Los autores desean agradecer el apoyo técnico de la Alegría Yao y el Dr. Karen Alt, así como la financiación de la NHMRC y NHF.

Todos los experimentos con animales fueron aprobados por el Medical Research Alfred y Comité de Educación del Distrito Electoral Animal Ética (E / 1534/2015 / B). Todas las manipulaciones quirúrgicas se realizaron bajo anestesia total y los animales no experimentan dolor en cualquier etapa. Todos los experimentos descritos son la no recuperación. 1. Preparación <ol><li> Cortar bandas delgadas de papel de filtro (1 mm x 2 mm). </li><li> Recién preparar 2 soluciones de cloruro férrico de 4% (w / v) y 6% (w / v) diluido en agua desionizada. Prepare la solución de rodamina 6G 0,3% en PBS, se filtra a través de 0,22 micras. </li><li> Corte un pedazo pequeño (5 mm x 1 cm) de la envoltura de plástico blanco de jeringa. </li></ol> 2. Mesenterio Arteriola formación de trombos Observado por Microscopia intravital <ol><li> Pesar un 10-12 semanas de edad C57BL / 6 de tipo salvaje del ratón y en consecuencia anestesiar con una mezcla de ketamina (100 mg / kg) y xilazina (10 mg / kg), aunque la inyección intraperitoneal. Supervisar la profundidad de la anestesia por la respuesta a los pies, la cola y / o pellizco de piel, la reactividad de pellizco córnea y los párpados, y la ausencia de movimiento bigotes. Si es necesario, inyectar una segunda dosis de ketamina (50 mg / kg) para mantener la anestesia del animal. Aplique un ungüento veterinario en los ojos para evitar la sequedad mientras que bajo anestesia. Coloca el ratón en una pequeña placa de Petri colocado sobre una almohadilla eléctrica ajustada a 37 ° C. NOTA: Aunque la inyección IP puede resultar en dolor para los animales, la duración de este malestar será mínimo (menos de 3 segundos). El dolor y las molestias asociadas con las inyecciones se reducirán al mínimo mediante el uso de personal experimentado y competente y tamaño de la aguja apropiada (25 G). Después de todos los procedimientos, la eutanasia de todos los animales utilizando una sobredosis de ketamina y xilazina seguido por dislocación cervical. </li><li> Realizar una incisión en la línea media abdominal de aproximadamente 3 cm en la piel y cortar cuidadosamente el peritoneo. </li><li> Coloque el ratón en el lado de la mascotaplato ri, exteriorizar suavemente sus intestinos y se extendió cuidadosamente el mesenterio con 2 bastoncillos de algodón para llevar un recipiente adecuado a la superficie de la placa de Petri. Seque bien con un limpiador delicado. NOTA: Para limitar el movimiento de los vasos mesentéricos, papaverina puede usarse para inhibir la peristalsis intestinal. </li><li> Remojar la cola del ratón en agua caliente para dilatar los vasos e inyectar 30 l de Rodamina 6G (0,3%) en la vena de la cola del ratón con una jeringa de 29 G para etiquetar los leucocitos y las plaquetas. </li><li> Coloque la placa de Petri bajo un microscopio invertido y se centran en la arteriola elegido utilizando el canal de campo claro. </li><li> Remojar una banda de papel de filtro con 6% (w / v) de cloruro de hierro (III) y aplicar el papel de filtro en la arteriola con dos pinzas; la primera en celebrar el papel de filtro, el segundo para presionar suavemente sobre el área de interés. Observar la formación de trombos en el primero 10 seg después de la deposición del papel de filtro. NOTA: Es muy commpor lo tanto, a dañar la microvasculatura rodeado y la precisión de la deposición del papel de filtro y la presión suavemente es importante limitar este problema. </li><li> Observar la formación de trombos por microscopía de fluorescencia (canal TRITC: excitación pico de 557 nm, pico de emisión 576 nm), a través del papel de filtro. Por lo tanto, observar los leucocitos y las plaquetas circulantes que han tenido hasta la Rodamina 6G y su agregación en el trombo es fácil de identificar. </li><li> Toma el papel de filtro después de 1 min de exposición al