Gene Expression In Placenta Research Articles

Hyperexpression of Inflammatory Genes in Placental Tissue in Patients with Hemoglobin SC Disease

Introduction: Pregnant women with sickle cell disease (SCD) are at higher risk of obstetrical complications, such as preeclampsia and preterm delivery. The placental circulation is susceptible to vaso-occlusion, which may contribute to fibrosis, necrosis and infarction of the placental villi. Restricted uteroplacental circulation is associated with low birth weight and fetal growth restriction. The mechanisms by which SCD affects placental physiology are largely unknown, and the chronic inflammatory state may be involved in this pathological process. The aim of this study was to evaluate the expression of mediators of inflammatory response in the placenta of pregnant women with SCD.Methods: This is an ongoing case control study that has recruited 4 pregnant patients with hemoglobin SC disease and 4 healthy control pregnant women paired for gestational age and route of delivery. Placental tissue was collected for total RNA extraction and synthesis of cDNA. Samples were run on a qPCR Array (Inflammatory Response and Autoimmunity RT2 Profiler, Qiagen), which includes 84 genes. Gene expression levels were normalized to housekeeping genes (ACTB, B2M, and RPLP0), and will be further validated by RT-qPCR.Results: Two patients underwent cesarean delivery and one patient delivered preterm. The analysis of placental gene expression showed that 16 genes were upregulated. The top 5 most upregulated genes were CCL24 (18.1 fold), TLR9 (8.6 fold), inducible nitric oxide synthase NOS2 (6.8 fold), IL17A (5.4 fold), and CCL16 (4.8 fold). Upregulated genes also included other chemokines and their receptors (CCL21, CCL22, CXCL3, and CCR7), interleukins (IL1RAP, IL5, and IL9), another toll-like receptor (TLR3), transcription factor BCL6, CD40 ligand (CD40LG), and kininogen 1 (KNG1). Interestingly, the two genes found to be downregulated were CRP (C reactive protein) and IFNG (interferon gamma).Conclusion: This is the first study of differential gene expression in placenta of patients with SCD. Despite patients with hemoglobin SC disease being clinically less severe than homozygous sickle cell anemia, we have found that placental tissue in this subset of patients displays increased expression of several genes involved in inflammatory response. Our data support local inflammatory changes in the placenta of these patients that might explain the development of complications in SCD pregnancies. Although the association between obstetric complications and SCD has been long described, our data should contribute to better define the mechanisms affecting the placental physiology that lead to complications, such as preeclampsia. DisclosuresNo relevant conflicts of interest to declare.

Analysis of gene expression and preliminary study of methylation about 11β-HSD2 gene in placentas of Chinese pre-eclampsia patients of Han ethnicity.

The aim of this study was to determine the promoter methylation status of type 2 isoform of 11β-hydroxysteroid dehydrogenase (11β-HSD2) and its regulatory correlation with 11β-HSD2 gene expression in placentas of pre-eclampsia (PE) patients of Chinese Han ethnicity. The pathological features of placental tissues were studied using hematoxylin-eosin staining and immunohistochemical staining. The 11β-HSD2 mRNA and protein expressions were detected by real-time polymerase chain reaction and Western blotting. The methylation of the 11β-HSD2 promoter sequence was examined by bisulfite sequencing polymerase chain reaction. Trophoblast hyperplasia and discontinuous syncytial layer were observed in the PE group, and the 11β-HSD2 was distributed irregularly and its immunoreactivity was weakened distinctly. The expressions of 11β-HSD2 mRNA and protein decreased significantly in the PE group compared with the control group. Unexpectedly, almost no 11β-HSD2 methylation was detected in PE placental tissue (two fragments, 0.6% vs 0%) or normal placental tissue (1% vs 0.6%). No significant difference in 11β-HSD2 promoter methylation was found between the two groups. The 11β-HSD2 expression decreased in PE women of Chinese Han ethnicity, but was not interrelated with the promoter methylation status.

