Expression Of Aromatic Hydrocarbon Receptor In Placenta Tissue And Its Effect On Placental Trophoblast Cell Proliferation

Objective:The aryl hydrocarbon receptor (AhR), a ligand-dependent transcription factor,mediates a variety of biological processes, including cell division,apoptosis,cell differentiation,immune system homeostasis, oxidative stress,development and remodeling of vascular, and reproduction. Nothern bloting analysis had confirmed that AhR was expressed in many human tissues, but the highest level of AhR mRNA was found in the placenta. Meanwhile, the AhR had been mainly observed in placental syncytiotrophoblast as well as endothelial cells of blood vessels. Upon binding its ligands, AhR regulates expression of target genes that includes the most important placental biotransformation enzymes cytochrome CYP1A1 and CYP1B1. In this study we examined the AhR protein expression in term human placentas from women with N and mPE pregnancies as well as in first trimester placentas using immunohistochemistry and Western blotting. The levels of AhR,CYP1A1 and CYP1B1 mRNA in first trimester and third trimester placentas were also detected by RT-PCR and real-time PCR. Then we use the human placental choriocarcinoma cell line JEG-3 and JAR as the models to reseach the effects of AhR on the human placental trophoblast proliferation and migration.Materials and Methods:1、Term placentas were obtained immediately after cesarean section delivery from women with N (n=14) and mPE (n=10) pregnancies. The first trimester placentas (n=12) were obtained from women with induced abortion at gestation time of 6-8 weeks.2、The expression of AhR protein in human placentas from women with normal and mild preeclamptic pregnancies, as well as placentas from first trimester by western blotting and immunohistochemistry.3、RT-PCR and real-time PCR analysis for AhR, CYP1A1 and CYP1B1 mRNA in human first trimester and third trimester placentas placentas.4、ITE treatment for JEG-3 and JAR in Culture. The cells were treated with increasing concentrations of ITE (the endogenous ligand for AhR) for 48h and cultured for increasing time with the same concentration of ITE (10uM), respectively.5、RT-PCR analysis for CYP1A1 mRNA in treated and control cells.6、Crystal Violet Staining Solution analysis for reproduction of treated and control cells.7、Flow Cytometry analysis for cell cycles of treated and control cells.8、The 24-Multiwell Insert System analysis for migration of treated and control cells.Results:1、AhR protein was mainly present in the cytoplasm and nuclei of syncytiotrophoblasts, but not in cytotrophoblast cells in first trimester placentas; Moderate AhR expression was observed in capillary endothelial cells within villi in the first trimester, but not term placentas; the distribution patterns of AhR protein in mPE placentas and N plcentas were similar.2、AhR protein expression in mPE placentas was increased as compared with N placentas, while the level was decreased in first trimester placentas as compared with N plcentas.3、AhR, CYP1A1 and CYP1B1 mRNA levels were decreased in first trimester placentas as compared with N plcentas.4、AhR mRNA expression was high in JEG-3, but which was handly expressed in JAR.5、CYP1A1 mRNA expression was activiated by ITE in JEG-3, but not in JAR.6、Reproduction of JEG-3 was inhibited by ITE, but not JAR. Cell cycles were changed by ITE in JEG-3,but not in JAR.7、Migration of JAR was not effected by ITE while JEG-3 hardly migrated.Conclusion:1、AhR protein played an important role in the develepment and growth of placenta, which might involve in promoting and maintaining syncytiotrophoblast differentiation and placental hormonal synthesis, particularly during late pregnancy, to support pregnancy and embryo/fetal development in uteri.2、AhR protein level was higher in mPE placentas than that in N and sPE placentas, suggesting that AhR might play a critical role in syncytiotrophoblasts and pathology of mPE, the latter of which may assist in dissecting and/or predicting the different pathologic processes between mPE and sPE pregnancies.3、AhR played a big role in trophoblast cells. ITE induced S-phase cell cycle arrest and inhibited its reproduction in JEG-3,but not in JAR.