M.-A. Deutsch is supported by Dr. Rusche Forschungsprojekt (2014) Deutsche Stiftung f&#252;r Herzforschung and Deutsche Gesellschaft f&#252;r Thorax- Herz- und Gef&#228;&#223;chirurgie. M. Krane is supported by Deutsche Stiftung f&#252;r Herzforschung (F/37/11), Deutsches Zentrum f&#252;r Herz Kreislauf Forschung (DZHK B 13-050A; DZHK B 14-013SE) and Deutsche Forschungsgemeinschaft (KR3770/7-1; KR3770/9-1)

Todos os experimentos foram aprovados pelo alemão Heart Center Munique eo Governo da Baviera, Alemanha (Gz. 55.2-1-54-2532.3-17-13). Os animais foram alojados em uma instalação livre de patógenos específicos e recebeu cuidados humanos em conformidade com os &quot;Princípios de Laboratório Animal Care&quot; formuladas pela Sociedade Nacional de Pesquisa Médica e do &quot;Guia para o Cuidado e Uso de Animais de Laboratório&quot; fornecido pela Academia Nacional de Ciências e publicado pelos Institutos Nacionais de Saúde (NIH publicação 86-23 no., revista, 1985). NOTA: Um conjunto completo de instrumentos de microcirurgia estéreis, incluindo duas pinças curvas e dilatadores dos vasos, é necessário para o procedimento (Figura 2a). Profilaxia com antibióticos perioperatórios não é rotineiramente necessária, mas pode ser indicada em animais imunocomprometidos. Os animais foram sacrificados usando inalação de isoflurano terminal. Doador de Preparação e Graft Colheita &lt;ol&gt; <li> Anestesiar o animal por via intraperitoneal (ip) de injecção de midazolam (5,0 mg / kg), medetomidina (0,5 mg / kg) e fentanil (0,05 mg / kg de peso corporal). Posicione o mouse em decúbito dorsal sobre o campo operatório e confirmar tolerância cirúrgico pelo reflexo interdigital. </li><li> Desinfectar a pele três vezes com clorexidina e realizar uma incisão na linha média abdominal. Recolha os intestinos com gaze húmida para a esquerda para expor a veia cava inferior. </li><li> Injectar 50 UI de heparina diluído em 0,4 ml de solução salina através da veia cava inferior para a anticoagulação. Faça uma incisão na embarcação 1 min após heparinização. </li><li> Abra o diafragma e executar uma toracotomia bilateral para expor o coração. A parede anterior do tórax é refletido superiormente e mantido no lugar com um grampo. </li><li> Remova o timo e sem rodeios dissecar o tecido conjuntivo entre aorta e tronco pulmonar. Faça uma incisão na veia cava superior direita. </li><li> Perfurar a aorta ascendente mobilizado no nível doTBE com uma agulha G 30, montado sobre uma seringa de 5 ml e perfundir lentamente o coração retrogradually sem pressão excessiva, com 3 ml de 4 ° C, solução cardioplégica a frio. Evitar a entrada de ar para dentro da circulação coronária. Largar solução salina gelada para o enxerto para aumentar ainda mais a parada cardíaca. </li><li> Transecto a aorta proximal à origem do tronco braquiocefálico. </li><li> Ligadura da veia inferior e ambos cavas superior perto do coração com 8-0 seda e transecto-los para distal para as ligaduras. </li><li> Dissecar livre do tronco pulmonar como distalmente quanto possível. Ligadura das artérias pulmonares perto da bifurcação com 8-0 seda e transecto-los para distal. </li><li> Ligadura das veias pulmonares todos juntos com um único ligadura 8-0 seda e transecto-los. </li><li> Tiraram o coração do doador e armazená-lo em solução cardioplégica gelada até a implantação. </li></ol> 2. Destinatário Preparação <ol><li> Prepararos punhos para anastomoses por corte do tubo de poliimida com um bisturi n ° 11 ou uma tesoura ao microscópio. A braçadeira é constituída por um corpo cilíndrico e uma extensão (1/3 da circunferência) para a colocação da pinça vascular, cada uma com um comprimento de 1 mm (Figura 2b). </li><li> Anestesiar o animal e confirmar tolerância cirúrgico, tal como descrito acima. Injectar 0,25 ml de soro fisiológico morno solução por via subcutânea (sc) para a reposição de líquidos. </li><li> Remover os pêlos da região cervical direita usando um barbeador elétrico e colocar o animal