Human Chorionic Gonadotropin Secretion Research Articles

2-Arachidonoylglycerol impairs human cytotrophoblast cells syncytialization: Influence of endocannabinoid signalling in placental development

A balanced cytotrophoblast cell turnover is crucial for placental development and anomalies in this process associated with gestational diseases. The endocannabinoid system (ECS) has emerged as a new player in several biological processes. However, its influence during placental development is still unknown. We report here the expression of the endocannabinoid 2-arachidonoylglycerol (2-AG) main metabolic enzymes in human cytotrophoblasts and syncytiotrophoblast. We also showed that 2-AG induced a decrease in placental alkaline phosphatase activity, human chorionic gonadotropin secretion and Leptin mRNA levels. Moreover, 2-AG reduced glial cell missing 1 and syncytin-2 transcription and the number of nuclei in syncytium. These effects were mediated by cannabinoid receptors and may result from 2-AG inhibition of the cAMP/PKA signalling pathway. Our data suggest that 2-AG may interfere with the biochemical and morphological differentiation of human cytotrophoblasts, through a CB receptor-dependent mechanism, shedding light on a role for the ECS in placental development.

Disruption of the hormonal network and the enantioselectivity of bifenthrin in trophoblast: maternal-fetal health risk of chiral pesticides.

Endocrine-disrupting chemicals (EDCs) can interfere with normal hormone signaling to increase health risks to the maternal-fetal system, yet few studies have been conducted on the currently used chiral EDCs. This work tested the hypothesis that pyrethroids could enantioselectively interfere with trophoblast cells. Cell viability, hormone secretion, and steroidogenesis gene expression of a widely used pyrethroid, bifenthrin (BF), were evaluated in vitro, and the interactions of BF enantiomers with estrogen receptor (ER) were predicted. At low or noncytotoxic concentrations, both progesterone and human chorionic gonadotropin secretion were induced. The expression levels of progesterone receptor and human leukocyte antigen G genes were significantly stimulated. The key regulators of the hormonal cascade, GnRH type-I and its receptor, were both upregulated. The expression levels of selected steroidogenic genes were also significantly altered. Moreover, a consistent enantioselective interference of hormone signaling was observed, and S-BF had greater effects than R-BF. Using molecular docking, the enantioselective endocrine disruption of BF was predicted to be partially due to enantiospecific ER binding affinity. Thus, BF could act through ER to enantioselectively disturb the hormonal network in trophoblast cells. These converging results suggest that the currently used chiral pesticides are of significant concern with respect to maternal-fetal health.

Intermediate Conductance Ca2+-Activated K+ Channels Modulate Human Placental Trophoblast Syncytialization

Regulation of human placental syncytiotrophoblast renewal by cytotrophoblast migration, aggregation/fusion and differentiation is essential for successful pregnancy. In several tissues, these events are regulated by intermediate conductance Ca2+-activated K+ channels (IKCa), in part through their ability to regulate cell volume. We used cytotrophoblasts in primary culture to test the hypotheses that IKCa participate in the formation of multinucleated syncytiotrophoblast and in syncytiotrophoblast volume homeostasis. Cytotrophoblasts were isolated from normal term placentas and cultured for 66 h. This preparation recreates syncytiotrophoblast formation in vivo, as mononucleate cells (15 h) fuse into multinucleate syncytia (66 h) concomitant with elevated secretion of human chorionic gonadotropin (hCG). Cells were treated with the IKCa inhibitor TRAM-34 (10 µM) or activator DCEBIO (100 µM). Culture medium was collected to measure hCG secretion and cells fixed for immunofluorescence with anti-IKCa and anti-desmoplakin antibodies to assess IKCa expression and multinucleation respectively. K+ channel activity was assessed by measuring 86Rb efflux at 66 h. IKCa immunostaining was evident in nucleus, cytoplasm and surface of mono- and multinucleate cells. DCEBIO increased 86Rb efflux 8.3-fold above control and this was inhibited by TRAM-34 (85%; p<0.0001). Cytotrophoblast multinucleation increased 12-fold (p<0.05) and hCG secretion 20-fold (p<0.05), between 15 and 66 h. Compared to controls, DCEBIO reduced multinucleation by 42% (p<0.05) and hCG secretion by 80% (p<0.05). TRAM-34 alone did not affect cytotrophoblast multinucleation or hCG secretion. Hyposmotic solution increased 86Rb efflux 3.8-fold (p<0.0001). This effect was dependent on extracellular Ca2+, inhibited by TRAM-34 and 100 nM charybdotoxin (85% (p<0.0001) and 43% respectively) but unaffected by 100 nM apamin. In conclusion, IKCa are expressed in cytotrophoblasts and their activation inhibits the formation of multinucleated cells in vitro. IKCa are stimulated by syncytiotrophoblast swelling implicating a role in syncytiotrophoblast volume homeostasis. Inappropriate activation of IKCa in pathophysiological conditions could compromise syncytiotrophoblast turnover and volume homeostasis in pregnancy disease.

