CT foi apoiada pela National Science Foundation suíço (SCORE concessão 3232230-126233). O estudo foi financiado pela Fundação Artères, a Astellas Foundation Europe e da Fundação Boninchi.

1. Operação doador <ol><li> Anestesiar ratos machos, pesando 200 ± 20 gramas, com isoflurano em cone máscara (3% para a indução, 2% durante a operação, um fluxo de ar l / min, FiO2 70%). </li><li> Realize uma laparotomia mediana e grande transversal e cauterização dos vasos epigástricos em ambos os lados. </li><li> Cortar o ligamento falciforme, e separar a veia esquerdo do diafragma a partir da veia cava suprehepatic. Finalmente, dividir a veia do diafragma entre duas ligaduras de seda 7-0. </li><li> Corte o triangular esquerdo e os ligamentos gastro-hepáticas. </li><li> Divida o ligamento hepato-esofágico e artéria entre 7-0 ligaduras (coagulação é uma alternativa válida). </li><li> Isolar o infrahepatic veia cava para baixo para a veia renal esquerda. Separa-se a veia renal direita a partir dos tecidos circundantes e dividi-lo entre um 10-0 e uma ligadura 7-0, nas suas extremidades proximal e distai, respectivamente. </li><li> Divida a veia supra-renal direita entre 7-0 ligatures e livre o fígado de seus ligamentos posteriores cortando sob leve tração. </li><li> Isolar a veia gastroesplênico e dividi-lo entre um 10-0 e uma ligadura 7-0, nas suas extremidades proximal e distai, respectivamente. </li><li> Divida a artéria hepática própria entre 7-0 laços. </li><li> Isolar a veia duodeno pâncreas e dividiu-a entre um 10-0 e uma ligadura 7-0, em sua extremidade proximal e distal, respectivamente. </li><li> Insira um 22G, 3,5 mm de stent no ducto biliar comum através da prática de uma pequena incisão de 1 cm de distância a partir do hilo hepático. Fixe o stent em posição com uma ligadura 7-0. </li><li> Injectar 10 UI de heparina, diluiu-se em 1 ml de solução salina normal, através da veia dorsal do pénis. </li><li> Canular a porta cava (tanto quanto possível a partir do hilo) com uma agulha de 21G e suavemente lavar o fígado com 20 ml de lactato de Ringer frio (ou outras soluções de conservação). Ao mesmo tempo, cortar a veia cava inferior das veias renais para permitir uma adequada outflow. Perfusão de fígado frio deve durar entre um e dois minutos. </li><li> Corte a porta abaixo da veia cava duodeno pâncreas. </li><li> Corte o duto biliar comum distal à ligadura ao redor do stent. </li><li> Complete a incisão da veia cava infrahepatic. </li><li> Cortar o supra-hepática da veia cava desnatado para o diafragma e remover o fígado. </li><li> Coloque o fígado em uma bacia cheia de ringer lactato fria e colocar a bacia em uma almofada de gelo. </li></ol> 2. Preparação do Enxerto <ol><li> Insira o porta veia em seu punho, então everter o navio ao redor da borda para expor a íntima. Fixe o segmento evertido no punho com uma ligadura de 7-0. </li><li> Repita a mesma operação no infrahepatic veia cava. </li><li> Posicionar uma braçadeira de micro-(4-6 mm de comprimento) na parte proximal do infrahepatic veia cava. Esta operação evita a perda de sangue após a reperfusão do portal. </li><li> Derrubar o fígado e inserir duas prolene 8-0 suturasnas bordas opostas laterais da supra-hepática da veia cava, de fora para dentro. </li><li> Guarde o enxerto (imersos em solução de ringer lactato) na geladeira a 0-4 ° C. </li></ol> 3. Operação destinatário <ol><li> Anestesiar ratos machos, pesando 200 ± 20 gramas, com isoflurano em cone máscara (3% para a indução, 2% durante a operação, um fluxo de ar l / min, FiO2 70%) e manter em uma almofada de aquecimento. De nota, pesos receptores e os doadores devem ser combinados (± 40 gramas). </li><li> Injectar 10 ml de soro fisiológico por via subcutânea e 0,03 g de piperacilina / tazobactam intramuscular antes da laparotomia. </li><li> Realize uma laparotomia xipho-púbica na linha média. </li><li> Inverter o processo xifóide com uma pinça de auto-estáticas, a fim de melhor expor o fígado. </li><li> Corte os ligamentos anterior e dividir a veia diafragmática esquerda entre dois laços 7-0. </li><li> Divida o ligamento hepato-esofágico e artéria entre 7-0 ligaduras (coagulation é uma alternativa válida). Não é mostrado no vídeo (idêntica à operação de dador). </li><li> Isolar o infrahepatic veia cava até a veia renal direita. </li><li> Dividir a veia supra-renal direita entre 7-0 ligaduras e fígado o livre a partir dos seus ligamentos posteriores cortando sob uma ligeira tracção (não mostrado no vídeo (idêntica à operação de dador)). </li><li> Coloque uma ligadura 7-0 ao redor do ducto biliar comum, logo abaixo de sua divisão. Dito de outra ligadura um milímetro abaixo e cortar no meio. </li><li> Isolar a artéria hepática no hilo e dividi-lo entre 7-0 ligaduras. </li><li> Gentilmente separar ramos portais direito e esquerdo. Preste atenção para não danificar a artéria hepática comum ou a veia gastro-esplênica. </li><li> Posicione o anel ao redor da cava da veia cava, cranialmente até a veia renal direita. Em seguida, fixar a parede do vaso para o anel através da colocação de quatro cardinais 9-0 suturas. </li><li> Repita a mesma operação com o anel de portal na porta cava. </li&gt; <li> Injectar 10 UI de heparina, diluiu-se em 1 ml de solução salina normal, através da veia dorsal do pénis. </li><li> Definir a anestesia em 0,25% ea