该研究工作得到由Innovatief Actieprogramma格罗宁根（IAG-3），扬德Kornelis屌斯蒂廷和提基亚Huizingafonds规定，所有在荷兰的资助。我们感谢所​​有荷兰移植协调员识别潜在的肝脏丢弃并取得知情同意。

该协议已获得医学伦理委员会的格罗宁根大学医学中心的（Medisch Ethische Toetsingscommissie），荷兰。 1.制备灌注流体注意：根据这个协议准备常温灌注机的灌注流体的总体积是2233毫升和灌注流体的目标容量渗透摩尔浓度是302 mOsmol /升。 <ol><li>从表1中描述的灌注流体的部件，保持人聚集的红血细胞，新鲜冰冻血浆和人白蛋白分离。混合各组分的其余部分以无菌的方式，并存储到手术室（OR）的溶液在无菌袋中运输。在无菌环境中（理想的是良好生产规范设施）或在层流柜中的培养室做到这一点。 </li></ol><table border="0" cellpadding="0" cellspacing="0"><tbody><tr><td> 组件 </td><td fo:width="2in" style="width:80px"> 数量 </td></tr><tr><td>聚集的红血细胞（血细胞比容60％） </td><td style="text-align:right"> 840毫升</td></tr><tr><td>新鲜冰冻血浆</td><td style="text-align:right"> 930毫升</td></tr><tr><td>人白蛋白为200g / L（Albuman，Sanquin） </td><td style="text-align:right">百毫升</td></tr><tr><td>修改肠外营养（Clinimix N17G35E，百特国际公司） </td><td style="text-align:right"> 7.35毫升</td></tr><tr><td>多种维生素输液（Cernevit，百特国际有限公司） </td><td style="text-align:right"> 7微升</td></tr><tr><td>集中微量元素输液（Nutritrace，贝朗Melsungen的AG） </td><td style="text-align:right"> 7.35毫升</td></tr><tr><td>甲硝唑用于静脉给药（5毫克/毫升）（灭滴灵，赛诺菲 - 安万特） </td><td style="text-align:right">40毫升</td></tr><tr><td>头孢唑林1000毫克瓶5毫升粉用于静脉内给药（Servazolin，山德士） </td><td style="text-align:right"> 2毫升</td></tr><tr><td>速效胰岛素（100IU / ml）的（Actrapid®，诺和诺德公司） </td><td style="text-align:right"> 20毫升</td></tr><tr><td>葡乳醛酸钙，静脉内溶液10％，137.5毫克/毫升（Sandoz公司） </td><td style="text-align:right"> 40毫升</td></tr><tr><td>无菌水2 O </td><td style="text-align:right"> 51.3毫升</td></tr><tr><td> 0.9％的NaCl溶液</td><td style="text-align:right"> 160毫升</td></tr><tr><td>碳酸氢钠8.4％的溶液</td><td style="text-align:right"> 31毫升</td></tr><tr><td>肝素5000 IE /毫升静脉给药</td><td style="text-align:right"> 4毫升</td></tr><tr><td> 总 </td><td style="text-align:right"> 2233毫升</td></tr></tbody></table> 表1：的直流电阻灌注的液体TS 16。 <ol start="2"><li>转让人聚集的红血细胞（840毫升），新鲜冷冻血浆（930毫升），人白蛋白为200g / L（100ml）中，并在步骤1.1到OR制备的溶液被施用于灌注设备。 </li></ol> 2.灌注灌注设备的<ol><li>添加灌注流体的部件，包括人类堆积的红血细胞，新鲜冷冻血浆，人白蛋白和经由连接器上的充氧的顶部在步骤1.1中制备的溶液到机器并从管道的所有气泡。 </li><li>交换机上的静脉泵，并按照屏幕上的制造商的说明。然后打开动脉泵，并按照屏幕上的制造商的说明。 </li><li>按照屏幕上的说明空对抗大气压的压力米。这确保了在perfus测得的压力离子是在门静脉和肝动脉的水平的实际压力。 </li><li>启动用氧合卡波金（95％O 2 + 5％CO 2）以4升/分钟的流速。空气流将在两个充氧器之间进行分配（每个氧合2升/分钟），这应导致约60千帕的PO 2（或450毫米汞柱）在灌注流体。对于较长的灌注，最好使用单独的氧气和二氧化碳的来源。这允许在将O 2 / CO 2的比率小的调整，其可用于调节pH值和灌注流体的pCO 2。 </li><li>取一个灌注样品用于血液气体测量15-20分钟设备已催芽后，并相应地监测pH值和电解质。 注：确保服用样品前丢弃约3ml灌注流体，因为这种流体是在外围管道，并不代表在系统中的灌注流体。添加8.4％的碳酸氢钠solution为缓冲能力，旨在为生理pH（7.35-7.45）。例如，添加25-35毫升的8.4％碳酸氢钠溶液和通过取样品用于血液气体测量定期检查在灌注流体的pH值和碳酸氢盐水平。 </li></ol> 3.采购和供肝的制备注：使用就地冷却的标准技术采购机关和冲洗与冷保存液（0-4°C）19。