XHTW es un WM Keck Foundation Distinguished Scholar jóvenes en la investigación médica, y también es apoyado por el NIH / NHLBI subvenciones R01-R01 y HL089598 R01HL091947, y Asociación de Distrofia Muscular de subvención # 69238. RJvO es el destinatario de la beca 2008-2010 American Physiological Society postdoctoral en genómica fisiológica. Este trabajo también es apoyado en parte por la Fundación Alianza Leducq de CaMKII en el Corazón de señalización.

Parte 1: Preparación del campo operatorio <ol><li> El campo de operación se desinfecta con un 75% de alcohol isopropílico. </li><li> Asegúrese de que el cojín de la calefacción está encendida y en la temperatura correcta. Un sistema recomendado es un Gaymar bomba de circulación de agua conectados a una plataforma de terapia que se mantiene a 37 ° C ± 1 ° C. Es importante mantener la temperatura corporal normal durante la cirugía para evitar una rápida disminución de la frecuencia cardíaca. </li><li> Los instrumentos quirúrgicos se esterilizan en un esterilizador de calor de cuentas antes de la cirugía. Para este procedimiento, tendrá los siguientes instrumentos quirúrgicos: tijeras de punta roma, por supuesto curva X 2 pinzas, pinzas finas en ángulo de 45 ° X 2, tijeras en ángulo de primavera, retractor en el pecho, y soporte de la aguja. </li><li> Aplicadores de algodón se debe a la mano en caso de hemorragia. </li></ol> Parte 2: Preparación y la intubación de los ratones <ol><li> Los ratones son anestesiados en una cámara de inducción con isoflurano 2% mezclado con 0.5 -1.0 L / min 100% de O 2. </li><li> Cortar el pelo se utilizan para afeitar la piel del escote a la altura del pecho medio. </li><li> El ratón es colocado en posición supina sobre un cojín de la calefacción con el fin de mantener la temperatura corporal. </li><li> Una banda de goma se coloca sobre los dientes delanteros del animal s para extender el cuello. El uso de pinzas curvas en una mano, la lengua es manipulado suavemente a un lado. Con la otra mano, la intubación endotraqueal se realiza con tubería de PE 90. El tubo endotraqueal se conecta a un volumen de Harvard de ciclo en ciclo del ventilador roedores 125-150 respiraciones / minuto y un volumen corriente de 0.1-0.3 ml. Durante la intervención quirúrgica, la anestesia se mantiene en 1,5 a 2% de isoflurano con 0,5 a 1,0 L / min 100% de O 2. Nivel correcto de la anestesia se verifica mediante la aplicación de presión sobre el lecho de la uña del ratón (dedo del pie-una pizca de reflejos). </li><li> El campo quirúrgico se desinfecta con una solución de betadine seguido por 70% de alcohol. Este procedimiento se repite tres veces. </li><li> Para evitar la contaminación del campo quirúrgico durante la operación, un campo estéril se coloca sobre el ratón dejando sólo el campo de operaciones al descubierto. </li><li> Un par de guantes estériles se utiliza para cada ratón individual. </li></ol> Parte 3: La ligadura de la aorta transversa <ol><li> Toracotomía parcial de la segunda costilla se realiza bajo un microscopio quirúrgico y el esternón se retractó de usar un separador en el pecho. </li><li> Punta fina pinza en ángulo de 45 ° se utilizan para separar suavemente el tejido del timo y la grasa del arco aórtico. </li><li> Tras la identificación de la aorta transversa, una pequeña pieza de una sutura de seda 6.0 se coloca entre las arterias carótidas innominada y la izquierda (Figura 1). </li><li> Dos nudos sueltos se atan alrededor de la aorta transversa y una pequeña pieza de un 27 ½ aguja roma de calibre se coloca paralelo a la aorta transversa. El primer nudo es atado rápidamente en contra de la aguja, seguida por la segunda y la aguja se eliminan sin demora con el fin de producir una contracción de 0,4 mm de diámetro. En los ratones control con tratamiento simulado, todo el procedimiento es idéntico, excepto