FiO2 em 100%. </li><li> Prenda o infrahepatic veia cava logo acima da veia renal direita, em seguida, apertar a veia porta logo acima da veia gastro-esplênica. Finalmente prender o supra-hepática da veia cava não se esquecendo de incluir pelo menos 2 mm do diafragma. </li><li> Corte o supra-hepática da veia cava próxima ao parênquima. </li><li> Corte a esquerda e os ramos direito do portal e dividir o septo entre os dois. </li><li> Corte o infrahepatic veia cava próximo ao parênquima, prestando atenção para não incisar qualquer dos pontos previamente posicionados. Remover o fígado e colocar o enxerto para dentro da cavidade abdominal. </li><li> Realizar uma anastomose ponta a ponta entre a cava supra-hepática veia do enxerto e do receptor. Comece com a sutura da parede posterior da esquerda para a direita, em seguida, proceder na parede anterior da direita para a leManter o pé de sutura anterior solto até ao fim, a fim de permitir que um rubor generosa do lúmen interior antes de apertar as extremidades. </li><li> Insira o ramo caudal da aproximador no punho anel portal, em seguida, insira o porta cava do enxerto na fenda do ramo cranial da aproximador, com a braçadeira portal encontrar sua posição no slot. </li><li> Feche o aproximador durante a lavagem do destinatário veia porta com solução salina normal e permitir que a braçadeira para inserir dentro do vaso. Fixe o destinatário da veia porta em torno de seu punho com um empate 7-0 circunferencial. </li><li> Remover o aproximador. Abra o grampo na veia cava supra-hepática ea veia porta em ordem: fluxo portal está restabelecida. </li><li> Retire o anel portal cortando as ligaduras 9-0 com extra-finas tesoura. </li><li> Inserir o ramo caudal do aproximador no punho anel cava; então inserir o infrahepatic veia cava do enxerto na fenda do crânio<html> ramo da aproximador, com a braçadeira cava encontrar sua posição no slot. </li><li> Feche o aproximador durante a lavagem do destinatário infrahepatic veia cava com solução salina normal e permitir que a braçadeira para inserir dentro do vaso. Fixe o destinatário da veia cava em torno de seu punho com um empate 7-0 circunferencial. </li><li> Remover o aproximador. Abra as microclamps no infrahepatic veia cava do enxerto e do receptor em ordem: fluxo venoso está agora restabelecida. </li><li> Realizar uma incisão parcial do ducto do destinatário biliar comum, aplique uma leve tração para a cabeça, e insira o lado livre do stent previamente inserido no ducto biliar do enxerto. Fixe o stent com um empate 7-0 circunferencial. </li><li> Feche o animal por camadas com 5-0. Permitir que a água livre e comida de despertar. </li><li> Administrar analgesia eficaz de acordo com protocolos institucionais locais. Uma duração mínima de 24 h é recomendada. </li></ol>&gt; _content &quot;Digno de nota: fim-de-final da anastomose da artéria hepática pode ser realizada, quando exigido pelo projeto experimental (a artéria não é necessário para a sobrevivência a longo prazo após o transplante de fígado de rato). 4. Os resultados representativos A técnica descrita pode ser dominado com sucesso pela nossa equipe após cerca de 10 cirurgias de treinamento. Ele permitiu que 100% sobrevivência a longo prazo (&gt; 21 dias) em 36 consecutivo Lewis-a-Lewis e Agouti-a-Lewis Trevas transplantes, após o período de formação de curta duração (um número de receptores de enxertos singênicos fígado foram mantidos vivos por mais de 12 meses agora). Pós-transplante singénicos dia 1 testes de função hepática mostrou aspartato transaminase mediana (AST): 84 U / l (51,7-92,7) alanina transaminase e mediana (ALT): 172,5 U / l (127,5-240,2). A fase anepática média (de veia porta de fixação de enxerto re-perfusão) foi de 14 ± 2 min, com o maior passo de ser o supra-hepática da veia cava anastomose (executando suture) tempo de 9 ± 2 min. O sistema quick-linker permitiu o posicionamento de 1,55 mm de calibre algemas em veia porta e 2,40 mm de calibre punhos na veia cava (destinatários pesando 200 ± 20 g). Em contraste, quando tentámos transplante utilizando a técnica de Kamada, eles não podia exceder 1,40 milímetros e 2,16 milímetros, respectivamente (ratos correspondentes para a estirpe e peso). Como mostrado na Figura 7, rápido-linker anéis são concebidos para manter os vasos receptoras sobre óptima alongamento, permitindo paquímetro mínimo e desperdício de comprimento, o que resulta em mais perto-a-fisiológicas resultados hemodinâmicos. <img src="/files/ftp_upload/4143/4143fig1.jpg" alt="A Figura 1" /> Figura 1. Medidas anéis. <img src="/files/ftp_upload/4143/4143fig2.jpg" alt="A Figura 2" /> Figura 2. Medidas Microclamps. <img src="/files/ftp_upload/4143/4143fig3.jpg" alt="A Figura 3" /> Figura 3. Quick-linker kit. <img src="/files/ftp_upload/4143/4143fig4.jpg" alt="A Figura 4" /> Figura 4. Quick-linker armado. <img src="/files/ftp_upload/4143/4143fig5.jpg" alt="A Figura 5" /> Figura 5. Quick-linker fechado. <img src="/files/ftp_upload/4143/4143fig6.jpg" alt="A Figura 6" /> Figura 6. Braçadeira SHVC. <img src="/files/ftp_upload/4143/4143fig7.jpg" alt="A Figura 7" /> Figura 7. Clássico contra quick-linker assistidas manguito anastomose-hemodinâmica. <img src="/files/ftp_upload/4143/4143fig8.jpg" alt="A Figura 8" /> Figura 8. Doadores e preparação destinatário pele e incisão.

