Nos gustaría dar las gracias a la Sra. Angela Freund por su inestimable asistencia técnica con la incrustación y la producción de diapositivas. También nos gustaría agradecer al Sr. Tadeusz Stopinski en el Instituto de Ciencia de Animales de Laboratorio y Cirugía Experimental por su perspicaz ayuda con el trabajo veterinario.

Los experimentos se realizaron de conformidad con la ley alemana de bienestar animal (TSchG) y la Directiva 2010/63/UE relativa a la protección de los animales utilizados con fines científicos. La aprobación oficial para este estudio fue otorgada por el Comité Gubernamental de Cuidado y Uso de Animales (Protocolo No.: AZ 87-51.04.2010.A065; Landesamt für Natur, Umwelt und Verbraucherschutz Nordrhein-Westfalen, Recklinghausen, Alemania). El protocolo de estudio cumplió con la Guía para el Cuidado y Uso de Animales de Laboratorio. El tratamiento del dolor postoperatorio se basa en las recomendaciones de la Sociedad Alemana para la Ciencia de Animales de Laboratorio (GV-SOLAS), así como en la Iniciativa de Terapia Veterinaria del Dolor.
1. Técnicas básicas y procedimientos comunes
<ol><li>Use apoE homocigótico-/- Ratas Sprague-Dawley. Identificar el genotipo de cada animal mediante métodos estándar17.</li>
	<li>Mantener a los animales en condiciones idénticas (21 °C ± 2 °C, 60% ± 5% de humedad y un ciclo de luz/oscuridad de 12 h) y garantizar el libre acceso al agua y a los alimentos.</li>
	<li>Lleve a cabo todos los procedimientos en condiciones limpias pero no esteriles.</li>
	<li>Una vez que la rata es anestesiada, realice todos los procedimientos bajo un microscopio quirúrgico a un aumento de 16x.</li>
	<li>Use hisopos de algodón para la hemostasia de compresión. Los hisopos de gasa (5 cm x 5 cm) empapados con solución de ringer lactato son útiles para mantener la ingle húmeda.</li>
	<li>Siga las regulaciones de eliminación de residuos para eliminar los materiales usados.</li>
</ol>2. Preparaciones antes de la cirugía
<ol><li>Prepare los medicamentos veterinarios antes de comenzar la operación. Mantenga todas las soluciones a temperatura ambiente, a menos que se indique lo contrario.</li>
	<li>
Treinta minutos antes del procedimiento, administre 0.03-0.05 mg/kg de buprenorfina por vía subcutánea.
</li>
	<li>Anestesiar a la rata con una inyección intraperitoneal de 100 mg/kg de peso corporal (BW) (S)-ketamina y 8 mg/kg de BW xilazina.</li>
	<li>Evalúe el peso de la rata usando una báscula de pesaje.</li>
	<li>Coloque la rata en una almohadilla térmica y fije las extremidades superiores e inferiores con cinta adhesiva médica. Coloque a la rata con su miembro trasero izquierdo completamente extendido y lo más en línea con su columna vertebral como sea posible para crear una línea recta entre la arteria femoral y la aorta. Esto facilitará el avance del stent montado en un balón a través de la bifurcación aórtica.</li>
	<li>Mantener la anestesia con inhalación de isoflurano al 1,5% vol. en oxígeno al 97,5% a un caudal de 2 L/min. NOTA: Permita que la rata respire espontáneamente, sin intubación.</li>
	<li>Aplique ungüento para los ojos para prevenir el daño ocular durante la inconsciencia.</li>
	<li>Afeitar el pelaje de la ingle y el área inferior del abdomen de la rata y esterilizar la piel correspondiente con una solución de povidona-yodo.</li>
	<li>Antes de comenzar la cirugía, verifique la profundidad adecuada de la anestesia pellizcando la punta de la cola y el tejido interdigital.</li>
</ol>3. Cirugía
<ol><li>Haga una incisión medial de ~ 0.5 \ u20121 cm en la ingle izquierda para abrir la piel y la fascia subyacente.</li>
	<li>Diseccione y sondee embotado embotado en las profundidades hasta que la arteria femoral izquierda que pulsa pueda ser identificada.</li>
	<li>Usando fórceps muy finos, prepare la arteria femoral eliminando suavemente el tejido conectivo circundante. Tenga cuidado de no dañar ni el nervio femoral ni la vena femoral, que es medial a la arteria.</li>
	<li>Preparar aproximadamente 1 cm de la arteria femoral. Coloque cuidadosamente la punta de las fórceps debajo del recipiente para levantarlo suavemente.</li>
	<li>Hilo piezas de 4-0 sutura de seda bajo las partes distales y proximales de la arteria y forman eslingas. Sujete los extremos de cada una de las dos eslingas de hilo entre las ramas de una abrazadera quirúrgica. Use las abrazaderas quirúrgicas para controlar la arteria. Estire y levante suavemente las eslingas para interrumpir temporalmente el flujo sanguíneo. NOTA: Trabaje rápido para evitar un torniquete prolongado que pueda llevar al daño tisular.</li>
	<li>Usando micro tijeras afiladas, realice una arteriotomía en el medio de la arteria femoral.</li>
	<li>Introduzca un cable guía a través de la arteriotomía. Al llegar a la eslinga del hilo proximal, suelte la tensión del hilo moviendo la abrazadera quirúrgica y avance el alambre guía más hacia la aorta abdominal. NOTA: Corte el alambre guía con un cortador de alambre para facilitar la manipulación.</li>