Effect of intrauterine injection of human chorionic gonadotropin before embryo transfer on clinical pregnancy rates from in vitro fertilisation cycles: a prospective study

BackgroundThe implantation process after embryo transfer depends on the embryo quality and endometrial receptivity. It is estimated that fifty to seventy-five per cent of pregnancies are lost due to a failure of implantation. There is evidence that there is an early secretion of human chorionic gonadotrophin before embryo implantation, and this secretion has been linked to an important function in angiogenesis and the inflammatory response that promotes the implantation process. Our objective was to determine the effects of intrauterine injection of human chorionic gonadotropin (hCG) before the embryo transfer in an in vitro fertilisation cycle.MethodsA prospective randomised study was conducted in Reproductive Medicine Centre PROCREA in Mexico City. Infertile patients who had a medical indication for in vitro fertilisation were studied. Two groups were included (n 210); the intervention group received an intrauterine injection of 500 IU of hCG before the embryo transfer (n 101). The control group (n 109) did not receive hCG. Comparisons were performed using a chi-square test.ResultsThe clinical pregnancy rate (CPR) was our principal outcome. The implantation rate was a secondary outcome. The implantation rate was significantly higher in the hCG group compared to the control group (52.4% vs 35.7%, p 0.014). The clinical pregnancy rate was also significantly higher (50.4 vs 33.0%, p 0.010). No adverse effects were observed.ConclusionsThe intrauterine injection of hCG before embryo transfer showed a significant increase in the clinical pregnancy rate. More clinical trials are needed to reproduce these results on this promising intervention. The live birth rate must be included in subsequent studies.

Mid-luteal phase injection of subcutaneous leuprolide acetate improves live delivered pregnancy and implantation rates in younger women undergoing in vitro fertilization-embryo transfer (IVF-ET)

Purpose To see if the single injection of one mg of the gonadotropin releasing hormone agonist (GnRHa) leuprolide acetate given in the mid-luteal phase can increase live delivered pregnancy and implantation rates. Furthermore the purpose was to determine if improvement was found, did the mechanism involve increased secretion of human chorionic gonadotropin (hCG). Materials and methods A prospective study was conducted in women aged 35 who were undergoing in vitro fertilization-embryo transfer (IVF-ET). They were advised of data from Tesarik et al. and a previous pilot study conducted in the present IVF center showing improved pregnancy rates with the injection of a GnRHa three days after embryo transfer. They were offered the option of returning for a one-mg injection s.c. of leuprolide acetate or not. Clinical and live delivered pregnancy rates were compared according to those taking or not the leuprolide acetate one-mg injection. Chi-square analysis was used for statistical comparisons. Serum beta-hCG levels were compared between those conceiving with or without the extra injection of leuprolide. Results There was a non-significant trend for higher live delivered pregnancy rates in those taking leuprolide (47.8%, 64/134) vs. those not taking it (38.6%, 76/197). For those pregnant there was no difference in hCG levels according to taking the GnRHa or not. Conclusions The 25% increased live delivered pregnancy rate per transfer was insufficiently powered to detect a significant difference. The results do justify continuing the study. Perhaps the difference could be wider using a slightly older age group whose embryos are frequently less hearty.

