Todos los procedimientos descritos se llevan a cabo de acuerdo con los Institutos Nacionales de Salud directrices para el cuidado y uso de animales en la investigación y todos los animales protocolos fueron aprobados por la Oregon Health &amp; Science de la Universidad de Cuidado de Animales institucional y el empleo. 1. Preparación quirúrgica <ol><li> Pesar el ratón. El procedimiento descrito se lleva a cabo en ratones C57BL / 6 que pesan entre 20 y 25 g. La anestesia es inducida en una caja de inducción con isoflurano 3.4%, y posteriormente mantuvo con 1,5-2,5% de isoflurano en la mezcla de aire / oxígeno. </li><li> Lubricar los ojos y la posición supina de los animales en un cojín de la calefacción. Inmovilizar cuatro extremidades con cinta adhesiva. Las patas traseras, se puede grabar en una posición neutral, sin embargo, las patas delanteras deben ser asegurados lo más cerca posible de la pared torácica como sea posible para permitir excursión de la pared torácica durante las compresiones torácicas. </li><li> Lubricar y colocar una sonda de temperatura rectal. La temperatura se controla mediante una almohadilla eléctrica y la lámpara conectada a un controlador electrónico de temperatura (Digi-Sense, Cole Parmer, Vernon Hills IL), que contempla el mantenimiento de 37,0 ° C. Debido a que es posible que un gradiente de temperatura en el animal puede desarrollar durante el estado de no-flujo, es importante que la temperatura se mide y controla cerca del órgano de interés. </li><li> Intubar la tráquea con un 2.5 de 22 cm ga catéter de teflón (Insyte-W, BD, Franklin NJ) y el extremo del corte distal de un introductor en ángulo (Frova Introductor, Médico Cook, Bloomington IN). Otros métodos de intubación traqueal son aceptables, sin embargo, el uso del introductor en ángulo permite un posicionamiento adecuado en el cuello en una posición ligeramente extendida, que optimiza la exposición quirúrgica para la colocación del catéter intravenoso. La conexión del catéter endotraqueal se asegura con un lazo de sutura para el incisivo y se mantiene con una ligera tensión para inmovilizar la cabeza durante las compresiones torácicas. </li><li> Ventilar mecánicamente el ratón con un ventilador de roedores conjunto a 140 l, 150 respiraciones / minuto. Esto no es ajustado por el peso de este protocolo se realiza siempre en 20-25 g animales. No se exceda en profundizar la anestesia. No coloque el ventilador para aplicar la presión espiratoria final, ya que perjudica la resucitación. </li><li> Utilizando una técnica estéril y de un microscopio, un lugar pre-lavado PE-10 del catéter en la vena yugular. Cualquiera de las partes puede ser utilizado, pero el uso de la parte más cercana al operador reduce la posibilidad de desprendimiento del catéter durante las compresiones en el pecho u otras manipulaciones. 0,5% bupivicaína, 0,1 mL es infitrated en bordes de la herida para controlar el dolor postoperatorio. </li><li> Asegure el catéter PE-10 en el cierre de la piel con un adhesivo de cianoacrilato quirúrgico. </li><li> Lugar subcutánea electrodos de EKG y conectarse a un dispositivo de vigilancia. Atención a la trayectoria de la señal y la maximización de la propia señal es crucial para el éxito de la reanimación. Asegúrese de que todos los cables se fijan a la superficie de trabajo, reducir al mínimo los cruces de la señal, y minimizar los aislantes (como el aire de agujas huecas o cable trenzado) en el camino de la señal. Agujas sólidas conectado a cable nonstranded puede ser comprado o hecho en el laboratorio. Una vez conectado, optimizar la señal de ECG en el monitor. </li></ol> 2. Paro cardíaco <ol><li> Asegúrese de que el ratón es normotérmica, que se define como la temperatura rectal de 36,5 º C - 37,5 ° C. Administrar 40 ml de la temperatura ambiente 0,5 M de cloruro de potasio por vía intravenosa y observar isoeléctrico trazado en el