Receptors In Human Placenta Research Articles

Placental Leptin Receptor Isoforms in Normal and Pathological Pregnancies

Alternate mRNA splicing of human leptin receptor generates four membrane isoforms with different C-terminal sequences. They differ by the length of their intracellular domain which include specific motifs crucial for the specificity of leptin signalling. As a step towards functional studies, we have characterized leptin receptors in human placenta from normal pregnancies and pregnancies associated with diabetes and pre-eclampsia. Leptin and leptin receptors were visualized by immunohistochemistry of placentas obtained from first and third trimester pregnancies. Antibodies against N and C-terminal epitopes showed signals in the apical membrane of the syncytiotrophoblast in early and term placental villi as well as in JAr and BeWo derived trophoblast cells. In addition, a distinct isoform recognized by its extracellular juxtamembrane epitope was exclusively localized in cytotrophoblast cells and likely stains the soluble receptor. At contrast with the transmembrane receptors, the expression of this isoform is increased in placentas of pre-eclamptic and diabetic women which synthesize more leptin than placenta from uncomplicated pregnancy.These data demonstrate that short and long transmembrane leptin receptors are expressed in the trophoblast and indicate that leptin synthetized within the placenta can act locally through both receptor isoforms. Being also accessible to leptin from maternal origin, these transmembrane receptors may signal differently in pregnancy with normal and increased leptin production. The co-localization of leptin and the soluble receptor isoform suggests that this isoform serves for modulating maternal free leptin levels through modification of leptin binding capacities.

Expression of the Second Isoform of GnRH (Chicken II Type) and Its Receptor in Human Placenta

Objectives: Many studies have reported that GnRH is expressed in the extrahypothalamic tissues such as placenta, gonads and breast. A second isoform of GnRH (GnRH-II), also called chicken II type GnRH is widely distributed in most of the vertebrate animals including mammals. In the literature, however, there has been no study on GnRH-II in human placenta. Herein, we report for the first time the presence of second isoform of GnRH and its receptor in human placenta.

11 -Hydroxysteroid Dehydrogenase Type II and Mineralocorticoid Receptor in Human Placenta

11 -Hydroxysteroid Dehydrogenase Type II and Mineralocorticoid Receptor in Human Placenta

Cannabinoid receptors and their role in the regulation of the serotonin transporter in human placenta

Objective: We sought to investigate the expression of cannabinoid receptors in human placenta and BeWo choriocarcinoma cells and study their role in the regulation of the serotonin transporter. Study Design: Expression of the 2 types of cannabinoid receptors (CB1 and CB2) in human placenta and BeWo cells was investigated by reverse transcriptase–polymerase chain reaction and Northern blot analysis. The involvement of the receptors in the regulation of the serotonin transporter expression was studied by using a cannabinoid receptor agonist (WIN 55212-2). BeWo cells were treated with the agonist in the presence or absence of forskolin, and the serotonin transporter activity was measured by assessing paroxetine-sensitive serotonin transport. Serotonin transporter density in cell membranes was monitored by measuring paroxetine-sensitive binding of RTI-55, a specific high-affinity ligand for the transporter. Agonist-induced changes in intracellular levels of cyclic adenosine monophosphate were also monitored. Results: Reverse transcriptase–polymerase chain reaction and Northern blot analysis demonstrated unequivocally that human placenta and BeWo cells express both types of cannabinoid receptors. Treatment of BeWo cells with the receptor agonist blocked the activity of the constitutive, as well as the forskolin-induced, serotonin transporter without affecting the serotonin transporter density. This effect is not mediated by alterations in intracellular cyclic adenosine monophosphate levels. Conclusion: The results show that cannabinoid receptors are expressed in human placenta and BeWo cells and play a role in the regulation of the serotonin transporter activity. Human placenta is therefore a direct target for cannabinoids, and marijuana use during pregnancy is likely to affect the placental clearance of serotonin through the serotonin transporter. (Am J Obstet Gynecol 1999;181:491-7.)