Placental infection by Salmonella enterica typhimurium in a murine model: mechanisms of pathogenesis and role of inflammatory cell death

Placental infection by Salmonella enterica typhimurium in a murine model: mechanisms of pathogenesis and role of inflammatory cell death

High-salt intake during pregnancy modifies placenta phenotype at least until the third generation

High-salt intake during pregnancy modifies placenta phenotype at least until the third generation

Sex-specific epigenetic impact of preconceptional maternal weight loss on foeto-placental development

Sex-specific epigenetic impact of preconceptional maternal weight loss on foeto-placental development

Methylation status of human chorionic gonadotropin beta subunit promoter and TFAP2A expression as factors regulating CGB gene expression in placenta

To evaluate mechanisms regulating the expression of CGB genes in placental tissues from uncomplicated pregnancies and chorionic samples from spontaneous miscarriages. Molecular analyses in human samples. Laboratory of molecular biology. Nine placental samples from term deliveries and 21 chorionic samples from miscarriages at 7-13weeks of gestation. None. The expression level of CGBs and genes encoding SP1, SP3, and AP2 transcription factors was analyzed using quantitative polymerase chain reaction (qPCR). The methylation status of the CGBs' promoter regions was determined using methylation-specific PCR. The experiments showed significant differences in CGBs' expression and their regulation between placental and chorionic tissues. In placental tissues and chorionic tissues from 7 to 9weeks of gestation, the expression level of CGBs was shown to be associated with the amount of TFAP2A transcripts. It was also demonstrated that variation in the expression level of CGB genes relies on changes in methylation of CGB3-9 and CGB1-2 promoter sequences. During pregnancy, regulation of hCG beta subunit genes expression correlates with both methylation of their promoters and TFAP2A expression level. The results suggest that these factors may be very influential in the early stages of pregnancy and may be associated with pregnancy outcome.

Na+/I− Symporter and Type 3 Iodothyronine Deiodinase Gene Expression in Amniotic Membrane and Placenta and Its Relationship to Maternal Thyroid Hormones

Placental type 3 iodothyronine deiodinase (D3) potentially protects the fetus from the elevated maternal thyroid hormones. Na(+)/I(-) symporter (NIS) is a plasma membrane glycoprotein, which mediates active iodide uptake. Our objectives were to establish the distribution of NIS and D3 gene expressions in the placenta and the amniotic membrane and to investigate the relationship between placental D3 and NIS gene expressions and maternal iodine, selenium, and thyroid hormone status. Thyroid hormones, urinary iodine concentration (UIC), and selenium levels were measured in 49 healthy term pregnant women. NIS and D3 gene expressions were studied with the total mRNA RT-PCR method in tissues from maternal placenta (n = 49), fetal placenta (n = 9), and amniotic membrane (n = 9). NIS and D3 gene expressions were shown in the fetal and maternal sides of the placenta and amniotic membrane. Mean blood selenium level was 66 ± 26.5 μg/l, and median UIC was 143 μg/l. We could not demonstrate any statistically significant relationship of spot UIC and blood selenium with NIS and D3 expression (p > 0.05). Positive correlations were found between NIS and thyroxine-binding globulin (TBG) (r = 0.3, p = 0.042) and between D3 and preoperative glucose levels (r = 0.4, p = 0.006). D3 and NIS genes are expressed in term placenta and amniotic membrane; thus, in addition to placenta, amniotic membrane contributes to regulation of maternofetal iodine and thyroid hormone transmission. Further studies are needed to clarify the relationship between maternal glucose levels and placental D3 expression and between TBG and placental NIS expression.

Widespread DNA hypomethylation at gene enhancer regions in placentas associated with early-onset pre-eclampsia.