<ol>
	<li>Starzl, T. E., Marchioro, T. L., Porter, K. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+and+clinical+observation+after+homotransplantation+of+the+whole+liver.">Experimental and clinical observation after homotransplantation of the whole liver.</a> Revue internationale d'hepatologie. 15, 1447 (1965).</li><li>Starzl, T. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evolution+of+liver+transplantation.">Evolution of liver transplantation.</a> Hepatology. 2, 614S-636S (1982).</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 (1979).</li><li>Kashfi, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+review+of+various+techniques+of+orthotopic+liver+transplantation+in+the+rat.">A review of various techniques of orthotopic liver transplantation in the rat.</a> Transplantation proceedings. 37, 185-188 (2005).</li><li>Hori, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comprehensive+and+innovative+techniques+for+liver+transplantation+in+rats:+A+surgical+guide.">Comprehensive and innovative techniques for liver transplantation in rats: A surgical guide.</a> World Journal of Gastroenterology: WJG. 16, 3120 (2010).</li><li>Oldani, G. <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+rat+liver+transplantation+using+Quick+Linker+system:+a+preliminary+result.">A novel technique for rat liver transplantation using Quick Linker system: a preliminary result.</a> Journal of Surgical Research. 149, 303-309 (2008).</li><li>Nosaka, T., Bowers, J. L., Cay, O., Clouse, M. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biliary+complications+after+orthotopic+liver+transplantation+in+rats.">Biliary complications after orthotopic liver transplantation in rats.</a> Surg. Today. 29, 963-965 (1999).</li><li>Li, G. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+incidence+of+biliary+complications+in+rat+liver+transplantation:+can+we+avoid+it.">High incidence of biliary complications in rat liver transplantation: can we avoid it.</a> World J. Gastroenterol. 17, 3140-3144 (2011).</li><li>Peng, Y., Gong, J. P., Yan, L. N., Li, S. B., Li, X. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improved+two-cuff+technique+for+orthotopic+liver+transplantation+in+rat.">Improved two-cuff technique for orthotopic liver transplantation in rat.</a> Hepatobiliary Pancreat. Dis. Int. 3, 33-37 (2004).</li><li>Urata, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Decreased+survival+in+rat+liver+transplantation+with+extended+cold+preservation:+role+of+portal+vein+clamping+time.">Decreased survival in rat liver transplantation with extended cold preservation: role of portal vein clamping time.</a> Hepatology. 28, 366-373 (1998).</li></ol>