ECG. Ajuste de la dosis para el peso no es necesario dentro del rango de 20 a 25 g. Iniciar el temporizador de detención. </li><li> Desconecte el respirador artificial. Interrumpir el vapor anestésico. Apague el cojín de la calefacción y cualquier otro equipo que produce el ruido electrónico que puede interferir con la monitorización ECG. Coloque una capa aislante sobre el ratón. </li><li> Registrar la temperatura cada minuto durante el paro cardiaco. Si es necesario, una lámpara de calor se puede utilizar para llevar la temperatura central hasta el rango normotérmica. Durante un paro cardiaco, preparar materiales y equipos (por ejemplo, la jeringa de epinefrina) para la reanimación. Una lista de verificación puede ser útil para garantizar la reanimación sin interrupciones, lo cual es esencial para la supervivencia. </li><li> Después de 7 minutos y 30 segundos de un paro cardíaco, vuelva a conectar el ventilador y aumentar la tasa a 180 respiraciones / min, manteniendo el volumen de marea en la fase previa al arresto. </li><li> A los 8 minutos, iniciar compresiones de pecho a 300 BPM. Artefactos de movimiento en el ECG se puede utilizar para juzgar la tasa de RCP. Compresiones torácicas debe ser entregado con el dedo índice, de 5 mm por encima de la apófisis xifoides y ligeramente a la izquierda de la línea media. El pecho debe ser comprimido 1.3 a 1.2 de la distancia anteroposterior y el retroceso completo se debe permitir que entre las compresiones. Posición de los dedos y la presión de compresión óptimo son absolutamente críticos. La imposibilidad de lograr la supervivencia en este modelo es casi siempre debido a las óptimas CPR. </li><li> Infundir 0,5 ml de epinefrina, diluida a 15 mg / ml, en los primeros 30 segundos de RCP. Dosis de adrenalina total es de 8.12 mg. Observe cuidadosamente el ECG para el retorno de la circulación espontánea (ROSC). Estrecha complejo QRS se ven entre los artefactos de compresión. ROSC ocurre generalmente entre 90 segundos y 2 minutos después de comenzar la RCP. Reanimación es abandonada si ROSC no se produce por 3 minutos. Contracciones ventriculares prematuras frecuentes y los cambios en el ECG eje se observan en los primeros 2 minutos después de ROSC, y casi siempre se resuelven en la taquicardia sinusal estable en 2 minutos. </li><li> Registrar el tiempo total de reanimación y la dosis de epinefrina. Registran la temperatura cada minuto durante 10 minutos después de ROSC. </li><li> Lleva EKG puede ser removido cuando la respiración espontánea se inicia, por lo general en 12-15 minutos después de ROSC. </li><li> Retirar la intubación de la tráquea cuando la frecuencia respiratoria espontánea&gt; 60/min. </li><li> Coloque el ratón en una jaula de recuperación en una superficie de temperatura controlada conjunto a 37 ° C durante las primeras 2 horas después del procedimiento, lo que garantiza un fácil acceso a los alimentos y el agua. La jaula se puede trasladar a las condiciones estándar de vivienda postoperatorio cuando el ratón se ha recuperado completamente de la anestesia y está activo. </li></ol> 3. Exanguinación / perfusión-fijación y la cosecha del Riñón <ol><li> 24 horas después de la CA / RCP, inducir la anestesia con isoflurano al 3-4% y seguro que el animal en decúbito supino sobre una superficie quirúrgico dentro de una campana extractora que es apropiado para su uso con formol .. </li><li> Profundizar la anestesia, aumentando gradualmente la concentración de vapor anestésico a un 5% y garantizar que el ratón está profundamente anestesiados como se evidencia por el cese de la respiración espontánea. </li><li> Realizar una toracotomía cubierta y exsanguinate el ratón a través de la punción apical del ventrículo cardíaco izquierdo de acuerdo con técnicas estándar. </li><li> A través de la misma aguja, administrar salina al 0,9% en infusión lenta. Para acclerate perfusión / desangrado, nick de la orejuela derecha con unas tijeras. Realizar una laparotomía y observar los riñones. Cuando se blanquean, solución salina puede ser cambiado a un 4% de formol para su fijación. Los riñones son luego incluidos en parafina y teñidas con B fluoro-jade para la cuantificación de la necrosis del epitelio tubular. </li></ol> 4. Los resultados representativos: Cuando se induce CA, la presión arterial media (PAM) y la regional del flujo renal sanguíneo (RRBCF) gota a cerca de cero y permanece estable hasta la reanimación comienza (Figura 1). 