	<li>Coloque el extremo proximal del alambre guía entre el diafragma y las arterias renales. NOTA: Avanzar demasiado el alambre guía conlleva el riesgo de lesión aórtica o cardíaca. Recomendamos abrir el abdomen para asegurar la colocación adecuada del alambre guía y del stent al menos para los primeros animales.</li>
	<li>Introduzca un stent coronario engarzado y montado en un balón que mida 2,25 mm x 8 mm (máx. 2,5 mm x 8 mm) sobre el alambre guía en la arteria femoral y avance hacia la aorta abdominal.</li>
	<li>Coloque el stent justo encima de la bifurcación aórtica pero debajo de las arterias renales. Despliegue el stent inflando el catéter del globo a 12 atm por 15 s usando un sistema de la jeringuilla del inflación.</li>
	<li>Desinflar el catéter con balón y mantener la presión negativa de acuerdo con las recomendaciones del fabricante para el stent en uso.</li>
	<li>Retire lentamente el catéter desinflado mientras deja el stent en su lugar.</li>
	<li>Justo antes de sacar el catéter, cree tensión en el lazo del hilo por encima de la incisión con la abrazadera quirúrgica para interrumpir el flujo sanguíneo nuevamente. Luego retire el catéter con balón y ligar directamente el vaso proximally.</li>
	<li>Ate los lazos próximos y distales del hilo de rosca para ligar la arteria femoral y para confirmar hemostasis adecuado de la arteriotomía. Las arterias colaterales asegurarán la perfusión adicional al miembro.</li>
	<li>Cierre el músculo que borda la arteria, así como la incisión de la piel mediante el uso de 10-0 suturas no reabsorbibles.</li>
</ol>4. Cuidado de animales después de la implantación de stent
<ol><li>Inmediatamente después de la operación, permita que la rata se recupere durante 60 minutos en una jaula de unidad de cuidados intensivos especial con aire calentado (30 \ u201235 ° C) y un suministro de oxígeno.</li>
	<li>Observe a los animales cuidadosamente hasta que se recuperen por completo. Después, mueva las ratas a una jaula normal. Proporcionar acceso ad libitum al agua y los alimentos.</li>
	<li>Administrar analgesia postoperatoria cada 6-12 horas con 0,03-0,05 mg/kg de buprenorfina (s.c., en 500μl de NaCl) durante un total de 72 horas bajo evaluación clínica.</li>
	<li>Mezcle el alimento con clopidogrel (15 mg/kg) para evitar la trombosis del stent implantado.</li>
	<li>Para mejorar las condiciones hipercolesterolémicas y la formación de placa, comience la alimentación con dieta occidental a las 6 semanas después del nacimiento y continúe hasta la eutanasia. Si se desea, una cohorte de animales alimentados con ratas normales puede servir como control.</li>
</ol>5. Recolección y procesamiento de tejidos
<ol><li>Antes de comenzar la explantación de tejido en el punto de tiempo designado, eutanasiar al animal de acuerdo con las directrices de IACUC. Cosechar la aorta con stent para el análisis histológico al final del período de observación.</li>
	<li>Abra el abdomen mediante una incisión en la línea media y retire el segmento con stent de la aorta, así como las partes adyacentes de la aorta sin stent, que miden 0,5 cm cada una.</li>
	<li>Coloque el tejido en una solución de formol tamponado al 4% durante 24 h para la fijación.</li>
	<li>Incrustar el tejido arterial con stent en plástico y realizar tinción histológica e inmunohistoquímica de acuerdo con los protocolos estándar18,19.</li>
</ol>6. Análisis histomorfométrico
<ol><li>Realizar análisis histomorfométricos de secciones secuenciales de la parte proximal, media y distal de la aorta con stent por medio de un microscopio conectado a una computadora con un software de análisis de imágenes apropiado.</li>
	<li>Traza los contornos de la lámina elástica externa (EEL, entre adventitia y media), la lámina elástica interna (IEL, entre media y neointima), y la luz con una tableta de dibujo gráfico. A partir de estos valores, calcule el área EEL, el área IEL y el área de lumen con el software.</li>
	<li>Calcule el porcentaje de reestenosis en stent (ISR) del área transversal: <img alt="Equation 1" src="/files/ftp_upload/61442/61442equ01.jpg">
</li>
	<li>Calcular el área neointimal total (Ai): <img alt="Equation 2" src="/files/ftp_upload/61442/61442equ02.jpg">
</li>
	<li>Mida el grosor neointimal (NIT) sobre cada puntal de stent como la distancia entre el puntal y la luz. Mida el NIT entre los puntales del stent como la distancia entre el IEL y la luz. Nota: Alternativamente, calcular NIT como <img alt="Equation 3" src="/files/ftp_upload/61442/61442equ03.jpg"> donde PL y PIEL son el lumen y el perímetro interno de la lámina elástica, respectivamente20.</li>
	<li>Realizar análisis adicionales de acuerdo con los requerimientos del estudio.</li>
</ol>

