The authors have nothing to disclose.

注:本研究通过授权伦理委员会数CEEA34.PB.103.12。 1.使用仔猪作为动物模型<ol><li>执行在两个组（每组6只），匹配性别，年龄（15±3个月）和重量（30±10公斤）（对照组和慢性缺氧（CH）基团）的条款。房子的CH组3周低压氧舱（0.4大气压），并在房子常压定期（1大气压）的条件下，对​​照组。 </li><li>使用低压室 ​​组成的有机玻璃箱的与2平方米足迹和1.6米的高度， 如图1。 </li><li>连续监测和保持为18℃的温度和0.4大气压的压力。确保有足够的通风用真空泵，使8 立方米每小时的空气更新率。 </li><li>将两种动物在适当的枯枝落叶室。每48小时，返回在框中的压力为常压一半条件一个小时清洗中，在两种动物的存在下，与盒保持closedto防止它们逃逸。诱导性肺动脉高压的这种方法已被广泛证实13。 </li><li>麻醉动物的<ol><li>麻醉的仔猪具有初始静脉注射硫喷妥钠（10毫克/公斤），并通过连续吸入异氟烷（1.5至3.5％）的维持麻醉。应用两滴卡波姆凝胶进入小猪的眼睛，以防止角膜干燥。 </li><li>将动物在其前腿绑在一弯曲位置以露出胸部左侧卧位。清洁用肥皂和水的皮肤，然后用刮电动剃须刀，以消除任何毛发，可以通过胸部妨碍超声波的穿透力。 </li></ol></li><li>超声心动图</li><li>通过超声心动图监测PAH的发展纵向和非侵入性。每周使用3兆赫传感器执行回声。建议奥德心电图通过将3个电极上的右侧和左侧爪子和右侧胸部。 </li><li>在使用多普勒探头三种不同的发生率（纵，短轴和心尖）记录二维和M模式图像数据。持探针在第四和第五左侧肋间空间之间的右手和地点。慢慢地移动探头上下，旋转它左，右，直到一个良好的图像分辨率获得用于不同心脏结构（ 即该隔膜）。声学窗口稍有不同从一个动物到另一个取决于心脏的胸部的位置。 </li><li>记录的心电图，同时为每个发生率和至少10个心动周期，以使离线分析。舒张末期被定义为在所述心动周期与心电图上的Q波的开始重合的点。收缩末期恰逢T波的发生。 </li><li>衡量一个二尖瓣和三尖瓣血流鉴于皮卡尔多普勒超声。将多普勒探头在胸骨柄sterni的尖端;采样区位于刚刚阀门上面记录血流渡阀。 </li><li>心动周期多普勒血流仪中记录的血流速度，以获得速度 - 时间积分血流通过二尖瓣和三尖瓣和主动脉和肺动脉瓣的。使用ECHOGRAPH的视频图形阵列在25每秒的帧速率来存储，以获得由ECHOGRAPH提供固定的图像或图像序列的图像。 </li><li>测量的尺寸和使用的测量仪器的心脏腔的表面面积，并建议通过集成到ECHOGRAPH按照国际建议14的软件的表面区域的轮廓。 </li><li>舒张和收缩由TM（时间运动）记录在短轴发病14期间测量的右心室游离壁的厚度。 </li><li> MEA确保肺动脉根直径在上短轴视图14的肺动脉瓣的尖端。 </li><li>测量隔膜和左心室舒张TM中运动上在心电图上和在收缩在心电图上14的T波的末端的Q波的开始的纵向视图中后壁。 </li><li>测量左室舒张末期内径（LVEDD），左室收缩末期内径（LVESD）。计算使用公式FS（％）=（LVEDD-LVESD）/ LVEDD短轴缩短率（FS）。测量主动脉的直径和右和左心房的表面积。 </li><li>从速度-时间积分二尖瓣和三尖瓣血流量，测量以下:E和A峰，E峰减速半时间的最大振幅，时间流动，速度，时间指数14。 </li><li>从肺动脉多普勒测量，测量以下:最大速度，提升半的时间，流量持续时间和积分14。 </li><li>评估使用辛普森15的方法左心室容积。 </li><li>存储数据在对整个集团后续的统计分析数据库的每个仔猪。 </li></ol> 2.右心导管<ol><li>动物准备<ol><li>脉动导管放置之前，猪3星期置于低压室，以诱导肺动脉高压。 </li><li>快动物手术前24小时（24小时为固体食物，8〜12小时的水）。 </li><li> 24小时前RHC，有一支训练有素的兽医进行麻醉前的临床检查，评估粘膜颜色，毛细血管再充盈时间，全球肺癌和使用听诊器心脏功能和体温用直肠体温计。 </li><li>感应Ó之前施用的咪达唑仑（肌内，0.5毫克/千克）和盐酸吗啡（肌内，0.1毫克/千克）15至30分钟的注射˚F麻醉。感应每4至6小时期间重复注射吗啡盐酸盐（0.05至0.5毫克/千克，肌肉注射）的。 </li><li>通过初始推注5mg / kg的然后通过局部注射施用硫喷妥钠（10毫克/公斤，静脉内），直到它是有效的。由包按照此和面罩通气，直至气管插管。 </li><li>仔猪插管<ol><li>润滑与pramocaine硅胶管。