Abstract

Prostaglandin (PG) production by human fetal membranes (amnion and chorion laeve) may be important in the onset and progression of labour, cervical ripening and membrane rupture. Prostaglandin H synthase (PGHS) is a key enzyme in PG formation and has two isoforms, a constitutive form (PGHS-1) and an inducible form (PGHS-2). The present study examined the cellular distribution of the PGHS-2 enzyme and PGHS-2 mRNA in term human fetal membranes and decidua prior to and following labour, using immunohistochemistry and in situ hybridization with an ³⁵S-labelled oligonucleotide probe. The PGHS-2 protein was found to be localized in amnion epithelial cells and chorion laeve trophoblast, but was absent or at low levels in the decidual stroma in most tissues, although cells surrounding some of the blood vessels in the decidua did express PGHS-2. In situ hybridization demonstrated that PGHS-2 mRNA had a similar distribution and was localized to amnion epithelial cells, cells in the amnion-chorion mesenchyme, chorion laeve trophoblast and, occasionally, to cells surrounding blood vessels in the decidua. Of particular note was the high mRNA expression in some cells and low expression in other cells, particularly in the chorion, and the low level of PGHS-2 mRNA in decidua. There was no observable difference in the cellular localization of PGHS-2 protein or PGHS-2 mRNA in tissues obtained prior to and following labour. The studies indicate that, at term, the inducible form of PGHS, PGHS-2, is expressed at a high level in fetal tissues in a number of different cell types rather than in the maternal decidua.

Journal of Endocrinology (1996) 150, 497–503

Abstract

The human fetal membranes and decidua may be important in the onset and/or progression of human labor by providing prostaglandins for this process. Glucocorticoids have been implicated in the regulation of prostaglandin production by these tissues but to date there is no direct evidence for glucocorticoid receptors (GRs) being present in human intrauterine tissues. The purpose of the present study was to determine, using immunohistochemistry, whether the human fetal membranes and decidua contained GRs; to determine the localization of receptors to the cytoplasm or nuclei, and to examine the content and distribution of the GRs in tissues obtained during pregnancy following preterm labor (<37 weeks) and at term prior to and following term labor. Term tissues were obtained prior to labor by elective Cesarian section (n=9) or following vaginal delivery (n=9). Tissues from 14 patients who delivered preterm but with no clinical evidence of infection were also examined. Cryostat sections were thaw-mounted onto microscope slides. The immunoreactive GRs were visualized with an Elite Vectastain ABC Kit using a polyclonal antibody prepared against a synthetic peptide corresponding to amino acids 346–367 of the human GR. At term, nuclear GRs were found in amnion epithelial cells, mesenchyme and the chorion laeve. GRs were present, but were less defined, in the decidua. A similar distribution was found in the preterm tissues. However, nuclear staining in the amnion epithelial cells, mesenchymal cells, chorion and decidua was more pronounced in tissues obtained following preterm labor. This study provides direct evidence for the presence of GRs in human fetal membranes and decidua, and suggests the possible importance of multiple cell types in the action of glucocorticoids in these tissues.

Journal of Endocrinology (1996) 149, 243–248

Increased prostaglandin production by tissues in the sheep uterus and placenta are thought to be important for the onset of parturition. In the sheep placenta, this is most likely due to increased expression of prostaglandin synthase type-2 (PGHS-2) rather than prostaglandin synthase type-1 (PGHS-1). However, there is no information concerning expression of PGHS isoenzymes in maternal uterine tissues during pregnancy. Therefore, the purpose of the present study was to examine the expression of PGHS-1 and PGHS-2 in the sheep myometrium and endometrium during late gestation using in situ hybridization and immunohistochemistry. Using (35)S-labelled oligonucleotide probes, which give specific hybridization signals in other tissues, we localized PGHS-2 mRNA to endometrial epithelium, and apparently to other cells in both endometrium and myometrium. This artefactual signal was still present with 100-fold excess unlabelled oligonucleotide probe and with sense probes, but was resolved with the use of (33)P-oligonucleotides. Using (33)P-labelled oligonucleotide probes we could not detect either PGHS-1 or PGHS-2 mRNA in myometrium, and found expression only of PGHS-2 mRNA in endometrium. PGHS-2 mRNA localized to the endometrial epithelium and was undetectable in glandular epithelium. The level of PGHS-2 expression rose significantly between days 80 and 85 of pregnancy and term, and this corresponded to the appearance of immunoreactive PGHS-2 protein, measured by immunohistochemistry, in the endometrial epithelium. Therefore we conclude that (33)P-labelled probes are preferred for detection of mRNAs encoding PGHS-2 in ovine uterine tissues. Expression of PGHS-2 mRNA is greater than that of PGHS-1, increases during gestation, and predominates in the endometrial epithelium, consistent with the site of PGHS-2 protein localization.