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Este protocolo describe la implantación de stents coronarios de tamaño humano en la aorta abdominal de las ratas del apoE-/-. Vale la pena destacar varios aspectos técnicos. En primer lugar, se debe evitar un desajuste entre el tamaño del stent y el tamaño de la aorta. La colocación de un stent demasiado pequeño puede llevar a la malaposición del stent, mientras que la implantación de un stent que es demasiado grande para la aorta puede causar estiramiento excesivo, desgarro y lesión del vaso. Por lo...

La intervención coronaria percutánea (ICP), combinada con el despliegue de un stent coronario, representa el patrón oro en el tratamiento intervencionista de la enfermedad arterial coronaria1. El éxito a largo plazo de los stents, sin embargo, puede verse limitado por la aparición de reestenosis en stent (ISR) que está determinada por una proliferación excesiva de tejido neointimal dentro del stent2,3. La ISR puede requerir una reintervención con revascularización coronaria o re-ICP. Se han sugerido una variedad de modelos animales para el estudio de la ISR, cada uno de ellos con ventajas y deficiencias. Los principales inconvenientes de los modelos de arterias ilíacas coronarias y conejos más comúnmente utilizados, aunque desarrollando lesiones marcadamente similares a las de los humanos después de la implantación del stent4,5,son los grandes costos de animales y alojamiento que traen consigo dificultades logísticas, especialmente en estudios a largo plazo, así como limitaciones en el manejo y el equipo. Además, la disponibilidad de anticuerpos contra las proteínas celulares de cerdos y conejos es limitada. Por otro lado, los modelos murinos proporcionan las principales ventajas de un alto rendimiento y reproducibilidad, así como facilidad de manejo, alojamiento y, por lo tanto, rentabilidad. Además, un mayor número de anticuerpos están disponibles. Sin embargo, mientras que los ratones con deficiencia de Apolipoproteína E (apoE-/-)han sido ampliamente utilizados para el estudio de la aterosclerosis6,7,8,no son adecuados para el estudio de la ISR, ya que los stents tienen que ser miniaturizados para ser implantados en ratones, lo que potencialmente cambia las propiedades mecánicas de los stents. Por otra parte, la pared aórtica de los ratones mide entre 50 μm en ratones jóvenes y 85 μm en ratones viejos9,y los stents tienen que ser desplegados utilizando niveles de presión tan bajos como 2 atm, lo que podría conducir a la malaposición del stent10. Las ratas, sin embargo, permiten la implantación de stents coronarios humanos disponibles en el mercado, y demuestran un curso de curación vascular similar a los animales más grandes después de la implantación del stent aórtico, reportado por primera vez por Langeveld et al.11. Esta técnica requirió originalmente un acceso transporte-abdominal, que hizo necesario una constricción física de la aorta para alcanzar una interrupción temporal del flujo de sangre. Para evitar la lesión potencialmente asociada a los vasos y las reacciones inflamatorias, la técnica se perfeccionó posteriormente mediante la introducción de un acceso trans ilíaco, lo que además resultó en una mayor tasa de supervivencia de los animales12.
Debido a que las ratas de tipo salvaje no desarrollan lesiones ateroscleróticas13,las ratas apoE-/- se han generado ratas utilizando técnicas de nucleasa como la Ncleasa Efectora Activadora de Transcripción (TALEN)14,las repeticiones palindrómicas cortas agrupadas regularmente interespaciadas (CRISPR/Cas9)15,y zinc finger (ZF)16. ApoE-/- las ratas han estado disponibles comercialmente desde 2011. Proporcionando un fondo atherogenic, las ratas del apoE-/- permiten una evaluación más realista de stents coronarios de tamaño humano, especialmente con respecto a ISR.
Adjunto, describimos el método vía la ruta de acceso transfemoral y usando un stent droga-que eluting del fino-puntal del fino-cromo disponible en el comercio (DES), sin embargo, puede también ser aplicado para el estudio de otros tipos stents, tales como stents del metal desnudo (BMS) o stents biodegradables.