以促进动脉插管，离开附着到肝动脉（ 图2A）的supratruncal主动脉的片段。 <ol><li>冲洗出来的胆管与保存液（ 即威斯康星解决方案，大学）。结扎胆囊管与外科缝合线。 </li><li>包和存储在一个标准的无菌供体器官袋器官和碎冰后续运输到MP中心框。 </li><li>开始表背过程中的手术室供肝抵达后立即。 <ol><li>取为至少10毫升保存液用于微生物测试的样品。 </li><li>取出膈肌附着物肝脏的裸露区以及从腔静脉手术剪的上袖任何剩余心肌。 </li><li>解剖动脉和门静脉解剖用剪刀和结扎侧枝使用手术缝合线或hemoclips。 </li><li>使用不可吸收单丝缝合线（ 例如，3-0 Prolene线）关闭supratruncal主动脉段的远端。插入动脉插管进入supratruncal主动脉的近端和带缝线（ 图2A）固定。使用由灌注设备的制造商提供设置在一次性包插管。 </li><li>插入在门静脉的静脉插管，并与缝合线固定。使用在dispos提供的套管能够包。肝静脉仍然uncannulated。 </li><li>冲洗出胆管与保存液。插入硅导管进入胆管并用缝线固定。 注意：不要插入导管过深地进入胆管，因为这可能会造成伤害胆管上皮细胞。 </li><li>通过门静脉插管冲洗用0.9％的NaCl溶液中的肝如下： <ol><li>如果接枝已在威斯康星溶液作为保存溶液大学被保存下来，冲洗出2,000毫升冷（0-4℃）的0.9％NaCl溶液中的肝接着500ml温水（37℃）的0.9％NaCl解决方案。 </li><li>如果接枝已经在组氨酸 - 色氨酸酮戊二酸溶液，作为保存液被保存下来，冲洗出1000毫升冷（0-4℃）的0.9％NaCl溶液，然后用500ml温水（37℃）0.9肝脏％的NaCl溶液。暖冲洗的目的是为了防止在的温度的显著下降灌注液。 </li><li>立即肝脏连接到灌注设备之前执行的热烈齐平。 注意：始终保持温暖和冲洗NMP开始小于1-2分钟之间的持续时间。 </li></ol></li></ol></li></ol><table border="0" cellpadding="0" cellspacing="0"><tbody><tr><td> 捐赠者的特征（N = 12） </td><td> 数（％）或中值（IQR） </td></tr><tr><td>年龄（岁） </td><td> 61（50-64） </td></tr><tr><td>性别男） </td><td> 8（67％） </td></tr><tr><td>捐赠者类型 DCD，马斯特里赫特III型 DBD </td><td> 10（83％） 2（17％） </td></tr><tr><td>身体质量指数（BMI） </td><td> 27（25-35） </td></tr><tr><td>拒绝理由 DCD +年龄&gt; 60岁 DCD +高体重指数 DCD +种种原因* 严重的脂肪变性</td><td> 5（41％） 3（25％） 2（17％） <br<html> /&gt; 2（17％） </td></tr><tr><td>保存液 UW液 HTK液</td><td> 6（50％） 6（50％） </td></tr><tr><td>在DCD供体热缺血时间（分钟） </td><td> 14（17 - 20） </td></tr><tr><td>冷缺血时间（min） </td><td> 389（458-585） </td></tr><tr><td>供体风险指数（DRI） </td><td> 2.35（2.01-2.54） </td></tr></tbody></table> 表2：供体特性 *静脉吸毒一移植供体的历史和长期捐助SO 2 &lt;撤销生命支持再移植后的30％。 缩写：DCD，循环死后捐赠; DBD，脑死亡后捐献;威斯康星州的华盛顿大学，大学; HTK，组氨酸 - 色氨酸酮戊二酸<html> g2.jpg“/&gt; 图2：（A）图片已准备对表背和人类供体移植物（B - D）随后灌注normothermically。 （A）中的动脉插管插入surpratruncal主动脉和静脉插管插入门静脉。胆管插管与硅胆汁导管。 （B）的肝脏被定位在器官腔与它的前表面朝下，插管连接到灌注设备的管路。 （C）30分钟的常温灌注机开始后。 （D）的常温灌注机开始后6小时。在操作过程中该器官腔覆盖有透明盖，以保持无菌湿润环境对肝脏（在这些照片中未示出）。“&gt;点击此处查看该图的放大版本。 4.常温灌注机<ol><li>定位在器官腔肝脏与前表面朝下。立即通过门静脉插管连接到灌注装置的门户流入管和动脉插管装置的动脉流入管连接的肝脏到底漆灌注设备。 </li><li>按照屏幕上的制造商的说明书开始灌注这两种门户和动脉侧。设定在70毫米汞柱的平均动脉压和在11毫米汞柱的平均门静脉压。 </li><li>取灌注流体样品每30分钟为的血气参数直接分析（PO 2，的pCO 2，SO 2，HCO 2 -和pH值），并使用常规的血液气体分析生化参数（葡萄糖，钙，乳酸钙，钾和钠） 。一定要丢弃约3ml灌注取的样品中，因为该流体在外围管道，并不代表在系统中的灌注流体之前流体。 <ol><li>取这些样品吸用1ml注射器从采样连接器，可在一次性管路灌注装置的一部分的灌注流体。对于每个样品使用新的注射器，并立即从注射器在灌注流体抽吸除去任何气泡。然后插入在血液气体分析仪的注射器，然后按照分析仪的手册中提供的制造商的说明。 </li></ol></li><li>从灌注液收集血浆，冷冻并储存在-80℃用于测定碱性磷酸酶（ALKP），γ-谷氨酰转移酶（γ-GT），丙氨酸转氨酶（ALT），尿素和总胆红素。 5分钟的灌注流体的离心在1500×g离心，4℃后收集血浆。 </li></ol>

