Botulinum neurotoxins A and E undergo retrograde axonal transport in primary motor neurons.
Botulinum neurotoxins A and E undergo retrograde axonal transport in primary motor neurons.
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DOI：
10.1371/journal.ppat.1003087
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发表时间：
2012-12
期刊：
影响因子：
6.7
通讯作者：
Schiavo G
中科院分区：
文献类型：
--
作者：
Restani L;Giribaldi F;Manich M;Bercsenyi K;Menendez G;Rossetto O;Caleo M;Schiavo G
破伤风和肉毒中毒的临床症状之间的显著差异被归因于一旦内化在运动神经元中，亲本神经毒素的不同命运。已知破伤风毒素（TeNT）经历转胞吞作用进入抑制性中间神经元并阻断脊髓中抑制性神经递质的释放，从而引起痉挛性麻痹。相反，肉毒杆菌神经毒素（BoNT）阻断神经肌肉接头处的乙酰胆碱释放，因此诱导弛缓性麻痹。虽然公开的实验证据支持分类TeNT的轴突逆行运输途径，最近的研究结果挑战既定的观点，BoNT贩运是限制在神经肌肉接头突出这些神经毒素引起的中枢作用。这些结果表明，一个更复杂的情况下，BoNTs也从事远程贩运机制。然而，这一过程背后的细胞内途径仍不清楚。我们试图填补这一空白，通过使用初级运动神经元，无论是在大规模培养或分化的微流控装置直接监测内吞和轴突运输的全长BoNT/A和BoNT/E和它们的重组结合片段。我们发现，BoNT/A和BoNT/E被脊髓运动神经元内化，并进行快速轴突逆行运输。BoNT/A和BoNT/E在非酸性轴突载体中内化，所述载体与含有TeNT的载体部分重叠，遵循在很大程度上独立于刺激的突触囊泡内分泌的过程。在体内肌肉注射后，BoNT/A和TeNT显示具有相似的时程的中枢效应。TeNT的中枢活动延迟于周围痉挛性麻痹，而BoNT/A的中枢活动延迟于弛缓性麻痹。这些结果表明，快速轴突逆行运输室是由多功能的运输细胞器精心策划的同时转移的不同货物从神经末梢的索马，并代表了一个一般的网关交付的毒力因子和病原体的中枢神经系统。植物神经毒素是人类已知的毒性最大的分子，因此，目前被列为顶级生物威胁之一。然而，它们特异性结合神经元的能力及其对调节分泌的抑制作用促使它们在以肌张力增加为特征的病理学中的临床应用，例如肌张力障碍和各种形式的痉挛，或异常分泌，例如流口水和过度出汗，仅举几例。因此，肉毒杆菌神经毒素A（其是人类治疗中最常用的血清型）已成为用于日益增多的病理性和非病理性（例如美容）病症的治疗选择。所有目前的适应症表明，肉毒杆菌神经毒素A的全身作用和毒性通过特定的给药途径（局部注射）和该分子在组织中的低扩散而最小化。然而，最近的报告表明，与这种普遍的看法相反，肉毒杆菌神经毒素A能够到达体内的远端部位，并且可能在中枢神经系统中产生先前未预料到的影响。在这项研究中，我们证明，肉毒杆菌神经毒素A和E进入替代内吞途径（S）除了突触囊泡回收，并在脊髓运动神经元的非降解隔室进行长距离运输。我们的研究结果表明，轴突逆行运输是一种常见的传播途径，在中枢神经系统的病原体和毒力因子的重要人类和动物的健康。
The striking differences between the clinical symptoms of tetanus and botulism have been ascribed to the different fate of the parental neurotoxins once internalised in motor neurons. Tetanus toxin (TeNT) is known to undergo transcytosis into inhibitory interneurons and block the release of inhibitory neurotransmitters in the spinal cord, causing a spastic paralysis. In contrast, botulinum neurotoxins (BoNTs) block acetylcholine release at the neuromuscular junction, therefore inducing a flaccid paralysis. Whilst overt experimental evidence supports the sorting of TeNT to the axonal retrograde transport pathway, recent findings challenge the established view that BoNT trafficking is restricted to the neuromuscular junction by highlighting central effects caused by these neurotoxins. These results suggest a more complex scenario whereby BoNTs also engage long-range trafficking mechanisms. However, the intracellular pathways underlying this process remain unclear. We sought to fill this gap by using primary motor neurons either in mass culture or differentiated in microfluidic devices to directly monitor the endocytosis and axonal transport of full length BoNT/A and BoNT/E and their recombinant binding fragments. We show that BoNT/A and BoNT/E are internalised by spinal cord motor neurons and undergo fast axonal retrograde transport. BoNT/A and BoNT/E are internalised in non-acidic axonal carriers that partially overlap with those containing TeNT, following a process that is largely independent of stimulated synaptic vesicle endo-exocytosis. Following intramuscular injection in vivo, BoNT/A and TeNT displayed central effects with a similar time course. Central actions paralleled the peripheral spastic paralysis for TeNT, but lagged behind the onset of flaccid paralysis for BoNT/A. These results suggest that the fast axonal retrograde transport compartment is composed of multifunctional trafficking organelles orchestrating the simultaneous transfer of diverse cargoes from nerve terminals to the soma, and represents a general gateway for the delivery of virulence factors and pathogens to the central nervous system. Botulinum neurotoxins are the most toxic molecules known to mankind, and as a result, are currently listed among the top bio-threats. However, their ability to bind specifically to neurons and their inhibitory effects on regulated secretion prompted their clinical use in pathologies characterised by increased muscular tone, such as dystonia and various forms of spasticity, or abnormal secretion, such as drooling and excessive sweating, to cite a few. As a consequence, botulinum neurotoxin A, which is the serotype most commonly used in human therapy, has become the treatment of choice for an ever-expanding number of pathological and non-pathological (e.g. cosmetic) conditions. All current indications show that the systemic effects and toxicity of botulinum neurotoxin A are minimised by the specific route of administration (local injection) and the low diffusion of this molecule in tissues. However, recent reports suggest that in contrast to this common belief, botulinum neurotoxin A is able to reach distal sites in the body and may have previously unanticipated effects in the central nervous system. In this study, we demonstrate that botulinum neurotoxin A and E enter alternative endocytic pathway(s) in addition to synaptic vesicle recycling, and undergo long-range transport in a non degradative compartment in spinal cord motor neurons. Our results show that axonal retrograde transport is a common pathway for the dissemination in the central nervous system of pathogens and virulence factors important for human and animal health.
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影响因子：
56.9
作者：
Dong, M;Yeh, F;Chapman, ER
通讯作者：
Chapman, ER
影响因子：
11.4
作者：
Granata, Alessandra;Koo, Seong Joo;Warner, Thomas T.
通讯作者：
Warner, Thomas T.
DOI：
10.1073/pnas.0812839106
发表时间：
2009-02-03
影响因子：
11.1
作者：
Fischer, Audrey;Nakai, Yuya;Montal, Mauricio
通讯作者：
Montal, Mauricio
影响因子：
8.6
作者：
Gracies, JM
通讯作者：
Gracies, JM
影响因子：
4.8
作者：
Bohnert, S;Schiavo, G
通讯作者：
Schiavo, G