title: Toll-like receptor profiling of seven trophoblast cell lines warrants caution for translation to primary trophoblasts.
abstract: BACKGROUND
  Excessive placental inflammation is associated with pregnancy complications. Toll-like receptors (TLRs) are sensors for danger signals from infections and damaged tissue and initiate inflammation. Trophoblasts in the placenta broadly express TLRs. Trophoblast cell lines are used as surrogates for primary trophoblasts for in vitro studies, but the inflammatory translatability of trophoblast cell lines warrants examination. We aimed to assess TLR1-10 gene expression and activation in seven trophoblast cell lines and compare this to primary trophoblasts.
  
  
  METHODS
  The five choriocarcinoma trophoblast cell lines BeWo, JAR, JEG-3, AC1M-32 and ACH-3P, and the two SV40 transfected trophoblast cell lines HTR-8/SVneo and SGHPL-5 were included and compared to primary first trimester trophoblasts (n = 6). TLR1-10 gene expression was analyzed by RT-qPCR. Cells were stimulated by specific TLR1-9 ligands for 24 h and cytokine release was measured by a 10-plex immunoassay.
  
  
  RESULTS
  All choriocarcinoma cell lines demonstrated broad TLR gene expression, but lacked functional cytokine response to TLR ligand activation. In contrast, SV40 transfected cell lines showed restricted TLR gene expression, but SGHPL-5 cells displayed significantly increased levels of interleukin (IL)-6, IL-8, IL-12 and vascular endothelial growth factor A after TLR3 and/or TLR4 activation (P < 0.01), while TLR2 activation increased IL-6 and IL-8 levels (P < 0.05). HTR8/SVneo cells responded to TLR3 activation by increased IL-6 and interferon (IFN)-γ (P < 0.05). The SGHPL-5 TLR profile most closely resembled primary trophoblast.
  
  
  CONCLUSIONS
  The characterized trophoblast cell line TLR profiles serve as a reference and warrant caution when selecting trophoblast cell lines as in vitro models for immune responses in primary trophoblasts.
authors.affiliation: Centre of Molecular Inflammation Research, Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.
authors.forename: L M
authors.initials: LM
authors.lastname: Gierman
authors.affiliation: Centre of Molecular Inflammation Research, Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway; St. Olavs Hospital, Trondheim University Hospital, Norway.
authors.forename: G S
authors.initials: GS
authors.lastname: Stødle
authors.affiliation: Centre of Molecular Inflammation Research, Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway; St. Olavs Hospital, Trondheim University Hospital, Norway.
authors.forename: L H
authors.initials: LH
authors.lastname: Tangerås
authors.affiliation: St. Olavs Hospital, Trondheim University Hospital, Norway; Department of Circulation and Medical Imaging, NTNU, Trondheim, Norway.
authors.forename: M
authors.initials: M
authors.lastname: Austdal
authors.affiliation: Centre of Molecular Inflammation Research, Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.
authors.forename: G D
authors.initials: GD
authors.lastname: Olsen
authors.affiliation: Department of Public Health and General Practice, NTNU, Trondheim, Norway.
authors.forename: T
authors.initials: T
authors.lastname: Follestad
authors.affiliation: Centre of Molecular Inflammation Research, Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.
authors.forename: B
authors.initials: B
authors.lastname: Skei
authors.affiliation: Department of Laboratory Medicine, Children's and Women's Health, NTNU, Trondheim, Norway.
authors.forename: K
authors.initials: K
authors.lastname: Rian
authors.affiliation: Centre of Molecular Inflammation Research, Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.
authors.forename: A S
authors.initials: AS
authors.lastname: Gundersen
authors.affiliation: Centre of Molecular Inflammation Research, Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway.
authors.forename: R
authors.initials: R
authors.lastname: Austgulen
authors.affiliation: Centre of Molecular Inflammation Research, Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology (NTNU), Trondheim, Norway. Electronic address: ann-charlotte.iversen@ntnu.no.
authors.forename: A C
authors.initials: AC
authors.lastname: Iversen
mesh_terms: D002460:Cell Line / Q000378:metabolism*; D016207:Cytokines / Q000378:metabolism; D006801:Humans; D051193:Toll-Like Receptors / Q000378:metabolism*; D014327:Trophoblasts / Q000378:metabolism*
chemical_list: D016207:Cytokines; D051193:Toll-Like Receptors
keywords: Cell line; Cytokine; Inflammation; Placenta; Toll-like receptor; Trophoblast