cloruro de hierro (III) y continuará supervisando la formación del trombo. Lave el recipiente con PBS. </li><li> Observar y registrar la formación dinámica del trombo se destaca con el etiquetado de las plaquetas y leucocitos con la Rodamina 6G. Capturar las imágenes y medir el tamaño del trombo. Las imágenes que aquí se presenta se obtuvieron con un microscopio intravital invertida, aunque un objetivo 4X, en el canal de fluorescencia TRITC. </li><li> Fespués de todos los procedimientos, la eutanasia del animal utilizando una sobredosis de ketamina y xilazina seguido por dislocación cervical. </li></ol> 3. carótida trombo Arteria Formación evaluados por Sangre Medición de Velocidad de Flujo <ol><li> Pesar un 10-12 semanas de edad C57BL / 6 de ratón y en consecuencia anestesiar con una mezcla de ketamina (100 mg / kg) y xilazina (10 mg / kg), aunque la inyección intraperitoneal. Aplique un ungüento veterinario en los ojos para evitar la sequedad mientras que bajo anestesia. </li><li> Fijar el ratón bajo un microscopio quirúrgico con cinta adhesiva en las piernas, en una almohadilla eléctrica ajustada a 37 ° C. </li><li> Hacer una pequeña almohada de un limpiador y la cinta bajo la cabeza del ratón para elevar la cabeza ligeramente. Utilice un bucle de hilo con pinzas para sacar el hocico hacia abajo (utilizar dientes superiores). Esto expondrá la región de la arteria carótida para un fácil acceso. </li><li> Realizar una pequeña 5 mm de profundidad de la incisión de la piel directamente debajo de la mandíbula, hasta el esternón. </li><li> Diseccionar la fascia y aislar un fragmento de cualquiera de la arteria carótida común izquierda o hacia la derecha por encima de la bifurcación. </li><li> Introducir cuidadosamente pinzas en-entre la arteria y el nervio para separarlos. No molestar el nervio corriendo cerca de la arteria y evitar tocar demasiado de la arteria carótida como fuere causa daños a la embarcación. Aislar una sección de un menos 5 mm de la arteria. </li><li> Seque el área de la arteria correctamente con limpiaparabrisas para evitar que cualquier líquido interfiere con el FeCl3. </li><li> Ponga la pequeña pieza de plástico blanco debajo de la parte aislada de la carótida común por lo que el FeCl3 no sumergirse en los tejidos circundantes. Para este propósito, utilizar un segundo fórceps para llevar la pieza a la primera uno luego lentamente deslice el papel de plástico debajo de la arteria. </li><li> Remojar un trozo de papel de filtro con 4% (w / v) o 6% (w / v) de cloruro férrico y lo coloca todo alrededor de la arteria. </li><li> Después de 3 minutos de exposición, despegar el papel de filtro, enjuagar con PBS y seque el área wlimpiaparabrisas i. </li><li> Coloque la sonda de flujo Doppler en todo el buque en la zona lesionada y empezar a grabar los cambios en el flujo. En la carótida común saludable de ratones adultos, el flujo es por lo general alrededor de 1 ml / min. ¡Precaución! el contacto de la sonda con cloruro férrico dañará la sonda por lo que cualquier contacto debe evitarse. Los datos presentados en este documento se obtuvieron con un metro Transonic System Inc. Flow, TS420 módulo perivascular equipado con una sonda de flujo Doppler Nano 0.5 PBS. NOTA: La concentración de cloruro férrico puede modificar la cinética de la formación de trombos resultantes en diferentes momentos de oclusión. Por lo tanto, la exposición a un 6% (w / v) de cloruro férrico da una oclusión más rápida que la exposición al 4% (w / v) de cloruro férrico. </li><li> Después de todos los procedimientos, la eutanasia de todos los animales mediante una sobredosis de ketamina y xilazina seguido por dislocación cervical y limpiar cuidadosamente la sonda Doppler. </li></ol>

<ol>
