Receptors In Human Placenta Research Articles

Expression of Mas1 receptor in human placenta and its effect on the function of trophoblast cells in pre-eclampsia patients

Objective: To investigate the expression change of the Mas1 receptor in the placenta of healthy pregnant women during different gestation periods, analyze the expression level of the Mas1 receptor in the placenta of pre-eclampsia (PE) patients, and its biological function in trophoblast cells. Methods: Placental villous tissues were collected from normal pregnant women in early, mid and late pregnancy. Human trophoblast stem cells were isolated and cultured from early pregnancy villous tissues. The expression of the Mas1 receptor was detected by fluorescence immunoassay and real-time fluorescence quantitative PCR. In a case-control study, patients with full-term PE were selected as the case group and healthy women with full-term pregnancy were selected as the control group. Placental villus tissues were collected from both groups. Immunofluorescence chemistry and immunoprotein blotting were used to study the changes in Mas1 receptor expression in PE. Mas1 receptor agonists and blockers induced HTR8/Svneo cells and BeWo cells, and the effects of the Mas1 receptor on the proliferation and migration of trophoblast cells were detected by the CCK8 proliferation test and scratch test. Results: Eight cases were included in early pregnancy, seven cases in mid-pregnancy and six cases in late pregnancy. Mas1 receptors in normal placental villi tissue were mainly expressed in human trophoblast stem cell membranes and cytoplasm, and the expression of Mas1 receptor mRNA in villi tissue was significantly higher in late pregnancy than in mid-pregnancy. There were 24 cases included in the case group and 12 cases in the control group. Mas1 receptor expression in placental villi was significantly lower in the case group compared to the control group; Activation/inhibition of the Mas1 receptor had no significant effect on the proliferation of HTR8/Svneo cells and BeWo cells. Activated Mas1 receptor had no significant effect on the migration ability of HTR8/Svneo cells. Conclusion: Mas1 receptors are expressed in placental villous tissue and their expression varies with gestation. Mas1 receptor expression is reduced in PE patients, but it does not affect the value-added or migratory function of trophoblast cells.

Apelin and apelin receptor in human placenta: Expression, signalling pathway and regulation of trophoblast JEG‑3 and BeWo cells proliferation and cell cycle.

Placentation requires the production of numerous growth factors, hormones and transcription factors. Many of them, like the adipose tissue-derived leptin or adiponectin, have been identified in the placenta and their role has been established in the proliferation and subsequent development of the placenta. Apelin is another adipokine known for proliferative effects in different cell types. PCR, immunoblotting and immunocytochemistry were used to study mRNA and protein expression of apelin and its receptor (APJ) in syncytiotrophoblast (BeWo) and cytotrophoblast (JEG-3) cells as well in immunohistochemistry in human normal placenta slides. The effect of apelin on cell proliferation study was investigated by alamarBlue® and Cell Counting Kit-8 assays, the cell cycle by the flow cytometry method and the protein expression of cyclins and phosphorylation level of extracellular signal-regulated kinases (ERK)1/2, phosphatidylinositol 3′-kinase/protein kinase B (Akt), signal transducer and activator of transcription 3 (Stat3) and 5′-monophosphate-activated protein kinase (AMPKα) were studied by western blotting. Apelin was increased in JEG-3 compared with in BeWo cells, while APJ was the same in both placenta cell lines. Immunocytochemical analyses revealed high cytoplasmic and/or membrane apelin localisation in JEG-3, while BeWo cells exhibited markedly weaker apelin signal in the cytoplasm. Apelin increased cell proliferation as well as the percentage of cells in the G2/M phase of the cell cycle, cyclin proteins and the expression of all kinases mentioned above. In conclusion, apelin by promotion of trophoblast cell proliferation by APJ and ERK1/2, Stat3 and AMPKα signalling could be a new important adipokine in the regulation of early placental development.

