作者要感谢比亚焦彼得拉博士为他以前的工作在我们的实验室。

所有动物饲养的无病原体条件下，在根据IACUC批准科罗拉多巴巴拉戴维斯中心动物设施大学和照顾根据美国国立卫生研究院指南。 麻醉深度是由脚趾捏最初并遵守呼吸速率判断，一旦程序已经开始。 1，供心收获<ol><li>注射戊巴比妥（60毫克/公斤的IP）麻醉供体小鼠。 </li><li>通过4路限制固定鼠标和剪辑皮毛。用酒精擦拭皮肤。 </li><li>解剖远离皮肤上后，通过使横切口正好逊色于膜片打开腹腔。 </li><li>切开膈肌后到沿海插入和解除前胸壁前上方暴露心脏。 </li><li>延伸的完整厚度切开后外侧胸腔在胸部的左侧和右侧。 </li><li>隔离下腔静脉（IVC）和周围放置IVC松散5-0丝线缝合邻近其插入到右心房。 </li><li>注入1.0毫升4℃肝素生理盐水（200 U /立方厘米）到下腔静脉，然后结扎血管用5-0丝线缝合和鸿沟。 </li><li>隔离右上腔静脉（rSVC）以类似的方式，并结扎用5-0丝线缝合和鸿沟。 </li><li>轻轻摇动心脏对动物的右侧和隔离左上腔静脉（lSVC），结扎用5-0丝线缝合，分化露出左肺动脉。 </li><li>下湿纱布保护心脏，而胸腺钝解剖远离主肺动脉（PA）和左，右PA分支机构及升主动脉弓。 </li><li>钝性解剖主动脉弓游离周围组织的。微型剪刀是用来划分主动脉近端右肱动脉头侧， 即应该有心脏和部门之间没有分支点的主动脉。 </li><li>本节中的主动脉弓形成动脉压脉袋的植入过程。 </li><li>反映主动脉袖口inferiorly，露出树干的PA和PA的左，右分支。 </li><li>钝解剖左和右PA分支离周围组织从心脏尽可能远。这允许从周围组织中的PA的躯干的容易清扫。 </li><li>划分PA作为主干远端越好，刚好接近其分叉。本节中的肺动脉的形成静脉压脉袋的植入过程。 </li><li>广场周围的心脏和领带的基础5-0丝线缝合。的心脏，然后切成免费在该基地，并置于4℃生理盐水。总收获时间是约。 10-15分钟。 </li></ol> 2，心脏植入技术<ol><li>麻醉受体小鼠用戊巴比妥（60毫克/公斤的IP的起始剂量，如需要25毫克/公斤的IP补充剂量）。 </li><li>剪辑的皮毛和通过4路限制固定鼠标和预习用聚维酮碘和悬垂在一个无菌的方式在皮肤上。 </li><li>做2厘米的垂直中线腹部切口，进入腹腔。 </li><li>收回肠优和外在的胸部。请将包裹在无菌湿纱布（无菌生理盐水）整个情况。 </li><li>下面孤立肾血管腹主动脉和下腔静脉（IVC），并放置主动脉周围4-0棉关系，下腔静脉，然后优越劣于吻合部位。 </li><li>找出任何腰椎血管内场和结扎与10-0尼龙线。 </li><li>结棉关系，首先劣质其次是优越的。在这样一些血液被保持在使织脉切开术容易主动脉。 </li><li>用30号针头形成织脉切开术进入主动脉的管腔。延长切口用细显微剪到大约2毫米的长度。此切口在沿长直线该船只的itudinal轴。 </li><li>做一个结束捐助主动脉在以下时尚收件人主动脉端侧吻合。放置一个10-0尼龙线缝住在供体主动脉和切口在收件人主动脉和领带的下角。将第二个10-0尼龙对面先在供体和主动脉切口在腹主动脉和领带的优越角落。 </li><li>使正在运行的缝合线从优去劣于主动脉侧壁和领带对先前放置的住宿针。一定要带的intimas（容器内表面）一起为你缝合。然后缝合在一个正在运行的时装和领带内侧。每一侧的第一个和最后缝应尽可能靠近逗留针越好。然后目标是有3均匀间隔即总共5针这两者之间的针迹。 </li><li>做一个结束捐助肺动脉收件人IVC以下时尚侧吻合。 PunctURE下腔静脉用30号针头，并延长切口约。 2mm的细显微剪。此切口在沿容器的纵向轴线成一直线。 </li><li>领带的捐助肺动脉切口的下腔静脉的下角落10-0尼龙。将第二个10-0尼龙对面的第一供体动脉和切口下腔静脉和领带的优越角落。 </li><li>使肺小动脉及下腔静脉和领带之间的连续缝合线。每一侧的第一个和最后缝应尽可能靠近逗留针越好。然后的目标是有5个均匀分布的制作共7针两针之间。 </li><li>松开远端4-0棉领带重新建立静脉血流。 </li><li>一旦静脉吻合术的止血已经观察到近端4-0棉布领带逐渐松动和动脉吻合观察止血。当两个吻合口被认为是安全的，删除C奥多栓从鼠标。 </li><li>返回肠道腹部。腹壁封闭在用5-0丝线缝合在运行方式两层。 </li><li>管理一个1.0毫升的静脉团无菌，温暖的生理盐水到腹部为收盘时液体复苏和0.8 ml生理盐水皮下注射手术后。手术时，不需要任何其他支援措施。恢复动物的升温毯。共植入时间约为90-120分钟，适合初学者，45-60分钟，使用经验。管理丁丙诺啡镇痛，0.05毫克/千克，资深大律师，0.1-0.2毫升在程序的开始，每6-12小时72小时后运。 </li></ol> 3，嫁接评估<ol><li>每天通过经腹部触诊评估肾功能。 </li><li>按住鼠标，就好像是被给予腹腔注射。 </li><li>轻轻地压在腹壁食指的前端，并确定打浆STRength的移植性和规律性。 </li><li>给触诊质量从4的分数（正常幅度和频率）设​​置为0（非打浆拒绝接枝）。 </li></ol>重要注意事项： 所有仪器进行灭菌，无菌手套戴在整个过程和无菌区被保持。供体和受体手术是通过使用在手术显微镜进行。确保吻合是“干净”。即，将针时背面壁没有被捕获。这将导致一个显著收缩流动在一个失败的移植物，在极端情况下，这将很可能导致以后肢瘫痪。这也是非常重要的，全厚度通过包括血管外膜和缝合针的内膜得以实现。边缘的Evertion也确保有内膜至中膜接触，这有助于密封和吻合口愈合。另一个非常重要的事实或者是，确保在吻合缝合线的张力也是最优的。太松，就会出现不可逆的泄漏，过紧和狭窄流动将导致。如果动脉侧，这将导致移植物灌注不良，如果在静脉侧拥挤心脏将导致。

