Name: investigating alterations in caecum microbiota after traumatic brain injury in mice
Uploaded: 2019-09-19T13:00:00
Description: Watch this Scientific Journal Video about Investigating Alterations in Caecum Microbiota After Traumatic Brain Injury in Mice at JoVE.com

作者没有什么可透露的。

所有程序均经浙江大学实验动物伦理委员会批准。手术中使用的所有仪器和材料都是无菌的。TBI 的代理大约需要 20 分钟。
1. 动物护理
<ol><li>在本实验中使用5至6周大的雄性C57BL/6J小鼠（20-25克重量）。</li>
	<li>使小鼠保持在12小时/12小时光/暗周期，并确保它们获得食物和水。在整个研究中，为假人和TBI组提供相同数量的食物和水。</li>
	<li>竭尽全力尽量减少动物的痛苦和不适...

<ol>
	<li>Cheng, 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=.">.</a> Trends in traumatic brain injury mortality in China, 2006-2013: A population-based longitudinal study. 14, e1002332 (2017).</li><li>Maas, A. I. R., 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=Traumatic+brain+injury:+integrated+approaches+to+improve+prevention,+clinical+care,+and+research.">Traumatic brain injury: integrated approaches to improve prevention, clinical care, and research.</a> The Lancet Neurology. 16, 987-1048 (2017).</li><li>Gaddam, S. S., Buell, T., Robertson, C. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Systemic+manifestations+of+traumatic+brain+injury.">Systemic manifestations of traumatic brain injury.</a> Handbook of Clinical Neurology. 127, 205-218 (2015).</li><li>Kabadi, S. V., 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=Fluid-percussion-induced+traumatic+brain+injury+model+in+rats.">Fluid-percussion-induced traumatic brain injury model in rats.</a> Nature Protocols. 5, 1552-1563 (2010).</li><li>Fung, T. C., Olson, C. A., Hsiao, E. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interactions+between+the+microbiota,+immune+and+nervous+systems+in+health+and+disease.">Interactions between the microbiota, immune and nervous systems in health and disease.</a> Nature Neuroscience. 20, 145-155 (2017).</li><li>Collins, S. M., Surette, M., Bercik, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+interplay+between+the+intestinal+microbiota+and+the+brain.">The interplay between the intestinal microbiota and the brain.</a> Nature Reviews Microbiology. 10, 735-742 (2012).</li><li>Cryan, J. F., Dinan, T. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mind-altering+microorganisms:+the+impact+of+the+gut+microbiota+on+brain+and+behaviour.">Mind-altering microorganisms: the impact of the gut microbiota on brain and behaviour.</a> Nature Reviews Neuroscience. 13, 701-712 (2012).</li><li>Nicholson, S. E., 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=Moderate+Traumatic+Brain+Injury+Alters+the+Gastrointestinal+Microbiome+in+a+Time-Dependent.">Moderate Traumatic Brain Injury Alters the Gastrointestinal Microbiome in a Time-Dependent.</a> Shock. , (2018).</li><li>Houlden, 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=Brain+injury+induces+specific+changes+in+the+caecal+microbiota+of+mice+via+altered+autonomic+activity+and+mucoprotein+production.">Brain injury induces specific changes in the caecal microbiota of mice via altered autonomic activity and mucoprotein production.</a> Brain, Behavior, and Immunity. 57, 10-20 (2016).</li><li>Treangen, T. 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=Traumatic+Brain+Injury+in+Mice+Induces+Acute+Bacterial+Dysbiosis+Within+the+Fecal+Microbiome.">Traumatic Brain Injury in Mice Induces Acute Bacterial Dysbiosis Within the Fecal Microbiome.</a> Frontiers in Immunology. 9, 2757 (2018).</li><li>Alder, J., Fujioka, W., Lifshitz, J., Crockett, D. P., Thakker-Varia, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lateral+fluid+percussion:+model+of+traumatic+brain+injury+in+mice.">Lateral fluid percussion: model of traumatic brain injury in mice.</a> Journal of Visualized Experiments. , (2011).</li><li>Thompson, H. 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=Lateral+fluid+percussion+brain+injury:+a+15-year+review+and+evaluation.">Lateral fluid percussion brain injury: a 15-year review and evaluation.</a> Journal of Neurotrauma. 22, 42-75 (2005).</li><li>Pang, W., Vogensen, F. K., Nielsen, D. S., Hansen, A. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Faecal+and+caecal+microbiota+profiles+of+mice+do+not+cluster+in+the+same+way.">Faecal and caecal microbiota profiles of mice do not cluster in the same way.</a> Laboratory Animals. 46, 231-236 (2012).</li></ol>

这里介绍的是一个简单而有效的协议，用于确定小鼠TBI后头亚微生物群的变化。脑损伤的诱导和采集的囊中内容样本是协议的关键部分。
尽管研究人员研究了TBI后肠道微生物群的变化，但这些研究中使用的脑损伤是CCI-8和体重下降/撞击引起的模型9。然而，CCI模型主要复制脑挫伤，而降重模型可能会受到一些不准确的影响。在现有的脑损伤?...

