Name: rat heterotopic abdominal heart/single-lung transplantation in a volume-loaded configuration
Uploaded: 2015-05-29T13:00:00.000+00:00
Duration: 6 min 16 s
Description: Watch this Scientific Journal Video about Rat Heterotopic Abdominal Heart/Single-lung Transplantation in a Volume-loaded Configuration at JoVE.com

马克J.卡恩斯接收来自UBC临床医生调查程序（通过医生和加拿大皇家外科学院授予），以及UBC 4YF博士生奖学金的支持。 约翰·博伊德H.是国家疗养院协会和迈克尔·史密斯基金会健康研究学者。通过资金CIHR。作者要感谢M.阿拉德博士和Richard Wamboldt他们与安装和维护设备的灌注援助。

所有的动物饲养和照顾按照国家和护理和使用实验动物的制度准则。伦理批准该协议被授予由不列颠哥伦比亚省的动物护理委员会的大学。男，SD大鼠的体重300 - 450克的用于本协议。 1.供体手术<ol><li>有有约100ml心麻痹（RT）的一个圆筒形烧瓶通过三路活塞相连长静脉（IV）导管管材。用支架提升烧瓶约80厘米工作表面上方，靠重力使心脏停搏交付。 </li><li>为了可视化结构充分，使用一对手术放大镜或解剖显微镜。 注:我们目前使用的是双目手术显微镜3.4 - 21.3X倍。 </li><li>将供体的麻醉室，诱导麻醉与4 - 5％异氟醚。 </l我&gt; <li>老鼠传输到操作平台，并维持麻醉用鼻锥与1 - 2％异氟醚。适用兽医药膏动物的眼睛，以防止干燥。辖咪达唑仑（2毫克/千克）腹膜内与地下25针。 </li><li>使用手术剪，刮捐助从剑突下颌骨来。剃左腹股沟的一个小区域（用于后续静脉通路）。应用脱毛剂对手术的表面，等待大约5分钟，然后取出头发用一块纱布。 </li><li>准备手术部位用碘伏或chlorexidine的解决方案（我们只用洗必泰）。渗透切口部位用0.1 - 0.5％利多卡因皮下。 </li><li>固定前肢和后肢留下的操作平台用胶带，留下正确的后肢免费监测麻醉深度和生命体征。 </li><li>在确保适当的麻醉深度踏板掐，用做左侧腹股沟折痕切口22刀片手术刀。受钝性分离，暴露左股静脉和静脉获得访问如下: <ol><li>轻轻缩回组织覆盖的股静脉和导管插入带24克四静脉。四，将连接到的盐水充注式IV油管短的长度，并用胶带固定管到位。 注:在大鼠股静脉插管的程序由耶斯佩森和同事14别处所述。 </li><li>连接盐水充注10毫升注射器的静脉输液管，轻轻地吸入血液，以确保IV的正确定位。 </li><li>注入300 - 500 IU普通肝素通过静脉注射管，并随后冲洗管与3 - 用5ml盐水。 </li></ol></li><li>接下来，tracheotomize捐助如下: <ol><li>使用22刀片手术刀的颈静脉切迹和下颌之间的软组织正中切口。穿透甲状腺的胶囊中使用梅森中线鲍姆剪刀，和其单独使用叶钝性分离。 </li><li>使用钝性分离，分离颈部的带状肌在中线以暴露气管的前表面。 </li><li>用弯钳直言不讳地剖析围绕气管圆周面。包围4-0丝质领带气管。 </li><li>使用虹膜剪刀，作一横切口气管前，大约5毫米逊色于甲状软骨。轻轻地介绍了气管插管（一地下14 IV），并使用4-0真丝领带固定到位。 </li><li>气管插管连接到一个机械呼吸器。重定向氧和异氟醚通过呼吸回路的流量，并通风施主的速率和潮气量通过其自重13预测。 </li></ol></li><li>做一个切口在胸（使用22刀片手术刀）的中线，延长切口在颈静脉切迹至xipisternum下方。 </li><li>住在中线，执行使用骨刀正中胸骨切开术。缩回胸骨的边缘与一个自保持牵开器。进入心包和胸膜腔。 </li><li>执行胸腺切除术。它是最简单的第一直截了当地划分在中线胸腺，然后将其从使用钝性和锐性剥离的组合周围结构分离。 注:胸廓内动脉的起源可以解剖胸腺距上级胸骨边缘时受伤。防止出血，止血夹可在这些点除去胸腺之前被应用。 </li><li>使用梅岑鲍姆剪刀和/或尖锐的劳尔，剖析周围静脉脂肪距下腔静脉（IVC）。确保IVC是相对自由的脂肪和结缔组织从CAVO心房交界处优，在隔膜下方。 </li><li>使用锋利的劳尔，在圆周免费上腔静脉（SVC）和4-0丝质领带包围它。 </li><li>接下来，解剖左腔静脉不受周围结构，并且近端和远端用4-0丝质领带结扎它。切除静脉的中间部分，以暴露左锁骨下动脉。 </li><li>周向自由使用锋利的劳尔主动脉弓血管。适用近端和远端手术夹到无名动脉，并把它剪辑之间。离开左颈总动脉和左锁骨下动脉的未裁剪。 </li><li>接下来，导管插入SVC与24克静脉导管。固定导管与先前放置4-0真丝领带。 </li><li>准备一个权宜之计，而是温柔的移植收获。 </li><li>用锋利的剪刀，划分IVC只是优于膈面。划分SVC优于其插管部位，确保气道附近的结构没有受伤。 </li><li>关闭呼吸机并拔下气管插管。近端样气管。 </li><li>抓住气管横断用钳子，取出心脏和肺整块。这将需要温和牵引带锐性剥离作为心脏 - 肺移植物被除去。从底层食道分离气管。避免伤及降主动脉，使得它的一个长的部分保持收获后完好无损。 </li></ol> 2.准备心肺移植的注:在完成该过程的这一部分，助手应该是麻醉收件人啮齿动物和准备移植术。 <ol><li>放置在手术纱布的心脏，肺移植与朝下心脏的前表面。用锐性剥离，去除残留的食道，避免损伤后气管。 </li><li>找到胸降主动脉，并插入一个为16G钝头插管。 4-0真丝领带固定套管到位。 </li><li>定位主动脉弓血管，并应用一个手术夹子的左锁骨下动脉。离开左侧颈总动脉未裁剪子相继式的排气。 </li><li>由管搏连接到为16G主动脉插管管理停跳的首次剂量。颈动脉必须暂时堵塞用钳子以确保有足够的交付心脏停搏。 注:利益具有连接主动脉插管是可以搏根据需要不断地和/或间歇给药。对于间歇给药，我们通常提供停跳每10 - 5毫升丸药15分钟30 - 45秒​​。 </li><li>使用梅岑鲍姆剪刀和/或锋利的劳尔，解剖周围主动脉脂肪远离主动脉。确保主动脉是相对自由的脂肪和结缔组织从左锁骨下动脉（远端拱）与其插管部位。 </li><li>接下来，揭露使用锋利的劳尔左主支气管，并用4-0的丝质领带结扎近端它。采用样带虹膜剪左支气管远端结扎。结扎左肺动脉进行左全肺切除动脉和静脉4-0真丝领带。样远端结扎并取出左肺。 </li><li>如上述，删除所有其他肺叶除外右上和中间凸角。避免伤及IVC在执行肺叶切除术的过程。 </li><li>在心脏停搏管连接主动脉插管和管理不断停跳而接收者正在准备。将心脏，肺移植在无菌容器（ 如烧杯）。 </li></ol> 3.收件人操作<ol><li>麻醉收件人按照捐赠者的操作，上面。使用兽医软膏，以保护动物的眼睛干涩从。通过踏板经常捏麻醉确认。 </li><li>动物定位为每个供体手术，这一次离开的权利前肢免费监测生命体征和麻醉深度。 </li><li>剃腹部从剑突至阴茎。剃左腹股沟的一个小区域（用于后续静脉通路）。应用脱毛剂的表面工作，等待大约5分钟，然后取出头发用一块纱布。 </li><li>预习的手术部位以碘伏或chlorexidine基溶液。渗透切口部位用0.1 - 0.5％利多卡因皮下。 </li><li> Tracheotomize和通风收件人为导向上方。另外，在保持鼻锥麻醉收件人。 </li><li>引入导管插入股静脉，如上所述。注入300 - 500 IU普通肝素通过静脉注射管，并随后冲洗管与3 - 用5ml盐水。 </li><li>通过中线腹部切口从剑突22刀片手术刀的阴茎进行剖腹手术。使用自固牵开器缩回腹壁。接下来，优和老鼠的左侧收回的肠子。他们包裹在温暖的盐水浸泡过的纱布。 </li><li>通过上覆再大幅解剖暴露下腔静脉和腹主动脉TRO-腹腔脂肪。 </li><li>有一个弯曲血管钳可用，并准备。 </li><li>圆周上释放IVC和主动脉近端和远端，并与4-0丝质领带包围他们。应该有大约2 - 的这些位点之间的空间3厘米。 </li><li>小心施加弯曲血管钳，确保两个IVC和主动脉的足够部分被上述夹具的钳口露出。 </li><li>使在腔静脉的前壁的切口与地下25针连接到生理盐水填充1ml注射器。延长切口与帕兹剪刀以匹配供体的IVC孔口的长度。 </li><li>从容器中取出心脏，肺脏移植，并从停搏断开。将收件人的腹部移植中的最佳位置进行静脉吻合。 注:心脏，肺移植最终会略微倾斜导向，以对腹部的心尖指向左下象限。 </li><li>固定脚跟和脚趾用9-0尼龙线缝合吻合的端部。扎一个安全结的两端，而使附着到缝合线的长臂的针，并且缝合线的短端将在后面绑。管理停跳的剂量。 </li><li>在运行方式，完成一半的缝合线，系着领带的相对的短缝线臂。管理停跳的剂量。 </li><li>完成缝合线的另一半和领带下来。静脉吻合完成。管理停跳的剂量</li><li>东方的心脏，肺移植的心脏的心尖指向腹部的左下腹。确保静脉吻合不扭结或扭曲。 </li><li>评估将需要到达收件人主动脉，既保持和下腔静脉吻合主动脉在无张力和未扭结配置供体主动脉的长度。 </li><li>划分降主动脉（远端左锁骨下动脉）与虹膜剪。 </li><li>做一个incisio中的n收件人主动脉与地下25针连接到生理盐水填充1ml注射器的前壁。延长切口与帕兹剪刀以匹配供体主动脉孔口的长度。 </li><li>锚捐助主动脉趾（上级方面）收件人主动脉用9-0尼龙缝合。然后，通过针对主动脉的内侧面，并完成二分之一吻合的运行方式。 </li><li>在主动脉吻合的脚跟，扭转缝合线（在缝合方向的转变）的方向，并完成吻合术的侧向半部在运行方式。 </li><li>慢慢在SVC除去IV导管，并结扎SVC与手术夹子。 </li><li>去空气中的主动脉通过定位的未裁剪左颈总动脉。握住动脉开口，在直立位置，这将允许空气自由地撤离。 </li><li>简要地打开弯曲血管钳的钳口，并允许颈动脉出血（四电子空气）为2 - 3秒。重新应用血管钳。 </li><li>申请手术夹子的颈动脉。卸下弯曲血管钳。 </li><li>检查在缝线止血。如果存在，应用温和的压缩与手术纱布或维修用9-0尼龙线（根据出血的严重程度）的长度。 </li><li>供体心脏应该恢复分钟内击败。 </li><li>狠抓收件人的生命体征，麻醉深度，并根据需要管理的容量复苏。 </li><li>根据实验终点，要么关闭动物的腹部，并让它从麻醉中恢复，或准备动物心血管评估。 注:接枝评估的实例包括在负载依赖性和负载无关的血流动力学体内措施的功能中的Langendorff 体外措施和工作心脏模式，以及（在存活手术）超声心动图或MRI调查。 </li><li>在急性终端实验结束时，将动物放血实施安乐死。 </li></ol>