The regulation of oxytocin and oxytocin receptor in human placenta according to gestational age

Oxytocin (OXT) is a peptide hormone that plays a central role in the regulation of parturition and lactation. OXT signaling is mediated by OXT receptor (OXTR), which shows species- and tissue-specific expressions and gene regulation. In the present study, we examined the synthesis of OXT and OXTR in human placenta tissue according to gestational age. A total of 48 placentas were divided into early preterm, late preterm and term groups depending on gestational age, and expression of OXT and OXTR was evaluated. First, OXT and OXTR mRNA and protein were detected in normal placenta tissue via Q-PCR, Dot-blot and Western blot assay. Both OXT and OXTR levels in normal placenta increased gradually in the late stage of pregnancy, suggesting that local OXT may play a critical role in the function of the placenta. To determine the regulatory mechanism of OXT, placental BeWo cells were administrated estrogen (E2) or progesterone (P4), and expression of OXT and OXTR was tested. The mRNA and protein levels of OXT and OXTR were upregulated by E2 but blocked by co-treatment with P4 In order to confirm the estrogen receptor (ESR)-mediated signaling, we administrated ESR antagonists together with E2 to BeWo cells. As a result, both OXT and OXTR were significantly altered by ESR1 antagonist (MPP) while moderately regulated by ESR2 antagonist (PHTPP). These results suggest that OXT and OXTR are controlled mainly by E2 in the placenta via ESR1 and thus may play physiological functions in the human placenta during the late stage of pregnancy.

Leptin and its Receptors in Human Placenta of Small, Adequate, and Large for Gestational Age Newborns.

Alterations in birth weight impact postnatal outcome and adult metabolic health. Therefore, fetal growth regulation is crucial for preventing chronic metabolic diseases. Leptin has been suggested to play an important role in placental and fetal growth, albeit its specific mechanisms of action have not been elucidated. The aim of this study was to analyze leptin concentrations in placenta, cord blood, and maternal blood of SGA, AGA, and LGA (small, adequate and large for gestational age, respectively) newborns, as well as placental leptin receptor (LEPRa and LEPRb) protein expression. We performed a cross-sectional comparative study in 3 groups of healthy mothers and their term newborns at delivery (SGA, AGA, and LGA, n=20 per group). Placental, maternal blood, and cord blood leptin content were measured by ELISA. Placental LEPRa and LEPRb protein expression were determined by Western Blot. Maternal leptin concentrations correlated positively with maternal weight before and at the end of gestation, without differences between groups. Cord leptin is higher in LGA and lower in SGA, whereas placental leptin is higher in SGA. Placental leptin was inversely correlated with placental weight, independently from maternal weight and gestational age. Both LEPRa and LEPRb expression are lower in SGA, while LEPRa positively correlated with placental weight and birthweight. The current findings indicate that placental leptin and its receptors are differentially expressed in SGA, AGA, and LGA newborns. We suggest that placental leptin and LEPR protein expression may influence placental growth and thus, birth weight.

Transient receptor potential vanilloid 1 is expressed in human cytotrophoblasts: Induction of cell apoptosis and impairment of syncytialization

The normal development of placenta relies essentially on a balanced proliferation, differentiation and apoptosis of cytotrophoblasts. These processes are tightly regulated by several hormones, cytokines, lipids and other molecules and anomalies in these events are associated with gestational complications. The cation channel transient receptor potential vanilloid 1 (TRPV1) is expressed in several organs and tissues and it participates in cellular events like nociception, inflammation and cell death. However, the expression and importance of this receptor in human placenta still remains unknown. In this work, we found that TRPV1 is expressed in human cytotrophoblasts and syncytiotrophoblasts. Furthermore, the TRPV1 agonists capsaicin and anandamide decreased cytotrophoblast viability and induced morphological alterations, such as chromatin condensation and fragmentation, which suggest the occurrence of apoptosis. Also, both TRPV1 agonists induced a loss of mitochondrial membrane potential and an increase of caspase 3/7 activity and production of reactive species of oxygen and nitrogen. Furthermore, capsaicin (10μM) impaired the spontaneous in vitro differentiation of cytotrophoblasts into syncytiotrophoblasts by triggering TRPV1, as observed by the decrease in placental alkaline phosphatase activity and in human chorionic gonadotropin secretion. On the other hand, anandamide decreased placental alkaline phosphatase activity via a TRPV1-independent mechanism but did not influence the secretion of human chorionic gonadotropin. In conclusion, we showed that TRPV1 is expressed in human cytotrophoblasts and syncytiotrophoblasts and also reported the involvement of this receptor in cytotrophoblast apoptosis and differentiation.

