The authors are solely responsible for the design and conduct of the presented study. Dr. Marbacher was supported by a grant from the Swiss National Science Foundation (PBSKP3-123454). The authors declare no conflict of interests. 
Author contributions to the study and manuscript preparation include the following. Conception and design: SM, JM, JF. Acquisition of data: SM, EA, JF. Analysis and interpretation of data: SM, JF, JM. Drafting the article: SM, JF, JM. Critically revising the article: JH, MN. Statistical analysis: SM, JF. Study supervision: JF, JH, MN.

NOTA: Masculino ratos Wistar (peso médio: 356 ± 44 g; 10-14 semanas de idade) foram alojados em sala de animal a 22-24 ° C e 12 horas ciclo claro / escuro com livre acesso a pellet dieta, a água da torneira normal e também recebeu atendimento humanizado, em conformidade com as diretrizes institucionais. Os experimentos foram analisados ​​e aprovados pelo Comitê de Bem-Estar Animal da Universidade de Helsinki, na Finlândia. NOTA: Para o seguinte demonstração nosso método cirúrgico é como se segue: Anestesiar ratos, por injecção subcutânea de adaptação ao peso de cloridrato de medetomidina (0,5 mg / kg) e a injecção intraperitoneal de cloridrato de cetamina (50 mg / kg). Teste para a falta de um reflexo de igual pitada de confirmar que o rato é completamente anestesiados. Aplicar pomada, prender o local cirúrgico, e limpar a pele com um desinfectante apropriado, por exemplo clorexidina, ou em álcool ou água. Lavar as mãos, vestir roupas de proteção, uma tampa da cabeça e da facemáscara e luvas cirúrgicas estéreis. Tenha uma ajuda de um auxiliar cirúrgico para manter condições de cirurgias assépticas e documentar as características cirúrgicas (conforme listado na Tabela 1). Monitore a profundidade da anestesia a cada 15 minutos durante a cirurgia, seguindo a freqüência respiratória, freqüência cardíaca, ea reação à estimulação nociceptiva (teste pitada tep). A injecção subcutânea de buprenorfina (0,03 mg / kg) foi dada para analgesia pós-operatória e repetidos, se necessário, a cada 12 h. 1. Hardware, Consumíveis e Posicionamento <ol><li> Mantenha o animal pequeno da sala de cirurgia tranquila, asséptica, e manter a temperatura ambiente em 23 +/- 3 ° C. Para realizar a cirurgia de aneurisma experimental o seguinte equipamento mínimo é necessário: <ol><li> Use uma tabela top microscópio cirúrgico idealmente equipado com um escopo assistente e câmera microscópio digital. Use não porosa superfície operacional reutilizável e lavável superfície instrumento para proteger a bancada do laboratório. </li><li&gt; Prepare consumíveis, tais como desinfetantes da pele, soro fisiológico, compressas de gaze menores e maiores e material de sutura, incluindo 9-0 micro-suturas, um 5-0 não absorvível e sutura absorvível 3-0. </li><li> Utilize os seguintes instrumentos cirúrgicos padrão: tesouras cirúrgicas, fórceps tecido, espalhador de tecidos moles ou autoportantes retrator e duas pinças cirúrgicas Mosquito. </li><li> Use um conjunto básico de microcirurgia instrumento que inclui: titular microagulhas Curvo, uma curva e dois micropinças retas, e uma microtesoura reta ou curva. </li><li> Manter os instrumentos de microcirurgia em um prato de rim preenchido com solução salina estéril para manter o instrumento não colante e limpo durante a cirurgia. O prato de rim é preenchido com tapete de borracha ou uma luva cirúrgica para evitar danos nas pontas dos micro-instrumentos. Certifique-se de que todas as fontes são estéreis e o procedimento é realizado com técnica asséptica de acordo com as recomendações atuais para cirurgia de sobrevivência em animais de laboratório <sup&gt; 8-10. </li><li> Além disso, utilizar um aplicador de clipe vascular e três grampos vasculares temporárias atraumáticas. É importante que os grampos utilizados têm um poder de fecho baixa para evitar lesão à parede muito fina da aorta de ratos. Também preparar um governante com barras de meio milímetro de escala, uma pequena almofada de borracha colorida, e uma agulha romba curta. </li><li> Coloque os ratos em decúbito dorsal, imobilizar frente e nas patas traseiras com esparadrapo sem aplicar estiramento ou compressão sobre a pele, e dobrar a sua volta com um marcador grosso ou caneta cautério, colocando-o sob a região de madeira serrada das costas. Isso é importante para se obter o máximo de lordose da coluna lombar como possível, a fim de melhorar a exposição retroperitoneal e o acesso à aorta infra-renal o que facilita a anastomose de microcirurgia. Este