ABSTRACT
Cardiac neural crest cells migrate into the pharyngeal arches where they support development of the pharyngeal arch arteries. The pharyngeal endoderm and ectoderm both express high levels of FGF8. We hypothesized that FGF8 is chemotactic for cardiac crest cells. To begin testing this hypothesis, cardiac crest was explanted for migration assays under various conditions. Cardiac neural crest cells migrated more in response to FGF8. Single cell tracing indicated that this was not due to proliferation and subsequent transwell assays showed that the cells migrate toward an FGF8 source. The migratory response was mediated by FGF receptors (FGFR) 1 and 3 and MAPK/ERK intracellular signaling. To test whether FGF8 is chemokinetic and/or chemotactic in vivo, dominant negative FGFR1 was electroporated into the premigratory cardiac neural crest. Cells expressing the dominant negative receptor migrated slower than normal cardiac neural crest cells and were prone to remain in the vicinity of the neural tube and die. Treating with the FGFR1 inhibitor, SU5402 or an FGFR3 function-blocking antibody also slowed neural crest migration. FGF8 over-signaling enhanced neural crest migration. Neural crest cells migrated to an FGF8-soaked bead placed dorsal to the pharynx. Finally, an FGF8 producing plasmid was electroporated into an ectopic site in the ventral pharyngeal endoderm. The FGF8 producing cells attracted a thick layer of mesenchymal cells. DiI labeling of the neural crest as well as quail-to-chick neural crest chimeras showed that neural crest cells migrated to and around the ectopic site of FGF8 expression. These results showing that FGF8 is chemotactic and chemokinetic for cardiac neural crest adds another dimension to understanding the relationship of FGF8 and cardiac neural crest in cardiovascular defects.
Subject(s)
Cell Movement/drug effects , Chemotaxis/drug effects , Fibroblast Growth Factor 8/pharmacology , Neural Crest/cytology , Animals , Apoptosis/drug effects , Butadienes/pharmacology , Cell Proliferation/drug effects , Chick Embryo , Dose-Response Relationship, Drug , Enzyme Inhibitors/pharmacology , Fibroblast Growth Factor 8/genetics , Fibroblast Growth Factor 8/metabolism , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Heart/embryology , Immunohistochemistry , In Situ Hybridization , Mesoderm/embryology , Mesoderm/metabolism , Mitogen-Activated Protein Kinases/antagonists & inhibitors , Mitogen-Activated Protein Kinases/metabolism , Myocardium/cytology , Myocardium/metabolism , Neural Crest/embryology , Neural Crest/metabolism , Nitriles/pharmacology , Pharynx/embryology , Pharynx/metabolism , Pyrroles/pharmacology , Receptor, Fibroblast Growth Factor, Type 1/antagonists & inhibitors , Receptor, Fibroblast Growth Factor, Type 1/genetics , Receptor, Fibroblast Growth Factor, Type 1/metabolism , Signal TransductionABSTRACT
Cardiac neural crest cells represent a unique subpopulation of cranial neural crest cells that are specified, delaminate and migrate from the developing neural tube to the caudal pharynx where they support aortic arch artery development. From the caudal pharynx, a subset of these cells migrates into the cardiac outflow tract where they are needed for outflow septation. Many signaling factors are known to be involved in specifying and triggering the migration of neural crest cells. These factors have not been specifically studied in cardiac crest but are assumed to be the same as for the other regions of crest. Signaling factors like Ephs and Semaphorins guide the cells into the caudal pharynx. Support of the cells in the pharynx is from endothelin, PDGF and the TGFbeta/BMP signaling pathways. Mutants in the TGFbeta/BMP pathway show abnormal migration or survival in the pharynx, whereas the migration of the neural crest cells into the outflow tract is orchestrated by Semaphorin/Plexin signaling. Although TGFbeta family members have been well studied and show defective neural crest function in outflow septation, their mechanism of action remains unclear.