	<li>Leadley, R. J., Chi, L., Rebello, S. S., Gagnon, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contribution+of+in+vivo+models+of+thrombosis+to+the+discovery+and+development+of+novel+antithrombotic+agents.">Contribution of in vivo models of thrombosis to the discovery and development of novel antithrombotic agents.</a> J Pharmacol Toxicol Methods. 43 (2), 101-116 (2000).</li><li>Johnson, G. J., Griggs, T. R., Badimon, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+utility+of+animal+models+in+the+preclinical+study+of+interventions+to+prevent+human+coronary+artery+restenosis:+analysis+and+recommendations.+On+behalf+of+the+Subcommittee+on+Animal,+Cellular+and+Molecular+Models+of+Thrombosis+and+Haemostasis+of+the+Scientific+and+Standardization+Committee+of+the+International+Society+on+Thrombosis+and+Haemostasis.">The utility of animal models in the preclinical study of interventions to prevent human coronary artery restenosis: analysis and recommendations. On behalf of the Subcommittee on Animal, Cellular and Molecular Models of Thrombosis and Haemostasis of the Scientific and Standardization Committee of the International Society on Thrombosis and Haemostasis.</a> Thromb Haemost. 81 (5), 835-843 (1999).</li><li>Day, S. M., Reeve, J. L., Myers, D. D., Fay, W. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Murine+thrombosis+models.">Murine thrombosis models.</a> Thromb Haemost. 92 (3), 486-494 (2004).</li><li>Sachs, U. J. H., Nieswandt, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+thrombus+formation+in+murine+models.">In vivo thrombus formation in murine models.</a> Circ Res. 100 (7), 979-991 (2007).</li><li>Furie, B., Furie, B. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms+of+thrombus+formation.">Mechanisms of thrombus formation.</a> N Engl J Med. 359 (9), 938-949 (2008).</li><li>Pierangeli, S. S., Liu, X. W., Barker, J. H., Anderson, G., Harris, E. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Induction+of+thrombosis+in+a+mouse+model+by+IgG,+IgM+and+IgA+immunoglobulins+from+patients+with+the+antiphospholipid+syndrome.">Induction of thrombosis in a mouse model by IgG, IgM and IgA immunoglobulins from patients with the antiphospholipid syndrome.</a> Thromb Haemost. 74 (5), 1361-1367 (1995).</li><li>Kikuchi, S., Umemura, K., Kondo, K., Saniabadi, A. R., Nakashima, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Photochemically+induced+endothelial+injury+in+the+mouse+as+a+screening+model+for+inhibitors+of+vascular+intimal+thickening.">Photochemically induced endothelial injury in the mouse as a screening model for inhibitors of vascular intimal thickening.</a> Arterioscler Thromb Vasc Biol. 18 (7), 1069-1078 (1998).</li><li>Rosen, E. D., Raymond, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Laser-induced+noninvasive+vascular+injury+models+in+mice+generate+platelet-+and+coagulation-dependent+thrombi.">Laser-induced noninvasive vascular injury models in mice generate platelet- and coagulation-dependent thrombi.</a> Am J Pathol. 158, 1613-1622 (2001).</li><li>Kurz, K. D., Main, B. W., Sandusky, 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=Rat+model+of+arterial+thrombosis+induced+by+ferric+chloride.">Rat model of arterial thrombosis induced by ferric chloride.</a> Thromb Res. 60 (4), 269-280 (1990).</li><li>Eckly, A., Hechler, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms+underlying+FeCl3-induced+arterial+thrombosis.">Mechanisms underlying FeCl3-induced arterial thrombosis.</a> J Thromb Haemost. 9 (4), 779-789 (2011).</li><li>Darbousset, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Involvement+of+neutrophils+in+thrombus+formation+in+living+mice.">Involvement of neutrophils in thrombus formation in living mice.</a> Pathol Biol (Paris). 62 (1), 1-9 (2014).</li><li>Denis, C., Methia, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+mouse+model+of+severe+von+Willebrand+disease:+defects+in+hemostasis+and+thrombosis).">A mouse model of severe von Willebrand disease: defects in hemostasis and thrombosis).</a> Proc Natl Acad Sci U S A. 95 (16), 9524-9529 (1998).</li><li>Wang, X., Hagemeyer, C. