title: A new luciferase-based quantitative assay for the evaluation of human trophoblast fusion.
abstract: The syncytiotrophoblast is a multinuclear cell layer maintained through fusion events with cytotrophoblasts and plays a key role in the properties of the placenta. Monitoring fusion in this cell layer is important in studies aimed at understanding its function. We herein propose a new fusion assay based on the transactivating potential of the human immunodeficiency virus type 1 (HIV-1) Tat protein on its promoter present in the long terminal repeat (LTR) region. We used 2 BeWo cell populations, one stably transfected with the HIV-1 LTR positioned upstream of the luciferase gene and the other stably transfected with a Tat expression vector. Both stable cell lines were responsive to Tat-mediated LTR transactivation and demonstrated normal fusion and human chorionic gonadotropin (hCG) secretion upon stimulation. When both BeWo cell lines were cocultured, forskolin-mediated induction of fusion led to an increase in luciferase activity, which was sensitive to anti-syncytin 1 and -2 antibodies and syncytin 2 small interfering RNAs (siRNAs). Similar results were obtained in primary trophoblasts. Our results highlight the effectiveness and accuracy of this new quantification assay for trophoblast fusion.
authors.affiliation: Département des Sciences Biologiques and Centre de recherche BioMed, Université du Québec à Montréal, Montréal, Québec, Canada.
authors.forename: Amandine
authors.initials: A
authors.lastname: Vargas
authors.forename: Émilie
authors.initials: É
authors.lastname: Larocque
authors.forename: Chirine
authors.initials: C
authors.lastname: Toufaily
authors.forename: Éric
authors.initials: É
authors.lastname: Rassart
authors.forename: Julie
authors.initials: J
authors.lastname: Lafond
authors.forename: Benoit
authors.initials: B
authors.lastname: Barbeau
mesh_terms: D045744:Cell Line, Tumor; D006063:Chorionic Gonadotropin / Q000378:metabolism; D018920:Coculture Techniques; D005576:Colforsin / Q000494:pharmacology; D005260:Female; D016325:HIV Long Terminal Repeat; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism*; D018613:Microscopy, Confocal; D010920:Placenta / Q000166:cytology / Q000502:physiology*; D011247:Pregnancy; D011401:Promoter Regions, Genetic; D014162:Transfection; D014327:Trophoblasts / Q000166:cytology / Q000502:physiology*; D054322:tat Gene Products, Human Immunodeficiency Virus / Q000235:genetics
chemical_list: D006063:Chorionic Gonadotropin; D054322:tat Gene Products, Human Immunodeficiency Virus; D005576:Colforsin; D008156:Luciferases