posicionamento é recomendado para a criação do aneurisma da aorta abdominal, mas não é necessário para a colheita do enxerto torácica. </li></ol></li></ol> 2. Graft colheita <ol><li> Uanestesia geral nder, abrir a cavidade torácica (início enxerto tempo de isquemia). Aplique uma pitada toe nocivo para confirmar que o rato não responde antes de prosseguir com os passos seguintes. Cortar através da parede abdominal médioventral, identificar o diafragma logo acima do fígado, e cortar o tecido conjuntivo na base do diafragma para permitir o acesso à caixa torácica. Com grandes tesouras, lado sem corte para baixo, cortam costelas apenas um centímetro para a esquerda e direita da linha média da caixa torácica e abrir a cavidade torácica. Os pulmões são mobilizados para o lado direito do coração. Sacrificar os ratos por overdose com injeção intracardíaca de cetamina hydrochloride.Pulmonary tronco, veia subclávia esquerda, veia cava cranial esquerda, ea veia ázigos interferem com a colheita da aorta torácica proximal. </li><li> Caudal para as veias proeminentes há um bom ponto de entrada para iniciar a dissecção da aorta torácica descendente usando micro-tesouras e micropinças. </li><li> Trace o torácica aorta back da parede dorsal do tórax, para cima, para o arco aórtico por retracção sem corte suave e dissecção com o grampo cirúrgico do mosquito. </li><li> Grampo e depois cortar as veias com uma tesoura. Manter os grampos nas veias e utilizar como um retractor para expor o arco aórtico subjacente. </li><li> Coloque um não-absorvível 6-0 seda ligadura apenas acima da primeira artéria intercostal deixando a aorta. </li><li> Corte a aorta descendente, imediatamente abaixo da artéria subclávia esquerda e, em seguida, abaixo da ligadura. A compensação pode ser feito de modo a obter uma geometria normalizada aneurisma perpendicular, ou, se necessário, para obter um determinado ângulo entre o eixo do aneurisma da aorta e. Medir o enxerto na sua largura e comprimento. NOTA: Colhida enxertos podem ser imediatamente transplantadas em ratos receptores ou tratados posteriormente para alcançar descelulação da parede do enxerto. Decelularizados podem ser armazenados a -4 graus Celsius até re-implantação em uma data posterior (Figura 1). The descelularização da parede do aneurisma foi mostrado para predispor o aneurisma para ampliar 11. </li></ol> 3. Graft Descelularização <ol style="margin-left: 40px;"><li> Congelar enxertos de dadores em solução salina tamponada com fosfato, a -4 ° C. </li><li> No dia seguinte, descongelar os enxertos, enxaguar com água destilada e desionizada, à temperatura ambiente, e incubar durante 10 horas a 37 ° C em 0,1% de dodecil sulfato de sódio. </li><li> Finalmente, lava as tratadas com enxertos de dodecil sulfato de sódio, três vezes, com agitação suave, recongelar em tampão fosfato salino, e manter a -4 ° C até à sua utilização. </li></ol> 4. Aneurysm Criação <ol style="margin-left: 40px;"><li> Dissecção da Aorta Abdominal <ol><li> Depois o animal foi anestesiado, o clipe de pele a partir do local da cirurgia e limpar a pele com um desinfectante apropriado. Teste para a falta de uma resposta de igual pitada antes da incisão na pele. Comece a incisão proximal um centímetro para os órgãos genitais in posição médioventral (hora de início de operação). Cuidadosamente separar a pele a partir dos músculos subjacentes. Fim dissecção 1-2 centímetros abaixo do esterno. </li><li> Com cuidado, mas com firmeza puxar para cima os músculos abdominais subjacentes para evitar danos aos órgãos subjacentes. Estender o corte longitudinal ao longo da linha alba para cima, para o processo xifóide e terminar em direção caudal ao nível da bexiga. </li><li> Delicadamente, aplique pressão na bexiga para esvaziá-lo, a fim de facilitar o acesso ao espaço retroperitoneal. </li><li> Mova o intestino delgado e do ceco de destaque para a direita ou a esquerda. Identificar o intestino grosso ou seja, do cólon descendente no canto inferior esquerdo da cavidade abdominal. </li><li> Cortar o ligamento entre o intestino delgado e o cólon descendente em direcção craniana para permitir uma maior exposição da parede do corpo dorsal. Coloque um afastador auto-retenção para segurar os intestinos à parte. </li><li> A localização ideal de end-to-side anastomose aneurisma is encontrados no nível entre o iliolombar renal e veias. A aorta abdominal reside retroperitoneal incorporado no tecido adiposo. Durante uma dissecção de prestar especial atenção aos pares quase transparentes ureteres