<ol>
	<li>Abbott, C. P., Lindsey, E. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+technique+for+heart+transplantation+in+the+rat.">A technique for heart transplantation in the rat.</a> Arch Surg. 89 (1964), 649-652 (1964).</li><li>Ma, Y., Wang, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+2+heterotopic+heart+transplant+techniques+in+rats:+cervical+and+abdominal+heart.">Comparison of 2 heterotopic heart transplant techniques in rats: cervical and abdominal heart.</a> Exp Clin Transplant. 9 (2), 128-133 (2011).</li><li>Wiedemann, D., Boesch, F., Schneeberger, S., Kocher, A., Laufer, G., Semsroth, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Graft+function+after+heterotopic+rat+heart+transplant+with+an+isolated+reperfused+working+heart:+a+methodic+consideration.">Graft function after heterotopic rat heart transplant with an isolated reperfused working heart: a methodic consideration.</a> Exp Clin Transplant. 10 (2), 154-157 (2012).</li><li>Chen, Z. H., Xia, S. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+technique+of+heterotopic+heart-lung+transplantation+in+the+rat.">The technique of heterotopic heart-lung transplantation in the rat.</a> J Tongji Med Univ. 6 (2), 67-70 (1986).</li><li>Ibrahim, M., Navaratnarajah, 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=Heterotopic+abdominal+heart+transplantation+in+rats+for+functional+studies+of+ventricular+unloading.">Heterotopic abdominal heart transplantation in rats for functional studies of ventricular unloading.</a> J Surg Res. 179 (1), e31-e39 (2013).</li><li>Asfour, B., Hare, J. 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=A+simple+new+model+of+physiologically+working+heterotopic+rat+heart+transplantation+provides+hemodynamic+performance+equivalent+to+that+of+an+orthotopic+heart.">A simple new model of physiologically working heterotopic rat heart transplantation provides hemodynamic performance equivalent to that of an orthotopic heart.</a> J Heart Lung Transplant. 18 (10), 927-936 (1999).</li><li>Figueiredo, J. -. L., Nahrendorf, M., Sosnovik, D. E., Weissleder, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=MRI+of+a+novel+murine+working+heart+transplant+model.">MRI of a novel murine working heart transplant model.</a> Circ Heart Fail. 2 (3), 272-274 (2009).</li><li>Wen, P., Wang, X., 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+simple+technique+for+a+new+working+heterotopic+heart+transplantation+model+in+rats.">A simple technique for a new working heterotopic heart transplantation model in rats.</a> Transplant Proc. 45 (6), 2522-2526 (2013).</li><li>Didi&#233;, M., Biermann, 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=Preservation+of+left+ventricular+function+and+morphology+in+volume-loaded+versus+volume-unloaded+heterotopic+heart+transplants.">Preservation of left ventricular function and morphology in volume-loaded versus volume-unloaded heterotopic heart transplants.</a> A.Am J Physiol Heart Circ Physiol. 305 (4), H533-H541 (2013).</li><li>Gali&#241;anes, M., Zhai, X., Hearse, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+load+on+atrophy,+myosin+isoform+shifts+and+contractile+function:+studies+in+a+novel+rat+heart+transplant+preparation.">The effect of load on atrophy, myosin isoform shifts and contractile function: studies in a novel rat heart transplant preparation.</a> J Mol Cell Cardiol. 27 (1), 407-417 (1995).</li><li>Yokoyama, H., Ohmi, M., Murata, S., Nakame, T., Tabayashi, K., Mohri, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Proposal+of+a+working+left+heart+model+with+a+heterotopic+transplantation+technique+in+rats.">Proposal of a working left heart model with a heterotopic transplantation technique in rats.</a> J Heart Lung Transplant. 14 (4), 706-712 (1995).</li><li>Maruyama, T., Swartz, M. T., McBride, L. R., Pennington, D. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Working+heart+model+of+heterotopic+heart-lung+transplantation+in+rats.">Working heart model of heterotopic heart-lung transplantation in rats.</a> J Thorac Cardiovasc Surg. 107 (1), 210-215 (1994).</li><li>Pacher, P., Nagayama, T., Mukhopadhyay, P., B&#225;tkai, S., Kass, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measurement+of+cardiac+function+using+pressure-volume+conductance+catheter+technique+in+mice+and+rats.">Measurement of cardiac function using pressure-volume conductance catheter technique in mice and rats.</a> Nat Protoc. 3 (9), 1422-1434 (2008).</li><li>Jespersen, B., Knupp, L., Northcott, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Femoral+arterial+and+venous+catheterization+for+blood+sampling,+drug+administration+and+conscious+blood+pressure+and+heart+rate+measurements.">Femoral arterial and venous catheterization for blood sampling, drug administration and conscious blood pressure and heart rate measurements.</a> J Vis Exp. (59), 3496-3410 (2012).</li><li>Habertheuer, A., Kocher, 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=Innovative,+simplified+orthotopic+lung+transplantation+in+rats).">Innovative, simplified orthotopic lung transplantation in rats).</a> J Surg Res. 185 (1), 419-425 (2013).</li></ol>