Pre-eclampsia is a serious complication of pregnancy that can affect both maternal and fetal outcomes. Early-onset pre-eclampsia (EOPET) is a severe form of pre-eclampsia that is associated with altered physiological characteristics and gene expression in the placenta. DNA methylation is a relatively stable epigenetic modification to DNA that can reflect gene expression, and can provide insight into the mechanisms underlying such expression changes. This case–control study focused on DNA methylation and gene expression of whole chorionic villi samples from 20 EOPET placentas and 20 gestational age-matched controls from pre-term births. DNA methylation was also assessed in placentas affected by late-onset pre-eclampsia (LOPET) and normotensive intrauterine growth restriction (nIUGR). The Illumina HumanMethylation450 BeadChip was used to assess DNA methylation at >480 000 cytosine-guanine dinucleotide (CpG) sites. The Illumina HT-12v4 Expression BeadChip was used to assess gene expression of >45 000 transcripts in a subset of cases and controls. DNA methylation analysis by pyrosequencing was used to follow-up the initial findings in four genes with a larger cohort of cases and controls, including nIUGR and LOPET placentas. Bioinformatic analysis was used to identify overrepresentation of gene ontology categories and transcription factor binding motifs. We identified 38 840 CpG sites with significant (false discovery rate <0.01) DNA methylation alterations in EOPET, of which 282 had >12.5% methylation difference compared with the controls. Significant sites were enriched at the enhancers and low CpG density regions of the associated genes and the majority (74.5%) of these sites were hypomethylated in EOPET. EOPET, but not associated clinical features, such as intrauterine growth restriction (IUGR), presented a distinct DNA methylation profile. CpG sites from four genes relevant to pre-eclampsia (INHBA, BHLHE40, SLC2A1 and ADAM12) showed different extent of changes in LOPET and nIUGR. Genome-wide expression in a subset of samples showed that some of the gene expression changes were negatively correlated with DNA methylation changes, particularly for genes that are responsible for angiogenesis (such as EPAS1 and FLT1). Results could be confounded by altered cell populations in abnormal placentas. Larger sample sizes are needed to fully address the possibility of sub-profiles of methylation within the EOPET cohort. Based on DNA methylation profiling, we conclude that there are widespread DNA methylation alterations in EOPET that may be associated with changes in placental function. This property may provide a useful tool for early screening of such placentas. This study identifies DNA methylation changes at many loci previously reported to have altered gene expression in EOPET placentas, as well as in novel biologically relevant genes we confirmed to be differentially expressed. These results may be useful for DNA- methylation-based non-invasive prenatal diagnosis of at-risk pregnancies.

Abnormal regulation for progesterone production in placenta with prenatal cocaine exposure in rats

Cocaine abuse in pregnant women is currently a significant public hygiene problem and is tightly associated with elevated risk for preterm delivery. Placental steroidogenesis especially progesterone production was essential for success and maintenance of pregnancy in humans and rodents. In the present study, we determined the impact of prenatal cocaine exposure on pathways of placental progesterone synthesis in rats. Pregnant rats were treated cocaine twice daily (15 mg/kg/day) during the third trimester, and the maternal and fetal plasma progesterone and pregnenolone concentrations were detected. We also examined both the protein and mRNA expression of some key enzymes and regulators for progesterone production in placenta. Results showed that, after maternal cocaine use during pregnancy, progesterone and pregnenolone concentrations in both maternal and fetal rats were significantly decreased. Although prenatal cocaine exposure had no effects on placental 3β-hydroxysteroid dehydrogenase type 1 (3βHSD1) expression, protein and mRNA expression of the cholesterol side-chain cleavage enzyme (P450scc/CYP11a) in placenta was significantly inhibited. Moreover, protein and mRNA expressions of MLN64 that regulating cholesterol transport and activating protein 2γ (AP2γ/Tfap2c) that controlling P450scc/CYP11a gene expression in placenta were both decreased following maternal cocaine use in pregnancy. Collectively, this study suggested that prenatal cocaine exposure could insult the placental progesterone production in rats possibly associated with the high risk for preterm delivery.

Aberrant placenta gene expression pattern in bovine pregnancies established after transfer of cloned or in vitro produced embryos

In the present study, we used the global transcriptome profile approach to identify dysregulated genes, molecular pathways, and molecular functional alterations in bovine placentas derived from somatic cell nuclear transfer (SCNT) and in vitro embryo production (IVP) pregnancies compared with their artificial insemination (AI) counterparts at day 50 of gestation. For this, day 7 blastocysts derived from AI, IVP, or SCNT were transferred to oestrus-synchronized cows. The pregnant animals were slaughtered at day 50 of gestation, and the placentas were then recovered and used for transcriptome analysis using Affymetrix GeneChip bovine genome array. Results showed the SCNT placenta to be different from its AI counterpart in the expression of 1,196 transcripts. These genes were found to be associated with alterations in key biological processes and molecular pathways in SCNT placenta, and the dysregulation of 9% (n = 110) of these genes was due to transcriptional reprogramming error. IVP placenta also displayed alterations in the expression of 72 genes, of which 58 were common to SCNT placenta. Gene enrichment analysis revealed that the expression of genes involved in organ development, blood vessel development, extracellular matrix organization, and the immune system was affected in both SCNT and IVP placentas. However, 96% of the affected genes in SCNT were not significantly altered in IVP groups. Thus, the higher transcriptome dysregulation in SCNT placenta followed by IVP would reflect the degree of placental abnormality in SCNT and IVP pregnancies at day 50 of the gestation, which may have a profound effect on subsequent fetal development and health of the offspring.