Expression of genes coding for proangiogenic factors and their receptors in human placenta complicated by preeclampsia and intrauterine growth restriction

The aim of the study was to investigate the expression of genes coding for vascular endothelial growth factor (VEGF) and placenta growth factor (PlGF) as well as their receptors, fms-like tyrosine kinase receptor 1 (VEGFR-1/Flt-1) and VEGF receptor 2 (VEGFR-2/KDR) in the placentae of patients with pregnancies complicated by preeclampsia (PE) and intrauterine growth restriction (IUGR). Tissue samples were collected from placentae of women with PE (n=31) and IUGR syndrome (n=25) as well as of healthy control women (n=31). Total RNA was extracted and purified, mRNA reversely transcribed, and amplified using real-time PCR. Expression of the examined genes was normalized to β-actin. Higher levels of PlGF (p<0.001) and Flt-1 (p<0.05) transcription were found in PE placentae compared to normal ones. A positive correlation between PlGF and Flt-1 expression was revealed in the PE patients. In conclusion, the presented data indicate the upregulation of both PlGF and Flt-1 in placentae of women with PE, which could be induced by a pathological process possibly due to endothelial dysfunction.

206: Melanocortin binding activity in term and preterm human placenta

term and preterm human placenta J. Newton, Minying Cai, Hua Xu, Kathryn Reed University of Arizona, Obstetrics and Gynecology, Tucson, AZ, University of Arizona, Chemistry, Tucson, AZ, University of Arizona, Pediatrics, Tucson, AZ OBJECTIVE: The purposes of this study were to compare melanocortin binding activity and melanocortin receptor subtype expression in human placenta samples collected in four clinical settings: preterm notin-labor, preterm labor, term not-in-labor, and term labor. STUDY DESIGN: Following IRB approval, placenta samples were collected at delivery in each of the four clinical settings listed above. Melanocortin binding activity using 125I-NDP-MSH was determined with 1 mg of total membrane protein from each sample. Competition binding assays between the non-selective 125I-NDP-MSH and -MSH analogs that have highly specific receptor subtype agonist activity were used to identify receptor subtype activity. Lastly, 500 ng of total placental RNA was used as original template for quantitative RT-PCR using validated TaqMan probes for melanocortin receptors 1 and 3 (MC1R and MC3R). Average MSH binding activities were compared between groups using two-tailed Students t-test. Relative expression of MC1R and MC3R were compared using the Pfaffl method. RESULTS: Human placenta samples express MSH-binding activity. This activity is significantly higher in the setting of labor compared to not-in-labor, both at term and preterm. Competition binding assays with highly selective receptor agonists showed that MC3R is the predominant melanocortin receptor in human placenta. In the quantitative RT-PCR studies, both MC1R and MC3R are detected at varying expression levels. While MC1R was expressed at a relatively constant level in this sample set, MC3R expression levels were significantly higher in term labor samples compared to preterm not-in-labor samples. CONCLUSION: Melanocortin receptors, which have important physiologic functions including roles in energy homeostasis and inflammation, are expressed in human placenta. The key melanocortin receptor in placental MSH binding activity appears to be the MC3R subtype based on both the RT-PCR and competitive binding assays. Finally, the level of melanocortin binding activity and MC3R gene expression varies with gestational age and in the setting of term and preterm labor. 