作者什么都没有透露。

成功与此处所描述的技术将被断定几个因素。其中的关键是确保捐助国和受援国动物的稳定性，采用细致的手术技术，是安全的和最小的失血，确保完成心脏停搏均匀移植冷却，最大限度地减少总缺血时间，并适当去宣扬移植。如上文承认，该技术的技术复杂性是其主要缺点。 我们拥有先进的先前HAHLT技术在几个方面。在供体和受体的操作中描述的变型提供执行以受控和有效...

The first rodent model of heterotopic abdominal heart transplantation (HAHT) was described by Abbott and colleagues in 19641. This technique, and subsequent modifications have been widely applied to characterize transplant graft function and immunologic status. The majority of HAHT techniques described involve a non-volume loaded heart2,3. Models of HAHT involving volume-loaded ventricles have been described, but they are frequently limited in one or more respects.
Heterotopic abdominal heart-lung transplantation (HAHLT) with a volume-loaded left ventricle (LV) has been described previously. Chen and colleagues4, and subsequently Ibrahim and colleagues5 described HAHLT with a single aorto-aortic (donor ascending to recipient abdominal aorta) anastomosis. The only volume load presented to the ventricle in this circulation is the coronary venous return. Asfour and colleagues described a HAHT technique in which the lung circuit was eliminated by anastomosing donor pulmonary artery (PA) to donor left atrium (LA)6. In this circulation, venous inflow to right ventricle (RV) occurs via a donor SVC to recipient IVC anastomosis, and the subsequent LV load is ejected into the aorto-aortic anastomosis. Cardiac function was partially assessed in vivo, and also in vitro using a Langendorff rig. Figueiredo and colleagues described a HAHLT model similar to our own7, but in mice. Venous inflow to the RV occurs via donor SVC to recipient IVC anastomosis. Blood subsequently passes through the single lung circulation and LV load is ejected into the aorto-aortic anastomosis. Cardiac function in their study was assessed by magnetic resonance imaging (MRI). Wen and colleagues described a unique HAHT technique in which the LV is loaded by means of a recipient aorta to donor LA anastomosis8. The LV, therefore, fills at systemic pressures. Cardiac function, and whether LV stroke volume is ejected antegradely in their model was not assessed.
Many of the techniques referenced above involve non-physiologic LV loading conditions, including the techniques whose partial LV load is represented only by coronary venous return. On the other hand, many techniques do approach physiologic LV loading. The majority of these techniques, as with the technique of Asfour and colleagues, omit the pulmonary circulation and utilize a donor PA to donor LA anastomosis6,9. The circulation described by Galinanes and colleagues10 employs a direct recipient cava to donor LA anastomosis, omitting the pulmonary circulation and the right heart. Yokoyama and colleagues achieve the same effect by ligating the donor PA and creating an interatrial communication in the donor heart (omitting donor lung and right heart circulations)11. The circulation of Maruyama and colleagues12 involves an anastomosis between donor left PA and recipient Ao, which permits LV filling via the pulmonary circulation as a conduit, but effectively excludes the right heart.
In cases where near physiologic loading conditions were met, we advance the technique of HAHLT in 2 major respects. First, to our knowledge, the exact configuration we report has not been described in rats. It is possibly the most versatile circulation for investigators wishing to study the physiology, structure, and immunology of the transplanted heart-lung graft. Second, we describe how the function of the transplant graft can be directly characterized in vivo. For this application, pressure-volume conductance catheters can be introduced directly into the LV apex of the transplant graft, which allows for complete cardiac functional characterization. 
The technique described here can be applied to both acute and chronic studies of transplant graft function, while the functional assessment may be performed either in vivo or in vitro. We present a model in which the loading conditions can be near physiologic, however the degree of ventricular loading may be manipulated both acutely and chronically by diverting venous return towards or away from the graft. Afterload conditions can also be manipulated. Because the lung and its airway are retained in this transplant configuration, investigators can re-ventilate the donor lung acutely. Uniquely, lung re-ventilation changes the composition of blood perfusing the transplant coronary arteries. Under non-ventilated conditions, blood ejected from the donor aorta is deoxygenated, and mixes with oxygenated blood in the recipient aorta. Under acutely ventilated conditions, ejected blood becomes oxygenated. Thus, transplant graft function can be compared under ventilated and non-ventilated conditions, and also under variably loaded conditions.
The protocol below describes important modifications to previously described HAHLT donor and recipient operations. It also describes an optimal technique for protecting the transplant graft throughout the period of ischemia (time between donor explant and recipient implant). Advantages of this technique include physiologic conditions potentially approaching that of an orthotopically transplanted graft, and a wide range of investigative applications. An important limitation is its technical complexity. With adequate mentoring and practice, the advantages of this technique will likely outweigh the challenges in adopting it.

