Chorionic Trophoblast Cells Research Articles

Regulation of 15-hydroxy prostaglandin dehydrogenase by corticotrophin-releasing hormone through a calcium-dependent pathway in human chorion trophoblast cells.

Prostaglandins (PGs) play a crucial role in mediating parturition events, and their synthesis and metabolism are regulated by PG H synthase and 15-hydroxy-PG dehydrogenase (PGDH), respectively. Within the chorion tissue, it is the actions of PGDH that predominate. Throughout gestation, the fetal membranes secrete increasing amounts of CRH. We hypothesized that CRH, produced locally in the chorion, could act to modulate PGDH activity throughout gestation. To investigate this, we obtained Percoll-purified human chorion and placental trophoblast cells from uncomplicated term pregnancies and cultured them for 72 h. Activity of PGDH was assessed by incubation (4 h) with PGF(2alpha) (282 nM) and measurement of conversion to 13,14-dihydro-15-keto PG F(2alpha). Dose-response curves were constructed for the chorion cell cultures with CRH or 8-bromo-cAMP. To investigate the role of CRH and calcium, cells were treated with either astressin, a CRH antibody, BAPTA, or EGTA. CRH (0-1 micro M) had no effect on PGDH activity; however, cells treated with astressin (10 micro M), with or without exogenous CRH (1 micro M), and cells treated with a CRH antibody showed a significant decrease in PGDH activity. 8-Bromo-cAMP (0-1 mM) had no effect on 13,14-dihydro-15-keto PG F(2alpha) output in chorion trophoblast cells but significantly decreased output from placental trophoblast cells. Cells treated with either BAPTA-AM or EGTA had significantly reduced PGDH activity; and, at intermediate concentrations of chelator, exogenous CRH restored PGDH activity. We suggest that, in chorion trophoblast cells, endogenously produced CRH exerts a tonic stimulatory effect on PGDH activity and may help maintain a metabolic barrier, preventing the transfer of bioactive PGs from the chorioamnion to the myometrium.

Expression of Matrix Metalloproteinase (MMP)-2 and MMP-9 in Human Placenta and Fetal Membranes in Relation to Preterm and Term Labor

Extensive extracellular matrix (ECM) remodeling is found in many processes during human parturition at term and preterm. These include cervical ripening, fetal membrane rupture, and placental detachment from the maternal uterus. Matrix metalloproteinases (MMPs) are the main mediators of ECM degradation. The present study was designed to investigate the expression of MMP-2 and MMP-9 in human fetal membranes (FMs) and placental (PL) tissues with or without labor at preterm and term parturition. Both zymography and Western blot analysis showed that MMP-9 was significantly (P < 0.01) increased in preterm and term labor FM, compared with nonlabor. Term labor PL also had a much higher (P < 0.05) level of MMP-9 than that of term nonlabor. No significant difference in MMP-2 expression was found between labor and nonlabor tissues. Immunolocalization studies revealed a specific distribution pattern for MMP-2 and MMP-9. MMP-2 was localized to the amnion mesenchyme, chorion laeve trophoblast, decidua parietalis, and blood vessels in PL villi. MMP-9 was localized mainly to amnion epithelia, chorion laeve trophoblast, decidua parietalis, and PL syncytiotrophoblasts. Separate cell culture from different layers of FM and culture of purified PL trophoblast cells showed that PL syncytiotrophoblast and amnion epithelial cells exclusively produced MMP-9; chorion trophoblast cells secreted both MMP-2 and MMP-9, but amnion mesenchymal cells produced only MMP-2. We concluded that MMP-2 and MMP-9 exhibited cell-specific expression in the human PL. An increase in MMP-9 expression may contribute to degradation of the ECM in the FM and PL, thereby facilitating FM rupture and PL detachment from the maternal uterus at labor, both preterm and term.

Expression of matrix metalloproteinase (MMP)-2 and MMP-9 in human placenta and fetal membranes in relation to preterm and term labor.