The Alternative Epac/cAMP Pathway and the MAPK Pathway Mediate hCG Induction of Leptin in Placental Cells

Pleiotropic effects of leptin have been identified in reproduction and pregnancy, particularly in the placenta, where it works as an autocrine hormone. In this work, we demonstrated that human chorionic gonadotropin (hCG) added to JEG-3 cell line or to placental explants induces endogenous leptin expression. We also found that hCG increased cAMP intracellular levels in BeWo cells in a dose-dependent manner, stimulated cAMP response element (CRE) activity and the cotransfection with an expression plasmid of a dominant negative mutant of CREB caused a significant inhibition of hCG stimulation of leptin promoter activity. These results demonstrate that hCG indeed activates cAMP/PKA pathway, and that this pathway is involved in leptin expression. Nevertheless, we found leptin induction by hCG is dependent on cAMP levels. Treatment with (Bu)2cAMP in combination with low and non stimulatory hCG concentrations led to an increase in leptin expression, whereas stimulatory concentrations showed the opposite effect. We found that specific PKA inhibition by H89 caused a significant increase of hCG leptin induction, suggesting that probably high cAMP levels might inhibit hCG effect. It was found that hCG enhancement of leptin mRNA expression involved the MAPK pathway. In this work, we demonstrated that hCG leptin induction through the MAPK signaling pathway is inhibited by PKA. We observed that ERK1/2 phosphorylation increased when hCG treatment was combined with H89. In view of these results, the involvement of the alternative cAMP/Epac signaling pathway was studied. We observed that a cAMP analogue that specifically activates Epac (CPT-OMe) stimulated leptin expression by hCG. In addition, the overexpression of Epac and Rap1 proteins increased leptin promoter activity and enhanced hCG. In conclusion, we provide evidence suggesting that hCG induction of leptin gene expression in placenta is mediated not only by activation of the MAPK signaling pathway but also by the alternative cAMP/Epac signaling pathway.

PAGE4 Positivity Is Associated with Attenuated AR Signaling and Predicts Patient Survival in Hormone-Naive Prostate Cancer

Aberrant activation of the androgen receptor (AR) plays a key role during prostate cancer (PCa) development and progression to castration-resistant prostate cancer (CR-PCa) after androgen deprivation therapy, the mainstay systemic treatment for PCa. New strategies to abrogate AR activity and biomarkers that predict aggressive tumor behavior are essential for improved therapeutic intervention. PCa tissue microarrays herein reveal that prostate-associated gene 4 (PAGE4), an X-linked cancer/testis antigen, is highly up-regulated in the epithelium of preneoplastic lesions compared with benign epithelium, but subsequently decreases with tumor progression. We show that AR signaling is attenuated in PAGE4-expressing cells both in vitro and in vivo, most likely via impaired androgen-induced AR nuclear translocation and subsequently reduced AR protein stabilization and phosphorylation at serines 81 and 213. Consistently, epithelial PAGE4 protein levels inversely correlated with AR activation status in hormone-naive and CR-PCa clinical specimens. Moreover, PAGE4 impaired the development of CR-PCa xenografts, and strong PAGE4 immunoreactivity independently predicted favorable patient survival in hormone-naive PCa. Collectively, these data suggest that dysregulation of epithelial PAGE4 modulates AR signaling, thereby promoting progression to advanced lethal PCa and highlight the potential value of PAGE4 as a prognostic and therapeutic target.