Estrogen-Related Receptor γ (ERRγ) Regulates Oxygen-Dependent Expression of Voltage-gated Potassium (K+) Channels and Tissue Kallikrein during Human Trophoblast Differentiation

Estrogen-related receptor γ (ERRγ) serves a critical O2-dependent regulatory role in the differentiation of human cytotrophoblasts to syncytiotrophoblast. In this study, we investigated expression of genes encoding tissue kallikrein (KLK1) and voltage-gated K(+) channels (KV7) during differentiation of human trophoblasts in culture and the roles of ERRγ and O2 tension in their regulation. Expression of KLK1 and the KV7 channel subunits, KCNQ1, KCNE1, KCNE3, and KCNE5, increased during differentiation of cultured human trophoblast cells in a 20% O2 environment. Notably, together with ERRγ, expression of KLK1, KCNQ1, KCNE1, KCNE3, and KCNE5 was markedly reduced when cells were cultured in a hypoxic environment (2% O2). Moreover, upon transduction of trophoblast cells with short hairpin RNAs for endogenous ERRγ, KLK1, KCNQ1, KCNE1, and KCNE3 expression was significantly decreased. Promoter and site-directed mutagenesis studies in transfected cells identified putative ERRγ response elements within the KLK1 and KCNE1 5'-flanking regions required for ERRγ-stimulated transcriptional activity. Binding of endogenous ERRγ to these ERRγ response elements increased during trophoblast differentiation in culture and was inhibited by hypoxia. The KV7 blocker linopirdine reduced human chorionic gonadotropin secretion and aggregation of cultured human trophoblasts, suggesting a possible role of KV7 channels in cell fusion and differentiation. Illumina gene expression arrays of cultured human trophoblast cells revealed several genes upregulated during syncytiotrophoblast differentiation and downregulated upon ERRγ knockdown involved in cell differentiation, adhesion, and synthesis of steroid and peptide hormones required for placental development and function. Collectively, these findings suggest that ERRγ mediates O2-dependent expression of genes involved in human trophoblast differentiation, function, and vascular homeostasis.

The effect of triclosan on hormone secretion and viability of human choriocarcinoma JEG-3 cells

Triclosan is an antimicrobial agent frequently used in pharmaceuticals and personal care products. We analyzed triclosan for its action on placental secretion of progesterone, estradiol and human chorionic gonadotropin in vitro in the JEG-3 cells. We also investigated its action on cell viability, proliferation and apoptosis. The JEG-3 cells were cultured with increasing doses of triclosan (1×10−9–1×10−4M) for 24, 48 and 72h. Triclosan was found to increase estradiol and progesterone secretion after short- and long-term exposure. The stimulatory effect was observed up to 10μM after short- and long-term exposure to triclosan. In addition, triclosan caused an adverse effect on β-hCG secretion. The highest doses of triclosan (50 and 100μM) showed a strong cytotoxic effect. Anti proliferative and pro-apoptotic effects were also observed. Overall, this study demonstrates that triclosan may indirectly disrupt steroidogenesis which may, in turn, affect placental development and consequently fetal growth.

A New Luciferase-Based Quantitative Assay for the Evaluation of Human Trophoblast Fusion

The syncytiotrophoblast is a multinuclear cell layer maintained through fusion events with cytotrophoblasts and plays a key role in the properties of the placenta. Monitoring fusion in this cell layer is important in studies aimed at understanding its function. We herein propose a new fusion assay based on the transactivating potential of the human immunodeficiency virus type 1 (HIV-1) Tat protein on its promoter present in the long terminal repeat (LTR) region. We used 2 BeWo cell populations, one stably transfected with the HIV-1 LTR positioned upstream of the luciferase gene and the other stably transfected with a Tat expression vector. Both stable cell lines were responsive to Tat-mediated LTR transactivation and demonstrated normal fusion and human chorionic gonadotropin (hCG) secretion upon stimulation. When both BeWo cell lines were cocultured, forskolin-mediated induction of fusion led to an increase in luciferase activity, which was sensitive to anti-syncytin 1 and -2 antibodies and syncytin 2 small interfering RNAs (siRNAs). Similar results were obtained in primary trophoblasts. Our results highlight the effectiveness and accuracy of this new quantification assay for trophoblast fusion.

Dysgerminoma and gonadal dysgenesis: The need for a new diagnosis tree for suspected ovarian tumours