<table><tbody><tr><td>Celsior Cardioplegic Soution</td><td>Genzyme</td><td></td><td>The solution is kept on ice throughout the procedure. We prepare our own solution, with slight modifications.</td></tr><tr><td>Rodent Ventilator</td><td>Harvard Apparatus</td><td>Model 683</td><td></td></tr><tr><td>Vital Sign Monitor</td><td>Nonin</td><td>Model 9847V</td><td>Displays SpO&lt;sub&gt;2&lt;/sub&gt; and heart rate.</td></tr><tr><td>IV Cannulae</td><td>Jelco</td><td>3063</td><td>24 - 26 G x 3/4&quot; cannulae.</td></tr><tr><td>IV Tubing</td><td>CareFusion</td><td>MP9259-C</td><td>Short-length connector tubing (18 cm).</td></tr><tr><td>Surgical Clips</td><td>Teleflex Medical</td><td>001204</td><td>Horizon titanium ligating clips.</td></tr><tr><td>Sutures</td><td>Ethicon, Sharpoint</td><td>LA54G, AK-0107</td><td>3-0 silk reel, and 9-0 prolene suture (single-armed, DR5 needle).</td></tr><tr><td>Surgical Instruments</td><td>Not Applicable</td><td>Not Applicable</td><td>The instruments used are generic, and can be purchased from any surgical supply company.</td></tr></tbody></table>

rat heterotopic abdominal heart/single-lung transplantation in a volume-loaded configuration

Herein, we describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The configuration of the transplant graft involves anastomosis of donor inferior vena cava (IVC) to recipient IVC, and donor ascending aorta (Ao) to recipient abdominal Ao. The right upper and middle lung lobes are preserved and function as conduits for blood flow from right heart to left heart.
There are several advantages to using this technique, and it lends itself to a broad range of applications. Because the graft is transplanted in a configuration that allows for dyamic volume-loading, cardiac function may be directly assessed in vivo. The use of pressure-volume conductance catheters permits characterization of load-dependent and load-independent hemodynamic parameters. The graft may be converted to a loaded configuration by applying a clamp to the recipient&#8217;s infra-hepatic IVC. We describe modified surgical techniques for both donor and recipient operations, and an ideal myocardial protection strategy. Depending on the experimental aim, this model may be adapted for use in both acute and chronic studies of graft function, immunologic status, and variable ventricular loading conditions. The conducting airways to the transplanted lung are preserved, and allow for acute lung re-ventilation. This facilitates analysis of the effects of the mixed venous and arterial blood providing coronary perfusion to the graft.
A limitation of this model is its technical complexity. There is a significant learning curve for new operators, who should ideally be mentored in the technique. A surgical training background is advantageous for those wishing to apply this model. Despite its complexity, we aim to present the model in a clear and easily applicable format. Because of the physiologic similarity of this model to orthotopic transplantation, and its broad range of study applications, the effort invested in learning the technique is likely to be worthwhile.

上述HAHLT技术是高度技术性，需要密切注意细节。 表1列出了一些与成功与不成功程序关联的关键因素，并且可以用作用于排除故障的技术困难的指南。 经过收件人主动脉松开，移植冠状动脉应被视为填补氧合血。因此，心肌应该成为粉红色和灌注良好。在技​​术上是成功的实验，心脏就会开始移植再灌注后不久击败。接枝应留在无负载状态（与收件人IVC夹紧?...