创伤性脑损伤（TBI）是一个全球性的公共卫生问题，是青少年死亡和残疾的主要原因。TBI每年导致许多死亡，幸存者经历各种身体、精神、情感和认知障碍。因此，TBI 是患者家庭和社会资源的沉重负担。TBI 既包括创伤时发生的原发性脑损伤，也涉及在最初受伤后数小时至数月出现的任何继发性脑损伤。继发性脑损伤由几个生化级联介，不仅对大脑有害，而且对各种器官系统，包括胃肠道系统3有显著的负面影响。
目前，动物实验中诱导TBI的模型有三种：流体撞击损伤、控制皮质冲击（CCI）和降压加速度。横向流体打击损伤（LFPI）是建立扩散性脑损伤（DAI）最常用的模型4。该设备通过颅骨切除术对完好的杜拉施加短暂的流体压力脉冲来产生脑损伤。此脉冲由钟摆的敲击产生。LFPI是TBI研究的一种可重复、可控的建模方法。
微生物群被定义为存在于人体中的所有微生物的集体基因组。特别是肠道微生物不仅在肠道平衡和功能中起着重要作用，而且调节宿主生理和其他器官功能的许多方面。近年来，越来越多的证据表明，肠道微生物群通过脑-肠轴6调节大脑发育和功能。肠道微生物群的中断与几个大脑功能障碍有关，包括帕金森病、情绪障碍和自闭症7。最近，临床前研究也报道，急性脑损伤可诱发肠道微生物群8、9的变化。
Treangen等人10年的一项研究发现，在CCI诱导的TBI之后，三种微生物物种明显减少，两种微生物物种增加。这一证据表明，肠道微生物群的调制可能是TBI管理的一种治疗方法。然而，脑损伤引起的肠道微生物群变化背后的机制仍然未知。因此，需要一个相对简单有效的模型来研究TBI后肠道微生物群的变化。因此，本研究提出了一种检查小鼠TBI后肠道微生物群变化的协议。

<table border="0" cellpadding="0" cellspacing="0" style="width:958px;" width="959">
	<tbody>
		<tr height="40">
			<td height="40" style="height:40px;width:222px;">DNA isolation kit&#160;</td>
			<td style="width:212px;">QIAGEN</td>
			<td style="width:354px;">51604</td>
			<td style="width:171px;">For fast purification of genomic DNA from stool samples</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Gene analysis service</td>
			<td>GENEWIZ</td>
			<td></td>
			<td>Gene analyse service</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Heating pad</td>
			<td>Shanghai SAFE Biotech Co.</td>
			<td>TR-200</td>
			<td>heating pad</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Injector</td>
			<td style="width:212px;">The First Affiliated Hospital, School of Medicine, Zhejiang University</td>
			<td></td>
			<td>injector</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">LFPI device</td>
			<td style="width:212px;">Virginia 
			Commonwealth University</td>
			<td>FP302</td>
			<td>LFPI device</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Micro cranial drill</td>
			<td>RWD Life Science</td>
			<td>78061</td>
			<td>Micro cranial drill</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Povidone Iodine</td>
			<td style="width:212px;">The First Affiliated Hospital, School of Medicine, Zhejiang University</td>
			<td></td>
			<td>Povidone Iodine</td>
		</tr>
	</tbody>
</table>

investigating alterations in caecum microbiota after traumatic brain injury in mice

越来越多的证据表明，微生物-胃脑轴在脑疾病的发病机制中起着重要的作用。几项研究还表明，创伤性脑损伤导致肠道微生物群的变化。然而，大脑-肠轴的双向调节背后的机制仍然未知。目前，研究创伤性脑损伤后肠道微生物群变化的模型很少。因此，提出的研究结合了利用横向流体打击装置诱导创伤性脑损伤的规程，并分析损伤后的卡库姆样本，以研究肠道微生物群的改变。创伤性脑损伤后肠道微生物群的改变使用16S-rDNA测序确定。本方案为研究肠道微生物与创伤性脑损伤的关系提供了一种有效的方法。

TBI 的建立如图1所示。麻醉和消毒后，头皮被切口（图1A）。颅骨切除术（直径3毫米）用电钻在右皮质的颅骨上被挖到头骨中，杜拉保持完好无损（图1B，C）。在骨窗上放置了一个塑料损伤管，并用牙科丙烯酸固定在头骨上（图1D）。
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Watch this Scientific Journal Video about Investigating Alterations in Caecum Microbiota After Traumatic Brain Injury in Mice at JoVE.com

Investigating Alterations in Caecum Microbiota After Traumatic Brain Injury in Mice

这里介绍的是一种使用横向流体打击装置诱导弥漫性创伤性脑损伤的协议，然后收集胆囊成分进行肠道微生物群落分析。

研究小鼠创伤性脑损伤后凯库姆微生物群的改动

Increasing evidence shows that the microbiota-gut-brain axis plays an important role in the pathogenesis of brain diseases. Several studies also ...