Purpose Diagnosis of dysgerminoma in the paediatric age group is uncommon, and most cases arise from dysgenetic gonads of 46, XY pure gonadal dysgenesis (PGD) patients. Bilateral gonadectomy is mandatory in these patients. So, the preoperative diagnosis of PGD is important in order to avoid multiple surgical procedures and delayed patient information in the case of a suspected ‘ovarian’ tumour. Our aim was to discuss preoperative clues that can lead to suspicion of dysgerminoma in the context of PGD. Materials and methods We reviewed the charts of six patients treated for dysgerminoma associated with 46, XY PGD. We focused on particularities of clinical and biological evaluations. Results Median age at diagnosis was 11 years. Pubertal development was absent or incomplete even at late ages. Dysgerminoma was associated with gonadoblastoma foci in all cases. Tumoral marker profile was a normal alfafetoprotein level, a high lactate dehydrogenase level and normal or moderate human chorionic gonadotropin (βHCG) secretion, except for one patient who had a mixed tumour with notably a choriocarcinoma share (high βHCG). Follicle-stimulating hormone (FSH) level was very high in all patients tested and, interestingly, also in one prepubertal patient. Conclusions In the case of a suspected ovarian tumour, delayed pubertal development, moderate βHCG level and elevated FSH level are clinical and biological clues to a diagnosis of dysgerminoma in the context of PGD and should prompt karyotype analysis before surgery. Because FSH is an efficient indirect marker of this condition, we suggest including this analysis in the management of gonadal tumours.

Factors affecting cytotrophoblast cell viability and differentiation: Evidence of a link between syncytialisation and apoptosis

A relationship between cytotrophoblast differentiation (syncytialisation) and apoptosis is hypothesised to exist, but has not been clearly determined. To address this, we explored the effects of cAMP, an inducer of syncytialisation, on human choriocarcinoma cell differentiation and viability under three different culture conditions related to diverse survival status: no serum, 10% fetal calf serum or 10% charcoal-stripped fetal calf serum. 8-Br-cAMP increased BeWo cell viability in culture media without serum, but viability was decreased in a dose- and time-dependent manner when serum was present. The appearance of apoptotic nuclei fragments were only observed when BeWo cells were cultured in media containing serum combined with 8-Br-cAMP treatment. In addition, the ratio of FasL to Fas expression following treatment with 8-Br-cAMP increased by 20-fold in 10% charcoal-stripped fetal calf serum media and 65-fold 10% fetal calf serum media, and activation of caspase-3 also required media with serum. The markers of syncytialisation (syncytin 1 expression and human chorionic gonadotropin secretion) were induced significantly by 8-Br-cAMP, and were higher in 10% fetal calf serum media than in 10% charcoal-stripped fetal calf serum media, than in the absence of serum. Syncytia formation was stimulated by 8-Br-cAMP and this required serum in the media. We now show that factors contained within serum are necessary for cAMP-stimulated cytotrophoblast differentiation, that syncytialisation involves apoptotic events, and that a lack of serum based factors could switch the cellular program away from differentiation.

Interplay of cAMP and MAPK pathways in hCG secretion and fusogenic gene expression in a trophoblast cell line

Differentiation of human placental mononuclear trophoblasts into a multinucleate syncytium involves up-regulation of key proteins promoting cell fusion and increased capacity for placental hormonogenesis. It is well established that the activation of adenylyl cyclase leads to increased expression of trophoblast fusogenic gene machinery and human chorionic gonadotropin (hCG) secretion. We used the forskolin-induced syncytialisation of BeWo choriocarcinoma cells as a model to characterise in detail the signalling pathway downstream of adenylyl cyclase. Forskolin treatment induced a rapid and potent ERK1/2 and p38MAPK phosphorylation; this cascade required PKA–AKAP interactions and led to downstream CREB-1/ATF-1 phosphorylation via ERK1/2-dependent but p38MAPK-independent mechanisms. Interestingly both p38MAPK and ERK1/2 were involved in forskolin-induced hCG-secretion, suggesting the presence of additional p38MAPK-dependent but CREB-1/ATF-1-independent pathways. Forskolin treatment of BeWo cells significantly up-regulated the expression of various fusogenic gene mRNAs, including syncytin-1 and -2 (by 3- and 10-fold, respectively) the transcription factors old astrocyte specifically induced substance (OASIS) and glial cells missing a (GCMa) (by 3- and 6-fold, respectively) and the syncytin-2 receptor, major facilitator superfamily domain containing 2 (MFSD2) (by 2-fold). Up-regulation of AKAP79 and AKAP250 (by 2.5- and 4-fold, respectively) was also identified in forskolin-treated BeWo cells. Forskolin effects on all these genes were suppressed by chemical inhibition of p38MAPK whereas only specific genes were sensitive to ERK1/2 inhibition. This data provide novel insights into the signalling molecules and mechanisms regulating fusogenic gene expression by the adenylyl cyclase pathway.

