Name: transplantation of olfactory ensheathing cells to evaluate functional recovery after peripheral nerve injury
Uploaded: 2014-02-23T16:00:00
Description: Watch this Scientific Journal Video about Transplantation of Olfactory Ensheathing Cells to Evaluate Functional Recovery after Peripheral Nerve Injury at JoVE.com

作者要感谢ADIR（爱德住所辅助Insuffisants Respiratoires）和基金会去l&#39;Avenir酒店提供的财政支持，以及博士查波斯巴纳比 - 海德编辑稿件。

1。嗅粘膜和嗅球的原代培养<ol><li>此前解剖<ol><li>使介质，50毫升，加入44毫升的无钙的Dulbecco改进的Eagle / Ham氏F12培养基（DMEM/F12），补充用5毫升胎牛血清（FBS）和1毫升青霉素/链霉素的。 </li><li>外套75 厘米的烧瓶用聚-L-赖氨酸（50微克/毫升，1.5微克/厘米2），1小时，在室温下进行。 </li><li>冲洗瓶用PBS 1X。 </li><li>储存在冰箱中的涂层烧瓶达1周。 </li></ol></li><li>解剖<ol><li>安乐死?...

<ol>
	<li>Barraud, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neural+crest+origin+of+olfactory+ensheathing+glia.">Neural crest origin of olfactory ensheathing glia.</a> Proc. Natl. Acad. Sci. U.S.A. 107, 21040-21045 (2010).</li><li>Guerout, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparative+gene+expression+profiling+of+olfactory+ensheathing+cells+from+olfactory+bulb+and+olfactory+mucosa.">Comparative gene expression profiling of olfactory ensheathing cells from olfactory bulb and olfactory mucosa.</a> Glia. 58, 1570-1580 (1002).</li><li>Honore, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation,+characterization,+and+genetic+profiling+of+subpopulations+of+olfactory+ensheathing+cells+from+the+olfactory+bulb.">Isolation, characterization, and genetic profiling of subpopulations of olfactory ensheathing cells from the olfactory bulb.</a> Glia. 60, 404-413 (2012).</li><li>Franssen, E. H., de Bree, F. M., Verhaagen, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Olfactory+ensheathing+glia:+their+contribution+to+primary+olfactory+nervous+system+regeneration+and+their+regenerative+potential+following+transplantation+into+the+injured+spinal+cord.">Olfactory ensheathing glia: their contribution to primary olfactory nervous system regeneration and their regenerative potential following transplantation into the injured spinal cord.</a> Brain Res. Rev. 56, 236-258 (2007).</li><li>Lakatos, A., Franklin, R. J., Barnett, S. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Olfactory+ensheathing+cells+and+Schwann+cells+differ+in+their+in+vitro+interactions+with+astrocytes.">Olfactory ensheathing cells and Schwann cells differ in their in vitro interactions with astrocytes.</a> Glia. 32, 214-225 (2000).</li><li>Woodhall, E., West, A. K., Chuah, M. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cultured+olfactory+ensheathing+cells+express+nerve+growth+factor,+brain-derived+neurotrophic+factor,+glia+cell+line-derived+neurotrophic+factor+and+their+receptors.">Cultured olfactory ensheathing cells express nerve growth factor, brain-derived neurotrophic factor, glia cell line-derived neurotrophic factor and their receptors.</a> Brain Res. Mol. Brain Res. 88, 203-213 (2001).</li><li>Dombrowski, M. A., Sasaki, M., Lankford, K. L., Kocsis, J. D., Radtke, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Myelination+and+nodal+formation+of+regenerated+peripheral+nerve+fibers+following+transplantation+of+acutely+prepared+olfactory+ensheathing+cells.">Myelination and nodal formation of regenerated peripheral nerve fibers following transplantation of acutely prepared olfactory ensheathing cells.</a> Brain Res. 1125, 1-8 (2006).</li><li>Guerout, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transplantation+of+olfactory+ensheathing+cells+promotes+axonal+regeneration+and+functional+recovery+of+peripheral+nerve+lesion+in+rats.">Transplantation of olfactory ensheathing cells promotes axonal regeneration and functional recovery of peripheral nerve lesion in rats.</a> Muscle Nerve. 43, 543-551 (2011).</li><li>Guntinas-Lichius, O., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transplantation+of+olfactory+ensheathing+cells+stimulates+the+collateral+sprouting+from+axotomized+adult+rat+facial+motoneurons.">Transplantation of olfactory ensheathing cells stimulates the collateral sprouting from axotomized adult rat facial motoneurons.</a> Exp. Neurol. 172, 70-80 (2001).</li><li>Makoukji, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lithium+enhances+remyelination+of+peripheral+nerves.">Lithium enhances remyelination of peripheral nerves.</a> Proc. Natl. Acad. Sci U.S.A. 109, 3973-3978 (2012).</li><li>Guerout, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Co-transplantation+of+olfactory+ensheathing+cells+from+mucosa+and+bulb+origin+enhances+functional+recovery+after+peripheral+nerve+lesion.">Co-transplantation of olfactory ensheathing cells from mucosa and bulb origin enhances functional recovery after peripheral nerve lesion.</a> PloS one. 6, 22816 (2011).</li><li>Paviot, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Efficiency+of+laryngeal+motor+nerve+repair+is+greater+with+bulbar+than+with+mucosal+olfactory+ensheathing+cells.">Efficiency of laryngeal motor nerve repair is greater with bulbar than with mucosal olfactory ensheathing cells.</a> Neurobiol. Dis. 41, 688-694 (2011).</li><li>Girard, S. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolating+nasal+olfactory+stem+cells+from+rodents+or+humans.">Isolating nasal olfactory stem cells from rodents or humans.</a> J. Vis. Exp. , (2011).</li><li>Nash, H. H., Borke, R. C., Anders, J. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=New+method+of+purification+for+establishing+primary+cultures+of+ensheathing+cells+from+the+adult+olfactory+bulb.">New method of purification for establishing primary cultures of ensheathing cells from the adult olfactory bulb.</a> Glia. 34, 81-87 (2001).</li><li>Bianco, J. I., Perry, C., Harkin, D. G., Mackay-Sim, A., Feron, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neurotrophin+3+promotes+purification+and+proliferation+of+olfactory+ensheathing+cells+from+human+nose.">Neurotrophin 3 promotes purification and proliferation of olfactory ensheathing cells from human nose.</a> Glia. 45, 111-123 (2004).</li><li>Paviot, A., Bon-Mardion, N., Duclos, C., Marie, J. P., Guerout, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Although+olfactory+ensheathing+cells+have+remarkable+potential+to+sustain+nerve+regeneration,+they+cannot+be+applied+to+a+severe+vagus+nerve+section/resection+model.">Although olfactory ensheathing cells have remarkable potential to sustain nerve regeneration, they cannot be applied to a severe vagus nerve section/resection model.</a> Muscle Nerve. 43, 919-920 (2011).</li></ol>