e vasos testiculares. </li><li> Se for necessária uma retração dos intestinos, use swaps gaze maiores. A lordose da coluna lombar induzida durante o posicionamento colocando uma caneta cautério ou um objecto semelhante sob a parte inferior das costas do rato, reduz significativamente a necessidade de retracção do intestino. </li><li> A superfície ventral da parede dorsal do corpo é coberto com uma fina peritoneu parietal. Uma vez que este é aberto, visualize a aorta logo abaixo. Durante a dissecção romba afiado e cuidado da aorta abdominal de grandes veias adjacentes agarrar apenas a adventícia para evitar danos à parede do vaso. </li><li> Raramente, as pequenas artérias lombares surgem como vaso segmentar a partir da superfície dorsal da aorta abdominal e interferir com a preparação. Ligadura e corte do navio é needed para evitar escorrendo retrógrado durante a sutura aneurisma. O uso de micro-pinças curvas pode facilitar a colocação de ligadura na profundidade. </li></ol></li><li> End-to-side Anastomose <ol><li> Coloque uma almofada de borracha colorida debaixo da aorta abdominal e estofar o com uma pequena compressa de gaze. Remover o tecido conjuntivo frouxo e adventícia, a nível do local da anastomose planeada. </li><li> Prenda a aorta abdominal, a anastomose em primeiro lugar, em seguida, proximal (iniciar tempo de pinçamento aórtico). Isto assegura um enchimento firme do recipiente e facilita arteriotomia subsequente. </li><li> Realize a arteriotomia usando ou micro-tesouras retas ou curvas. Um micro-pinças mantém-se um pequeno pedaço da parede do vaso para cortar uma forma elíptica. </li><li> Lave a artéria completamente com soro fisiológico em ambos os sentidos utilizando uma agulha de ponta romba. </li><li> Colocar os dois primeiros suturas deste anastomose término-lateral na extremidade proximal e distal da arteriotomia (início da anastomose). Evitar agarrar da parede do vaso, com os micro-fórceps sempre que possível. Certifique-se de que cada fio de sutura é colocado através de todas as camadas da parede do vaso. </li><li> Realizar a sutura seja como suturas contínuas ou interrompidas. Se sutura interrompida é escolhido em seguida, coloque a parte de trás do lado de nove o`clock sutura primeiro. Suturas subsequentes podem ser espaçadas de partida adjacente ao primeiro fio de sutura. Segure a adventícia com cuidado. Evite apertar / apreensão da íntima. </li><li> Quando a parede de trás estiver concluída, verifique a parte endoluminal da anastomose para suturas extraviado. Executar os mesmos procedimentos descritos na mesma ordem no lado da frente. Certifique-se que o primeiro de um total de três nós por sutura é firme, mas não muito apertado. </li></ol></li><li> Hemostasia e fechamento <ol><li> Depois do fim-anastomose está completa (o tempo anastomose término), lavar o local com soro fisiológico e remover o grampo distal primeiro a permitir o refluxo (aórtica final de aperto tempo e enxerto final isquemia time). </li><li> Se o sangramento óbvio ocorre por refluxo um ponto adicional pode ser necessária (hora de início da hemostasia). Em caso de menor exsudação, hemostasia, com uma leve pressão no local da hemorragia utilizando um pequeno pedaço de um cotonete gaze. </li><li> Retire o grampo vascular proximal, lavar o local da anastomose, mais uma vez, e cortar as pontas restantes da ligadura na cúpula aneurisma. </li><li> Confirme a permeabilidade do aneurisma pela observação do aumento do volume do aneurisma durante a onda de pulso arterial de pico. Avaliar desobstrução da artéria abdominal distal através do &quot;teste de ordenha&quot; direta. </li><li> Retire a folha de plástico e um pequeno chumaço de gaze por baixo. O turbilhão de sangue pulsando dentro do aneurisma criado é claramente visível. </li><li> Cubra linhas de sutura em torno da anastomose com pequenos pedaços de tecido adiposo ou Spongostan para hemostasia adicional se pequeno escorrendo ainda está presente (tempo final da hemostasia). </li><li> Remover o disseminador e gaze cotonetes de tecidos moles.Coloque o intestino delgado, ceco, ea massa de gordura na sua posição correta. </li><li> Feche os músculos abdominais de linha média, utilizando 5-0, 4-0, 3-0 ou suturas absorvíveis ou não absorvíveis seguidos de pele fechamento da ferida utilizando a técnica de sutura contínua com 3-0 reabsorvível sutura polyfilament (tempo de operação final). Nota: Em nossos ratos experiência tolerado suturas reabsorvíveis melhores do que as suturas monofilamento não absorvível. </li></ol></li></ol>