em decúbito dorsal sobre uma almofada quente. Prenda os pés com fios de fita, evitando overstretching os membros dianteiros, pois isso pode comprometer a respiração. Cubra os olhos usando pomada oftálmica para prevenir o ressecamento. Desinfetar o local da cirurgia três vezes com clorexidina. </li><li> Faça uma incisão transversa da pele a partir do ângulo da mandíbula direito ao entalhe jugular. </li><li> Sem rodeios mobilizar a extErnal veia jugular. Cauterizar ramos laterais com um fórceps bipolares e transecto-los. </li><li> Dividir a veia jugular externa entre ligaduras no nível de confluência com a veia a partir da glândula submandibular. </li><li> Puxe a veia jugular externa através do manguito e oclusão do vaso proximal, colocando um grampo vascular sobre a extensão do punho. Remova a ligadura e irrigar o lúmen do vaso com 01:10 salina heparinizada. </li><li> Evert parede do vaso através da braçadeira e fixe-o com uma predefinição circunferencial 8-0 seda ligadura. </li><li> Remova a parte superficial do músculo esternocleidomastóideo. </li><li> Mobilizar a artéria carótida comum direita e transecto embarcação entre ligaduras abaixo da bifurcação carotídea. </li><li> Puxe a artéria carótida através do manguito e oclusão do vaso proximal, colocando um grampo Yasargil sobre a extensão do punho. Remova a ligadura e irrigar o lúmen do vaso com 01:10 salina heparinizada. </li><li> Gentilmente dilate do lúmen arterial, com um dilatador navio, everter parede do vaso através da braçadeira e fixe-o com uma predefinição circunferencial 8-0 seda ligadura. </li><li> Para ganhar espaço suficiente para a implantação do enxerto, remover o lobo direito da glândula submandibular. </li><li> Mantenha o campo operatório úmido até a implantação. </li></ol> 3. Enxerto Implantação <ol><li> Coloque o enxerto na região cervical do receptor de cabeça para baixo com a aorta orientada medialmente e a artéria pulmonar lateralmente. </li><li> Puxe a aorta do enxerto sobre o manguito com a artéria carótida e garantir a anastomose com uma ligadura circunferencial 8-0 seda. </li><li> Inciso do tronco pulmonar no seu aspecto anterior e puxe-o sobre o manguito com a veia jugular externa. Novamente garantir a anastomose com uma ligadura circunferencial 8-0 seda. </li><li> Remover o grampo na veia jugular externa, seguindo por desclampeamento da artéria carótida. Os corações enche imediatamente com sangue. Para aumentar reperfusion cair 37 ° C soro fisiológico morno para o enxerto. Normalmente, o coração se desenvolve em ritmo sinusal dentro de um minuto. Remover o manguito extensão no local venoso. </li><li> Assegurar a posição correta do enxerto, anastomoses deve ser livre de tensão e vasos sem torção. Não vasos cauterizados da pele uma vez que este pode prejudicar a cicatrização de feridas. Sangramento geralmente pára espontaneamente. Fechar a pele com uma sutura contínua única 8-0. </li></ol> 4. Cuidados Pós-Operatórios <ol><li> Injectar 0,25 ml quente sc solução salina para a reposição de líquidos. </li><li> Para antagonizar narcóticos injectar a naloxona (1,2 mg / kg), o flumazenil (0,5 mg / kg) e atipamezol (2,5 mg / kg de peso corporal) SC esquerdo do animal na almofada quente até ter recuperado a consciência suficiente para manter decúbito esternal. Observar o animal durante pelo menos uma hora adicional para registar qualquer comportamento anormal. Não devolva um animal que tenha sido submetido a cirurgia para a companhia de outros animais até que esteja totalmente Recovevermelho. </li><li> Fornecer analgesia preemptiva com buprenorfina (0,05 mg / kg sc três vezes por dia) ou uma analgesia alternativa de acordo com um protocolo institucional local por um período mínimo de 72 horas após o procedimento. </li><li> O cirurgião e um veterinário deve ver os animais regularmente. Em caso de perda de peso&gt; 15%, a apatia, inchaço ou infecção lado cirúrgicos animais são sacrificados usando inalação de isoflurano terminal. </li></ol>

