This work was funded by the American Association of Plastic Surgeons 2014 Academic Scholar Award.

所有的手术过程均符合美国约翰霍普金斯大学和农业部的美国农业部和公共卫生服务的要求完成。该协议遵循美国约翰霍普金斯大学动物护理和使用委员会，机构审查委员会批准的准则（协议号M013M490）。最终存活数据被记录为下面描述的外科手术。供体和受体动物接收使用丁丙诺啡先发制人麻醉以0.1mg /千克SC手术前和在受体动物丁丙诺啡之一小时重新施用同样剂量移植后，并根据需要在最初48小时内重新给药手术后。 1.供体移植恢复注意:开始移植40分钟的捐助部分早于接受者的移植，以尽量减少受援国的麻醉时间，便于同时结束时间或略有耳利尔结束时间与收件人的准备。 <ol><li>使用标准的无菌显微器械和无菌手套的程序。我们的实验室使用显微仪器高压灭菌。 </li><li>麻醉用异氟醚感应蒸发器在4％的供体小鼠（男）。使用无创伤机械剪除去从颈椎，胸椎头发，腹部区域。将动物在仰卧位，并通过鼻锥保持异氟烷上1-2％。通过定期评估脚趾捏撤回反射确保整个过程充分麻醉。 </li><li>在此之前的皮肤切口，广泛应用聚维酮碘消毒剂其次是异丙醇用消毒棉签准备工作。 </li><li>开始时先用浅横皮肤切口与整个颈部和腹部的皮肤剪刀。连接两个切口双边沿腋线。 </li><li>采用显微镊解剖颈部两侧识别，结扎和划分颈外静脉用6-0丝线缝合和剪刀。然后使用电划分胸锁乳突肌，露出颈内静脉和颈总动脉，双侧。通过在左路和右路颈总动脉和颈内静脉散装时尚6-0丝线缝合。 注:它们将被捆绑，后来在步骤1.9分。 </li><li>急剧划分带状肌和相关疏松蜂窝组织，位于前部的气管，用剪刀释放颈椎区的其余附件。 </li><li>用双极电和锐性剥离，分胸大肌肌肉和锁骨露出锁骨下血管和结扎（6-0丝线缝合），并划分近端。 </li><li>接下来，轻轻的，把握和撤回动物的阴茎。沿阴茎的背形象化阴茎的背静脉，和消毒区域用异丙醇。使用地下30针，注入30000单位肝素静脉内通过背静脉并允许阴茎反冲回到其原来的位置。可能会发生局部漏肝素溶液到周围组织的。 </li><li>使用周围颈总动脉和颈内静脉的前面放置散装的联系，结扎和划分结构，两侧。 </li><li>接下来用剪刀作一横切口intrabdominal。剔骨肠内以暴露肝下下腔静脉，并注入2毫升冷欧洲柯林斯心麻痹液进入肝下下腔静脉。确保正确注射肝变色和停止心跳的推进到下一步骤之前，可视化。 注:欧洲Collins液是在我们的实验室制备，见表特定的试剂和仪器。 </li><li>用剪刀通过双边DIAPH进入胸腔内空腔ragmatic切口从露出腹部。延伸穿过肋间肌肉和肋头侧的切口。反映胸壁暴露心脏，胸腺和大血管的同时，确保保护沿胸壁胸廓内血管。 </li><li>注入肝上下腔静脉用4ml冷欧洲柯林斯心麻痹液。 </li><li>识别主动脉根部和远侧跟踪到降主动脉。大幅削减降主动脉（保留最大长度）。 </li><li>确定肺动脉主干及分刚好接近其分支点（保留最大长度）。然后，使用2毫升冷欧元柯林斯停搏液，通过放置一个软塑料尖导管进入肺动脉的管腔冲洗肺动脉干和心脏。 </li><li>用6-0丝线缝合，结扎和划分下腔静脉，肺静脉汇合，和双侧上腔静脉辅助分支。然后提升并沿主干支气管和气管小心不要进入呼吸道附件解剖心脏头侧。使用锋利的和双极电解剖胸壁，胸腺和心脏完全从供体小鼠释放它。 </li><li>最后，修整同种异体移植胸壁体外到更小的尺寸，用剪刀沿胸骨和横向肋脉，小心不要破坏胸廓血管（ 图1A）。为了下面的血运重建尽量减少出血，使用沿着osteomusculocutaneous胸骨边界双极电灼。 </li><li>将在10毫升冷（4度摄氏度）的移植欧洲-科林斯的解决方案，如果收款人没有为插入做好准备。然而，如果接收方是准备插页，同种异体移植物转移直到接收者手术视野。 </li></ol> 2.