Expression of angiogenesis‐related factors and inflammatory cytokines in placenta and umbilical vessels in pregnancies with preeclampsia and chorioamnionitis/funisitis

We hypothesized that gene expression in placenta and umbilical vessels are affected by intrauterine environment and some of the expression in umbilical vessels originating from the fetus could reflect fetal condition of these complicated pregnancies. Expression of angiogenesis-related factors and inflammatory cytokines were examined in placenta and umbilical vessels to clarify the effects of intrauterine environment of pregnancies complicated by preeclampsia and chorioamnionitis/funisitis. Forty-six preterm cesarean section deliveries were classified into three groups based on maternal condition during prenatal monitoring: preeclampsia (PE) (n = 11), chorioamnionitis/funisitis (CAM) (n = 8), and preterm control (PC) (n = 27). Angiogenesis-related factors and inflammatory cytokines in placentas, umbilical arteries and umbilical veins were analyzed by RT-PCR and immunohistochemistry. We demonstrated that Ang-2, Tie-2, and Dll4 increase in the placentas of PE compared to PC for the first time, and we confirmed the findings of previous reports showing the high expression of HIF-1α, sFlt-1, endoglin, leptin, and AT1R. Expression of angiogenesis-related factors, including HIF-1α, VEGF, angiopoietin, and TGF-β systems, and inflammatory cytokines, such as TNF-α and IL-6, increased in umbilical vessels of PE. Umbilical veins of CAM showed a higher Dll4 level than did PC. In preeclampsia, abnormal expressions of angiogenesis-related factors related to lifestyle diseases in adulthood were seen in the placenta and umbilical vessels as compared to PC. Chorioamnionitis/funisitis showed only upregulation of DII4 in umbilical veins.

Differential Expression of HrtA1 and ADAM12 in Placentas from Preeclamptic and Normotensive Pregnancies.

High temperature requirement factor A 1 (HtrA1) and a disintegrin and metalloproteinase 12 (ADAM12), which play roles in placental implantation and placental growth, have been implicated in the pathogenesis of preeclampsia. We investigated relative mRNA expression of both genes in placental tissues from women with preeclampsia (N=18) (average gestational age 36 weeks) and an equal number of women with normotensive pregnancies (average gestational age 39 weeks). Real-time polymerase chain reaction was used to measure mRNA extracted from term placental biopsies. Differential gene expression was evaluated using Student's T-test and fold change analyses. Statistically significant increases in placental HtRA1 (1.69-fold, p=0.030) and ADAM12 (1.48-fold, p=0.010) mRNA expression were observed among preeclamptic cases as compared with normotensive controls. HtrA1 expression was correlated with maternal age (p-value <0.01) among preeclampsia cases. Increases in HtRA1 and ADAM12 placental gene expression in placentas from preeclamptic pregnancies are consistent with some earlier reports of altered serum protein concentrations in preeclamptic pregnancies. This adds to the literature suggesting that defects in placentation (e.g. involving trophoblast invasion) are of etiologic importance in preeclampsia.

Expression profiling of vitamin D receptor in placenta, decidua and ovary of pregnant mice

ObjectivesThe presence of vitamin D receptor (VDR) and the identification of localized vitamin D3 synthesis in placenta and decidua implicate the importance of vitamin D3 in reproductive function. There is, however, no data on the expression profile of VDR in the mouse placenta and endometrium throughout the pregnancy period. Study designIn the present work expression of VDR in reproductive tissues of pregnant mice at different gestational phases has been addressed. Expression of VDR was determined by semi-quantitative RT-PCR, Western blotting and immunohistochemistry. ResultsThe results showed that VDR mRNA and protein were expressed in decidua, placenta and ovary throughout the pregnancy. VDR gene expression in placenta was significantly elevated in late pregnancy when compared to that of mid pregnancy. Additionally, VDR expression level in decidua rose significantly as pregnancy progressed from early to mid stages. VDR expression in decidua of pregnant mice was higher in comparison to endometrium of non-pregnant mice. Immunohistochemical analysis revealed that VDR protein is consistently expressed by luminal and glandular epithelial cells of decidua, giant cells, glycogen rich cells and labyrinth cells of placenta and by almost all follicular cell types of ovary. Surveying the expression of VDR at the protein level by Western blotting confirmed PCR results. ConclusionIt seems that expression of VDR in reproductive organs is finely tuned during pregnancy indicating its eminent role in reproductive biology.