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N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessment+of+graft+function+in+rodent+models+of+heart+transplantation.">Assessment of graft function in rodent models of heart transplantation.</a> Microsurgery. 28, 565-570 (2008).</li><li>Schramm, 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=The+subepicardial+microcirculation+in+heterotopically+transplanted+mouse+hearts:+an+intravital+multifluorescence+microscopy+study.">The subepicardial microcirculation in heterotopically transplanted mouse hearts: an intravital multifluorescence microscopy study.</a> J Thorac Cardiovasc Surg. 134, 210-217 (2007).</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 : JoVE. 238, (2007).</li><li>Gong, 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=Introduction+of+modified+cervical+cardiac+transplant+model+in+mice.">Introduction of modified cervical cardiac transplant model in mice.</a> Exp Clin Transplant. 10, 158-162 (2012).</li><li>Matsuura, A., Abe, T., Yasuura, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Simplified+mouse+cervical+heart+transplantation+using+a+cuff+technique.">Simplified mouse cervical heart transplantation using a cuff technique.</a> Transplantation. 51, 896-898 (1991).</li><li>Chen, H., Zhang, Y., Zheng, D., Praseedom, R. K., Dong, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Orthotopic+kidney+transplantation+in+mice:+technique+using+cuff+for+renal+vein+anastomosis.">Orthotopic kidney transplantation in mice: technique using cuff for renal vein anastomosis.</a> PloS one. 8, e77278 (2013).</li><li>Maglione, 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+technique+for+heterotopic+vascularized+pancreas+transplantation+in+mice+to+assess+ischemia+reperfusion+injury+and+graft+pancreatitis.">A novel technique for heterotopic vascularized pancreas transplantation in mice to assess ischemia reperfusion injury and graft pancreatitis.</a> Surgery. 141, 682-689 (2007).</li><li>Sucher, 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=Orthotopic+hind-limb+transplantation+in+rats.">Orthotopic hind-limb transplantation in rats.</a> J Vis Exp : JoVE. , (2010).</li><li>Kamada, N., Calne, R. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Orthotopic+liver+transplantation+in+the+rat.+Technique+using+cuff+for+portal+vein+anastomosis+and+biliary+drainage.">Orthotopic liver transplantation in the rat. Technique using cuff for portal vein anastomosis and biliary drainage.</a> Transplantation. 28, 47-50 (1979).</li><li>Oldani, G., Lacotte, S., Morel, P., Mentha, G., Toso, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Orthotopic+liver+transplantation+in+rats.">Orthotopic liver transplantation in rats.</a> J Vis Exp : JoVE. , (2012).</li><li>Okazaki, 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+mouse+model+of+orthotopic+vascularized+aerated+lung+transplantation.">A mouse model of orthotopic vascularized aerated lung transplantation.</a> Am J Transplant. 7, 1672-1679 (2007).</li><li>Suzuki, H., Fan, L., Wilkes, 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=Development+of+obliterative+bronchiolitis+in+a+murine+model+of+orthotopic+lung+transplantation.">Development of obliterative bronchiolitis in a murine model of orthotopic lung transplantation.</a> J Vis Exp : JoVE. , (2012).</li><li>Zou, Y., 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=Mouse+model+of+venous+bypass+graft+arteriosclerosis.">Mouse model of venous bypass graft arteriosclerosis.</a> Am J Pathol. 153, 1301-1310 (1998).</li><li>Zhou, Y., Gu, X., Xiang, J., Qian, S., Chen, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+comparative+study+on+suture+versus+cuff+anastomosis+in+mouse+cervical+cardiac+transplant.">A comparative study on suture versus cuff anastomosis in mouse cervical cardiac transplant.</a> Exp Clin Transplant. 8, 245-249 (2010).</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+3000+operations+by+one+surgeon.">The technique for heterotopic cardiac transplantation in mice: experience of 3000 operations by one surgeon.</a> J Heart Lung Transplant. 20, 1123-1128 (2001).</li><li>Arras, M., Autenried, P., Rettich, A., Spaeni, D., Rulicke, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optimization+of+intraperitoneal+injection+anesthesia+in+mice:+drugs,+dosages,+adverse+effects,+and+anesthesia+depth.">Optimization of intraperitoneal injection anesthesia in mice: drugs, dosages, adverse effects, and anesthesia depth.</a> Comp Med. 51, 443-456 (2001).</li></ol>

The authors have nothing to disclose.