收件人准备注意:要尽量减少收件人麻醉时间，在一个单独的操作站开始接收方准备前的供体移植收获完成约40分钟。 <ol><li>使用单独的一套标准的无菌显微仪器和程序无菌手套。 </li><li>麻醉用异氟醚感应蒸发器在4％的受体小鼠（男或女）。使用无创机械快船移除右颈部和胸部的头发。 </li><li>鼠标放置在仰卧位置和角度的右上肢稍稍下方形成的头部和右上肢之间的110度角。通过鼻锥维持麻醉上1-2％异氟醚。 </li><li>放置石油眼药膏上使用棉签鼠标眼睛。在此之前的皮肤切口，用聚维酮碘防腐剂接着异丙醇广泛准备手术部位。 </li><li>用剪刀，做一个皮肤切口沿右下博尔中线下颌骨DER和infero，横向延伸的切口右胸部。使用钝性分离微血管钳，调动了颈外静脉被周向无血管的软组织和外膜。除以使用电各分公司，并删除使用锐性剥离和电灼对移植的自由空间颌下腺右叶。 </li><li>确保足够的长度的颈外静脉的外翻在一个压脉袋，并结扎外颈静脉用6-0丝线缝合。通过预切聚酰亚胺箍的内腔插入静脉并使用牛头犬微血管钳固定到位容器袖口复杂。然后用剪刀，近侧划分外部颈静脉，外翻在袖口，以及在适当位置10-0尼龙缝合线固定。 （图1B） </li><li>划分右胸锁乳突肌双极电以暴露颈总动脉。周向暴徒ilize颈部区域内头侧向最远侧点的动脉。这是使用钳船只钝性分离，除去软组织和周围的外膜完成。 </li><li>用6-0丝线缝合，结扎并划分颈总动脉。通过预切割的聚酰亚胺袖口的管腔通过动脉和其固定用的牛头犬微血管钳尽量靠近胸部入口越好。除以血管远端，轻轻地扩张用显微扩张血管，埃弗特在袖口，并在适当位置10-0尼龙缝合修复。 （图1B） 注:具体显微扩张器在特定的试剂和仪器表所述。 </li></ol> 3.移植插图<ol><li>保持无菌的标准仪器和无菌手套把收件人颈部区域内的移植在倒置和倾斜位置。 </li><li>下一步，将捐助下降AOR抽动管腔在收件人的动脉袖套构建体和其固定用10-0尼龙缝合线（图1C 和1D）。 </li><li>时尚相同吻合作为供体肺动脉和受体小鼠（图1C 和1D）的外翻颈外静脉袖套构建体之间步骤3.2。 </li><li>首先拆下静脉微血管钳（颈外静脉夹），然后松开动脉夹（颈总动脉夹）。在动脉灌注，检查移植解决任何出血的全部内容。如果出血可视化，重新动脉夹，以减少出血量及减少使用双极电出血的来源。 </li><li>检查移植物，并确保止血。释放并彻底清除动脉微血管夹。观察心脏表现出再灌注的迹象，这将是快速扩容的心脏室立即显现出来，并等待对B吃开始在0.5-1分钟。用温盐水（35摄氏度），以滋润的心脏。 </li><li>悬垂胸壁到解剖位置，以免诱发的吻合任何的扭曲和紧张局势。关闭手术伤口的是用6-0尼龙连续缝线（图1E）的皮肤。 </li></ol> 4.术后护理<ol><li>辖0.3 ml生理盐水腹腔内液体推注，术后立即进行补液。 </li><li>然后经皮下疼痛和感染预防，分别注入丁丙诺啡（0.1mg的/ kg）和恩诺沙星（5毫克/公斤）。 </li><li>放置在热灯下的动物直到从麻醉觉醒，并返回到胸骨斜卧。在恢复过程中，检查颈部以可视化的移植保证足够的移植物灌注的原纤化的心跳。 </li><li>一旦醒来，在卧位，返回鼠标到一个单独的笼子（无其他老鼠的公司）在那里可以领取食物和水自由采食。由于右上肢任何临时次要限制性运动，留在笼子的地板上的明胶的食物来源。 </li><li>观察受体小鼠1小时，术后，然后将其返回到笼子设备，在它能够接收的食物和水随意并进行检查，一天三次的第一个24小时的活性和营养摄取。根据需要为先72小时全天监控小鼠的痛苦和困扰，并重新剂量丁丙诺啡（0.1毫克/千克）的迹象皮下的两倍。检查动物此后每天，每星期体重它们。 </li><li>与兽医工作人员咨询，如果任何小鼠显示的痛苦，痛苦的迹象，或采食量下降。考虑早期安乐死（在我们的协议安乐死技术采用的 CO 2过量为7分钟，接着通过颈椎脱位）。 </li><li>同种异体移植心脏跳动的停止被定义为一个特定端点促使鼠标为sacrificed。 </li></ol>