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O presente modelo de transplante de fígado de rato pode ser facilmente estabelecida. Em nossa experiência, alguns pontos-chave ajudar a minimizar o risco de morte (≤ 20%). Mortes precoces, que ocorrem dentro das primeiras três horas de reperfusão, são mais frequentemente devido a um enfarte do intestino, como consequência de um portal longo de aperto. Portanto, é recomendável para manter a fase anepática dentro do limite ideal de 15 minutos (20 minutos ainda aceitável). Central embolia aérea também pode ser uma causa de morte súbita e é obrigatório para descarregar todas as veias antes da anastomose. Óbitos ocorridos entre o dia pós-operatório um e três são muitas vezes devido à insuficiência hepática ou trombose da veia cava. Deve-se, portanto, evitar qualquer enxerto agressivo de lavagem antes do explante no doador (que lavar com 20 ml durante aproximadamente 60 segundos) e garantir uma adequada preservação frio (imersão total em 0-4 ° C solução ringer lactato). O risco de trombosepode ser diminuída através da minimização da manipulação da íntima e preservar a sua integridade. Nós não recomendamos o uso de anticoagulação após a cirurgia. As mortes entre o dia pós-operatório de quatro e nove estão muitas vezes relacionados com colangite bacteriana 7. Com base na nossa experiência, uma dose única de piperacilline / tazobactam (0,03 g) administrado antes da laparotomia é suficiente para minimizar o risco de infecção biliar comum. A morte é um evento raro após o dia 10. No entanto, devido à ausência de fluxo arterial, alguns problemas de condutas biliares pode ser observado com cholastasis 8. Dependendo das necessidades do investigador uma arterialização pode ser considerado 9. Digno de nota, um tempo de isquemia fria até 24 horas pode ser considerado como 10 aceitável. O sistema quick-linker foi originalmente concebido para realizar transplantes de fígado com três anastomoses manguito (fase anepática muito curto). No entanto, o te três bainha-condutahaver descrição prévia requer o uso de pinças muito pequenas para a cava supra-hepática veia braçadeira (SHVC) e não tem sido largamente aceite, devido ao risco de problemas de fígado de saída 11-14. Nós, portanto, a favor de uma rápida linker técnica de três manguito assistida somente quando os tempos de isquemia quente com menos de dez minutos são necessários. Na maioria dos estudos no entanto, as fases de implantação até 15-18 minutos são aceitáveis, permitindo a utilização da técnica de dois bainha-conduta, e uma drenagem mais fisiológico SHVC. Como mostrado neste papel de vídeo, que rotineiramente usar o sistema rápido-ligante na segunda parte da implantação só. Embora algum tempo extra cirúrgico é necessário para posicionar as alças rápido-ligante, esta parte do processo está associado com riscos mínimos e permite a um encurtamento das graças anepática fase crítica para inserções mais fáceis golas. Outras vantagens do sistema rápido-ligante são um melhor alinhamento vaso enxerto-receptor e da utilização de punhos mais largos, com melhores resultados hemodinâmicos em comparação com otécnicas anteriormente descritas (incluindo a descrita por Kamada et ai.) 6 (Fig. 7).

<table border="1"><tbody><tr><td> Ferramenta / Reagente </td><td> Companhia </td><td> Catálogo n. </td><td> Comentar </td></tr><tr><td> Manguito Portal </td><td> <a href="http://altecweb.com" target="_blank">altecweb.com</a> ou <a href="http://smiths-medical.com" target="_blank">ferreiros-medical.com</a> </td><td> Bobina 01-96-1729 800/100/420 </td><td> Calibre: 1,55 milímetros Parede: 0,30 milímetros Comprimento: 3,5 mm </td></tr><tr><td> Infrahepatic v manguito cava </td><td> <a href="http://altecweb.com" target="_blank">altecweb.com</a> ou <a href="http://smiths-medical.com" target="_blank">ferreiros-medical.com</a> </td><td> Bobina 01-96-1733 800/100/540 </td><td> Calibre: 2,40 milímetros Parede: 0,30 milímetros <br/&gt; Comprimento: 3,5 mm </td></tr><tr><td> Stent biliar comum </td><td> comum 22G venflon </td><td></td><td></td></tr><tr><td> 7-0 laços de seda </td><td> <a href="http://georgetiemann.com" target="_blank">georgetiemann.com</a> </td><td> 160-1226-7 / 0 </td><td></td></tr><tr><td> 10-0 prolene </td><td> <a href="http://ethicon.com" target="_blank">ethicon.com</a> </td><td></td><td></td></tr><tr><td> 9-0 prolene </td><td> <a href="http://ethicon.com" target="_blank">ethicon.com</a> </td><td></td><td></td></tr><tr><td> 8-0 prolene </td><td> <a href="http://ethicon.com" target="_blank">ethicon.com</a> </td><td></td><td></td></tr><tr><td> Retas fórceps micro </td><td> <a href="http://s-and-t.net" target="_blank">-s e-t</a>. Net </td><td> FRC-15RM-8 </td><td></td></tr><tr><td> Curvas fórceps micro </td><td> <a href="http://s-and-t.net" target="_blank">-s e-t.net</a> </td><td> FRS-15 RM-8 </td><td></td></tr><tr><td> Dissecção porta-agulhas </td><td> <a href="http://s-and-t.net" target="_blank">-s e-t.net</a> </td><td> B-15-8,2 </td><td></td></tr><tr><td> Anastomose porta-agulhas </td><td> <a href="http://rumex.net" target="_blank">rumex.net</a> </td><td> 8-021T </td><td></td></tr><tr><td> Grampos Micro </td><td> <a href="http://finescience.com" target="_blank">finescience.com</a> </td><td> 18055-03 </td><td> 6 x 1 milímetro </td></tr><tr><td> Aplicador de grampos Micro </td><td> <a href="http://finescience.com" target="_blank">finescience.com</a> </td><td> 18057-14 </td><td></td></tr&gt; <tr><td> Supra-hepática v. cava grampo </td><td> <a href="http://pharmap.ch" target="_blank">pharmap.ch</a> </td><td> 13.349.14 </td><td></td></tr><tr><td> Quick-Linker anéis </td><td> <a href="http://dltchiropody.co.uk" target="_blank">dltchiropody.co.uk</a> </td><td> PAR 10 </td><td> modificados lâminas de bisturi </td></tr><tr><td> Quick-Linker aproximador </td><td> <a href="http://pharmap.ch" target="_blank">pharmap.ch</a> </td><td> UD-04-275 </td><td> fórceps modificado Kocher </td></tr></tbody></table>