24 horas después de la CA / RCP, los índices de suero de la función renal (nitrógeno ureico en sangre, creatinina y BUN) son significativamente elevados en relación con operación simulada animales. AST / ALT también son elevados, ofreciendo evidencia de la profunda de todo el cuerpo isquemia (Figura 2). Neutrófilos gelatinasa lipocalina asociada (NGAL), un indicador sensible de la lesión isquémica renal es masivamente upregulated después de 24 CA / CPR (Figura 3). Por último, microfotografías demuestran la necrosis parcheada, medular típico de lesión renal isquémica con el adelgazamiento del epitelio tubular y el relleno luminal de hematoxilina y eosina (Figura 4, panel a) y una extensa muerte celular en el epitelio tubular medular cuando se tiñen con B fluoro-jade ( Figura 4, panel b). <img src="/files/ftp_upload/3116/3116fig1.jpg" alt="Figura 1" /> Figura 1. Paro cardíaco induce la pérdida inmediata de la presión de perfusión, representado aquí como la presión arterial media (PAM), medido en la arteria femoral, lo que resulta en casi completo cese de la regional del flujo sanguíneo renal cortical (RRCBF) durante todo el período de paro cardíaco (la sombra área). Reanimación con masaje cardíaco y epinefrina retorna a la normalidad y MAPA RRCBF constantemente se levanta en el periodo de reanimación post. Reproducido con permiso del 14. <img src="/files/ftp_upload/3116/3116fig2.jpg" alt="Figura 2" /> Figura 2. 24h después del procedimiento, nitrógeno ureico en sangre (BUN), creatinina sérica, y la extensión de la muerte de las células tubulares son significativamente más elevados en los animales que hayan sido CA / RCP en comparación con los animales tratados con un procedimiento simulado. CA / RCP induce una lesión isquémica pan-de organismos, aquí se evidencia por la elevación masiva de enzimas de función hepática alanina aminotransferasa (ALT) y aspartato aminotransferasa (AST) en CA / RCP ratones en comparación con el simulacro de los animales tratados. <img src="/files/ftp_upload/3116/3116fig3.jpg" alt="Figura 3" /> Figura 3. Western blot realizado con anticuerpo policlonal de neutrófilos gelatinasa lipocalina asociada (NGAL), un indicador sensible de la lesión isquémica renal. Las muestras de orina fueron obtenidas inmediatamente antes (&quot;pre&quot;) y 24 horas después (&quot;24&quot;) CA / CPR en 4 animales (a, b, c, d, y más arriba). NGAL es enormemente aumentada en ratonesorina después de la CA / RCP. Reproducido con permiso del 14. <img src="/files/ftp_upload/3116/3116fig4.jpg" alt="Figura 4" /> Figura 4. A) hematoxilina y eosina de una sección de eje corto de 24 hiliares tejido renal después de CA / RCP. Hay un daño irregular pero claro medular y corticomedular túbulos con tubular taponamiento. Las flechas señalan los túbulos dañados con núcleos hinchados, picnóticos en la unión corticomedular. B) B-jade flúor mancha de la misma región en el mismo animal, 24 horas después de CA / RCP. Jade fluoro-manchas B células necróticas de color verde brillante, mostrando irregular necrosis tubular corticomedular. Las flechas señalan los túbulos brillantes manchas dañado en la unión corticomedular. Estos resultados son sustancialmente similares a hallazgos de la biopsia renal de los seres humanos que desarrollan IRA, a diferencia de las producidas por otros modelos animales de IRA.