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Os autores não têm qualquer interesse financeiro ou comercial em qualquer um dos medicamentos, materiais ou equipamentos utilizados.

O melhor conhecimento da biologia complexa de aneurisma da artéria cerebral sacular depende da análise de dados epidemiológicos e clínicos, complementado por estudos de laboratório em amostras de pacientes e trabalhos experimentais em modelos animais 3,12,13. Pequenos animais como o rato está intrinsecamente associado com menores custos de experiências e habitação, e menor necessidade de equipamentos especializados. Um tempo de operação total médio de menos de 60 minut...

A ruptura de um aneurisma da artéria cerebral sacular provoca hemorragia ameaçadora da vida levando a acidente vascular cerebral, danos neurológicos permanentes ou morte. A rotura pode ser evitada por qualquer recorte microcirúrgico ou oclusão endovascular de aneurisma. Um tratamento médico para evitar o crescimento de aneurisma e ruptura não foi ainda estabelecida. Modelos experimentais para os saculares são necessários para estudar a biologia de aneurismas arteriais e para o teste de equipamentos terapêuticos e estratégias. Para estes fins, de vários modelos diferentes em espécies diferentes foram desenvolvidos e publicado em 1. Os modelos de aneurisma maiores em suínos, cães e coelhos são preferencialmente utilizadas para testar inovações endovasculares em arquitetura aneurisma complexo 1,2. Modelos murinos aneurisma, por outro lado, permite testar questões de pesquisa de espécies geneticamente modificadas 3,4 e facilitar a clarificação de aneurisma biologia celular e molecular nonível muito melhor do que as espécies maiores 1. Embora endovascular trans-carótida e a implantação do dispositivo de trans-ilíaca é limitado a ratos maiores (&gt; 400-500 g) e stents menor do que 2,0 mm e 1,5 mm de diâmetro de 5,6, os stents podem ser colocados através de inserção directa para a aorta abdominal segmento abrigar os aneurismas experimentais. Trabalhos anteriores, utilizando o modelo abdominal microcirúrgica aneurisma da aorta de ratos lateral demonstrou sua viabilidade em dispositivos embólicos testes novela e seu uso no estudo da base molecular de aneurisma biologia 3,7. Muitos dos modelos aneurisma sacular experimentais publicados atualmente exigem equipamentos caros, ambientes especiais (por exemplo, salas de operação estéreis com capacidade de fluoroscopia), a competência radiologia intervencionista, ou o uso de espécies caros. Essas exigências limitam o uso em larga escala desses modelos, e levar ao uso de diferentes modelos, em laboratórios diferentes, que makes comparação de dados e meta-análise difícil, se não impossível. Um modelo experimental simples, robusto e de baixo custo é necessário, como uma ferramenta normalizada que pode ser usada de uma maneira normalizada em vários laboratórios de forma a obter resultados comparáveis ​​de diferentes instituições. Para isso, criamos a aorta parede lateral modelo aneurisma arterial saccular rato. O objetivo deste relatório é apresentar instruções de procedimento passo a passo, informações sobre equipamentos necessários, além de discutir as características anatômicas e cirúrgicos importantes para a criação de microcirurgia sucesso de aneurisma da parede lateral da aorta abdominal em ratos.

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			<td height="21" style="height:21px;width:231px;">Name of Material/ Equipment</td>
			<td style="width:236px;">Company</td>
			<td style="width:119px;">Catalog Number</td>
			<td style="width:167px;">Comments/Description</td>
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			<td height="21" style="height:21px;width:231px;">Medetomidine</td>
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			<td>Any genericon</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Ketamin</td>
			<td style="width:236px;"></td>
			<td style="width:119px;"></td>
			<td>Any genericon</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Buprenorphine</td>
			<td style="width:236px;"></td>
			<td style="width:119px;"></td>
			<td>Any genericon</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Phosphate buffered saline</td>
			<td style="width:236px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Sodium dodecyl sulfate (0.1%)</td>
			<td style="width:236px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">3-0 resorbable suture</td>
			<td style="width:236px;">Ethicon Inc., USA</td>
			<td><a name="RANGE!C7">VCP824G</a></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">5-0 non absorbable suture</td>
			<td style="width:236px;">Ethicon Inc., USA</td>
			<td>8618G</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">6-0 non absorbable silk suture</td>
			<td style="width:236px;">B. Braun, Germany</td>
			<td style="width:119px;">C0761060</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">9-0 nylon micro suture</td>
			<td style="width:236px;">B. Braun, Germany</td>
			<td style="width:119px;">G1118471</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Spongostan</td>
			<td style="width:236px;">Ethicon Inc., USA</td>
			<td style="width:119px;">MS0002</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Operation microscope</td>
			<td style="width:236px;">Leica , Germany</td>
			<td style="width:119px;">M651</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Digital microscope camera</td>
			<td style="width:236px;">Sony, Japan</td>
			<td style="width:119px;">SSC-DC58AP</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Standard surgical instruments</td>
			<td style="width:236px;">B. Braun, Germany</td>
			<td style="width:119px;">Multiple</td>
			<td>See protocol 1.4</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Microsurgical instruments</td>
			<td style="width:236px;">B. Braun, Germany</td>
			<td style="width:119px;">Multiple</td>
			<td>See protocol 1.5</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Vascular clip applicator</td>
			<td style="width:236px;">B. Braun, Germany</td>
			<td style="width:119px;">FT495T</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Temporary vascular clamps</td>
			<td style="width:236px;">B. Braun, Germany</td>
			<td style="width:119px;">FT250T</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;">Graph Pad Prism statistical software&#160;</td>
			<td style="width:236px;">GraphPad Software, San Diego, California, USA</td>
			<td style="width:119px;"></td>
			<td>V 6.02 for Windows</td>
		</tr>
	</tbody>
</table>

