Name: veno-venous extracorporeal membrane oxygenation in a mouse
Uploaded: 2018-10-24T13:00:00
Description: Watch this Scientific Journal Video about Veno-Venous Extracorporeal Membrane Oxygenation in a Mouse at JoVE.com

该项目得到了德意志科技的肯德基311赠款的支持。

对雄性 C57BL/6 小鼠进行了实验, 年龄为12周。这项研究是按照《 TSA 16/2250 议定书》规定的德国动物法准则进行的。
1. 材料制备
注: 所有步骤均在清洁、非灭菌条件下执行。如果动物在术后存活, 将需要无菌条件。
<ol><li>使用16X 放大显微镜下的手术刀片, 将 3 fenestrations 引入2阻燃聚氨酯管中。 注意: 所有 fenestrations 必须位于套管的远端第三, 以确保最佳?...

<ol>
	<li>Hill, J. 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=Prolonged+Extracorporeal+Oxygenation+for+Acute+Post-Traumatic+Respiratory+Failure+(Shock-Lung+Syndrome).">Prolonged Extracorporeal Oxygenation for Acute Post-Traumatic Respiratory Failure (Shock-Lung Syndrome).</a> New England Journal of Medicine. 286 (12), 629-634 (1972).</li><li>Noah, M. 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=Referral+to+an+Extracorporeal+Membrane+Oxygenation+Center+and+Mortality+Among+Patients+With+Severe+2009+Influenza+A(H1N1).">Referral to an Extracorporeal Membrane Oxygenation Center and Mortality Among Patients With Severe 2009 Influenza A(H1N1).</a> Journal of the American Medical Association. 306 (15), 1659 (2011).</li><li>Maslach-Hubbard, A., Bratton, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extracorporeal+membrane+oxygenation+for+pediatric+respiratory+failure:+History,+development+and+current+status.">Extracorporeal membrane oxygenation for pediatric respiratory failure: History, development and current status.</a> World Journal of Critical. Care Medicine. 2 (4), 29-39 (2013).</li><li>Langer, T., 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="Awake"+extracorporeal+membrane+oxygenation+(ECMO):+pathophysiology,+technical+considerations,+and+clinical+pioneering.">"Awake" extracorporeal membrane oxygenation (ECMO): pathophysiology, technical considerations, and clinical pioneering.</a> Critical Care. 20 (1), 150 (2016).</li><li>Esper, S. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extracorporeal+Membrane+Oxygenation.">Extracorporeal Membrane Oxygenation.</a> Advances in Anesthesia. 35 (1), 119-143 (2017).</li><li>Millar, J. E., Fanning, J. P., McDonald, C. I., McAuley, D. F., Fraser, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+inflammatory+response+to+extracorporeal+membrane+oxygenation+(ECMO):+a+review+of+the+pathophysiology.">The inflammatory response to extracorporeal membrane oxygenation (ECMO): a review of the pathophysiology.</a> Critical Care. 20 (1), 387 (2016).</li><li>Lubnow, M., 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=Technical+complications+during+veno-venous+extracorporeal+membrane+oxygenation+and+their+relevance+predicting+a+system-exchange--retrospective+analysis+of+265+cases.">Technical complications during veno-venous extracorporeal membrane oxygenation and their relevance predicting a system-exchange--retrospective analysis of 265 cases.</a> Public Library of Science One. 9 (12), e112316 (2014).</li><li>Passmore, M. 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=Inflammation+and+lung+injury+in+an+ovine+model+of+extracorporeal+membrane+oxygenation+support.">Inflammation and lung injury in an ovine model of extracorporeal membrane oxygenation support.</a> American Journal of Physiology - Lung Cellular and Molecular Physiology. 311 (6), L1202-L1212 (2016).</li><li>Vaquer, S., de Haro, C., Peruga, P., Oliva, J. C., Artigas, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Systematic+review+and+meta-analysis+of+complications+and+mortality+of+veno-venous+extracorporeal+membrane+oxygenation+for+refractory+acute+respiratory+distress+syndrome.">Systematic review and meta-analysis of complications and mortality of veno-venous extracorporeal membrane oxygenation for refractory acute respiratory distress syndrome.</a> Annals of Intensive Care. 7 (1), 51 (2017).</li><li>Houser, S. 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=Animal+Models+of+Heart+Failure+A+Scientific+Statement+From+the+American+Heart+Association.">Animal Models of Heart Failure A Scientific Statement From the American Heart Association.</a> Circulation Research. 111 (1), 131-150 (2012).</li><li>Russell, J. C., Proctor, S. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Small+animal+models+of+cardiovascular+disease:+tools+for+the+study+of+the+roles+of+metabolic+syndrome,+dyslipidemia,+and+atherosclerosis.">Small animal models of cardiovascular disease: tools for the study of the roles of metabolic syndrome, dyslipidemia, and atherosclerosis.</a> Cardiovascular Pathology. 15 (6), 318-330 (2006).</li><li>Madrahimov, 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=Novel+mouse+model+of+cardiopulmonary+bypass.">Novel mouse model of cardiopulmonary bypass.</a> European Journal of Cardio-thoracic Surgery. 53 (1), 186-193 (2017).</li><li>Madrahimov, 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=Cardiopulmonary+Bypass+in+a+Mouse+Model:+A+Novel+Approach.">Cardiopulmonary Bypass in a Mouse Model: A Novel Approach.</a> J. Journal of Visualized Experiments. (127), (2017).</li></ol>