Nós completamos mais de 200 transplantes cardíacos com uma taxa de sucesso superior a 90% em várias configurações experimentais. Com a técnica descrita aqui, o procedimento completo pode ser feito no prazo de 60 min por um cirurgião experiente. Ao preparar os vasos cervicais antes de colheita de órgãos, o tempo total de isquemia do enxerto pode ser limitada a 20 min. Tempo para o implante (tempo de isquemia quente) é significativamente mais curta utilizando a técnica de braçadeira em comparação com a técn...

O modelo de transplante cardíaco é freqüentemente usada para investigar I / R-lesão, rejeição do enxerto, doença vascular pós-transplante ea eficiência dos agentes imunomoduladores. Mouse modelos combinam muitas vantagens como o imunológica conhecida e origens genéticas, a disponibilidade de muitas estirpes puras e transgénicas experimentais e custos relativamente baixos. Em 1973, uma técnica de transplante cardíaco abdominal em ratinhos foi descrita pela primeira vez por Corry et al 1. Neste modelo enxertos são revascularizada pelo ponta-a-lado anastomose da aorta ascendente a aorta abdominal do receptor e da principal artéria pulmonar à veia cava inferior. Em 1991, a técnica de sutura para o transplante cardíaco cervical em ratos foi descrita por Chen et al 2. Neste modelo a perfusão coronária retrógrada é estabelecido por anastomose da artéria carótida do destinatário à aorta ascendente do grpopa. Os drenos de sangue venoso, através do seio coronário no átrio direito, ventrículo direito e é ejetado através da artéria pulmonar para o destinatário veia jugular externa (Figura 1). Em comparação com a implantação do enxerto abdominal modelo cervical é menos invasiva ea localização superficial do enxerto permite um acesso muito fácil para exames de acompanhamento adicionais (por exemplo palpação, ecocardiografia, microscopia de fluorescência intravital) 3,4. Apesar das melhorias técnicas 5,6, uma utilização mais generalizada deste modelo tem sido limitada devido a dificuldades técnicas para realizar anastomoses suturadas de pequenos vasos com alta incidência de vazamento de anastomose e trombose. Em 1991, Matsuura et ai. introduziu o manguito técnica em que a parede do vaso é revirado sobre um cilindro de síntese, o que facilita grandemente o modelo de transplante de coração 7 do colo do útero. A técnica foi further adotado para vários modelos experimentais de transplante renal, incluindo 8, 9 pancreático, membro 10, hepáticos 11,12, 13,14 e transplante de pulmão vascular 15. Em comparação com a técnica de sutura o tempo necessário para a anastomose e, assim, o tempo de isquemia quente é mais curto e existem complicações vasculares significativamente menor, utilizando a técnica do manguito 16. Além disso, ele permite que os cirurgiões com pouco treinamento em microcirurgia para executar a operação com uma elevada taxa de sucesso. Uma desvantagem da técnica de braçadeira é pré-requisito para a ligação da artéria carótida comum, o que pode alterar a perfusão cerebral. Neste artigo, apresentamos um vídeo técnica manguito melhorado e simplificado para o transplante cardíaco cervical em camundongos.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Forceps JFC-7.2</td>
			<td>S&#38;T</td>
			<td>P-00036</td>
			<td>Curved tip 0.20 mm</td>
		</tr>
		<tr>
			<td>Vessel dilatator D-5a</td>
			<td>S&#38;T</td>
			<td>S-00124</td>
			<td></td>
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		<tr>
			<td>Adventita scissor</td>
			<td>S&#38;T</td>
			<td>S-00102</td>
			<td></td>
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		<tr>
			<td>Vascular clamp B-1V</td>
			<td>S&#38;T</td>
			<td>S-00396</td>
			<td></td>
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		<tr>
			<td>Yasargil Clip</td>
			<td>Peter Lazic</td>
			<td>65,097</td>
			<td>3.5 mm lenght</td>
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		<tr>
			<td>Bipolar forceps</td>
			<td>Micromed</td>
			<td>148-100-011</td>
			<td>tip 0.25 mm</td>
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			<td>Polyimide tubes</td>
			<td>River Tech Medical</td>
			<td></td>
			<td>19-24 gauge</td>
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			<td>8.0 silk ligatures</td>
			<td>Catgut</td>
			<td></td>
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			<td>Custodiol HTK solution</td>
			<td>Essential Pharmaceuticals</td>
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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.

