这项工作是支持的，部分由一个津贴从美国国立卫生研究院（RO1 HL098228）到CKB。

注:所有动物的程序批准了全国儿童医院实验动物管理和使用委员会。 1，肺动脉心脏瓣膜收获从心脏瓣膜供体小鼠<ol><li>高压灭菌所有的外科手术工具手术前:1倍罚款剪刀，3微型镊子，微型2倍血管钳，1x夹紧应用镊子，1个微型针器，1个弹簧剪刀，1卷收器。 </li><li> 6-8周龄雌性C57BL / 6小鼠作为肺动脉心脏瓣膜捐助。从笼中取出小鼠并称重，然后用安乐死氯胺酮/甲苯​​噻嗪鸡尾酒（氯胺酮200 mg / kg和甲苯噻嗪，20毫克/千克，IP）过量。 </li><li>夹在胸部，然后将鼠标在垫背部靠着位置。然后进行开胸手术。暴露心脏，请在右心房一小截，并灌注左心室与冰冷的生理盐水。 </li><li>说白了从升一解剖肺动脉（PA）奥尔塔。切出肺动脉瓣（PV）以及肺动脉2毫米袖口。处置的心脏的其余部分。 </li><li>存储在PV中冷肝素和生理盐水溶液（100单位/ ml）。注意:该PV可以保持在溶液中进行移植前两小时到供体心脏。 </li></ol> 2，供心准备<ol><li> 6-8周龄雌性C57BL / 6小鼠作为供体心脏。从笼中取出小鼠并称重，然后用安乐死氯胺酮/甲苯​​噻嗪鸡尾酒（氯胺酮200 mg / kg和甲苯噻嗪，20毫克/千克，IP）过量。这是一个终端程序。 </li><li>夹在胸部，然后将鼠标在垫背部靠着位置。然后进行开胸手术。说白了分离心脏，下腔静脉（IVC），上腔静脉（SVC），升主动脉，PA和肺静脉。灌注下腔静脉与冰冷的无菌生理盐水。 </li><li>结扎下腔静脉，SVC和肺静脉用6-0丝线缝合再切优于连字。 </li><li>切开主动脉和​​PA为2 mm袖带。 </li><li>切出的PV和处置。 </li></ol> 3，心脏瓣膜移植到供心<ol><li>步骤2.5之后，立即将心脏瓣膜步骤1.5到供体心脏和定向心脏瓣膜。 </li><li>固定光伏与右侧用10-0单丝缝线上锥形针的阀的针脚，并开始与从PV的另一侧5-6拆线连续缝合。 </li><li>整理的前侧之后，水平旋转的心脏，并开始到PV的背面侧缝合到供体心脏。 </li><li>在寒冷的无菌肝素/生理盐水存储的心脏。注:供体心脏能保持在溶液中对植入前两小时到受体小鼠。 </li></ol> 4，异位心脏移植到一个受体小鼠<ol><li> 6-8周龄雌性C57BL / 6小鼠作为一个RECIpient。从笼中取出小鼠并称重，然后用氯胺酮/甲苯​​噻嗪鸡尾酒（氯胺酮100 mg / kg和甲苯噻嗪10毫克/公斤）麻醉。酮洛芬（5毫克/千克）作为麻醉前镇痛。 </li><li>检查镇静乘尾水平捏后，夹在腹部和胸部的头发。润滑眼睛用无菌眼药膏，然后将鼠标在垫背部靠着位置。消毒腹部与优碘和酒精垫。然后覆盖鼠标，用无菌的悬垂性和唯一暴露在切口区域。 </li><li>使从剑突至耻骨上区域下方中线剖腹切口，然后插入一个自固定牵引器。在包裹蘸生理盐水纱布的肠子。说白了定义下腹主动脉和腔静脉。 </li><li>将两个6-0丝线缝合近端和远端主动脉周围及下腔静脉抑制血液循环。 </li><li>将供体心脏在右侧腹主动脉并用无菌GAU覆盖它泽。用生理盐水滋润它。 </li><li>使一个织脉切开术中使用30号针头的腹主动脉和延伸开口用剪刀给施主主动脉的大小。 </li><li>使用在锥形针无菌10-0单丝缝合线进行端至端吻合术。确保供体主动脉与一个线圈上的开口在腹主动脉的近端，并开始与从腹主动脉的远端4-5拆线连续缝合。 </li><li>翻转的心脏的左侧，用生理盐水注入纱布覆盖它，并开始与从腹主动脉的远端4-5拆线连续缝合。 </li><li>使用30号针头在IVC一个venotomy并延长开放供体肺动脉的大小。 </li><li>使用在锥形针无菌10-0单丝缝合线执行端侧吻合。固定施主PA​​与一个线圈上的开口中的下腔静脉的近端，并开始与4-5拆线连续缝合下腔静脉的远端。这一次，因为主动脉的方式，确保捐赠者的​​功率放大器的左壁上的缝合是对下腔静脉的内侧。 </li><li>冲洗腔静脉管腔用肝素和生理盐水溶液（100单位/ ml）。通过不断地缝合他们到远端关闭捐助PA和接受者下腔静脉的右侧壁。 </li><li>删除远端结扎并通过应用外用无菌可吸收止血剂控制出血。当出血完全停止后，取出近端缝合线和控制出血的方法相同。 </li><li>返回的肠子，用6-0黑色尼龙单丝缝合关闭腹部肌肉和皮肤的两层。 </li><li>注射0.5毫升皮下注射生理盐水，然后将鼠标在一个变暖垫恢复笼，直到鼠标是完全移动。恢复后，返回鼠标到一个新的笼子纸床上用品。给予止痛药（布洛芬，30毫克/千克，生活饮用水）48小时。做不会返回已经动过手术，以其他动物的公司，直到完全康复的动物。 </li></ol>