Extensive extracellular matrix (ECM) remodeling is found in many processes during human parturition at term and preterm. These include cervical ripening, fetal membrane rupture, and placental detachment from the maternal uterus. Matrix metalloproteinases (MMPs) are the main mediators of ECM degradation. The present study was designed to investigate the expression of MMP-2 and MMP-9 in human fetal membranes (FMs) and placental (PL) tissues with or without labor at preterm and term parturition. Both zymography and Western blot analysis showed that MMP-9 was significantly (P < 0.01) increased in preterm and term labor FM, compared with nonlabor. Term labor PL also had a much higher (P < 0.05) level of MMP-9 than that of term nonlabor. No significant difference in MMP-2 expression was found between labor and nonlabor tissues. Immunolocalization studies revealed a specific distribution pattern for MMP-2 and MMP-9. MMP-2 was localized to the amnion mesenchyme, chorion laeve trophoblast, decidua parietalis, and blood vessels in PL villi. MMP-9 was localized mainly to amnion epithelia, chorion laeve trophoblast, decidua parietalis, and PL syncytiotrophoblasts. Separate cell culture from different layers of FM and culture of purified PL trophoblast cells showed that PL syncytiotrophoblast and amnion epithelial cells exclusively produced MMP-9; chorion trophoblast cells secreted both MMP-2 and MMP-9, but amnion mesenchymal cells produced only MMP-2. We concluded that MMP-2 and MMP-9 exhibited cell-specific expression in the human PL. An increase in MMP-9 expression may contribute to degradation of the ECM in the FM and PL, thereby facilitating FM rupture and PL detachment from the maternal uterus at labor, both preterm and term.

Cortisol/progesterone antagonism in regulation of 15-hydroxysteroid dehydrogenase activity and mRNA levels in human chorion and placental trophoblast cells at term.

PGs mediate parturition events. 15-Hydroxyprostaglandin dehydrogenase (PGDH) catalyzes the first step in the metabolism of PGs to render them inactive. We have reported previously that cortisol (F) decreases PGDH activity and progesterone (P(4)) maintains PGDH in human chorion and placenta at term. To study the interaction of P(4) and F on the regulation of PGDH, we treated chorion and placental trophoblast cells in culture with combinations of F, dexamethasone, P(4), trilostane, and medroxyprogesterone acetate (MPA). Following a 24-h steroid treatment period and 4-h PGF(2alpha) challenge, culture media and cells were collected for measurement of PGF(2alpha) levels and PGDH mRNA by RIA and Northern blotting analysis. F and dexamethasone decreased PGDH activity and mRNA levels. Exogenous P(4) did not significantly alter PGDH activity or mRNA levels; however, MPA significantly stimulated PGDH activity. Trilostane decreased P(4) production by more than 90% and also decreased PGDH activity and expression. Coincubation with P(4) or MPA reversed trilostane inhibition of PGDH, consistent with a stimulatory role for endogenous P(4) on PGDH. MPA significantly reversed F inhibition of PGDH activity and mRNA levels. In the presence of trilostane, P(4) at equimolar concentration to F reversed F inhibition of PGDH mRNA levels. These findings suggest that F may be acting as an endogenous inhibitor of P(4) action in the regulation of PGDH at term.

Cortisol/Progesterone Antagonism in Regulation of 15-Hydroxysteroid Dehydrogenase Activity and mRNA Levels in Human Chorion and Placental Trophoblast Cells at Term

Cortisol/Progesterone Antagonism in Regulation of 15-Hydroxysteroid Dehydrogenase Activity and mRNA Levels in Human Chorion and Placental Trophoblast Cells at Term

The fetal placental hypothalamic–pituitary–adrenal (HPA) axis, parturition and post natal health

