Este trabalho foi financiado, em parte, por uma concessão do NIH (RO1 HL098228) para CKB.

Nota: Todos os procedimentos com animais foram aprovados pelo Comitê Institucional Hospital Animal Care e Use do Nationwide Children. 1. Pulmonar Válvula Cardíaca Colheita de um doador de válvula do coração mouse <ol><li> Autoclave todos os instrumentos cirúrgicos antes da cirurgia: 1x tesoura fina, micro fórceps 3x, pinças vasculares micro 2x, braçadeira 1x fórceps de aplicação, suporte da agulha micro 1x, tesoura primavera 1x, 1x afastador. </li><li> Um 6-8 semanas de idade fêmea C57BL / 6 de rato é usado como um doador de válvula cardíaca pulmonar. Remover o rato da sua gaiola e pesá-lo, em seguida, a eutanásia com um cocktail de cetamina / xilazina (cetamina 200 mg / kg e xilazina, 20 mg / kg, IP) sobredosagem. </li><li> Clipe da área do peito e coloque o mouse em uma posição de decúbito dorsal sobre uma almofada. Em seguida, fazer a toracotomia. Expor o coração, fazer um pequeno corte no átrio direito, e aspergir o ventrículo esquerdo com gelo salina fria. </li><li> Sem rodeios dissecção da artéria pulmonar (AP) a partir da ascensão de umorta. Cortar a válvula pulmonar (PV), juntamente com dois milímetros punho da artéria pulmonar. Elimine o resto do coração. </li><li> Armazenar o PV em heparina frio e solução salina (100 unidades / ml). Nota: O PV pode ser mantido na solução durante duas horas antes do transplante para o coração de dador. </li></ol> 2. Preparação coração doador <ol><li> Um 6-8 semanas de idade fêmea C57BL / 6 de rato é usado como um doador de coração. Remover o rato da sua gaiola e pesá-lo, em seguida, a eutanásia com um cocktail de cetamina / xilazina (cetamina 200 mg / kg e xilazina, 20 mg / kg, IP) sobredosagem. Este é um procedimento terminal. </li><li> Clipe da área do peito e coloque o mouse em uma posição de decúbito dorsal sobre uma almofada. Em seguida, fazer a toracotomia. Sem rodeios separar o coração, veia cava inferior (VCI), veia cava superior (VCS), aorta ascendente, PA, e veia pulmonar. Perfundir da VCI com gelo frio salina estéril. </li><li> Ligadura da VCI, SVC, e veia pulmonar com seda 6-0 sutura em seguida, corte superior aoligaduras. </li><li> Corte a aorta e PA com 2 milímetros punho. </li><li> Recorte o PV e descartá-la. </li></ol> 3. Válvula de Transplante de Coração em um coração do doador <ol><li> Imediatamente após o passo 2.5, coloque a válvula cardíaca a partir do passo 1.5 para o coração do doador e orientar a válvula cardíaca. </li><li> Fixe o PV com um ponto no lado direito da válvula usando 10-0 suturas monofilamento em agulhas cônicos e começar a sutura contínua com 5-6 stiches do outro lado do PV. </li><li> Depois de terminar o lado da frente, girar o coração na horizontal e começa a suturar o lado de trás do PV para o coração de dador. </li><li> Armazenar o coração em solução de heparina / solução salina estéril fria. Nota: O coração do dador podem ser mantidas na solução durante duas horas antes da implantação até o rato destinatário. </li></ol> 4. Heterotópico Transplante de Coração para um destinatário Rato <ol><li> Um 6-8 semanas de idade fêmea C57BL / 6 de ratinho foi utilizado como um recipient. Remover o rato da sua gaiola e pesá-lo, depois