The increase in plasma 11-hydroxycorticosteroid (11-OHCS) levels following surgery has been well documented (Mattingly & Tyler, 1965; Plumpton, Besser & Cole, 1969) but it was noted that these accounts made no mention of the state of fluid balance of the subjects. As this may vary considerably in surgical patients, the effect of an intravenous infusion on the degree of change of plasma 11-OHCS levels associated with cholecystectomy was studied in 34 otherwise healthy subjects. Informed consent was obtained from each patient. Patients were allocated at random to one of three groups. Group 1 (n = 9) received no infusion, group 2 (n = 13) an infusion during and after operation, and group 3 (n = 12) an infusion before, during and after operation. The pre-operative infusion consisted of 2l physiological saline given over 12 h. Over the operative period, 1l of compound sodium lactate solution was given in 1·5–2 h, followed by

In the late-gestation sheep, increased fetal plasma cortisol concentration and placental oestradiol (E(2)) output contribute to fetal organ maturation, in addition to the onset of parturition. Both cortisol and E(2) are believed to regulate the enzyme 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1), which interconverts bioactive 11-hydroxy glucocorticoids and their inactive 11-keto metabolites. 11beta-HSD1, abundantly expressed in fetal liver, operates primarily as a reductase enzyme to produce bioactive cortisol and thus regulates local hepatic glucocorticoid concentrations. Cortisol acts through the glucocorticoid receptor (GR) present in the liver. In this study, we examined the effects of cortisol and E(2) on hepatic 11beta-HSD1 and GR in the liver of chronically catheterized sheep fetuses treated with saline (n=5), cortisol (1.35 mg/h; n=5), saline+4-hydroxyandrostendione, a P450 aromatase inhibitor (4-OHA; 1.44 mg/h; n=5), or cortisol+4-OHA (n=5). Cortisol infusion resulted in increased plasma concentrations of fetal cortisol and E(2); concurrent administration of 4-OHA attenuated the increase in plasma E(2) concentrations. Using immunohistochemistry, we showed that fetal hepatocytes expressed both 11beta-HSD1 and GR proteins. Cortisol treatment increased GR in both cytosol and nuclei of hepatocytes; concurrent administration of 4-OHA was associated with distinct nuclear GR staining. Western blot revealed that cortisol, in the absence of increased E(2) concentrations, significantly increased concentrations of 11beta-HSD1 (34 kDa) and GR (95 kDa) proteins. 11beta-HSD1 enzyme activity was measured in the liver microsomal fraction in the presence of [(3)H]cortisone (10(-)(6) M) or [(3)H]cortisol (10(-)(6) M) and NADPH (reductase activity) or NADP(+) (dehydrogenase activity) respectively. 11beta-HSD1 reductase activity was significantly greater in the presence of cortisol. In summary, we found that, in sheep during late gestation, cortisol increased both 11beta-HSD1 and GR in the fetal liver, and these effects were accentuated in the absence of increased E(2).

ABSTRACT

The plasma concentrations of LH and prolactin and various parameters of ovarian function were examined in cows on known days of the oestrous cycle during May and June (autumn and winter) and during October (spring).

Luteinizing hormone peak frequency and plasma prolactin concentrations were significantly higher in October than during the May–June period (LH, P<0·05; prolactin, P<0·01). The mean diameters of large healthy follicles (≥8 mm diameter) and the dominant oestrogen-secreting follicles were significantly larger (P<0·01 for both follicle types) and each follicle contained more granulosa cells (both P<0·01) in May–June than in October. The LH responsiveness of theca interna with respect to androstenedione production and the levels of aromatase activity in granulosa cells did not differ with time of year. The corpora lutea were heavier (P<0·05) and secreted more progesterone (P<0·01) in May–June than in October.

It is concluded that seasonal differences in ovarian activity exist in cows and that these differences are probably the consequence of seasonal differences in gonadotrophin secretion.

J. Endocr. (1984) 102, 189–198