插入金属手写笔入管变硬并有利于气管插管的过程。 </li><li>轻触眼睑，以确保深麻醉。由喉部直接可视化使用喉镜，以提高舌头，避免伤到声带进行气管插管。充气管球囊，以防止反流有关的问题。 </li></ol></li><li>控制呼吸频率为每分钟10-12次呼吸; 7-10毫升/千克当前卷，25〜30厘米的H 2 O和2秒的吸气阶段带有正Ê吹入压力5厘米H 2 O的第二呼气压力</li><li>麻醉动物用异氟烷在100％的氧气（感应3-5％的2至3升/分钟，维持在1.5-2.5％的氧气流以1升/分钟氧气流量）。适用卡波姆凝胶角膜如步骤1.5.1。 </li><li>插入一个肝素导管进入尾耳动脉经皮下输注绿色套管固定就位用缝合针（5毫升0.9％盐水的掺有5000国际单位/毫升肝素）。 </li><li>注入乳酸林格氏溶液（10-20毫升/公斤/小时）。 </li><li>将动物在一个稍微倾斜的体检表。保持头部稍微倾斜向下，促进唾液流动。 </li></ol></li><li>监控<ol><li>每隔5分钟，检查并在个别麻醉病例报告记录下列值:粘膜颜色和毛细血管再充盈时间，下颌肌张力和眼球位置，瞳孔缩小/瞳孔散大，眼睑反射。 </li><li>持续监测心脏和respira保守党率，脉搏血氧饱和度，体温和心电图。插入动脉插管插入股动脉。插入一个大口径长10厘米的导管，插入股动脉和监控系统血压。 </li></ol></li><li>设置脉动导管 注意:此型医疗装置包括两个导管放置侧由端和焊接在一起。第一个的末端部分被连接到一个标准的气球，直径为20毫米和5毫升的最大体积容量。第二导管能够插入电线以方便定位在肺动脉。该装置为750mm长，0.035内部表和12 Fr的外径<ol><li>对于该实验，用小哈佛683动物呼吸机，其中放气和正压期间施加有源真空到气囊在充气期间，为2.5毫升每脉冲的体积进行脉动。使用氦气作为喷射气体的球OON泵，以防止气体栓塞。 </li><li>连续记录心电图表面的描记协议期间记录任何心脏节律紊乱。在左股动脉中，连接的传感器设备，以一个20号的导管，连接到血液动力学监测系统。 </li></ol></li><li>右心脏导管插入术<ol><li>洗脸，刮胡子动物的脖子。用清洁纱布敷用皮肤消毒液（聚维酮碘擦洗）皮肤。划定的手术部位，放置无菌单周围右肩和胸骨柄sterni之间的右颈。 </li><li>使4cm的纵行切口用无菌剪刀右肩和柄sterni之间的中途。 </li><li>小心地用钳子取出皮肤和肌肉层。然后轻轻取出周围静脉超过大约5厘米长的结缔组织。片段中的远侧，以防止出血。将粘合剂绕丝PRoximal侧，以便能够控制后半部分中的静脉的开口。 </li><li>使用特定小很尖凿，切静脉半横向。确保切口边缘整齐。使用细沫，提高切口的一边，轻轻地按压导管插入静脉的近侧。控制出血与粘合剂线。 </li><li>引入导管进入颈内静脉，并通过上腔静脉，右心房，右心室，最后，肺动脉相继推。 </li><li>记录的压力超过10稳定的心动周期，每个心腔和肺动脉（T 0）。测量心脏血液流动三次以1分钟的间隔。 </li><li>定位的气囊导管在根据放射线控制肺动脉。充气和放气的气球（脉动）用氦气1厘米3。继续脉动10分钟。超过10稳定心动周期记录的压力，在每个心脏腔和肺ARTERY和测量10分钟后（T1）的心脏的血流。 </li><li>再次测量心脏血流搏动协议（T2），30分钟后。 </li></ol></li></ol> 3.无测量<ol><li>连接道格拉斯袋呼出的呼吸气体出口管，直到它被完全填充。将它放置在管内一个没有气息分析仪。 </li><li>慢慢地，不断地通过压缩弹性道格拉斯包迫使呼出的空气进入分析仪。测量袋流出通过分析仪的流量计，以保持恒定的流量。 </li></ol> 4.组织学测量<ol><li>在麻醉下，注入30毫升Dolethal的（安乐死注射液）到搏动导管置于心脏。 </li><li>立即安乐死后，打开胸部纵向锯开胸骨柄，稍微动心脏按住右肺部。然后用解剖剪刀，切两2〜3厘米的MI 3个样品肺ddle叶。 </li><li>卡扣冷冻在液氮中一个样品并储存在-80℃。固定在3.7％多聚甲醛中24小时的第二样品，然后嵌入石蜡进行后续组织学分析。 </li><li>执行如先前所述16组织学测量。测量PA壁厚如下:2次测量/动脉血管10 /仔猪和6头/组。 </li><li>进行统计分析。数值表示为倍数变化±SEM表示。 <ol><li>为了比较两种方法，使用非配对t检验。为了比较两个以上的装置，可使用单向ANOVA，然后通过一个唐恩的测试。在P &lt;0.05为差异有显著（*）。 </li></ol></li></ol>