TRAIL (Apo-2L) and TRAIL Receptors in Human Placentas: Implications for Immune Privilege

Mechanisms accounting for protection of the fetal semiallograft from maternal immune cells remain incompletely understood. In other contexts, interactions between TRAIL (TNF-related apoptosis-inducing ligand/Apo-2L) and its receptors kill activated lymphocytes. The purpose of this study was therefore to investigate the potential of the TRAIL/TRAIL-R system to protect the placenta against immune cell attack. Analysis by Northern blotting demonstrated mRNAs encoding TRAIL as well as the four TRAIL receptors (DR4, DR5, DcR1/TRID, DcR2/TRUNDD) in human placentas. Immunohistochemical experiments demonstrated that TRAIL protein is prominent in syncytiotrophoblast, an uninterrupted placental cell layer that is continuously exposed to maternal blood, as well as in macrophage-like placental mesenchymal cells (Hofbauer cells). Studies on cell lines representing trophoblasts (Jar, JEG-3 cells) and macrophages (U937, THP-1 cells) showed that both lineages contained TRAIL mRNA and that steady state levels of transcripts were increased 2- to 11-fold by IFN-gamma. By contrast, cell lineage-specific differences were observed in expression of the TRAIL-R genes. Although all four lines contained mRNA encoding the apoptosis-inducing DR5 receptor, only trophoblast cells contained mRNA encoding the DcR1 decoy receptor and only macrophages contained DcR2 decoy receptor transcripts. DR4 mRNA was present only in THP-1 cells and was the only TRAIL-R transcript increased by IFN-gamma. Cytotoxicity assays revealed that the two trophoblast cell lines were resistant, whereas the two macrophage lines were partially susceptible to killing by rTRAIL. Collectively, the results are consistent with a role for the TRAIL/TRAIL-R system in the establishment of placental immune privilege.

Gestational Age-Dependant Expression of Interleukin -10 (IL-10) and Its Receptor in Human Placenta

Gestational Age-Dependant Expression of Interleukin -10 (IL-10) and Its Receptor in Human Placenta

Gonadotropin-releasing hormone mRNA, gonadotropin-releasing hormone peptide, and variants of gonadotropin-releasing hormone receptor in human placenta

Gonadotropin-releasing hormone mRNA, gonadotropin-releasing hormone peptide, and variants of gonadotropin-releasing hormone receptor in human placenta

Localization of leukemia inhibitory factor and its receptor in human placenta throughout pregnancy.

Mice in which the gene that encodes the receptor (R) for leukemia inhibitory factor (LIF) has been deleted show abnormal growth and development of the placenta. This indicates that LIF plays an important role in placental development. The expression of LIF-R and LIF was examined in human trophoblast and decidua using in situ hybridization and immunocytochemistry. LIF-R mRNA and immunoreactivity was localized in villous and extravillous trophoblast throughout pregnancy, and in endothelial cells of the fetal villi. Strong expression of mRNA encoding LIF was detected in decidual leukocytes, which are abundant at the implantation site. Extravillous trophoblast, which invades the maternal decidua, therefore expresses LIF-R as it moves past decidual leukocytes, which express LIF mRNA. The effect of LIF on cultured human trophoblast was examined in vitro. Recombinant human LIF had no effect on [3H]thymidine incorporation by purified extravillous trophoblast, nor on expression of integrins alpha1, alpha5, or beta1 by isolated trophoblast. These results identify fetal endothelial cells and all cells of the trophoblast lineage as targets for the action of LIF in human placenta. Although its effects on trophoblast are not yet clear, LIF appears to mediate interactions between maternal decidual leukocytes and invading trophoblast. LIF may also play a critical role in controlling angiogenesis in the placental villi, since human fetal endothelial cells express LIF-R, and mice lacking a functional LIF receptor gene show altered vascular development in the placenta.

Studies on the localization of 5-hydroxytryptamine and its receptors in human placenta

The localization of 5-hydroxytryptamine (5-HT) and its receptors in human placenta was studied under light and electron microscopy using immunohistochemistry and in situ hybridization. The syncytiotrophoblast, cytotrophoblast, stromal cells and decidual cells in human placentae all appeared to be 5-HT immunoreactive. The 5-HT immunoreactive material was distributed in cytoplasm with negative nuclei. The 5-HT immunoreactive material was also found in capillary endothelium. Trophoblast cells cultured in serum-free medium also showed 5-HT immunoreactivity in the cytoplasm. Fetal white blood cells and both syncytiotrophoblast and cytotrophoblast in human placenta showed 5-HT receptor immunoreactivity and 5-HT 1A receptor mRNA hybridized signal was also detected in cytoplasm. The stromal cells and capillary endothelium in placental villi and maternal decidual cells all showed 5-HT receptor immunoreactivity in cytoplasm. The small flattened vesicles and large dense cored vesicles within trophoblast cells showed electron-dense 5-HT receptor immunoreactivity using immunoelectron microscopy. These results suggest that human placenta may not only produce 5-HT but also be a 5-HT target organ, and that 5-HT may not only play roles in placental development and pregnancy maintenance by paracrine and autocrine interactions but may also take part in regulating fetal development.