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No hay conflictos de interés declarado.

El modelo normotérmica de un paro cardíaco y resucitación cardiopulmonar en el ratón ofrece múltiples posibilidades de la evaluación en un modelo que reproduce la fisiopatología y la morfología de la causa clínica más frecuente de lesión renal aguda en todo el cuerpo hipoperfusión. Prueba de hipótesis puede ser ayudado por el acceso a un abanico de técnicas de manipulación genética y la anatomía bien entendido y se caracteriza y la fisiología de los ratones de laboratorio. ...

<table border="1"><tbody><tr><th> Nombre del reactivo </th><th> Empresa </th><th> Número de catálogo </th></tr><tr><td> Cloruro de Potasio </td><td> Abbott Medical </td><td> 06653-5 </td></tr><tr><td> Isoflurano </td><td> Abbott Medical </td><td> 05260-05 (y otros) </td></tr><tr><td> De clorhidrato de epinefrina </td><td> Múltiple </td><td> Múltiple </td></tr><tr><td> Digi-Sense controlador de temperatura </td><td> Cole-Palmer </td><td> EW-89.000-00 </td></tr><tr><td> Angiocath IV Cath </td><td> VWR </td><td> 381134 </td></tr><tr><td> Frova introductor vía aérea en ángulo </td><td> Cocinar </td><td> G27282 </td></tr><tr><td> Ventilador MicroVent de ratones </td><td> Harvard Apparatus </td><td> 733591 </td></tr></tbody></table>

normothermic cardiac arrest and cardiopulmonary resuscitation: a mouse model of ischemia-reperfusion injury

La lesión renal aguda (IRA) es un común, altamente letal, complicación de la enfermedad grave que tiene una alta mortalidad 1-4 y que es más frecuentemente causada por la hipoperfusión de todo el cuerpo. 5,6 reproducción exitosa de la hipoperfusión en todo el cuerpo-en modelos de roedores ha estado plagado de dificultades. 7-9,9,10 modelos que emplean la isquemia focal han demostrado repetidamente que los resultados no se traducen en la práctica clínica, y más modelos animales que permiten acceder a la falta de cuerpo entero hipoperfusión al conjunto de herramientas completo de la manipulación genética posible en el ratón. 11,12 Sin embargo, en los últimos años un modelo de ratón de un paro cardíaco y resucitación cardiopulmonar se ha convertido en lo que se puede adaptar el modelo de la IRA. 13 Este modelo reproduce de forma fiable los resultados fisiológicos y funcionales, anatómicos e histológicos visto en la práctica clínica AKI , es repetible rápidamente, y ofrece todas las ventajas de un modelo murino quirúrgica, incluido el acceso a técnicas de manipulación genética, en relación de bajo costo para animales de gran tamaño, y facilidad de uso. Nuestro grupo ha desarrollado una amplia experiencia con el uso de este modelo para evaluar una serie de órganos específicos en los resultados AKI 14,15.

Watch this Scientific Journal Video about Normothermic Cardiac Arrest and Cardiopulmonary Resuscitation: A Mouse Model of Ischemia-Reperfusion Injury at JoVE.com

Normothermic Cardiac Arrest and Cardiopulmonary Resuscitation: A Mouse Model of Ischemia-Reperfusion Injury

Un modelo de gran alcance para los cuidados perioperatorios y críticos relacionados con la insuficiencia renal aguda se presenta. Utilizando todo el cuerpo hipoperfusión inducida por paro cardíaco, es posible reproducir casi los cambios histológicos y funcionales de la clínica AKI.

Arresto normotérmica cardiaca y reanimación cardiopulmonar: un modelo de ratón de la lesión por isquemia-reperfusión

Acute Kidney Injury (AKI) is a common, highly lethal, complication of critical illness which has a high mortality1-4 and which is most 
frequently caused ...

normothermic-cardiac-arrest-cardiopulmonary-resuscitation-mouse-model

Research

JoVE Journal

Medicine

17.9K vistas.  Oregon Health & Sciences University. Un modelo de gran alcance para los cuidados perioperatorios y críticos relacionados con la insuficiencia renal aguda se presenta. Utilizando todo el cuerpo hipoperfusión inducida por paro cardíaco, es posible reproducir casi los cambios histológicos y funcionales de la clínica AKI.

Video: Normothermic Cardiac Arrest and Cardiopulmonary Resuscitation: A Mouse Model of Ischemia-Reperfusion Injury

Ischemia-reperfusion Model of Acute Kidney Injury and Post Injury Fibrosis in Mice

Ischemia-reperfusion induced acute kidney injury (IR-AKI) is widely used as a model of AKI in mice, but results are often quite variable with high, often unreported mortality rates that may confound analyses. Bilateral renal pedicle clamping is commonly used to induce IR-AKI, but differences between effective clamp pressures and/or renal responses to ischemia between kidneys often lead to more variable results. In addition, shorter clamp times are known to induce more variable tubular injury, and while mice undergoing bilateral injury with longer clamp times develop more consistent tubular injury, they often die within the first 3 days after injury due to severe renal insufficiency. To improve post-injury survival and obtain more consistent and predictable results, we have developed two models of unilateral ischemia-reperfusion injury followed by contralateral nephrectomy. Both surgeries are performed using a dorsal approach, reducing surgical stress resulting from ventral laparotomy, commonly used for mouse IR-AKI surgeries. For induction of moderate injury BALB/c mice undergo unilateral clamping of the renal pedicle for 26 min and also undergo simultaneous contralateral nephrectomy. Using this approach, 50-60% of mice develop moderate AKI 24 hr after injury but 90-100% of mice survive. To induce more severe AKI, BALB/c mice undergo renal pedicle clamping for 30 min followed by contralateral nephrectomy 8 days after injury. This allows functional assessment of renal recovery after injury with 90-100% survival. Early post-injury tubular damage as well as post injury fibrosis are highly consistent using this model.

