Migration and differentiation of transplanted enteric neural crest-derived cells in murine model of Hirschsprung's disease
Migration and differentiation of transplanted enteric neural crest-derived cells in murine model of Hirschsprung's disease
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DOI：
10.1007/s10616-014-9754-8
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发表时间：
2015-08-01
期刊：
影响因子：
2.2
通讯作者：
Morikawa, Yasuhide
中科院分区：
文献类型：
--
作者：
Nishikawa, Ryuhei;Hotta, Ryo;Morikawa, Yasuhide
干细胞治疗提供了在先天性巨结肠患者的无神经节肠段重建肠神经系统(ENS)的潜力。取神经脊来源细胞表达EGFP的P0-CRE/FLOXED-EGFP小鼠，获得ENS干/祖细胞。将ENS干细胞/祖细胞移植到Ret(-/-)小鼠的肠道中，Ret(-/-)小鼠是先天性巨结肠的动物模型。进行免疫组织化学分析，以确定移植细胞是否在受体肠道中长出神经元。表达EGFP的神经沟来源细胞占胃肠道细胞总数的2.52%。用流式细胞仪分离ENS干/祖细胞，在无血清培养条件下扩增为神经球样小体(NLB)。NLB中部分细胞表达神经脊标记物p75和Sox10以及神经干/祖细胞标记物Nestin和Musashi1。分离的ENS干/祖细胞在培养中分化为神经元、神经胶质细胞和成肌成纤维细胞时，检测其多能性。当与Ret(-/-)小鼠的后肠外植体共同培养时，ENS干/祖细胞迁移到无神经节的肠段并长出神经元。这项研究中使用的ENS干细胞/祖细胞似乎是治疗先天性巨结肠的细胞疗法中临床相关的供体细胞，能够在受影响的肠道定植并产生神经元。
Stem cell therapy offers the potential of rebuilding the enteric nervous system (ENS) in the aganglionic bowel of patients with Hirschsprung's disease. P0-Cre/Floxed-EGFP mice in which neural crest-derived cells express EGFP were used to obtain ENS stem/progenitor cells. ENS stem/progenitor cells were transplanted into the bowel of Ret(-/-) mouse, an animal model of Hirschsprung's disease. Immunohistochemical analysis was performed to determine whether grafted cells gave rise to neurons in the recipient bowel. EGFP expressing neural crest-derived cells accounted for 7.01 +/- A 2.52 % of total cells of gastrointestinal tract. ENS stem/progenitor cells were isolated using flow cytometry and expanded as neurosphere-like bodies (NLBs) in a serum-free culture condition. Some cells in NLBs expressed neural crest markers, p75 and Sox10 and neural stem/progenitor cells markers, Nestin and Musashi1. Multipotency of isolated ENS stem/progenitor cells was determined as they differentiated into neurons, glial cells, and myofibloblasts in culture. When co-cultured with explants of hindgut of Ret(-/-) mice, ENS stem/progenitor cells migrated into the aganglionic bowel and gave rise to neurons. ENS stem/progenitor cells used in this study appear to be clinically relevant donor cells in cell therapy to treat Hirschsprung's disease capable of colonizing the affected bowel and giving rise to neurons.