<ol>
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这个手稿的作者有没有利益冲突披露。

此视频提供了使用设备，使压力控制双灌注通过肝动脉和门静脉一步的人供体肝脏的常温灌注机步协议。虽然按照此协议，灌注机的技术故障并没有出现，所有移植物灌注良好和良好的氧化。 易地肝脏灌注了血流动力学稳定，并为代谢活跃，所生产的胆16,17定义。 这是一个公认的协议，用于人类供体肝脏的灌注机。该技术具有超过SCS 21的常规方法的几?...

器官保存在肝移植目前的方法是冲洗出来，并与供肝的后续存储在寒冷的（0-4°C）保存液（如威斯康星溶液或组氨酸，色氨酸酮戊二酸溶液大学）。这种方法被称为静态冷库（SCS）。尽管肝脏在0-4℃下的代谢率是很低的，仍有0.27微摩尔氧/分钟/克肝组织，这是不能SCS 1期间提供的需求。 SCS的常规方法，因此，导致在一定程度的供体肝脏的（附加的）损伤。虽然这一数额保存损伤是不是质量好供肝的一个问题，它可以成为次优的肝脏已经遭受一定程度损伤的供体的一个关键和限制因素。出于这个原因，肝脏与次优的质量或所谓的扩展标准供体（ECD）肝脏经常拒绝用于移植的风险ö˚F早期移植失败被认为是太高了。对移植肾功能延迟，主要的非功能和非吻合口胆管狭窄（NAS）的高利率已经在肝脏接受者循环死亡（DCD），旧的捐助者或脂肪肝移植2收件人描述后从捐赠。 NAS是发病率和死亡率的肝脏移植后的一个主要原因。 NAS可能发生在两个预算外和捐助者的肝内胆管，可伴有导管内胆泥和铸造形成3,4。尽管NAS的病因被认为是多因素的，在移植保存和移植胆管缺血/再灌注损伤已被确定为主要的潜在机制2,5。一个DCD移植物的移植已被确定为最强的危险因素为NAS的发展中的一个。一段暖缺血的一个DCD供体，器官保存在冷缺血随后再灌注的组合，和在收件人损伤被认为是负责胆管可逆损伤，这在与胆管的不良再生能力结合，导致纤维化瘢痕和胆管的变窄肝移植2,5-之后。 NAS已经报道了高达接收DCD肝6-8 30％的患者。它已经很清楚，SCS的用于移植的当前方法肝移植是不足以preinjured ECD肝脏如来自DCD供体。需要其他方法来增加和优化利用ECD肝脏移植。 机灌注（MP）是器官保存，其可以提供供体器官的保存较好，相比SCS的方法。 MP可以是特别相关的ECD移植物的保存。 MP的一项重要优点是有可能向在保存期间提供氧气的移植物。 MP可以在各种温度下进行，已被列为低温（0-10℃），subnormothermic（10-36℃）和常温（36-37℃），MP（NMP）。取决于用于MP的温度，灌注流体的类型必须调整并随着温度的升高更多的氧气应提供。 MP在人肝移植的第一个临床应用是基于低温灌注未经灌注流体9,10的活性氧。在动物模型中，低温氧化的MP（0-10℃）已经显示出具有抗缺血的保护作用/肝的再灌注损伤移植物11，并提供更好的保存胆汁的胆管周围血管丛的管道12。 Subnormothermic含氧的MP在20℃或30℃下也进行了研究的动物模型，并且显示出提供早期恢复DCD肝脏移植物功能，相比SCS 13,14。人类肝脏subnormothermic充氧MP的可行性RECently报道了七人丢弃的供肝15。 NMP（37℃）允许接枝生存能力和现有功能的评估移植16,17。此外，MP允许肝移植物的逐渐复温移植前，这已被证明有利于恢复的移植物18和复苏。 在当前协议肝灌注机中使用的灌注装置能够双重灌注（通过门静脉和肝动脉）使用两个离心泵，即提供连续的门脉血流和脉动的动脉血流。该系统的压力进行控制，从而自动调节流经肝脏，取决于肝内阻力。两个中空纤维膜氧合允许肝移植物的氧合作用，以及用于除去的CO 2。该温度可根据期望的类型的MP（最小Temperat .. [温度设定<html> 10℃URE）。流量，压力和温度的显示装置实时地允许灌注过程的连续控制上。一种新的一次性无菌集油管，水库及氧合可用于每个移植（ 图1）的灌注。 这个视频文章的目的是提供一个一步一步的协议，使用这种新开发的肝脏灌注机人供体肝脏的易地常温灌注机。 <img alt="图1" src="/files/ftp_upload/52688/52688fig1.jpg" /> 图1：（A）的概略图，（B）中的灌注机的照片，（C）的人工肺的更近的视图，以及（D）用于人类供体肝脏的常温灌注离心泵。获得=“_空白”&gt;点击此处查看该图的放大版本。

