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OBJECTIVE: The purpose of this study was to identify the placental expression of adrenomedullin and adrenomedullin receptor messenger ribonucleic acid and compare them between placentas from pregnancies associated with oligohydramnios as a result of uteroplacental insufficiency and placentas from normal pregnancies. STUDY DESIGN: Total ribonucleic acid was extracted from the amnion, chorion, cotyledon, umbilical vein, and umbilical artery in 5 normal placentas and 3 placentas from pregnancies complicated by oligohydramnios. A cell line known to express messenger ribonucleic acid of adrenomedullin and its receptor was used to optimize the polymerase chain reaction and served as a positive control preparation in all experiments. Semiquantitative reverse transcriptase-polymerase chain reaction results for adrenomedullin and adrenomedullin receptor were compared between tissues as densitometric ratios of adrenomedullin or adrenomedullin receptor messenger ribonucleic acid to beta(2)-microglobulin messenger ribonucleic acid. Results were analyzed with a Kruskal-Wallis 1-way analysis of variance. Immunohistochemical staining with an antibody to human adrenomedullin was used to localize adrenomedullin in all tissue types. RESULTS: Messenger ribonucleic acid sequences for adrenomedullin and adrenomedullin receptor genes were identified in all tested placental tissue components. Within the normal placentas the expressions of adrenomedullin and adrenomedullin receptor messenger ribonucleic acid sequences did not differ statistically between the tissue components. Within placentas from patients with oligohydramnios the expressions of adrenomedullin and adrenomedullin receptor messenger ribonucleic acid did not differ statistically between the tissue components. When normal placentas were compared with placentas from pregnancies complicated by oligohydramnios, however, a 5-fold increase in adrenomedullin messenger ribonucleic acid and a 3-fold increase in adrenomedullin receptor messenger ribonucleic acid were seen in placentas from patients with oligohydramnios. Adrenomedullin immunoreactivity was present in all tissues studied. CONCLUSION: The expression of messenger ribonucleic acid for both adrenomedullin and its receptor in these tissue components implies that placental tissues function in both synthesis and action of adrenomedullin. The increased adrenomedullin messenger ribonucleic acid expression in the umbilical artery and the elevated adrenomedullin receptor messenger ribonucleic acid expression in the cotyledons of placentas from patients with oligohydramnios may represent a local fetoplacental physiologic adaptive response to vascular compromise.

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BACKGROUND: The authors tested the effects of the antiestrogenic agent tamoxifen on telomerase activity and cell proliferation in MCF-7 and MDA-MB-231 breast carcinoma cell lines. MCF-7 cells belong to a known estrogen receptor positive cell line, whereas MDA-MB-231 cells, previously thought to be estrogen receptor negative, are now shown to contain estrogen receptor-beta. METHODS: Both cell lines were grown in the presence of tamoxifen 10(-6), 10(-7), 10(-8), and 10(-9) M for 10 days. Cells in separate flasks were harvested daily for determination of total cell number, protein was extracted for determination of telomerase activity, and RNA was extracted for reverse transcriptase-polymerase chain reaction analysis to measure expression levels of the telomerase components (the RNA component and the catalytic subunit) and estrogen receptors. RESULTS: Total cell counts and telomerase activity levels of both cell lines with 10(-8) M tamoxifen treatment were lower than control cells and other tamoxifen treatments. Changes in the expression of individual telomerase components correlated with telomerase activity. Estrogen receptor status did not correlate with telomerase activity. CONCLUSIONS: Tamoxifen strongly affected both cell count and telomerase activity within the 10(-8) M concentration of both cell lines. Cells were able to overcome drug inhibition at all other doses after 4 days. Telomerase activity and cell proliferation were correlated in both cell lines and depended on drug concentration. Tamoxifen showed long term effects on cell proliferation of the MCF-7 cells.

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Sex hormone binding globulin (SHBG) is a high affinity binding protein for estrogens and androgens. SHBG has been found in breast tissue and cell lines through immunostaining. The goal of this series of experiments was to determine whether mRNA for SHBG is expressed in breast cancer cell lines and tumor tissue. Reverse transcriptase-polymerase chain reaction (RT-PCR) was used to detect SHBG and beta-2 microglobulin (control for tissue extractions). Three breast cancer cell lines, ZR-75-1, MCF-7, and MDA-MB-231 and 56 breast tissue samples were collected and analysed for SHBG mRNA expression. mRNA was successfully extracted from 30 of these breast tissue samples. SHBG mRNA was detected in ZR-75-1, MCF-7 and MDA-MB-231 cells, and in 11 of the breast tissue samples. Two PCR products were routinely amplified from the breast cancer cell line RNA, one at approximately 500 bp and another at approximately 300 bp. The DNA sequence of the 300 bp PCR produce was consistent with alternate splicing of the SHBG mRNA, where exon 7 is deleted, and is accompanied by a point deletion at the beginning of exon 8. SHBG protein production from the three breast cancer cell lines was detected by immunoprecipitation using an affinity purified SHBG antibody. SHBG mRNA was found in 11 of 30 samples of breast tissue. Some samples expressed only the 500 bp or the 300 bp PCR product, whereas others expressed both PCR products. The presence of SHBG mRNA in these samples was not associated with either the presence or absence of steroid receptors. SHBG mRNA is thus expressed in breast cancer cell lines, and in some breast tissue samples.