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F. <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+pulmonary+arterial+hypertension:+the+hope+for+etiological+discovery+and+pharmacological+cure.">Animal models of pulmonary arterial hypertension: the hope for etiological discovery and pharmacological cure.</a> Am J Physiol Lung Cell Mol Physiol. 297 (6), L1013-L1032 (2009).</li><li>Torre-Amione, G., 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=Reversal+of+secondary+pulmonary+hypertension+by+axial+and+pulsatile+mechanical+circulatory+support.">Reversal of secondary pulmonary hypertension by axial and pulsatile mechanical circulatory support.</a> J Heart Lung Transplant. 29 (2), 195-200 (2010).</li></ol>

The authors would like to thank the INRA institute, which housed the piglets, and Cardio Innovating systems for the funding.

对于第一次，它已被证明其在变化肺脉动流被因果相关的PAH的继发慢性缺氧暴露的发展。这种转换方法提供的证据表明诱导人工增加使用一个特别设计的导管肺脉动流改善肺动脉高血压，可能是通过增加NO的产生。 这些发现不仅原来，他们是伟大的治疗意义也证明了内源性NO的产生可以机械，安全地刺激了肺循环中，而不会影响系统的功能。研究设计符合最新的临床前研究建?...

肺动脉高血压是一种威胁生命的疾病，影响肺血管。有协议认为增加血管收缩剂（内皮素，血清素）和血管扩张剂的减少（NO，前列环素）之间的不平衡有助于PAH的发展领域。随着时间的推移，这种亲缩窄型演变成一个复杂的亲增殖和抗凋亡表型，促进血管病变的发展1。 长时间暴露于血管收缩导致的[Ca 2+]一个显著和持续增加i的肺动脉平滑肌细胞，允许几个钙调节转录因子，如NFAT 2-4的活化，促进PASMC增殖和耐一细胞凋亡表型5。这种表型导致肺血管病变，有助于增加在两个PA压和肺动脉水库istance，最终导致致命右心脏衰竭6。 目前，存在着逆转PAH虽然有几个是改善生活7患者的生存质量提供任何治疗。在这些处理中，吸入NO治疗的有效性已经被证实，但由于其半衰期短的，很难在临床实践中使用。出于这个原因，更稳定和持久的治疗方法已被优选的，如前列环素类似物，或内皮素受体 ​​阻断剂7。为了开发更好的治疗方法，有必要改进和扩大PAH的病理生理学的知识。 搏动是公知的刺激激活切应力诱发的血管扩张，保护从高压侧流损伤8,9-非弹性远端动脉。在PAH的二次模型来体肺分流手术，努尔等证明肺内剪STRESS介导的血管内皮功能的增强10。几个研究已经证明，NO，前列环素和ET-1的表达是紧密的变化脉动流调节。实际上，适度增加脉动流增加eNOS活性和前列环素的水平，这两者都降低了肺动脉高压。脉动流调制可能牵连PAH的病因和人为地增加这是一个有吸引力的和新颖的肺循环中增加NO和前列环素的生产方式。 本研究旨在评估10分钟搏动的影响，用流动的血流动力学测量了新开发的脉动导管在肺动脉高压（PH）模型在仔猪在其中缺氧已致。据推测，增加肺动脉搏动引起肺动脉的血管舒张，因此降低肺动脉压力。 右心脏猫heterization（RHC）是诊断的关键临床干预和随访PAH患者。事实上，它是诊断肺动脉高压，并允许医生评估血管反应性11,12以及疾病进展的最可靠的方法。其实每一个PAH患者接受RHC几次。本研究在大型动物的目的是展示脉动导管的疗效和安全性评估和有规律RHC过程期间处理的PAH。因为脉动导管已经上市和RHC是在PAH患者常规进行，该研究提供了所有能够迅速进行临床试验所需的信息。

<table><tbody><tr><td>Drugs for anesthesia</td><td></td><td></td><td></td></tr><tr><td>sodium thiopental, THIOPENTAL SODIUM</td><td>Abbott, France</td><td>0000071-73-8</td><td>powder&lt;br/&gt; 3 place Gustave Eiffe 94518 RUNGIS CEDEX.</td></tr><tr><td>&amp;nbsp;isoflurane, FORANE</td><td>Abbott, France</td><td>05260-05</td><td>glass bottle 250 ml&lt;br/&gt; 3 place Gustave Eiffe 94518 RUNGIS CEDEX.</td></tr><tr><td>midazolam, Hypnovel</td><td>Accord Healthcare&amp;nbsp;</td><td>Vidal</td><td>injectable ampoules 1mg/ml&lt;br/&gt; 45 Rue du Faubourg de Roubaix 59000 Lille France</td></tr><tr><td>pramocaine,TRONOTHANE 1%&amp;nbsp;</td><td>Laboratoires LISAPHARM</td><td>Vidal</td><td>Gel appl locale T/30g&lt;br/&gt; 3, rue Scheffer. 75016 Paris.</td></tr><tr><td>morphine chlohydrate Lavoisier</td><td>CMD Lavoisier Laboratoires CHAIX et DU MARAIS</td><td>Vidal</td><td>injectable ampoules&amp;nbsp;&lt;br/&gt; 7, rue Labie -75017 Paris - France</td></tr><tr><td>Acrylates Copolymer-Carbopol&amp;reg; Aqua SF-1 Polymer</td><td>Lubrizol</td><td></td><td>gel appl local&lt;br/&gt; Elys&amp;eacute;es La D&amp;eacute;fense 19 le Parvis 92073 Paris la d&amp;eacute;fense</td></tr><tr><td>Material&amp;nbsp;</td><td></td><td></td><td></td></tr><tr><td>Ventilateur Harvard 683</td><td>Harvard apparatus</td><td>Harvard apparatus</td><td>DRIM 75 rue des Anglais - 78700 Conflans Ste Honorine &amp;nbsp;&amp;nbsp;</td></tr><tr><td>Echographe Voluson E8 with a 3.5 MHz probe</td><td>General Electric</td><td>GEHealthcare</td><td>DRIM 75 rue des Anglais - 78700 Conflans Ste Honorine &amp;nbsp;&amp;nbsp;</td></tr><tr><td>Pulsatil Catheter</td><td>Cardio inovating system</td><td></td><td>Cardio innovative systems, 33 rue Vivienne, Paris, France 75002</td></tr><tr><td>NO breath Analyseur</td><td>Respur</td><td>Respur</td><td>26 rue Felix Rouget 95490 Vaureal France</td></tr></tbody></table>