The authors do not have any conflicts of interest or financial disclosures to declare.

有众多的现象因入同种异体移植，其中包括的免疫学调查，但不限于:急性和慢性排斥反应，直接和间接的抗原呈递，收件人敏化，或混合嵌合体诱导的机制。19动物模型已成为的黄金标准移植免疫学的研究中，和小鼠模型被普遍，由于其成本低，转基因和基因敲除小鼠的可用性，市售的单克隆抗体，相对降低兽医和外壳的要求，并易于复制的实现。迄今为止，多种心脏移植模型已经用于研究固体器官移植。19,22-27同样，过多的小鼠模型已被用于研究血管复合同种异体移植28然而，结合实体器官和血管的复合同种异体移植的研究是有限的，和技术还没有建立在小鼠中。整块胸壁，胸腺，和心脏移植的小鼠这里提出的模型是研究的影响，结合实体器官和血管复合同种异体移植的免疫学机制的可靠和可复制的工具。 为了进一步推进移植领域，通过新的治疗方式延长移植物存活和尽量减少免疫抑制的承诺，必须继续实施。一种这样的方法是通过混合嵌合体（供体造血细胞在受体的局部植入），这可能会导致免疫抑制自由供者特异性耐受的诱导，即使在某些情况下，嵌合体不能持续。29,30骨髓输血/移植搭配实体器官31或血管的复合allotransplanation 32,33需要大量的预处理构成一个显著的挑战。血管的骨髓中，血管化复合移植构建第一部分，可以解决这个问题。血管骨髓移植允许供体骨髓细胞的供体保留微环境中的不间断传送，并且被认为是优于单细胞的骨髓移植耐受性和减少的免疫抑制的要求的诱导。34-36此外，掺入与血管骨髓移植胸腺组织已证明能增加供体来源的T细胞嵌合体，最终打在诱导和维持嵌合体的支撑作用。延长移植物存活和尽量减少免疫抑制的基础概念化的2,9上述策略组合实体器官，胸腺和血管复合同种异体移植小鼠模型。 异位整块胸壁，胸腺和心脏移植是多个的合并其torical动物模型。用小鼠非袖口缝合颈部异位心脏移植已经确立，并认为比异位腹部微血管心脏移植少技术要求高。19事实上，袖口技术已在多个其他动物移植模型中实现37-44异位在大鼠胸骨移植于1999年推出由圣地亚哥等人的研究血管骨髓移植的替代方法。1他们能够表现出长期的外围嵌合体，耐受性和生存以下免疫抑制术后当天停止30. 1 Bozkurt的等人随后开发了一个大鼠模型于2013年合并胸腺和胸壁的骨肌部的充分程度。应当指出，然而，这种模式不同于我们的在多个方面的模型。这包括:（1）他们的模型是没有任何固体器官，（2），使用传统的显微外科技术正在完成大鼠，（3）供体收获期间的胸廓血管结扎，（4）通过一个共同的颈动脉和外颈静脉，和执行一个单方面的，单椎弓根（5 ）移植的同种异体移植到腹股沟区域。2尽管如此，他们的模型是能够证明与单独血管的骨髓移植相比，捐助者出身的胸腺起着不仅为嵌合体隆胸也为嵌合维护显著的作用。2共移植的胸腺和心脏的后续猪模型已经显示在心脏同种异体移植物存活的优良效果。每个动物模型10,11的优点，但缺乏机械体内研究关于结合实体器官，胸腺和血管复合同种异体移植促使我们组设计这个模型。 在经历前ecuting这种新颖的模式表现出一定的经验教训，要求我们的团队，制定修改，以达到更好的动物的生存。这种模式试图与一家运营商，最终延长了手术和麻醉时间超过3-4小时，还延长冷缺血时间。动物不会醒来之后终止程序。的两个团队的方式实施削减总的手术和麻醉时间90分钟。这反映在受体小鼠的60分钟麻醉时间，和0-10分钟同种异体冷缺血时间。期间同种异体移植灌注，鼠标易受出血，这可能限制其在不久的围手术期的生存能力。我们提倡细致的检查移植体外潜在的出血来源，以及斗牛犬微血管钳移植再灌注过程中温和释放。通过将同种异体移植物中的反射位置是更容易识别的特定网站出血。此外，这种接枝插画位置促进血管的最符合人体工学的捻最小化容器扭结的风险。最后，与第一个48小时的恢复，运动的受体小鼠的右上肢范围可能阻碍与问候攀爬笼子，以获得食物和水。因此，我们建议明胶营养源的位置沿笼，方便营养的摄入。通常由3术后第二天，是右上肢内恢复全套动作。 虽然有一些限制这种模式，其中包括需要显微外科技术娴熟，两个同时显微镜可用性和两个团队的方式的要求，它仍然证明是一个成功的方法来执行相关的机理免疫学研究结合实体器官和血管复合同种异体移植。其更广泛的应用程序可能会进一步有助于开发新的与免疫osuppressive协议，急性和慢性排斥反应研究力学，和实施潜力的策略的诱导和维持嵌合，并延长同种异体移植物存活。