the helsinki rat microsurgical sidewall aneurysm model

Modelos de aneurisma sacular experimentais são necessários para testar novas opções e dispositivos de tratamento cirúrgico e endovascular, antes de serem introduzidos na prática clínica. Além disso, os modelos experimentais são necessários para elucidar a biologia aneurisma complexo levando à ruptura de aneurismas saculares. Vários tipos diferentes de modelos experimentais para os saculares foram estabelecidas em diferentes espécies. Muitos deles, no entanto, exigem habilidades especiais, equipamentos caros, ou ambientes especiais, o que limita a sua utilização generalizada. Um modelo experimental simples, robusto e de baixo custo é necessário, como uma ferramenta normalizada que pode ser usada de uma maneira normalizada em diversas instituições. O modelo de aneurisma da aorta abdominal lateral rato microcirúrgica combina a possibilidade de estudar duas novas estratégias de tratamento endovascular ea base molecular da biologia aneurisma em um padronizado e baratomaneira. Enxertos normalizado por meio de forma, tamanho e geometria são colhidos a partir da aorta torácica descendente de um rato doador e depois transplantadas para um rato destinatário singénico. Os aneurismas são suturados de ponta a lado com contínuo ou interrompido 9-0 suturas de nylon para a aorta abdominal infra-renal. Apresentamos instruções processuais passo-a-passo, informações sobre o equipamento necessário, e discutir detalhes anatômicos e cirúrgicos importantes para a criação de microcirurgia sucedido de um aneurisma da aorta abdominal lateral no rato.

Uma série piloto era composto por 14 ratos. Posteriormente, um total de 84 animais foram operados de acordo com o protocolo apresentado em vários projectos de investigação, entre Março e Setembro de 2012. adicionais 29 animais serviram como doadores de enxertos arteriais saculares. Os demais experimentos foram realizados utilizando enxertos pré-tratados colhidas e armazenadas a partir de experiências anteriores com ratos do mesmo sexo, tensão, peso e idade. O peso corporal, o tempo t...

Watch this Scientific Journal Video about The Helsinki Rat Microsurgical Sidewall Aneurysm Model at JoVE.com

The Helsinki Rat Microsurgical Sidewall Aneurysm Model

Microsurgical sidewall aneurysms in rats are created by end-to-side anastomosis of an aortic graft to the abdominal aorta. We present step-by-step instructions and discuss anatomical and surgical details for successful experimental saccular aneurysm creation.

O Helsinki Rat microcirúrgica Sidewall Aneurysm Modelo

Experimental saccular aneurysm models are necessary for testing novel surgical and endovascular treatment options and devices before they are introduced ...

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Medicine

12.1K visualizações.  University of Helsinki, Helsinki, Finland. Microsurgical sidewall aneurysms in rats are created by end-to-side anastomosis of an aortic graft to the abdominal aorta. We present step-by-step instructions and discuss anatomical and surgical details for successful experimental saccular aneurysm creation.

Vídeo: The Helsinki Rat Microsurgical Sidewall Aneurysm Model

Microsurgical Venous Pouch Arterial-Bifurcation Aneurysms in the Rabbit Model: Technical Aspects

For ruptured human cerebral aneurysms endovascular embolization has become an equivalent alternative to aneurysm clipping.1 However, large clinical trials have shown disappointing long-term results with unacceptable high rates of aneurysm recanalization and delayed aneurysm rupture.2 To overcome these problems, animal experimental studies are crucial for the development of better endovascular devices.3-5
Several animal models in rats, rabbits, canines and swine are available.6-8 Comparisons of the different animal models showed the superiority of the rabbit model with regard to hemodynamics and comparability of the coagulation system and cost-effectiveness.9-11 
The venous pouch arterial bifurcation model in rabbits is formed by a venous pouch sutured into an artificially created true bifurcation of both common carotid arteries (CCA). The main advantage of this model are true bifurcational hemodynamics.12 The major drawbacks are the sofar high microsurgical technical demands and high morbidity and mortality rates of up to 50%.13 These limitations have resulted in less frequent use of this aneurysm model in the recent years. These shortcomings could be overcome with improved surgical procedures and modified peri- and postoperative analgetic management and anticoagulation.14-16 Our techniques reported in this paper demonstrate this optimized technique for microsurgical creation of arterial bifurcation aneurysms.

An optimized technique for the microsurgical creation of arterial bifurcation aneurysms mimicking bifurcation human cerebral aneurysms is described. A venous pouch is sutured into an artificially created true bifurcation of both common carotid arteries. Facilitated microsurgical techniques and aggressive postoperative anticoagulation and analgesia lead to minimized morbidity rates and high aneurysm patency rates.

Microcirúrgica venoso Bolsa Bifurcação Arterial-aneurismas no Modelo de Coelho: Aspectos Técnicos

For ruptured human cerebral aneurysms endovascular embolization has become an equivalent alternative to aneurysm clipping.1 However, large clinical trials ...