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Liver Assist</td>
			<td>Organ Assist</td>
			<td>OA.Li.Li.140</td>
			<td>Perfusion device</td>
		</tr>
		<tr>
			<td>Liver Assist disposable package</td>
			<td>Organ Assist</td>
			<td>OA.Li.DP.540</td>
			<td>Disposable set and cannulas</td>
		</tr>
		<tr>
			<td>Meredith No.8</td>
			<td>Vygon Nederlands B.V.</td>
			<td>1362082</td>
			<td>Bile duct cannula</td>
		</tr>
		<tr>
			<td>Human albumin 200g/l / ALBUMAN</td>
			<td>Sanquin</td>
			<td>15522598</td>
			<td>100 ml</td>
		</tr>
		<tr>
			<td>Modified parenteral nutrition</td>
			<td>Baxter Nederland B.V.</td>
			<td>N14G30E</td>
			<td>7.35 ml</td>
		</tr>
		<tr>
			<td>Multivitamins for infusion / CERNEVIT</td>
			<td>Baxter International Inc.</td>
			<td>9800927</td>
			<td>7 ul</td>
		</tr>
		<tr>
			<td>Concentrated trace elements for infusion / NUTRITRACE</td>
			<td>B. Braun Melsungen AG</td>
			<td>14811332</td>
			<td>7.35 ml</td>
		</tr>
		<tr>
			<td>Metronidazole 5mg/ml</td>
			<td>Baxter Nederland B.V.</td>
			<td>98181882</td>
			<td>40 ml</td>
		</tr>
		<tr>
			<td>Cefazoline / SERVAZOLIN</td>
			<td>Sandoz B.V.</td>
			<td>15611337</td>
			<td>2 ml</td>
		</tr>
		<tr>
			<td>Fast acting insulin&#160;</td>
			<td>various vendors</td>
			<td></td>
			<td>20 ml</td>
		</tr>
		<tr>
			<td>Calcium glubionate, intravenous solution 10%, 137.5 mg/mL</td>
			<td>Sandoz</td>
			<td>97038695</td>
			<td>40 ml</td>
		</tr>
		<tr>
			<td>Sterile H2O</td>
			<td>Fresenius Kabi Nederland B.V.</td>
			<td>98084453</td>
			<td>51.3 ml</td>
		</tr>
		<tr>
			<td>NaCl 0.9%</td>
			<td>Baxter Nederland B.V.</td>
			<td>15262510</td>
			<td>160 ml</td>
		</tr>
		<tr>
			<td>Heparin 5000 IE/ml for i.v. administration</td>
			<td>LEO Pharma B.V.</td>
			<td>98026178</td>
			<td>4 ml</td>
		</tr>
		<tr>
			<td>Sodium bicarbonate 8,4%</td>
			<td>B. Braun Melsungen AG</td>
			<td>97973874</td>
			<td>The amount depends on the pH</td>
		</tr>
		<tr>
			<td>Packed red blood cell (in SAGM)</td>
			<td>Blood bank (Sanquin)</td>
			<td>N0012000</td>
			<td>750 ml</td>
		</tr>
		<tr>
			<td>Fresh frozen plasma</td>
			<td>Blood bank (Sanquin)</td>
			<td>N04030A0/N04030B0</td>
			<td>900 ml</td>
		</tr>
	</tbody>
</table>