<table>
	<tbody>
		<tr>
			<td>Diet</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>SNIFF High Fat diet + Clopidogrel (15 mg/kg)</td>
			<td>SNIFF Spezialdi&#228;ten GmbH, Soest</td>
			<td>custom prepared</td>
			<td>Western Diet</td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Drugs and Anesthetics</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Buprenorphine</td>
			<td>Essex Pharma</td>
			<td>997.00.00</td>
			<td></td>
		</tr>
		<tr>
			<td>ISOFLO (Isoflurane Vapor) vaporiser</td>
			<td>Eickemeyer</td>
			<td>4802885</td>
			<td></td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Forene Abbott</td>
			<td>B 506</td>
			<td></td>
		</tr>
		<tr>
			<td>Isotonic (0.9%) NaCl solution</td>
			<td>DeltaSelect GmbH</td>
			<td>PZN 00765145</td>
			<td></td>
		</tr>
		<tr>
			<td>Ringer&#39;s lactate solution</td>
			<td>Baxter Deutschland GmbH</td>
			<td>3775380</td>
			<td></td>
		</tr>
		<tr>
			<td>(S)-ketamine</td>
			<td>CEVA Germany</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Xylazine</td>
			<td>Medistar Germany</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Consumable supplies</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>10 mL syringes</td>
			<td>BD Plastipak</td>
			<td>4606108V</td>
			<td></td>
		</tr>
		<tr>
			<td>2 mL syringes</td>
			<td>BD Plastipak</td>
			<td>4606027V</td>
			<td></td>
		</tr>
		<tr>
			<td>6-0 prolene suture</td>
			<td>ETHICON</td>
			<td>N-2719K</td>
			<td></td>
		</tr>
		<tr>
			<td>4-0 silk suture</td>
			<td>Seraflex</td>
			<td>IC 158000</td>
			<td></td>
		</tr>
		<tr>
			<td>Bepanthen Eye and Nose Ointment</td>
			<td>Bayer Vital GmbH</td>
			<td>6029009.00.00</td>
			<td></td>
		</tr>
		<tr>
			<td>Cotton Gauze swabs</td>
			<td>Fuhrmann GmbH</td>
			<td>32014</td>
			<td></td>
		</tr>
		<tr>
			<td>Durapore silk tape</td>
			<td>3M</td>
			<td>1538-1</td>
			<td></td>
		</tr>
		<tr>
			<td>Poly-Alcohol Skin Desinfection Solution</td>
			<td>Antiseptica GmbH</td>
			<td>72PAH200</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterican needle 18 G</td>
			<td>B. Braun</td>
			<td>304622</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterican needle 27 3/4 G</td>
			<td>B.Braun</td>
			<td>4657705</td>
			<td></td>
		</tr>
		<tr>
			<td>Tissue Paper</td>
			<td>commercially available</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical instruments</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Graefe forceps curved x1</td>
			<td>Fine Science Tools Inc.</td>
			<td>11151-10</td>
			<td></td>
		</tr>
		<tr>
			<td>Graefe forceps straight</td>
			<td>Fine Science Tools Inc.</td>
			<td>11050-10</td>
			<td></td>
		</tr>
		<tr>
			<td>Needle holder Mathieu</td>
			<td>Fine Science Tools Inc.</td>
			<td>12010-14</td>
			<td></td>
		</tr>
		<tr>
			<td>Scissors</td>
			<td>Fine Science Tools Inc.</td>
			<td>14074-11</td>
			<td></td>
		</tr>
		<tr>
			<td>Semken forceps</td>
			<td>Fine Science Tools Inc.</td>
			<td>11008-13</td>
			<td></td>
		</tr>
		<tr>
			<td>Small surgical scissors curved</td>
			<td>Fine Science Tools Inc.</td>
			<td>14029-10</td>
			<td></td>
		</tr>
		<tr>
			<td>Small surgical scissors straight</td>
			<td>Fine Science Tools Inc.</td>
			<td>14028-10</td>
			<td></td>
		</tr>
		<tr>
			<td>Standard pattern forceps</td>
			<td>Fine Science Tools Inc.</td>
			<td>11000-12</td>
			<td></td>
		</tr>
		<tr>
			<td>Vannas spring scissors</td>
			<td>Fine Science Tools Inc.</td>
			<td>15000-08</td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Equipment</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Dissecting microscope</td>
			<td>Leica MZ9</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Temperature controlled heating pad</td>
			<td>Sygonix</td>
			<td>26857617</td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Equipment for stent implantation</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Drug-eluting stent Xience 2,25mm x 8mm</td>
			<td>Abbott Vascular USA</td>
			<td>1009544-18</td>
			<td></td>
		</tr>
		<tr>
			<td>Guide wire Fielder XT PTCA guide wire: 0.014&#34; x 300cm</td>
			<td>ASAHI INTECC CO., LTD Japan</td>
			<td>AGP140302</td>
			<td></td>
		</tr>
		<tr>
			<td>Inflation syringe system</td>
			<td>Abbott 20/30 Priority Pack</td>
			<td>1000186</td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Tissue processing and analysis</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>30% H2O2</td>
			<td>Roth</td>
			<td>9681</td>
			<td>Histology</td>
		</tr>
		<tr>
			<td>Ethanol</td>
			<td>Roth</td>
			<td>K928.1</td>
			<td>Histology</td>
		</tr>
		<tr>
			<td>Giemsas Azur-Eosin-Methylenblau</td>
			<td>Merck</td>
			<td>109204</td>
			<td>Histology</td>
		</tr>
		<tr>
			<td>Graphic Drawing Tablet</td>
			<td>WACOM Europe GmbH</td>
			<td>CTL-6100WLK-S</td>
			<td></td>
		</tr>
		<tr>
			<td>Roti Histofix, Formaldehyd 4% buffered</td>
			<td>Roth</td>
			<td>P087</td>
			<td>Histology</td>
		</tr>
		<tr>
			<td>Technovit 9100</td>
			<td>Morphisto</td>
			<td>12225.K1000</td>
			<td>Histology</td>
		</tr>
	</tbody>
</table>

