Characterisation and transplantation of enteric nervous system progenitor cells.
OBJECTIVE
Enteric nervous system (ENS) progenitor cells have been postulated to be an appropriate source of cells for the treatment of Hirschsprung's disease. In order for this to be successful, the techniques previously used for the isolation of rodent ENS progenitor cells need to be adapted for postnatal human tissue. In this paper, we describe a method suitable for the preparation of both mouse and human postnatal ENS progenitor cells and assess their transplantation potential.


METHODS
Single cell suspensions were isolated from 11.5 days post-coitum embryonic mouse caecum and postnatal human myenteric plexus. These cells were cultured under non-adherent conditions to generate neurospheres which were implanted into aganglionic embryonic mouse hindgut explants. Cell proliferation, migration and differentiation were observed using immunofluorescence microscopy.


RESULTS
Neurospheres generated from both mouse and human tissues contained proliferating neural crest-derived cells that could be expanded in tissue culture to generate both glial cells and neurons. When implanted into aganglionic murine gut, cells migrated from the neurospheres using pathways appropriate for cells derived from the neural crest, and differentiated to become glia and neurons expressing neuronal phenotypic markers characteristic of the ENS including nitric oxide synthase and vasoactive intestinal polypeptide.


CONCLUSIONS
We have developed a technique for the isolation and expansion of ENS progenitor cells from human neonates. These cells have the ability to differentiate into neurons and glia when transplanted into aganglionic gut, this demonstration being a necessary first step for their autologous transplantation in the treatment of Hirschsprung's disease.
Almond
Sarah
S
Institute of Child Health, Royal Liverpool University Children's Hospital, Alder Hey, Liverpool, UK.
Lindley
Richard M
RM
Kenny
Simon E
SE
Connell
M Gwen
MG
Edgar
David H
DH
D000818:Animals; D002432:Cecum / Q000166:cytology; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D002465:Cell Movement; D049109:Cell Proliferation; D017615:Enteric Nervous System / Q000166:cytology*; D006627:Hirschsprung Disease / Q000628:therapy*; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D051379:Mice; D008856:Microscopy, Fluorescence; D009432:Neural Crest / Q000166:cytology* / Q000637:transplantation; D009457:Neuroglia / Q000166:cytology; D009474:Neurons / Q000166:cytology; D033581:Stem Cell Transplantation / Q000379:methods*; D046509:Tissue Culture Techniques; D014183:Transplantation, Heterologous