Preferential entry of botulinum neurotoxin A Hc domain through intestinal crypt cells and targeting to cholinergic neurons of the mouse intestine.
Preferential entry of botulinum neurotoxin A Hc domain through intestinal crypt cells and targeting to cholinergic neurons of the mouse intestine.
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DOI：
10.1371/journal.ppat.1002583
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发表时间：
2012
期刊：
影响因子：
6.7
通讯作者：
Popoff MR
中科院分区：
文献类型：
--
作者：
Couesnon A;Molgó J;Connan C;Popoff MR
肉毒中毒的特征是弛缓性麻痹，通常是由于肉毒杆菌神经毒素（BoNT）从消化道吸收穿过上皮屏障，然后通过血液循环扩散到靶向自主神经和运动神经末梢。BoNT/A通过肠屏障的运输途径尚未完全了解。我们报告，肠腔内给药纯化的BoNT/A到小鼠回肠段受损的自发性肌肉收缩，并取消由电场刺激引起的平滑肌收缩。BoNT/A进入小鼠小肠上部用荧光HcA（重链的半C-末端结构域）监测，其与细胞表面受体相互作用。我们发现，HcA优先识别的神经内分泌肠隐窝细胞，这可能是通过肠屏障的毒素的入口网站的一个子集，然后在粘膜下层和后来（90-120分钟）在肌层的特定神经元为目标。HcA主要与粘膜下层和肌间神经丛的某些胆碱能神经元结合，但也识别（尽管程度较低）其他神经元细胞，包括粘膜下层中的胆碱能和胆碱能神经元。HcA靶向肠胆碱能神经元可以解释在肉毒中毒中观察到的肠蠕动和分泌的抑制，但靶向非胆碱能神经元的后果仍有待确定。肉毒杆菌中毒是人类和动物的一种严重且通常致命的疾病，其特征是驰缓性瘫痪。肉毒梭菌产生一种有效的神经毒素（肉毒神经毒素），负责肉毒中毒的所有症状。肉毒杆菌中毒最常见的是通过摄入受污染食物中预先形成的肉毒杆菌神经毒素或在肠内定植C。肉毒杆菌在某些情况下，如婴儿肉毒杆菌中毒，和毒素的生产在肠道。这种疾病的第一步是肉毒杆菌神经毒素通过肠屏障，这一点仍然知之甚少。我们研究了贩运的肉毒杆菌神经毒素在小鼠肠循环模型，使用荧光HcA（半C-末端结构域的重链）。我们观察到HcA优先识别神经内分泌肠隐窝细胞，其可能代表毒素通过肠屏障的进入位点，然后靶向粘膜下层中的特定神经元，主要是胆碱能神经元，随后（90-120分钟）在肌层中导致局部麻痹作用，例如抑制肠痉挛。这些结果代表了对肉毒杆菌中毒初始步骤的理解的重要进展，可以作为开发新的针对肉毒杆菌中毒的具体对策的基础。
Botulism, characterized by flaccid paralysis, commonly results from botulinum neurotoxin (BoNT) absorption across the epithelial barrier from the digestive tract and then dissemination through the blood circulation to target autonomic and motor nerve terminals. The trafficking pathway of BoNT/A passage through the intestinal barrier is not yet fully understood. We report that intralumenal administration of purified BoNT/A into mouse ileum segment impaired spontaneous muscle contractions and abolished the smooth muscle contractions evoked by electric field stimulation. Entry of BoNT/A into the mouse upper small intestine was monitored with fluorescent HcA (half C-terminal domain of heavy chain) which interacts with cell surface receptor(s). We show that HcA preferentially recognizes a subset of neuroendocrine intestinal crypt cells, which probably represent the entry site of the toxin through the intestinal barrier, then targets specific neurons in the submucosa and later (90–120 min) in the musculosa. HcA mainly binds to certain cholinergic neurons of both submucosal and myenteric plexuses, but also recognizes, although to a lower extent, other neuronal cells including glutamatergic and serotoninergic neurons in the submucosa. Intestinal cholinergic neuron targeting by HcA could account for the inhibition of intestinal peristaltism and secretion observed in botulism, but the consequences of the targeting to non-cholinergic neurons remains to be determined. Botulism is a severe and often fatal disease in man and animals characterized by flaccid paralysis. Clostridium botulinum produces a potent neurotoxin (botulinum neurotoxin) responsible for all the symptoms of botulism. Botulism is most often acquired by ingesting preformed botulinum neurotoxin in contaminated food or after intestinal colonization by C. botulinum under certain circumstances, such as in infant botulism, and toxin production in the intestine. The first step of the disease consists in the passage of the botulinum neurotoxin through the intestinal barrier, which is still poorly understood. We investigated the trafficking of the botulinum neurotoxin in a mouse intestinal loop model, using fluorescent HcA (half C-terminal domain of the heavy chain). We observed that HcA preferentially recognizes neuroendocrine intestinal crypt cells, which likely represent the entry site of the toxin through the intestinal barrier, then targets specific neurons, mainly cholinergic neurons, in the submucosa, and later (90–120 min) in the musculosa leading to local paralytic effects such as inhibition of intestinal peristaltism. These results represent an important advance in the understanding of the initial steps of botulism intoxication and can be the basis for the development of new specific countermeasures against botulism.