implantation of human-sized coronary stents into rat abdominal aorta using a trans-femoral access

La intervención coronaria percutánea (ICP), combinada con el despliegue de un stent coronario, representa el patrón oro en el tratamiento intervencionista de la enfermedad arterial coronaria. La reestenosis en stent (ISR) está determinada por una proliferación excesiva de tejido neointimal dentro del stent y limita el éxito a largo plazo de los stents. Una variedad de modelos animales se han utilizado para aclarar los procesos fisiopatológicos subyacentes a la reestenosis en stent (ISR), siendo los modelos porcinos coronarios y de la arteria ilíaca del conejo los más frecuentemente utilizados. Los modelos murinos proporcionan las ventajas de un alto rendimiento, facilidad de manejo y alojamiento, reproducibilidad y una amplia disponibilidad de marcadores moleculares. El modelo de ratón deficiente en apolipoproteína E (apoE-/-) ha sido ampliamente utilizado para estudiar enfermedades cardiovasculares. Sin embargo, los stents deben ser miniaturizados para ser implantados en ratones, lo que implica cambios importantes de sus propiedades mecánicas y (potencialmente) biológicas. El uso de apoE-/- las ratas pueden superar estas deficiencias como apoE-/- las ratas permiten la evaluación de stents coronarios de tamaño humano mientras que al mismo tiempo proporcionan un fenotipo atherogenic. Esto los convierte en un modelo excelente y confiable para investigar la ISR después de la implantación del stent. Aquí, describimos, detalladamente, la implantación de stents coronarios humanos disponibles en el comercio en la aorta abdominal de ratas con un apoE-/- fondo usando un acceso transporte-femoral.

Este protocolo describe la implantación de stent en la aorta abdominal de ratas utilizando una vía de acceso transfemoral (Figura 1). El primer punto central de este modelo animal es que permite el despliegue de stents coronarios de tamaño humano. Un stent coronario engarzado y montado en el globo disponible en el comercio se puede colocar en la aorta abdominal de ratas. Así, además, se puede aplicar el mismo principio de despliegue de stent que en humanos. Otra ventaja del uso de ratas...

Watch this Scientific Journal Video about Implantation of Human-Sized Coronary Stents into Rat Abdominal Aorta Using a Trans-Femoral Access at JoVE.com

Implantation of Human-Sized Coronary Stents into Rat Abdominal Aorta Using a Trans-Femoral Access

Este protocolo describe la implantación de stents coronarios humanos en la aorta abdominal de ratas con un apoE-/- fondo usando un acceso transporte-femoral. En comparación con otros modelos animales, los modelos murinos tienen las ventajas de un alto rendimiento, reproducibilidad, facilidad de manejo y alojamiento, y una amplia disponibilidad de marcadores moleculares.

Implantación De Stents Coronarios De Tamaño Humano En La Aorta Abdominal De Rata Usando Un Acceso Trans-Femoral

Percutaneous coronary intervention (PCI), combined with the deployment of a coronary stent, represents the gold standard in interventional treatment of ...

implantation-human-sized-coronary-stents-into-rat-abdominal-aorta

Research

JoVE Journal

Medicine

4.1K vistas.  University Hospital RWTH Aachen. Este protocolo describe la implantación de stents coronarios humanos en la aorta abdominal de ratas con un apoE-/- fondo usando un acceso transporte-femoral. En comparación con otros modelos animales, los modelos murinos tienen las ventajas de un alto rendimiento, reproducibilidad, facilidad de manejo y alojamiento, y una amplia disponibilidad de marcadores moleculares.