Cardiac transplantation is the treatment of choice for end-stage heart failure. Both technical advancements and pharmacological innovations have propelled the field to early graft acceptance rates above 90%.12,13 Despite this, 60-80% 5-year graft survival is at a standstill and chronic rejection, characterized by transplant vasculopathy, remains inevitable.14-16 Furthermore, patients are subjected to multiple surgical procedures and lifelong immunosuppression, which are associated with chest wall deformities and medical sequelae and toxicities, respectively. The need for innovative approaches to extend allograft survival, minimize the immunosuppressive requirements, and offer reconstructive options for anatomical deformities is pressing.
Vascularized composite allotransplantation offers a unique strategy for improving heart transplant outcomes both from an immunological aspect as well as a reconstructive perspective.17 Vascularized composite allografts are also unique in a way that they have an inherent source of donor-derived hematopoietic stem cells which has shown a favorable ability to reduce immunosuppression and induce and sustain mixed chimerism.1-8 Additionally, co-transplantation of the thymus has shown to prolong survival of both, solid organ transplants and vascularized composite allografts.2,9-11 Combining these strategies with heart transplantation offers a novel solution to the aforementioned challenges facing heart transplantation.18
Murine models serve as excellent platforms for mechanistic in vivo investigation because of the availability of antibodies and well-defined inbred and knockout strains.19-21 Although heart transplantation in mice is commonly studied using a heterotopic intraabdominal microsurgical suture transplant model22-25, a heterotopic, cervical, non-suture cuff technique model has shown to be extremely replicable, reliable, and carries fewer rates of thrombosis.19,26,27 The goal of this study is to develop a heterotopic en bloc osteomyocutaneous chest wall, thymus, and heart transplant technique in mice to study the immunological mechanisms of combined solid organ and vascularized composite allotransplantation using a cervical non-suture cuff technique. This cluster allograft is perfused through the anastomosis of the donor descending aorta to the right common carotid artery and the donor pulmonary artery to the right external jugular vein. Preservation of the internal thoracic vessels and associated thymus branches is paramount to perfusing the chest wall (sternum, ribs, muscles, and skin) and thymus.