increasing pulmonary artery pulsatile flow improves hypoxic pulmonary hypertension in piglets

肺动脉高压（PAH）是一种疾病，影响远端肺动脉（PA）。这些动脉发生变形，导致右心衰竭。目前的治疗是有限的。生理学上，脉动血流是不利的脉管。响应于持续脉动应力，血管释放一氧化氮（NO）来诱导血管舒张自我保护。基于这一观察，本研究开发的协议，以评估一个人工肺脉动血流是否能诱导肺动脉压的编号为依赖性减少。一组小猪暴露于慢性缺氧3周和相比仔猪的对照组。每周一次，仔猪进行超声心动图检查，以评估PAH的严重程度。在缺氧暴露结束时，仔猪经使用一种脉动导管脉动协议。之后被麻醉并为手术准备，​​仔猪的颈静脉分离并在CAtheter通过右心房，右心室和肺动脉，下放射线控制引入。肺动脉压（PAP）的前（T0）测定，（T1）和后（T 2）的搏动协议30分钟后立即使用。已证明，这种脉动协议是经由NO依赖性机制诱导显著减少平均PAP的安全和有效的方法。这些数据开辟PAH的临床管理的新途径。

增加肺动脉脉动流改善大鼠慢性缺氧性肺动脉高压的仔猪之前暴露的动物的增加脉动流，超声被用于非侵入性地检查该仔猪已经开发肺动脉高压。 如图2三周慢性缺氧诱发的肺动脉高压的仔猪，其特征在于一个显著降低肺动脉加速时间（多普勒），并增加了右心室肥大（M型）的开发。在RHC，双方右心室收缩压（RVSP），平均PA压微创测量证实肺动脉高压的慢性缺氧?...

Watch this Scientific Journal Video about Increasing Pulmonary Artery Pulsatile Flow Improves Hypoxic Pulmonary Hypertension in Piglets at JoVE.com

增加肺动脉脉动流量可改善仔猪缺氧性肺动脉高压医学|视频

Increasing Pulmonary Artery Pulsatile Flow Improves Hypoxic Pulmonary Hypertension in Piglets

Pulmonary hypertension is associated with a significant reduction in pulmonary artery pulsatility, contractility and elasticity, contributing to an increase in pulmonary artery pressure and pulmonary resistance. Using a hypoxic piglet model, this study demonstrated that improving pulmonary artery plasticity using a newly developed pulsatile catheter improves hypoxic pulmonary hypertension.

增加肺动脉脉动流改善缺氧性肺动脉高压的仔猪

Pulmonary arterial hypertension (PAH) is a disease affecting distal pulmonary arteries (PA). These arteries are deformed, leading to right ventricular ...

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14.1K 次观看.  Laval University. Pulmonary hypertension is associated with a significant reduction in pulmonary artery pulsatility, contractility and elasticity, contributing to an increase in pulmonary artery pressure and pulmonary resistance. Using a hypoxic piglet model, this study demonstrated that improving pulmonary artery plasticity using a newly developed pulsatile catheter improves hypoxic pulmonary hypertension.

视频: Increasing Pulmonary Artery Pulsatile Flow Improves Hypoxic Pulmonary Hypertension in Piglets