anestesiados com um cocktail de cetamina / xilazina (cetamina 100 mg / kg de xilazina e 10 mg / kg). O cetoprofeno (5 mg / kg) é utilizado como pré-anestésica analgésico. </li><li> Depois de verificar o nível de sedação pela cauda beliscar, prenda o cabelo abdominal e torácica. Lubrifique os olhos com pomada oftálmica estéril, e coloque o mouse em uma posição de decúbito dorsal sobre uma almofada. Desinfetar o abdômen com Betadine e álcool pads. Em seguida, cubra o mouse com um campo estéril e expor apenas a área da incisão. </li><li> Faça uma laparotomia mediana incisão abaixo do xifóide até a região suprapúbica, e inserir um afastador auto de retenção. Enrole os intestinos em salina umedecido gaze. Sem rodeios definir a aorta infra-renal e veia cava. </li><li> Coloque duas suturas de seda 6-0 proximal e distal ao redor da aorta e veia cava inferior para conter a circulação sanguínea. </li><li> Coloque o coração do doador, no lado direito da aorta abdominal e cobri-lo com gau estérilze. Hidrate-lo com soro fisiológico. </li><li> Realizar uma Aortotomia na aorta abdominal com uma agulha G 30, e estender a abertura com uma tesoura para o tamanho da aorta do doador. </li><li> Realizar um fim-de lado anastomose usando estéreis suturas 10-0 monofilamento as agulhas cônicos. Fixar a aorta do dador com um ponto na extremidade proximal da abertura na aorta abdominal e iniciar a sutura contínua com 4-5 stiches a partir da extremidade distal da aorta abdominal. </li><li> Virar o coração para o lado esquerdo, cobri-lo com uma solução salina infundida gaze, e iniciar a sutura contínua com 4-5 stiches a partir da extremidade distal da aorta abdominal. </li><li> Fazer uma venotomia na VCI usando uma agulha G 30 e estender a abertura para o tamanho da artéria pulmonar do dador. </li><li> Realizar um fim a anastomose lado usando estéreis suturas 10-0 monofilamento as agulhas cônicos. Fixar o doador PA com um ponto na extremidade proximal da abertura na VCI e iniciar a sutura contínua com 4-5 de stichesa extremidade distal da veia cava inferior. Desta vez, por causa da aorta é na forma, certifique-se de sutura da parede esquerda da PA do dador está no interior da veia cava inferior. </li><li> Lave o lúmen da VCI com heparina e solução salina (100 unidades / ml). Fechar a parede direita do doador e do receptor PA IVC suturando-os continuamente para a extremidade distal. </li><li> Remover a ligadura distal e controlar a hemorragia através da aplicação de um agente hemostático estéril absorvível tópica. Quando a hemorragia parar completamente, retirar a sutura proximal e controlar a hemorragia da mesma maneira. </li><li> Retorne os intestinos e fechar a musculatura abdominal e pele em duas camadas usando uma sutura de monofilamento 6-0 poliamida preto. </li><li> Injetar 0,5 ml por via subcutânea de solução salina e coloque o rato em uma gaiola de recuperação em um bloco de aquecimento até que o mouse é totalmente móvel. Após a recuperação, o retorno do mouse para uma nova gaiola com roupa de cama de papel. Dar medicação para dor (Ibuprofeno, 30 mg / kg, a água potável) para 48 horas. Fazernão devolver um animal que passou por uma cirurgia para a companhia de outros animais até que esteja totalmente recuperado. </li></ol>

