Unbalanced Placental Expression of Imprinted Genes in Human Intrauterine Growth Restriction

University of Toronto, Toronto, Ontario, Canada
Placenta (Impact Factor: 2.71). 06/2006; 27(6-7):540-9. DOI: 10.1016/j.placenta.2005.07.004
Source: PubMed


Imprinted genes control fetal and placental growth in mice and in rare human syndromes, but the role of these genes in sporadic intrauterine growth restriction (IUGR) is less well-studied. We measured the ratio of mRNA from a maternally expressed imprinted gene, PHLDA2, to that from a paternally expressed imprinted gene, MEST, by Northern blotting in 38 IUGR-associated placentae and 75 non-IUGR placentae and found an increase in the PHLDA2/MEST mRNA ratio in IUGR (p=0.0001). Altered expression of PHLDA2 and MEST was not accompanied by changes in DNA methylation within their imprinting centers, and immunohistochemistry showed PHLDA2 protein appropriately restricted to villous and intermediate cytotrophoblast in the IUGR placentae. We next did a genome-wide survey of mRNA expression in 14 IUGR placentae with maternal vascular under-perfusion compared to 15 non-IUGR placentae using Affymetrix U133A microarrays. In this series six imprinted genes were differentially expressed by ANOVA with a Benjamini-Hochberg false discovery rate of 0.05, with increased expression of PHLDA2 and decreased expression of MEST, MEG3, GATM, GNAS and PLAGL1 in IUGR placentae. At lower significance, we found IGF2 mRNA decreased and CDKN1C mRNA increased in the IUGR cases. We confirmed the significant reduction in MEG3 non-translated RNA in IUGR placentae by Northern blotting. In addition to imprinted genes, the microarray data highlighted non-imprinted genes acting in endocrine signaling (LEP, CRH, HPGD, INHBA), tissue growth (IGF1), immune modulation (INDO, PSG-family genes), oxidative metabolism (GLRX), vascular function (AGTR1, DSCR1) and metabolite transport (SLC-family solute carriers) as differentially expressed in IUGR vs. non-IUGR placentae.