A Swine Model of Neonatal Asphyxia

Annually more than 1 million neonates die worldwide as related to asphyxia. Asphyxiated neonates commonly have multi-organ failure including hypotension, perfusion deficit, hypoxic-ischemic encephalopathy, pulmonary hypertension, vasculopathic enterocolitis, renal failure and thrombo-embolic complications. Animal models are developed to help us understand the patho-physiology and pharmacology of neonatal asphyxia. In comparison to rodents and newborn lambs, the newborn piglet has been proven to be a valuable model. The newborn piglet has several advantages including similar development as that of 36-38 weeks human fetus with comparable body systems, large body size (&tilde;1.5-2 kg at birth) that allows the instrumentation and monitoring of the animal and controls the confounding variables of hypoxia and hemodynamic derangements.
We here describe an experimental protocol to simulate neonatal asphyxia and allow us to examine the systemic and regional hemodynamic changes during the asphyxiating and reoxygenation process as well as the respective effects of interventions. Further, the model has the advantage of studying multi-organ failure or dysfunction simultaneously and the interaction with various body systems. The experimental model is a non-survival procedure that involves the surgical instrumentation of newborn piglets (1-3 day-old and 1.5-2.5 kg weight, mixed breed) to allow the establishment of mechanical ventilation, vascular (arterial and central venous) access and the placement of catheters and flow probes (Transonic Inc.) for the continuously monitoring of intra-vascular pressure and blood flow across different arteries including main pulmonary, common carotid, superior mesenteric and left renal arteries. Using these surgically instrumented piglets, after stabilization for 30-60 minutes as defined by Z&lt;10% variation in hemodynamic parameters and normal blood gases, we commence an experimental protocol of severe hypoxemia which is induced via normocapnic alveolar hypoxia. The piglet is ventilated with 10-15% oxygen by increasing the inhaled concentration of nitrogen gas for 2h, aiming for arterial oxygen saturations of 30-40%. This degree of hypoxemia will produce clinical asphyxia with severe metabolic acidosis, systemic hypotension and cardiogenic shock with hypoperfusion to vital organs. The hypoxia is followed by reoxygenation with 100% oxygen for 0.5h and then 21% oxygen for 3.5h. Pharmacologic interventions can be introduced in due course and their effects investigated in a blinded, block-randomized fashion.

Large animal models have good translational values in the examination of physiology and pharmacology of neonatal asphyxia. Using newborn piglets, we develop an experimental protocol to simulate neonatal asphyxia which has advantages of studying the systemic and regional hemodynamics, oxygen transport with biochemical and pathologic pathways and correlations.

新生儿窒息了猪模型

Annually more than 1 million neonates die worldwide as related to asphyxia. Asphyxiated neonates commonly have multi-organ failure including hypotension, ...

科研

医学

Chronic Thromboembolic Pulmonary Hypertension and Assessment of Right Ventricular Function in the Piglet

An original piglet model of Chronic Thromboembolic Pulmonary Hypertension (CTEPH) associated with chronic Right Ventricular (RV) dysfunction is described. Pulmonary Hypertension (PH) was induced in 3-week-old piglets by a progressive obstruction of the pulmonary vascular bed. A ligation of the left Pulmonary Artery (PA) was performed first through a mini-thoracotomy. Second, weekly embolizations of the right lower pulmonary lobe were done under fluoroscopic guidance with n-butyl-2-cyanoacrylate during 5 weeks. Mean Pulmonary Arterial Pressure (mPAP) measured by ritght heart catheterism, increased progressively, as well as Right Atrial pressure and Pulmonary Vascular Resistances (PVR) after 5 weeks compared to sham animals. Right Ventricular (RV) structural and functional remodeling were assessed by transthoracic echocardiography (RV diameters, RV wall thickness, RV systolic function). RV elastance and RV-pulmonary coupling were assessed by Pressure-Volume Loops (PVL) analysis with conductance method. Histologic study of the lung and the right ventricle were also performed. Molecular analyses on RV fresh tissues could be performed through repeated transcutaneous endomyocardial biopsies. Pulmonary microvascular disease in obstructed and unobstructed territories was studied from lung biopsies using molecular analyses and pathology. Furthermore, the reliability and the reproducibility was associated with a range of PH severity in animals. Most aspects of the human CTEPH disease were reproduced in this model, which allows new perspectives for the understanding of the underlying mechanisms (mitochondria, inflammation) and new therapeutic approaches (targeted, cellular or gene therapies) of the overloaded right ventricle but also pulmonary microvascular disease.

Chronic Thromboembolic Pulmonary Hypertension (CTEPH) and Right Ventricular (RV) dysfunction were induced in piglets by progressive obstruction of the pulmonary arteries. Consequences were remarkably similar to those observed in CTEPH patients. This animal model would be a very useful tool for pathophysiology and therapeutic experiments on CTEPH and RV failure.

慢性血栓栓塞性肺动脉高压和右心室功能的仔猪评估

An original piglet model of Chronic Thromboembolic Pulmonary Hypertension (CTEPH) associated with chronic Right Ventricular (RV) dysfunction is described. ...

Hemodynamic Characterization of Rodent Models of Pulmonary Arterial Hypertension

Pulmonary arterial hypertension (PAH) is a rare disease of the pulmonary vasculature characterized by endothelial cell apoptosis, smooth muscle proliferation and obliteration of pulmonary arterioles. This in turn results in right ventricular (RV) failure, with significant morbidity and mortality. Rodent models of PAH, in the mouse and the rat, are important for understanding the pathophysiology underlying this rare disease. Notably, different models of PAH may be associated with different degrees of pulmonary hypertension, RV hypertrophy and RV failure. Therefore, a complete hemodynamic characterization of mice and rats with PAH is critical in determining the effects of drugs or genetic modifications on the disease. 
Here we demonstrate standard procedures for assessment of right ventricular function and hemodynamics in both rat and mouse PAH models. Echocardiography is useful in determining RV function in rats, although obtaining standard views of the right ventricle is challenging in the awake mouse. Access for right heart catheterization is obtained by the internal jugular vein in closed-chest mice and rats. Pressures can be measured using polyethylene tubing with a fluid pressure transducer or a miniature micromanometer pressure catheter. Pressure-volume loop analysis can be performed in the open chest. After obtaining hemodynamics, the rodent is euthanized. The heart can be dissected to separate the RV free wall from the left ventricle (LV) and septum, allowing an assessment of RV hypertrophy using the Fulton index (RV/(LV+S)). Then samples can be harvested from the heart, lungs and other tissues as needed.