We describe models of moderate and severe ischemia-reperfusion-induced kidney injury in which the mice undergo unilateral renal pedicle clamping followed by simultaneous or delayed contralateral nephrectomy, respectively. These models consistently give rise to renal dysfunction and post-injury fibrosis, but injury severity and survival are dependent on mouse background, age and surgical equipment.

La isquemia-reperfusión modelo de lesión renal aguda y post lesión fibrosis en ratones

Ischemia-reperfusion induced acute kidney injury (IR-AKI) is widely used as a model of AKI in mice, but results are often quite variable with high, often ...

Investigación

Medicina

Induction and Assessment of Ischemia-reperfusion Injury in Langendorff-perfused Rat Hearts

The biochemical events surrounding ischemia reperfusion injury in the acute setting are of great importance to furthering novel treatment options for myocardial infarction and cardiac complications of thoracic surgery. The ability of certain drugs to precondition the myocardium against ischemia reperfusion injury has led to multiple clinical trials, with little success. The isolated heart model allows acute observation of the functional effects of ischemia reperfusion injury in real time, including the effects of various pharmacological interventions administered at any time-point before or within the ischemia-reperfusion injury window. Since brief periods of ischemia can precondition the heart against ischemic injury, in situ aortic cannulation is performed to allow for functional assessment of non-preconditioned myocardium. A saline filled balloon is placed into the left ventricle to allow for real-time measurement of pressure generation. Ischemic injury is simulated by the cessation of perfusion buffer flow, followed by reperfusion. The duration of both ischemia and reperfusion can be modulated to examine biochemical events at any given time-point. Although the Langendorff isolated heart model does not allow for the consideration of systemic events affecting ischemia and reperfusion, it is an excellent model for the examination of acute functional and biochemical events within the window of ischemia reperfusion injury as well as the effect of pharmacological intervention on cardiac pre- and postconditioning. The goal of this protocol is to demonstrate how to perform in situ aortic cannulation and heart excision followed by ischemia/reperfusion injury in the Langendorff model.

The isolated rat heart is an enduring model for ischemia reperfusion injury. Here, we describe the process of harvesting the beating heart from a rat via in situ aortic cannulation, Langendorff perfusion of the heart, simulated ischemia-reperfusion injury, and infarct staining to confirm the extent of ischemic insult.

Inducción y Evaluación de la isquemia-reperfusión en perfundidos Langendorff corazones de ratas

The biochemical events surrounding ischemia reperfusion injury in the acute setting are of great importance to furthering novel treatment options for ...

Murine Model of Intestinal Ischemia-reperfusion Injury

Intestinal ischemia is a life-threatening condition associated with a broad range of clinical conditions including atherosclerosis, thrombosis, hypotension, necrotizing enterocolitis, bowel transplantation, trauma and chronic inflammation. Intestinal ischemia-reperfusion (IR) injury is a consequence of acute mesenteric ischemia, caused by inadequate blood flow through the mesenteric vessels, resulting in intestinal damage. Reperfusion following ischemia can further exacerbate damage of the intestine. The mechanisms of IR injury are complex and poorly understood. Therefore, experimental small animal models are critical for understanding the pathophysiology of IR injury and the development of novel therapies.
Here we describe a mouse model of acute intestinal IR injury that provides reproducible injury of the small intestine without mortality. This is achieved by inducing ischemia in the region of the distal ileum by temporally occluding the peripheral and terminal collateral branches of the superior mesenteric artery for 60 min using microvascular clips. Reperfusion for 1 hr, or 2 hr after injury results in reproducible injury of the intestine examined by histological analysis. Proper position of the microvascular clips is critical for the procedure. Therefore the video clip provides a detailed visual step-by-step description of this technique. This model of intestinal IR injury can be utilized to study the cellular and molecular mechanisms of injury and regeneration.