Medicina

A New Murine Model of Endovascular Aortic Aneurysm Repair

Endovascular aneurysm exclusion is a validated technique to prevent aneurysm rupture. Long-term results highlight technique limitations and new aspects of Abdominal aortic aneurysm (AAA) pathophysiology. There is no abdominal aortic aneurysm endograft exclusion model cheap and reproducible, which would allow deep investigations of AAA before and after treatment. We hereby describe how to induce, and then to exclude with a covered coronary stentgraft an abdominal aortic aneurysm in a rat. The well known elastase induced AAA model was first reported in 19901 in a rat, then described in mice2. Elastin degradation leads to dilation of the aorta with inflammatory infiltration of the abdominal wall and intra luminal thrombus, matching with human AAA. Endovascular exclusion with small covered stentgraft is then performed, excluding any interactions between circulating blood and the aneurysm thrombus. Appropriate exclusion and stentgraft patency is confirmed before euthanasia by an angiography thought the left carotid artery. Partial control of elastase diffusion makes aneurysm shape different for each animal. It is difficult to create an aneurysm, which will allow an appropriate length of aorta below the aneurysm for an easy stentgraft introduction, and with adequate proximal and distal neck to prevent endoleaks. Lots of failure can result to stentgraft introduction which sometimes lead to aorta tear with pain and troubles to stitch it, and endothelial damage with post op aorta thrombosis. Giving aspirin to rats before stentgraft implantation decreases failure rate without major hemorrhage. Clamping time activates neutrophils, endothelium and platelets, and may interfere with biological analysis.

Histological and biochemical modifications after aneurysm endograft exclusion are still unclear. We describe a new model of endograft implantation on a murine aneurysm. Through thrombus analysis with and without persistant circulating blood stream interface, and new endovascular biomaterials evaluation, this model will help to better understand endovascular aneurysm exclusion pathobiology.

Um novo modelo murino de reparação endovascular de aneurisma da aorta

Endovascular aneurysm exclusion is a validated technique to prevent aneurysm rupture. Long-term results highlight technique limitations and new aspects of ...

Demonstration of the Rat Ischemic Skin Wound Model

The propensity for chronic wounds in humans increases with ageing, disease conditions such as diabetes and impaired cardiovascular function, and unrelieved pressure due to immobility. Animal models have been developed that attempt to mimic these conditions for the purpose of furthering our understanding of the complexity of chronic wounds. The model described herein is a rat ischemic skin flap model that permits a prolonged reduction of blood flow resulting in wounds that become ischemic and resemble a chronic wound phenotype (reduced vascularization, increased inflammation and delayed wound closure). It consists of a bipedicled dorsal flap with 2 ischemic wounds placed centrally and 2 non-ischemic wounds lateral to the flap as controls. A novel addition to this ischemic skin flap model is the placement of a silicone sheet beneath the flap that functions as a barrier and a splint to prevent revascularization and reduce contraction as the wounds heal. Despite the debate of using rats for wound healing studies due to their quite distinct anatomic and physiologic differences compared to humans (i.e., the presence of a panniculus carnosus muscle, short life-span, increased number of hair follicles, and their ability to heal infected wounds) the modifications employed in this model make it a valuable alternative to previously developed ischemic skin flap models.

The rat, due to its size, availability, and rather docile behavior, has been utilized as a research model for many years. The goal of this protocol is to utilize the rat as an ischemic skin wound healing model to provide valuable insight into the pathophysiology of chronic wounds.

Demonstração da Wound modelo de pele de rato Isquêmico

The propensity for chronic wounds in humans increases with ageing, disease conditions such as diabetes and impaired cardiovascular function, and ...

The Dimethylnitrosamine Induced Liver Fibrosis Model in the Rat

Four to six week old, male Wistar rats were used to produce animal models of liver fibrosis. The process requires four weeks of administration of 10 mg/kg dimethylnitrosamine (DMN), given intraperitoneally for three consecutive days per week. Intraperitoneal injections were performed in the fume hood as DMN is a known hepatoxin and carcinogen. The model has several advantages. Firstly, liver changes can be studied sequentially or at particular stages of interest. Secondly, the stage of liver disease can be monitored by measurement of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) enzymes. Thirdly, the severity of liver damage at different stages can be confirmed by sacrifice of animals at designated time points, followed by histological examination of Masson&#39;s Trichome stained liver tissues. After four weeks of DMN dosing, the typical fibrosis score is 5 to 6 on the Ishak scale. The model can be reproduced consistently and has been widely used to assess the efficacy of potential anti-fibrotic agents.

We describe a method to produce an animal model of liver fibrosis in the rat, and assess the degree of fibrosis by histological examination of the liver. The model can be used to study the development of liver disease as well as to test the efficacy of potential anti-fibrotic agents.