Pulmonary arterial hypertension (PAH) is a disease of pulmonary arterioles that leads to their obliteration and the development of right ventricular failure. Rodent models of PAH are critical in understanding the pathophysiology of PAH. Here we demonstrate hemodynamic characterization, with right heart catheterization and echocardiography, in the mouse and rat.

肺动脉高压的啮齿动物模型的血流动力学特性

Pulmonary arterial hypertension (PAH) is a rare disease of the pulmonary vasculature characterized by endothelial cell apoptosis, smooth muscle ...

Shunt Surgery, Right Heart Catheterization, and Vascular Morphometry in a Rat Model for Flow-induced Pulmonary Arterial Hypertension

In this protocol, PAH is induced by combining a 60 mg/kg monocrotalin (MCT) injection with increased pulmonary blood flow through an aorto-caval shunt (MCT+Flow). The shunt is created by inserting an 18-G needle from the abdominal aorta into the adjacent caval vein. Increased pulmonary flow has been demonstrated as an essential trigger for a severe form of PAH with distinct phases of disease progression, characterized by early medial hypertrophy followed by neointimal lesions and the progressive occlusion of the small pulmonary vessels. To measure the right heart and pulmonary hemodynamics in this model, right heart catheterization is performed by inserting a rigid cannula containing a flexible ball-tip catheter via the right jugular vein into the right ventricle. The catheter is then advanced into the main and the more distal pulmonary arteries. The histopathology of the pulmonary vasculature is assessed qualitatively, by scoring the pre- and intra-acinar vessels on the degree of muscularization and the presence of a neointima, and quantitatively, by measuring the wall thickness, the wall-lumen ratios, and the occlusion score.

This protocol describes a surgical procedure to create a model for flow-induced pulmonary arterial hypertension (PAH) in rats and the procedures to analyze the principle hemodynamic and histological end-points in this model.

分流手术，右心导管，血管形态计量学在大鼠模型中的流量引起的肺动脉高压

In this protocol, PAH is induced by combining a 60 mg/kg monocrotalin (MCT) injection with increased pulmonary blood flow through an aorto-caval shunt ...

A Large Animal Model for Pulmonary Hypertension and Right Ventricular Failure: Left Pulmonary Artery Ligation and Progressive Main Pulmonary Artery Banding in Sheep

Decompensated right ventricular failure (RVF) in pulmonary hypertension (PH) is fatal, with limited medical treatment options. Developing and testing novel therapeutics for PH requires a clinically relevant large animal model of increased pulmonary vascular resistance and RVF. This manuscript discusses the latest development of the previously published ovine PH-RVF model that utilizes left pulmonary artery (PA) ligation and main PA occlusion. This model of PH-RVF is a versatile platform to control not only the disease severity but also the RV&#39;s phenotypic response.
Adult sheep (60-80 kg) underwent left PA (LPA) ligation, placement of main PA cuff, and insertion of RV pressure monitor. PA cuff and RV pressure monitor were connected to subcutaneous ports. Subjects underwent progressive PA banding twice per week for 9 weeks with sequential measures of RV pressure, PA cuff pressures, and mixed venous blood gas (SvO2). At the initiation and endpoint of this model, ventricular function and dimensions were assessed using echocardiography. In a representative group of 12 animal subjects, RV mean and systolic pressure increased from 28 &#177; 5 and 57 &#177; 7 mmHg at week 1, respectively, to 44 &#177; 7 and 93 &#177; 18 mmHg (mean &#177; standard deviation) by week 9. Echocardiography demonstrated characteristic findings of PH-RVF, notably RV dilation, increased wall thickness, and septal bowing. The longitudinal trend of SvO2 and PA cuff pressure demonstrates that the rate of PA banding can be titrated to elicit varying RV phenotypes. A faster PA banding strategy led to a precipitous decline in SvO2 &#60; 65%, indicating RV decompensation, whereas a slower, more paced strategy led to the maintenance of physiologic SvO2 at 70%-80%. One animal that experienced the accelerated strategy developed several liters of pleural effusion and ascites by week 9. This chronic PH-RVF model provides a valuable tool for studying molecular mechanisms, developing diagnostic biomarkers, and enabling therapeutic innovation to manage RV adaptation and maladaptation from PH.

This manuscript describes the surgical technique and experimental approach to develop severe right ventricular pressure overload to model their adaptive and maladaptive phenotypes.

肺动脉高压与右心室衰竭的大型动物模型:绵羊左肺动脉结扎与进行性主肺动脉束带

Decompensated right ventricular failure (RVF) in pulmonary hypertension (PH) is fatal, with limited medical treatment options. Developing and testing ...