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Novel+single-chain+antibody-targeted+microbubbles+for+molecular+ultrasound+imaging+of+thrombosis:+validation+of+a+unique+noninvasive+method+for+rapid+and+sensitive+detection+of+thrombi+and+monitoring+of+success+or+failure+of+thrombolysis+in+mice.">Novel single-chain antibody-targeted microbubbles for molecular ultrasound imaging of thrombosis: validation of a unique noninvasive method for rapid and sensitive detection of thrombi and monitoring of success or failure of thrombolysis in mice.</a> Circulation. 125 (25), 3117-3126 (2012).</li><li>Wang, X., Smith, P. L., Hsu, M. -. Y., Ogletree, M. L., Schumacher, W. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Murine+model+of+ferric+chloride-induced+vena+cava+thrombosis:+evidence+for+effect+of+potato+carboxypeptidase+inhibitor.">Murine model of ferric chloride-induced vena cava thrombosis: evidence for effect of potato carboxypeptidase inhibitor.</a> J Thromb Haemost. 4 (2), 403-410 (2006).</li><li>Karatas, H., Erdener, S. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thrombotic+distal+middle+cerebral+artery+occlusion+produced+by+topical+FeCl(3)+application:+a+novel+model+suitable+for+intravital+microscopy+and+thrombolysis+studies.">Thrombotic distal middle cerebral artery occlusion produced by topical FeCl(3) application: a novel model suitable for intravital microscopy and thrombolysis studies.</a> J Cereb Blood Flow Metab. 31 (6), 1452-1460 (2011).</li><li>Jirouskov&#225;, M., Chereshnev, I., V&#228;&#228;n&#228;nen, H., Degen, J. L., Coller, B. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antibody+blockade+or+mutation+of+the+fibrinogen+gamma-chain+C-terminus+is+more+effective+in+inhibiting+murine+arterial+thrombus+formation+than+complete+absence+of+fibrinogen.">Antibody blockade or mutation of the fibrinogen gamma-chain C-terminus is more effective in inhibiting murine arterial thrombus formation than complete absence of fibrinogen.</a> Blood. 103 (6), 1995-2002 (2004).</li><li>Dubois, C., Panicot-Dubois, L., Merrill-Skoloff, G., Furie, B., Furie, B. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glycoprotein+VI-dependent+and+-independent+pathways+of+thrombus+formation+in+vivo.">Glycoprotein VI-dependent and -independent pathways of thrombus formation in vivo.</a> Blood. 107 (10), 3902-3906 (2006).</li><li>Navarrete, A. -. M., Casari, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+murine+model+to+characterize+the+antithrombotic+effect+of+molecules+targeting+human+von+Willebrand+factor.">A murine model to characterize the antithrombotic effect of molecules targeting human von Willebrand factor.</a> Blood. 120 (13), 2723-2732 (2012).</li><li>Rayes, J., Hollestelle, M. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mutation+and+ADAMTS13-dependent+modulation+of+disease+severity+in+a+mouse+model+for+von+Willebrand+disease+type+2B.">Mutation and ADAMTS13-dependent modulation of disease severity in a mouse model for von Willebrand disease type 2B.</a> Blood. 115 (23), 4870-4877 (2010).</li><li>Lamrani, L., Lacout, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hemostatic+disorders+in+a+JAK2V617F-driven+mouse+model+of+myeloproliferative+neoplasm.">Hemostatic disorders in a JAK2V617F-driven mouse model of myeloproliferative neoplasm.</a> Blood. 124 (7), 1136-1145 (2014).</li><li>Zhang, Y., Ye, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increased+platelet+activation+and+thrombosis+in+transgenic+mice+expressing+constitutively+active+P2Y12.">Increased platelet activation and thrombosis in transgenic mice expressing constitutively active P2Y12.</a> J Thromb Haemost. 10 (10), 2149-2157 (2012).</li><li>Sch&#228;fer, K., Konstantinides, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Different+mechanisms+of+increased+luminal+stenosis+after+arterial+injury+in+mice+deficient+for+urokinase-+or+tissue-type+plasminogen+activator.">Different mechanisms of increased luminal stenosis after arterial injury in mice deficient for urokinase- or tissue-type plasminogen activator.