A fibrose do fígado Modelo dimetilnitrosamina induzida no Rato

Four to six week old, male Wistar rats were used to produce animal models of liver fibrosis. The process requires four weeks of administration of 10 mg/kg ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

Medição de contratilidade isométrica da artéria mesentérica Mouse usando fio miografia

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

Corneal Epithelial Abrasion with Ocular Burr As a Model for Cornea Wound Healing

The murine cornea provides an excellent model to study wound healing. The cornea is the outermost layer of the eye, and thus is the first defense to injury. In fact, the most common type of eye injury found in clinic is a corneal abrasion. Here, we utilize an ocular burr to induce an abrasion resulting in removal of the corneal epithelium in vivo on anesthetized mice. This method allows for targeted and reproducible epithelial disruption, leaving other areas intact. In addition, we describe the visualization of the abraded epithelium with fluorescein staining and provide concrete advice on how to visualize the abraded cornea. Then, we follow the timeline of wound healing 0, 18, and 72 h after abrasion, until the wound is re-epithelialized. The epithelial abrasion model of corneal injury is ideal for studies on epithelial cell proliferation, migration and re-epithelialization of the corneal layers. However, this method is not optimal to study stromal activation during wound healing, because the ocular burr does not penetrate to the stromal cell layers. This method is also suitable for clinical applications, for example, pre-clinical test of drug effectiveness.

This protocol describes a method to inflict an abrasion to the ocular surface of the mouse, and to follow the wound healing process thereafter. The protocol takes advantage of an ocular burr to partially remove the surface epithelium of the eye in anaesthetized mice.

Cicatrização de feridas abrasão epitelial da córnea com Burr Ocular como um modelo para córnea

The murine cornea provides an excellent model to study wound healing. The cornea is the outermost layer of the eye, and thus is the first defense to ...

An Ex Vivo Tissue Culture Model for Fibrovascular Complications in Proliferative Diabetic Retinopathy

Diabetic retinopathy (DR) is the most common microvascular complication of diabetes and one of the leading causes of blindness in working-age adults. No current animal models of diabetes and oxygen-induced retinopathy develop the full-range progressive changes manifested in human proliferative diabetic retinopathy (PDR). Therefore, understanding of the disease pathogenesis and pathophysiology has relied largely on the use of histological sections and vitreous samples in approaches that only provide steady-state information on the involved pathogenic factors. Increasing evidence indicates that dynamic cell-cell and cell-extracellular matrix (ECM) interactions in the context of three-dimensional (3D) microenvironments are essential for the mechanistic and functional studies towards the development of new treatment strategies. Therefore, we hypothesized that the pathological fibrovascular tissue surgically excised from eyes with PDR could be utilized to reliably unravel the cellular and molecular mechanisms of this devastating disease and to test the potential for novel clinical interventions. Towards this end, we developed a novel method for 3D ex vivo culture of surgically-excised patient-derived fibrovascular tissue (FT), which will serve as a relevant model of human PDR pathophysiology. The FTs are dissected into explants and embedded in fibrin matrix for ex vivo culture and 3D characterization. Whole-mount immunofluorescence of the native FTs and end-point cultures allows thorough investigation of tissue composition and multicellular processes, highlighting the importance of 3D tissue-level characterization for uncovering relevant features of PDR pathophysiology. This model will allow the simultaneous assessment of molecular mechanisms, cellular/tissue processes and treatment responses in the complex context of dynamic biochemical and physical interactions within the PDR tissue architecture and microenvironment. Since this model recapitulates PDR pathophysiology, it will also be amenable for testing or developing new treatments.

Here, we present a protocol to study the pathophysiology of proliferative diabetic retinopathy by using patient-derived, surgically-excised, fibrovascular tissues for three-dimensional native tissue characterization and ex vivo culture. This ex vivo culture model is also amenable for testing or developing new treatments.

Um modelo de cultura de tecidos de Ex Vivo para fibrovasculares complicações retinopatia diabética proliferativa

Diabetic retinopathy (DR) is the most common microvascular complication of diabetes and one of the leading causes of blindness in working-age adults. No ...

Murine Surgical Model of Topical Elastase Induced Descending Thoracic Aortic Aneurysm

According to the Center for Disease Control, aortic aneurysms (AAs) were considered a leading cause of death in all races and both sexes from 1999-2016. An aneurysm forms as a result of progressive weakening and eventual dilation of the aorta, which can rupture or tear once it reaches a critical diameter. Aneurysms of the descending aorta in the chest, called descending thoracic aortic aneurysms (dTAA), make up a large proportion of aneurysm cases in the United States. Uncontained dTAA rupture is almost universally lethal, and elective repair has a high rate of morbidity and mortality. The purpose of our model is to study dTAA specifically, to elucidate the pathophysiology of dTAA and to search for molecular targets to halt the growth or reduce the size of dTAA. By having a murine model to study thoracic pathology precisely, targeted therapies can be developed to specifically test dTAA. The method is based on the placement of porcine pancreatic elastase (PPE) directly on the outer murine aortic wall after surgical exposure. This creates a destructive and inflammatory reaction, which weakens the aortic wall and allows for aneurysm formation over weeks to months. Though murine models possess limitations, our dTAA model produces robust aneurysms of predictable size. Furthermore, this model can be used to test genetic and pharmaceutical targets which may arrest dTAA growth or prevent rupture. In human patients, interventions such as these could help avoid aneurysm rupture, and difficult surgical intervention.