para la ligadura de la aorta. </li><li> El retractor se quita el pecho y la salida del ventilador apagado apretado durante 2 segundos para volver a inflar los pulmones. </li><li> La caja torácica se cierra con una sutura de prolene 6.0 con un patrón de sutura interrumpida. </li><li> La piel se cierra con una sutura de prolene 6.0 con un patrón de sutura continua. </li></ol> Parte 4: recuperación post-operatoria <ol><li> Para la analgesia postoperatoria, el ratón se inyecta con buprenorfina (0,1 mg / kg) por vía intraperitoneal. Si hay signos de deshidratación después de la cirugía, se le da una solución salina estéril por vía intraperitoneal. </li><li> La anestesia es rebajarse gradualmente a la posición de apagado y el tubo endotraqueal eliminado cuando los signos de respiración espontánea se producen. </li><li> El ratón se mueve a la posición prona y les permitió recuperarse de una almohadilla eléctrica. </li></ol> Parte 5: Confirmación de la ligadura de éxito de la aorta transversa <ol><li> Una semana después de TAC, el ratón se re-anestesia para determinar el grado de sobrecarga de presión inducida por la ligadura de la aorta transversa. Como se describió anteriormente, la anestesia se mantiene en 1,5 a 2% de isoflurano con 0,5 a 1,0 L / min 100% de O2 y la temperatura corporal se mantuvo a 37 ° C ± 1 ° C. </li><li> A 20 MHz Doppler sonda se coloca en los lados izquierdo y derecho del cuello en un ángulo de 45 ° para detectar velocidades de flujo. Un ordenador basado en procesador de señales Doppler (Indo Instruments, Houston, TX) se utiliza para mostrar y almacenar señales de Doppler. 5 </li><li> Dependiendo del grado de sobrecarga de presión necesario para el protocolo experimental, los ratones sólo con una carótida derecha (RC) / carótida izquierda (LC) la relación de flujo dentro de cierto rango se incluyen para su posterior análisis. Por ejemplo, un moderado grado de sobrecarga de presión lleva a una proporción de 5-8, mientras que una contracción más estrecha que resulta en una sobrecarga de presión severa lleva tOA cociente de 8.10 (Figura 5.3A). Un animal simulado (operado, pero no ligado), sin embargo, se espera que tenga una proporción de ~ 1 (Figura 5.3b). </li></ol> 6: Resultados de Representante La supervivencia típica quirúrgica después de la TAC en ratones de tipo salvaje es alrededor de 80-90%. La ligadura quirúrgica exitosa de la aorta transversa dará lugar a una relación de velocidad Doppler del flujo entre la arteria carótida derecha y la izquierda (RC / LC) de 10.5 (Figura 1). En comparación con el simulacro que funciona con los ratones (Figura 2A), los ratones con sobrecarga de presión se espera que se desarrolle la hipertrofia cardiaca en 1-2 semanas, y la dilatación cardiaca después de 6-8 semanas, dependiendo de la tensión de la contracción (Figura 2B). La mortalidad durante el primer mes después de la TAC es más baja (&lt;20%) en ratones de tipo salvaje, a pesar de ratones modificados genéticamente pueden presentar diferentes tasas de supervivencia 6,7. <img src="/files/ftp_upload/1729/1729fig1tmb.jpg" alt="Figura 1" border="0" /> Figura 1. La confirmación de la ligadura de la aorta transversal con Doppler de medición de velocidad. Representante de las señales de velocidad Doppler de la derecha (CD) y la arteria carótida izquierda (LC) 6 días después de la TAC. Estas grabaciones demuestran una ligadura de éxito con un ratio de flujo de RC / LC de ~ 6.8. Por favor, <a href="https://www.jove.com/files/ftp_upload/1729/1729fig1.jpg">haga clic aquí</a> para ver una versión ampliada de la figura 1. <img src="/files/ftp_upload/1729/1729fig2.jpg" alt="Figura 2" /> Figura 2. Imágenes representativas de todo corazón Después de TAC. A, B. Sham-operado del corazón salvaje del ratón tipo.. Corazón salvaje del ratón tipo 16 semanas después de la TAC. Cada línea = 1 mm.