<ol>
	<li>Polito, 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=Increased+morbidity+and+mortality+in+very+preterm+VLBW+infants+with+congenital+heart+disease.">Increased morbidity and mortality in very preterm VLBW infants with congenital heart disease.</a> Intens Care Med. 39, 1104-1112 (2013).</li><li>Wren, C., Reinhardt, Z., Khawaja, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Twenty+year+trends+in+diagnosis+of+life+threatening+neonatal+cardiovascular+malformations.">Twenty year trends in diagnosis of life threatening neonatal cardiovascular malformations.</a> Arch Dis Child Fetal. 93, F33-F35 (2008).</li><li>Vacanti, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Beyond+transplantation+Third+annual+Samuel+Jason+Mixter+lecture.">Beyond transplantation Third annual Samuel Jason Mixter lecture.</a> Archives of surgery. 123, 545-549 (1960).</li><li>Tudorache, I., 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=Orthotopic+replacement+of+aortic+heart+valves+with+tissue-engineered+grafts.">Orthotopic replacement of aortic heart valves with tissue-engineered grafts.</a> Tissue engineering Part A. 19, 1686-1694 (2013).</li><li>van Geldorp, M. W., 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=Patient+outcome+after+aortic+valve+replacement+with+a+mechanical+or+biological+prosthesis+weighing+lifetime+anticoagulant+related+event+risk+against+reoperation+risk.">Patient outcome after aortic valve replacement with a mechanical or biological prosthesis weighing lifetime anticoagulant related event risk against reoperation risk.</a> The Journal of thoracic and cardiovascular surgery. 137, 881-886 (2009).</li><li>El Oakley, R., Kleine, P., Bach, D. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Choice+of+prosthetic+heart+valve+in+today's+practice.">Choice of prosthetic heart valve in today's practice.</a> Circulation. 117, 253-256 (2008).</li><li>Quinn, R. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Animal+Models+for+Bench+to+Bedside+Translation+of+Regenerative+Cardiac+Constructs.">Animal Models for Bench to Bedside Translation of Regenerative Cardiac Constructs.</a> Progress in Pediatric cardiology. 35, (2013).</li><li>Shinoka, T., 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=Tissue+engineering+heart+valves+valve+leaflet+replacement+study+in+a+lamb+model.">Tissue engineering heart valves valve leaflet replacement study in a lamb model.</a> The Annals of thoracic surgery. 60, S513-S516 (1995).</li><li>Sodian, R., 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=Tissue+engineering+of+heart+valves+in+vitro+experiences.">Tissue engineering of heart valves in vitro experiences.</a> The Annals of thoracic surgery. 70, 140-144 (2000).</li><li>Shinoka, T., 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=Tissue+engineered+heart+valves.+Autologous+valve+leaflet+replacement+study+in+a+lamb+model.">Tissue engineered heart valves. Autologous valve leaflet replacement study in a lamb model.</a> Circulation. 94, II164-II168 (1996).</li><li>Sodian, R., 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=Evaluation+of+biodegradable+three+dimensional+matrices+for+tissue+engineering+of+heart+valves.">Evaluation of biodegradable three dimensional matrices for tissue engineering of heart valves.</a> ASAIO journal (American Society for Artificial Internal Organs. 46, 107-110 (1992).</li><li>Sodian, R., 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=Early+in+vivo+experience+with+tissue-engineered+trileaflet+heart+valves).">Early in vivo experience with tissue-engineered trileaflet heart valves).</a> Circulation. 102, III22-III29 (2000).</li></ol>