orthotopic liver transplantation in rats

Progresso clínico no campo de transplante de fígado tem sido largamente suportado por 1,2 modelos animais. Desde a publicação do transplante hepático primeiro rato em 1979 por Kamada et al. 3, este modelo permanece o padrão ouro apesar de várias propostas de técnicas alternativas 4. No entanto, sua maior utilização é limitada pela sua curva de aprendizagem 5. Neste trabalho de vídeo, vamos mostrar uma revisão simples e fácil de estabelecer de técnica Kamada de dois manguito. A veia cava supra-hepática anastomose é realizada manualmente com uma sutura contínua, ea porta cava e anastomoses infrahepatic veia cava são realizados utilizando um sistema de manguito rápida linker 6. Fabricar o kit rápido-ligante é mostrado em um papel de vídeo separada.

Watch this Scientific Journal Video about Orthotopic Liver Transplantation in Rats at JoVE.com

Orthotopic Liver Transplantation in Rats

Nós apresentamos uma revisão fácil de estabelecer a técnica de dois manguito clássica para transplante hepático em ratos.

Transplante hepático ortotópico experimental em ratos

Clinical progress in the field of liver transplantation has been largely supported by animal models1,2. Since the publication of the first orthotopic rat ...

Research

JoVE Journal

Medicine

29.7K visualizações.  University of Geneva Hospitals. Nós apresentamos uma revisão fácil de estabelecer a técnica de dois manguito clássica para transplante hepático em ratos.

Vídeo: Orthotopic Liver Transplantation in Rats

Development of Obliterative Bronchiolitis in a Murine Model of Orthotopic Lung Transplantation

Orthotopic lung transplantation in rats was first reported by Asimacopoulos and colleagues in 1971 1. Currently, this method is well accepted and standardized not only for the study of allo-rejection but also between syngeneic strains for examining mechanisms of ischemia-reperfusion injury after lung transplantation. Although the application of the rat and other large animal model 2 contributed significantly to the elucidation of these studies, the scope of those investigations is limited by the scarcity of knockout and transgenic rats. Due to no effective therapies for obliterative bronchiolitis, the leading cause of death in lung transplant patients, there has been an intensive search for pre-clinical models that replicate obliterative bronchiolitis. The tracheal allograft model is the most widely used and may reproduce some of the histopathologic features of obliterative bronchiolitis 3. However, the lack of an intact vasculature with no connection to the recipient's conducting airways, and incomplete pathologic features of obliterative bronchiolitis limit the utility of this model 4. Unlike transplantation of other solid organs, vascularized mouse lung transplants have only recently been reported by Okazaki and colleagues for the first time in 2007 5. Applying the basic principles of the rat lung transplant, our lab initiated the obliterative bronchiolitis model using minor histoincompatible antigen murine orthotopic single-left lung transplants which allows the further study of obliterative bronchiolitis immunopathogenesis6.

Obliterative bronchiolitis is the key impediment to the long-term survival of lung transplant recipients and the lack of a robust preclinical model precludes examining obliterative bronchiolitis immunopathogenesis. Unlike other solid organ transplants, vascularized mouse lung transplantation has only recently been developed. Here we show our independently developed obliterative bronchiolitis model after murine orthotopic single-lung transplantation.

Desenvolvimento de bronquiolite obliterante em um modelo murino de transplante de pulmão ortotópico

Orthotopic lung transplantation in rats was first reported by Asimacopoulos and colleagues in 1971 1. Currently, this method is well accepted and ...