Expression and localization of the Th2-type cytokine interleukin-13 and its receptor in the placenta during human pregnancy.

To investigate the expression of the Th2-type cytokine interleukin (IL)-13 and its receptor in human placenta during gestation. Expression of IL-13 and its receptor was analyzed by reverse transcriptase (RT)-polymerase chain reaction (PCR), in situ hybridization and immunohistochemistry using human placental samples. IL-13 mRNA was detected by RT-PCR in placental extracts from all stages of gestation. In situ hybridization revealed IL-13 mRNA in first trimester cytotrophoblast and syncytiotrophoblast. Few positive cells were found within decidual sections from the same pregnancy. Immunohistochemistry revealed a similar pattern to in situ hybridization. This was largely absent in second- and third-trimester placentae. IL-13 receptor alpha chain (IL-13R alpha) was detected by immunofluorescence on the surface of leukocytes in first-trimester villous core and decidua. The expression of IL-13 is spatially and temporally regulated in the placenta, and associated with the close proximity of a receptor-bearing target leukocyte population.

Gene expression of oxytocin and oxytocin receptor in human placenta

Cyclooxygenase catalysis by prostaglandin H synthase (PGHS)-1 and -2 involves reaction of a peroxide-induced Tyr385 radical with arachidonic acid (AA) to form an AA radical that reacts with O2. The potential for isomeric AA radicals and formation of an alternate tyrosyl radical at Tyr504 complicate analysis of radical intermediates. We compared the EPR spectra of PGHS-1 and -2 reacted with peroxide and AA or specifically deuterated AA in anaerobic, single-turnover experiments. With peroxide-treated PGHS-2, the carbon-centered radical observed after AA addition was consistently a pentadienyl radical; a variable wide-singlet (WS) contribution from mixture of Tyr385 and Tyr504 radicals was also present. Analogous reactions with PGHS-1 produced EPR signals consistent with varying proportions of pentadienyl and tyrosyl radicals, and two additional EPR signals. One, insensitive to oxygen exposure, is the narrow singlet tyrosyl radical with clear hyperfine features found previously in inhibitor-pretreated PGHS-1. The second type of EPR signal is a narrow singlet lacking detailed hyperfine features that disappeared upon oxygen exposure. This signal was previously ascribed to an allyl radical, but high field EPR analysis indicated that ~ 90% of the signal originates from a novel tyrosyl radical, with a small contribution from a carbon-centered species. The radical kinetics could be resolved by global analysis of EPR spectra of samples trapped at various times during anaerobic reaction of PGHS-1 with a mixture of peroxide and AA. The improved understanding of the dynamics of AA and tyrosyl radicals in PGHS-1 and -2 will be useful for elucidating details of the cyclooxygenase mechanism, particularly the H-transfer between tyrosyl radical and AA.

Identification of L-type calcium channels associated with kappa opioid receptors in human placenta.

Transduction pathways of kappa receptor activation are not fully understood. Human placenta at term expresses only this type of opioid receptors and therefore offers a unique advantage for such investigations. It has previously been postulated that kappa receptors-mediated modulation of acetylcholine and placental lactogen release from human placentas require the influx of extracellular calcium and into the cells, possibly via voltage-dependent channels. We report here that another opioid-regulated placental function, the release of human chorionic gonadotropin (hCG), depends on extracellular calcium and the modality of its influx via L-type channels. Data presented demonstrated that the stimulation of hCG secretion by the kappa-selective agonist U69,593 was abolished in presence of either EGTA or the calcium channel blocker nifedipine. Results obtained on the combined effect of opioids and dihydropyridines indicated that placental kappa opioid receptors could be directly coupled to L-type calcium channels. The identification of the latter in villus membrane preparations, reported here for the first time, further contributes to the hypothesis that, in human placenta, kappa receptors-linked transduction mechanisms involve calcium and its conductance across villus membranes.