<ol>
	<li>Corry, R. J., Winn, H. J., Russell, P. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heart+transplantation+in+congenic+strains+of+mice.">Heart transplantation in congenic strains of mice.</a> Transplant Proc. 5 (1), 733 (1973).</li><li>Corry, R. J., Winn, H. J., Russell, P. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Primarily+vascularized+allografts+of+hearts+in+mice.+The+role+of+H-2D,+H-2K,+and+non-H-2+antigens+in+rejection.">Primarily vascularized allografts of hearts in mice. The role of H-2D, H-2K, and non-H-2 antigens in rejection.</a> Transplantation. 16 (4), 343 (1973).</li><li>Niimi, M. <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+for+heterotopic+cardiac+transplantation+in+mice:+experience+of+3,000+operations+by+one+surgeon.">The technique for heterotopic cardiac transplantation in mice: experience of 3,000 operations by one surgeon.</a> J Heart Lung Transplant. 20 (10), 1123 (2001).</li><li>Plenter, R. J., Zamora, M. R., Grazia, T. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Four+decades+of+vascularized+heterotopic+cardiac+transplantation+in+the+mouse.">Four decades of vascularized heterotopic cardiac transplantation in the mouse.</a> J Invest Surg. 26 (4), 223 (2013).</li><li>Su, S., 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=Modified+suture+technique+in+a+mouse+heart+transplant+model.">Modified suture technique in a mouse heart transplant model.</a> Asian J Surg. 34 (2), 86 (2011).</li><li>Mao, 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+novel+and+knotless+technique+for+heterotopic+cardiac+transplantation+in+mice.">A novel and knotless technique for heterotopic cardiac transplantation in mice.</a> J Heart Lung Transplant. 28 (10), 1102 (2009).</li><li>Csencsits, K. L., Bishop, D. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contrasting+alloreactive+CD4++and+CD8++T+cells:+there's+more+to+it+than+MHC+restriction.">Contrasting alloreactive CD4+ and CD8+ T cells: there's more to it than MHC restriction.</a> Am J Transplant. 3 (2), 107 (2003).</li><li>Hasegawa, T., Visovatti, S. H., Hyman, M. C., Hayasaki, T., Pinsky, 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=Heterotopic+vascularized+murine+cardiac+transplantation+to+study+graft+arteriopathy.">Heterotopic vascularized murine cardiac transplantation to study graft arteriopathy.</a> Nat Protoc. 2 (3), 471 (2007).</li><li>Linfert, D., Chowdhry, T., Rabb, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lymphocytes+and+ischemia-reperfusion+injury.">Lymphocytes and ischemia-reperfusion injury.</a> Transplant Rev. 23 (1), (2009).</li><li>Grazia, T. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acute+cardiac+allograft+rejection+by+directly+cytotoxic+CD4+T+cells:+parallel+requirements+for+Fas+and+perforin.">Acute cardiac allograft rejection by directly cytotoxic CD4 T cells: parallel requirements for Fas and perforin.</a> Transplantation. 89 (1), 33 (2010).</li><li>Liu, F., Kang, S. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heterotopic+heart+transplantation+in+mice.">Heterotopic heart transplantation in mice.</a> J Vis Exp. (6), (2007).</li><li>Plenter, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heterotopic+heart+transplantation+in+mice.">Heterotopic heart transplantation in mice.</a> European Society for Surgical Research. 45th Annual Congress. , (2010).</li></ol>