<ol>
	<li>Rockman, H. A., Ross, R. S., Harris, A. N., Knowlton, K. U., Steinhelper, M. E., Field, L. J., Ross, J., Chein, K. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Segregation+of+atrial-specific+and+inducible+expression+of+an+atrial+natriuretic+factor+transgene+in+an+in+vivo+murine+model+of+cardiac+hypertrophy.">Segregation of atrial-specific and inducible expression of an atrial natriuretic factor transgene in an in vivo murine model of cardiac hypertrophy.</a> Proc. Natl. Acad. Sci. USA. 88, 8277-8281 (1991).</li><li>Heineke, J., Molkentin, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Regulation+of+cardiac+hypertrophy+by+intracellular+signaling+pathways.">Regulation of cardiac hypertrophy by intracellular signaling pathways.</a> Nat. Rev. Mol. Cell. Biol. 7, 589-600 (2006).</li><li>Hartley, C. J., Reddy, A. K., Madala, S., Michael, L. H., Entman, M. L., Taffett, G. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Doppler+estimation+of+reduced+coronary+flow+reserve+in+mice+with+pressure+overload+cardiac+hypertrophy.">Doppler estimation of reduced coronary flow reserve in mice with pressure overload cardiac hypertrophy.</a> Ultrasound Med. Biol. 34, 892-901 (2008).</li><li>Reddy, A. K., Taffett, G. E., Li, Y. -. H., Lim, S. -. W., Pham, T. T., Pocius, J. S., Entman, M. L., Michael, L. H., Hartley, C. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Doppler+signal+processing+for+use+in+mice:+applications.">Doppler signal processing for use in mice: applications.</a> IEEE Trans. Biomed. Eng. 52, 1771-1783 (2005).</li><li>Li, Y. -. H., Reddy, A. K., Ochoa, L. N., Pham, T. T., Hartley, C. J., Micheal, L. H., Entman, M. L., Taffett, G. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+age+on+peripheral+vascular+response+to+transverse+aortic+banding+in+mice.">Effect of age on peripheral vascular response to transverse aortic banding in mice.</a> J. Gerontol. 58A, 895-899 (2003).</li><li>Bourajjaj, M., Armand, A. S., Martins, d. a. C. o. s. t. a., Weijts, P. A., Nagel, B. V. a. n. d. e. r., Heeneman, R., Wehrens, S., Windt, X. H. D. e., J, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NFATc2+is+a+necessary+mediator+of+calcineurin-dependent+cardiac+hypertrophy+and+heart+failure.">NFATc2 is a necessary mediator of calcineurin-dependent cardiac hypertrophy and heart failure.</a> J. Biol. Chem. 28, 22295-22303 (2008).</li><li>Oort, R. J. v. a. n., Respress, J. L., Li, N., Reynolds, C., Almeida, A. C. D. e., Skapura, D. G., Windt, L. J. D. e., Wehrens, X. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Accelerated+development+of+pressure+overload+induced+cardiac+hypertrophy+and+dysfunction+in+an+RyR2-R176Q+knockin+mouse+model.">Accelerated development of pressure overload induced cardiac hypertrophy and dysfunction in an RyR2-R176Q knockin mouse model.</a> Hypertension. 55, (2010).</li><li>Tarnavski, O., McMullen, J. R., Schinke, M., Nie, Q., Kong, S., Izumo, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+cardiac+surgery:+comprehensive+techniques+for+the+generation+of+mouse+models+of+human+diseases+and+their+application+for+genomic+studies.">Mouse cardiac surgery: comprehensive techniques for the generation of mouse models of human diseases and their application for genomic studies.</a> Physiol Genomics. 16, 349-360 (2004).</li></ol>

TAC, que imita a la estenosis aórtica humana, es un método común para inducir la hipertrofia e insuficiencia cardiaca en ratones. Sitios alternativos para la constricción de la aorta incluyen la aorta ascendente y abdominal. Constricción de la aorta ascendente proporciona una sobrecarga extrema y más rápido en el ventrículo izquierdo (VI). Por el contrario, la constricción de la aorta abdominal deja intacta una porción más grande de la circulación como medio de compensación posible. 8 Por lo tant...