Microscopic and ultramicroscopic localizations and quantitative analysis of 5-HT receptors in human placentas

The localizations of 5-hydroxytryptamine receptor (5-HTR) at light and electron microscopic levels and its quantitative analysis in human placentas were studied by using immunohistochemistry andin situ hybridization. Both syncytiotrophoblast and cytotrophoblast in placental villi and fetal white blood cells in villose capillary cavity showed 5-HT receptor immunoreactivity, with 5-HTIA receptor mRNA hybridized signal detected in cytoplasm. But the stromal cells and capillary endothelium in placental villi showed 5-HT receptor immunoreactivity in cytoplasm, without 5-HT1A receptor mRNA hybridized signal detected. This suggested that two layers of trophoblast cells may produce 5-HT1 and 5-HT2 receptors, that the stromal cells and capillary endothelium in placental villi may only produce 5-HT2 receptor. By immunohistochemistry at electron microscopic level, the small flattened vesicles and large dense cored vesicle within trophoblast cells showed 5-HT receptor immunoreactivity. This suggested that it may be the result of 5-HT receptors internalization and transportion. Using a quantitative immunohistochemical method, the contents of 5-HT receptor in placenta were higher during the 6th week of gestation, and decreased in 7th and 8th. reoccurred the second peak in the 9th. reduced gradually during the 10th, 20th and 40th of the gestation period. These changes paralleled the contents of 5-HT in the authors’ studies, reflecting that 5-HT may be one of the most important bioactive substances in placental self-regulation.

Interleukin-1 receptors in human placenta and their role in the regulation of the serotonin transporter gent expression

Interleukin-1 receptors in human placenta and their role in the regulation of the serotonin transporter gent expression

Nucleotide sequence analyses predict that human pituitary and human placental gonadotropin-releasing hormone receptors have identical primary structures.

Gonadotropin-releasing hormone (GnRH) interacts with a putative receptor in human placenta to cause the dose-dependent release of human chorionic gonadotropin (hCG) in a manner analogous to hypothalamic GnRH stimulation of luteinizing hormone (LH), and follicle-stimulating hormone (FSH) by the pituitary gland. However, GnRH agonists bind a placental binding site at a lower affinity than they bind the pituitary GnRH receptor, suggesting the two sites differ. To address this issue, human placental GnRH receptor mRNA from an 8-wk placental sample was amplified using primers based on the sequence of the human pituitary receptor, cloned, and sequenced. The sequence of the placental transcript was found to be identical to its pituitary counterpart. Thus, as with the pituitary receptor, this placental receptor transcript appears to encode a single polypeptide chain with seven intramembranous domains and two glycosylation sites. However, it is possible these receptors differ in their posttranslational processing or there is more than one placental GnRH receptor. In addition, subsequent gene amplification studies, modified to increase the sensitivity of the method, revealed a band of the predicted size (approximately 1051 bp) in a 6-wk and two 8-wk placental samples, but not in 5-wk and 7-wk samples. The failure to amplify GnRH receptor mRNA at these gestational ages did not appear to be an issue of cell health or sample degradation, since GnRH, alpha hCG, and beta hCG mRNA were amplified from each of these tissues. Instead these data suggest GnRH receptor mRNA may have a short half-life or the mRNA may be translated as rapidly as it is transcribed, with the result that it has not accumulated in the cell.

Association of the transferrin receptor in human placenta with HFE, the protein defective in hereditary hemochromatosis.