作者什么都没有透露。

这种手术方法是不容易掌握，但一旦掌握是一个强有力的研究工具。研究者/外科医生是由技术的一致性和通过注重细节的回报。在学习阶段耐心是关键。正如新实3发布，与一个基于视频的学习工具花费平均为11努力实现第一次成功的过程和78企图达到90％的成功率的帮助。视频已成为一个重要的教学工具在手术11,12。 故障排除从吻合口出血，可能会发生，这可能是由于要么缺乏正确的张力的缝合线，或太少缝合。虽然凝血诱导剂，如明胶海绵可减少泄漏非常有用，我们建议医生应该依靠良好的技术。拥挤的非跳动的心脏是最常见的原因为吻合口太紧，特别是在静脉侧。非BEATI纳克，非灌注移植物通常是由一个空气泡已行进到冠状动脉中的一个引起的。它保持了潮湿对湿字段以避免气泡进入血管是非常重要的。 该技术的局限性这种技术是不适合，如果一个研究者想研究一个全面运作的心脏效应。这就需要原位移植术，迄今已证明不可能执行。 相对于现有方法的意义如果要研究对小鼠完全血管化，实体器官移植的效果，那么心脏模型可能是最简单的掌握。肺，肾和肝移植的小鼠模型确实存在，但更难学习和完善。 关键步骤在议定书内这是非常重要的全厚度P驴包括血管外膜和缝合针的内膜得以实现。边缘的Evertion也确保有内膜至中膜接触，这有助于密封和吻合口愈合。另一个非常重要的因素是确保吻合缝合线的张力也是最优的。太松，就会出现不可逆的泄漏，过紧和狭窄流动将导致。如果动脉侧，这将导致移植物灌注不良，如果在静脉侧拥挤心脏将导致。 总之，重复，程序和持续注重细节的一致性，将产生很大的成绩，可资助和发布的数据。