Dual-specificity phosphatase 23 mediates GCM1 dephosphorylation and activation

Glial cells missing homolog 1 (GCM1) is a transcription factor essential for placental development. GCM1 promotes syncytiotrophoblast formation and placental vasculogenesis by activating fusogenic and proangiogenic gene expression in placenta. GCM1 activity is regulated by multiple post-translational modifications. The cAMP/PKA-signaling pathway promotes CBP-mediated GCM1 acetylation and stabilizes GCM1, whereas hypoxia-induced GSK-3β-mediated phosphorylation of Ser322 causes GCM1 ubiquitination and degradation. How and whether complex modifications of GCM1 are coordinated is not known. Here we show that the interaction of GCM1 and dual-specificity phosphatase 23 (DUSP23) is enhanced by PKA-dependent phosphorylation of GCM1 on Ser269 and Ser275. The recruitment of DUSP23 reverses GSK-3β-mediated Ser322 phosphorylation, which in turn promotes GCM1 acetylation, stabilization and activation. Supporting a central role in coordinating GCM1 modifications, knockdown of DUSP23 suppressed GCM1 target gene expression and placental cell fusion. Our study identifies DUSP23 as a novel factor that promotes placental cell fusion and reveals a complex regulation of GCM1 activity by coordinated phosphorylation, dephosphorylation and acetylation.

Placental Gene Expression in a Rat ‘Model’ of Placental Insufficiency

Objective Placental insufficiency is a major factor associated with pregnancy complications such as miscarriages, intrauterine growth restriction and pre-eclampsia. Recent studies have identified the Brown Norway (BN) rat as a natural ‘model’ of placental insufficiency associated with decreased trophoblast remodeling of maternal uterine arteries. Hypothesis Genetic pathways involved in angiogenesis and immune cell regulation are dysregulated in the placenta of BN rats. Methods Global gene expression in placentas from BN rats were compared with that from Sprague–Dawley (SD) controls at 17.5 days of gestation using the Affimetrix Rat 1.0 microarray chip, and results confirmed with real-time PCR and immunoblotting. Results We found significant differences in 272 genes with 108 being up-regulated and 164 down-regulated in BN placentas compared to SD placentas. BN placentas overexpressed genes involved in the renin-angiotensin system (RAS) such as Ace, Ace2, Agtr1a, Nox4, and Ephx2, while key genes involved in angiogenesis, such as Mmp1, Mmp10, Fgfbp1, Esr1, Itga2, Rgs5, and Ccnb1 were down-regulated. We also observed increased expression of Timd2, Itm2a, Irak3, and Csf1r, and decreased expression of Slpi, Ncam1, and Igsf3 in BN placentas. In addition, we observed lower placental weights in BN males compared to BN females, together with increased expression of Cyp1a1 in BN males, as compared to BN females. Conclusions The present study demonstrates differential expression of genes involved in blood pressure, angiogenesis and immune cell regulation in BN placenta, and suggests that the RAS may be involved in the pathogenesis of placental dysfunction observed in BN rats.

GCM1 Regulation of the Expression of Syncytin 2 and Its Cognate Receptor MFSD2A in Human Placenta1

Syncytin 2 is a newly identified placental membrane protein with fusogenic and immunosuppressive activities. Major facilitator superfamily domain containing 2A (MFSD2A) is the cognate receptor for syncytin 2-mediated cell-cell fusion. Both syncytin 2 and MFSD2A are highly expressed in placenta. In this study to understand the regulation of syncytin 2 and MFSD2A expression in placenta, we found that syncytin 2 gene is epigenetically silenced in nonplacental cells by cytosine-phosphate-guanine (CpG) dinucleotide methylation and that expression of syncytin 2 and MFSD2A genes are regulated by the placental transcription factor GCM1 in placental cells. Functional GCM1-binding sites were identified in syncytin 2 and MFSD2A promoters based on electrophoretic mobility shift assay and chromatin immunoprecipitation assay. Because GCM1 activity is decreased in hypoxic placental cells, we further confirmed that expression of MFSD2A is downregulated in hypoxic BeWo choriocarcinoma cells. Interestingly, ectopic expression of GCM1 activated syncytin 2 and MFSD2A expression in MCF-7 breast cancer cells and facilitated MCF-7 cell fusion. The expression of syncytin 2 in MCF-7 cells was partly attributed to CpG demethylation in the syncytin 2 promoter in the presence of GCM1. Our results suggest that GCM1 is a critical factor in controlling placental cell fusion through transcriptional regulation of syncytin 2 and MFSD2A gene expression in placenta. In addition, GCM1 may also play an important role in the epigenetic regulation of syncytin 2 gene expression.