ex situ normothermic machine perfusion of donor livers

相对于传统的静态低温保存（0-4°C）， 易地灌注机可以提供更好的保护供肝。器官的连续灌流提供了机会，改善器官的质量，并允许移植前供肝易地可行性评估。这个视频文章提供了分步协议易地人供体肝脏的常温灌注机（37°C）使用的设备，提供了肝动脉和门静脉的连续灌流的压力和温度控制脉冲灌流。灌注流体由两个中空纤维膜氧合充氧，温度可以在10°C和37℃下进行调节。在灌注期间，肝脏的代谢活性以及损伤的程度可以通过从灌注流体取样的生化分析进行评估。灌注机是一个非常有前途的工具增加肝脏适合于移植的数量。

这是由于各种原因拒绝移植12人的肝脏获得知情同意从捐赠者家庭的调研后使用。捐赠者的特征是通过使用本文中描述的协议表2中描述的人类供体肝脏灌流normothermically 6小时。肝移植物的质量被监测肝灌注（ - D图2A）的宏观均匀性进行评价。肝脏的血流动力学通过监测变化的动脉和门的流动进行评估。在肝动脉和门静脉流和流的随后稳定的初始增加进行观察，结果为256±1...

Watch this Scientific Journal Video about Ex Situ Normothermic Machine Perfusion of Donor Livers at JoVE.com

Ex Situ Normothermic Machine Perfusion of Donor Livers

在这里，我们提出描述体肝移植含氧易地灌注机的协议。本文包含一步步协议采购和准备肝移植的灌注机，准备灌注液，素灌注机，并执行肝移植的含氧常温灌注机。

供肝易地常温灌注机

In contrast to conventional static cold preservation (0-4 &#176;C), ex situ machine perfusion may provide better preservation of donor livers. Continuous ...

ex-situ-normothermic-machine-perfusion-of-donor-livers

Research

JoVE Journal

Medicine

Ex Situ Normothermic Machine Perfusion of Donor Livers

11.1K 次观看.  University of Groningen, University Medical Center Groningen. 在这里，我们提出描述体肝移植含氧易地灌注机的协议。本文包含一步步协议采购和准备肝移植的灌注机，准备灌注液，素灌注机，并执行肝移植的含氧常温灌注机。 

视频: Ex Situ Normothermic Machine Perfusion of Donor Livers

Normothermic Ex Vivo Kidney Perfusion for the Preservation of Kidney Grafts prior to Transplantation

Kidney transplantation has become a well-established treatment option for patients with end-stage renal failure. The persisting organ shortage remains a serious problem. Therefore, the acceptance criteria for organ donors have been extended leading to the usage of marginal kidney grafts. These marginal organs tolerate cold storage poorly resulting in increased preservation injury and higher rates of delayed graft function. To overcome the limitations of cold storage, extensive research is focused on alternative normothermic preservation methods.
Ex vivo normothermic organ perfusion is an innovative preservation technique. The first experimental and clinical trials for ex vivo lung, liver, and kidney perfusions demonstrated favorable outcomes.
In addition to the reduction of cold ischemic injury, the method of normothermic kidney storage offers the opportunity for organ assessment and repair. This manuscript provides information about kidney retrieval, organ preservation techniques, and isolated ex vivo normothermic kidney perfusion (NEVKP) in a porcine model. Surgical techniques, set up for the perfusion solution and the circuit, potential assessment options, and representative results are demonstrated.

Normothermic Ex Vivo Kidney Perfusion for the Preservation of Kidney Grafts prior to Transplantation

The severe organ shortage has resulted in increased use of marginal kidney grafts for transplantation. This has triggered interest in alternative storage methods, since marginal grafts especially tolerate cold storage poorly. The technique of normothermic ex vivo kidney perfusion (NEVKP) represents a novel preservation method for kidney grafts prior to &#160;transplantation.

常温<em&gt;离体</em&gt;肾脏的灌注肾的移植保护移植前

Kidney transplantation has become a well-established treatment option for patients with end-stage renal failure. The persisting organ shortage remains a ...