生物工程

Invasive Hemodynamic Monitoring of Aortic and Pulmonary Artery Hemodynamics in a Large Animal Model of ARDS

One of the leading causes of morbidity and mortality in patients with heart failure is right ventricular (RV) dysfunction, especially if it is due to pulmonary hypertension. For a better understanding and treatment of this disease, precise hemodynamic monitoring of left and right ventricular parameters is important. For this reason, it is essential to establish experimental pig models of cardiac hemodynamics and measurements for research purpose. 
This article shows the induction of ARDS by using oleic acid (OA) and consequent right ventricular dysfunction, as well as the instrumentation of the pigs and the data acquisition process that is needed to assess hemodynamic parameters. To achieve right ventricular dysfunction, we used oleic acid (OA) to cause ARDS and accompanied this with pulmonary artery hypertension (PAH). With this model of PAH and consecutive right ventricular dysfunction, many hemodynamic parameters can be measured, and right ventricular volume load can be detected. 
All vital parameters, including respiratory rate (RR), heart rate (HR) and body temperature were recorded throughout the whole experiment. Hemodynamic parameters including femoral artery pressure (FAP), aortic pressure (AP), right ventricular pressure (peak systolic, end systolic and end diastolic right ventricular pressure), central venous pressure (CVP), pulmonary artery pressure (PAP) and left arterial pressure (LAP) were measured as well as perfusion parameters including ascending aortic flow (AAF) and pulmonary artery flow (PAF). Hemodynamic measurements were performed using transcardiopulmonary thermodilution to provide cardiac output (CO). Furthermore, the PiCCO2 system (Pulse Contour Cardiac Output System 2) was used to receive parameters such as stroke volume variance (SVV), pulse pressure variance (PPV), as well as extravascular lung water (EVLW) and global end-diastolic volume (GEDV). Our monitoring procedure is suitable for detecting right ventricular dysfunction and monitoring hemodynamic findings before and after volume administration.

We present a protocol of creating right ventricular dysfunction in a pig model by inducing ARDS. We demonstrate invasive monitoring of left and right ventricular cardiac output using flow probes around the aorta and the pulmonary artery, as well as blood pressure measurements in the aorta and pulmonary artery.

ards 大型动物模型主动脉和肺动脉血流动力学的侵袭血流动力学监测

One of the leading causes of morbidity and mortality in patients with heart failure is right ventricular (RV) dysfunction, especially if it is due to ...

Induction and Phenotyping of Acute Right Heart Failure in a Large Animal Model of Chronic Thromboembolic Pulmonary Hypertension

The development of acute right heart failure (ARHF) in the context of chronic pulmonary hypertension (PH) is associated with poor short-term outcomes. The morphological and functional phenotyping of the right ventricle is of particular importance in the context of hemodynamic compromise in patients with ARHF. Here, we describe a method to induce ARHF in a previously described large animal model of chronic PH, and to phenotype, dynamically, right ventricular function using the gold standard method (i.e., pressure-volume PV loops) and with a non-invasive clinically available method (i.e., echocardiography). Chronic PH is first induced in pigs by left pulmonary artery ligation and right lower lobe embolism with biological glue once a week for 5 weeks. After 16 weeks, ARHF is induced by successive volume loading using saline followed by iterative pulmonary embolism until the ratio of the systolic pulmonary pressure over systemic pressure reaches 0.9 or until the systolic systemic pressure decreases below 90 mmHg. Hemodynamics are restored with dobutamine infusion (from 2.5 &#181;g/kg/min to 7.5 &#181;g/kg/min). PV-loops and echocardiography are performed during each condition. Each condition requires around 40 minutes for induction, hemodynamic stabilization and data acquisition. Out of 9 animals, 2 died immediately after pulmonary embolism and 7 completed the protocol, which illustrates the learning curve of the model. The model induced a 3-fold increase in mean pulmonary artery pressure. The PV-loop analysis showed that ventriculo-arterial coupling was preserved after volume loading, decreased after acute pulmonary embolism and was restored with dobutamine. Echocardiographic acquisitions allowed to quantify right ventricular parameters of morphology and function with good quality. We identified right ventricular ischemic lesions in the model. The model can be used to compare different treatments or to validate non-invasive parameters of right ventricular morphology and function in the context of ARHF.

We present a protocol to induce and phenotype an acute right heart failure in a large animal model with chronic pulmonary hypertension. This model can be used to test therapeutic interventions, to develop right heart metrics&#160;or to improve the understanding of acute right heart failure pathophysiology.

慢性血栓栓塞性肺动脉高压大型动物模型中急性右心衰竭的诱导和表型分析

The development of acute right heart failure (ARHF) in the context of chronic pulmonary hypertension (PH) is associated with poor short-term outcomes. The ...