这里介绍的技术使嗅鞘细胞，以研究细胞移植在周围神经损伤模型一个有用的模型。细胞培养协议相对简单，可以很容易地进行。在另一方面，外科手术，在喉返神经的特定部分/吻合，需要经验的，必须由合格的人员。 在这个协议中描述的方法强调重要因素集中，以便获得可能的最佳结果。首先，有机质分解过程中，我们建议仔细解剖，以获得唯一的嗅觉，而不是呼吸道上?...

主嗅觉系统包括两个不同的部分;嗅粘膜（OM）在中枢神经系统外周神经系统和嗅球（OB）。主嗅觉系统的特征在于主嗅觉神经元（PON）的自我更新整个生命中哺乳动物物种的能力。这种能力是由可能的，因为神经干细胞在OM的存在。 PON的生长和再生从OM到OB是由专门的神经胶质细胞称为嗅鞘细胞（嗅鞘细胞）促进。嗅鞘细胞是其从OM的PON到OB 1的神经发生创造了有利的微环境神经嵴细胞。因此，嗅鞘细胞可以在OM和OB构成细胞2,3的不同亚群被发现。嗅鞘细胞的不同属性有铅科学家将其用于细胞移植在几个神经系统病变范式4。事实上，嗅鞘细胞产生生长因子，减少胶质瘢痕，公关表参轴突再生，并且可以自由交融与星形胶质细胞5,6。然而，这些研究绝大多数是基于对脊髓损伤（SCI），其中一些已经用于嗅鞘细胞周围神经损伤（PNI）7,8之后。 虽然周围神经系统有很大的容量神经损伤后再生，科室齐全诱发异常的轴突再生。事实上，面部或喉返神经（RLN）完全横断后误传的轴突产生肌肉cocontractions叫联带运动。因此，它是最重要的，提出PNI的一个模型，不仅量化轴突再生长，但也评估肌肉收缩的效率。在文献中描述了最常见的模式是基于面部神经病变9,10。在这个模型中，功能性评估是基于对晶须运动10的恢复。然而，它是复杂的，以证明MOV的效率对此语句和歧视的肌肉cocontractions现象。在这里我们提出基于一个喉返神经损伤模型。这种模式不仅允许轴突再生和声带运动，但也是这些运动细胞移植11,12后的效率和功能的评价。该协议提供了一个喉返神经节/吻合模型一步步的程序，以文化和嗅鞘细胞移植从OM和OB和术后评估动物。

