP-2-like IGFBP in pig sera while increasing expression of pIGFBP-3.

Journal of Endocrinology (1991) 128, 175–180

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A Stoica, M Saceda, VL Doraiswamy, C Coleman and MB Martin

The role of epidermal growth factor (EGF) in the regulation of estrogen receptor-alpha (ER-alpha) gene expression in the human breast cancer cell line MCF-7 was investigated. Treatment of cells with 0.4 ng/ml EGF resulted in an approximately 60% decrease in ER-alpha protein concentration by 6 h and the amount of receptor remained suppressed for 24 h. Ligand binding assays demonstrated that the decrease in ER-alpha protein corresponded to a similar decrease (approximately 50%) in estradiol binding sites. Although EGF treatment resulted in a decrease in the number of binding sites, it had no effect on the binding affinity of ER-alpha. The dissociation constant of the estradiol-ER-alpha complex in the presence or absence of EGF was the same (K(d)=2.3x10(-)(10) M in control cells versus K(d)=1.98x10(-)(10) M in EGF-treated cells). The decrease in ER-alpha protein concentration paralleled a decrease in the steady-state amount of ER-alpha mRNA. By 9 h there was an approximately 60% decrease in ER-alpha mRNA. The amount of ER-alpha mRNA remained suppressed for 48 h. Transcription run-on experiments demonstrated that there was a decrease of approximately 70% in ER-alpha gene transcription upon EGF treatment, suggesting that the mechanism by which EGF regulates ER-alpha gene expression is transcriptional. In addition to regulating the amount of ER-alpha, EGF affected the activity of the receptor. At high concentrations, EGF induced progesterone receptor. Estradiol and high concentrations of EGF had an additive effect on progesterone receptor. In contrast to high concentrations, low concentrations of EGF had no effect on progesterone receptor and blocked estradiol induction. The effects of EGF on ER-alpha expression were inhibited by tyrophostins and wortmannin, suggesting that the effects of the growth factor are mediated by the EGF receptor and protein kinase B. When the cells were placed in serum-free medium and then treated with EGF, there was no effect on ER-alpha protein concentration or activity. However, increasing concentrations of serum restored the effects of EGF on ER-alpha, suggesting that an additional serum factor was required for the EGF-mediated effect on the decrease in ER-alpha protein concentration.

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P. V. BERTRAND, J. R. COLEMAN, A. C. CROOKE, M. C. MACNAUGHTON and I. H. MILLS

SUMMARY

Eight women with secondary amenorrhoea were treated, four at each of two centres. They were each given four preparations of gonadotrophin having different ratios of follicle-stimulating hormone (FSH) to luteinizing hormone (LH) and the order of treatment with the preparations was determined at each centre by a latin square design. At the completion of these treatments each patient received a fifth preparation of gonadotrophin. The dosage of FSH was determined for each woman by a previous sensitivity test and remained constant for each of her 5 months of treatment. Each gonadotrophin preparation was given in approximately equal daily injections on days 1, 2 and 3 of each course of treatment and was invariably followed by a single injection of 12000 i.u. human chorionic gonadotrophin on the 10th day. Treatment was assessed by the measurement of the excretion of oestriol and pregnanediol, by vaginal cytology, by the degree of ferning of cervical mucus and by the basal body temperature (BBT). The administration of gonadotrophin with a ratio of FSH:LH < 1·0 produced a greater excretion of oestriol than gonadotrophin with a ratio of FSH:LH > 4·0. There was no significant loss of sensitivity to treatment with time. The karyopyknotic index was correlated with the excretion of oestriol but neither was of use in predicting the other because of the wide scatter. There was no correlation between the degree of ferning and the excretion of oestriol. Assessments of ovulation by the pregnanediol response and by the BBT were in good agreement. These results are discussed.

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JA Keelan, KW Marvin, TA Sato, LM McCowan, M Coleman, LW Evans, NP Groome and MD Mitchell

To investigate labour-associated changes in production of activin and related hormones by gestational tissues we prepared extracts from amnion, choriodecidual and placental tissues delivered at term before labour (TNL; n=15), at term after spontaneous labour (TSL; n=15) or preterm (PTD; n=31) and measured concentrations of inhibin A, activin A and follistatin by ELISA. Activin concentrations in placental tissues were significantly (Mann-Whitney U-test; P<0.05) elevated with term labour (pg/mg protein, median; 1313 vs 2591), but in the PTD tissues concentrations were lower than those delivered spontaneously at term (3650 vs 2649). Inhibin concentrations also increased with term labour in the placenta (480 vs 686), but paradoxically decreased in amnion (188 vs 64) and choriodecidua (657 vs 358). Little or no significant changes in follistatin concentrations were observed. Concentrations of all three proteins were significantly correlated between amnion and choriodecidual tissues, and were significantly correlated with each other in most tissues (Spearman's ranked correlation; P<0.05). The activin:inhibin ratio in term amnion and choriodecidual tissues was increased 2 to 3-fold (P<0.0005 by Mann-Whitney U-test) after term labour, with similar trends also observed in the activin:follistatin ratio in placental tissue. These data suggest that a modest increase in placental activin and inhibin production may occur with labour at term. In addition, an increase in activin bioactivity may occur with labour, potentiating any paracrine effects of activin during parturition. The data, however, do not support an association between increased intrauterine activin biosynthesis and preterm delivery.