Fluorescence Visualization of the Enteric Nervous Network in a Chemically Induced Aganglionosis Model.
Fluorescence Visualization of the Enteric Nervous Network in a Chemically Induced Aganglionosis Model.
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DOI：
10.1371/journal.pone.0150579
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发表时间：
2016
期刊：
影响因子：
3.7
通讯作者：
Kuroda T
中科院分区：
文献类型：
--
作者：
Fujimura T;Shibata S;Shimojima N;Morikawa Y;Okano H;Kuroda T
胃肠动力障碍，特别是严重的变异，仍然是儿科手术的治疗挑战。缺乏源自神经嵴细胞的肠神经节细胞是运动障碍的主要原因。然而，目前可用的运动障碍动物模型的局限性继续阻碍新疗法的开发。事实上，现有的运动障碍遗传模型的寿命短和/或遗传性差，禁止有前途的方法，如干细胞替代策略的功能评价。在这里，我们诱导无神经节细胞症模型，局部苯扎氯铵在P0-Cre/GFP转基因小鼠，其中神经嵴谱系标记的绿色荧光。在化学损伤后2-8周评价胃肠道的病理学异常和功能变化。剖腹术结合荧光显微镜检查允许直接可视化的肠神经网络在体内。免疫组化进一步证实了神经节细胞、胶质细胞和Cajal间质细胞的不可逆性消失。剩余粪便重量和珠粒排出时间特别支持这种化学诱导的无神经节细胞症模型的病理生理学相关性。有趣的是，我们发现肠神经节细胞的化学消融与较长的寿命有关。通过结合神经嵴衍生物的遗传标记和肠神经节细胞的化学消融，我们开发了一种新的定制的无神经节细胞症模型。我们的研究结果表明，这种无神经节细胞症模型表现出降低胃肠动力，并显示出足够的生存功能评价。这种模型可能被证明是有用的，为未来的治疗对运动障碍的发展。
Gastrointestinal motility disorders, severe variants in particular, remain a therapeutic challenge in pediatric surgery. Absence of enteric ganglion cells that originate from neural crest cells is a major cause of dysmotility. However, the limitations of currently available animal models of dysmotility continue to impede the development of new therapeutics. Indeed, the short lifespan and/or poor penetrance of existing genetic models of dysmotility prohibit the functional evaluation of promising approaches, such as stem cell replacement strategy. Here, we induced an aganglionosis model using topical benzalkonium chloride in a P0-Cre/GFP transgenic mouse in which the neural crest lineage is labeled by green fluorescence. Pathological abnormalities and functional changes in the gastrointestinal tract were evaluated 2–8 weeks after chemical injury. Laparotomy combined with fluorescence microscopy allowed direct visualization of the enteric neural network in vivo. Immunohistochemical evaluation further confirmed the irreversible disappearance of ganglion cells, glial cells, and interstitial cell of Cajal. Remaining stool weight and bead expulsion time in particular supported the pathophysiological relevance of this chemically-induced model of aganglionosis. Interestingly, we show that chemical ablation of enteric ganglion cells is associated with a long lifespan. By combining genetic labeling of neural crest derivatives and chemical ablation of enteric ganglion cells, we developed a newly customized model of aganglionosis. Our results indicate that this aganglionosis model exhibits decreased gastrointestinal motility and shows sufficient survival for functional evaluation. This model may prove useful for the development of future therapies against motility disorders.