title: Human fetal and adult epicardial-derived cells: a novel model to study their activation.
abstract: The epicardium, a cell layer covering the heart, plays an important role during cardiogenesis providing cardiovascular cell types and instructive signals, but becomes quiescent during adulthood. Upon cardiac injury the epicardium is activated, which includes induction of a developmental gene program, epithelial-to-mesenchymal transition (EMT) and migration. However, the response of the adult epicardium is suboptimal compared to the active contribution of the fetal epicardium to heart development. To understand the therapeutic value of epicardial-derived cells (EPDCs), a direct comparison of fetal and adult sources is paramount. Such analysis has been hampered by the lack of appropriate culture systems.
  
  
  
  Human fetal and adult EPDCs were isolated from cardiac specimens obtained after informed consent. EPDCs were cultured in the presence of an inhibitor of the TGFβ receptor ALK5. EMT was induced by stimulation with 1 ng/ml TGFβ. PCR, immunofluorescent staining, scratch assay, tube formation assay and RT2-PCR for human EMT genes were performed to functionally characterize and compare fetal and adult EPDCs.
  
  
  
  In this study, a novel protocol is presented that allows efficient isolation of human EPDCs from fetal and adult heart tissue. In vitro, EPDCs maintain epithelial characteristics and undergo EMT upon TGFβ stimulation. Although similar in several aspects, we observed important differences between fetal and adult EPDCs. Fetal and adult cells display equal migration abilities in their epithelial state. However, while TGFβ stimulation enhanced adult EPDC migration, it resulted in a reduced migration in fetal EPDCs. Matrigel assays revealed the ability of adult EPDCs to form tube-like structures, which was absent in fetal cells. Furthermore, we observed that fetal cells progress through EMT faster and undergo spontaneous EMT when TGFβ signaling is not suppressed, indicating that fetal EPDCs more rapidly respond to environmental changes.
  
  
  
  Our data suggest that fetal and adult EPDCs are in a different state of activation and that their phenotypic plasticity is determined by this activation state. This culture system allows us to establish the cues that determine epicardial activation, behavior, and plasticity and thereby optimize the adult response post-injury.
authors.affiliation: Department of Molecular Cell Biology, Leiden University Medical Center, P.O Box 9600, Postzone S-1-P, 2300RC, Leiden, The Netherlands.
authors.forename: Asja T
authors.initials: AT
authors.lastname: Moerkamp
authors.affiliation: Department of Molecular Cell Biology, Leiden University Medical Center, P.O Box 9600, Postzone S-1-P, 2300RC, Leiden, The Netherlands.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Lodder
authors.affiliation: Department of Molecular Cell Biology, Leiden University Medical Center, P.O Box 9600, Postzone S-1-P, 2300RC, Leiden, The Netherlands.
authors.forename: Tessa
authors.initials: T
authors.lastname: van Herwaarden
authors.affiliation: Department of Molecular Cell Biology, Leiden University Medical Center, P.O Box 9600, Postzone S-1-P, 2300RC, Leiden, The Netherlands.
authors.forename: Esther
authors.initials: E
authors.lastname: Dronkers
authors.affiliation: Department of Molecular Cell Biology, Leiden University Medical Center, P.O Box 9600, Postzone S-1-P, 2300RC, Leiden, The Netherlands.
authors.forename: Calinda K E
authors.initials: CK
authors.lastname: Dingenouts
authors.affiliation: Department of Molecular Cell Biology, Leiden University Medical Center, P.O Box 9600, Postzone S-1-P, 2300RC, Leiden, The Netherlands.
authors.forename: Fredrik C
authors.initials: FC
authors.lastname: Tengström
authors.affiliation: Department of Cardiothoracic Surgery, Leiden University Medical Center, P.O Box 9600, Postzone S-1-P, 2300RC, Leiden, The Netherlands.
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: van Brakel
authors.affiliation: Department of Molecular Cell Biology, Leiden University Medical Center, P.O Box 9600, Postzone S-1-P, 2300RC, Leiden, The Netherlands. M.J.Goumans@lumc.nl.
authors.forename: Marie-José
authors.initials: MJ
authors.lastname: Goumans
authors.affiliation: Department of Molecular Cell Biology, Leiden University Medical Center, P.O Box 9600, Postzone S-1-P, 2300RC, Leiden, The Netherlands. A.M.Smits@lumc.nl.
authors.forename: Anke M
authors.initials: AM
authors.lastname: Smits
mesh_terms: D002465:Cell Movement / Q000502:physiology; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D004338:Drug Combinations; D058750:Epithelial-Mesenchymal Transition / Q000502:physiology; D005333:Fetus / Q000166:cytology* / Q000378:metabolism; D006321:Heart / Q000502:physiology; D006801:Humans; D007797:Laminin / Q000378:metabolism; D038081:Organogenesis / Q000502:physiology; D010496:Pericardium / Q000166:cytology* / Q000378:metabolism; D011509:Proteoglycans / Q000378:metabolism; D018125:Receptors, Transforming Growth Factor beta / Q000378:metabolism
chemical_list: D004338:Drug Combinations; D007797:Laminin; D011509:Proteoglycans; D018125:Receptors, Transforming Growth Factor beta; C059485:matrigel; D003094:Collagen
keywords: Cardiac development; Epicardium; Epithelial-to-mesenchymal transition; Human EPDCs; In vitro model; TGFβ