Nossa equipe pode executar com êxito a técnica descrita após cerca de 20 cirurgias de formação. A parte mais desafiadora é a everter parede do vaso da artéria carótida sobre o punho, porque facilmente vira para trás e múltiplas tentativas podem levar a rasgar do muro. Após o período de treinamento, o tempo cirúrgico médio total foi de 60 ± 8 min. Cerca de 20 min são necessários para a colheita doador, 25 min para a preparação dos vasos cervicais e 15 minutos para anastomoses e fechamento da pele. ...

Watch this Scientific Journal Video about Heterotopic Cervical Heart Transplantation in Mice at JoVE.com

Heterotopic Cervical Heart Transplantation in Mice

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.

Heterotopic Cervical Transplante Cardíaco em Ratos

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

heterotopic-cervical-heart-transplantation-in-mice

Research

JoVE Journal

Medicine

11.1K visualizações.  German Heart Center Munich, Technische Universität München. 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.

Vídeo: Heterotopic Cervical Heart Transplantation in Mice

Controlled Cervical Laceration Injury in Mice

Use of genetically modified mice enhances our understanding of molecular mechanisms underlying several neurological disorders such as a spinal cord injury (SCI). Freehand manual control used to produce a laceration model of SCI creates inconsistent injuries often associated with a crush or contusion component and, therefore, a novel technique was developed. Our model of cervical laceration SCI has resolved inherent difficulties with the freehand method by incorporating 1) cervical vertebral stabilization by vertebral facet fixation, 2) enhanced spinal cord exposure, and 3) creation of a reproducible laceration of the spinal cord using an oscillating blade with an accuracy of &plusmn;0.01 mm in depth without associated contusion. Compared to the standard methods of creating a SCI laceration such as freehand use of a scalpel or scissors, our method has produced a consistent lesion. This method is useful for studies on axonal regeneration of corticospinal, rubrospinal, and dorsal ascending tracts.

A novel technique to create a reproducible in vivo model of cervical spinal cord laceration injury in the mouse is described. This technique is based on spine stabilization by fixation of the cervical facets and laceration of the spinal cord using an oscillating blade with an accuracy of &plusmn;0.01 mm.

Controlada por laceração do colo do útero em camundongos

Use of genetically modified mice enhances our  understanding of molecular mechanisms underlying several neurological disorders  such as a spinal cord ...

Medicina

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.

Murino Cervical Modelo de coração Transplante usando uma técnica Cuff Modificado

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.

Um método modificado para heterotópico Rato coração transplante

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 ...

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.

Técnica de transplante heterotópico Murino Coração

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.

O transplante de válvula pulmonar Usando um modelo do rato de Heterotópica Transplante de Coração

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.

Rat heterotópico abdominal coração / pulmão Transplante Single-carregado em uma configuração-Volume

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

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.

A Microsurgical Modelo Novel para heterotópico, em bloco da parede torácica, Thymus, e Transplante Cardíaco em Ratos

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

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.

Um modelo simplificado para transplante de válvula cardíaca heterotópica em roedores

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.

Um Protocolo de Transplante de Coração Heterotópico Porcino para Entrega de Terapêutica a um Aoenxerto Cardíaco

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.

Técnica de Cuff Modificada para Modelo de Transplante Cardíaco Heterotópico Cervical:

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

Methods Article

Heterotopic Cervical Transplante Cardíaco em Ratos