Medicina

Orthotopic Small Bowel Transplantation in Rats

Small bowel transplantation has become an accepted clinical option for patients with short gut syndrome and failure of parenteral nutrition (irreversible intestinal failure). In specialized centers improved operative and managing strategies have led to excellent short- and intermediate term patient and graft survival while providing high quality of life 1,3. Unlike in the more common transplantation of other solid organs (i.e. heart, liver) many underlying mechanisms of graft function and immunologic alterations induced by intestinal transplantation are not entirely known6,7. Episodes of acute rejection, sepsis and chronic graft failure are the main obstacles still contributing to less favorable long term outcome and hindering a more widespread employment of the procedure despite a growing number of patients on home parenteral nutrition who would potentially benefit from such a transplant. The small intestine contains a large number of passenger leucocytes commonly referred to as part of the gut associated lymphoid system (GALT) this being part of the reason for the high immunogenity of the intestinal graft. The presence and close proximity of many commensals and pathogens in the gut explains the severity of sepsis episodes once graft mucosal integrity is compromised (for example by rejection). To advance the field of intestinal- and multiorgan transplantation more data generated from reliable and feasible animal models is needed. The model provided herein combines both reliability and feasibility once established in a standardized manner and can provide valuable insight in the underlying complex molecular, cellular and functional mechanisms that are triggered by intestinal transplantation. We have successfully used and refined the described procedure over more than 5 years in our laboratory 8-11. The JoVE video-based format is especially useful to demonstrate the complex procedure and avoid initial pitfalls for groups planning to establish an orthotopic rodent model investigating intestinal transplantation.

Small bowel transplantation has become an accepted treatment option for patients with irreversible intestinal failure. Our experimental model of orthotopic small bowel transplantation in rats serves as a reliable tool to address underlying immunologic and inflammatory processes that complicate intestinal transplantation.

Transplante de intestino delgado de ratos ortotópico

Small  bowel transplantation has become an accepted clinical option for patients with  short gut syndrome and failure of parenteral nutrition ...

Orthotopic Aortic Transplantation in Mice for the Study of Vascular Disease

Vascular procedures involving anastomoses in the mouse are generally thought to be difficult and highly dependent on the skill of the individual surgeon. This is largely true, but there are a number of important principles that can reduce the difficulty of these procedures and enhance reproducibility. Orthotopic aortic transplantation is an excellent procedure in which to learn these principles because it involves only two end-to-end anastomoses, but requires good suturing technique and handling of the vessels for consistent success. This procedure begins with the procurement of a length of abdominal aorta from a donor animal, followed by division of the native aorta in the recipient. The procured aorta is then placed between the divided ends of the recipient aorta and sutured into place using end-to-end anastomoses. To accomplish this objective successfully requires a high degree of concentration, good tools, a steady hand, and an appreciation of how easily the vasculature of a mouse can be damaged, resulting in thrombosis. Learning these important principles is what occupies most of the beginner's time when learning microsurgery in small rodents. Throughout this protocol, we refer to these important points. This model can be used to study vascular disease in a variety of different experimental systems1-8. In the context shown here, it is most often used for the study of post-transplant vascular disease, a common long-term complication of solid organ transplantation in which intimal hyperplasia occurs within the allograft. The primary advantage of the model is that it facilitates quantitative morphometric analyses and the transplanted vessel lies contiguous to the endogenous vessel, which can serve as an additional control9. The technique shown here is most often used for mice weighing 18-25 grams. We have accumulated most of our experience using the C57BL/6J, BALB/cJ, and C3H/HeJ strains.

We describe a technique in which a section of the abdominal aorta from a mouse is transplanted orthotopically to just below the renal arteries in an allogeneic or syngeneic recipient. This technique can be useful in studies in which transplantation of large arteries of uniform size is deemed advantageous.

Transplante ortotópico de aorta em camundongos para o Estudo da Doença Vascular

Vascular procedures involving anastomoses in the mouse are generally thought to be difficult and highly dependent on the skill of the individual surgeon. ...

Surgical Procedures for a Rat Model of Partial Orthotopic Liver Transplantation with Hepatic Arterial Reconstruction