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Total, as well as free, T4 and T3 levels were obtained over four seasons for young male infantry soldiers assigned to interior Alaska. Significant seasonal variations were found in both T3 and T4. Total T4 and T3 levels were highest in winter, while free T4 and T3 levels were highest in early spring. Correlations with melatonin levels from a concurrent study showed an association between late day (17.00) mean spot melatonin levels during the preceding summer and T3 levels in winter and spring. Differences in seasonal T4 and T3 levels between indigenous and newly arrived people in the sub-Arctic may be related not only to cold acclimation but also to light.

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Sex hormone-binding globulin (SHBG) is a glycoprotein whose production has been demonstrated to be regulated by both sex steroids, as well as by thyroid hormone and peptide hormones such as insulin. However, none of these regulatory factors would explain the marked decrease in serum SHBG seen throughout the prepubertal and pubertal time period in both boys and girls. Furthermore, current in vitro data show that both androgens and estrogens can stimulate SHBG production by the human hepatoblastoma cell line Hep G2; yet, in vivo androgens appear to suppress SHBG levels, while estrogens are associated with elevated levels. This study was undertaken to determine possible mechanisms to explain this phenomenon. Hep G2 cell cultures were incubated with insulin-like growth factor I (IGF-I), epidermal growth factor (EGF), transforming growth factor alpha (TGF-alpha), or dehydroepiandrosterone (DHEA). Significant decreases in the level of SHBG in the culture medium relative to control cultures occurred for each of the growth factors (P less than .01), whereas an increase in SHBG levels was observed in the medium of DHEA-treated cells. When cells were coincubated with IGF-I and thyroxine (T4), which alone stimulates SHBG production both in vivo and in vitro, the SHBG response to T4 was blunted. These results suggest that growth factors, as well as insulin, may be important determinants in SHBG production.

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Sex hormone-binding globulin (SHBG) production in humans has been thought to be stimulated by estrogens and thyroid hormone and inhibited by androgens. However, recent data indicate that SHBG production in vitro is stimulated by both androgens and estrogens. This study was designed to determine what other hormonal factors regulate SHBG production. Since hyperinsulinemia and hyperprolactinemia both occur in disease states in which low serum SHBG levels are found, the effects of insulin and PRL were compared to and/or studied in combination with estradiol (E2), T4, and testosterone (T) in a human hepatoma cell line (Hep G2). Hep G2 cells were grown to near confluence in medium including 10% fetal calf serum, and then 72-h experimental incubations were carried out which used only fetal calf serum-free medium. Compared to control incubations, both insulin (10(-8) mol/L) and PRL (10(-8) mol/L) decreased SHBG production from 65.0 +/- 0.6 (+/- SE) to 46.8 +/- 1.1 and 46.8 +/- 1.2 nmol/10(6) cells, respectively (P less than 0.01). Insulin also inhibited both E2 and T4-stimulated SHBG production. T stimulated SHBG production to the same degree as E2. Finally, both E2 and insulin significantly increased cell number, an important consideration when expressing the effect of a hormone on SHBG production in cultured cells. We conclude that insulin and PRL inhibit SHBG production and confirm that T4, T, and E2 stimulate SHBG production in vitro. These findings suggest that insulin and PRL may be important factors in the regulation of SHBG production in vivo.

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The following study was undertaken to determine the presence of an androgen binding protein (ABP) in the ejaculate. Seminal fluid was obtained from ten fertile subjects. Sperm counts ranged from 20 x 106/cc to 80 x 106/cc. Each sample was preincubated with tritiated dihydrotestosterone (DHT) and examined by polyacrylamide gel electrophoresis (PAGE). Second, after preincubation of the samples with tritiated DHT, the samples were mixed with Concanavalin-A bound to sepharose-D (C-S) and the supernate separated and counted. Finally, saturation analysis of the samples using dextran-coated charcoal for separation and counted. Finally, saturation analysis of the samples using dextran-coated charcoal for separation was performed and Scatchard analysis was used to determine the concentration of ABP present. PAGE demonstrated ABP in seven of ten samples studied. The bound counts of DHT were removed by C-S. The concentrations of ABP ranged from 0.132 to 0.468 nanomoles ABP/dl semen. These data indicate there is an ABP in seminal fluid similar to that from the Sertoli cell.

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Semen samples were collected from 35 men and the levels of prolactin in semen and seminal plasma were measured. There was no significant difference in prolactin concentrations between the two fluids (t = 0.333, P greater than 0.7). There was also no correlation between the prolactin concentration and the kinematic viscosity of the semen (r = 0.065, P greater than 0.7).

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