A general characteristic of fetal endocrine maturation across different species is the enhanced activity of the fetal hypothalamic–pituitary–adrenal (HPA) axis during late gestation. Precocious activation of this axis may occur when the fetus is exposed to an adverse intra-uterine environment, such as hypoxemia. HPA development is associated with increased levels of ACTH 1–39 and adrenal corticosteroids (cortisol in sheep and human) in the fetal circulation, and increased expression of mRNA encoding corticotrophin releasing hormone (CRH) in the hypothalamus, proopiomelanocortin (POMC) in the pituitary, and key steroidogenic enzymes in the fetal adrenal. At term, increased levels of cortisol act on the placenta/trophoblast derived cells to increase expression of prostaglandin synthase Type II (PGHS-II). In human gestation, cortisol also decreases expression of 15-hydroxyprostaglandin dehydrogenase (PGDH) in chorionic trophoblast cells. Increased synthesis and decreased metabolism of prostaglandin (PG) results, during late gestation, in enhanced output of primary PG, which in turn increases the activity of 11β-hydroxysteroid dehydrogenase (11β HSD) in the human fetal membranes. Increased chorionic 11β HSD-1 results in increased local generation of cortisol from cortisone, with further paracrine/autocrine stimulation of PG output. Increased fetal cortisol contributes to the maturation of organ systems required for postnatal extra-uterine survival. However, excessive levels of feto-placental glucocorticoid, derived from maternal administration of synthetic corticosteroids or sustained endogenous fetal cortisol production, results in intrauterine growth restriction. Fetal sheep, exposed to maternal betamethasone in late gestation, develop insulin resistance and exaggerated adrenal responses to HPA stimulation by 6–12 months postnatal life. Thus, the level of fetal HPA activity is crucial not only for determining gestation length, but may also predict pathophysiologic adjustments in later life.

Prostaglandin F2alpha potentiates cortisol production by stimulating 11beta-hydroxysteroid dehydrogenase 1: a novel feedback loop that may contribute to human labor.

In human pregnancy, cortisol and PGs are involved in the onset of labor and play an important role in the mechanisms leading to parturition. Recent studies have shown that at term, cortisol increases PG synthesis and decreases PG metabolism in chorion trophoblast (CT) cells. In CT, 11 beta-hydroxysteroid oxidase type 1 (11 beta-HSD1) converts biologically inactive cortisone to cortisol to regulate cortisol availability. In the present study, we have investigated whether 11 beta-HSD1 activity could be influenced by PGs. We have shown that in CT, PGF2alpha rapidly increased 11 beta-HSD1 reductase activity in a dose-dependent manner via the PGF2alpha receptor, localized in the fetal membranes. PGF2alpha stimulated 11 beta-HSD1 activity through increased intracellular calcium mobilization, activation of PKC, and the phosphorylation of the 11 beta-HSD enzyme. We propose that within CT there is a novel feed forward loop by which PGF2alpha acts to promote cortisol production from cortisone through increases in 11beta-HSD1, and this in turn leads to further net PG output for the onset of labor and birth.

Prostaglandin F2 Potentiates Cortisol Production by Stimulating 11 -Hydroxysteroid Dehydrogenase 1: A Novel Feedback Loop That May Contribute to Human Labor

In human pregnancy, cortisol and PGs are involved in the onset of labor and play an important role in the mechanisms leading to parturition. Recent studies have shown that at term, cortisol increases PG synthesis and decreases PG metabolism in chorion trophoblast (CT) cells. In CT, 11 beta-hydroxysteroid oxidase type 1 (11 beta-HSD1) converts biologically inactive cortisone to cortisol to regulate cortisol availability. In the present study, we have investigated whether 11 beta-HSD1 activity could be influenced by PGs. We have shown that in CT, PGF2alpha rapidly increased 11 beta-HSD1 reductase activity in a dose-dependent manner via the PGF2alpha receptor, localized in the fetal membranes. PGF2alpha stimulated 11 beta-HSD1 activity through increased intracellular calcium mobilization, activation of PKC, and the phosphorylation of the 11 beta-HSD enzyme. We propose that within CT there is a novel feed forward loop by which PGF2alpha acts to promote cortisol production from cortisone through increases in 11beta-HSD1, and this in turn leads to further net PG output for the onset of labor and birth.

The glial cells missing-1 protein is essential for branching morphogenesis in the chorioallantoic placenta.