以前, 我们描述了一个成功的模型的 CPB 在鼠标12,13。为了实现急性或终末期肺部疾病的这种模式, 我们开发了一个易于使用的静脉-静脉 ECMO 电路的小鼠。不同于 CPB 模型, 静脉静脉 ECMO 不需要复杂的手术程序, 如胸骨和夹紧的主动脉, 从而减少伤口出血的风险在一个完全肝素化的动物。为避免肺血栓栓塞, 每只动物可获得 2.5 IU 肝素/千克。这种剂量是基于?...

体外膜氧化 (ECMO) 是一种临时的生命支持系统, 接管肺和心脏的功能, 以允许适当的气体交换和灌注。希尔 et al1描述了1972年首次使用 ECMO 患者;然而, 它仅在22009年在甲型h1n1 流感大流行期间成功应用后才得到广泛使用。今天, ECMO 通常被用作在末期心脏病和肺部疾病3的救生程序。静脉静脉 ECMO 被越来越多地用于在清醒, 非插管, 自发性呼吸衰竭患者4的侵入性机械通气的替代。
尽管其广泛采用, ECMO5,6,7报告了各种各样的复杂化。ECMO 患者可能经历的并发症包括出血、血栓、败血症、血小板减少症、与设备有关的故障和空气栓塞。此外, 全身炎症反应综合征 (sir) 导致多器官损害是很好地描述了临床和实验研究8,9。在长期 ECMO 治疗的患者中也经常报告神经并发症, 如脑梗死。混淆问题, 通常很难区分并发症是由 ECMO 本身引起的, 还是源于伴随着急性和终末期疾病的基础紊乱。
为了具体研究 ECMO 对健康机体的影响, 必须建立可靠的实验动物模型。很少有关于 ECMO 在小动物身上的性能的报告, 都只限于老鼠。迄今为止, 文献中没有描述 ECMO 的小鼠模型。由于大量的转基因小鼠菌株的可用性, 建立一个小鼠 ECMO 模型将允许进一步调查涉及 ECMO 相关并发症的分子机制10,11。
根据我们先前描述的小鼠体外循环 (CPB) 模型12, 我们开发了一种稳定的静脉-静脉 ECMO 的方法, 在非插管, 自发呼吸鼠。ECMO 电路 (图 1), 包含流出和流入套管, 蠕动泵, 肺和气陷油藏, 类似于我们以前描述的小鼠 CPB12模型, 除了有一个较小的启动容积 (0.5 毫升)。该协议演示了在成功的 ECMO 过程中涉及的详细技术、生理监测和血气分析。