207 Role of foramen ovale in the regulation of fetal cardiac output: a fetal sheep study with acute ductus arteriosus occlusion Jason Hashima, David Sahn, Muhammad Ashraf, Vanessa Rogers, Stephen Langley, Lowell Davis, Roger Hohimer, Juha Rasanen Oregon Health and Science University, Obstetrics and Gynecology, Portland, OR, Oregon Health and Science University, Pediatric Cardiology, Portland, OR, Oregon Health and Science University, Pediatric Surgery, Portland, OR, Oregon Health & Science University, Maternal Fetal Medicine, Portland, OR OBJECTIVE: We hypothesized that during acute fetal ductus arteriosus (DA) occlusion (DO) foramen ovale (FO) volume blood flow increases, in order to maintain adequate systemic cardiac output. STUDY DESIGN: Nine ewes with singleton pregnancies underwent surgery at 120-126 gestational days (term 145 days) for the placement of a vascular occluder around fetal DA. Fetal carotid artery and jugular vein were cannulated. After a 5-day recovery, fetal heart rate (FHR), right (RVCO) and left (LVCO) ventricular cardiac outputs were measured by ultrasonography. Pulmonary (Qp) and foramen ovale (Qfo) volume blood flows, and systemic CO (combined cardiac output-Qp) were calculated at baseline, 15 and 60 minutes after DO, and 15 minutes after release of DO. Fetal mean arterial (mABP) and central venous (CVP) pressures, and pO2 and oxygen saturation were monitored. RESULTS: All the data are presented as means (SD). * p 0.05, compared with baseline. CONCLUSION: Acute DO in fetal sheep leads to decreased RVCO and increased LVCO, however systemic CO decreases. The rise in LVCO is caused by increased Qp with no change in the Qfo. Fetal CVP increases and arterial oxygen saturation decreases during DO. These findings suggest that fetal foramen ovale has a limited capacity to increase its volume blood flow, at least in response to acutely increased right ventricular afterload. Poster Session I Clinical Obstetrics, Medical-Surgical-Disease, Neonatology, Physiology-Endocrinology www.AJOG.org

Expression of Adiponectin Receptors in Human Placenta and Its Possible Implication in Gestational Diabetes

Problem statement: Similar to obese patients and type 2 diabetic patients, adiponectin levels are reduced in former Gestational Diabetes Mellitus (GDM) patients and are lower in GDM women during late pregnancy compared with pregnant control subjects matched for BMI. Diabetic insult at later stages in gestation, such as may occur in gestational diabetes, will foremost lead to short-term changes in a variety of molecules for key functions including gene expression in the placenta. Approach: In this study we assessed the expression of adiponectin receptors in human placenta to identify the site (s) of expression and to clarify the effect of gestational diabetes in this expression. This study was carried on 10 normoglycemic pregnant women and 20 GDM women. The placental tissue was collected immediately after delivery and tissue biopsies were taken from both fetal and maternal sides of each placenta. One step-RT-PCR for ADIPOR1 and ADIPOR2 was done by Real Time PCR using Syber Green technique. Relative quantification of mRNA of the ADIPOR1 and ADIPOR2 genes was measured using ABI7900 Real Time machine. Results: Both types of Adiponectin Receptors (ADIPOR1 and ADIPOR2) are expressed in human placenta. ADIPOR1 is more highly expressed than ADIPOR2 in both fetal and maternal sides of GDM cases and normal pregnant women. ADIPOR1 mRNA expression was significantly up regulated in GDM women compared to normal pregnant women, whereas no significant difference in the expression of ADIPOR2 was detected between the two groups. There was no evidence of maternal-fetal side difference in the expression of adiponectin receptors in GDM cases but in normal pregnant women there is a statistically significant difference between both sides in the expression of both ADIPOR1 and ADIPOR2. Conclusion: We concluded that adiponectin plays an important role in mediation the glucose metabolism in fetal tissues through its receptors, mainly Adiponectin Receptor 1 (ADIPOR1).