Video: Implantation of Human-Sized Coronary Stents into Rat Abdominal Aorta Using a Trans-Femoral Access

The C-seal: A Biofragmentable Drain Protecting the Stapled Colorectal Anastomosis from Leakage

Colorectal anastomotic leakage (AL) is a serious complication in colorectal surgery leading to high morbidity and mortality rates1. The incidence of AL varies between 2.5 and 20% 2-5. Over the years, many strategies aimed at lowering the incidence of anastomotic leakage have been examined6, 7. 
The cause of AL is probably multifactorial. Etiological factors include insufficient arterial blood supply, tension on the anastomosis, hematoma and/or infection at the anastomotic site, and co-morbid factors of the patient as diabetes and atherosclerosis8. Furthermore, some anastomoses may be insufficient from the start due to technical failure.
Currently a new device is developed in our institute aimed at protecting the colorectal anastomosis and lowering the incidence of AL. This so called C-seal is a biofragmentable drain, which is stapled to the anastomosis with the circular stapler. It covers the luminal side of the colorectal anastomosis thereby preventing leakage.
The C-seal is a thin-walled tube-like drain, with an approximate diameter of 4 cm and an approximate length of 25 cm (figure 1). It is a tubular device composed of biodegradable polyurethane. Two flaps with adhesive tape are found at one end of the tube. These flaps are used to attach the C-seal to the anvil of the circular stapler, so that after the anastomosis is made the C-seal can be pulled through the anus. The C-seal remains in situ for at least 10 days. Thereafter it will lose strength and will degrade to be secreted from the body together with the gastrointestinal natural contents.
The C-seal does not prevent the formation of dehiscences. However, it prevents extravasation of faeces into the peritoneal cavity. This means that a gap at the anastomotic site does not lead to leakage.
Currently, a phase II study testing the C-seal in 35 patients undergoing (colo-)rectal resection with stapled anastomosis is recruiting. The C-seal can be used in both open procedures as well as laparoscopic procedures. The C-seal is only applied in stapled anastomoses within 15cm from the anal verge. In the video, application of the C-seal is shown in an open extended sigmoid resection in a patient suffering from diverticular disease with a stenotic colon.

The C-seal is a biofragmentable drain protecting the stapled colorectal anastomosis. In this video, details of the manufacturing, deployment and use are shown.

El C-sello: Un drenaje Biofragmentable protección de la anastomosis colorrectal grapada de fuga

Colorectal anastomotic leakage (AL) is a serious complication in colorectal surgery leading to high morbidity and mortality rates1. The incidence of AL ...

Investigación

Medicina

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.

Rata heterotópico abdominal Corazón / Trasplante de un solo pulmón en una configuración de volumen cargado

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

Creation of Abdominal Adhesions in Mice

Abdominal adhesions consist of fibrotic tissue that forms in the peritoneal space in response to an inflammatory insult, typically surgery or intraabdominal infection. The precise mechanisms underlying adhesion formation are poorly understood. Many compounds and physical barriers have been tested for their ability to prevent adhesions after surgery with varying levels of success. The mouse and rat are important models for the study of abdominal adhesions. Several different techniques for the creation of adhesions in the mouse and rat exist in the literature. Here we describe a protocol utilizing abrasion of the cecum with sandpaper and sutures placed in the right abdominal sidewall. The mouse is anesthetized and the abdomen is prepped. A midline laparotomy is created and the cecum is identified. Sandpaper is used to gently abrade the surface of the cecum. Next, several figure-of-eight sutures are placed into the peritoneum of the right abdominal sidewall. The abdominal cavity is irrigated, a small amount of starch is applied, and the incision is closed. We have found that this technique produces the most consistent adhesions with the lowest mortality rate.

Abdominal adhesions that form after surgery are a major cause of pain, infertility, and hospitalization and reoperation for small bowel obstruction. Our surgical procedure for creating abdominal adhesions in mice is a reliable tool to study the mechanisms underlying the formation of adhesions.

Creación de abdominales adherencias en ratones

Abdominal adhesions consist of fibrotic tissue that forms in the peritoneal space in response to an inflammatory insult, typically surgery or ...

A Model of Cardiac Remodeling Through Constriction of the Abdominal Aorta in Rats

Heart failure is one of the leading causes of death worldwide. It is a complex clinical syndromethat includes fatigue, dyspnea, exercise intolerance, and fluid retention. Changes in myocardial structure, electrical conduction, and energy metabolism develop with heart failure, leading to contractile dysfunction, increased risk of arrhythmias, and sudden death. Hypertensive heart disease is one of the key contributing factors of cardiac remodeling associated with heart failure. The most commonly-used animal model mimicking hypertensive heart disease is created via surgical interventions, such as by narrowing the aorta. Abdominal aortic constriction is a useful experimental technique to induce a pressure overload, which leads to heart failure. The surgery can be easily performed, without the need for chest opening or mechanical ventilation. Abdominal aortic constriction-induced cardiac pathology progresses gradually, making this model relevant to clinical hypertensive heart failure. Cardiac injury and remodeling can be observed 10 weeks after the surgery. The method described here provides a simple and effective approach to produce a hypertensive heart disease animal model that is suitable for studying disease mechanisms and for testing novel therapeutics.