Orthotopic liver transplantation (OLT) in rats using a whole or partial graft is an indispensable experimental model for transplantation research, such as studies on graft preservation and ischemia-reperfusion injury 1,2, immunological responses 3,4, hemodynamics 5,6, and small-for-size syndrome 7. The rat OLT is among the most difficult animal models in experimental surgery and demands advanced microsurgical skills that take a long time to learn. Consequently, the use of this model has been limited. Since the reliability and reproducibility of results are key components of the experiments in which such complex animal models are used, it is essential for surgeons who are involved in rat OLT to be trained in well-standardized and sophisticated procedures for this model.
While various techniques and modifications of OLT in rats have been reported 8 since the first model was described by Lee et al. 9 in 1973, the elimination of the hepatic arterial reconstruction 10 and the introduction of the cuff anastomosis technique by Kamada et al. 11 were a major advancement in this model, because they simplified the reconstruction procedures to a great degree. In the model by Kamada et al., the hepatic rearterialization was also eliminated. Since rats could survive without hepatic arterial flow after liver transplantation, there was considerable controversy over the value of hepatic arterialization. However, the physiological superiority of the arterialized model has been increasingly acknowledged, especially in terms of preserving the bile duct system 8,12 and the liver integrity 8,13,14.
In this article, we present detailed surgical procedures for a rat model of OLT with hepatic arterial reconstruction using a 50% partial graft after ex vivo liver resection. The reconstruction procedures for each vessel and the bile duct are performed by the following methods: a 7-0 polypropylene continuous suture for the supra- and infrahepatic vena cava; a cuff technique for the portal vein; and a stent technique for the hepatic artery and the bile duct.

Orthotopic liver transplantation in rats is an indispensable experimental model for biomedical research. Here we present our surgical procedures for orthotopic rat liver transplantation with hepatic arterial reconstruction using a 50% partial graft.

Procedimentos cirúrgicos para um modelo de rato de Transplante de Fígado parcial ortotópico com reconstrução arterial hepática

Orthotopic liver transplantation (OLT) in rats using a whole or partial graft is an indispensable experimental model for transplantation research, such as ...

Technique of Porcine Liver Procurement and Orthotopic Transplantation using an Active Porto-Caval Shunt

The success of liver transplantation has resulted in a dramatic organ shortage. Each year, a considerable number of patients on the liver transplantation waiting list die without receiving an organ transplant or are delisted due to disease progression. Even after a successful transplantation, rejection and side effects of immunosuppression remain major concerns for graft survival and patient morbidity. 
Experimental animal research has been essential to the success of liver transplantation and still plays a pivotal role in the development of clinical transplantation practice. In particular, the porcine orthotopic liver transplantation model (OLTx) is optimal for clinically oriented research for its close resemblance to human size, anatomy, and physiology. 
Decompression of intestinal congestion during the anhepatic phase of porcine OLTx is important to guarantee reliable animal survival. The use of an active porto-caval-jugular shunt achieves excellent intestinal decompression. The system can be used for short-term as well as long-term survival experiments. The following protocol contains all technical information for a stable and reproducible liver transplantation model in pigs including post-operative animal care.

Experimental animal research plays a pivotal role in the development of clinical transplantation practice. The porcine orthotopic liver transplantation model (OLTx) closely resembles human conditions and is frequently used in clinically oriented research. The following protocol contains all information for a reliable porcine OLTx model using an active porto-caval-jugular shunt.

Técnica para as aquisições de fígado de porco e Transplante ortotópico usando um Active Porto-Caval Shunt

The success of liver transplantation has resulted in a dramatic organ shortage. Each year, a considerable number of patients on the liver transplantation ...

Orthotopic Hind Limb Transplantation in the Mouse

In vivo animal model systems, and in particular mouse models, have evolved into powerful and versatile scientific tools indispensable to basic and translational research in the field of transplantation medicine. A vast array of reagents is available exclusively in this setting, including mono- and polyclonal antibodies for both diagnostic and interventional applications. In addition, a vast number of genotyped, inbred, transgenic, and knock out strains allow detailed investigation of the individual contributions of humoral and cellular components to the complex interplay of an immune response and make the mouse the gold standard for immunological research. 
Vascularized Composite Allotransplantation (VCA) delineates a novel field of transplantation using allografts to replace "like with like" in patients suffering traumatic or congenital tissue loss. This surgical methodological protocol shows the use of a non-suture cuff technique for super-microvascular anastomosis in an orthotopic mouse hind limb transplantation model. The model specifically allows for comparison between established paradigms in solid organ transplantation with a novel form of transplants consisting of various different tissue components. Uniquely, this model allows for the transplantation of a viable vascularized bone marrow compartment and niche that have the potential to exert a beneficial effect on the balance of immune acceptance and rejection. This technique provides a tool to investigate alloantigen recognition and allograft rejection and acceptance, as well as enables the pursuit of functional nerve regeneration studies to further advance this novel field of transplantation.

This novel model for orthotopic hind limb transplantation in the mouse, applying a non-suture cuff technique for super-microvascular anastomosis, provides a powerful tool for in&#160;vivo mechanistic immunological research related to vascularized composite allotransplantation (VCA).

Ortotópico Hind Limb Transplantation no mouse

In vivo animal model systems, and in particular mouse models, have evolved into powerful and versatile scientific tools indispensable to basic and ...

