Optimizing neurogenic potential of enteric neurospheres for treatment of neurointestinal diseases

• Published in: The Journal of surgical research Vol. 206; no. 2; pp. 451 - 459
• Main Authors: Cheng, Lily S., Graham, Hannah K., Pan, Wei Hua, Nagy, Nandor, Carreon-Rodriguez, Alfonso, Goldstein, Allan M., Hotta, Ryo
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.12.2016
• Subjects: Adolescent; Animals; Cell Differentiation; Cell Movement; Cell Proliferation; Cells, Cultured; Child; Endothelin-3; Enteric nervous system; Enteric Nervous System - cytology; Enteric Nervous System - physiology; Enteric neural stem cells; Female; Gastrointestinal Diseases - therapy; Glial cell line-derived neurotrophic factor; Hirschsprung Disease - therapy; Humans; Infant; Intestine, Large - cytology; Intestine, Large - physiology; Intestine, Small - cytology; Intestine, Small - physiology; Male; Mice; Mice, Inbred C57BL; Neural Stem Cells - physiology; Neural Stem Cells - transplantation; Neurogenesis - physiology; Neurospheres; Surgery; Young Adult; Enteric nervous system; Glial cell line-derived neurotrophic factor; Enteric neural stem cells; Endothelin-3; Neurospheres
• ISSN: 0022-4804; 1095-8673; 1095-8673
• DOI: 10.1016/j.jss.2016.08.035

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. 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. Mouse small intestine–derived neurospheres contained significantly more P75-expressing neural crest-derived cells (49.9 ± 15.3% versus 21.6 ± 11.9%, P < 0.05) and gave rise to significantly more Tuj1-expressing neurons than colon-derived neurospheres (69.9 ± 8.6% versus 46.2 ± 15.6%, P < 0.05). A similar pattern was seen in neurospheres isolated from human small and large intestine (32.6 ± 17.5% versus 10.2 ± 8.2% neural crest cells, P < 0.05; 29.7 ± 16.4% versus 16.0 ± 13.5% enteric neurons, P < 0.05). The addition of GDNF to the culture media further improved the neurogenic potential of small intestinal neurospheres (75.9 ± 4.0% versus 67.8 ± 5.8%, P < 0.05) whereas endothelin-3 had no effect. 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.