<table>
	<tbody>
		<tr>
			<td>Sterofundin</td>
			<td>B.Braun Petzold GmbH</td>
			<td>PZN:8609189</td>
			<td>in 1:1 with Tetraspan</td>
		</tr>
		<tr>
			<td>Tetraspan 6% Solution</td>
			<td>B. Braun Melsungen AG</td>
			<td>PZN: 05565416</td>
			<td>in 1:1 with Sterofundin</td>
		</tr>
		<tr>
			<td>Heparin Natrium 25.000</td>
			<td>Ratiopharm GmbH</td>
			<td>PZN: 3029843</td>
			<td>2,5 IU per ml of priming</td>
		</tr>
		<tr>
			<td>NaHCO3 8,4% Solution</td>
			<td>B. Braun Melsungen AG</td>
			<td>PZN: 1579775</td>
			<td>3% in priming solution</td>
		</tr>
		<tr>
			<td>Carprofen</td>
			<td>Zoetis Inc., USA</td>
			<td>PZN:00289615</td>
			<td>5mg/kg/BW</td>
		</tr>
		<tr>
			<td>1 Fr PU Catheter</td>
			<td>Instechlabs INC., USA</td>
			<td>C10PU-MCA1301</td>
			<td>carotide artery</td>
		</tr>
		<tr>
			<td>2 Fr PU Catheter</td>
			<td>Instechlabs INC., USA</td>
			<td>C20PU-MJV1302</td>
			<td>jugular vein</td>
		</tr>
		<tr>
			<td>8-0 Silk suture braided</td>
			<td>Ashaway Line &#38; Twine Co., USA</td>
			<td>75290</td>
			<td>ligature</td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Piramal Critical Care GmbH</td>
			<td>PZN:9714675</td>
			<td>narcosis</td>
		</tr>
		<tr>
			<td>Spring Scissors - 6mm Blades</td>
			<td>Fine Science Tools GmbH</td>
			<td>15020-15</td>
			<td>instruments</td>
		</tr>
		<tr>
			<td>Spring Scissors - 2mm Blades</td>
			<td>Fine Science Tools GmbH</td>
			<td>15000-03</td>
			<td>instruments</td>
		</tr>
		<tr>
			<td>Halsted-Mosquito Hemostat</td>
			<td>Fine Science Tools GmbH</td>
			<td>13009-12</td>
			<td>instruments</td>
		</tr>
		<tr>
			<td>Dumont #55 Forceps</td>
			<td>Fine Science Tools GmbH</td>
			<td>11295-51</td>
			<td>instruments</td>
		</tr>
		<tr>
			<td>Castroviejo Micro Needle Holder - 9cm</td>
			<td>Fine Science Tools GmbH</td>
			<td>12060-02</td>
			<td>instruments</td>
		</tr>
		<tr>
			<td>Micro Serrefines</td>
			<td>Fine Science Tools GmbH</td>
			<td>18555-01</td>
			<td>instruments</td>
		</tr>
		<tr>
			<td>Bulldog Serrefine</td>
			<td>Fine Science Tools GmbH</td>
			<td>18050-28</td>
			<td>instruments</td>
		</tr>
		<tr>
			<td>Isoflurane Vaporizer Drager 19.1</td>
			<td>Dr&#228;gerwerk AG &#38; Co. KGaA</td>
			<td></td>
			<td>anesthesia 1,3 -2,5%</td>
		</tr>
		<tr>
			<td>Multichannel Data Aquisition Device with ISOHEART Software</td>
			<td>Hugo Sachs Elektronik GmbH, Germany</td>
			<td></td>
			<td>invasive pressure, ECG, t &#176;C</td>
		</tr>
		<tr>
			<td>i-STAT portable device</td>
			<td>Abbott Laboratories, Lake Bluff, Illinois, USA</td>
			<td></td>
			<td>blood gas analysis</td>
		</tr>
		<tr>
			<td>i-STAT CG4+ and CG8+ cartridges</td>
			<td>Abbott Laboratories, Lake Bluff, Illinois, USA</td>
			<td></td>
			<td>blood gas analysis</td>
		</tr>
		<tr>
			<td>C57Bl/6 mice, male, 30 g, 14 weeks old</td>
			<td>Charles River Laboratories</td>
			<td></td>
			<td>housed 1 week before</td>
		</tr>
	</tbody>
</table>

