Cheng Lily S, Graham Hannah K, Pan Wei Hua, Nagy Nandor, Carreon-Rodriguez Alfonso, Goldstein Allan M, Hotta Ryo
Department of Pediatric Surgery, Massachusetts General Hospital, Harvard Medical School, 55 Fruit St., Boston, MA, 02114, USA.
Department of Surgery, University of California San Francisco, 500 Parnassus Ave., San Francisco, CA, 94143, USA.
J Surg Res. 2016 Dec;206(2):451-459. doi: 10.1016/j.jss.2016.08.035. Epub 2016 Aug 12.
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.
源自出生后肠道的肠神经球是细胞替代疗法治疗先天性巨结肠病和其他神经肠道疾病的一条有前景的途径。我们描述了一种简单的方法来提高自发形成的肠道源性神经球的神经元产量。
从小鼠和人类受试者的小肠和大肠中形成肠神经球。分析了培养中神经球大小、神经嵴细胞含量、细胞迁移、神经元分化和神经元增殖情况。还评估了包括胶质细胞系源性神经营养因子（GDNF）和内皮素-3在内的补充神经营养因子的作用。
源自小鼠小肠的神经球含有显著更多表达P75的神经嵴衍生细胞（49.9±15.3%对21.6±11.9%，P<0.05），并且比源自结肠的神经球产生显著更多表达Tuj1的神经元（69.9±8.6%对46.2±15.6%，P<0.05）。在从人类小肠和大肠分离的神经球中也观察到类似模式（神经嵴细胞为32.6±17.5%对10.2±8.2%，P<0.05；肠神经元为29.7±16.4%对16.0±13.5%，P<0.05）。向培养基中添加GDNF进一步提高了小肠神经球的神经发生潜能（75.9±4.0%对67.8±5.8%，P<0.05），而内皮素-3没有作用。
由小肠形成并补充GDNF的肠神经球产生了富集的神经嵴衍生祖细胞群体，并产生高密度的肠神经元。