Expression of Androgen Receptor in Human Placentas from Normal and Preeclamptic Pregnancies

To elucidate the expression of androgen receptor (AR) in human placenta from normal and preeclamptic (PE) pregnancies. This study was performed at the Division of Obstetrics and Gynecology, Kaohsiung Chang Gung Memorial Hospital, Academic Medical Center. Forty normal pregnant women at 37.0 +/- 0.89 weeks' gestation and 40 women with preeclampsia at 35.9 +/- 1.33 weeks' gestation were included in this study. Serum testosterone was measured before delivery. Immunochemical methodology was employed in formalin-fixed, paraffin-embedded sections from the placentas of all subjects. AR expression was evaluated in placentas collected from 15 normal pregnant women and 12 PE women. Women with preeclampsia had an elevated serum testosterone level (0.52 +/- 0.13 vs. 0.34 +/- 0.11 ng/mL; p < 0.01). The expression of AR in both placentas was confirmed using immunohistochemistry and was primarily present in the nucleus and cytoplasm of syncytiotrophoblasts and stromal cells. There was a significant increase in AR mRNA expression in PE placentas compared with normal placentas. These data suggest that the increased AR expression may alter AR-mediated function on syncytiotrophoblasts and stromal cells in PE placentas, and be a possible mechanism for its association with preeclampsia.

Expression of Pleiotrophin and its Receptors in Human Placenta Suggests Roles in Trophoblast Life Cycle and Angiogenesis

Pleiotrophin (PTN) is a heparin-binding protein with multiple activities in cell growth, migration and differentiation mediated through multiple receptors. In mammals, PTN expression in trophoblast is found exclusively in the human and in some of the apes in which an endogenous retrovirus upstream of the first coding exon generates a phylogenetically new trophoblast-specific promoter associated with exon UV3. To understand the functions of ERV promoter-mediated trophoblastic PTN expression in pregnancy, we correlated the expression of PTN and its receptors anaplastic lymphoma kinase (ALK), receptor protein tyrosine phosphatase beta/zeta (RPTPbeta/zeta), and Syndecan-1 and Syndecan-3 (SDC1 and SDC3) with key developmental processes in first-trimester human placentation. In an extensive survey of cell lines and primary tissues, we found that trophoblastic transcription of PTN is initiated exclusively from the ERV promoter, whereas decidual expression is initiated at the phylogenetically ancient U1 exon-associated promoter. Using immunohistochemistry, we found that different patterns of overlapping expression of PTN and its receptors occur in different trophoblast subtypes. Notably, a role in angiogenesis is supported by expression of PTN and its receptors in villous mesenchyme, fetal macrophages and villus core fetal vessels. PTN staining of extravillous cytotrophoblasts and the syncytial microvillous membrane is consistent with increasing levels of PTN, as measured by ELISA, in the maternal bloodstream as pregnancy progresses.

Effect of impaired glucose tolerance during pregnancy on the expression of VEGF receptors in human placenta

The aim of the present study was to determine the expression of vascular endothelial growth factor (VEGF) receptors VEGFR-1, VEGFR-2 and VEGFR-3 in placentas from pregnancies complicated by altered glycaemia. Placentas from women with physiological pregnancies (Group 1), pregnancies complicated by minor degree of glucose intolerance (MDGI, Group 2) and by gestational diabetes mellitus (GDM) treated with insulin (Group 3) were collected. Immunohistochemistry, RT-PCR and western blot were employed to evaluate receptor expression. In the three study groups, VEGFR-1 immunoreactivity was detected in all the placental components. VEGFR-2 immunoreactivity was observed in the vessels of all the placentas from Groups 1 and 2, but only in some placentas of Group 3. VEGFR-3 reactivity was observed in all the components of Group 1; in Groups 2 and 3 reactivity was observed in some portions of the trophoblast or the whole trophoblast, and in the stroma. VEGFR-1 and VEGFR-2 mRNA levels in Groups 2 and 3 were significantly higher compared with Group 1, whereas those of VEGFR-3 were significantly lower. Receptor protein levels were significantly lower in Groups 2 and 3 compared with Group 1. These findings demonstrated dysregulation of expression of the three placental receptors, both in GDM and in MDGI.