veno-venous extracorporeal membrane oxygenation in a mouse

近年来, 体外膜氧合 (ECMO) 的使用大幅度增加。ECMO 已成为急性和终末期肺部疾病的可靠和有效的治疗方法。随着临床需求的增加和 ECMO 的长期使用, 对多器官损伤的程序优化和预防具有重要的意义。本协议的目的是在非插管、自发呼吸的小鼠中提出静脉静脉 ECMO 的详细技术。本协议展示了 ECMO 和手术步骤的技术设计。这种小鼠 ECMO 模型将有助于研究与 ECMO 相关的病理生理学 (如炎症, 出血和血栓栓塞事件)。由于基因修饰小鼠的丰富性, ECMO 相关并发症的分子机制也可以被解剖。

本协议描述了小鼠静脉 ECMO 的方法。这个模型是可靠的和可重现的, 并与我们先前描述的体外循环呼吸和循环阻滞模型12,13, 它是不太技术上要求建立。
ECMO 在静脉系统中的流动保持在1.5 和5毫升/分钟之间。通过在 ECMO 电路中加入额外的启动液, 将平均动脉压力保持在70到85毫米汞柱之?...

Watch this Scientific Journal Video about Veno-Venous Extracorporeal Membrane Oxygenation in a Mouse at JoVE.com

Veno-Venous Extracorporeal Membrane Oxygenation in a Mouse

本文介绍了一种非插管、自发呼吸小鼠静脉-静脉体外膜氧合 (ECMO) 技术的协议。这种小鼠 ECMO 模型可在急性和终末期肺部疾病的实验研究中得到有效的实施。

小鼠静脉内膜氧化术

The use of extracorporeal membrane oxygenation (ECMO) has increased substantially in recent years. ECMO has become a reliable and effective therapy for ...

这种方法可以帮助回答肺研究领域的一些关键问题，例如与急性和慢性晚期肺病的治疗相关的问题。该技术的主要优点是，它有利于在长期体外膜氧合期间启动多器官损伤的分子和病理机制，作为一种拯救生命的过程。在清洁、非无菌条件下，使用手术刀片和解剖显微镜将三个芬斯特引入两个法国聚氨酯导管的远点三分之一，其放大倍率为 16 倍。接下来，将流出的管罐放入注油溶液中，打开近源泵以填充体外膜氧合（ECMO）机，以每分钟一毫米的流量对电路进行注油。30分钟后，向氧机添加每分钟0.5升100%氧气，并确认麻醉成年小鼠对手趾捏缺乏反应。在动物的眼睛上涂下软膏后，在颈部左侧做一个四毫米的侧皮切口，以可视化壶静脉，并使用微钳子和棉签进一步暴露血管。使用 8-0丝缝合，以打上暴露静脉的端端，并在近端放置一个滑结。用微切口将静脉前壁插入，通过26量球的麸皮将每克肝素2.5个国际单位注射到血管静脉中，将手术垫的头部端提高30度，以避免在插入过程中过度失血。小心地将两根法国聚氨酯管插入壶静脉的近端，稍微旋转管，同时将管杆推到四厘米深，以达到劣质维纳卡瓦的腹腔分叉。轻轻推管很重要，但连续不使用额外的力。如果遇到阻力，将管拉回并再次插入。用 8 - 0 固定管丝结，并暴露正确的壶静脉，刚刚证明。用一个法国聚氨酯管，将右壶静脉，轻轻地将管状动脉向右中庭移动五毫米，并用 8-0 固定管状丝绸结。用另一个法国聚氨酯导管将左股动脉连接起来，用于侵入性压力监测和血气分析，并将连接到数据采集装置的心电图针头插入两个前肢和左胸壁。然后插入连接到数据采集设备的直肠温度计。要启动静脉体外膜氧合，请将泵打开，初始流量为每分钟 0.1 毫升。两分钟后，将流量调整为每分钟三到五毫升。在稳定流量下，通过数据采集设备实时监控关键参数，持续检查静脉排水的回流，监测空气陷阱储液罐中的血液水平。使用装有 24 量表的麸皮的一毫升注射器收集任何血液泄漏，然后通过空气陷印储液罐将血液返回到 ECMO 回路。对于血气分析，在ECMO启动后10分钟，使用血液采样盒从氧气器前、氧气器后和股动脉采集约75微升动脉血液。ECMO启动后30分钟和60分钟，仅从股动脉采集血液。启动后 45 分钟和 90 分钟，施用 0.1 毫升的注油溶液，以补偿通过空气疏水器的内脏液体损失。启动两小时后，从氧气器、劣质静脉和股动脉中采集血液。上次血液采集后，在五分钟内降低泵的流量，从而停止 ECMO，同时继续记录重要参数 10 分钟。在一个典型的实验中，动物的生理参数每10分钟记录一次。在这个具有代表性的实验中，血液学参数表明在ECMO期间有相关的血液调节，尽管不需要输血来补偿中度贫血。氧合参数在氧空气混合物中以 1.0 的受启发氧气的一小部分显示适当的氧机性能。此外，ECMO期间的代谢变化包括实验开始时和实验结束时的呼吸碱化。没有进行额外的血液缓冲。使用适当的造血技术吸收通过ECMO回路的血液流动，通过血气分析监测氧合参数，用额外的注血溶液取代血液采样造成的任何失血，这一点很重要。这项技术为肺病领域的研究人员铺平了道路，通过使用80多个可用的仿敲鼠标模型，显著改善了挽救生命的治疗方案。