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

Não temos nada a divulgar.

A taxa de mortalidade do presente processo é de cerca de 20%, que foi principalmente causada por hemorragia no local da transplante de PV e anastomose sobre a aorta do doador para a aorta abdominal do destinatário. Na maioria dos casos, a taxa de mortalidade diminui significativamente 48 horas pós cirurgia. Os ratos de sobrevivência mostraram fortes batimentos cardíacos e do fluxo de sangue através do PV implantado. Todo o processo leva quatro horas para um cirurgião experiente micro. Levará cerca de 250 ratos p...

Defeitos cardiovasculares congênitos são uma das principais causas de mortalidade infantil no mundo 1,2 ocidental. Entre eles, a estenose da válvula pulmonar e os defeitos da válvula aórtica bicúspide são uma forma que ocorre com freqüência 3. Cirurgia de substituição da válvula cardíaca é uma escolha rotina de cirurgias reconstrutivas; no entanto, complicações, incluindo estenose e calcificação da válvula cardíaca, ea dependência ao longo da vida em anticoagulantes são uma fonte significativa de problemas de saúde crônica e morte 4-7. Além disso, a falta de potencial crescimento requer cirurgias de revisão, que aumenta ainda mais a mortalidade dos pacientes jovens 4,8,9. Numa tentativa para desenvolver uma válvula cardíaca de substituição funcional com um potencial de crescimento, Shinoka et al. Células autólogas semeadas em uma válvula cardíaca sintética biodegradável 8. A válvula sintético transformado em uma válvula cardíaca nativa como a estrutura com potenciômetro crescimentoai. Preliminares grandes estudos com animais demonstraram a viabilidade da utilização desta metodologia para criar uma válvula cardíaca funcional 10. No entanto, estudos de implantação de longo prazo demonstraram pouca durabilidade devido ao espessamento progressivo da válvula neotissue resultando em estreitamento da válvula cardíaca. Trabalhe a partir de Sodian et al. Utilizada a metodologia Shinoka, mas finalmente substituiu a matriz PGA com um elastômero biodegradável, que deu as propriedades biomecânicas da válvula de engenharia de tecidos construir um perfil mais fisiológico 9,11,12. No estudo in vivo, apesar do sucesso da implantação, um revestimento de células endoteliais confluentes não foi formado, o que poderia limitar o sucesso a longo prazo deste andaime 12. A fim de projetar racionalmente uma válvula cardíaca sintético de segunda geração melhorada, um modelo murino de transplante de válvula cardíaca foi criado para investigar os mecanismos celulares e moleculares underlyinformação g neotissue, espessamento da válvula, e o desenvolvimento de estenose. Modelos murinos oferecem uma vasta gama de reagentes moleculares, incluindo transgénicos, que não estão facilmente disponíveis em outras espécies 7. Neste modelo de transplante de coração válvula, um vivo substituição da válvula cardíaca pulmonar ex singênico foi realizada pela primeira vez; e, em seguida, o coração, com a válvula de coração implantado foi implantado heterotopicamente no hospedeiro singeneico usando uma técnica de microcirurgia. Este modelo permite a substituição da válvula do coração sem a necessidade de circulação extracorpórea. Neste trabalho, uma explicação detalhada de uma colheita de válvula cardíaca, preparações de coração doador, o transplante de válvula cardíaca e transplante cardíaco heterotópico é descrito. Os resultados mostraram uma pulsação contínua a partir do coração do doador, que era independente do batimento cardíaco destinatário. O fluxo de sangue através da válvula pulmonar implantado foi medida usando um sistema de ultra-som de alta freqüência com uma onda pulsada Fazerppler.