当它可以在小鼠进行肾，肺，肝脏和胰腺移植的时代，基本的器官移植和免疫学研究自1973年以来的基石1-4保持在鼠标异位心脏移植模型。在随后的几年一些论文已经发表，详细改进/优化5,6到这个程序。 随着实体器官主要血管移植模型这个程序是首屈一指的。一旦掌握了这种方法适合于研究分为异体排斥反应7，慢性血管病变8和缺血再灌注损伤9机制的发展。 成功地学习这个过程的关键是，就像任何其他手术，耐心的教练和学员，注重细节的部分。在该过程开始时，新的外科医生会发现吨哎会花很多时间在每个移植。随着经验的不断积累，手术时间，因此缺血，会大大减少。注重每一个步骤的详细迟早会走向成功。 而教师可以尽自己所能传递，并预测，所有可能在这些手术可能遇到的可能坑瀑布，“创意”实习生将有可能找到一些自己的！ 是程序的基本过程如下：捐助升主动脉弓端 - 侧吻合到受体腹主动脉与供体肺动脉端 - 侧吻合到受体腹下腔静脉（IVC）。从逆行时尚收件人主动脉通过捐助主动脉冠状动脉血流量。一旦血已通过冠状系统它漏入右心房通过冠状窦流出，被泵入右心室和然后通过肺动脉进入收件人IVC。以这种方式，冠状动脉系统供给的动脉血和窦性心律返回到接枝内灌注1-2分钟。由于心脏的左厢房正在加载基本上压左室游离壁就会萎缩随着时间的推移。

<table border="0" cellpadding="0" cellspacing="0" style="width:608px;" width="607">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="17">
			<td height="17" style="height:17px;width:247px;">Instrument</td>
			<td style="width:109px;">Roboz #</td>
			<td style="width:147px;">Fine Science Tools #</td>
			<td style="width:105px;">Arosurgical #</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Straight micro-dissecting forcep #5</td>
			<td>RS-5015</td>
			<td>11295-51</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Curved micro-dissecting forcep #7</td>
			<td>RS-5047</td>
			<td>11297-00</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Curved serrated forcep</td>
			<td>RS-5137</td>
			<td>11052-10</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Vannas micro-dissecting scissors, short</td>
			<td>RS-5610</td>
			<td></td>
			<td>09.140.08</td>
		</tr>
		<tr height="17">
			<td colspan="2" height="17" style="height:17px;">Micro-dissecting scissors, straight, sharp, long</td>
			<td></td>
			<td>11.602.11</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Micro spring handle needle holder</td>
			<td></td>
			<td></td>
			<td>11.549.15</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Straight mosquito forcep</td>
			<td></td>
			<td>91308-12</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Micro-dissecting scissors, straight, blunt</td>
			<td>RS-5962</td>
			<td>14078-10</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Micro-dissecting scissors, curved, blunt</td>
			<td>RS-5981</td>
			<td>14079-10</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Micro retractor</td>
			<td>RS-6540</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Instrument tray, 10&#8221; x 6 &#189;&#8221; x &#190;&#8221;</td>
			<td>RT-1350S</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Silk suture, 5/0, 22.5m spool</td>
			<td></td>
			<td>18020-50</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;"></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Suture</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">10/0 nylon</td>
			<td></td>
			<td></td>
			<td>T4A10Q07</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">5/0 silk</td>
			<td></td>
			<td></td>
			<td>E19A05N</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;"></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Gloves</td>
			<td></td>
			<td>Drapes</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Biogel from Medex Supply</td>
			<td></td>
			<td>Precept, #64-9012-9</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;"></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Syringes</td>
			<td></td>
			<td>Cotton applicators</td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">B-D 1cc insulin, #329424</td>
			<td></td>
			<td colspan="2">Fisher-brand, #23-400-100</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;"></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">Povidone-Iodine swabs</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">PDI, #B40600</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;"></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="17">
			<td height="17" style="height:17px;">4/0 Cotton ties</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="17">
			<td colspan="2" height="17" style="height:17px;">Domestic cotton autoclaved with instruments</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