我们什么都没有透露。

死亡率此程序是接近20％，这主要是由出血在PV移植部位吻合捐助主动脉到受体腹主动脉造成的。在大多数情况下，死亡率显著减小后48小时的手术。存活小鼠表现出通过植入光伏强劲的心脏跳动和血液的流动。整个过程需要四个小时有经验的外科医生微。这将需要大约250小鼠掌握的技术。异位心脏移植是比较直截了当相比，光伏植入的供体心脏。其中一个成功的高压移植中最关键的步骤是收获从?...

先天性心血管缺陷的婴儿死亡率在西方世界1,2的主要原因之一。其中，肺动脉瓣狭窄和主动脉瓣二瓣缺陷是一个经常出现的形式3。心脏瓣膜置换手术是修复手术的常规选择;然而，并发症，包括狭窄的心脏瓣膜的钙化，以及终身抗凝药物的依赖是慢性疾病和死亡4-7一个显著来源。此外，缺乏增长潜力，需要重新进行手术，这进一步增加了这些年轻患者4,8,9的死亡率。 在试图开发一种功能置换心脏瓣膜的增长潜力，Shinoka 等。种子自体细胞到可生物降解的合成心脏瓣膜8。合成阀转换成一个原生心脏瓣膜状结构与增长电位人。初步大型动物的研究表明使用这种方法来创建一个功能心脏瓣膜10的可行性。然而，长期植入的研究表明耐用性差，由于该阀neotissue导致心脏瓣膜变窄的逐渐增厚。从Sodian工作等使用的Shinoka方法，但最终换成了PGA矩阵，可生物降解弹性体，这给了组织工程瓣膜的生物力学特性构建一个更符合生理轮廓9,11,12。在体内研究中，尽管植入的成功，一个融合的内皮细胞衬里没有形成这可能限制了长期的成功这个支架12。 为了合理设计改进的第二代人造心脏瓣膜，建立心脏瓣膜移植的小鼠模型来研究underlyin的细胞和分子机制克neotissue形成，瓣膜增厚，管腔狭窄和发展。小鼠模型提供了分子试剂繁多，包括转基因，这是不是在其他物种7一应俱全。在这种心脏瓣膜移植模型， 离体同源的肺心脏瓣膜置换首先执行;然后与植入心脏瓣膜的心脏被植入异位成利用显微外科技术一同源主机。通过此模型，心脏瓣膜置换，无需体外循环。 在本文中，心脏瓣膜收获的详细说明，供体心脏的准备工作，心脏瓣膜移植和异位心脏移植模型描述。结果表明，连续的心跳从供体心脏，这是独立于接收者的心跳。通过植入肺动脉瓣的血流使用高频超声系统用脉冲波执行测量ppler。

