Optimizing neurogenic potential of enteric neurospheres for treatment of neurointestinal diseases

Abstract Background Enteric neurospheres derived from postnatal intestine represent a promising avenue for cell replacement therapy to treat Hirschsprung disease and other neurointestinal diseases. We describe a simple method to improve the neuronal yield of spontaneously formed gut-derived neurospheres. Materials and methods Enteric neurospheres were formed from the small and large intestines of mouse and human subjects. Neurosphere size, neural crest cell content, cell migration, neuronal differentiation, and neuronal proliferation in culture were analyzed. The effect of supplemental neurotrophic factors, including glial cell line-derived neurotrophic factor (GDNF) and endothelin-3, was also assessed. Results Mouse small intestine–derived neurospheres contained significantly more P75-expressing neural crest-derived cells (49.9 ± 15.3% versus 21.6 ± 11.9%, P versus 46.2 ± 15.6%, P versus 10.2 ± 8.2% neural crest cells, P versus 16.0 ± 13.5% enteric neurons, P versus 67.8 ± 5.8%, P Conclusions Enteric neurospheres formed from small intestine and supplemented with GDNF yield an enriched population of neural crest-derived progenitor cells and give rise to a high density of enteric neurons.