科研

医学

A Small Animal Model of Ex Vivo Normothermic Liver Perfusion

There is a significant shortage of liver allografts available for transplantation, and in response the donor criteria have been expanded. As a result, normothermic ex vivo liver perfusion (NEVLP) has been introduced as a method to evaluate and modify organ function. NEVLP has many advantages in comparison to hypothermic and subnormothermic perfusion including reduced preservation injury, restoration of normal organ function under physiologic conditions, assessment of organ performance, and as a platform for organ repair, remodeling, and modification. Both murine and porcine NEVLP models have been described. We demonstrate a rat model of NEVLP and use this model to show one of its important applications &#8212; the use of a therapeutic molecule added to liver perfusate. Catalase is an endogenous reactive oxygen species (ROS) scavenger and has been demonstrated to decrease ischemia-reperfusion in the eye, brain, and lung. Pegylation has been shown to target catalase to the endothelium. Here, we added pegylated-catalase (PEG-CAT) to the base perfusate and demonstrated its ability to mitigate liver preservation injury. An advantage of our rodent NEVLP model is that it is inexpensive in comparison to larger animal models. A limitation of this study is that it does not currently include post-perfusion liver transplantation. Therefore, prediction of the function of the organ post-transplantation cannot be made with certainty. However, the rat liver transplant model is well established and certainly could be used in conjunction with this model. In conclusion, we have demonstrated an inexpensive, simple, easily replicable NEVLP model using rats. Applications of this model can include testing novel perfusates and perfusate additives, testing software designed for organ evaluation, and experiments designed to repair organs.

A Small Animal Model of Ex Vivo Normothermic Liver Perfusion

There is a significant liver donor shortage, and criteria for liver donors have been expanded. Normothermic ex vivo liver perfusion (NEVLP) has been developed to evaluate and modify organ function. This study demonstrates a rat model of NEVLP and tests the ability of pegylated-catalase, to mitigate liver preservation injury.

体外常温肝灌注的小动物模型

There is a significant shortage of liver allografts available for transplantation, and in response the donor criteria have been expanded. As a result, ...

Normothermic Ex Situ Heart Perfusion in Working Mode: Assessment of Cardiac Function and Metabolism

The current standard method for organ preservation (cold storage, CS), exposes the heart to a period of cold ischemia that limits the safe preservation time and increases the risk of adverse post-transplantation outcomes. Moreover, the static nature of CS does not allow for organ evaluation or intervention during the preservation interval. Normothermic ex situ heart perfusion (ESHP) is a novel method for preservation of the donated heart that minimizes cold ischemia by providing oxygenated, nutrient-rich perfusate to the heart. ESHP has been shown to be non-inferior to CS in the preservation of standard-criteria donor hearts and has also facilitated the clinical transplantation of the hearts donated after the circulatory determination of death. Currently, the only available clinical ESHP device perfuses the heart in an unloaded, non-working state, limiting assessments of myocardial performance. Conversely, ESHP in working mode provides the opportunity for comprehensive evaluation of cardiac performance by assessment of functional and metabolic parameters under physiologic conditions. Moreover, earlier experimental studies have suggested that ESHP in working mode may result in improved functional preservation. Here, we describe the protocol for ex situ perfusion of the heart in a large mammal (porcine) model, which is reproducible for different animal models and heart sizes. The software program in this ESHP apparatus allows for real-time and automated control of the pump speed to maintain desired aortic and left atrial pressure and evaluates a variety of functional and electrophysiological parameters with minimal need for supervision/manipulation.

Normothermic ex situ heart perfusion (ESHP), preserves the heart in a beating, semi-physiologic state. When performed in a working mode, ESHP provides the opportunity to perform sophisticated assessments of donor heart function and organ viability. Here, we describe our method for myocardial performance evaluation during ESHP.

工作模式下的心功能和代谢评估

The current standard method for organ preservation (cold storage, CS), exposes the heart to a period of cold ischemia that limits the safe preservation ...

Normothermic Negative Pressure Ventilation Ex Situ Lung Perfusion: Evaluation of Lung Function and Metabolism

Lung transplantation (LTx) remains the standard of care for end-stage lung disease. A shortage of suitable donor organs and concerns over donor organ quality exacerbated by excessive geographic transportation distance and stringent donor organ acceptance criteria pose limitations to current LTx efforts. Ex situ lung perfusion (ESLP) is an innovative technology that has shown promise in attenuating these limitations. The physiologic ventilation and perfusion of the lungs outside of the inflammatory milieu of the donor body affords ESLP several advantages over traditional cold static preservation (CSP). There is evidence that negative pressure ventilation (NPV) ESLP is superior to positive pressure ventilation (PPV) ESLP, with PPV inducing more significant ventilator-induced lung injury, pro-inflammatory cytokine production, pulmonary edema, and bullae formation. The NPV advantage is perhaps due to the homogenous distribution of intrathoracic pressure across the entire lung surface. The clinical safety and feasibility of a custom NPV-ESLP device have been demonstrated in a recent clinical trial involving extender criteria donor (ECD) human lungs. Herein, the use of this custom device is described in a juvenile porcine model of normothermic NPV-ESLP over a 12 h duration, paying particular attention to management techniques. Pre-surgical preparation, including ESLP software initialization, priming, and de-airing of the ESLP circuit, and the addition of anti-thrombotic, anti-microbial, and anti-inflammatory agents, is specified. The intraoperative techniques of central line insertion, lung biopsy, exsanguination, blood collection, cardiectomy, and pneumonectomy are described. Furthermore, particular focus is paid to anesthetic considerations, with anesthesia induction, maintenance, and dynamic modifications outlined. The protocol also specifies the custom device's initialization, maintenance, and termination of perfusion and ventilation. Dynamic organ management techniques, including alterations in ventilation and metabolic parameters to optimize organ function, are thoroughly described. Finally, the physiological and metabolic assessment of lung function is characterized and depicted in the representative results.