<table border="0">
	<tbody>
		<tr>
			<td>Euro-Collins Solution</td>
			<td></td>
			<td></td>
			<td>The solution is not commercially purchased but rather prepared in the laboratory. To make a 500ml solution add the ingredient listed below to a 330ml of double distilled water. Mix well, and then fill in the rest of the 170ml of double distilled water into the solution to a final volume of 500ml. 
			Ingredients: 1.02g KH2PO4, 3.66g K2HPO4, 0.56g KCl, 0.42g NaHCO3, and 17.52g of glucose.</td>
		</tr>
		<tr>
			<td>Suture</td>
			<td>Ethilon</td>
			<td>MWI 72667</td>
			<td>6-0 Ethilon</td>
		</tr>
		<tr>
			<td>Suture</td>
			<td>Microsurgery Instruments, Inc.</td>
			<td>10-0 Nylon</td>
			<td>10-0 Nylon</td>
		</tr>
		<tr>
			<td>Polyimide Cuff Vein (21G)</td>
			<td>Vention Medical</td>
			<td>141-0043</td>
			<td>http://www.ventionmedical.com/products-and-services/polyimide-tubing/</td>
		</tr>
		<tr>
			<td>Polyimide Cuff Artery (24G)</td>
			<td>Vention Medical</td>
			<td>141-0027</td>
			<td>http://www.ventionmedical.com/products-and-services/polyimide-tubing/</td>
		</tr>
		<tr>
			<td>Soft plastic tip catheter</td>
			<td>Terumo</td>
			<td>SR*OX2419CA</td>
			<td>24G x 3/4&#34;&#160;</td>
		</tr>
		<tr>
			<td>Microsurgical dilator</td>
			<td>S&#38;T</td>
			<td>D-5a.1</td>
			<td>Dilator, 11cm, FH, 0.1mm AT10d</td>
		</tr>
	</tbody>
</table>

a novel microsurgical model for heterotopic, en bloc chest wall, thymus, and heart transplantation in mice

Exploration of novel strategies in organ transplantation to prolong allograft survival and minimizing the need for long-term maintenance immunosuppression must be pursued. Employing vascularized bone marrow transplantation and co-transplantation of the thymus have shown promise in this regard in various animal models.1-11 Vascularized bone marrow transplantation allows for the uninterrupted transfer of donor bone marrow cells within the preserved donor microenvironment, and the incorporation of thymus tissue with vascularized bone marrow transplantation has shown to increase T-cell chimerism ultimately playing a supportive role in the induction of immune regulation. The combination of solid organ and vascularized composite allotransplantation can uniquely combine these strategies in the form of a novel transplant model. Murine models serve as an excellent paradigm to explore the mechanisms of acute and chronic rejection, chimerism, and tolerance induction, thus providing the foundation to propagate superior allograft survival strategies for larger animal models and future clinical application. Herein, we developed a novel heterotopic en bloc chest wall, thymus, and heart transplant model in mice using a cervical non-suture cuff technique. The experience in syngeneic and allogeneic transplant settings is described for future broader immunological investigations via an instructional manuscript and video supplement.

同系C57BL / 6移植实现长期生存。同种异体移植物（图1）的设计中被证明是成功的，从动物的生存透视，并评价正在进行移植物存活的能力。这是通过覆盖皮肤表明剩余可行，活性持续同种异体移植物的毛发生长，和心跳能够用可视化和手指触诊进行评估。存活数据被表示在图2中进行同源移植小鼠。平均存活时间大于109天。基于存活数据，可以合理推断，移植的同种异体移植物的技术方面的目的是灌注胸壁，胸腺和心脏的全部。此外，同系动物的生存长期的能力进一步支持了这种小鼠模型不仅是可行的，但可以复制​​。这证明了概念整块胸壁，胸腺，和心脏移植验证次Ë小鼠模型来研究结合实体器官和血管复合同种异体移植。 <img alt="图1" src="/files/ftp_upload/53442/53442fig1.jpg" /> 图1.术中的照片。 （一）胸壁，胸腺，和心脏移植的成功回收，整理和可视化体外从后方面。双侧胸廓容器被保留。（B）中的收件人外部颈静脉（箭头）和颈总动脉（箭头）是外翻在准备吻合血管固定过聚酰亚胺袖口。（C）的同种异体移植血管吻合完成。箭头示出了供体肺动脉和受体外颈静脉之间的吻合。箭头显示了捐助降主动脉与受体颈总动脉之间的吻合。星号标识胸腺和反射胸壁被可视化，可欣赏莹胸腺。 （四）较高的放大倍数显示了微血管袖口吻合。 （五）完成移植插图。 <a href="https://www.jove.com/files/ftp_upload/53442/53442fig1large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图2" src="/files/ftp_upload/53442/53442fig2.jpg" /> 图2.整块胸壁，胸腺，和心脏移植物存活的整块胸壁，胸腺，和心脏异体移植在同系C57BL / 6小鼠的Kaplan-Meier生存曲线（N = 3;平均生存时间大于109天）。 <a href="https://www.jove.com/files/ftp_upload/53442/53442fig2large.jpg" target="_blank">请点击此处查看该图的放大版本。</a>

