Localization and endothelin-3 dependence of stem cells of the enteric nervous system in the embryonic colon.
OBJECTIVE
The aganglionosis in a variable length of the distal gut found in Hirschsprung's disease results from the abnormal prenatal development of neural crest-derived stem cells of the enteric nervous system. The cytokine endothelin-3 is necessary for successful colonization of the distal gut, but the location of this interaction with neural crest-derived stem cells remains to be established. The hypothesis tested here is that the stem cells of the enteric nervous system (ENS) in the colon are located at the leading edge of the migrating wave of neural crest-derived stem cells and that these cells require colonic endothelin-3 for complete colonization of the gut.


METHODS
Explants of 11.5-day-old embryonic intact mouse gut and isolated colon were cultured for 72 hours in the presence and absence of the endothelin-B receptor antagonist, BQ788. Specimens then were sectioned and stained by immunohistochemistry to assess enteric nervous system development.


RESULTS
Isolated colon contained a very low number (mean, 73 cells; range, 37 to 106; n = 8) of neural crest-derived stem cells, which had just entered its proximal end at the leading edge of neural crest cell migration. After 72 hours of culture, progeny of these few neural crest-derived stem cells had colonized the colon at an equivalent ganglionic density to those in intact gut. Furthermore, neuronal differentiation, as shown by the appearance of nitric oxide synthase positive neurons, also was equivalent to intact gut. Blockade of the endothelin-B receptor produced terminal aganglionosis in both isolated colons and intact gut.


CONCLUSIONS
The very small number of cells that first enter the proximal colon at the leading edge of neural crest cell migration have the ability to colonize the entire colon normally in an ET-3-dependent manner. These cells therefore have the functional characteristics expected of the stem cells of the colonic enteric nervous system. Furthermore, the normal development of these cells is dependent on the endothelin-3 expressed by the mesenchymal cells of the colon itself.
Sidebotham
Emma L
EL
Liverpool, England.
Woodward
Mark N
MN
Kenny
Simon E
SE
Lloyd
David A
DA
Vaillant
Camille R
CR
Edgar
David H
DH
D000818:Animals; D002465:Cell Movement / Q000187:drug effects; D002478:Cells, Cultured; D003106:Colon / Q000166:cytology / Q000196:embryology / Q000294:innervation*; D004195:Disease Models, Animal; D019334:Endothelin-3 / Q000032:analysis / Q000502:physiology*; D017615:Enteric Nervous System / Q000166:cytology / Q000196:embryology* / Q000502:physiology; D006627:Hirschsprung Disease / Q000196:embryology / Q000503:physiopathology; D051379:Mice; D009432:Neural Crest / Q000166:cytology* / Q000196:embryology*; D009842:Oligopeptides / Q000494:pharmacology; D010880:Piperidines / Q000494:pharmacology; D013234:Stem Cells / Q000737:chemistry / Q000166:cytology* / Q000187:drug effects
D019334:Endothelin-3; D009842:Oligopeptides; D010880:Piperidines; C086539:BQ 788