Trophoblast cells of the placenta are established at the blastocyst stage and differentiate into specialized subtypes after implantation. In mice, the outer layer of the placenta consists of trophoblast giant cells that invade the uterus and promote maternal blood flow to the implantation site by producing cytokines with angiogenic and vasodilatory actions. The innermost layer, called the labyrinth, consists of branched villi that provide a large surface area for nutrient transport and are composed of trophoblast cells and underlying mesodermal cells derived from the allantois. The chorioallantoic villi develop after embryonic day (E) 8.5 through extensive folding and branching of an initially flat sheet of trophoblast cells, the chorionic plate, in response to contact with the allantois. We show here that Gcm1, encoding the transcription factor glial cells missing-1 (Gcm1), is expressed in small clusters of chorionic trophoblast cells at the flat chorionic plate stage and at sites of chorioallantoic folding and extension when morphogenesis begins. Mutation of Gcm1 in mice causes a complete block to branching of the chorioallantoic interface, resulting in embryonic mortality by E10 due to the absence of the placental labyrinth. In addition, chorionic trophoblast cells in Gcm1-deficient placentas do not fuse to form syncytiotrophoblast. Abnormal development of placental villi is frequently associated with fetal death and intrauterine growth restriction in humans, and our studies provide the earliest molecular insight into this aspect of placental development.

Activity and expression of 15-hydroxyprostaglandin dehydrogenase in cultured chorionic trophoblast and villous trophoblast cells and in chorionic explants at term with and without spontaneous labor

Objective: The aim of this study was to investigate whether any changes occurred at term before and with labor in the 15-hydroxyprostaglandin dehydrogenase messenger ribonucleic acid level and in the 15-hydroxyprostaglandin dehydrogenase activity in cultured chorionic and villous trophoblast cells and in chorionic explants. Study Design: Twelve placentas (labor group [vaginal delivery], n = 6; nonlabor group [elective cesarean delivery], n = 6) were collected. Chorionic trophoblast and villous trophoblast cells and chorionic disks were obtained, cultured, and incubated with 282-nmol/L prostaglandin F2α. Medium was collected to measure the 13,14-dihydro-15-keto metabolite of prostaglandin F2α, and the cells and disks were snap-frozen to quantify 15-hydroxyprostaglandin dehydrogenase messenger ribonucleic acid expression by Northern blot analysis. Results: The formation of the 13,14-dihydro-15-keto metabolite of prostaglandin F2α was significantly lower in the labor group than in the nonlabor group for both sets of cultured cells and for chorionic explants. 15-Hydroxyprostaglandin dehydrogenase messenger ribonucleic acid expression was lower in the chorionic trophoblast cells and chorionic disks of the labor group than those of the nonlabor group. However, the 15-hydroxyprostaglandin dehydrogenase messenger ribonucleic acid level in the villous trophoblast cells did not differ between the labor and nonlabor groups. Conclusion: Prostaglandin metabolic activity in the chorion is reduced significantly at the time of labor. (Am J Obstet Gynecol 2000;182:221-6.)

Human Placenta, Chorion, Amnion and Decidua Express Different Variants of Corticotropin-Releasing Factor Receptor Messenger RNA