Dihydrofolate Reductase Transcript Level Is Not Suitable for Methotrexate-Resistance Prediction in Choriocarcinoma Cell Line

To assess whether dihydrofolate reductase (DHFR) expression is correlated with the concentration of methotrexate (MTX) exposure and the resistance index during the establishment of MTX-resistant human choriocarcinoma JeG-3 cell line. Two series of different MTX-resistant JeG-3 sublines were established by intermittent- and consecutive-inducing methods. Quantitative real-time reverse transcription polymerase chain reaction was used to measure DHFR messenger RNA expression in the sublines during different concentrations of MTX exposure. [beta]-Human chorionic gonadotropin ([beta]-HCG) secretion in these cell lines were detected by using chemoluminescence method. In the 2 series of MTX-resistant cell lines, the same dynamic profiling of [beta]-HCG secretion and DHFR transcript level were found: At a certain range of concentration, the upward [beta]-HCG secretion and the messenger RNA level of DHFR were correlated with the dose of MTX exposing. However, once the concentration of MTX exceeded this range, the expression would not increase but decrease sharply. According to the concentration of MTX exposure from low to high, the multiples of [beta]-HCG increases were 1.86-, 5.15-, 8.38-, 64.17-, 2.6-, and 0.78-fold for intermittent induction and 22.05-, 6.12-, 5.80-, and 1.77-fold for continuous induction; the multiples of DHFR gene expression increases were 12.84-, 99.02-, 129.01-, 177.63-, 420.78-, and 86.09-fold for intermittent induction and 9.59-, 20.56-, 96.05-, and 58.72-fold for continuous induction. The transcript level of DHFR gene only correlated with MTX drug resistance at a certain phase. It would be of limited use as a biomarker for the MTX-based chemoresistance prediction.

Differential Triiodothyronine Responsiveness and Transport by Human Cytotrophoblasts from Normal and Growth-Restricted Pregnancies

Context: Abnormal placentation in human pregnancy is associated with intrauterine fetal growth restriction (IUGR). Our group has previously reported the association between severe IUGR, lower fetal circulating concentrations of thyroid hormones (THs), and altered expression of TH receptors and TH transporters within human placental villi. We postulate that altered TH bioavailability to trophoblasts may contribute to the pathogenesis of IUGR.Design and Objective: Cytotrophoblasts were isolated from normal and IUGR human placentae to compare their responsiveness to T3 and their capability for T3 transport.Results: Compared with normal cytotrophoblasts, the viability of IUGR cytotrophoblasts (assessed by methyltetrazoleum assay) was significantly reduced (P < 0.001), whereas apoptosis (assessed using caspase 3/7 activity and M30 immunoreactivity) was significantly increased after T3 treatment for 48 h (P < 0.001 and P < 0.01, respectively). The secretion of human chorionic gonadotropin was significantly increased by IUGR cytotrophoblasts compared with normal cytotrophoblasts (P < 0.001), independently of T3 treatment. Net transport of [125I]T3 was 20% higher by IUGR cytotrophoblasts compared with normal cytotrophoblasts (P < 0.001), and this was accompanied by a 2-fold increase in the protein expression of the TH transporter, monocarboxylate transporter 8, as assessed by Western immunoblotting (P < 0.01).Conclusions: IUGR cytotrophoblasts demonstrate altered responsiveness to T3 with significant effects on cell survival and apoptosis compared with normal cytotrophoblasts. Increased monocarboxylate transporter 8 expression and intracellular T3 accumulation may contribute to the altered T3 responsiveness of IUGR cytotrophoblasts.

Modulation of endocrine and transport functions in human trophoblasts by saquinavir and nelfinavir

Objectives The distribution of drugs to the maternal–fetal interface is influenced by the expression of various efflux transporters. Among these transporters, P-glycoprotein (P-gp) is responsible for the efflux of a great number of drugs such as protease inhibitors of the human immunodeficiency virus, thus reducing the chemical exposure of the fetus. Study design The effects of saquinavir and nelfinavir were evaluated on human trophoblast functions and integrity by investigating their effect on human chorionic gonadotropin (hCG) secretion and on P-gp expression and functionality. Results Nelfinavir significantly reduced hCG secretion by 30% after a 48-h treatment but it had no effect on syncytia formation. Saquinavir had no effect on hCG secretion but significantly increased both expression (to a 2-fold extent) and functionality (by 17.9%) of P-gp, whereas nelfinavir only increased functionality (by 23.1%) with a dissociation of P-gp from caveolin-1. Conclusion These results suggest that the effects of saquinavir and nelfinavir differ on trophoblast functions.

