title: A functional CFTR assay using primary cystic fibrosis intestinal organoids.
abstract: We recently established conditions allowing for long-term expansion of epithelial organoids from intestine, recapitulating essential features of the in vivo tissue architecture. Here we apply this technology to study primary intestinal organoids of people suffering from cystic fibrosis, a disease caused by mutations in CFTR, encoding cystic fibrosis transmembrane conductance regulator. Forskolin induces rapid swelling of organoids derived from healthy controls or wild-type mice, but this effect is strongly reduced in organoids of subjects with cystic fibrosis or in mice carrying the Cftr F508del mutation and is absent in Cftr-deficient organoids. This pattern is phenocopied by CFTR-specific inhibitors. Forskolin-induced swelling of in vitro-expanded human control and cystic fibrosis organoids corresponds quantitatively with forskolin-induced anion currents in freshly excised ex vivo rectal biopsies. Function of the CFTR F508del mutant protein is restored by incubation at low temperature, as well as by CFTR-restoring compounds. This relatively simple and robust assay will facilitate diagnosis, functional studies, drug development and personalized medicine approaches in cystic fibrosis.
authors.affiliation: Department of Pediatric Pulmonology, Wilhelmina Children's Hospital, University Medical Center, Utrecht, The Netherlands.
authors.forename: Johanna F
authors.initials: JF
authors.lastname: Dekkers
authors.forename: Caroline L
authors.initials: CL
authors.lastname: Wiegerinck
authors.forename: Hugo R
authors.initials: HR
authors.lastname: de Jonge
authors.forename: Inez
authors.initials: I
authors.lastname: Bronsveld
authors.forename: Hettie M
authors.initials: HM
authors.lastname: Janssens
authors.forename: Karin M
authors.initials: KM
authors.lastname: de Winter-de Groot
authors.forename: Arianne M
authors.initials: AM
authors.lastname: Brandsma
authors.forename: Nienke W M
authors.initials: NW
authors.lastname: de Jong
authors.forename: Marcel J C
authors.initials: MJ
authors.lastname: Bijvelds
authors.forename: Bob J
authors.initials: BJ
authors.lastname: Scholte
authors.forename: Edward E S
authors.initials: EE
authors.lastname: Nieuwenhuis
authors.forename: Stieneke
authors.initials: S
authors.lastname: van den Brink
authors.forename: Hans
authors.initials: H
authors.lastname: Clevers
authors.forename: Cornelis K
authors.initials: CK
authors.lastname: van der Ent
authors.forename: Sabine
authors.initials: S
authors.lastname: Middendorp
authors.forename: Jeffrey M
authors.initials: JM
authors.lastname: Beekman
mesh_terms: D000818:Animals; D001681:Biological Assay / Q000379:methods; D002478:Cells, Cultured; D003550:Cystic Fibrosis / Q000175:diagnosis / Q000235:genetics / Q000473:pathology*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D005260:Female; D057809:HEK293 Cells; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism / Q000473:pathology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018181:Mice, Inbred CFTR; D050505:Mutant Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D009940:Organoids / Q000378:metabolism / Q000473:pathology*; D061251:Primary Cell Culture / Q000379:methods; D014867:Water / Q000378:metabolism
chemical_list: D050505:Mutant Proteins; D014867:Water; D019005:Cystic Fibrosis Transmembrane Conductance Regulator