Human placenta is a major source of corticotropin-releasing factor (CRF), and local effects of CRF in fetal membranes and placenta have been shown, i.e., adrenocorticotropic hormone (ACTH) and oxytocin release from cultured placental cells, as well as prostaglandin release from amnion, chorion and decidua. Two distinct CRF receptors (CRF-R1 and CRF-R2) have been characterized: CRF-R1 consists of two isoforms (CRF-R1α and CRF-R1β) while CRF-R2 has at least three different splice variants (CRF-R2α, CRF-R2β and CRF-R2γ). To date, CRF-R1 receptor has been identified in human placenta and in pregnant myometrium, while no evidence for placental CRF-R2 receptor isoforms has been provided. The present study investigated whether the different isoforms of CRF-R1 and CRF-R2 receptor mRNA are expressed in fetal membranes and placenta. Tissues were collected after spontaneous vaginal delivery (38–40 weeks) or elective caesarean section (39–41 weeks). The gene expression of CRF receptors was first studied by reverse transcriptase-polymerase chain reaction (RT-PCR), and the presence of CRF-R1α, but not of CRF-R1β, in human placental trophoblast, amnion/chorion and decidua was shown. In addition, among the three CRF-R2 splice variants, only CRF-R2β mRNA was expressed by trophoblast and fetal membranes. By using in situ hybridization, CRF-R1 and CRF-R2 probes positively hybridized trophoblast and related membranes. CRF-R1 was localized in the syncytiotrophoblast cells, chorionic trophoblast and decidua with a small amount in the amnion. CRF-R2 probe mainly hybridized syncytiotrophoblast cells, but cytotrophoblast also contained discreet amounts of CRF-R2 mRNA signal. The CRF-R2 hybridization signal was also observed within the structure of the villi (blood vessels), chorionic trophoblast and decidual cells, but it was faint or absent in the amniotic epithelium. There was no significant difference in the distribution of CRF-R1 or CRF-R2 mRNA signal between placentas collected from vaginal delivery or caesarean section. The evidence that intrauterine tissues differently express CRF-R1α and CRF-R2β supports possible different local roles of CRF and related peptides into intrauterine tissues during pregnancy.

Apoptotic cell death induced by actinomycin D arisen at different levels and cell cycle stages according to human cultured cell species

Nine kinds of human cultured cells, including fetus cells (smooth chorion trophoblast cells, amnion epithelial cells and HE-21), adult non-carcinoma cells (HCF), and carcinoma cells (KATO-III, COLO 201, Lu-134-AH, SK-OV-3 and SKG-3a) were stimulated with Actinomycin (Act.) D for 24 h. Apoptosis induction was investigated by agarose gel electrophoresis for DNA fragmentation analysis and by flow cytometric analysis of stained cells using in situ terminal deoxynucleotidyl-transferase (TdT)-mediated dUTP nick-end labeling TUNEL) staining techniques for the quantification of apoptosis, and simultaneously using propidium iodide for the gain of some information about cell cycle. By agarose gel electrophoresis, DNA fragmentation of these cells except amnion epithelial and SKG-3a cells was detected, depending on concentration of Act. D. Using flow cytometric analysis, these cells were separated into four groups according to the information about cell cycle. Group 1 included amnion epithelial and SKG-3a cells, which were TUNEL negative. In group 2, all cell populations at G0/G1 and G2/M phases of HCC, KATO-III and SK-OV-3 were TUNEL staining positive. A portion of each G0/G1 or G2/M phase cell of Lu-134-AH and COLO 201 in group 3 was TUNEL stain positive. In group 4, G2/M phase cells of smooth chorion trophoblast cells and HE-21 were mostly stained and a small population of G0/G1 phase cells were also TUNEL stain positive. These results show that the stages of the cell cycle at which apoptosis was arisen by Act. D stimulation were significantly different depending on the cells types.

Interleukin-10 modifies the effects of interleukin-1beta and tumor necrosis factor-alpha on the activity and expression of prostaglandin H synthase-2 and the NAD+-dependent 15-hydroxyprostaglandin dehydrogenase in cultured term human villous trophoblast and chorion trophoblast cells.