<table border="0" cellpadding="0" cellspacing="0" width="644">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">DPBS</td>
			<td style="width:167px;">gibco</td>
			<td style="width:133px;">14190-144</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:176px;">Microscope</td>
			<td style="width:167px;">Leica</td>
			<td style="width:133px;">M80</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:176px;">C57BL/6J (H-2b), Female</td>
			<td style="width:167px;">Jackson Laboratories&#160;</td>
			<td style="width:133px;">664</td>
			<td style="width:168px;">8-12 weeks</td>
		</tr>
		<tr height="39">
			<td height="39" style="height:39px;width:176px;">Ketamine Hydrochloride Injection</td>
			<td style="width:167px;">Hospira Inc.</td>
			<td style="width:133px;">NDC 0409-2053</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:176px;">Xylazine Sterile Solution</td>
			<td style="width:167px;">Akorn Inc.</td>
			<td style="width:133px;">NADA# 139-236</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="36">
			<td height="36" style="height:36px;width:176px;">ketoprofen</td>
			<td style="width:167px;">Fort Dodge Animal Health</td>
			<td style="width:133px;">NDC 0856-4396-01</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="36">
			<td height="36" style="height:36px;width:176px;">Ibuprofen</td>
			<td style="width:167px;">PrecisionDose</td>
			<td style="width:133px;">NDC 68094-494-59</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="37">
			<td height="37" style="height:37px;width:176px;">Heparin Sodium</td>
			<td style="width:167px;">Sagent Pharmaceticals</td>
			<td style="width:133px;">NDC 25021-400</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="39">
			<td height="39" style="height:39px;width:176px;">Saline solution (Sterile 0.9% Sodium Chloride)</td>
			<td style="width:167px;">Hospira Inc.</td>
			<td style="width:133px;">NDC 0409-0138-22</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="39">
			<td height="39" style="height:39px;width:176px;">0.9% Sodium Chloride Injection</td>
			<td style="width:167px;">Hospira Inc.</td>
			<td style="width:133px;">NDC 0409-4888-10</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="41">
			<td height="41" style="height:41px;width:176px;">Petrolatum Ophthalmic Ointment</td>
			<td style="width:167px;">Dechra Veterinary Products</td>
			<td style="width:133px;">NDC 17033-211-38</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:176px;">Iodine Prep Pads</td>
			<td style="width:167px;">Triad Disposables, Inc.</td>
			<td style="width:133px;">NDC 50730-3201-1</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:176px;">Alcohol Prep Pads</td>
			<td style="width:167px;">McKesson Corp.</td>
			<td style="width:133px;">NDC 68599-5805-1</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:176px;">Cotton tipped applicators</td>
			<td style="width:167px;">Fisher Sientific</td>
			<td style="width:133px;">23-400-118</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:176px;">Fine Scissor</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">14028-10</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:176px;">Micro-Adson Forcep</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">11018-12</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:176px;">Clamp Applying Forcep</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">00072-14</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:176px;">S&#38;T Vascular Clamp</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">00396-01</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:176px;">Spring Scissors</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">15008-08</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Colibri Retractors</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">17000-04</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Dumont #5 Forcep</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">11251-20&#160;</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Dumont #7 - Fine Forceps</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">11274-20</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Dumont #5/45 Forceps</td>
			<td style="width:167px;">FST</td>
			<td style="width:133px;">11251-35</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Tish Needle Holder/Forceps</td>
			<td style="width:167px;">Micrins</td>
			<td style="width:133px;">MI1540</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="38">
			<td height="38" style="height:38px;width:176px;">Black Polyamide Monofilament Suture, 10-0</td>
			<td style="width:167px;">AROSurgical Instruments Corporation</td>
			<td style="width:133px;">TI638402</td>
			<td style="width:168px;">For sutureing the graft</td>
		</tr>
		<tr height="37">
			<td height="37" style="height:37px;width:176px;">Black Polyamide Monofilament Suture, 6-0</td>
			<td style="width:167px;">AROSurgical Instruments&#160;</td>
			<td style="width:133px;">SN-1956</td>
			<td style="width:168px;">For musculature and skin closure</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Non-Woven Songes</td>
			<td style="width:167px;">McKesson Corp.</td>
			<td style="width:133px;">94442000</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Absorbable hemostat</td>
			<td style="width:167px;">Ethicon</td>
			<td style="width:133px;">1961</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:176px;">1 ml Syringe</td>
			<td style="width:167px;">BD</td>
			<td style="width:133px;">309659</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:176px;">3 ml Syringe</td>
			<td style="width:167px;">BD</td>
			<td style="width:133px;">309657</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;width:176px;">10 ml Syringe</td>
			<td style="width:167px;">BD</td>
			<td style="width:133px;">309604</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">18G 1 1/2 in, Needle</td>
			<td style="width:167px;">BD</td>
			<td style="width:133px;">305190</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">25G 1 in., Needle</td>
			<td style="width:167px;">BD</td>
			<td style="width:133px;">305125</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">30G 1 in., Needle</td>
			<td style="width:167px;">BD</td>
			<td style="width:133px;">305106</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:176px;">Warm Water Recircultor</td>
			<td style="width:167px;">Gaymar</td>
			<td style="width:133px;">TP-700</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:176px;">Warming Pad</td>
			<td style="width:167px;">Gaymar</td>
			<td style="width:133px;">TP-22G</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:176px;">Trimmer</td>
			<td style="width:167px;">Wahl</td>
			<td style="width:133px;">9854-500</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="53">
			<td height="53" style="height:53px;width:176px;">VEVO2100 HIGH-FREQUENCY ULTRASOUND</td>
			<td>VisualSonics</td>
			<td style="width:133px;"><a href="http://www.visualsonics.com/vevo2100">http://www.visualsonics.com/vevo2100</a></td>
			<td style="width:168px;">The catalog number and pricing can be acquired from the sales representatives.</td>
		</tr>
		<tr height="38">
			<td height="38" style="height:38px;width:176px;">Ultrasound transmission gel</td>
			<td style="width:167px;">PARKER LABORATORIES, 
			&#160;INC.</td>
			<td style="width:133px;">01-02</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="38">
			<td height="38" style="height:38px;width:176px;">Table Top Laboratory Animal Anesthesia System</td>
			<td style="width:167px;">VetEquip, INC.</td>
			<td style="width:133px;">901806&#160;</td>
			<td style="width:168px;"></td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:176px;">Isoflurane</td>
			<td style="width:167px;">Baxter</td>
			<td style="width:133px;">1001936060</td>
			<td style="width:168px;"></td>
		</tr>
	</tbody>
</table>

transplantation of pulmonary valve using a mouse model of heterotopic heart transplantation