murine heterotopic heart transplant technique

现在是四十多年来，因为这种技术最早由科里，永利和罗素。虽然花了一些年，其他实验室成为精通并利用该技术，目前已广泛应用于世界各地的许多实验室。一个显著细化到原来的技术的开发和新实报道于2001年。这里描述的是已经在三个外科医生（Plenter，格拉齐亚，彼得拉）在我中心手中发展了十余年的技术。这些技术现在被传递到年轻一代的外科医生和研究人员。 主要基于新实经验，所使用的程序已经进化中的细节 - 细节，我们将尽力这里涉及这样一种方式，其他人可能能够使用这个非常有用的模型。像新实，我们已经发现，视频辅助学习是初学者一个无价的资源。

<html>这种手术技术的使用率无论是简单的移植物的存活/排斥反应的研究，还是蛮复杂的实验方案开辟了道路。在下面的图中简要介绍了研究中，我们力图确定的参与，如果有的话，Fas和/或穿孔为CD4 + T细胞介导的​​心脏排斥反应机制。这之所以成为可能，通过小鼠品系目前可用的非凡的数组。结果表明，直接抑制心脏移植的CD4效应T细胞的需要接枝Fas表达和T细胞穿孔素表达的替代性的贡献。据我们所知，这是首次证明了由CD4 + T细胞杀伤活性可以玩初级急性排斥反应在体内的专性作用。 1.JPG“/&gt; 图1穿孔素和Fas专性代表和CD4 + T细胞介导的心肌抑制维生素B6，B6 PFPKO（穿孔素基因敲除），和B6 GLD 平行路径 （FAS配体不足）的CD4 T细胞被用来重建B6 抹布- / -收件人C3H野生型或FAS缺陷C3H LPR心脏移植。去除的Fas单独的（♦，P = NS与控制体重的C3H + B6的CD4 + T细胞）从供体心脏或罢免或穿孔单纯从CD4 + T细胞（■，P = NS与控制体重的C3H + B6的CD4 + T细胞）没有废除排斥。有趣的是，从效应子CD4 + T细胞的FasL蛋白去除的那样延迟排斥显著（●，P &lt;0.02与重量C3H + B6的CD4 + T细胞，P &lt;0.01相对于体重的C3H + B6 PFPKO CD4 + T细胞，和P &lt;0.01，与。C3H LPR + B6的CD4 + T细胞）。然而，大多数移植仍然拒绝（4）。显著，同时去除捐助国Fas和CD4 + T细胞穿孔素完全消除排斥反应（○，P &lt;0.002相对于对照重量C3H + B6的CD4 + T细胞，重量C3H + B6 PFPKO CD4 + T细胞，和C3H 的lpr + B6的CD4 + T细胞）。这废止是显著比单独去除CD4 + T细胞FasL的更强大（○，P &lt;0.003与控制体重的C3H + B6 GLD CD4 + T细胞）。从格拉齐亚等[10]。经许可后转载。

Watch this Scientific Journal Video about Murine Heterotopic Heart Transplant Technique at JoVE.com

Murine Heterotopic Heart Transplant Technique

这份手稿描述了在小鼠进行异位心脏移植手术所需的步骤。

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 ...

murine-heterotopic-heart-transplant-technique

Research

JoVE Journal

Medicine

16.7K 次观看.  University of Colorado Denver. 这份手稿描述了在小鼠进行异位心脏移植手术所需的步骤。 

视频: Murine Heterotopic Heart Transplant Technique

A Modified Heterotopic Swine Hind Limb Transplant Model for Translational Vascularized Composite Allotransplantation (VCA) Research