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			<td height="21" style="height:21px;width:240px;">DMEM/F12</td>
			<td style="width:155px;">Invitrogen</td>
			<td style="width:197px;">E3521T</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">FBS</td>
			<td>Invitrogen</td>
			<td>E3387M</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Penicilin/streptomycin</td>
			<td>Invitrogen</td>
			<td>1152-8876</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">HBSS</td>
			<td>Invitrogen</td>
			<td>M3467Y</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Trypsin-EDTA</td>
			<td>Invitrogen</td>
			<td>M3513P</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Cacodylate</td>
			<td>Merck</td>
			<td>1.03256.0100</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">DDSA</td>
			<td>BIOVALLEY</td>
			<td>00563-450</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">MNA</td>
			<td>BIOVALLEY</td>
			<td>00886-450</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">BDMA</td>
			<td>BIOVALLEY</td>
			<td>00141-100</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">POLYBED 812</td>
			<td>BIOVALLEY</td>
			<td>08791-500</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">PE anti-mouse&#160;</td>
			<td>BD Bioscience&#160;</td>
			<td>550589</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Matrigel GFR</td>
			<td>BD Bioscience&#160;</td>
			<td>356231</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Collagenase A</td>
			<td>Roche</td>
			<td>10103586001</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Mouse anti P75</td>
			<td>Chemicon</td>
			<td>MAB 365</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">11.0 Wire</td>
			<td>Ethicon&#160;</td>
			<td>FG 2881</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Toluidine Blue&#160;</td>
			<td>RAL DIAGNOSTICS</td>
			<td>361590-0025</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Centrifuge</td>
			<td>Sigma&#160;</td>
			<td>Sigma 2-16PK</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Incubator&#160;</td>
			<td>Thermo scientific</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Laminar flow hood</td>
			<td>Faster</td>
			<td style="width:197px;">BH-EN 2003 S</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Flow cytometer</td>
			<td>BD Bioscience&#160;</td>
			<td>FACSCalibur</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Microscope</td>
			<td>Zeiss</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Videolaryngoscope</td>
			<td>Karl Storz Endoskope</td>
			<td>Telecam SL NLSC 20212120</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Acquisition system</td>
			<td>ADInstruments</td>
			<td>Powerlab system</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Pyramitome Ultramicrotomy System&#160;</td>
			<td style="width:155px;">Leica</td>
			<td>Ultracut S</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Image analysis system</td>
			<td>Explora Nova</td>
			<td>Mercator</td>
		</tr>
	</tbody>
</table>

transplantation of olfactory ensheathing cells to evaluate functional recovery after peripheral nerve injury

<html>嗅鞘细胞（嗅鞘细胞）是神经嵴细胞，使增长的主要嗅觉神经元和再生。事实上，主嗅觉系统的特征在于，其以产生新的神经细胞即使在成年动物的能力。这种特殊的能力，部分原因是嗅鞘细胞的存在而对神经营造良好的微环境。嗅鞘细胞的这种特性已被用于细胞移植，如脊髓损伤模型。虽然外周神经系统有较大容量的神经损伤比对中枢神经系统后再生，完整的章节后，面部喉神经切断术的过程中，特别轴突再生长诱导错误路由。具体来说，喉返神经（RLN）的全切片诱导导致声带联带运动异常轴突再生。在这种特定的模型中，我们发现，嗅鞘细胞移植有效地增加轴突再生。 的同时存在于嗅黏膜（OM-嗅鞘细胞）和嗅球（OB-嗅鞘细胞）亚群的几个ve_content“&gt;嗅鞘细胞构成的。我们在座细胞移植的基础上在利用嗅鞘细胞的这些不同亚群的典范喉返神经损伤模型，利用这种模式，OB-嗅鞘细胞和OM-嗅鞘细胞的原代培养的喉返神经第吻合后移植于基底膜。术后2个月，我们评估移植的动物通过互补分析的基础上videolaryngoscopy，肌电图（EMG）和组织学研究。首先，videolaryngoscopy使我们能够评估喉功能，特别是肌肉cocontractions现象，然后，肌电图的分析表明丰富性和肌肉的活动同步。最后，基于甲苯胺蓝染色的组织学研究中所允许的数量和分布的量化有髓纤维。 总之，我们在这里描述了如何分离，培养，IDEntify和嗅鞘细胞移植从OM和OB RLN段吻合术，以及如何评估和分析轴突的再生和喉功能这些移植细胞的效率之后。

插图与控制和神经支配（节/吻合）动物被选为结果可能会因执行（OM，OB或OM + OB）细胞移植而有所不同。 细胞培养 细胞粘附迅速到塑料表面，成为分隔成细胞，长形或锥形，三角形，多极，或纺锤状的平行分割行（ 图1A和1B）。经过8 天的体外流式细胞分析表明p75的阳性细胞的比率，在初级OB 71％，并且在初级培养物的OM 13％（ <strong...