Histamine H4 receptors in human placenta in diabetes-complicated pregnancy

Histamine H4 receptors in human placenta in diabetes-complicated pregnancy

Complex Expression Pattern of Wnt Ligands and Frizzled Receptors in Human Placenta and its Trophoblast Subtypes

Canonical Wingless (Wnt) signalling provoked by exogenous and endogenous Wnt ligands was recently shown to play a crucial role in the invasive differentiation of human trophoblasts. To gain insights into the expression pattern of the developmental regulators, we analysed all human Wnt ligands and their frizzled (FZD) receptors in the human placenta and different trophoblast model systems using semi-quantitative PCR. Fourteen out of 19 Wnt ligands and 8 out of 10 FZD receptors were detectable in placental tissues, however, expression patterns varied with gestational age and between different trophoblast subtypes suggesting cell-specific functions. Besides Wnt ligands acting through the canonical pathway, non-canonical ligands such as Wnt-5a, which may also activate alternative Wnt signalling pathways or inhibit canonical Wnt signalling, could be identified. Western blot analyses revealed secretion of Wnt-5a from primary trophoblast cultures and trophoblastic cell lines. To evaluate the potential role of Wnt-5a, SGHPL-5 trophoblast cells were transfected with luciferase reporter plasmids harbouring eight T-cell factor (TCF) DNA-recognition sequences which are exclusively activated through the canonical Wnt signalling pathway. Luciferase assays revealed that Wnt-3a-induced reporter activity was repressed by recombinant Wnt-5a indicating an antagonistic role in trophoblasts. The data suggest that a complex network of Wnt ligands and FZD receptors may regulate developmental processes of the human placenta.

Localization and expression of prostaglandin E 2 receptors in human placenta and corresponding fetal membranes with labor

The effects of prostaglandin E2 (PGE2) are mediated through G-protein coupled receptors, acting via different second messengers. The aim of this study was to characterize the temporal and tissue specific localization and expression of the PGE2 receptor subtypes (EP1-4) in uteroplacental tissues during human pregnancy. Placenta and fetal membranes were collected after delivery at preterm or term, each with or without labor. The localization and expression of the PGE2 receptor subtypes were determined by immunohistochemistry and Western blot. All 4 receptors were expressed in the placenta and fetal membranes; only EP3 was present in the syncytiotrophoblast layer. EP1 to EP4 were consistently expressed across gestation in the fetal membranes; however, a different cellular localization with labor was observed in the amnion for EP1, EP2, and EP4. The presence of these receptors in the placenta and fetal membranes may indicate autocrine roles for PGE2 in the signaling pathways associated with placental function and parturition.

Expression and Oxygen Regulation of Endocrine Gland-Derived Vascular Endothelial Growth Factor/Prokineticin-1 and Its Receptors in Human Placenta during Early Pregnancy