We describe a surgical protocol to consistently induce robust descending thoracic aortic aneurysms in mice. The procedure involves left thoracotomy, thoracic aorta exposure, and placement of a sponge soaked in porcine pancreatic elastase on the aortic wall.

Modelo cirúrgico murino de elastase tópica induzida por aneurisma de aorta torácica descendente

According to the Center for Disease Control, aortic aneurysms (AAs) were considered a leading cause of death in all races and both sexes from 1999-2016. ...

Porcine Model of Infrarenal Abdominal Aortic Aneurysm

Large animal models to study abdominal aortic aneurysms are sparse. The purpose of this model is to create reproducible, clinically significant infrarenal abdominal aortic aneurysms (AAA) in swine. To achieve this, we use a combination of balloon angioplasty, elastase and collagenase, and a lysyl oxidase inhibitor, called &#946;-aminopropionitrile (BAPN), to create clinically significant infrarenal aortic aneurysms, analogous to human disease.
Noncastrated male swine are fed BAPN for 7 days prior to surgery to achieve a steady state in the blood. A midline laparotomy is performed and the infrarenal aorta is circumferentially dissected. An initial measurement is recorded prior to aneurysm induction with a combination of balloon angioplasty, elastase (500 units)/collagenase (8000 units) perfusion, and topical elastase application. Swine are fed BAPN daily until terminal procedure on either postoperative day 7, 14, or 28, at which time the aneurysm is measured, and tissue procured. BAPN + surgery pigs are compared to pigs that underwent surgery alone.
Swine treated with BAPN and surgery had a mean aortic dilation of 89.9% &#177; 47.4% at day 7, 105.4% &#177; 58.1% at day 14, and 113.5% &#177; 30.2% at day 28. Pigs treated with surgery alone had significantly smaller aneurysms compared to BAPN + surgery animals at day 28 (p &#60; 0.0003). The BAPN + surgery group had macroscopic and immunohistochemical evidence of end stage aneurysmal disease.
Clinically significant infrarenal AAA can be induced using balloon angioplasty, elastase/collagenase perfusion and topical application, supplemented with oral BAPN. This model creates large, clinically significant AAA with hallmarks of human disease. This has important implications for the elucidation of AAA pathogenesis and testing of novel therapies and devices for the treatment of AAA. Limitations of the model include variation in BAPN ingested by swine, quality of elastase perfusion, and cost of BAPN.&#160;

This novel model creates robust infrarenal abdominal aortic aneurysms in swine using a combination of balloon angioplasty, elastase/collagenase perfusion, topical elastase application, and oral compound &#946;-aminopropionitrile administration, which interferes with collagen cross-linking.

Modelo suíno de aneurisma aórtica abdominal infra-renal

Large animal models to study abdominal aortic aneurysms are sparse. The purpose of this model is to create reproducible, clinically significant infrarenal ...

Vessel-Sparing Microsurgical Longitudinal Intussusception Vasoepididymostomy to Treat Epididymal Obstructive Azoospermia

The epididymis is a common site of obstruction in obstructive azoospermia (OA). Vasoepididymostomy has become an important method for the treatment of epididymal OA since 2000. There are two challenges in classic microscopic vasoepididymostomy. First, anastomosis of the vas deferens and epididymis is performed with double-needle sutures. However, there is a lack of good-quality and cost-effective double-needle sutures in China, which leads to increased difficulty and poor success rates of anastomosis. Second, the separation of the vas deferens does not retain vasculature, although the vas deferens vasculature plays an important role in the blood supply to the vas deferens, epididymis, and testis. This affects the blood supply to the anastomotic area and epididymis.
Therefore, this team has made innovative improvements to address these problems. Good-quality, cost-effective, single-needle sutures, which are easy to purchase in China and other countries, were used in microsurgical longitudinal intussusception vasoepididymostomy. This can optimize the operation procedure and shorten the operation time while ensuring the success rate of the anastomosis. The surgical method of preserving the vas deferens vessels was innovatively proposed because the etiology of epididymal OA is mostly inflammatory in China. The protection of the blood supply to the vas deferens and epididymis is maximized using microsurgical forceps to separate and protect the vasculature. Patency reached 81.7% in the postoperative follow-up, indicating a better surgical treatment effect.

The present protocol describes a vessel-sparing, longitudinal intussusception vasoepididymostomy using readily available single-needle sutures in China, a safe and effective procedure, which may improve patients' patency and natural pregnancy rates.

Vasoepididymostomia Microsúrgica E Microsúrgica De Vasoepididymostomia para Tratar Azoospermia Obstrutiva Epididímal

The epididymis is a common site of obstruction in obstructive azoospermia (OA). Vasoepididymostomy has become an important method for the treatment of ...