Here we describe the detailed procedure of intestinal ischemia-reperfusion in mice which results in reproducible injury without mortality to encourage the standardization of this technique across the field. This model of intestinal ischemia-reperfusion injury can be utilized to study the cellular and molecular mechanisms of injury and regeneration.

Modelo murino de isquemia-reperfusión intestinal Lesiones

Intestinal ischemia is a life-threatening condition associated with a broad range of clinical conditions including atherosclerosis, thrombosis, ...

A Mouse Model of Retinal Ischemia-Reperfusion Injury Through Elevation of Intraocular Pressure

Retinal ischemia-reperfusion (I/R) is a pathophysiological process contributing to cellular damage in multiple ocular conditions, including glaucoma, diabetic retinopathy, and retinal vascular occlusions. Rodent models of I/R injury are providing significant insights into mechanisms and treatment strategies for human I/R injury, especially with regard to neurodegenerative damage in the retinal neurovascular unit. Presented here is a protocol for inducing retinal I/R injury in mice through elevation of intraocular pressure (IOP). In this protocol, the ocular anterior chamber is cannulated with a needle, through which flows the drip of an elevated saline reservoir. Using this drip to raise IOP above systolic arterial blood pressure, a practitioner temporarily halts inner retinal blood flow (ischemia). When circulation is reinstated (reperfusion) by removal of the cannula, severe cellular damage ensues, resulting ultimately in retinal neurodegeneration. Recent studies demonstrate inflammation, vascular permeability, and capillary degeneration as additional elements of this model. Compared to alternative retinal I/R methodologies, such as retinal arterial ligation, retinal I/R injury by elevated IOP offers advantages in its&#160;anatomical specificity, experimental tractability, and technical accessibility, presenting itself as a valuable tool for examining neuronal pathogenesis and therapy in the retinal neurovascular unit.

This article describes a procedure for inducing retinal ischemia-reperfusion injury by elevated intraocular pressure in mice. Retinal ischemia-reperfusion injury by elevated intraocular pressure serves to model human pathologies characterized by compromised oxygen and nutrient delivery in the retina, enabling researchers to examine potential cellular mechanisms and treatments for human diseases of the retinal neurovascular unit.

Un modelo murino de isquemia retiniana-reperfusión mediante la elevación de la presión intraocular

Retinal ischemia-reperfusion (I/R) is a pathophysiological process contributing to cellular damage in multiple ocular conditions, including glaucoma, ...

Ultrasonographic Assessment During Cardiopulmonary Resuscitation

The US-CAB (Ultrasound, Circulation/Airway/Breathing) protocol integrates several sonographic techniques into a structured assessment of the circulation, airway, and breathing status of a patient during cardiopulmonary resuscitation (CPR) in an advanced life support-compliant manner. US-C provides a subxiphoid view of the heart, to look for potentially reversible causes of disease, such as pericardial effusion, pulmonary embolism, hypovolemia, and acute coronary thrombosis. Sonographic cardiac activity during CPR not only helps differentiate pseudo-pulseless electrical activity (PEA) from true PEA but also represents a higher chance of the return of spontaneous circulation (ROSC) and survival. Evaluation of the inferior vena cava (IVC) shows the fluid status of the patient and indicates the best methods to use for fluid resuscitation. If aortic dissection is suspected, a subxiphoid view of the aorta is suggested for identifying an intimal flap. Once intubation is done, tracheal ultrasound (US-A) at the suprasternal notch helps differentiate endotracheal intubation (one air-mucosal interface with one comet-tail) from esophageal intubation (double tract sign). Immediately following US-A, bilateral lung US (US-B) should be done to confirm proper bilateral ventilation using the lung sliding sign. In addition, US-C can be serially followed to see the dynamic changes in the cardiac chambers and IVC, or any cardiac contraction suggestive of ROSC. US-B can also detect coexisting lung or pleural pathologies without interfering with the performance of CPR. The main concern when implementing this method is maintaining high-quality CPR without delays in chest compressions when performing US-CAB. Rigorous training and continued practice are key to minimize any interruptions during resuscitation.

Here, we present a US-CAB (Ultrasound, Circulation/Airway/Breathing) protocol for use during cardiopulmonary resuscitation (CPR). US-C evaluates the subxiphoid view of the heart and inferior vena cava. After intubation, tracheal US (US-A) and lung US (US-B) help confirm endotracheal intubation and proper ventilation.