Prenatal Phenol Exposure and Expression of Imprinted Genes in Human Placenta

ISEE-0086 Background and Objective: Many phenols are hormonally active and are thought to impact human development. Alteration of imprinted gene expression in placenta may be one underlying mechanism. Methods: We measured 8 phenol metabolites in urine and amniotic fluid from 11 women enrolled in a prospective pregnancy cohort for assessing toxicant exposures in women undergoing amniocentesis. We examined the relation between analyte concentrations in urine adjusted for specific gravity and expression of imprinted genes in placentas. Results: 2,5-Dichlorophenol (25-DCP), methyl paraben (MPB), and ethyl paraben (EPB) were detected in all urine samples while triclosan was detected in 10 of 11 samples. Neither 25-DCP nor EPB were detected in amniotic fluid. Phenol metabolites most frequently detected in amniotic fluid were MPB and benzophenone-3 (BP-3). We observed no correlation between amniotic fluid and urinary concentrations of MPB; however, amniotic fluid and urine concentrations of BP-3 were highly correlated (R-square >0.7). Bisphenol-A (BPA) was detected in 3 of 3 urine samples from women who delivered large for gestational age babies as compared to 1 of 8 samples for average sized babies. The most biologically active analytes appear to be BP-3, BPA, and 25-DCP; associations between biomarker level and gene expression levels were observed for 20, 17, and 14 imprinted genes, respectively. The gene most sensitive to phenol exposure appears to be HYMA1, with higher levels of 5 biomarkers. BMPR2, WT1, MKRN3, and CTNNA3 were each associated with higher levels of 4 biomarkers. Conclusion: Biomarkers of phenol exposure were detectable in prenatal urine and amniotic fluid. Continued exploration of the effect of phenols on human development is warranted given the high rate of detection in this vulnerable population combined with the observed potential for dysregulating placental gene expression.

Different promoter usage for CYP19 gene expression in buffalo ovary and placenta

Aromatase is the key enzyme for estrogen biosynthesis and is coded by CYP19 gene. The expression of CYP19 gene is regulated in tissue-specific manner by alternate use of different promoters. In this study, we have analyzed tissue-specific expression and their regulation of CYP19 gene in preovulatory and postovulatory stages of buffalo ( Bubalus bubalis) ovary and placenta. RT-PCR analysis showed that the CYP19 gene expression was significantly ( p < 0.05) higher in granulosa cells of large follicles as compared to the other tissues. The transcript analysis and transcriptional start site identified by 5′-RLM RACE for CYP19 expression indicated different transcriptional start sites within the different sized follicles during folliculogenesis. Sequence analysis showed that the transcription start site of transcript isolated from buffalo granulosa cells of small follicles was 37 bases upstream of the transcript isolated from granulosa cells of large follicles. However, both the transcripts were found to be derived from proximal promoter II. Difference in the transcriptional start site indicates the different promoter sequence usage in granulosa cells of different sized follicles. Further, in silico analysis of the difference in promoter sequence based on the 5′-UTRs isolated from granulosa cells of small follicles (151 bases) and large follicles (114 bases) showed that consensus sequence for certain important trans-elements (viz., TATA binding protein, E2F and CAAT binding protein) would lie in the promoter sequence isolated from the granulosa cells of large follicles. These transcription factors may be involved in regulation of CYP19 gene expression in ovary, either directly or indirectly. The difference in the size of 5′-UTR among the granulosa cells of ovary reflects the possible mechanism for the differential regulation of CYP19 gene during development. The transcripts isolated from buffalo corpus luteum and placental cotyledons were having same 5′-UTR comprises of 168 bases and found to be derived from PI.1. Estimates of CYP19 gene transcript concentration in the different tissues revealed that the CYP19 gene expression in granulosa cells is predominantly regulated by PII and to a minor extent by PI.1. However, PI.1 is almost exclusively responsible for CYP19 gene expression in placenta and residual expression in corpus luteum. In order to understand the complex CYP19 gene regulation in these tissues, further studies are needed to elucidate the activity of different promoters and define regulatory elements for binding of transcription factors.