A rat model of abdominal aortic constriction that induces cardiac hypertrophy and remodeling is described. An efficient, highly-reproducible, and minimally-invasive method is used to provide a simple yet useful platform for research in myocardial hypertrophy and dysfunction.

Un modelo de remodelamiento cardiaco A través de la constricción de la aorta abdominal en ratas

Heart failure is one of the leading causes of death worldwide. It is a complex clinical syndromethat includes fatigue, dyspnea, exercise intolerance, and ...

Patch Angioplasty in the Rat Aorta or Inferior Vena Cava

Pericardial patches are commonly used in vascular surgery to close vessels. To facilitate studies of the neointimal hyperplasia that forms on the patch, we developed a rat model of patch angioplasty that can be used in either a vein or an artery, creating a patch venoplasty or a patch arterioplasty, respectively. Technical aspects of this model are discussed. The infra-renal IVC or aorta are dissected and then clamped proximally and distally. A 3 mm venotomy or arteriotomy is performed in the infrarenal inferior vena cava or aorta of 6 to 8 week-old Wistar rats. A bovine pericardial patch (3 mm x 1.5 mm x 0.6 mm) is then used to close the site using a 10-0 nylon suture. Compared to arterial patches, venous patches show increased neointimal thickness on postoperative day 7. This novel model of pericardial patch angioplasty can be used to examine neointimal hyperplasia on vascular biomaterials, as well as to compare the differences between the arterial and venous environments.

We have established a model of pericardial patch angioplasty that can be used in either small-diameter veins or arteries. This model can be used to compare venous and arterial neointimal hyperplasia formation.

Parche angioplastia en la aorta de rata o de la vena cava inferior

Pericardial patches are commonly used in vascular surgery to close vessels. To facilitate studies of the neointimal hyperplasia that forms on the patch, ...

Direct Re-implantation of Left Coronary Artery into the Aorta in Adults with Anomalous Origin of Left Coronary Artery from the Pulmonary Artery (ALCAPA)

Anomalous origin of the left coronary artery from the pulmonary artery (ALCAPA) is a rare congenital anomaly which is one of leading causes of myocardial ischemia and infarction in children. If left untreated, it results in a 90% mortality rate in the first year of life. In patients who survive to the adulthood, the coronary steal phenomenon and retrograde left-sided coronary flow provide a substrate for chronic subendocardial ischemia, which may lead to left ventricular dysfunction, ischemic mitral regurgitation, malignant ventricular arrhythmias, and sudden cardiac death. The average age of life-threatening presentation is 33 years and of sudden cardiac death 31 years. Therefore, surgical correction is highly recommended as soon as the diagnosis is made, regardless of age. In adult-type ALCAPA originating from the right-facing sinus of the pulmonary artery, direct re-implantation of the ALCAPA into the aorta is the more physiologically sound repair technique to re-establish the dual-coronary perfusion system and is recommended. This protocol describes the technique of direct re-implantation of adult-type ALCAPA into the aorta.

Direct Re-implantation of Left Coronary Artery into the Aorta in Adults with Anomalous Origin of Left Coronary Artery from the Pulmonary Artery ALCAPA

Surgical correction of ALCAPA is highly recommended, regardless of age or the degree of intercoronary collateralization. This protocol presents a technique for the direct re-implantation of adult-type ALCAPA into the aorta to re-establish the dual-coronary perfusion. Whenever feasible, direct re-implantation is preferred to other surgical correction techniques.

Directa reimplantación de la arteria coronaria izquierda en la aorta en adultos con origen anómalo de la arteria coronaria izquierda desde la arteria pulmonar (ALCAPA)

Anomalous origin of the left coronary artery from the pulmonary artery (ALCAPA) is a rare congenital anomaly which is one of leading causes of myocardial ...

Balloon-based Injury to Induce Myointimal Hyperplasia in the Mouse Abdominal Aorta

The use of animal models is essential for a better understanding of MH, one major cause for arterial stenosis.In this article, we demonstrate a murine balloon denudation model, which is comparable with established vessel injury models in large animals. The aorta denudation model with balloon catheters mimics the clinical setting and leads to comparable pathobiological and physiological changes. Briefly, after performing a horizontal incision in the aorta abdominalis, a balloon catheter will be inserted into the vessel, inflated, and introduced retrogradely. Inflation of the balloon will lead to intima injury and overdistension of the vessel. After removing the catheter, the aortic incision will be closed with single stiches. The model shown in this article is reproducible, easy to perform, and can be established quickly and reliably. It is especially suitable for evaluating expensive experimental therapeutic agents, which can be applied in an economical fashion. By using different knockout-mouse strains, the impact of different genes on MH development can be assessed.