A Rat Model of Orthotopic Liver Transplantation Using a Novel Magnetic Anastomosis Technique for Suprahepatic Vena Cava Reconstruction

The rat model of orthotopic liver transplantation (OLT) is essential for transplant research. It is a very sophisticated animal model and requires a steep learning curve. The introduction of the cuff technique for anastomosis of the portal vein (PV) and infrahepatic vena cava (IHVC) has significantly simplified the transplant procedure in rats. However, due to the short anterior wall of the recipients' suprahepatic vena cava (SHVC), the cuff technique is very difficult to use for the reconstruction of the SHVC. Most researchers in this field still use the hand-suture technique for SHVC reconstruction, which makes it the bottleneck step in rat orthotopic liver transplantation. The magnetic anastomosis technique (i.e., magnamosis) is a method of connecting two vessels using the attractive force between two magnets. Our recent study has shown that the magnetic anastomosis technique is superior to the hand-suture technique for SHVC reconstruction in rats. In this article, we show a step-by-step protocol for SHVC reconstruction in rats using the novel magnetic anastomosis technique. In this model, the reconstruction of the PV and IHVC was performed by the standard cuff technique, while the reconstruction of the bile duct (BD) was performed by a stent technique. The hepatic re-arterialization was not performed. The magnetic anastomosis technique made SHVC reconstruction much easier and significantly shortened the anphepatic phase. After a reasonable learning curve, even researchers without advanced microsurgical skills can produce reliable and reproducible results using this rat model of OLT.

The reconstruction of the suprahepatic vena cava (SHVC) remains a difficult step in rat orthotopic liver transplantation. In this article, we show a step-by-step protocol for SHVC reconstruction in rats using a novel magnetic anastomosis technique.

Um modelo do rato de ortotópico transplante hepático, usando uma técnica de anastomose magnético romance para reconstrução de veia Cava Suprahepatic

The rat model of orthotopic liver transplantation (OLT) is essential for transplant research. It is a very sophisticated animal model and requires a steep ...

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.

Um modelo pré-clínico porcino de transplante cardíaco ortotópico

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

Reduced Complications after Arterial Reconnection in a Rat Model of Orthotopic Liver Transplantation

The rat orthotopic liver transplantation (OLT) model is a powerful tool to study acute and chronic rejection. However, it is not a complete representation of human liver transplantation due to the absence of arterial reconnection. Described here is a modified transplantation procedure that includes the incorporation of hepatic artery (HA) reconnection, leading to a marked improvement in transplant outcomes. With a mean anhepatic time of 12 min and 14 s, HA reconnection results in improved perfusion of the transplanted liver and an increase in long-term recipient survival from 37.5% to 88.2%. This protocol includes the use of 3D-printed cuffs and holders to connect the portal vein and infrahepatic inferior vena cava. It can be implemented for studying multiple aspects of liver transplantation, from immune response and infection to technical aspects of the procedure. By incorporating a simple and practical method for arterial reconnection using a microvascular technique, this modified rat OLT protocol closely mimics aspects of human liver transplantation and will serve as a valuable and clinically relevant research model.

The goal of this study is to modify the rat orthotopic liver transplant model to better represent human liver transplantation and improve recipient survival. The presented method reestablishes hepatic arterial inflow by connecting the donor liver&#39;s common hepatic artery to the recipient liver&#39;s proper hepatic artery.

Complicações reduzidas após reconexão arterial em um modelo de rato de transplante de fígado ortotópico

The rat orthotopic liver transplantation (OLT) model is a powerful tool to study acute and chronic rejection. However, it is not a complete representation ...

Left Lung Orthotopic Transplantation in a Juvenile Porcine Model for ESLP

Lung transplantation is the gold-standard treatment for end-stage lung disease, with over 4,600 lung transplantations performed worldwide annually. However, lung transplantation is limited by a shortage of available donor organs. As such, there is high waitlist mortality. Ex situ lung perfusion (ESLP) has increased donor lung utilization rates in some centers by 15%-20%. ESLP has been applied as a method to assess and recondition marginal donor lungs and has demonstrated acceptable short- and long-term outcomes following transplantation of extended criteria donor (ECD) lungs. Large animal (in vivo) transplantation models are required to validate ongoing in vitro research findings. Anatomic and physiologic differences between humans and pigs pose significant technical and anesthetic challenges. An easily reproducible transplant model would permit the&#160;in vivo&#160;validation of current ESLP strategies and the preclinical evaluation of various interventions designed to improve donor lung function. This protocol describes a porcine model of orthotopic left lung allotransplantation. This includes anesthetic and surgical techniques, a customized surgical checklist, troubleshooting, modifications, and the benefits and limitations of the approach.

This protocol describes a juvenile porcine model of orthotopic left lung allotransplantation designed for use with ESLP research. Focus is made on anesthetic and surgical techniques, as well as critical steps and troubleshooting.

Transplante ortotópico de pulmão esquerdo em modelo suíno juvenil para PESL

Lung transplantation is the gold-standard treatment for end-stage lung disease, with over 4,600 lung transplantations performed worldwide annually. ...