Research

JoVE Journal

Medicine

Dual-mode Imaging of Cutaneous Tissue Oxygenation and Vascular Function

双模式成像，皮肤组织氧合和血管功能

Accurate assessment of cutaneous tissue oxygenation and vascular function is important for appropriate detection, staging, and treatment of many health ...

科研

医学

A Mouse Model of in Utero Transplantation

A Mouse Model of in Utero Transplantation

一个小鼠模型在子宫内移植

The transplantation of stem cells and viruses in utero has tremendous potential for treating congenital disorders in the human fetus.  For example, in ...

Multi-photon Imaging of Tumor Cell Invasion in an Orthotopic Mouse Model of Oral Squamous Cell Carcinoma

多光子成像在口腔鳞状细胞癌原位小鼠模型的肿瘤细胞的侵袭

Loco-regional invasion of head and neck cancer is linked to metastatic risk and presents a difficult challenge in designing and implementing patient ...

Hyperinsulinemic-euglycemic Clamps in Conscious, Unrestrained Mice

在自觉的，奔放的小鼠正常血糖高胰岛素 - 夹钳

Type 2 diabetes is characterized by a defect in insulin action. The hyperinsulinemic-euglycemic clamp, or insulin clamp, is widely considered the "gold ...

A Simple Method of Mouse Lung Intubation

一个简单的方法小鼠肺插管

A  simple procedure to intubate mice for pulmonary function measurements would  have several advantages in longitudinal studies with limited numbers or  ...

Carotid Artery Infusions for Pharmacokinetic and Pharmacodynamic Analysis of Taxanes in Mice

颈动脉输液的紫杉烷类化合物在小鼠体内的药代动力学和药效学分析

When proposing the use of a drug, drug combination, or drug delivery into a novel system, one must assess the pharmacokinetics of the drug in the study ...

Corneal Donor Tissue Preparation for Descemet's Membrane Endothelial Keratoplasty

角膜供体组织准备后弹力层内皮角膜移植术

Descemet&#8217;s Membrane Endothelial Keratoplasty (DMEK) is a form of corneal transplantation in which only a single cell layer, the corneal endothelium, ...

Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo

Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo

宫内遥测来衡量鼠标收缩压力<em&gt;在体内</em

A complex integration of molecular and electrical signals is needed to transform a quiescent uterus into a contractile organ at the end of pregnancy. ...

Cardiopulmonary Bypass in a Mouse Model: A Novel Approach

小鼠体外循环模型: 一种新方法

As prolonged cardiopulmonary bypass becomes more essential during cardiac interventions, an increasing clinical demand arises for procedure optimization ...

Measurement of Tissue Oxygenation Using Near-Infrared Spectroscopy in Patients Undergoing Hemodialysis

使用近红外光谱测量血液透析患者的组织氧合

Near-infrared spectroscopy (NIRS) has recently been applied as a tool to measure regional oxygen saturation (rSO2), a marker of tissue oxygenation, in ...