Normothermic Negative Pressure Ventilation Ex Situ Lung Perfusion: Evaluation of Lung Function and Metabolism

This paper describes a porcine model of negative pressure ventilation ex situ lung perfusion, including procurement, attachment, and management on the custom-made platform. Focus is made on anesthetic and surgical techniques, as well as troubleshooting.

非原位负压通气肺灌注：肺功能和代谢的评估

Lung transplantation (LTx) remains the standard of care for end-stage lung disease. A shortage of suitable donor organs and concerns over donor organ ...

Normothermic Ex Vivo Pancreas Perfusion for the Preservation of Pancreas Allografts before Transplantation

Pancreas transplantation (PTx) is a curative treatment for people who live with the burden of a diagnosis of diabetes mellitus (DM). However, due to organ shortages and increasing numbers of patients being listed for PTx, new strategies are needed to increase the number of available grafts for transplantation. 
Static cold storage (SCS) is considered the gold standard for standard criteria organs. However, standard criteria donors (SCD) are becoming scarce and new strategies that can increase the rate of organ acceptance from extended criteria donors (ECD) are urgently needed. 
Normothermic ex vivo perfusion (NEVP) is one of the strategies that has become increasingly popular over the past couple of decades. This preservation method has already been used successfully in other organs (liver, kidneys, and lungs) but has been minimally explored in pancreas transplantation. The few papers that describe the method for pancreas show little success, edema being one of the major issues. The following manuscript describes the successful NEVP method and setup developed by our group to perfuse swine pancreas.

Normothermic Ex Vivo Pancreas Perfusion for the Preservation of Pancreas Allografts before Transplantation

Normothermic ex vivo machine perfusion (NEVP) has scarcely been explored for the preservation of pancreas allografts. We present an innovative preservation technique for pancreas allografts before transplantation.

非原生离体胰腺灌注用于移植前胰腺同种异 体 移植物的保存

Pancreas transplantation (PTx) is a curative treatment for people who live with the burden of a diagnosis of diabetes mellitus (DM). However, due to organ ...

Lung Rapid Recovery Procurement Combined with Abdominal Normothermic Regional Perfusion in Controlled Donation after Circulatory Death

Controlled donation after circulatory death (cDCD) has contributed to increasing donor numbers all over the world. Experiences published in the last years confirm that the outcomes after lung transplantation from cDCD are similar to those from brain death donors; however, the utilization of lungs from asystole donors remains low. Several reasons may be involved: different legal frameworks among countries and centers with different premortem interventions, inadequate lung donor care before procurement, or even poor experience with cDCD procedures and protocols.
Initially, the rapid recovery technique was commonly employed for the procurement of thoracic and abdominal organs in cDCD, but, in the last decade, abdominal normothermic regional perfusion (ANRP) with extracorporeal membrane oxygenation devices has become a useful method to restore blood flow to abdominal organs, allowing their quality improvement and their functional assessment prior to transplantation. This makes the donation procedure more complex and generates doubts about injury to the grafts due to dual temperature.
The aim of this article is to describe a protocol based on a single center experience with Maastricht III donors combining lung cooling rapid recovery in the thorax and abdominal normothermic regional perfusion. Tips and tricks focused on premortem interventions and lung procurement procedure techniques are explained. This may help to minimize the reluctance among professionals to use this combined technique and encourage other donor centers to use it, despite the increased complexity of the procedure.

The protocol combines a lung cooling rapid recovery technique with abdominal normothermic regional perfusion for abdominal graft procurement in controlled asystole donors, which is a safe and useful method to expand the donor pool.

肺快速恢复采购联合腹部正常区域灌注在循环死亡后受控捐献中的应用

Controlled donation after circulatory death (cDCD) has contributed to increasing donor numbers all over the world. Experiences published in the last years ...