The concentrations of tumor necrosis factor-alpha (TNFalpha) and interleukin-1beta (IL-1beta), two inflammatory cytokines in amniotic fluid, have been shown to rise during chorioamnionitis. This is probably related to activation of the immune system in order to intensify the inflammatory process and to protect the maternal and fetal organism from infectious agents. These cytokines activate the PG biosynthetic pathway in several tissues, but few studies have examined effects on PG-metabolizing enzymes. When PGs are produced by increased synthesis and/or decreased metabolism at the chorio-decidual interface, labor can be induced. Interleukin-10 (IL-10) is known to act as an antiinflammatory cytokine. The goals of this study were to evaluate the interaction of IL-10 with IL-1beta and TNFalpha on PG synthesis and to determine the effects of IL-10, IL-1beta, and TNFalpha on PG metabolism using purified cultures of villous trophoblast and chorion trophoblast cells prepared from placentas of patients at term. Cells were treated with IL-1beta and TNFalpha with or without IL-10 for various times up to 24 h. Levels of messenger ribonucleic acid (mRNA) encoding PGH synthase-2 (PGHS-2) and NAD+-dependent 15-hydroxyprostaglandin dehydrogenase (PGDH) were quantified by Northern blotting, and PGE2 and 13,14-dihydro-15-keto-PGF2alpha (PGFM) output in the medium was measured by RIA. IL-1beta increased PGHS-2 mRNA and PGE2 output from villous and chorion trophoblasts and decreased PGDH mRNA in villous trophoblasts (all P < 0.05). These effects were reversed by IL-10. We found no change in PGHS-2 mRNA or PGE2 output in either trophoblast type treated with TNFalpha, but TNFalpha reduced PGDH mRNA in villous trophoblast, and this effect was reversed by IL-10 (both P < 0.05). We conclude that proinflammatory cytokines can influence PG output through effects on PG synthesis and metabolism and that these effects may be opposed by an antiinflammatory cytokine. These interactions may be important in the progression of preterm labor with underlying infection and in term labor in regions of the uterus where cytokine production is increased.

Interleukin-10 Modifies the Effects of Interleukin-1 and Tumor Necrosis Factor- on the Activity and Expression of Prostaglandin H Synthase-2 and the NAD+-Dependent 15-Hydroxyprostaglandin Dehydrogenase in Cultured Term Human Villous Trophoblast and Chorion Trophoblast Cells

Interleukin-10 Modifies the Effects of Interleukin-1 and Tumor Necrosis Factor- on the Activity and Expression of Prostaglandin H Synthase-2 and the NAD+-Dependent 15-Hydroxyprostaglandin Dehydrogenase in Cultured Term Human Villous Trophoblast and Chorion Trophoblast Cells

Cytokines and the regulation of apoptosis in reproductive tissues: a review.

To determine the role of apoptosis-regulating genes bax and bcl-2 in reproduction. Review of literature and current data. The bcl-2 family of apoptotic regulatory gene products interact and form dimers of anti- and pro-apoptotic proteins (e.g., bcl-2 and bax respectively), the ratio of which determines cell death or survival. Menses is associated with increased apoptosis in the glands, a change in bcl-2:bax ratio and increased levels of the pro-apoptotic cytokine TNFalpha. Apoptosis occurs in all placental cell types and increases from first to third trimester. Placental apoptosis is induced by TNFalpha in vitro and increased levels in utero characterize most failing pregnancies, intra-uterine growth restriction (IUGR) and labour. An increased bcl-2:bax ratio and apoptosis in the syncytiotrophoblast characterizes failing first trimester pregnancies. Apoptosis in the syncytiotrophoblast is also associated with IUGR. In a rat model, maternal vitamin A deficiency perturbs fetal development. This is associated with a placental infiltrate of TNFalpha positive neutrophils (day 20) and increased placental apoptosis in areas of infiltration. A similar infiltrate occurs in a mouse model of early pregnancy loss. In the fetal membranes, clusters of bcl-2 negative chorion trophoblast cells undergo apoptosis. This may allow passage of myometrial stimulatory factors that induce labour. The ratio of bcl-2:bax is crucial in the regulation of apoptosis, particularly in the human placenta. Changes in trophoblast apoptosis characterize (1) early pregnancy failure, (2) IUGR and (3) pre-term and term labour. Regardless of gestational age, TNFalpha plays a major role in the induction of placental apoptosis.

Local modulation by 11beta-hydroxysteroid dehydrogenase of glucocorticoid effects on the activity of 15-hydroxyprostaglandin dehydrogenase in human chorion and placental trophoblast cells.