Evaluación ultrasonográfica durante la reanimación cardiopulmonar

The US-CAB (Ultrasound, Circulation/Airway/Breathing) protocol integrates several sonographic techniques into a structured assessment of the circulation, ...

A Pre-clinical Rat Model for the Study of Ischemia-reperfusion Injury in Reconstructive Microsurgery

Ischemia-reperfusion injury is the main cause of flap failure in reconstructive microsurgery. The rat is the preferred preclinical animal model in many areas of biomedical research due to its cost-effectiveness and its translation to humans. This protocol describes a method to create a preclinical free skin flap model in rats with ischemia-reperfusion injury. The described 3 cm x 6 cm rat free skin flap model is easily obtained after the placement of several vascular ligatures and the section of the vascular pedicle. Then, 8 h after the ischemic insult and completion of the microsurgical anastomosis, the free skin flap develops the tissue damage. These ischemia-reperfusion injury-related damages can be studied in this model, making it a suitable model for evaluating therapeutic agents to address this pathophysiological process. Furthermore, two main monitoring techniques are described in the protocol for the assessment of this animal model: transit-time ultrasound technology and laser speckle contrast analysis.

Here, we describe a pre-clinical animal model for studying the pathophysiology of ischemia-reperfusion injury in reconstructive microsurgery. This free skin flap model based on the superficial caudal epigastric vessels in the rat may also allow for the evaluation of different therapies and compounds to counteract ischemia-reperfusion injury-related damage.

Un modelo preclínico de rata para el estudio de la lesión por isquemia-reperfusión en microcirugía reconstructiva

Ischemia-reperfusion injury is the main cause of flap failure in reconstructive microsurgery. The rat is the preferred preclinical animal model in many ...

Delayed Intramyocardial Delivery of Stem Cells after Ischemia Reperfusion Injury in a Murine Model

There is significant interest in the use of stem cells (SCs) for the recovery of cardiac function in individuals with myocardial injuries. Most commonly, cardiac stem cell therapy is studied by delivering SCs concurrently with the induction of myocardial injury. However, this approach presents two significant limitations: the early hostile pro-inflammatory ischemic environment may affect the survival of transplanted SCs, and it does not represent the subacute infarction scenario where SCs will likely be used. Here we describe a two-part series of surgical procedures for the induction of ischemia-reperfusion injury and delivery of mesenchymal stem cells (MSCs). This method of stem cell administration may allow for the longer viability and retention around damaged tissue by circumventing the initial immune response. A model of ischemia reperfusion injury was induced in mice accompanied by the delivery of mesenchymal stem cells (3.0 x 105), stably expressing the reporter gene firefly luciferase under the constitutively expressed CMV promoter, intramyocardially 7 days later. The animals were imaged via ultrasound and bioluminescent imaging for confirmation of injury and injection of cells, respectively. Importantly, there was no added complication rate when performing this two-procedure approach for SC delivery. This method of stem cell administration, collectively with the utilization of state-of-the-art reporter genes, may allow for the in vivo study of viability and retention of transplanted SCs in a situation of chronic ischemia commonly seen clinically, while also circumventing the initial pro-inflammatory response. In summary, we established a protocol for the delayed delivery of stem cells into the myocardium, which can be used as a potential new approach in promoting regeneration of the damaged tissue.

Stems cells are continuously investigated as potential treatments for individuals with myocardial damage, however, their decreased viability and retention within injured tissue can impact their long-term efficacy. In this manuscript we describe an alternative method for stem cell delivery in a murine model of ischemia reperfusion injury.

Retraso en la administración intramiocardio de células madre después de la lesión por reperfusión de isquemia en un modelo murino

There is significant interest in the use of stem cells (SCs) for the recovery of cardiac function in individuals with myocardial injuries. Most commonly, ...

A Rat Lung Transplantation Model of Warm Ischemia/Reperfusion Injury: Optimizations to Improve Outcomes

From our experience with rat lung transplantation, we have found several areas for improvement. Information in the existing literature regarding methods for choosing appropriate cuff sizes for the pulmonary vein (PV), pulmonary artery (PA), or bronchus (Br) are varied, thus making the determination of proper cuff size during rat lung transplantation an exercise of trial and error. By standardizing the cuffing technique to use the smallest effective cuff appropriate for the size of the vessel or bronchus, one can make the transplantation procedure safer, faster, and more successful. Since diameters of the PV, PA, and Br are related to the body weight of the rat, we present a strategy to choosing an appropriate size using a weight-based guide. Since lung volume is also related to body weight, we recommend that this relationship should also be considered when choosing the proper volume of air for donor lung inflation during warm ischemia as well as for the proper volume of PBS to be instilled during bronchoalveolar lavage (BAL) fluid collection. We also describe methods for 4th intercostal space dissection, wound closure, and sample collection from both the native and transplanted lobes.