Watch this Scientific Journal Video about Rat Heterotopic Abdominal Heart/Single-lung Transplantation in a Volume-loaded Configuration at JoVE.com

Rat Heterotopic Abdominal Heart/Single-lung Transplantation in a Volume-loaded Configuration

We describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The transplant configuration results in a partially loaded graft circulation, allowing direct functional assessment. This model may be employed for acute or chronic studies of function and immunologic status of the transplanted graft.

大鼠异位心脏腹部/在卷装配置单肺移植

Herein, we describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The configuration of the transplant graft ...

rat-heterotopic-abdominal-heartsingle-lung-transplantation-volume

Research

JoVE Journal

Medicine

8.3K 次观看.  UBC James Hogg Research Center, St. Paul's Hospital. We describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The transplant configuration results in a partially loaded graft circulation, allowing direct functional assessment. This model may be employed for acute or chronic studies of function and immunologic status of the transplanted graft.

视频: Rat Heterotopic Abdominal Heart/Single-lung Transplantation in a Volume-loaded Configuration

Heterotopic Heart Transplantation in Mice

The mouse heterotopic heart transplantation has been used widely since it was introduced by Drs. Corry and Russell in 1973. It is particularly valuable for studying rejection and immune response now that newer transgenic and gene knockout mice are available, and a large number of immunologic reagents have been developed. The heart transplant model is less stringent than the skin transplant models, although technically more challenging. We have developed a modified technique and have completed over 1000 successful cases of heterotopic heart transplantation in mice. When making anastomosis of the ascending aorta and abdominal aorta, two stay sutures are placed at the proximal and distal apexes of recipient abdominal aorta with the donor s ascending aorta, then using 11-0 suture for anastomosis on both side of aorta with continuing sutures. The stay sutures make the anastomosis easier and 11-0 is an ideal suture size to avoid bleeding and thrombosis.
When making anastomosis of pulmonary artery and inferior vena cava, two stay sutures are made at the proximal apex and distal apex of the recipient s inferior vena cava with the donor s pulmonary artery. The left wall of the inferior vena cava and donor s pulmonary artery is closed with continuing sutures in the inside of the inferior vena cava after, one knot with the proximal apex stay suture the right wall of the inferior vena cava and the donor s pulmonary artery are closed with continuing sutures outside the inferior vena cave with 10-0 sutures. This method is easier to perform because anastomosis is made just on the one side of the inferior vena cava and 10-0 sutures is the right size to avoid bleeding and thrombosis. In this article, we provide details of the technique to supplement the video.

The mouse heterotopic heart transplantation model has been proven by many investigators to be an important method for studying mechanisms of rejection and immune response. However, the techniques involved are still challenging. By modifying standard techniques we have had success with more than 1000 transplants.

小鼠异位心脏移植

The mouse heterotopic heart transplantation has been used widely since it was introduced by Drs. Corry and Russell in 1973. It is particularly valuable ...

科研

生物学

Heterotopic Auxiliary Rat Liver Transplantation With Flow-regulated Portal Vein Arterialization in Acute Hepatic Failure

In acute hepatic failure auxiliary liver transplantation is an interesting alternative approach. The aim is to provide a temporary support until the failing native liver has regenerated.1-3 The APOLT-method, the orthotopic implantation of auxiliary segments- averts most of the technical problems. However this method necessitates extensive resections of both the native liver and the graft.4 In 1998, Erhard developed the heterotopic auxiliary liver transplantation (HALT) utilizing portal vein arterialization (PVA) (Figure 1). This technique showed promising initial clinical results.5-6 We developed a HALT-technique with flow-regulated PVA in the rat to examine the influence of flow-regulated PVA on graft morphology and function (Figure 2).
A liver graft reduced to 30 % of its original size, was heterotopically implanted in the right renal region of the recipient after explantation of the right kidney.&#160; The infra-hepatic caval vein of the graft was anastomosed with the infrahepatic caval vein of the recipient. The arterialization of the donor&#8217;s portal vein was carried out via the recipient&#8217;s right renal artery with the stent technique. The blood-flow regulation of the arterialized portal vein was achieved with the use of a stent with an internal diameter of 0.3 mm. The celiac trunk of the graft was end-to-side anastomosed with the recipient&#8217;s aorta and the bile duct was implanted into the duodenum. A subtotal resection of the native liver was performed to induce acute hepatic failure. 7
In this manner 112 transplantations were performed. The perioperative survival rate was 90% and the 6-week survival rate was 80%. Six weeks after operation, the native liver regenerated, showing an increase in weight from 2.3&#177;0.8 g to 9.8&#177;1 g. At this time, the graft&#8217;s weight decreased from 3.3&#177;0.8 g to 2.3&#177;0.8 g.
We were able to obtain promising long-term results in terms of graft morphology and function. HALT with flow-regulated PVA reliably bridges acute hepatic failure until the native liver regenerates.

Auxiliary liver transplantation provides a temporary support in acute hepatic failure, until regeneration of the failing liver. The heterotopic auxiliary liver transplantation (HALT) with portal vein arterialization (PVA) renders sufficient liver function. We developed an analogous technique in the rat, to examine the influence of the portal vein arterialization on the morphology and function of the graft.

在急性肝衰竭异位辅助性大鼠肝移植流量调节门静脉动脉

In acute hepatic failure auxiliary liver transplantation is an interesting alternative approach. The aim is to provide a temporary support until the ...