<!DOCTYPE html PUBLIC "-//W3C//DTD XHTML 1.0 Transitional//EN" "http://www.w3.org/TR/xhtml1/DTD/xhtml1-transitional.dtd">
<html xmlns="http://www.w3.org/1999/xhtml" xml:lang="en" lang="en">	
		
		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Gaymar T/pump</td>
<td>&nbsp;</td>
<td>Harvard Apparatus</td>
<td>Model TP-500</td>
<td>Circulating water pump</td>
</tr>
<tr>
<td>Temp. therapy pads</td>
<td>&nbsp;</td>
<td>Harvard Apparatus</td>
<td>60-3414</td>
<td>&nbsp;</td>
<tr>
<td>Hot bead sterilizer</td>
<td>&nbsp;</td>
<td>Fine Science Tools</td>
<td>18000-45</td>
<td>&nbsp;</td>
<tr>
<td>Blunt scissors</td>
<td>&nbsp;</td>
<td>Roboz</td>
<td>RS-5980</td>
<td>&nbsp;</td>
<tr>
<td>Angled spring scissors</td>
<td>&nbsp;</td>
<td>Roboz</td>
<td>RS-5668</td>
<td>&nbsp;</td>
<tr>
<td>Chest retractor</td>
<td>&nbsp;</td>
<td>Fine Science Tools</td>
<td>17002-02</td>
<td>&nbsp;</td>
<tr>
<td>Course curved forceps</td>
<td>&nbsp;</td>
<td>Roboz</td>
<td>RS-5138</td>
<td>&nbsp;</td>
<tr>
<td>Dumont fine 45&deg; angled forceps</td>
<td>&nbsp;</td>
<td>Fine Science Tools</td>
<td>11253-25</td>
<td>&nbsp;</td>
<tr>
<td>Needle holder</td>
<td>&nbsp;</td>
<td>Fine Science Tools</td>
<td>12565-14</td>
<td>&nbsp;</td>
<tr>
<td>Electric shaver</td>
<td>&nbsp;</td>
<td>General Electric</td>
<td>&nbsp;</td>
<td>For shaving mouse fur</td>
</tr>
<tr>
<td>PE 90 tubing</td>
<td>&nbsp;</td>
<td>BD</td>
<td>427420</td>
<td>For intubation</td>
</tr>
<tr>
<td>MiniVent (ventilator)</td>
<td>&nbsp;</td>
<td>Harvard Apparatus</td>
<td>Type 845</td>
<td>&nbsp;</td>
<tr>
<td>Betadine</td>
<td>&nbsp;</td>
<td>Fisher</td>
<td>19-027132</td>
<td>May be purchased at medical supply store</td>
</tr>
<tr>
<td>Sterile gloves</td>
<td>&nbsp;</td>
<td>Mckesson</td>
<td>20-1565</td>
<td>&nbsp;</td>
<tr>
<td>Stereo microscope</td>
<td>&nbsp;</td>
<td>Unico</td>
<td>ZM186</td>
<td>&nbsp;</td>
<tr>
<td>6-0 silk suture</td>
<td>&nbsp;</td>
<td>Fine Science Tools</td>
<td>18020-60</td>
<td>&nbsp;</td>
<tr>
<td>27 &frac12; gauge needle</td>
<td>&nbsp;</td>
<td>BD</td>
<td>305109</td>
<td>&nbsp;</td>
<tr>
<td>6-0 prolene suture</td>
<td>&nbsp;</td>
<td>Mckesson</td>
<td>3286</td>
<td>&nbsp;</td>
<tr>
<td>Buprenorphine</td>
<td>&nbsp;</td>
<td>Institutional Animal Facility</td>
<td>&nbsp;</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