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			<td height="20" style="height:20px;width:176px;">DPBS</td>
			<td style="width:167px;">gibco</td>
			<td style="width:133px;">14190-144</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:176px;">Microscope</td>
			<td style="width:167px;">Leica</td>
			<td style="width:133px;">M80</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:176px;">C57BL/6J (H-2b), Female</td>
			<td style="width:167px;">Jackson Laboratories&#160;</td>
			<td style="width:133px;">664</td>
			<td style="width:168px;">8-12 weeks</td>
		</tr>
		<tr height="39">
			<td height="39" style="height:39px;width:176px;">Ketamine Hydrochloride Injection</td>
			<td style="width:167px;">Hospira Inc.</td>
			<td style="width:133px;">NDC 0409-2053</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:176px;">Xylazine Sterile Solution</td>
			<td style="width:167px;">Akorn Inc.</td>
			<td style="width:133px;">NADA# 139-236</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="36">
			<td height="36" style="height:36px;width:176px;">ketoprofen</td>
			<td style="width:167px;">Fort Dodge Animal Health</td>
			<td style="width:133px;">NDC 0856-4396-01</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="36">
			<td height="36" style="height:36px;width:176px;">Ibuprofen</td>
			<td style="width:167px;">PrecisionDose</td>
			<td style="width:133px;">NDC 68094-494-59</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="37">
			<td height="37" style="height:37px;width:176px;">Heparin Sodium</td>
			<td style="width:167px;">Sagent Pharmaceticals</td>
			<td style="width:133px;">NDC 25021-400</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="39">
			<td height="39" style="height:39px;width:176px;">Saline solution (Sterile 0.9% Sodium Chloride)</td>
			<td style="width:167px;">Hospira Inc.</td>
			<td style="width:133px;">NDC 0409-0138-22</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="39">
			<td height="39" style="height:39px;width:176px;">0.9% Sodium Chloride Injection</td>
			<td style="width:167px;">Hospira Inc.</td>
			<td style="width:133px;">NDC 0409-4888-10</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="41">
			<td height="41" style="height:41px;width:176px;">Petrolatum Ophthalmic Ointment</td>
			<td style="width:167px;">Dechra Veterinary Products</td>
			<td style="width:133px;">NDC 17033-211-38</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:176px;">Iodine Prep Pads</td>
			<td style="width:167px;">Triad Disposables, Inc.</td>
			<td style="width:133px;">NDC 50730-3201-1</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:176px;">Alcohol Prep Pads</td>
			<td style="width:167px;">McKesson Corp.</td>
			<td style="width:133px;">NDC 68599-5805-1</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:176px;">Cotton tipped applicators</td>
			<td style="width:167px;">Fisher Sientific</td>
			<td style="width:133px;">23-400-118</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:176px;">Fine Scissor</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">14028-10</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:176px;">Micro-Adson Forcep</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">11018-12</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:176px;">Clamp Applying Forcep</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">00072-14</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:176px;">S&#38;T Vascular Clamp</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">00396-01</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:176px;">Spring Scissors</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">15008-08</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Colibri Retractors</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">17000-04</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Dumont #5 Forcep</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">11251-20&#160;</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Dumont #7 - Fine Forceps</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">11274-20</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Dumont #5/45 Forceps</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">11251-35</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Tish Needle Holder/Forceps</td>
			<td style="width:167px;">Micrins</td>
			<td style="width:133px;">MI1540</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="38">
			<td height="38" style="height:38px;width:176px;">Black Polyamide Monofilament Suture, 10-0</td>
			<td style="width:167px;">AROSurgical Instruments Corporation</td>
			<td style="width:133px;">TI638402</td>
			<td style="width:168px;">For sutureing the graft</td>
		</tr>
		<tr height="37">
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transplantation of pulmonary valve using a mouse model of heterotopic heart transplantation

组织工程心脏瓣膜，特别是脱细胞瓣膜，也开始获得动力在临床使用重建手术的结果好坏参半。然而，neotissue发展，阀增厚和狭窄发展的细胞和分子机制尚未广泛研究。要回答上述问题，我们开发了一个小鼠异位心脏瓣膜移植模型。心脏瓣膜是从阀供体小鼠收获移植到心脏供体小鼠。用一个新的阀的心脏异位移植到受体小鼠。移植心脏表明自己的心跳，独立收件人的心跳。使用高频超声系统用脉冲多普勒血流进行定量。通过植入肺动脉瓣流量呈正向流量以最小的反流和洪峰流量接近100毫米/秒。心脏瓣膜移植的这个小鼠模型是highlÝ通用性，因此它可以被修改和调整以提供不同的血流动力学的环境和/或可用于各种转基因小鼠来研究neotissue发展在组织工程心脏瓣膜。