医学

Murine Heterotopic Heart Transplant Technique

It is now over forty years since this technique was first reported by Corry, Wynn and Russell. Although it took some years for other labs to become proficient in and utilize this technique, it is now widely used by many laboratories around the world. A significant refinement to the original technique was developed and reported in 2001 by Niimi. Described here are the techniques that have evolved over more than a decade in the hands of three surgeons (Plenter, Grazia, Pietra) in our center. These techniques are now being passed on to a younger generation of surgeons and researchers.
Based largely on the Niimi experience, the procedures used have evolved in the fine details - details which we will endeavor to relate here in such a way that others may be able to use this very useful model. Like Niimi, we have found that a video aid to learning is a priceless resource for the beginner.

The manuscript describes the steps required to perform the heterotopic heart transplant in the mouse.

小鼠异位心脏移植技术

It is now over forty years since this technique was first reported by Corry, Wynn and Russell. Although it took some years for other labs to become ...

An Immunological Model for Heterotopic Heart and Cardiac Muscle Cell Transplantation in Rats

Heterotopic heart transplantation in rats has been a commonly used model for diverse immunological studies for more than 50 years. Several modifications have been reported since the first description in 1964. After 30 years of performing heterotopic heart transplantation in rats, we have developed a simplified surgical approach, which can be easily taught and performed without further surgical training or background.
After dissection of the ascending aorta and the pulmonary artery and ligation of superior and inferior caval and pulmonary veins, the donor heart is harvested and subsequently perfused with ice-cold saline solution supplemented with heparin. After clamping and incising the recipient abdominal vessels, the donor ascending aorta and pulmonary artery are anastomosed to the recipient abdominal aorta and inferior vena cava, respectively, using continuous running sutures.
Depending on different donor-recipient combinations, this model allows analyses of either acute or chronic rejection of allografts. The immunological significance of this model is further enhanced by a novel approach of in-ear injection of vital cardiac muscle cells and subsequent analysis of draining cervical lymphatic tissue.

We describe a model of heterotopic abdominal heart transplantation in rats, implying modifications of current strategies, which lead to a simplified surgical approach. Additionally, we describe a novel rejection model by in-ear injection of vital cardiac muscle cells, allowing further transplant immunological analyses in rats.

大鼠异位心脏和心肌细胞移植的免疫模型

Heterotopic heart transplantation in rats has been a commonly used model for diverse immunological studies for more than 50 years. Several modifications ...

免疫与感染

Blood Circuit Reconstruction in an Abdominal Mouse Heart Transplantation Model

The surgical technique of heterotopic abdominal heart transplantation in mice is a standard model for research in transplantation immunology. Here, the established technique for a modified blood circuit reconstruction in a heterotopic abdominal heart transplantation model is presented. This method uses the intrathoracic inferior vena cava (IIVC) instead of the pulmonary artery of the donor heart for the anastomosis to the inferior vena cava of the recipient. It is facilitating and improving success rates for abdominal heart transplantation in mice.

A novel technique for blood circuit reconstruction in a heterotopic abdominal mouse heart transplantation model is demonstrated.

腹部小鼠心脏移植模型中的血液回路重建

The surgical technique of heterotopic abdominal heart transplantation in mice is a standard model for research in transplantation immunology. Here, the ...

A Simplified Model for Heterotopic Heart Valve Transplantation in Rodents

There is an urgent clinical need for heart valve replacements that can grow in children. Heart valve transplantation is proposed as a new type of transplant with the potential to deliver durable heart valves capable of somatic growth with no requirement for anticoagulation. However, the immunobiology of heart valve transplants remains unexplored, highlighting the need for animal models to study this new type of transplant. Previous rat models for heterotopic aortic valve transplantation into the abdominal aorta have been described, though they are technically challenging and costly. For addressing this challenge, a renal subcapsular transplant model was developed in rodents as a practical and more straightforward method for studying heart valve transplant immunobiology. In this model, a single aortic valve leaflet is harvested and inserted into the renal subcapsular space. The kidney is easily accessible, and the transplanted tissue is securely contained in a subcapsular space that is well vascularized and can accommodate a variety of tissue sizes. Furthermore, because a single rat can provide three donor aortic leaflets and a single kidney can provide multiple sites for transplanted tissue, fewer rats are required for a given study. Here, the transplantation technique is described, providing a significant step forward in studying the transplant immunology of heart valve transplantation.

This protocol describes a simple and efficient method for the transplantation of aortic valve leaflets under the renal capsule to allow for the study of alloreactivity of heart valves.

啮齿动物异位心脏瓣膜移植的简化模型

There is an urgent clinical need for heart valve replacements that can grow in children. Heart valve transplantation is proposed as a new type of ...