transverse aortic constriction in mice

Constricción de la aorta transversal (TAC) en el ratón es un modelo experimental de uso común para la presión inducida por sobrecarga de la hipertrofia cardíaca y la insuficiencia cardiaca.<sup&gt; 1</sup&gt; TAC inicialmente conduce a la hipertrofia compensada del corazón, que a menudo se asocia con un aumento temporal de la contractilidad cardíaca. Con el tiempo, sin embargo, la respuesta a la sobrecarga hemodinámica crónica se convierte en mala adaptación, lo que resulta en la dilatación cardíaca y la insuficiencia cardiaca.<sup&gt; 2</sup&gt; El modelo murino TAC fue validado por primera vez por Rockman<em&gt; Et al</em&gt;.<sup&gt; 1</sup&gt;, Y desde entonces ha sido ampliamente utilizado como una herramienta valiosa para imitar humanos las enfermedades cardiovasculares y dilucidar los procesos fundamentales de señalización implicadas en la respuesta hipertrófica y el desarrollo de insuficiencia cardíaca. En comparación con otros modelos experimentales de insuficiencia cardíaca, como la oclusión completa de la arteria descendente anterior (LAD) arteria coronaria, TAC proporciona un modelo más reproducible de la hipertrofia cardíaca y un curso de tiempo más gradual en el desarrollo de insuficiencia cardíaca. Aquí se describe un procedimiento paso a paso para realizar cirugía TAC en ratones. Para determinar el nivel de sobrecarga de presión producida por la ligadura de la aorta, una alta frecuencia de sonda Doppler se utiliza para medir la relación entre las velocidades del flujo sanguíneo en las arterias carótidas derecha e izquierda.<sup&gt; 3, 4</sup&gt; Con las tasas de supervivencia quirúrgica del 80-90%, aórtica transversal de bandas es una técnica eficaz para inducir la hipertrofia ventricular izquierda e insuficiencia cardiaca en ratones.

Watch this Scientific Journal Video about Transverse Aortic Constriction in Mice at JoVE.com

Transverse Aortic Constriction in Mice

Constricción de la aorta transversal (TAC) en el ratón es un modelo de uso experimental para estudiar mecanismos que subyacen a la hipertrofia cardíaca y el desarrollo de insuficiencia cardíaca. Aquí se describen los procedimientos para contraer la aorta para crear un grado de reproducibilidad de la hipertrofia cardiaca en ratones.

Constricción de la aorta transversa en ratones

Transverse aortic constriction (TAC) in the mouse is a commonly used experimental model for pressure overload-induced cardiac hypertrophy and heart ...

Research

JoVE Journal

Biology

66.5K vistas.  Baylor College of Medicine (BCM). Constricción de la aorta transversal (TAC) en el ratón es un modelo de uso experimental para estudiar mecanismos que subyacen a la hipertrofia cardíaca y el desarrollo de insuficiencia cardíaca. Aquí se describen los procedimientos para contraer la aorta para crear un grado de reproducibilidad de la hipertrofia cardiaca en ratones.

Video: Transverse Aortic Constriction in Mice

Aortic Ring Assay

Angiogenesis, the sprouting of blood vessels from preexisting vasculature is associated with both natural and pathological processes. Various angiogenesis assays involve the study of individual endothelial cells in culture conditions (1). The aortic ring assay is an angiogenesis model that is based on organ culture. In this assay, angiogenic vessels grow from a segment of the aorta (modified from (2)). Briefly, mouse thoracic aorta is excised, the fat layer and adventitia are removed, and rings approximately 1 mm in length are prepared. Individual rings are then embedded in a small solid dome of basement matrix extract (BME), cast inside individual wells of a 48-well plate. Angiogenic factors and inhibitors of angiogenesis can be directly added to the rings, and a mixed co-culture of aortic rings and other cell types can be employed for the study of paracrine angiogenic effects. Sprouting is observed by inspection under a stereomicroscope over a period of 6-12 days. Due to the large variation caused by the irregularities in the aortic segments, experimentation in 6-plicates is strongly advised. Neovessel outgrowth is monitored throughout the experiment and imaged using phase microscopy, and supernatants are collected for measurement of relevant angiogenic and anti-angiogenic factors, cell death markers and nitrite.