This article demonstrates a murine model to study the development of myointimal hyperplasia (MH) after aortic balloon injury.

Lesión basada en globo para inducir la hiperplasia Myointimal de la Aorta Abdominal de ratón

The use of animal models is essential for a better understanding of MH, one major cause for arterial stenosis.In this article, we demonstrate a murine ...

Ultrasound Imaging of the Thoracic and Abdominal Aorta in Mice to Determine Aneurysm Dimensions

Contemporary high-resolution ultrasound instruments have sufficient resolution to facilitate the measurement of mouse aortas. These instruments have been widely used to measure aortic dimensions in mouse models of aortic aneurysms. Aortic aneurysms are defined as permanent dilations of the aorta, which occur most frequently in the ascending and abdominal regions. Sequential measurements of aortic dimensions by ultrasound are the principal approach for assessing the development and progression of aortic aneurysms in vivo. Although many reported studies used ultrasound imaging to measure aortic diameters as a primary endpoint, there are confounding factors, such as probe position and cardiac cycle, that may impact the accuracy of data acquisition, analysis, and interpretation. The purpose of this protocol is to provide a practical guide on the use of ultrasound to measure the aortic diameter in a reliable and reproducible manner. This protocol introduces the preparation of mice and instruments, the acquisition of appropriate ultrasound images, and data analysis.

Ultrasound imaging has become a common modality to determine the luminal dimensions of thoracic and abdominal aortic aneurysms in mice. This protocol describes the procedure to acquire reliable and reproducible two-dimensional ultrasound images of the ascending and abdominal aorta in mice.

Imágenes por ultrasonido de la Aorta torácica y Abdominal en ratones para determinar dimensiones de aneurisma

Contemporary high-resolution ultrasound instruments have sufficient resolution to facilitate the measurement of mouse aortas. These instruments have been ...

A Murine Model of Hemodialysis Access-Related Hand Dysfunction

Chronic kidney disease is a major public health problem, and the prevalence of end-stage renal disease (ESRD) requiring chronic renal replacement therapies such as hemodialysis continues to increase. Autogenous arteriovenous fistula (AVF) placement remains a primary vascular access option for ESRD patients. Unfortunately, approximately half of the hemodialysis patients experience dialysis access-related hand dysfunction (ARHD), ranging from subtle paresthesia to digital gangrene. Notably, the underlying biologic drivers responsible for ARHD are poorly understood, and no adequate animal model exists to elucidate the mechanisms and/or develop novel therapeutics for the prevention/treatment of ARHD. Herein, we describe a new mouse model in which an AVF is created between the left common iliac artery and vein, thereby facilitating the assessment of limb pathophysiology. The microsurgery includes vessel isolation, longitudinal venotomy, creation of arteriovenous anastomosis, and venous reconstruction. Sham surgeries include all the critical steps except for AVF creation. Iliac AVF placement results in clinically relevant alterations in central hemodynamics, peripheral ischemia, and impairments in hindlimb neuromotor performance. This novel preclinical AVF model provides a useful platform that recapitulates common neuromotor perturbations reported by hemodialysis patients, allowing researchers to investigate the mechanisms of ARHD pathophysiology and test potential therapeutics.

This protocol details the surgical steps of murine common iliac arteriovenous fistula creation. We developed this model to study hemodialysis access-related limb pathophysiology.

Un modelo murino de disfunción de la mano relacionada con el acceso a la hemodiálisis

Chronic kidney disease is a major public health problem, and the prevalence of end-stage renal disease (ESRD) requiring chronic renal replacement ...

Robotic Cochlear Implantation for Direct Cochlear Access

Robot-assisted systems offer great potential for gentler and more precise cochlear implantation. In this article, we provide a comprehensive overview of the clinical workflow for robotic cochlear implantation using a robotic system specifically developed for a minimally invasive, direct cochlear access. The clinical workflow involves experts from various disciplines and requires training to ensure a smooth and safe procedure. The protocol briefly summarizes the history of robotic cochlear implantation. The clinical sequence is explained in detail, beginning with the assessment of patient eligibility and covering surgical preparation, preoperative planning with the special planning software, drilling of the middle ear access, intraoperative imaging to confirm the trajectory, milling of the inner ear access, insertion of the electrode array, and implant management. The steps that require special attention are discussed. As an example, the postoperative outcome of robotic cochlear implantation in a patient with advanced otosclerosis is presented. Finally, the procedure is discussed in the context of the authors' experience.

Robotic cochlear implantation is a procedure for minimally invasive inner ear access. Compared to conventional surgery, robotic cochlear implantation involves additional steps that need to be carried out in the operating room. In this article we give a description of the procedure and highlight the important aspects of robotic cochlear implantation.

Implantación coclear robótica para acceso coclear directo

Robot-assisted systems offer great potential for gentler and more precise cochlear implantation. In this article, we provide a comprehensive overview of ...