A Modified Cuff Technique for Mouse Cervical Heterotopic Heart Transplantation Model

Cardiac allograft rejection limits the long-term survival of patients after heart transplantation. A mouse heart transplantation model is ideal for investigating the mechanism of cardiac allograft rejection in preclinical studies because of their high homology with human genes. This understanding would help develop unique approaches to improving patients&#39; long-term survival treated with cardiac allografts. In a mouse model, abdominal donor heart implantation is commonly performed with an end-to-end&#160;anastomosis to the recipient&#39;s aorta and inferior vena cava using stitches. In this model, the donor&#39;s heart is implanted by end-to-end anastomosis to the recipient&#39;s carotid artery and jugular vein by the modified-Cuff technique. The transplantation surgery is performed without stitching and thus may increase the survival of the recipient since there is no interference with the blood supply and venous reflux of the lower body. This mouse model would help investigate the mechanisms underlying the immunological and pathological (acute/chronic) rejection of cardiac allografts.

In the present protocol, a mouse heart transplantation model is used for investigating the mechanism of cardiac allograft rejection. In this heterotopic heart transplantation model, operation efficiency is improved, and the survival of cardiac grafts is ensured by a cervical end-to-end anastomosis of heart implantation using a modified Cuff technique.

一种改进的袖带技术用于小鼠颈椎异位心脏移植模型

Cardiac allograft rejection limits the long-term survival of patients after heart transplantation. A mouse heart transplantation model is ideal for ...

A Heterotopic Rat Heart Transplantation Model using Circulatory Death Donor Hearts

The objective of this protocol is to set up a rat heterotopic heart transplantation model with donation after circulatory death (DCD) donor hearts. There are two setups for this protocol: heart donor setup and recipient setup. In the heart donor setup, Sprague Dawley rats are anesthetized, endotracheally intubated, and ventilated. The right carotid artery is cannulated to deliver heparin and the paralytic agent vecuronium-bromide. The DCD process is initiated by terminating the ventilation. After 20 min, the heart is exposed and the aorta distal to the brachiocephalic branch is clamped. At 25 min from terminating the ventilator, ice-cold University of Wisconsin (UW) solution is perfused through the carotid catheter to flush the heart. The heart is procured by dividing the aorta, pulmonary artery, venae cavae, and pulmonary veins and stored in UW solution for implantation. In the recipient setup, the Lewis rat is anesthetized with isoflurane. Slow-release buprenorphine is administered subcutaneously to facilitate a smooth postoperative recovery. Through a midline abdominal incision, the infra-renal aorta and the inferior vena cava are isolated and clamped with an atraumatic vascular clamp. The donor heart aorta and pulmonary artery are sutured to the recipient abdominal aorta and vena cava, respectively, with a running 8-0 Prolene. The vascular clamp is removed to reperfuse the heart. The abdominal wall is closed and the rat is recovered. After a set interval (24 h to 2 weeks), the recipient rat is anesthetized, the transplanted heart is exposed, and a balloon-tip-catheter is inserted into the left ventricle via the apex to record developed pressure and dP/dt using a data acquisition system. The heart tissue is collected for histology, immunology, or molecular analysis. A successful DCD donor rat heart transplantation model will allow further studies on the cardioprotective approaches to improve heart transplantation outcomes from DCD donors.

Here, we describe the preparation and technical details of murine heterotopic heart transplantation utilizing a circulatory death donor heart.

The objective of this protocol is to set up a rat heterotopic heart transplantation model with donation after circulatory death (DCD) donor hearts. There ...

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 ...

Modified Heterotopic Abdominal Heart Transplantation and a Novel Aortic Regurgitation Model in Rats

Over the past 50 years, many researchers have reported heterotopic abdominal heart transplantation in mice and rats, with some variations in the surgical technique. Modifying the transplantation procedure to strengthen the myocardial protection could prolong the ischemia time while preserving the donor&#39;s cardiac function. This technique&#39;s key points are as follows: transecting the donor&#39;s abdominal aorta before harvesting to unload the donor&#39;s heart; perfusing the donor&#39;s coronary arteries with a cold cardioplegic solution; and topical cooling of the donor&#39;s heart during the anastomosis procedure. Consequently, since this procedure prolongs the acceptable ischemia time, beginners can easily perform it and achieve a high success rate.
Moreover, a new aortic regurgitation (AR) model was established in this work using a technique different from the existing one,&#160;which is created by inserting a catheter from the right carotid artery and puncturing the native aortic valve under continuous echocardiographic guidance. A heterotopic abdominal heart transplantation was performed using the novel AR model. In the protocol, after the donor&#39;s heart is harvested, a stiff guidewire is inserted into the donor&#39;s brachiocephalic artery and advanced toward the aortic root. The aortic valve is punctured by pushing the guidewire further even after the resistance is felt, thus inducing AR. It is easier to damage the aortic valve using this method than with the procedure described in the conventional AR model. Additionally, this novel AR model does not contribute to the recipient&#39;s circulation; therefore, this method is expected to produce a more severe AR model than the conventional procedure.

This study demonstrates a reproducible heterotopic abdominal heart transplantation technique in rats that beginners can learn and perform. Additionally, a novel aortic regurgitation model in rats is generated by performing heterotopic abdominal heart transplantation and damaging the donor's aortic valve using a guidewire after harvesting.

改良异位腹心移植及新型大鼠主动脉瓣反流模型

Over the past 50 years, many researchers have reported heterotopic abdominal heart transplantation in mice and rats, with some variations in the surgical ...