NAD+-dependent 15-hydroxy-PG dehydrogenase (PGDH) is the major enzyme involved in the initial inactivation of PGs, and its activity is reduced by glucocorticoids, cortisol (F), and dexamethasone (DEX). In turn, glucocorticoid regulation of PGDH activity in placenta and chorion could be regulated indirectly by 11beta-hydroxysteroid dehydrogenase (11beta-HSD) activity. In the placenta, 11beta-HSD2 is the dominant isoform, acting as a dehydrogenase [F to cortisone (E)]; and in chorion, 11beta-HSD1 predominates as a reductase (E to F). The present study was designed to determine whether glucocorticoid regulation of PGDH activity in placenta and chorion could be regulated indirectly by 11beta-HSD activity. We obtained Percoll-purified human placental and chorion trophoblast cells from uncomplicated term pregnancies, cultured them for 72 h, then treated the cells with cortisol (100 nmol/L), cortisone (1 micromol/L), or DEX (100 nmol/L), in the presence or absence of carbenoxolone (CBX, 800 nmol/L), an 11beta-HSD inhibitor, for 24 h. Activity of PGDH was assessed by incubation (4 h) with PGF2alpha (282 nmol/L) and measurement of conversion to 13,14-dihydro-15-keto PGF2alpha. CBX alone had no effect on PGDH activity in either placenta or chorion trophoblast cells. In chorion, E significantly inhibited PGDH activity, and this effect was reversed by addition of CBX. F and DEX significantly inhibited PGDH, and this effect was unaltered by coadministration of CBX. In contrast, in placenta, there was no effect of E, or of E with CBX, on PGDH activity. However, F and DEX inhibited PGDH, and the effect of F (but not DEX) was greater in the presence of CBX. In conclusion, we suggest that effects of E and F on PGDH are modified by the tissue-specific expression of 11beta-HSD isoforms.

Local Modulation by 11 -Hydroxysteroid Dehydrogenase of Glucocorticoid Effects on the Activity of 15-Hydroxyprostaglandin Dehydrogenase in Human Chorion and Placental Trophoblast Cells

Local Modulation by 11 -Hydroxysteroid Dehydrogenase of Glucocorticoid Effects on the Activity of 15-Hydroxyprostaglandin Dehydrogenase in Human Chorion and Placental Trophoblast Cells

Steroid regulation of prostaglandin dehydrogenase activity and expression in human term placenta and chorio-decidua in relation to labor.

NAD+-dependent 15-hydroxyprostaglandin dehydrogenase (PGDH) is the key catabolic enzyme controlling levels of biologically active PGs. PGDH is localized to syncytiotrophoblast in placenta, and to trophoblast cells in chorion. To examine the regulation of PGDH by steroids and to determine any changes with labor, we obtained placenta and chorion from term elective cesarean section or spontaneous delivery and isolated trophoblast cells using a Percoll density gradient. Cells were treated with varying concentrations of cortisol, progesterone, the synthetic progestins R5020, and medroxyprogesterone acetate with or without RU486 or the specific progesterone receptor antagonist, onapristone, and the 3beta-hydroxysteroid dehydrogenase inhibitor, trilostane. The activity of PGDH was assessed by measurement of 13,14-dihydro-15-keto-PGF2alpha. PGDH messenger ribonucleic acid was quantified by in situ hybridization and computerized image analysis. The basal output of 13,14-dihydro-15-keto-PGF2alpha was lower in placenta or chorion collected at spontaneous labor than in that obtained at elective cesarean section. Cortisol had a significant dose-dependent inhibitory effect on PGDH activity in both placental and chorion trophoblast cells and significantly decreased levels of PGDH messenger ribonucleic acid. Responses were similar between tissues from laboring and nonlaboring women. PGDH activity was increased by R5020 and medroxyprogesterone acetate and was inhibited by RU486, onapristone, and trilostane. We conclude that cortisol inhibits PGDH activity and expression and that progestagens increase PGDH activity in human chorion and placenta.

Apoptosis and Fas expression in human fetal membranes.