大脑和肠道微生物群之间的同步性和线相互作用仍然未知。使用这种技术，可以检查小鼠脑外伤后肠道微生物群的改变。这种技术的主要优点是相对简单和高效。演示这个程序的将是文东你，一个研究生从我实验室。对于创伤性脑损伤诱导，确认麻醉小鼠对手趾捏缺乏反应，并给动物的眼睛涂抹软膏。剃须后，用70%乙醇对裸露的头皮皮肤进行消毒，然后再将头皮在下垂平面中倾斜。使用钳子，收回两侧的切口，并稍微分离周分。使用标记在头骨的右侧区域绘制一个三毫米直径的圆，该圆离中线两毫米远。使用电钻小心地穿透头骨，而不会损坏杜拉。取出骨瓣，露出三毫米骨窗，并在颅骨切除术上放置一个塑料损伤管。用牙科丙烯酸将牙管粘合到头骨后，使用五毫升注射器用无菌 0.9% 的正常盐水填充管，注意避免气泡。打开示波器和放大器。确认横向流体打击损伤装置的高压管无气泡，并提供约10个测试脉冲，直到该装置发出稳定信号。调整钟摆起始位置的角度，达到约两个标准大气的脉冲强度，将损伤管连接到横向流体打击损伤装置。扣动扳机释放钟摆，诱发脑损伤。然后通过整个封闭的、充满液体的管道系统获取脉冲并将其传输到 dura。脑损伤诱导后，取出牙管，缝合切口。然后将鼠标放在加热垫上，进行监测，直到它被流动，然后将动物返回到其家庭笼子，并获取食物和水。在指示的实验时间点，根据标准方案暴露肠道，并使用创伤钳轻轻地将肠子与其他肠道分离。当组织被定位时，用锋利的剪刀切割表，并手动将表骨内容物提取到无菌敷料上。然后将内装物转移到1.5毫升微离心管中，用于零下80摄氏度的储存，直到微生物群分析。16S重组DNA测序显示，在创伤性脑损伤三天后，小鼠的caecum微生物群多样性降低，图中所示的虚假和创伤性脑损伤组中，最丰富的分类。此外，非公制、多维结垢显示创伤性脑损伤后，小细胞微生物群的组成发生变化。记住在钻头骨时保持杜拉完好无损，在定位和分离其他肠道时要保持温和。按照这个程序，血氧素和 eosin 染色和分析可以执行，以检查创伤性脑损伤后肠道内的组织学变化和炎症反应。

Research

JoVE Journal

Neuroscience

Lateral Fluid Percussion: Model of Traumatic Brain Injury in Mice

侧向液压冲击：创伤性脑损伤小鼠模型

Traumatic brain injury (TBI) research has attained renewed momentum due to the increasing awareness of head injuries, which result in morbidity and ...

科研

神经科学

Investigations on Alterations of Hippocampal Circuit Function Following Mild Traumatic Brain Injury

调查轻度创伤性脑损伤海马电路功能的改变

Traumatic Brain Injury (TBI) afflicts more than 1.7 million people in the United States each year and even mild TBI can lead to persistent neurological ...

Laser Capture Microdissection of Enriched Populations of Neurons or Single Neurons for Gene Expression Analysis After Traumatic Brain Injury

激光捕获显微切割的丰富种群的创伤性脑损伤后神经元或单个神经元的基因表达分析

Long-term cognitive disability after TBI is associated with injury-induced neurodegeneration in the hippocampus-a region in the medial temporal lobe that ...

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

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 ...

A Method to Inflict Closed Head Traumatic Brain Injury in Drosophila

A Method to Inflict Closed Head Traumatic Brain Injury in Drosophila

方法对敌人造成闭合性颅脑创伤性脑损伤<em&gt;果蝇</em

Traumatic brain injury (TBI) affects millions of people each year, causing impairment of physical, cognitive, and behavioral functions and death. Studies ...

Imaging Neural Activity in the Primary Somatosensory Cortex Using Thy1-GCaMP6s Transgenic Mice

Imaging Neural Activity in the Primary Somatosensory Cortex Using Thy1-GCaMP6s Transgenic Mice

胸腺1-gmcmp6 转基因小鼠在原代体血源皮质中的成像神经活性

The mammalian brain exhibits marked symmetry across the sagittal plane. However, detailed description of neural dynamics in symmetric brain regions in ...

Automated Gait Analysis in Mice with Chronic Constriction Injury

慢性压迫性损伤小鼠步态的自动分析

The von Frey test is a classical method that has been widely used to examine the sensory function of neuropathic pain animals. However, it has some ...

A Novel In Vitro Model of Blast Traumatic Brain Injury

A Novel In Vitro Model of Blast Traumatic Brain Injury

一种新的脑外伤体损伤模型

Traumatic brain injury is a leading cause of death and disability in military and civilian populations. Blast traumatic brain injury results from the ...

Effects of Blast-induced Neurotrauma on Pressurized Rodent Middle Cerebral Arteries

爆炸性神经损伤对压生啮鼠中脑动脉的影响

Though there have been studies on the histopathological and behavioral effects of blast exposure, fewer have been dedicated to blast&#39;s cerebral ...