Caspases rather than calpains mediate remodelling of the fodrin skeleton during human placental trophoblast fusion

Fusion of cytotrophoblasts with the overlying syncytiotrophoblast is an integral step in differentiation of the human placental villous trophoblast. Multiple factors, such as growth factors, hormones, cytokines, protein kinases, transcription factors and structural membrane proteins, were described to modulate trophoblast fusion. However, the knowledge on remodelling of the membrane-associated cytoskeleton during trophoblast fusion is very limited. This study describes the link between remodelling of spectrin-like alpha-fodrin and intercellular trophoblast fusion. Experiments with primary trophoblasts isolated from term placentas and the choriocarcinoma cell line BeWo revealed a biphasic strategy of the cells to achieve reorganization of alpha-fodrin. Syncytialization of trophoblasts was accompanied by down-regulation of alpha-fodrin mRNA, whereas the full-length alpha-fodrin protein was cleaved into 120 and 150 kDa fragments. Application of calpeptin and calpain inhibitor III did not affect alpha-fodrin fragmentation in primary term trophoblasts and forskolin-treated BeWo cells, but decreased secretion of beta human chorionic gonadotropin. In contrast, inhibitors of caspases 3, 8 and 9 attenuated generation of the 120 kDa fragment and a general caspase inhibitor completely blocked fragmentation, suggesting an exclusive function of caspases in alpha-fodrin remodelling. Immunofluorescence double staining of human placenta revealed co-localization of active caspase 8 with alpha-fodrin positive vesicles in fusing villous cytotrophoblasts. These results suggest that caspase-dependent fragmentation of alpha-fodrin may be important for reorganization of the sub-membranous cytoskeleton during trophoblast fusion.

Lumbar Metastasis of Choriocarcinoma

A case of lumbar metastasis of a choriocarcinoma is presented. To present and review a rare case of metastatic choriocarcinoma in the lumbar spine. Choriocarcinoma is a highly anaplastic malignancy derived from trophoblastic cells characterized by the secretion of human chorionic gonadotropin (hCG) and early hematogenous metastasis. However, metastatic choriocarcinoma in the spine is extremely rare. Although 2 cases of metastasis in lumbar and/or sacral vertebra have been reported, the efficacy of surgical treatment for the spinal metastasis of choriocarcinoma is not yet known. The clinical course, radiologic features, pathology, and outcome of the treatment of metastatic choriocarcinoma of the lumbar spine is reported. A 38-year-old female patient with abnormal uterine bleeding 6 weeks after a normal-term delivery showed high serum levels of hCG. A whole body image analysis revealed a lesion in the L2 vertebra. After computed tomography-guided needle biopsy, a clinical and pathologic diagnosis of lumbar metastasis of choriocarcinoma was made. Surgical resection of the localized L2 vertebra lesion was performed by total en bloc spondylectomy after a poor response to initial chemotherapy with methotrexate. Postsurgically, the serum level of hCG explosively increased and local recurrences around the original L2 vertebra and epidural metastasis abruptly developed. Lung metastases also occurred concurrently and progressed and the patient eventually died to the disease. We have reported a rare case of lumbar metastasis of choriocarcinoma after a normal-term pregnancy. This is the first report of lumbar metastasis of choriocarcinoma treated by spinal surgery. Because surgical resection of a lumbar metastasis of choriocarcinoma involves a substantial risk of profuse hemorrhage, local recurrence and the spread of metastasis, multiagent chemotherapy in combination with radiotherapy should be preformed before surgical resection.

SHORT COMMUNICATION: Interleukin‐17 Increased Progesterone Secretion by JEG‐3 Human Choriocarcinoma Cells

Problem JEG‐3 choriocarcinoma cell line has previously been reported to express a receptor for interleukin (IL)‐17. The involvement of IL‐17 in the production of progesterone and human chorionic gonadotropin by placental trophoblast has not been investigated.Method of study The present study investigated the in vitro effect of IL‐17 on progesterone and human chorionic gonadotropin (hCG) secretion by JEG‐3 cells. Both hormones were quantified using enzyme‐linked immunosorbent assays.Results The results showed that IL‐17 significantly increased progesterone secretion at 6 (P &lt; 0.001) and 24 (P &lt; 0.01) hr, while this cytokine had no effect on hCG secretion.Conclusion Interleukin‐17 may regulate the function of JEG‐3 cells through increased progesterone secretion.