Vascularized Composite Allotransplantation (VCA) such as hand and face transplants represent a viable treatment option for complex musculoskeletal trauma and devastating tissue loss. Despite favorable and highly encouraging early and intermediate functional outcomes, rejection of the highly immunogenic skin component of a VCA and potential adverse effects of chronic multi-drug immunosuppression continue to hamper widespread clinical application of VCA. Therefore, research in this novel field needs to focus on translational studies related to unique immunologic features of VCA and to develop novel immunomodulatory strategies for immunomodulation and tolerance induction following VCA without the need for long term immunosuppression.
This article describes a reliable and reproducible translational large animal model of VCA that is comprised of an osteomyocutaneous flap in a MHC-defined swine heterotopic hind limb allotransplantation. Briefly, a well-vascularized skin paddle is identified in the anteromedial thigh region using near infrared laser angiography. The underlying muscles, knee joint, distal femur, and proximal tibia are harvested on a femoral vascular pedicle. This allograft can be considered both a VCA and a vascularized bone marrow transplant with its unique immune privileged features. The graft is transplanted to a subcutaneous abdominal pocket in the recipient animal with a skin component exteriorized to the dorsolateral region for immune monitoring.
Three surgical teams work simultaneously in a well-coordinated manner to reduce anesthesia and ischemia times, thereby improving efficiency of this model and reducing potential confounders in experimental protocols. This model serves as the groundwork for future therapeutic strategies aimed at reducing and potentially eliminating the need for chronic multi-drug immunosuppression in VCA.

A Modified Heterotopic Swine Hind Limb Transplant Model for Translational Vascularized Composite Allotransplantation VCA Research

Vascularized Composite Allotransplantations (VCA) have become a clinical reality. However, broad clinical application of VCA is limited by chronic multi-drug immunosuppression. The authors present a reliable and reproducible large animal model to translate novel immunomodulatory strategies that can minimize or potentially eliminate the need of immunosuppression in VCA.

平移血管复合同种异体移植（VCA）研究一种改进的异位猪后肢移植模型

Vascularized Composite Allotransplantation (VCA) such as hand and face transplants represent a viable treatment option for complex musculoskeletal trauma ...

科研

医学

Murine Cervical Heart Transplantation Model Using a Modified Cuff Technique

Mouse models are of special interest in research since a wide variety of monoclonal antibodies and commercially defined inbred and knockout strains are available to perform mechanistic in vivo studies. While heart transplantation models using a suture technique were first successfully developed in rats, the translation into an equally widespread used murine equivalent was never achieved due the technical complexity of the microsurgical procedure. In contrast, non-suture cuff techniques, also developed initially in rats, were successfully adapted for use in mice1-3. This technique for revascularization involves two major steps I) everting the recipient vessel over a polyethylene cuff; II) pulling the donor vessel over the formerly everted recipient vessel and holding it in place with a circumferential tie. This ensures a continuity of the endothelial layer, short operating time and very high patency rates4.
Using this technique for vascular anastomosis we performed more than 1,000 cervical heart transplants with an overall success rate of 95%. For arterial inflow the common carotid artery and the proximal aortic arch were anastomosed resulting in a retrograde perfusion of the transplanted heart. For venous drainage the pulmonary artery of the graft was anastomosed with the external jugular vein of the recipient5.
Herein, we provide additional details of this technique to supplement the video.

The murine cervical heart transplantation model is well suited for immunological as well as ischemia reperfusion injury studies. We modified the procedure using a non-suture cuff technique and performed more than 1,000 successful transplants with this approach.
Herein, we provide additional details of this technique to supplement the video.

用改进的袖套法小鼠颈部心脏移植模型

Mouse models are of special interest in research since a wide variety of monoclonal antibodies and commercially defined inbred and knockout strains are ...