图1说明了心脏瓣膜移植模型采用异位心脏移植的原理图。心脏瓣膜是从供体心脏收获，并从第二供体小鼠植入到心脏。然后用新的心脏瓣膜的心脏被植入到受体小鼠的腹部。 图2显示了植入心脏的插图上腹部空间（A），心脏移植后（B）的权利，移植后5分钟。在剔除对主动脉和下腔静脉的两侧缝线，心脏开始后打1-2分钟，成为平克与更多的血液?...

Watch this Scientific Journal Video about Transplantation of Pulmonary Valve Using a Mouse Model of Heterotopic Heart Transplantation at JoVE.com

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

为了了解neotissue形成和狭窄的发展在组织工程心脏瓣膜相关的细胞和分子机制，开发了异位心脏瓣膜移植的小鼠模型。使用异位心脏移植术肺动脉心脏瓣膜移植到收件人。

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 ...

transplantation-pulmonary-valve-using-mouse-model-heterotopic-heart

Research

JoVE Journal

Medicine

18.8K 次观看.  Nationwide Children's Hospital. 为了了解neotissue形成和狭窄的发展在组织工程心脏瓣膜相关的细胞和分子机制，开发了异位心脏瓣膜移植的小鼠模型。使用异位心脏移植术肺动脉心脏瓣膜移植到收件人。 

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

A Mouse Model of in Utero Transplantation

The transplantation of stem cells and viruses in utero has tremendous potential for treating congenital disorders in the human fetus. For example, in utero transplantation (IUT) of hematopoietic stem cells has been used to successfully treat patients with severe combined immunodeficiency.1,2 In several other conditions, however, IUT has been attempted without success.3 Given these mixed results, the availability of an efficient non-human model to study the biological sequelae of stem cell transplantation and gene therapy is critical to advance this field. We and others have used the mouse model of IUT to study factors affecting successful engraftment of in utero transplanted hematopoietic stem cells in both wild-type mice4-7 and those with genetic diseases.8,9 The fetal environment also offers considerable advantages for the success of in utero gene therapy. For example, the delivery of adenoviral10, adeno-associated viral10, retroviral11, and lentiviral vectors12,13 into the fetus has resulted in the transduction of multiple organs distant from the site of injection with long-term gene expression. in utero gene therapy may therefore be considered as a possible treatment strategy for single gene disorders such as muscular dystrophy or cystic fibrosis. Another potential advantage of IUT is the ability to induce immune tolerance to a specific antigen. As seen in mice with hemophilia, the introduction of Factor IX early in development results in tolerance to this protein.14 
In addition to its use in investigating potential human therapies, the mouse model of IUT can be a powerful tool to study basic questions in developmental and stem cell biology. For example, one can deliver various small molecules to induce or inhibit specific gene expression at defined gestational stages and manipulate developmental pathways. The impact of these alterations can be assessed at various timepoints after the initial transplantation. Furthermore, one can transplant pluripotent or lineage specific progenitor cells into the fetal environment to study stem cell differentiation in a non-irradiated and unperturbed host environment.
The mouse model of IUT has already provided numerous insights within the fields of immunology, and developmental and stem cell biology. In this video-based protocol, we describe a step-by-step approach to performing IUT in mouse fetuses and outline the critical steps and potential pitfalls of this technique.

A Mouse Model of in Utero Transplantation

The mouse model of in utero transplantation is a versatile tool that can be used to study the potential clinical applications of stem cell transplantation and gene therapy in the fetus. In this protocol, we present a general approach to performing this technique

一个小鼠模型在子宫内移植

The transplantation of stem cells and viruses in utero has tremendous potential for treating congenital disorders in the human fetus.  For example, in ...

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 ...

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 ...

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 ...

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.

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 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 ...

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 ...