title: Altered gene expression in human placentas after IVF/ICSI.
abstract: OBJECTIVE
  Is gene expression in placental tissue of IVF/ICSI patients altered when compared with a spontaneously conceived group, and are these alterations due to loss of imprinting (LOI) in the case of imprinted genes?
  
  
  CONCLUSIONS
  An altered imprinted gene expression of H19 and Pleckstrin homology-like domain family A member 2 (PHLDA2), which was not due to LOI, was observed in human placentas after IVF/ICSI and several biological pathways were significantly overrepresented and mostly up-regulated.
  
  
  BACKGROUND
  Genomic imprinting plays an important role in placental biology and in placental adaptive responses triggered by external stimuli. Changes in placental development and function can have dramatic effects on the fetus and its ability to cope with the intrauterine environment. An increased frequency of placenta-related problems as well as an adverse perinatal outcome is seen in IVF/ICSI derived pregnancies, but the role of placental epigenetic deregulation is not clear yet.
  
  
  METHODS
  In this prospective cohort study, a total of 115 IVF/ICSI and 138 control couples were included during pregnancy. After applying several exclusion criteria (i.e. preterm birth or stillbirth, no placental samples, pregnancy complications or birth defects), respectively, 81 and 105 placentas from IVF/ICSI and control pregnancies remained for analysis. Saliva samples were collected from both parents.
  
  
  METHODS
  We quantitatively analysed the mRNA expression of several growth-related imprinted genes [H19, insulin-like growth factor 2 (IGF2), PHLDA2, cyclin-dependent kinase inhibitor 1C (CDKN1C), mesoderm-specific transcript homolog (MEST) isoform α and β by quantitative PCR] after standardization against three housekeeping genes [Succinate dehydrogenase A (SDHA), YWHAZ and TATA-binding protein (TBP)]. A quantitative allele-specific expression analysis of the differentially expressed imprinted genes was performed to investigate LOI, independent of the mechanism of imprinting. Furthermore, a microarray analysis was carried out (n = 10 in each group) to investigate the expression of non-imprinted genes as well.
  
  
  RESULTS
  Both H19 and PHLDA2 showed a significant change, respectively, a 1.3-fold (P = 0.033) and 1.5-fold (P = 0.002) increase in mRNA expression in the IVF/ICSI versus control group. However, we found no indication that there is an increased frequency of LOI in IVF/ICSI placental samples. Genome-wide mRNA expression revealed 13 significantly overrepresented biological pathways involved in metabolism, immune response, transmembrane signalling and cell cycle control, which were mostly up-regulated in the IVF/ICSI placental samples.
  
  
  CONCLUSIONS
  Only a subset of samples was found to be fully informative, which unavoidably led to lower sample numbers for our LOI analysis. Our study cannot distinguish whether the reported differences in the IVF/ICSI group are exclusively attributable to the IVF/ICSI technique itself or to the underlying subfertility of the patients.
  
  
  CONCLUSIONS
  Whether these placental adaptations observed in pregnancies conceived by IVF/ICSI might be connected to an adverse perinatal outcome after IVF remains unknown. However, it is possible that these differences affect fetal development and long-term patterns of gene expression, as well as maternal gestational physiology.
  
  
  BACKGROUND
  Partly funded by an unrestricted research grant by Organon BV (now MSD BV) and GROW School for Oncology and Developmental Biology without any role in study design, data collection and analysis or preparation of the manuscript. No conflict of interests to declare.
  
  
  BACKGROUND
  Dutch Trial Registry (NTR) number 1298.
authors.affiliation: Department of Obstetrics and Gynaecology, GROW School for Oncology and Developmental Biology, Maastricht University Medical Centre, Maastricht, The Netherlands.
authors.forename: Ewka C M
authors.initials: EC
authors.lastname: Nelissen
authors.affiliation: Department of Obstetrics and Gynaecology, GROW School for Oncology and Developmental Biology, Maastricht University Medical Centre, Maastricht, The Netherlands.
authors.forename: John C M
authors.initials: JC
authors.lastname: Dumoulin
authors.affiliation: Laboratory for Epigenetics and Environment, CEA-Institut de Génomique, Centre National de Génotypage, Evry, France.
authors.forename: Florence
authors.initials: F
authors.lastname: Busato
authors.affiliation: MNHN CNRS UMR7196, Paris, France INSERM U565, Paris, France.
authors.forename: Loïc
authors.initials: L
authors.lastname: Ponger
authors.affiliation: Department of Bioinformatics-BiGCaT, Maastricht University, Maastricht, The Netherlands.
authors.forename: Lars M
authors.initials: LM
authors.lastname: Eijssen
authors.affiliation: Department of Obstetrics and Gynaecology, GROW School for Oncology and Developmental Biology, Maastricht University Medical Centre, Maastricht, The Netherlands.
authors.forename: Johannes L H
authors.initials: JL
authors.lastname: Evers
authors.affiliation: Laboratory for Epigenetics and Environment, CEA-Institut de Génomique, Centre National de Génotypage, Evry, France.
authors.forename: Jörg
authors.initials: J
authors.lastname: Tost
authors.affiliation: Department of Obstetrics and Gynaecology, GROW School for Oncology and Developmental Biology, Maastricht University Medical Centre, Maastricht, The Netherlands aafke.van.montfoort@mumc.nl.
authors.forename: Aafke P A
authors.initials: AP
authors.lastname: van Montfoort
mesh_terms: D000328:Adult; D050761:Cyclin-Dependent Kinase Inhibitor p57 / Q000235:genetics / Q000378:metabolism; D005260:Female; D005307:Fertilization in Vitro; D015870:Gene Expression; D020869:Gene Expression Profiling; D018392:Genomic Imprinting; D006801:Humans; D007335:Insulin-Like Growth Factor II / Q000235:genetics / Q000378:metabolism; D008297:Male; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D010920:Placenta / Q000378:metabolism*; D011247:Pregnancy; D011446:Prospective Studies
chemical_list: C493964:CDKN1C protein, human; D050761:Cyclin-Dependent Kinase Inhibitor p57; C035170:IGF2 protein, human; D009687:Nuclear Proteins; C109277:TSSC3 protein; D007335:Insulin-Like Growth Factor II
keywords: IVF/ICSI; gene expression; human; imprinting; placenta