Angiogenesis is a key process of dynamic tissue remodeling occurring during placentation. Compelling evidence indicates that vascular endothelial growth factor (VEGF) is an important mediator of placental angiogenesis and appears to be deregulated in preeclampsia. Recently a new angiogenic factor, endocrine gland-derived VEGF (EG-VEGF), also known as prokineticin 1 (PK1), has been identified, and its expression was shown to be restricted to endocrine glands, including the placenta. In this study we investigated the pattern of expression of EG-VEGF, its related factor Bv8/PK2, and their common receptors, PKR1 and PKR2, in human placenta during the first trimester of pregnancy. We also examined EG-VEGF and PKR1 regulation by oxygen tension in isolated trophoblast cells (TCs). Our results show that EG-VEGF, but not Bv8/PK2, is expressed in human placenta. EG-VEGF is mainly localized to the syncytiotrophoblast layer with the highest expression detected between the 8th and 10th wk of gestation. EG-VEGF expression within placental villi is different from that of VEGF, which is mainly localized in the cytotrophoblast and extravillous trophoblast cells. In TCs, PKR1 mRNA is about 80 times more abundant than PKR2 mRNA. Both EG-VEGF and PKR1 mRNAs appear to be regulated by hypoxia. These findings suggest that EG-VEGF has a direct effect on TCs via its receptor PKR1 and is likely to play an important role in human placentation. The expression pattern of EG-VEGF, its regulation by oxygen tension, and its complementary localization to that of VEGF suggest that this new factor might also be deregulated in preeclampsia.

Expression of Hepatocyte Growth Factor/Scatter Factor Receptor in IUGR Fetuses’ Placentas: An Immunohistochemical Analysis

Objective: We aimed to evaluate the expression sta tus of c-met, receptor for the hepatocyte growth factor (HGF), in the placentas of intrauterine growth retardation (IUGR) fetuses. Materials and Methods: Placentas were obtained during delivery from both third trimester normal and IUGR complicated pregnancies. Ultrasonographic estimation of fetal birth weight has been done and placentas of those under the fifth percentile for their gestational ages were enrolled in the study group. Eighteen fetuses with IUGR and 6 uncomplicated pregnancies were subjected to the study. Histological sections from placentas were immunohistochemically evaluated for the expression status of c-met. Results: Seventeen of the eighteen patients (94.4%) in the study group were found to have overexpression of the c-met while this figure was only 16.7% (1/6) in the control group (p < 0.0001). There was a strong statistically significant difference between the two groups regarding degree of c-met expression. Conclusion: Our present findings suggest that c-met is an important cell membrane receptor in human placenta. Deregulation of the interaction between HGF and its receptor c-met during placentation may be the cause responsible for the growth retardation of the fetus due to the impaired placental functions.

Expression of pituitary adenylate cyclase activating polypeptide and its type I receptor mRNAs in human placenta

Pituitary adenylate cyclase activating polypeptide (PACAP) was first isolated from ovine hypothalamus and was known to stimulate the release of growth factor in various cells. Recently, we reported the cellular localization of PACAP and its type I (PAC1) receptor in rat placenta during pregnancy. Placenta is a critical organ that synthesizes several growth factors and angiogenic factors for the fetal development and its own growth. However, there is little information regarding the cellular localization of PACAP and its receptor in human placenta at various gestations. The aim of the present study was to define the expression and distribution of PACAP and PAC1 receptor mRNAs in the human placenta during the pregnancy period. PACAP and PAC1 receptor mRNAs were expressed in stroma cells of stem villi and terminal villi. At the early stage, on 7 and 14 weeks, PACAP and PAC1 receptor genes were moderately expressed in stroma cells surrounding the blood vessels within stem villi. These genes were strongly expressed in stroma cells of stem villi and terminal villi on 24 and 38 weeks. The expression of these genes was increased as gestation advanced, and localized in the same areas. Localization of PACAP and PAC1 receptor demonstrate the evidence that PACAP may play an important role, as an autoregulator or pararegulator via its PAC1 receptor. In conclusion, our findings strongly suggest that PACAP may have a critical role in physiological function of the placenta for gestational maintenance and fetal growth.