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    • "Placental methylation is significantly lower compared to other somatic tissues242526and this has been associated with promoting fetal development throughout gestation27282930. Placental function and the intrauterine environment play critical roles in fetal program- ming313233, and different lines of evidence suggest a role for epigenetic mechanisms, including genomic imprinting and DNA methylation in this process[34,35]. Several animal models have also suggested that altering placental DNA methylation plays an important role in placental and fetal growth[36]. The aim of the current study was to investigate the genetic and epigenetic changes in placental IGF1/IGF2 and their seven binding proteins in order to understand the " net IGF bioavailability " in pregnancies with small for gestational age (SGA), large for gestational age (LGA) and appropriately grown neonates. "
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    ABSTRACT: Insulin-like growth factors 1 and 2 (IGF1 and IGF2) and their binding proteins (IGFBPs) are expressed in the placenta and known to regulate fetal growth. DNA methylation is an epigenetic mechanism which involves addition of methyl group to a cytosine base in the DNA forming a methylated cytosine-phosphate-guanine (CpG) dinucleotide which is known to silence gene expression. This silences gene expression, potentially altering the expression of IGFs and their binding proteins. This study investigates the relationship between DNA methylation of components of the IGF axis in the placenta and disorders in fetal growth. Placental samples were obtained from cord insertions immediately after delivery from appropriate, small (defined as birthweight <10th percentile for the gestation [SGA]) and macrosomic (defined as birthweight > the 90th percentile for the gestation [LGA]) neonates. Placental DNA methylation, mRNA expression and protein levels of components of the IGF axis were determined by pyrosequencing, rtPCR and Western blotting. In the placenta from small for gestational age (SGA) neonates (n = 16), mRNA and protein levels of IGF1 were lower and of IGFBPs (1, 2, 3, 4 and 7) were higher (p < 0.05) compared to appropriately grown neonates (n = 37). In contrast, in the placenta from large for gestational age (LGA) neonates (n = 20), mRNA and protein levels of IGF1 was not different and those of IGFBPs (1, 2, 3 and 4) were lower (p < 0.05) compared to appropriately grown neonates. Compared to appropriately grown neonates, CpG methylation of the promoter regions of IGF1 was higher in SGA neonates. The CpG methylation of the promoter regions of IGFBP1, IGFBP2, IGFBP3, IGFBP4 and IGFBP7 was lower in the placenta from SGA neonates as compared to appropriately grown neonates, but was unchanged in the placenta from LGA neonates. Our results suggest that changes in CpG methylation contribute to the changes in gene expression of components of the IGF axis in fetal growth disorders. Differential methylation of the IGF1 gene and its binding proteins is likely to play a role in the pathogenesis of SGA neonates.
    Full-text · Article · Dec 2016
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    • "A recent key study showed that PHLDA3, a homologue of PHLDA2, is in fact a P53 regulated repressor of AKT and the loss of the PHLDA3 genomic locus is frequently observed in primary lung cancers, suggesting a role of PHLDA3 in tumor suppression[28]. PHLDA3 is expressed in multiple fetal and adult tissues, while the PHLDA2 gene is expressed in the placenta and fetal liver, indicating that these two genes have nonoverlapping functions in development[22,28]. Since we found that PHLDA2 expression correlates positively with AKT activation in lung cancer we then analyzed the effect of PHLDA2 on AKT activity by modulating PHLDA2 expression in lung cancer cells via overexpression and knockdown studies. "
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    ABSTRACT: Pleckstrin homology-like domain family A member 2 (PHLDA2) is located within the tumor suppressor region of 11p15, and its expression is suppressed in several malignant tumor types. We recently identified PHLDA2 as a robustly induced, novel downstream target of oncogenic EGFR/ErbB2 signaling. In an immunohistochemical study, we find that PHLDA2 protein expression correlates positively with AKT activation in human lung cancers corroborating our data that PHLDA2 is induced upon oncogenic activation and might serve as a biomarker for AKT pathway activation. We show that PHLDA2 overexpression inhibits AKT phosphorylation while decreased PHLDA2 expression increases AKT activity. We further find that PHLDA2 competes with the PH domain of AKT for binding of membrane lipids, thereby directly inhibiting AKT translocation to the cellular membrane and subsequent activation. Indeed, PHLDA2 overexpression suppresses anchorage-independent cell growth and decreased PHLDA2 expression results in increased cell proliferation and reduced sensitivity to targeted agents of EGFR/ErbB2-driven cancers demonstrating functional relevance for this interaction. In summary, our studies demonstrate that PHLDA2 is strongly regulated by EGFR/ErbB2 signaling and inhibits cell proliferation via repressing AKT activation in lung cancers in a negative feedback loop. We highlight a novel action for PHLDA2 as a potential biomarker for AKT pathway activation.
    Full-text · Article · Apr 2015 · Oncotarget
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    • "In mice, hypermethylation in as few as one paternally expressed imprinted gene is sufficient to induce FGR (Murphy et al. 2001, McMinn et al. 2006, Dilworth et al. 2010). Additionally, global methylation changes and altered gene expression in non-imprinted genes have also been identified in FGR placentas (McCarthy et al. 2007, Einstein et al. 2010, Struwe et al. 2010). "
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    ABSTRACT: Fetal growth restriction (FGR) is a major obstetric complication stemming from poor placental development. Previously, we have shown that paternal obesity in mice is associated with impaired embryo development and significantly reduced fetal and placental weights. We hypothesised that the FGR observed in our rodent model of paternal diet-induced obesity is associated with alterations in metabolic, cell signalling and stress pathways. Male C57Bl/6 mice were fed either a normal or high fat diet for 10 weeks prior to sperm collection for IVF and subsequent embryo transfer. On embryonic day 14, placentas were collected and RNA extracted from both male and female placentas to assess mRNA expression of 24 target genes using custom RT-qPCR arrays. Peroxisome proliferator activated receptor alpha (Ppara) and caspase-12 (Casp12) expression were significantly altered in male placentas from obese fathers compared to normal (p<0.05), but not female placentas. PPARA and CASP12 protein was localised within the placenta to trophoblast giant cells by immunohistochemistry, and relative protein abundance was determined by Western Blot analysis. DNA was also extracted from the same placentas to determine methylation status. Global DNA methylation was significantly increased in female placentas from obese fathers compared to normal (p<0.05), but not male placentas. Here we demonstrate for the first time that paternal obesity is associated with changes in gene expression and methylation status of extraembryonic tissue in a sex-specific manner. These findings reinforce the negative consequences of paternal obesity prior to conception, and emphasise the need for more lifestyle advice for prospective fathers.
    Preview · Article · Feb 2015 · Reproduction (Cambridge, England)
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