Watch this Scientific Journal Video about A Novel Microsurgical Model for Heterotopic, En Bloc Chest Wall, Thymus, and Heart Transplantation in Mice at JoVE.com

A Novel Microsurgical Model for Heterotopic, En Bloc Chest Wall, Thymus, and Heart Transplantation in Mice

To study combined solid organ and vascularized composite allotransplantation, we describe a novel heterotopic en bloc chest wall, thymus, and heart transplant model in mice using a cervical non-suture cuff technique.

一种新的显微模型异位，整块胸壁，胸腺，和心脏移植小鼠

Exploration of novel strategies in organ transplantation to prolong allograft survival and minimizing the need for long-term maintenance immunosuppression ...

a-novel-microsurgical-model-for-heterotopic-en-bloc-chest-wall-thymus

Universidad San Sebastian

Research

JoVE Journal

Medicine

9.2K 次观看.  Johns Hopkins University School of Medicine. To study combined solid organ and vascularized composite allotransplantation, we describe a novel heterotopic en bloc chest wall, thymus, and heart transplant model in mice using a cervical non-suture cuff technique.

视频: A Novel Microsurgical Model for Heterotopic, En Bloc Chest Wall, Thymus, and Heart Transplantation in Mice

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 ...

科研

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 ...

Transplantation of Pulmonary Valve Using a Mouse Model of Heterotopic Heart Transplantation

Tissue engineered heart valves, especially decellularized valves, are starting to gain momentum in clinical use of reconstructive surgery with mixed results. However, the cellular and molecular mechanisms of the neotissue development, valve thickening, and stenosis development are not researched extensively. To answer the above questions, we developed a murine heterotopic heart valve transplantation model. A heart valve was harvested from a valve donor mouse and transplanted to a heart donor mouse. The heart with a new valve was transplanted heterotopically to a recipient mouse. The transplanted heart showed its own heartbeat, independent of the recipient&#8217;s heartbeat. The blood flow was quantified using a high frequency ultrasound system with a pulsed wave Doppler. The flow through the implanted pulmonary valve showed forward flow with minimal regurgitation and the peak flow was close to 100 mm/sec. This murine model of heart valve transplantation is highly versatile, so it can be modified and adapted to provide different hemodynamic environments and/or can be used with various transgenic mice to study neotissue development in a tissue engineered heart valve.

In order to understand the cellular and molecular mechanisms underlying neotissue formation and stenosis development in tissue engineered heart valves, a murine model of heterotopic heart valve transplantation was developed. A pulmonary heart valve was transplanted to recipient using the heterotopic heart transplantation technique.

肺动脉瓣使用异位心脏移植小鼠模型移植

Tissue engineered heart valves, especially decellularized valves, are starting to gain momentum in clinical use of reconstructive surgery with mixed ...

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 ...

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 ...

Heterotopic Renal Autotransplantation in a Porcine Model: A Step-by-Step Protocol

Kidney transplantation is the treatment of choice for patients suffering from end-stage renal disease. It offers better life expectancy and higher quality of life when compared to dialysis. Although the last few decades have seen major improvements in patient outcomes following kidney transplantation, the increasing shortage of available organs represents a severe problem worldwide. To expand the donor pool, marginal kidney grafts recovered from extended criteria donors (ECD) or donated after circulatory death (DCD) are now accepted for transplantation. To further improve the postoperative outcome of these marginal grafts, research must focus on new therapeutic approaches such as alternative preservation techniques, immunomodulation, gene transfer, and stem cell administration.
Experimental studies in animal models are the final step before newly developed techniques can be translated into clinical practice. Porcine kidney transplantation is an excellent model of human transplantation and allows investigation of novel approaches. The major advantage of the porcine model is its anatomical and physiological similarity to the human body, which facilitates the rapid translation of new findings to clinical trials. This article offers a surgical step-by-step protocol for an autotransplantation model and highlights key factors to ensure experimental success. Adequate pre- and postoperative housing, attentive anesthesia, and consistent surgical techniques result in favorable postoperative outcomes. Resection of the contralateral native kidney provides the opportunity to assess post-transplant graft function. The placement of venous and urinary catheters and the use of metabolic cages allow further detailed evaluation. For long-term follow-up studies and investigation of alternative graft preservation techniques, autotransplantation models are superior to allotransplantation models, as they avoid the confounding bias posed by rejection and immunosuppressive medication.