A Modified Method for Heterotopic Mouse Heart Transplantion

Mice are often used as heart transplant donors and recipients in studies of transplant immunology due to the wide range of transgenic mice and reagents available. A difficulty is presented due to the small size of the animal and the considerable technical challenges of the microsurgery involved in heart transplantation. In particular, a high rate of technical failure early after transplantation may result from recipient death and post-operative complications such as hind limb paralysis or a non-beating heart. Here, the complete technique for heterotopic mouse heart transplantation is demonstrated, involving harvesting the donor heart and its subsequent implantation into a recipient mouse. The donor heart is harvested immediately following in situ perfusion with cold heparinized saline and transection of the ascending aorta and pulmonary artery. The recipient operation involves preparation of the abdominal aorta and inferior vena cava (IVC), followed by end-to-side anastomosis of the donor aorta with the recipient aorta using a single running 10-0 microsuture and a similar anastomosis of the donor pulmonary artery with the recipient IVC. Following the operation the animal is injected with 0.6 ml normal saline subcutaneously and allowed to recover on a 37&#160;&#176;C heating pad. The results from 227 mouse heart transplants are summarized with a success rate at 48 hr of 86.8%. Of the 13.2% failures within 48 hr, 5 (2.2%) experienced hind limb paralysis, 10 (4.4%) had a non-beating heart due to graft ischemic injury and/or thrombosis, while 15 (6.6%) died within 48 hr.

A technique is demonstrated for the microsurgical procedure for heterotopic transplantation of hearts in mice, including simplified methods for donor harvesting and recipient vessel anastomosis.

对于异位小鼠心脏移植技术的一种改进方法

Mice are often used as heart transplant donors and recipients in studies of transplant immunology due to the wide range of transgenic mice and reagents ...

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.

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 ...

Murine Kidney Transplant Technique

The first mouse kidney transplant technique was published in 19731 by the Russell laboratory. 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 using the donor aorta to form the arterial anastomosis instead of the renal artery was developed and reported in 1993 by Kalina and Mottram 2 with a further advancement coming from the same laboratory in 1999 3. While one can become proficient in this model, a search of the literature reveals that many labs still experience a high proportion of graft loss due to arterial thrombosis. We describe here a technique that was devised in our laboratory that vastly reduces the arterial thrombus reported by others 4,5. This is achieved by forming a heel-and-toe cuff of the donor infra-renal aorta that facilitates a larger anastomosis and straighter blood flow into the kidney.

The goal of this manuscript is to describe the steps required to perform a kidney transplant in a mouse, paying particular attention to the details of the arterial anastomosis.

小鼠肾脏移植技术

The first mouse kidney transplant technique was published in 19731 by the Russell laboratory. Although it took some years for other labs to become ...

Heterotopic Cervical Heart Transplantation in Mice

The heterotopic cervical heart transplantation in mice is a valuable tool in transplant and cardiovascular research. The cuff technique greatly simplifies this model by avoiding challenging suture anastomoses of small vessels thereby reducing warm ischemia time. In comparison to abdominal graft implantation the cervical model is less invasive and the implanted graft is easily accessible for further follow-up examinations. Anastomoses are performed by pulling the ascending aorta of the graft over the cuff with the recipient&#8217;s common carotid artery and by pulling the main pulmonary artery over the cuff with the external jugular vein. Selection of appropriate cuff size and complete mobilization of the vessels are important for successful revascularization. Ischemia-reperfusion (I/R) injury can be minimized by perfusing the graft with a cardioplegic solution and by hypothermia. In this article, we provide technical details for a simplified and improved cuff technique, which should allow surgeons with basic microsurgical skills to perform the procedure with a high success rate.

The cuff technique shortens and facilitates the model of heterotopic cervical heart transplantation in mice by avoiding technically challenging suture anastomoses of small vessels. The technical details shown in this video paper should allow researches to establish this model in their laboratories.

颈椎异位心脏移植小鼠

The heterotopic cervical heart transplantation in mice is a valuable tool in transplant and cardiovascular research. The cuff technique greatly simplifies ...

Balloon-based Injury to Induce Myointimal Hyperplasia in the Mouse Abdominal Aorta

The use of animal models is essential for a better understanding of MH, one major cause for arterial stenosis.In this article, we demonstrate a murine balloon denudation model, which is comparable with established vessel injury models in large animals. The aorta denudation model with balloon catheters mimics the clinical setting and leads to comparable pathobiological and physiological changes. Briefly, after performing a horizontal incision in the aorta abdominalis, a balloon catheter will be inserted into the vessel, inflated, and introduced retrogradely. Inflation of the balloon will lead to intima injury and overdistension of the vessel. After removing the catheter, the aortic incision will be closed with single stiches. The model shown in this article is reproducible, easy to perform, and can be established quickly and reliably. It is especially suitable for evaluating expensive experimental therapeutic agents, which can be applied in an economical fashion. By using different knockout-mouse strains, the impact of different genes on MH development can be assessed.