Apoptosis (i.e. programmed cell death) plays a key role in maintaining reproductive function in the ovary, mammary and prostate glands, uterus, and testis. The purpose of the present report was to determine, based on biochemical and morphological parameters, whether cells in human fetal membranes undergo apoptosis and express Fas (CD95), a cell surface receptor that mediates apoptosis. Using the terminal deoxynucleotidyl transferase deoxy-UTP-nick end labeling immunohistochemical technique, apoptotic nuclei were identified in amnion epithelial, chorionic trophoblast, and decidua parietalis cell layers of human fetal membranes at term. Electron microscopy of fetal membranes revealed ultrastructural characteristics in amnion epithelium and chorion trophoblast cell layers consistent with apoptosis, including condensation of chromatin along the periphery of the nucleus and nuclear shrinkage. The apoptotic index (percentage of terminal deoxynucleotidyl transferase deoxy-UTP-nick end labeling-positive nuclei of the total nuclei) ranged from 8-29% in amnion epithelial, chorionic trophoblast, and decidual cell layers from women at 23-30, 31-36, and 37-42 weeks gestation. The apoptotic index was statistically greater in the 37-42 week group than in the 23-30 week group in chorionic trophoblast (P < 0.05) and decidual cell (P < 0.01) layers. In contrast, the apoptotic index in the amnion epithelial cell layer was statistically greater (P < 0.05) in the 23-30 week group than in the 31-36 week group, suggesting that apoptosis may be independently regulated in amnion epithelial, chorionic trophoblast, and decidual cell types. Based on the importance of Fas in mediating apoptosis, we investigated whether Fas was expressed by human fetal membrane cells. Immunohistochemical staining of fetal membranes with anti-Fas antibody localized Fas in amnion epithelial, chorionic trophoblast, and decidua parietalis cell layers. A 266-bp band corresponding to the cytoplasmic domain of Fas was detected in samples of amnion, chorion, decidua, and placenta by RT-PCR. Northern blotting revealed a molecular weight of approximately 1.9 kilobases for Fas messenger ribonucleic acid in amniotic tissue. These data suggest that apoptosis and Fas signaling may play a role in remodeling of fetal membrane architecture across gestation.

Differential regulation of 11 beta-hydroxysteroid dehydrogenase type 1 and 2 by nitric oxide in cultured human placental trophoblast and chorionic cell preparation.

Two types of 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD) have been identified in different tissues. Type 1 has both oxidase and reductase activities interconverting cortisol and cortisone, whereas type 2 has only oxidase activity converting cortisol to cortisone. It has been proposed that placental 11 beta-HSD controls the passage of maternal glucocorticoids to the fetal circulation. However, little is known about the regulation of 11 beta-HSD in the human placenta and fetal membranes. We cultured human term placental trophoblast and chorionic trophoblast cells to examine effects of nitric oxide donors, sodium nitroprusside (SNP) and S-nitroso-N-acetyl penicillamine (SNAP), on the activity and messenger RNA (mRNA) expression of 11 beta-HSD. At 72 h of culture, placental trophoblast formed syncytial clumps that were cytokeratin positive and displayed mainly type 2 oxidase activity, although some type 1 reductase activity was detectable. Chorion preparations contain greater than 90% trophoblast cells as demonstrated by immunostaining for cytokeratin and less than 5% vimentin positive cells. Type 1 reductase activity predominated in the chorionic trophoblast cells with barely detectable type 1 or type 2 oxidase activity. Both SNP (1-400 microM) and SNAP (1 mM) inhibited placental 11 beta-HSD type 2 oxidase activity but not type 1 reductase activity either in placental or chorionic cells. An inhibitory effect on type 2 oxidase activity was reproduced in part by 8-bromo cGMP, blocked partially by the guanylate cyclase inhibitor LY83583 (1 microM), but not by an ADP-ribosylation inhibitor N, N'-hexamethylene-bis-acetamide (HMBG) (10 mM). SNP also suppressed the expression of type 2 mRNA in cultured placental trophoblast in a dose-dependent manner, and this effect was also blocked by LY83583. We conclude that human placental trophoblast possesses predominantly 11 beta-HSD type 2 oxidase activity, whereas chorionic cells possess mainly type 1 reductase activity under the culture conditions employed. Nitric oxide specifically attenuated 11 beta-HSD type 2 oxidase activity as well as its mRNA expression in the placental trophoblast. The effect was mediated at least partially through the cGMP pathway, although an alternative pathway other than ADP-ribosylation may exist.