Angiogenesis, the sprouting of blood vessels from pre-existing vasculature, is associated with both natural and pathological processes. Here we demonstrate an aortic ring assay that allows angiogenic potentiators and inhibitors to be directly added to aortic rings in culture. Sprouting and neovessel outgrowth can be determined by inspecting the aortic rings over a period of 6-12 days.

Ensayo del anillo aórtico

Angiogenesis, the sprouting of blood vessels from preexisting vasculature is associated with both natural and pathological processes. Various angiogenesis ...

Investigación

Biología

Isolation and Primary Culture of Mouse Aortic Endothelial Cells

The vascular endothelium is essential to normal vascular homeostasis. Its dysfunction participates in various cardiovascular disorders. The mouse is an important model for cardiovascular disease research. This study demonstrates a simple method to isolate and culture endothelial cells from the mouse aorta without any special equipment. To isolate endothelial cells, the thoracic aorta is quickly removed from the mouse body, and the attached adipose tissue and connective tissue are removed from the aorta. The aorta is cut into 1 mm rings. Each aortic ring is opened and seeded onto a growth factor reduced matrix with the endothelium facing down. The segments are cultured in endothelial cell growth medium for about 4 days. The endothelial sprouting starts as early as day 2. The segments are then removed and the cells are cultured continually until they reach confluence. The endothelial cells are harvested using neutral proteinase and cultured in endothelial cell growth medium for another two passages before being used for experiments. Immunofluorescence staining indicated that after the second passage the majority of cells were double positive for Dil-ac-LDL uptake, Lectin binding, and CD31 staining, the typical characteristics of endothelial cells. It is suggested that cells at the second to third passages are suitable for in vitro and in vivo experiments to study the endothelial biology. Our protocol provides an effective means of identifying specific cellular and molecular mechanisms in endothelial cell physiopathology.

The vascular endothelial cells play a significant role in many important cardiovascular disorders. This article describes a simple method to isolate and expand endothelial cells from the mouse aorta without using any special equipment. Our protocol provides an effective means of identifying mechanisms in endothelial cell physiopathology.

Aislamiento y cultivo primario de células endoteliales aórticas de ratón

The vascular endothelium is essential to normal vascular homeostasis. Its dysfunction participates in various cardiovascular disorders. The mouse is an ...

Induction of Right Ventricular Failure by Pulmonary Artery Constriction and Evaluation of Right Ventricular Function in Mice

The mechanism of right ventricular failure (RVF) requires clarification due to the uniqueness, high morbidity, high mortality, and refractory nature of RVF. Previous rat models imitating RVF progression have been described. Compared with rats, mice are more accessible, economical, and widely used in animal experiments. We developed a pulmonary artery constriction (PAC) approach which is comprised of banding the pulmonary trunk in mice to induce right ventricular (RV) hypertrophy. A special surgical latch needle was designed that allows for easier separation of the aorta and the pulmonary trunk. In our experiments, the use of this fabricated latch needle reduced the risk of arteriorrhexis and improved the surgical success rate to 90%. We used different padding needle diameters to precisely create quantitative constriction, which can induce different degrees of RV hypertrophy. We quantified the degree of constriction by evaluating the blood flow velocity of the PA, which was measured by noninvasive transthoracic echocardiography. RV function was precisely evaluated by right heart catheterization at 8 weeks after surgery. The surgical instruments made inhouse were composed of common materials using a simple process that is easy to master. Therefore, the PAC approach described here is easy to imitate using instruments made in the lab and can be widely used in other labs. This study presents a modified PAC approach that has a higher success rate than other models and an 8-week postsurgery survival rate of 97.8%. This PAC approach provides a useful technique for studying the mechanism of RVF and will enable an increased understanding of RVF.

