Altered differentiation of enteric neural crest-derived cells from endothelin receptor-B null mouse model of Hirschsprung's disease.

Objective: Hirschsprung's disease (HD) is caused by a failure of enteric neural crest-derived cells (ENCC) to colonize the bowel, resulting in an absence of the enteric nervous system (ENS). Previously, we developed a Sox10 transgenic version of the Endothelin receptor-B (Ednrb) mouse to visualize ENCC with the green fluorescent protein, Venus. The aim of this study was to isolate Sox10-Venus+ cells, which are differentiated neurons and glial cells in the ENS, and analyze these cells using Sox10-Venus mice gut.

Methods: The mid-and hindgut of Sox10-Venus+/Ednrb +/+ and Sox10-Venus+/Ednrb -/- at E13.5 and E15.5 were dissected and cells were dissociated. Sox10-Venus+ cells were then isolated. Expression of PGP9.5 and GFAP were evaluated neurospheres using laser scanning microscopy.

Results: 7 days after incubation, Sox10-Venus+ cells colonized the neurosphere. There were no significant differences in PGP9.5 expressions on E13.5 and E15.5. GFAP was significantly increased in HD compared to controls on E15.5 (P < 0.05).

Conclusions: Our results suggest increased glial differentiation causes an imbalance in ENCC lineages, leading to a disruption of normal ENS development in this HD model. Isolation of ENCC provides an opportunity to investigate the ENS with purity and might be a useful tool for modeling cell therapy approaches to HD.