Engenharia de tecidos de válvulas cardíacas, válvulas especialmente descelularizados, estão começando a ganhar impulso no uso clínico de cirurgia reconstrutiva com resultados mistos. No entanto, os mecanismos celulares e moleculares do desenvolvimento neotissue, espessamento da válvula, e o desenvolvimento de estenose não são pesquisadas extensivamente. Para responder às perguntas acima, foi desenvolvido um modelo de transplante heterotópico murino coração válvula. A válvula cardíaca foi colhida a partir de um camundongo doador válvula e transplantadas para um doador de rato coração. O coração com uma nova válvula foi transplantado heterotopicamente a um rato destinatário. O coração transplantado mostrou o seu próprio batimento cardíaco, independente de batimentos cardíacos do destinatário. O fluxo de sangue foi quantificado usando um sistema de ultra-som de alta freqüência com Doppler pulsátil. O fluxo através da válvula pulmonar implantado mostrou a frente de fluxo com refluxo mínimo e pico de fluxo foi próximo a 100 mm / seg. Este modelo murino de transplante de válvula cardíaca é highly versátil, de modo que pode ser modificada e adaptada para fornecer diferentes ambientes hemodinâmicas e / ou pode ser usado com vários ratinhos transgénicos para estudar o desenvolvimento neotissue em uma válvula cardíaca de engenharia de tecidos.

A figura 1 ilustra o esquema do modelo de transplante de válvula cardíaca utilizando transplante cardíaco heterotópico. A válvula cardíaca foi colhida a partir de um doador e implantado em um coração de um segundo rato doador. Em seguida, o coração com a nova válvula cardíaca foi implantado no abdómen de um rato receptor. Figura 2 mostra uma ilustração do coração implantado no espaço abdominal (A), logo após o transplante de coração (B),</st...

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

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

De modo a compreender os mecanismos celulares e moleculares subjacentes neotissue formação e desenvolvimento de estenose em válvulas cardíacas de engenharia de tecidos, foi desenvolvido um modelo murino de transplante heterotópico de válvula cardíaca. A válvula cardíaca pulmonar foi transplantado para destinatário usando o coração técnica de transplante heterotópico.

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 ...

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

Research

JoVE Journal

Medicine

18.8K visualizações.  Nationwide Children's Hospital.  De modo a compreender os mecanismos celulares e moleculares subjacentes neotissue formação e desenvolvimento de estenose em válvulas cardíacas de engenharia de tecidos, foi desenvolvido um modelo murino de transplante heterotópico de válvula cardíaca. A válvula cardíaca pulmonar foi transplantado para destinatário usando o coração técnica de transplante heterotópico. 

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

A Mouse Model of in Utero Transplantation

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

A Mouse Model of in Utero Transplantation

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

Um modelo do rato de No Utero Transplante

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

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 ...

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 ...

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 ...

Heterotopic Cervical Heart Transplantation in Mice

The heterotopic cervical heart transplantation in mice is a valuable tool in transplant and cardiovascular research. The cuff technique greatly simplifies this model by avoiding challenging suture anastomoses of small vessels thereby reducing warm ischemia time. In comparison to abdominal graft implantation the cervical model is less invasive and the implanted graft is easily accessible for further follow-up examinations. Anastomoses are performed by pulling the ascending aorta of the graft over the cuff with the recipient&#8217;s common carotid artery and by pulling the main pulmonary artery over the cuff with the external jugular vein. Selection of appropriate cuff size and complete mobilization of the vessels are important for successful revascularization. Ischemia-reperfusion (I/R) injury can be minimized by perfusing the graft with a cardioplegic solution and by hypothermia. In this article, we provide technical details for a simplified and improved cuff technique, which should allow surgeons with basic microsurgical skills to perform the procedure with a high success rate.

The cuff technique shortens and facilitates the model of heterotopic cervical heart transplantation in mice by avoiding technically challenging suture anastomoses of small vessels. The technical details shown in this video paper should allow researches to establish this model in their laboratories.

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 ...

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 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 ...

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 ...