Expression of the 5-HT 2A Serotoninergic Receptor in Human Placenta and Choriocarcinoma Cells: Mitogenic Implications of Serotonin

Serotonin (5-hydroxytryptamine, 5-HT) has diverse physiological functions and acts as a mitogen in a variety of cell types, including bovine placental cells. It exerts its mitogenic effect by interacting with a wide range of 5-HT receptor types. Previous studies have reported the presence of 5-HT(2) binding sites in human placental trophoblastic cells, but this has never been confirmed at the molecular level. In this study, we demonstrated that the 5-HT(2A) receptor subtype is fully expressed in the human choriocarcinoma cell lines JEG-3 and BeWo as well as in normal human placental tissue. DNA sequencing has confirmed that the 5-HT(2A) receptor present in these cell lines and tissues is identical to the human 5-HT(2A) receptor found in platelets and in the cerebral cortex. This receptor was localized by immunofluorescence on the plasma membrane, in JEG-3 and BeWo cells. Furthermore, MTT proliferation assays revealed a positive effect of 5-HT on the proliferation of JEG-3 and BeWo cells. These results suggest that 5-HT constitutes a potent mitogen for neoplastic placental cells.

Differential expression of vascular endothelial growth factor (VEGF), endocrine gland derived-VEGF, and VEGF receptors in human placentas from normal and preeclamptic pregnancies.

Vascular endothelial growth factor (VEGF) is a potent regulator of placental vascular function. Endothelial dysfunction is a key factor associated with preeclampsia. In this study, we examined expression of VEGF, endocrine gland-derived VEGF (EG-VEGF), VEGF receptors 1 and 2 (VEGFR-1 and VEGFR-2), and neuropilin-1 and -2 (NP-1 and NP-2) in human placentas from women with normal and preeclamptic (PE) pregnancies using quantitative or semiquantitative PCR. We found that total VEGF mRNA expression was increased 2.8-fold (P < 0.05), along with increases in mRNA expression of VEGF121, 165, and 189 (P < 0.05; 1.7-, 1.9-, and 1.8-fold, respectively) in PE vs. normal placentas. Expression of VEGFR-1 mRNA, but not EG-VEGF and the other three VEGF receptors studied, was elevated (P < 0.05) 2.7-fold in PE vs. normal placentas. Protein expression of VEGF and its four receptors was determined using Western blot analysis. For VEGF, two major isoforms (VEGF165 and 189) were detected. For VEGFR-1, VEGFR-2, NP-1, and NP-2, one major band was observed at 180, 235, 130, and 130 kDa, respectively. All of these bands were corresponding to their positive controls. Of these five proteins studied, only VEGFR-1 levels were increased (P < 0.05; 1.7-fold) in PE placentas. The expression of VEGF and the four VEGF receptors was confirmed using immunohistochemistry. They were primarily present in syncytiotrophoblasts and endothelial cells of villous capillaries and large vessels. Thus, together with previous reports that VEGFR-1 mediates trophoblast function and inhibits VEGF-induced angiogenesis and endothelium-dependent vasodilation, these data suggest that the increased VEGFR-1 expression may alter VEGF- mediated function on trophoblast and endothelial cells in PE placentas.

Localization and variation of TRAIL and its receptors in human placenta during gestation

The localization of TRAIL and its receptors in human placenta was studied under light microscopy using immunohistochemistry method. The variation of TRAIL and its receptors with development was also detected by in situ semi-quantification. The syncytiotrophoblast, cytotrophoblast, stromal cells and the capillary endothelium cells in human placenta all appeared to be TRAIL immunoreactive and the immunoreactive material was distributed on membrane and in cytoplasm with negative nuclei. During whole gestation there was no obvious variation of the staining of TRAIL. Although DR4, DR5, DcR1 and DcR2 can also be detected in the placenta throughout pregnancy, DR4 and DR5 staining increased with development whereas DcR1 and DcR2 staining decreased. Interestingly, at the beginning of the gestation DR4 and DR5 staining distributed on the cytotrophoblast mainly, whereas DcR1 and DcR2 mainly located in the syncytiotrophoblast cells. Collectively, these results suggest that human placenta may not only produce TRAIL but also be a TRAIL target organ, and that TRAIL/TRAILR system could take part in the self-homeostasis of placenta during whole gestation.