Functional Assessment of the Donor Heart During Ex Situ Perfusion: Insights from Pressure-Volume Loops and Surface Echocardiography

Heart transplantation remains the gold standard treatment for advanced heart failure. However, the current critical organ shortage has resulted in the allocation of a growing number of donor hearts with extended criteria. These marginal grafts are associated with a high risk of primary graft failure and may benefit from ex situ perfusion before transplant. This technology allows for extended organ preservation using warm oxygenated blood perfusion with continuous metabolic monitoring. The only NESP device currently available for clinical practice perfuses the organ in an unloaded non-working state, which does not allow for functional assessment of the beating heart. We therefore developed an original platform of NESP in working mode conditions with adjustment of left ventricular preload and afterload. This protocol was applied in porcine hearts. Ex situ functional assessment of the heart was achieved with intracardiac conductance catheterization and surface echocardiography. Along with a description of the experimental protocol, we herein report the main results, as well as pearls and pitfalls associated with the acquisition of pressure-volume loops and myocardial power during NESP. Correlations between hemodynamic findings and ultrasound variables are of major interest, especially for further rehabilitation of donor hearts before transplantation. This protocol aims to improve the assessment of donor hearts to both increase the donor pool and reduce the incidence of primary graft failure.

Functional Assessment of the Donor Heart During Ex Situ Perfusion: Insights from Pressure-Volume Loops and Surface Echocardiography

A reliable noninvasive approach for functional assessment of the donor heart during normothermic ex situ heart perfusion (NESP) is lacking. We herein describe a protocol for ex situ assessment of myocardial performance using the epicardial echocardiography and conductance catheter method.

非原位灌注期间供体心脏的功能评估：来自压力-容量环和表面超声心动图的见解

Heart transplantation remains the gold standard treatment for advanced heart failure. However, the current critical organ shortage has resulted in the ...

Rat Model of Normothermic Ex-Situ Perfused Heterotopic Heart Transplantation

Heart transplantation is the most effective therapy for end-stage heart failure. Despite the improvements in therapeutic approaches and interventions, the number of heart failure patients waiting for transplantation is still increasing. The normothermic ex situ preservation technique has been established as a comparable method to the conventional static cold storage technique. The main advantage of this technique is that donor hearts can be preserved for up to 12 h in a physiologic condition. Moreover, this technique allows resuscitation of the donor hearts after circulatory death and applies required pharmacologic interventions to improve donor function after implantation. Numerous animal models have been established to improve normothermic ex situ preservation techniques and eliminate preservation-related complications. Although large animal models are easy to handle compared to small animal models, it is costly and challenging. We present a rat model of normothermic ex situ donor heart preservation followed by heterotopic abdominal transplantation. This model is relatively cheap and can be accomplished by a single experimenter.

Here, we present an assessment protocol of a heterotopically implanted heart after normothermic ex situ preservation in the rat model.

异位灌注异位心脏移植大鼠模型

Heart transplantation is the most effective therapy for end-stage heart failure. Despite the improvements in therapeutic approaches and interventions, the ...

<meta charset="utf8"></head><body>在大鼠 EVLP 模型中使用基于 PolyhHb 的灌注推进肺移植

Exploring Alternative Perfusion Solutions Using Next-Generation Polymerized Hemoglobin-Based Oxygen Carriers in a Model of Rat Ex Vivo Lung Perfusion

Lung transplantation is hampered by the lack of suitable donors. Previously, donors that were thought to be marginal or inadequate were discarded. However, new and exciting technology, such as ex vivo lung perfusion (EVLP), offers lung transplant providers extended assessment for marginal donor allografts. This dynamic assessment platform has led to an increase in lung transplantation and has allowed providers to use donors that were previously discarded, thus expanding the donor pool. Current perfusion techniques use cellular or acellular perfusates, and both have distinct advantages and disadvantages. Perfusion composition is critical to maintaining a homeostatic environment, providing adequate metabolic support, decreasing inflammation and cellular death, and ultimately improving organ function. Perfusion solutions must contain sufficient protein concentration to maintain appropriate oncotic pressure. However, current perfusion solutions often lead to fluid extravasation through the pulmonary endothelium, resulting in inadvertent pulmonary edema and damage. Thus, it is necessary to develop novel perfusion solutions that prevent excessive damage while maintaining proper cellular homeostasis. Here, we describe the application of a polymerized human hemoglobin (PolyhHb)-based oxygen carrier as a perfusate and the protocol in which this perfusion solution can be tested in a model of rat EVLP. The goal of this study is to provide the lung transplant community with key information in designing and developing novel perfusion solutions, as well as the proper protocols to test them in clinically relevant translational transplant models.

<meta charset="utf8"></head><body>作者聚焦：利用下一代聚合人血红蛋白改善大鼠供体肺的评估和保存

Here, we describe the application of a polymerized human hemoglobin (PolyhHb)-based oxygen carrier as a perfusate and the protocol in which this perfusion solution can be tested in a model of rat ex vivo lung perfusion.