The Left Pneumonectomy Combined with Monocrotaline or Sugen as a Model of Pulmonary Hypertension in Rats

In this protocol, we detail the correct procedural steps and necessary precautions to successfully perform a left pneumonectomy and induce PAH in rats with the additional administration of monocrotaline (MCT) or SU5416 (Sugen). We also compare these two models to other PAH models commonly used in research. In the last few years, the focus of animal PAH models has moved towards studying the mechanism of angioproliferation of plexiform lesions, in which the role of increased pulmonary blood flow is considered as an important trigger in the development of severe pulmonary vascular remodeling. One of the most promising rodent models of increased pulmonary flow is the unilateral left pneumonectomy combined with a "second hit" of MCT or Sugen. The removal of the left lung leads to increased and turbulent pulmonary blood flow and vascular remodeling. Currently, there is no detailed procedure of the pneumonectomy surgery in rats. This article details a step-by-step protocol of the pneumonectomy surgical procedure and post-operative care in male Sprague-Dawley rats. Briefly, the animal is anesthetized and the chest is opened. Once the left pulmonary artery, pulmonary vein, and bronchus are visualized, they are ligated and the left lung is removed. The chest then closed and the animal recovered. Blood is forced to circulate only on the right lung. This increased vascular pressure leads to a progressive remodeling and occlusion of small pulmonary arteries. The second hit of MCT or Sugen is used one week post-surgery to induce endothelial dysfunction. The combination of increased blood flow in the lung and endothelial dysfunction produces severe PAH. The primary limitation of this procedure is that it requires general surgical skills.

The rodent left pneumonectomy is a valuable technique in pulmonary hypertension research. Here, we present a protocol to describe the rat pneumonectomy procedure and postoperative care to ensure minimal morbidity and mortality.

左肺切除术联合单氯联氨酸或苏根为大鼠肺动脉高压的模型

In this protocol, we detail the correct procedural steps and necessary precautions to successfully perform a left pneumonectomy and induce PAH in rats ...

Isolation of Pulmonary Artery Smooth Muscle Cells from Neonatal Mice

Pulmonary hypertension is a significant cause of morbidity and mortality in infants. Historically, there has been significant study of the signaling pathways involved in vascular smooth muscle contraction in PASMC from fetal sheep. While sheep make an excellent model of term pulmonary hypertension, they are very expensive and lack the advantage of genetic manipulation found in mice. Conversely, the inability to isolate PASMC from mice was a significant limitation of that system. Here we described the isolation of primary cultures of mouse PASMC from P7, P14, and P21 mice using a variation of the previously described technique of Marshall et al.26 that was previously used to isolate rat PASMC. These murine PASMC represent a novel tool for the study of signaling pathways in the neonatal period. Briefly, a slurry of 0.5% (w/v) agarose + 0.5% iron particles in M199 media is infused into the pulmonary vascular bed via the right ventricle (RV). The iron particles are 0.2 &#956;M in diameter and cannot pass through the pulmonary capillary bed. Thus, the iron lodges in the small pulmonary arteries (PA). The lungs are inflated with agarose, removed and dissociated. The iron-containing vessels are pulled down with a magnet. After collagenase (80 U/ml) treatment and further dissociation, the vessels are put into a tissue culture dish in M199 media containing 20% fetal bovine serum (FBS), and antibiotics (M199 complete media) to allow cell migration onto the culture dish. This initial plate of cells is a 50-50 mixture of fibroblasts and PASMC. Thus, the pull down procedure is repeated multiple times to achieve a more pure PASMC population and remove any residual iron. Smooth muscle cell identity is confirmed by immunostaining for smooth muscle myosin and desmin.

We have developed a novel and reproducible technique to isolate primary cultures of pulmonary artery smooth muscle cells (PASMC) from mice as young as P7, thereby allowing better study of the signaling pathways involved in neonatal smooth muscle cell contraction and relaxation.

肺动脉平滑肌细胞从新生小鼠隔离

Pulmonary hypertension is a significant cause of morbidity and mortality in infants. Historically, there has been significant study of the signaling ...

生物学

Multiple Intravenous Bolus Dosing and Invasive Hemodynamic Assessment in a Hypoxia-Induced Mouse Pulmonary Artery Hypertension Model

Pulmonary arterial hypertension (PAH) is a progressive life-threatening disease, primarily affecting small pulmonary arterioles of the lung. Currently, there is no cure for PAH. It is important to discover new compounds that can be used to treat PAH. The mouse hypoxia-induced PAH model is a widely used model for PAH research. This model recapitulates human clinical manifestations of PAH Group 3 disease and is an important research tool to evaluate the effectiveness of new experimental therapies for PAH. Research using this model often requires the administration of compounds in mice. For a compound that needs to be given directly into the bloodstream, optimizing intravenous (IV) administration is a key part of the experimental procedures. Ideally, the IV injection system should permit multiple injections over a set time course. Although the mouse hypoxia-induced PAH model is very popular in many laboratories, it is technically challenging to perform multiple IV bolus dosing and invasive hemodynamic assessment in this model. In this protocol, we present step-by-step instructions on how to carry out multiple IV bolus dosing via mouse jugular vein and perform arterial and right ventricle catheterization for hemodynamic assessment in mouse hypoxia-induced PAH model.

This protocol provides a step-by-step procedure for executing multiple intravenous bolus dose administration and invasive hemodynamic monitoring in mice. Investigators can use this protocol for future therapeutic compound screening for pulmonary artery hypertension.

缺氧诱导小鼠肺动脉高压模型中的多次静脉推注给药和侵入性血流动力学评估

Pulmonary arterial hypertension (PAH) is a progressive life-threatening disease, primarily affecting small pulmonary arterioles of the lung. Currently, ...