Here, we present optimizations to a rat lung transplantation model that serve to improve outcomes. We provide a size guide for cuffs based on body weight, a measurement strategy to ascertain the 4th intercostal space, and methods of wound closure and BAL (bronchoalveolar lavage) fluid and tissue collection.

Un modelo de trasplante de pulmón de rata de lesión por isquemia/reperfusión caliente: optimizaciones para mejorar los resultados

From our experience with rat lung transplantation, we have found several areas for improvement. Information in the existing literature regarding methods ...

An Effective Mouse Model of Unilateral Renal Ischemia-Reperfusion Injury

Ischemia-reperfusion injury (IRI) is the leading cause of acute renal failure and is a significant contributor to delayed graft function. Animal models are the only available resources that mimic the complexities of the IRI-associated damage encountered in vivo. This paper describes an effective mouse model of unilateral renal IRI that delivers highly reproducible data. Ischemia is induced by occluding the right renal pedicle for 30 min followed by reperfusion. In addition to the surgical procedure, a sequential overview of the expected physiological and histopathological changes following renal IRI will be provided by comparing data from seven different reperfusion times (4 h, 8 h, 16 h, 1 day, 2 days, 4 days, and 7 days). Critical data for planning experiments ahead, such as mean surgical time, average anesthetic consumption, and body weight changes over time, will be shared. This work will help researchers implement a reliable renal IRI model and select the appropriate reperfusion time that aligns with their intended investigative goals.

Renal ischemia-reperfusion injury is associated with high morbidity and mortality in hospitalized patients. Here, we present a simple and effective mouse model of unilateral renal ischemia-reperfusion injury and provide a sequential overview of representative pathological changes observed in the kidney.

Un modelo de ratón eficaz de lesión unilateral por isquemia-reperfusión renal

Ischemia-reperfusion injury (IRI) is the leading cause of acute renal failure and is a significant contributor to delayed graft function. Animal models ...

A Piglet Perinatal Asphyxia Model to Study Cardiac Injury and Hemodynamics after Cardiac Arrest, Resuscitation, and the Return of Spontaneous Circulation

Neonatal piglets have been extensively used as translational models for perinatal asphyxia. In 2007, we adapted a well-established piglet asphyxia model by introducing cardiac arrest. This enabled us to study the impact of severe asphyxia on key outcomes, including the time taken for the return of spontaneous circulation (ROSC), as well as the effect of chest compressions according to alternative protocols for cardiopulmonary resuscitation. Due to the anatomical and physiological similarities between piglets and human neonates, piglets serve as good models in studies of cardiopulmonary resuscitation and hemodynamic monitoring. In fact, this cardiac arrest model has provided evidence for guideline development through research on resuscitation protocols, pathophysiology, biomarkers, and novel methods for hemodynamic monitoring. Notably, the incidental finding that a substantial fraction of piglets have pulseless electrical activity (PEA) during cardiac arrest may increase the applicability of the model (i.e., it may be used to study pathophysiology extending beyond the perinatal period). However, the model generation is technically challenging and requires various skill sets, dedicated personnel, and a fine balance of the measures, including the surgical protocols and the use of sedatives/analgesics, to ensure a reasonable rate of survival. In this paper, the protocol is described in detail, as well as experiences with adaptations to the protocol over the years.

This piglet model involves surgical instrumentation, asphyxiation until the cardiac arrest, resuscitation, and post-resuscitation observation. The model allows for multiple sampling per animal, and by using continuous invasive arterial blood pressure, ECG, and non-invasive cardiac output monitoring, it provides knowledge about hemodynamics and cardiac pathophysiology in perinatal asphyxia and neonatal cardiopulmonary resuscitation.

Un modelo de asfixia perinatal de lechón para estudiar la lesión cardíaca y la hemodinámica después de un paro cardíaco, reanimación y el retorno de la circulación espontánea

Neonatal piglets have been extensively used as translational models for perinatal asphyxia. In 2007, we adapted a well-established piglet asphyxia model ...