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.
Department of Pediatric Surgery, Juntendo University School of Medicine, Tokyo, Japan. naho@juntendo.ac.jp.|Department of Pediatric Surgery, Juntendo Nerima Hospital, Tokyo, Japan. naho@juntendo.ac.jp.
Naho
N
Fujiwara
Department of Pediatric Surgery, Juntendo University School of Medicine, Tokyo, Japan.
Katsumi
K
Miyahara
Department of Pediatric Surgery, Juntendo University School of Medicine, Tokyo, Japan.|Department of Pediatric Surgery, Juntendo Nerima Hospital, Tokyo, Japan.
Nana
N
Nakazawa-Tanaka
Department of Biochemistry and Biophysics, Graduate School of Health Care Science, Tokyo Medical and Dental University, Tokyo, Japan.
Chihiro
C
Akazawa
Department of Pediatric Surgery, Juntendo University School of Medicine, Tokyo, Japan.
Atsuyuki
A
Yamataka
D000818:Animals; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000502:physiology*; D004195:Disease Models, Animal; D017615:Enteric Nervous System / Q000196:embryology* / Q000503:physiopathology; D005455:Fluorescent Antibody Technique; D006627:Hirschsprung Disease / Q000196:embryology*; D007422:Intestines / Q000196:embryology / Q000503:physiopathology; D051379:Mice; D018345:Mice, Knockout; D009432:Neural Crest / Q000196:embryology* / Q000503:physiopathology; D009474:Neurons / Q000502:physiology; D044023:Receptor, Endothelin B / Q000502:physiology*
D044023:Receptor, Endothelin B
Enteric nervous system; Enteric neural crest-derived cell; Hirschsprung’s disease; Lineage segregation; Sox10-Venus mice
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