Hereditary hemochromatosis (HH) is a common autosomal recessive disease associated with loss of regulation of dietary iron absorption and excessive iron deposition in major organs of the body. Recently, a candidate gene for HH (also called HFE) was identified that encodes a novel MHC class I-like protein. Most patients with HH are homozygous for the same mutation in the HFE gene, resulting in a C282Y change in the HFE protein. Studies in cultured cells show that the C282Y mutation abrogates the binding of the recombinant HFE protein to beta2-microglobulin (beta2M) and disrupts its transport to the cell surface. The HFE protein was shown by immunohistochemistry to be expressed in certain epithelial cells throughout the human alimentary tract and to have a unique localization in the cryptal cells of small intestine, where signals to regulate iron absorption are received from the body. In the studies presented here, we demonstrate by immunohistochemistry that the HFE protein is expressed in human placenta in the apical plasma membrane of the syncytiotrophoblasts, where the transferrin-bound iron is normally transported to the fetus via receptor-mediated endocytosis. Western blot analyses show that the HFE protein is associated with beta2M in placental membranes. Unexpectedly, the transferrin receptor was also found to be associated with the HFE protein/beta2M complex. These studies place the normal HFE protein at the site of contact with the maternal circulation where its association with transferrin receptor raises the possibility that the HFE protein plays some role in determining maternal/fetal iron homeostasis. These findings also raise the question of whether mutations in the HFE gene can disrupt this association and thereby contribute to some forms of neonatal iron overload.

Identification, characterization and localization of corticotropin-releasing hormone receptors in human placenta.

The purpose of this work was to identify, characterise, and localise specific CRH binding sites in whole vaginally delivered term human placental membranes as well as in dispersed and purified trophoblastic membranes. We have found that whole placenta membranes contained specific and saturable CRH-binding sites. Maximum specific binding was obtained at pH 7.2-7.4, 22 C, for 2 h. The order of potency of CRH analogs was h/rCRH > alpha-helCRH > oCRH. Scatchard analysis revealed a single population of high affinity binding sites with a dissociation constant of 1.25 nM and maximum binding capacity of 119 fmol/mg of protein. Purified syncytiotrophoblast membranes contained high affinity CRH binding sites exhibiting the same dissociation constant and maximum binding capacity as whole placental membranes, suggesting that the CRH binding sites are expressed almost exclusively by the syncytiotrophoblast. Our findings indicate that CRH binding sites in the human placenta are exhibit similar characteristics to that described for anterior pituitary, brain and adrenals.

Down-regulation of angiotensin AT1 receptor by progesterone in human placenta.

Regulation of the angiotensin AT1 receptor in human placenta is poorly understood. In this study, we analyzed the time course of angiotensin AT1 receptor expression, internalization, and recycling by human trophoblast cells. We also studied the effects of estradiol, progesterone, and chloroquine on regulation of the angiotensin AT1 receptor in 48-h cell culture. The angiotensin II receptor expression increased with the time of incubation, reaching a level at 48 h of culture that was about 120% above the initial value. A large majority of angiotensin II receptors was of the AT1 subtype, as it was completely inhibited by losartan (1 mumol/L). The internalization of [125]angiotensin II binding and the angiotensin AT1 receptor recycling were also time dependent, with t1/2 values of 12 and 21 min, respectively. In human trophoblast cells exposed to progesterone (10 mumol/L) for 48 h, angiotensin AT1 receptor density was decreased by 49%, whereas estradiol (10 mumol/L) or chloroquine (100 mumol/L) treatment was ineffective. In the freshly isolated trophoblast cells initially treated with unlabeled angiotensin II (200 nmol/L) for 30 min, chloroquine was shown to decrease angiotensin AT1 receptor recycling by 73%, whereas estradiol and progesterone had no effect. These findings indicate that progesterone induces a down-regulation of the angiotensin AT1 receptor in human placenta and that the recycling of this receptor can be delayed by chloroquine.

Purification and characterization of 1,25(OH)2D3 receptor from human placenta.

To characterize the receptor for 1,25-dihydroxycholecalciferol [1,25-(OH)2D3], we purified it from nuclear fractions of human placentae. Human placental fractions were concentrated with ammonium sulfate, extracted from hydroxylapatite, and then chromatographed on Sepharcryl S-200 and DEAE-cellulose. The receptor for [1,25(OH)2D3] was purified approximately 1,500-fold. The molecular weight of the receptor was estimated to be 55 K dalton by gel filtration. The receptor fractions showed a dissociation constant (Kd) of 3.0 x 10(-10) mol/l, and adsorbed to the DNA cellulose column. D3 analogs, estradiol, and progesterone had almost no effect on 1,25(OH)2D3 binding. These properties of the 1,25(OH)2D3 receptor in human placenta are similar to those of the chicken intestinal 1,25(OH)2D3 receptor.

Expression of vascular endothelial growth factor receptor in human placenta

Expression of vascular endothelial growth factor receptor in human placenta