Here, we provide a useful approach for studying the mechanism of right ventricular failure. A more convenient and efficient approach to pulmonary artery constriction is established using surgical instruments made inhouse. In addition, methods to evaluate the quality of this approach by echocardiography and catheterization are provided.

Inducción de la insuficiencia ventricular derecha por constricción pulmonar de la arteria y evaluación de la función ventricular derecha en ratones

The mechanism of right ventricular failure (RVF) requires clarification due to the uniqueness, high morbidity, high mortality, and refractory nature of ...

Transverse Fracture of the Mouse Femur with Stabilizing Pin

Fracture repair is an essential function of the skeleton that cannot be reliably modeled in vitro. A mouse injury model is an efficient approach to test whether a gene, gene product or drug influences bone repair because murine bones recapitulate the stages observed during human fracture healing. When a mouse or human breaks a bone, an inflammatory response is initiated, and the periosteum, a stem cell niche surrounding the bone itself, is activated and expands. Cells residing in the periosteum then differentiate to form a vascularized soft callus. The transition from the soft callus to a hard callus occurs as the recruited skeletal progenitor cells differentiate into mineralizing cells, and the bridging of the fractured ends results in the bone union. The mineralized callus then undergoes remodeling to restore the original shape and structure of the healed bone. Fracture healing has been studied in mice using various injury models. Still, the best way to recapitulate this entire biological process is to break through the cross-section of a long bone that encompasses both cortices. This protocol describes how a stabilized, transverse femur fracture can be safely performed to assess healing in adult mice. A surgical protocol including detailed harvesting and imaging techniques to characterize the different stages of fracture healing is also provided.

This protocol describes a method to perform fractures on adult mice and monitor the healing process.

Fractura transversal del fémur del ratón con pasador estabilizador

Fracture repair is an essential function of the skeleton that cannot be reliably modeled in vitro. A mouse injury model is an efficient approach to test ...

A Modified Technique for Transverse Aortic Constriction in Mice

Transverse aortic constriction (TAC) is a frequently used surgery in research regarding heart failure and cardiac hypertrophy based on the formation of pressure overload in mouse models. The main challenge of this procedure is to clearly visualize the transverse aortic arch and precisely band the target vessel. Classical approaches perform a partial thoracotomy to expose the transverse aortic arch. However, it is an open-chest model that causes a rather large surgical trauma and requires a ventilator during the surgery. To prevent unnecessary trauma and simplify the operating procedure, the aortic arch is approached via the proximal proportion of the sternum, reaching and binding the target vessel using a small self-made retractor that contains a snare. This procedure can be conducted without entering the pleura cavity and does not need a ventilator or microsurgical operation, which leaves the mice with physiological breathing patterns, simplifies the procedure, and significantly reduces operation time. Due to the less invasive approach and less operation time, mice can undergo fewer stress reactions and recover rapidly.

The present protocol describes a modified and simplified technique with a minimally invasive transverse aortic constriction (TAC) procedure using a self-made retractor. This procedure can be conducted without a ventilator or microscope and introduces pressure overload, eventually leading to cardiac hypertrophy or heart failure.

Una técnica modificada para la constricción aórtica transversa en ratones

Transverse aortic constriction (TAC) is a frequently used surgery in research regarding heart failure and cardiac hypertrophy based on the formation of ...