Watch this Scientific Journal Video about Transplantation of Olfactory Ensheathing Cells to Evaluate Functional Recovery after Peripheral Nerve Injury at JoVE.com

Transplantation of Olfactory Ensheathing Cells to Evaluate Functional Recovery after Peripheral Nerve Injury

嗅鞘细胞（嗅鞘细胞）是神经嵴细胞，使初级嗅觉神经元的生长。此特定属性可用于蜂窝移植。我们在座细胞移植的基础上在喉上神经损伤模型中使用嗅鞘细胞的模型。

嗅鞘细胞移植周围神经损伤后，以评估功能恢复

Olfactory ensheathing cells (OECs) are neural crest cells which allow growth and regrowth of the primary olfactory neurons. Indeed, the primary olfactory ...

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Neuroscience

An Optic Nerve Crush Injury Murine Model to Study Retinal Ganglion Cell Survival

一个视神经挤压伤的小鼠模型，研究视网膜神经节细胞生存

Injury to the optic nerve can lead to axonal degeneration, followed by a gradual death of retinal ganglion cells (RGCs), which results in irreversible ...

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Dorsal Root Ganglia Neurons and Differentiated Adipose-derived Stem Cells: An In Vitro Co-culture Model to Study Peripheral Nerve Regeneration

Dorsal Root Ganglia Neurons and Differentiated Adipose-derived Stem Cells: An In Vitro Co-culture Model to Study Peripheral Nerve Regeneration

背根神经节神经元和区分脂肪干细胞：一种<em&gt;在体外</em&gt;共培养模型来研究周围神经再生

Dorsal root ganglia (DRG) neurons, located in the intervertebral foramina of the spinal column, can be used to create an in vitro system facilitating the ...

Functional Evaluation of Olfactory Pathways in Living Xenopus Tadpoles

Functional Evaluation of Olfactory Pathways in Living Xenopus Tadpoles

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Xenopus tadpoles offer a unique platform to investigate the function of the nervous system. They provide multiple experimental advantages, such as ...

Toluidine Blue Staining of Resin-Embedded Sections for Evaluation of Peripheral Nerve Morphology

甲苯胺蓝染色树脂嵌段评价周围神经形态学

Peripheral nerves extend throughout the body, innervating target tissues with motor or sensory axons. Due to widespread distribution, peripheral nerves ...

A Preclinical Model to Assess Brain Recovery After Acute Stroke in Rats

评估大鼠急性中风后脑恢复的临床前模型

The purpose of this study was to establish and validate an animal brain ischemia model in the recovery and sequela stages. A middle cerebral artery ...

3D Kinematic Analysis for the Functional Evaluation in the Rat Model of Sciatic Nerve Crush Injury

坐骨神经粉碎损伤大鼠模型中功能评估的3D运动学分析

Compared to the Sciatic Functional Index (SFI), kinematic analysis is a more reliable and sensitive method for performing functional evaluations of ...

Purification of Fibroblasts and Schwann Cells from Sensory and Motor Nerves in Vitro

从体外感觉和运动神经中纯化成纤维细胞和施万细胞

The principal cells in the peripheral nervous system are the Schwann cells (SCs) and the fibroblasts. Both these cells distinctly express the sensory and ...

Automated Gait Analysis to Assess Functional Recovery in Rodents with Peripheral Nerve or Spinal Cord Contusion Injury

自动步态分析，评估带外周神经或脊髓挫伤的啮齿动物的功能恢复

Peripheral and central nerve injuries are mostly studied in rodents, especially rats, given the fact that these animal models are both cost-effective and ...

Coculture of Axotomized Rat Retinal Ganglion Neurons with Olfactory Ensheathing Glia, as an In Vitro Model of Adult Axonal Regeneration

与嗅觉恩希廷格利亚合体培养的 Axotom 化大鼠视网膜甘利翁神经元，作为成人轴再生体外模型

Olfactory ensheathing glia (OEG) cells are localized all the way from the olfactory mucosa to and into the olfactory nerve layer (ONL) of the olfactory ...

Motor Dual-Tasks for Gait Analysis and Evaluation in Post-Stroke Patients

用于中风后患者步态分析和评估的运动双任务

Eighteen stroke patients were recruited for this study involving the evaluation of cognition and walking ability and multitask gait analysis. Multitask ...