This article demonstrates a murine model to study the development of myointimal hyperplasia (MH) after aortic balloon injury.

球囊损伤诱发小鼠腹主动脉 Myointimal 增生的研究

The use of animal models is essential for a better understanding of MH, one major cause for arterial stenosis.In this article, we demonstrate a murine ...

A Mouse Model of Vascularized Heterotopic Spleen Transplantation for Studying Spleen Cell Biology and Transplant Immunity

The spleen is a unique lymphoid organ that plays a critical role in the homeostasis of the immune and hematopoietic systems. Patients that have undergone splenectomy regardless of precipitating causes are prone to develop an overwhelming post-splenectomy infection and experience increased risks of deep venous thrombosis and malignancies. Recently, epidemiological studies indicated that splenectomy might be associated with the occurrence of cardiovascular diseases, suggesting that physiological functions of the spleen have not yet been fully recognized. Here, we introduce a mouse model of vascularized heterotopic spleen transplantation, which not only can be utilized to study the function and behavioral activity of splenic immune cell subsets in different biologic processes, but also can be a powerful tool to test the therapeutic potential of spleen transplantation in certain diseases. The main surgical steps of this model include donor spleen harvest, the removal of recipient native spleen, and spleen graft revascularization. Using congenic mouse strains (e.g., mice with CD45.1/CD45.2 backgrounds), we observed that after syngeneic transplantation, both donor-derived splenic lymphocytes and myeloid cells migrated out of the graft as early as post-operative day 1, concomitant with the influx of multiple types of recipient cells, thus generating a unique chimera. &#160;Despite relatively challenging techniques, this procedure can be performed with &#62;90% success rate. This model allows tracking the fate, longevity, and function of splenocytes during steady state and in a disease setting following a spleen transplantation, thereby offering a great opportunity to discover the distinct role for spleen-derived immune cells in different disease processes.

This protocol details the surgical steps of a mouse model of vascularized heterotopic spleen transplantation, a technically challenging model that can serve as a powerful tool in studying the fate and longevity of spleen cells, the mechanisms of distinct spleen cell populations in disease progression, and transplant immunity.

血管化异位脾移植小鼠模型在脾细胞生物学和移植免疫研究中的应用

The spleen is a unique lymphoid organ that plays a critical role in the homeostasis of the immune and hematopoietic systems. Patients that have undergone ...

Murine Cervical Aortic Transplantation Model using a Modified Non-Suture Cuff Technique

With the introduction of powerful immunosuppressive protocols, distinct advances are possible in the prevention and therapy of acute rejection episodes. However, only minor improvement in the long-term results of transplanted solid organs could be observed over the past decades. In this context, chronic allograft vasculopathy (CAV) still represents the leading cause of late organ failure in cardiac, renal and pulmonary transplantation.
Thus far, the underlying pathogenesis of CAV development remains unclear, explaining why effective treatment strategies are presently missing and emphasizing a need for relevant experimental models in order to study the underlying pathophysiology leading to CAV formation. The following protocol describes a murine heterotopic cervical aortic transplantation model using a modified non-suture cuff technique. In this technique, a segment of the thoracic aorta is interpositioned in the right common carotid artery. With the use of the non-suture cuff technique, an easy to learn and reproducible model can be established, minimizing the possible heterogeneity of sutured vascular micro anastomoses.

Here, we present a protocol of heterotopic aortic transplantation in mice using the non-suture cuff technique in a cervical murine model. This model can be used to study the underlying pathology of chronic allograft vasculopathy (CAV) and can help evaluate new therapeutic agents in order to prevent its formation.

使用改性非缝合铐技术的宫颈主动脉移植模型

With the introduction of powerful immunosuppressive protocols, distinct advances are possible in the prevention and therapy of acute rejection episodes. ...

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.