Does altered oxygenation or reactive oxygen species alter cell turnover of BeWo choriocarcinoma cells?

This study assessed the effect of 20 and 6% ambient oxygen (O 2) or 5–50 μmol/l hydrogen peroxide (H 2O 2) on apoptosis, necrosis, proliferation and fusion of BeWo cells. The expression of p53, Mdm2 and Bax was assessed by western blotting. Apoptosis was increased in cells cultured in 6% O 2 tension and 50 μmol/l H 2O 2 ( P < 0.05, P < 0.01 by ADP:ATP ratio). In the same conditions, cell viability as estimated by the MTT assay was decreased (6% O 2 P < 0.01, 50 μmol/l H 2O 2 P < 0.05). Human chorionic gonadotrophin secretion was decreased by culture in 6%O 2 and 50 μmol/l H 2O 2 ( P < 0.05). Cell fusion was also decreased by treatment with 50 μmol/l H 2O 2 ( P < 0.05). Treatment with 50 μmol/l H 2O 2 was associated with increased expression of p53 and decreased expression of Mdm2 ( P < 0.05). This study provides evidence that BeWo cell turnover is altered following exposure to hypoxia or ROS. It is concluded that BeWo cell culture is an appropriate model for investigating the regulation of trophoblast cell turnover. In addition, these data support a role for p53 in mediating altered trophoblast cell turnover in response to oxidative stress.

Detrimental effects of high levels of antioxidant vitamins C and E on placental function: Considerations for the Vitamins in Preeclampsia (VIP) trial

Supplementation with antioxidant vitamins has been proposed to reduce the risk of preeclampsia and perinatal complications. In a recent study, it has been shown that this supplementation to above physiological doses does not reduce the risk of preeclampsia, but increases the rate of low birthweight babies, suggesting a detrimental effect on placental function, given the lower birthweight. The aim of the present study was to investigate the effects of high levels of antioxidants vitamins C and E on placental cells in vitro. Isolated fresh human cytotrophoblasts were exposed to high concentrations of vitamins C and E for 48 h. Then the secretion of human chorionic gonadotropin (hCG) and the production of tumor necrosis factor-alpha (TNF-alpha) were assessed. High levels of vitamins C and E, separately or combined, decrease the secretion of hCG by cytotrophoblasts and increase their production of TNF-alpha. Exposure of cytotrophoblasts to high levels of antioxidant vitamins C and E may affect placental function, as reflected by decreased secretion of hCG; and placental immunity, as reflected by increased production of TNF-alpha. Such alterations are known to lead to endothelial dysfunction and adverse pregnancy outcomes, such as fetal growth restriction (FGR).

Comparison of cell cycle regulatory gene mRNA in three different types of human trophoblasts and effect of transforming growth factor

Identifying the factors responsible for reducing the proliferation, syncytialization, and invasiveness of trophoblast tissues, as seen with preeclampsia, intrauterine growth restriction, and spontaneous miscarriage, is a current challenge in reproductive biology. These factors, transforming growth factor (TGF)-beta as an example, can work by altering trophoblast differentiation or proliferation. We therefore investigated and compared specific markers of trophoblast proliferation and differentiation in three commonly used trophoblast tissue cell models, and also investigated the influence of TGF-beta on these markers. In this study, we isolated human trophoblasts from first trimester and term placentas, and additionally used human choriocarcinoma cells (JEG-3). Baseline values of human chorionic gonadotropin (hCG) secretion and relative mRNA levels of cell cycle regulators (cyclin E, p21, p27, and p57) were investigated for each cell type. We also investigated the influence of TGF-beta on these parameters. Quantitative and longitudinal production of hCG differed between the three cell types. Significantly different amounts of cyclin E, p21, p27, and p57 mRNA were demonstrated within each cell type, as well as between all the cell types, throughout the culture time period. Each trophoblast type demonstrated a reduction of hCG secretion in response to TGF-beta. TGF-beta did not show a consistent effect on the cell cycle mRNA of any of the cell types. We were able to characterize and compare the differential production of hCG, as well as the differential expression of cell cycle-associated mRNA of early trophoblasts, term trophoblasts, and choriocarcinoma cells. The production of hCG was altered by TGF-beta, although mRNA levels were not markedly altered by TGF-beta.