title: Ibuprofen slows migration and inhibits bowel colonization by enteric nervous system precursors in zebrafish, chick and mouse.
abstract: Hirschsprung Disease (HSCR) is a potentially deadly birth defect characterized by the absence of the enteric nervous system (ENS) in distal bowel. Although HSCR has clear genetic causes, no HSCR-associated mutation is 100% penetrant, suggesting gene-gene and gene-environment interactions determine HSCR occurrence. To test the hypothesis that certain medicines might alter HSCR risk we treated zebrafish with medications commonly used during early human pregnancy and discovered that ibuprofen caused HSCR-like absence of enteric neurons in distal bowel. Using fetal CF-1 mouse gut slice cultures, we found that ibuprofen treated enteric neural crest-derived cells (ENCDC) had reduced migration, fewer lamellipodia and lower levels of active RAC1/CDC42. Additionally, inhibiting ROCK, a RHOA effector and known RAC1 antagonist, reversed ibuprofen effects on migrating mouse ENCDC in culture. Ibuprofen also inhibited colonization of Ret+/- mouse bowel by ENCDC in vivo and dramatically reduced bowel colonization by chick ENCDC in culture. Interestingly, ibuprofen did not affect ENCDC migration until after at least three hours of exposure. Furthermore, mice deficient in Ptgs1 (COX 1) and Ptgs2 (COX 2) had normal bowel colonization by ENCDC and normal ENCDC migration in vitro suggesting COX-independent effects. Consistent with selective and strain specific effects on ENCDC, ibuprofen did not affect migration of gut mesenchymal cells, NIH3T3, or WT C57BL/6 ENCDC, and did not affect dorsal root ganglion cell precursor migration in zebrafish. Thus, ibuprofen inhibits ENCDC migration in vitro and bowel colonization by ENCDC in vivo in zebrafish, mouse and chick, but there are cell type and strain specific responses. These data raise concern that ibuprofen may increase Hirschsprung disease risk in some genetically susceptible children.
authors.affiliation: Department of Pediatrics, Washington University School of Medicine, 660 South Euclid Avenue, St. Louis, MO 63110, USA.
authors.forename: Ellen Merrick
authors.initials: EM
authors.lastname: Schill
authors.affiliation: Department of Pediatrics, Washington University School of Medicine, 660 South Euclid Avenue, St. Louis, MO 63110, USA.
authors.forename: Jonathan I
authors.initials: JI
authors.lastname: Lake
authors.affiliation: Department of Pediatrics, Washington University School of Medicine, 660 South Euclid Avenue, St. Louis, MO 63110, USA.
authors.forename: Olga A
authors.initials: OA
authors.lastname: Tusheva
authors.affiliation: Department of Pediatric Surgery, Massachusetts General Hospital, Harvard Medical School, 55 Fruit St., Boston, MA 02114, USA; Department of Human Morphology and Developmental Biology, Faculty of Medicine, Semmelweis University, Budapest, Hungary.
authors.forename: Nandor
authors.initials: N
authors.lastname: Nagy
authors.affiliation: Department of Pediatrics, The Children's Hospital of Philadelphia Research Institute and the Perelman School of Medicine at the University of Pennsylvania, Abramson Research Center, 3615 Civic Center Blvd, Philadelphia, PA 19104, USA.
authors.forename: Saya K
authors.initials: SK
authors.lastname: Bery
authors.affiliation: Department of Internal Medicine, Washington University School of Medicine, 660 South Euclid Avenue, St. Louis, MO 63110, USA.
authors.forename: Lynne
authors.initials: L
authors.lastname: Foster
authors.affiliation: Department of Pediatrics, Washington University School of Medicine, 660 South Euclid Avenue, St. Louis, MO 63110, USA.
authors.forename: Marina
authors.initials: M
authors.lastname: Avetisyan
authors.affiliation: Department of Genetics, Washington University School of Medicine, 660 South Euclid Avenue, St. Louis, MO 63110, USA.
authors.forename: Stephen L
authors.initials: SL
authors.lastname: Johnson
authors.affiliation: Department of Internal Medicine, Washington University School of Medicine, 660 South Euclid Avenue, St. Louis, MO 63110, USA.
authors.forename: William F
authors.initials: WF
authors.lastname: Stenson
authors.affiliation: Department of Pediatric Surgery, Massachusetts General Hospital, Harvard Medical School, 55 Fruit St., Boston, MA 02114, USA.
authors.forename: Allan M
authors.initials: AM
authors.lastname: Goldstein
authors.affiliation: Department of Pediatrics, The Children's Hospital of Philadelphia Research Institute and the Perelman School of Medicine at the University of Pennsylvania, Abramson Research Center, 3615 Civic Center Blvd, Philadelphia, PA 19104, USA. Electronic address: Heuckerothr@email.chop.edu.
authors.forename: Robert O
authors.initials: RO
authors.lastname: Heuckeroth
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D000818:Animals; D053148:Caspase 3 / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects; D002645:Chickens; D051545:Cyclooxygenase 1 / Q000172:deficiency / Q000378:metabolism; D051546:Cyclooxygenase 2 / Q000172:deficiency / Q000378:metabolism; D017615:Enteric Nervous System / Q000166:cytology*; D004789:Enzyme Activation / Q000187:drug effects; D007052:Ibuprofen / Q000494:pharmacology*; D007422:Intestines / Q000166:cytology*; D008565:Membrane Proteins / Q000172:deficiency / Q000378:metabolism; D008648:Mesoderm / Q000166:cytology; D051379:Mice; D008954:Models, Biological; D041681:NIH 3T3 Cells; D058953:Neural Stem Cells / Q000166:cytology* / Q000187:drug effects; D009474:Neurons / Q000166:cytology / Q000187:drug effects; D009924:Organ Culture Techniques; D047495:PPAR gamma / Q000378:metabolism; D011554:Pseudopodia / Q000187:drug effects / Q000378:metabolism; D015027:Zebrafish; D020830:rac1 GTP-Binding Protein / Q000378:metabolism; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D008565:Membrane Proteins; D047495:PPAR gamma; C077623:Ptgs2 protein, mouse; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; C496512:Ptgs1 protein, mouse; D054460:rho-Associated Kinases; D053148:Caspase 3; D020830:rac1 GTP-Binding Protein; D007052:Ibuprofen
keywords: Enteric nervous system development; Gene-environment interactions; Migration