A Calcium Phosphate-Induced Mouse Abdominal Aortic Aneurysm Model

An abdominal aortic aneurysm (AAA) is a life-threatening cardiovascular disease that occurs worldwide and is characterized by irreversible dilation of the abdominal aorta. Currently, several chemically induced murine AAA models are used, each simulating a different aspect of the pathogenesis of AAA. The calcium phosphate-induced AAA model is a rapid and cost-effective model compared to the angiotensin II- and elastase-induced AAA models. The application of CaPO4 crystals to the mouse aorta results in elastic fiber degradation, loss of smooth muscle cells, inflammation, and calcium deposition associated with aortic dilation. This article introduces a standard protocol for the CaPO4-induced AAA model. The protocol includes material preparation, the surgical application of the CaPO4&#160;to the adventitia of the infrarenal abdominal aorta, the harvesting of aortas to visualize aortic aneurysms, and histological analyses in mice.

This protocol describes a calcium phosphate-induced abdominal aortic aneurysm (AAA) mouse model to study the pathological features and molecular mechanisms of AAAs.

Un modelo de aneurisma aórtico abdominal de ratón inducido por fosfato de calcio

An abdominal aortic aneurysm (AAA) is a life-threatening cardiovascular disease that occurs worldwide and is characterized by irreversible dilation of the ...

Establishing a Diaphyseal Femur Fracture Model in Mice

Bones have a significant regenerative capacity. However, fracture healing is a complex process, and depending on the severity of the lesions and the age and overall health status of the patient, failures can occur, leading to delayed union or nonunion. Due to the increasing number of fractures resulting from high-energy trauma and aging, the development of innovative therapeutic strategies to improve bone repair based on the combination of skeletal/mesenchymal stem/stromal cells and biomimetic biomaterials is urgently needed. To this end, the use of reliable animal models is fundamental to better understanding the key cellular and molecular mechanisms that determine the healing outcomes. Of all the models, the mouse is the preferred research model because it offers a wide variety of transgenic strains and reagents for experimental analysis. However, the establishment of fractures in mice may be technically challenging due to their small size. Therefore, this article aims to demonstrate the procedures for the surgical establishment of a diaphyseal femur fracture in mice, which is stabilized with an intramedullary wire and resembles the most common bone repair process, through cartilaginous callus formation.

This protocol describes a surgical procedure for the establishment of a diaphyseal fracture in the femur of mice, which is stabilized with an intramedullary wire, for fracture healing studies.

Establecimiento de un modelo de fractura de fémur diafisario en ratones

Bones have a significant regenerative capacity. However, fracture healing is a complex process, and depending on the severity of the lesions and the age ...

Métodos mejorados para la obtención de imágenes de Primordia de hoja de maíz

Improved Methods for Preparing Transverse Sections and Unrolled Whole Mounts of Maize Leaf Primordia for Fluorescence and Confocal Imaging

In maize (Zea mays) and other grasses (Poaceae), the leaf primordia are deeply ensheathed and rolled within the leaf whorl, making it difficult to study early leaf development. Here, we describe methods for preparing transverse sections and unrolled whole mounts of maize leaf primordia for fluorescence and confocal imaging. The first method uses a wire stripper to remove the upper portions of older leaves, exposing the tip of the leaf primordium and allowing its measurement for more accurate transverse section sampling. The second method uses clear, double-sided nano tape to unroll and mount whole-leaf primordia for imaging. We show the utility of the two methods in visualizing and analyzing fluorescent protein reporters in maize. These methods provide a solution to the challenges presented by the distinctive morphology of maize leaf primordia and will be useful for visualizing and quantifying leaf anatomical and developmental traits in maize and other grass species.

Autor destacado: Métodos mejorados para preparar secciones transversales y desenrollar montajes enteros de Primordia de hoja de maíz para imágenes de fluorescencia y confocales  

Maize leaf primordia are deeply ensheathed and rolled, making them difficult to study. Here, we present methods for preparing transverse sections and unrolled whole mounts of maize leaf primordia for fluorescence and confocal imaging.

Métodos mejorados para preparar secciones transversales y montajes enteros desenrollados de primordios de hojas de maíz para imágenes de fluorescencia y confocales

In maize (Zea mays) and other grasses (Poaceae), the leaf primordia are deeply ensheathed and rolled within the leaf whorl, making it difficult to study ...