title: Cervical trophoblasts for non-invasive single-cell genotyping and prenatal diagnosis.
abstract: OBJECTIVE
  We aimed at developing a method to recover trophoblastic cells from the cervix through a completely non-invasive approach and obtaining a genetic proof of their fetal nature implying that they can be used for non-invasive prenatal diagnosis (NIPD).
  
  
  METHODS
  We studied obstetrical samples from 21 pregnant women between 8 and 12 weeks of gestation scheduled for chorionic villus sampling or undergoing elective termination of pregnancy. A cytobrush was used to extract cells from the external parts of the cervix and transferred to 10 ml of preservative solution. Cells were layered on filters with 8 microns pores using the ISET system (Isolation by SizE of Tumor/Trophoblastic cells) and stained. Putative fetal cells were collected by single cell laser-assisted microdissection and identified as fetal or maternal cells by Short Tandem Repeat genotyping. NIPD was blindly performed on 6 mothers at risk of having a fetus with Cystic Fibrosis or Spinal Muscular Atrophy.
  
  
  RESULTS
  Trophoblastic cells were recovered from all tested cervical samples with a frequency of 2-12 trophoblasts per 2 ml. NIPD was blindly obtained and verified in 6 mothers at risk of having a fetus with Cystic Fibrosis or Spinal Muscular Atrophy.
  
  
  CONCLUSIONS
  Although larger confirmation studies are required, this is the first report providing a solid proof of principle that trophoblasts can be consistently and safely recovered from cervical samples. Since they are a source of pure fetal DNA, i.e. fetal DNA not mixed with maternal DNA, they constitute an ideal target to develop NIPD of recessive diseases, which is a technical challenge for methods based on cell free DNA.
authors.affiliation: INSERM Unit 1151 - Team 13, Paris Descartes University, Paris, France.
authors.forename: I
authors.initials: I
authors.lastname: Pfeifer
authors.affiliation: Department of Gyncology, Antoine Béclère Hospital, Clamart, France.
authors.forename: A
authors.initials: A
authors.lastname: Benachi
authors.affiliation: INSERM Unit 1151 - Team 13, Paris Descartes University, Paris, France.
authors.forename: A
authors.initials: A
authors.lastname: Saker
authors.affiliation: Laboratory of Medical Genetics, Necker-Enfants Malades Hospital, Paris, France.
authors.forename: J P
authors.initials: JP
authors.lastname: Bonnefont
authors.affiliation: INSERM Unit 1151 - Team 13, Paris Descartes University, Paris, France.
authors.forename: H
authors.initials: H
authors.lastname: Mouawia
authors.affiliation: INSERM Unit 1151 - Team 13, Paris Descartes University, Paris, France.
authors.forename: L
authors.initials: L
authors.lastname: Broncy
authors.affiliation: Department of Reproduction, Foch Hospital, Suresnes, France.
authors.forename: R
authors.initials: R
authors.lastname: Frydman
authors.affiliation: Department of Maternity, Les Lilas, France.
authors.forename: M L
authors.initials: ML
authors.lastname: Brival
authors.affiliation: Laboratoiry of Biochemistry A, Necker-Enfants Malades Hospital, Paris, France.
authors.forename: B
authors.initials: B
authors.lastname: Lacour
authors.affiliation: Alfred Fournier Institute, Paris, France.
authors.forename: R
authors.initials: R
authors.lastname: Dachez
authors.affiliation: INSERM Unit 1151 - Team 13, Paris Descartes University, Paris, France; Laboratoiry of Biochemistry A, Necker-Enfants Malades Hospital, Paris, France. Electronic address: patriziapaterlini@gmail.com.
authors.forename: P
authors.initials: P
authors.lastname: Paterlini-Bréchot
mesh_terms: D002584:Cervix Uteri / Q000166:cytology*; D015193:Chorionic Villi Sampling / Q000379:methods; D003550:Cystic Fibrosis / Q000175:diagnosis / Q000235:genetics; D005260:Female; D005820:Genetic Testing / Q000379:methods; D060005:Genotyping Techniques / Q000379:methods*; D006801:Humans; D008297:Male; D009134:Muscular Atrophy, Spinal / Q000175:diagnosis / Q000235:genetics; D011247:Pregnancy; D011261:Pregnancy Trimester, First; D011296:Prenatal Diagnosis / Q000379:methods*; D059010:Single-Cell Analysis / Q000379:methods*; D014327:Trophoblasts / Q000166:cytology*
keywords: Cystic fibrosis; ISET(®); Non-invasive prenatal diagnosis; Spinal muscular atrophy; Trophoblasts