Porcine models of organ transplantation provide an important platform to study mechanisms of organ preservation. This article describes a heterotopic porcine renal autotransplantation model, which allows investigating new approaches to improve the outcome of transplantation using marginal kidney grafts.

异位自体肾移植的猪模型:有步骤，分步协议

Kidney transplantation is the treatment of choice for patients suffering from end-stage renal disease. It offers better life expectancy and higher quality ...

Cardiopulmonary Bypass in a Mouse Model: A Novel Approach

As prolonged cardiopulmonary bypass becomes more essential during cardiac interventions, an increasing clinical demand arises for procedure optimization and for minimizing organ damage resulting from prolonged extracorporal circulation. The goal of this paper was to demonstrate a fully functional and clinically relevant model of cardiopulmonary bypass in a mouse. We report on the device design, perfusion circuit optimization, and microsurgical techniques. This model is an acute model, which is not compatible with survival due to the need for multiple blood drawings. Because of the range of tools available for mice (e.g., markers, knockouts, etc.), this model will facilitate investigation into the molecular mechanisms of organ damage and the effect of cardiopulmonary bypass in relation to other comorbidities.

This paper describes how to perform cardiopulmonary bypass in mice. This novel model will facilitate the investigation of the molecular mechanisms involved in organ damage.

小鼠体外循环模型: 一种新方法

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

A Pre-Clinical Porcine Model of Orthotopic Heart Transplantation

Fifty-years following the first successful report, cardiac transplantation remains the gold-standard treatment for eligible patients with advanced heart failure. Multiple small-animal models of heart transplantation have been used to study the acute and long-term effects of novel therapies. However, few are tested and demonstrated success in clinical trials. It is of critical importance to evaluate new therapies in a clinically relevant large-animal model for efficient and reliable translation of basic studies&#39;&#160;findings. Here, we describe a pre-clinical large-animal (porcine) model of orthotopic heart transplantation that has been firmly established and previously used to investigate novel cardioprotective strategies. This procedure focuses on acute ischemia-reperfusion injury and is a reliable method to investigate novel interventions which have been tested and validated in smaller experimental models, such as the murine model. We demonstrate its usefulness in assessing cardiac performance during the early post-transplantation period and other potential possibilities enabled by the model.

Here, we describe a pre-clinical large-animal (porcine) model of orthotopic heart transplantation that has been firmly established and utilized to investigate novel cardioprotective strategies.

正交心脏移植的临床前猪模型

Fifty-years following the first successful report, cardiac transplantation remains the gold-standard treatment for eligible patients with advanced heart ...

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 Porcine Heterotopic Heart Transplantation Protocol for Delivery of Therapeutics to a Cardiac Allograft

Cardiac transplantation is the gold standard treatment for end-stage heart failure. However, it remains limited by the number of available donor hearts and complications such as primary graft dysfunction and graft rejection. The recent clinical use of an ex vivo perfusion device in cardiac transplantation introduces a unique opportunity for treating cardiac allografts with therapeutic interventions to improve function and avoid deleterious recipient responses. Establishing a translational, large-animal model for therapeutic delivery to the entire allograft is essential for testing novel therapeutic approaches in cardiac transplantation. The porcine, heterotopic heart transplantation model in the intraabdominal position serves as an excellent model for assessing the effects of novel interventions and the immunopathology of graft rejection. This model additionally offers long-term survival for the pig, given that the graft is not required to maintain the recipient's circulation. The aim of this protocol is to provide a reproducible and robust approach for achieving ex vivo delivery of a therapeutic to the entire cardiac allograft prior to transplantation and provide technical details to perform a survival heterotopic transplant of the ex vivo perfused heart.

We present a protocol for utilizing a normothermic ex vivo sanguinous perfusion system for the delivery of therapeutics to an entire cardiac allograft in a porcine heterotopic heart transplant model.

一种猪异位心脏移植方案，用于向心脏同种异体移植物提供治疗药物

Cardiac transplantation is the gold standard treatment for end-stage heart failure. However, it remains limited by the number of available donor hearts ...