Project description:Introduction: In human placenta, alteration in trophoblast differentiation has a major impact on placental maintenance and integrity. Moreover, abnormal syncytial fusion seems to be implicated in the development of many complications including pre-eclampsia and intra-uterine growth restriction (IUGR). However, little is known about the mechanisms that control cytotrophoblast fusion into syncytiotrophoblast. BeWo cell line is commonly used to study placental function since it can be induced to form syncytium and secretes hormones. This fusion is triggered by cAMP or forskolin treatments. In contrast, JEG-3 cell line fails to undergo substantial fusion. Therefore, we will use BeWo and JEG-3 cells to identify a set of genes responsible of trophoblast fusion. Methods: BeWo and JEG-3 cells were treated with forskolin for 48 to induce fusion. RNA was extracted, hybridized to Affymetrix HuGene ST1.0 arrays and analyzed using system biology. Trophoblast differentiation was evaluated by real time PCR and immunocytochemistry analysis. Moreover, some of the identified genes were validated by real time PCR. Results: Our results identified a list of 32 altered genes in fused BeWo cells compared to JEG-3 cells after forskolin treatment. Among these genes, 4 were validated by real-time PCR including salt inducible Kinase 1 (SIK1) gene which is specifically upregulated in BeWo cells upon fusion. Moreover, using system biology analysis, SIK1 showed to be at the center of many biological and functional processes which suggest that it might play a major role in trophoblast differentiation. Conclusion: This study identified new target genes implicated in trophoblast fusion; a process implicated in the etiology of many pregnancy-related diseases such as preeclampsia, IUGR, and gestational diabetes.
Project description:Trophoblast fusion is a central step in placental physiology. This mechanisms can be studied in the BeWO cell model that fuse following forskolin treatment that activates the c-AMP cascade.
Project description:BeWo trophoblast cells differentiate in response to expsure to cyclic adenosine monophosphate (cAMP) analogs. Differentiation includes syncytialization (fusion) and hormonogenesis. The goal of this study was to globally determine transcripts differentially expressed in BeWo trophoblast cells following a 24-h exposure to 250 uM 8-bromo-cAMP. 3 replicates undifferentiated BeWo trophoblast cells; and 3 replicates BeWo trophoblast cells treated with 250 uM 8-Br-cAMP for 24 h.
Project description:Invasive extravillous trophoblasts (EVTs) of the human placenta are critically involved in successful pregnancy outcome since they remodel the uterine spiral arteries to increase blood flow and oxygen delivery to the placenta and the developing fetus. To gain more insights into their biological role different primary cell culture models are commonly utilised. However, access to early placental tissue may be limited and primary trophoblasts rapidly cease proliferation in vitro impairing genetic manipulation. Hence, trophoblastic cell lines have been widely used as surrogates to study EVT function. Although the cell lines share some molecular marker expression with their primary counterpart, it is unknown to what extent they recapture the invasive phenotype of EVT. Therefore, we here report the first thorough GeneChip analyses of SGHPL-5, HTR-8/SVneo, BeWo, JEG-3 and the novel ACH-3P trophoblast cells in comparison to previously analysed primary villous cytrophoblasts and extravillous trophoblasts. To identify EVT-specific gene expression signatures in trophoblast cell lines, we calcuted differentially expressed genes between pre-defined groups based on the distinct origins of the five trophoblast cell lines under investigation. Comparison 1 comprised EVT, HTR-8/Svneo and SGHPL-5 vs choriocarcinoma cells (ACH-3P, BeWo, JEG-3). Comparison 2 comprised EVT, ACH-3P, BeWo, JEG-3 vs extravillous trophoblast cell lines (HTR-8/SVneo, SGHPL-5).
Project description:BeWo trophoblast cells differentiate in response to expsure to cyclic adenosine monophosphate (cAMP) analogs. Differentiation includes syncytialization (fusion) and hormonogenesis. The goal of this study was to globally determine transcripts differentially expressed in BeWo trophoblast cells following a 24-h exposure to 250 uM 8-bromo-cAMP.
Project description:Q fever is a zoonosis caused by Coxiella burnetii, an obligate intracellular bacterium usually found in myeloid cells. The infection is a source of severe obstetrical complications in humans and cattle, and of chronic evolution in pregnant women. As C. burnetii is found in the placenta of aborted foetuses in humans and ruminants, we wondered if it may infect trophoblasts. In this work, we showed that C. burnetii, infected JEG trophoblastic cells without replication and was localized within phagolysosomes. We analyzed gene expression programs induced by C. burnetii in JEG trophoblastic cell line and compared it with transcriptomic program of BeWo trophoblasts in which C. burnetii replicates. These transcriptomic programs induced by C. burnetii in JEG trophoblasts was poor and markedly different from that induced by C. burnetii in BeWo trophoblasts. Hence, the differences in transcriptomic programs may explain the different intracellular fate of C. burnetii in JEG and BeWo cells. Our results suggest that C. burnetii may use trophoblastic cells as a reservoir by interfering with gene expression. Comparaison between unstimulated JEG cell line and Coxiella burnetii stimulated JEG cell line (bacterial ratio 200:1) for 6 hours
Project description:Q fever is a zoonosis caused by Coxiella burnetii, an obligate intracellular bacterium usually found in myeloid cells. The infection is a source of severe obstetrical complications in humans and cattle, and of chronic evolution in pregnant women. As C. burnetii is found in the placenta of aborted foetuses in humans and ruminants, we wondered if it may infect trophoblasts. In this work, we showed that C. burnetii, infected JEG trophoblastic cells without replication and was localized within phagolysosomes. We analyzed gene expression programs induced by C. burnetii in JEG trophoblastic cell line and compared it with transcriptomic program of BeWo trophoblasts in which C. burnetii replicates. These transcriptomic programs induced by C. burnetii in JEG trophoblasts was poor and markedly different from that induced by C. burnetii in BeWo trophoblasts. Hence, the differences in transcriptomic programs may explain the different intracellular fate of C. burnetii in JEG and BeWo cells. Our results suggest that C. burnetii may use trophoblastic cells as a reservoir by interfering with gene expression.
Project description:We studied morphological and transcriptional changes of JEG-3 cells, a trophoblast model, using a hypoxia microchip and human leiomyoma-derived matrix Myogel that mimics human tumor microenvironment.
Project description:Transcriptional profiling comparison of human primary villous trophoblast and extra-villous trophoblast (VT and EVT respectively) cells, isolated from the placenta at 8 to 12 weeks gestation, with complimentary transcriptional profiling of choriocarcinoma cell lines JEG-3 and JAR. Based on phenotypic markers, JEG is frequently used as a model for EVT and JAR is used as a model for VT.
Project description:Invasive extravillous trophoblasts (EVTs) of the human placenta are critically involved in successful pregnancy outcome since they remodel the uterine spiral arteries to increase blood flow and oxygen delivery to the placenta and the developing fetus. To gain more insights into their biological role different primary cell culture models are commonly utilised. However, access to early placental tissue may be limited and primary trophoblasts rapidly cease proliferation in vitro impairing genetic manipulation. Hence, trophoblastic cell lines have been widely used as surrogates to study EVT function. Although the cell lines share some molecular marker expression with their primary counterpart, it is unknown to what extent they recapture the invasive phenotype of EVT. Therefore, we here report the first thorough GeneChip analyses of SGHPL-5, HTR-8/SVneo, BeWo, JEG-3 and the novel ACH-3P trophoblast cells in comparison to previously analysed primary villous cytrophoblasts and extravillous trophoblasts.