</a> Circulation. 106 (14), 1847-1852 (2002).</li><li>Wang, X., Palasubramaniam, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Towards+effective+and+safe+thrombolysis+and+thromboprophylaxis:+preclinical+testing+of+a+novel+antibody-targeted+recombinant+plasminogen+activator+directed+against+activated+platelets.">Towards effective and safe thrombolysis and thromboprophylaxis: preclinical testing of a novel antibody-targeted recombinant plasminogen activator directed against activated platelets.</a> Circ Res. 114 (7), 1083-1093 (2014).</li><li>Decrem, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ir-CPI,+a+coagulation+contact+phase+inhibitor+from+the+tick+Ixodes+ricinus,+inhibits+thrombus+formation+without+impairing+hemostasis.">Ir-CPI, a coagulation contact phase inhibitor from the tick Ixodes ricinus, inhibits thrombus formation without impairing hemostasis.</a> J Exp Med. 206 (11), 2381-2395 (2009).</li><li>Ma, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Desmolaris,+a+novel+factor+XIa+anticoagulant+from+the+salivary+gland+of+the+vampire+bat+(Desmodus+rotundus)+inhibits+inflammation+and+thrombosis+in+vivo.">Desmolaris, a novel factor XIa anticoagulant from the salivary gland of the vampire bat (Desmodus rotundus) inhibits inflammation and thrombosis in vivo.</a> Blood. 122 (25), 4094-4106 (2013).</li><li>Lei, X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anfibatide,+a+novel+GPIb+complex+antagonist,+inhibits+platelet+adhesion+and+thrombus+formation+in+vitro+and+in+vivo+in+murine+models+of+thrombosis.">Anfibatide, a novel GPIb complex antagonist, inhibits platelet adhesion and thrombus formation in vitro and in vivo in murine models of thrombosis.</a> Thromb Haemost. 111 (2), 279-289 (2014).</li><li>Waisberg, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Plasmodium+falciparum+infection+induces+expression+of+a+mosquito+salivary+protein+(Agaphelin)+that+targets+neutrophil+function+and+inhibits+thrombosis+without+impairing+hemostasis.">Plasmodium falciparum infection induces expression of a mosquito salivary protein (Agaphelin) that targets neutrophil function and inhibits thrombosis without impairing hemostasis.</a> PLoS Pathog. 10 (9), e1004338 (2014).</li><li>Owens, A. P., Lu, Y., Whinna, H. C., Gachet, C., Fay, W. P., Mackman, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Towards+a+standardization+of+the+murine+ferric+chloride-induced+carotid+arterial+thrombosis+model.">Towards a standardization of the murine ferric chloride-induced carotid arterial thrombosis model.</a> J Thromb Haemost. 9 (9), 1862-1863 (2011).</li><li>Wang, X., Xu, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+optimized+murine+model+of+ferric+chloride-induced+arterial+thrombosis+for+thrombosis+research.">An optimized murine model of ferric chloride-induced arterial thrombosis for thrombosis research.</a> Thromb Res. 115 (1-2), 95-100 (2005).</li><li>Tseng, M. T., Dozier, A., Haribabu, B., Graham, U. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transendothelial+migration+of+ferric+ion+in+FeCl3+injured+murine+common+carotid+artery.">Transendothelial migration of ferric ion in FeCl3 injured murine common carotid artery.</a> Thromb Res. 118 (2), 275-280 (2006).</li><li>Bonnard, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Leukocyte+mimetic+polysaccharide+microparticles+tracked+in+vivo+on+activated+endothelium+and+in+abdominal+aortic+aneurysm.">Leukocyte mimetic polysaccharide microparticles tracked in vivo on activated endothelium and in abdominal aortic aneurysm.</a> Acta Biomater. 10 (8), 3535-3545 (2014).</li><li>Boulaftali, Y., Lamrani, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+mouse+dorsal+skinfold+chamber+as+a+model+for+the+study+of+thrombolysis+by+intravital+microscopy.">The mouse dorsal skinfold chamber as a model for the study of thrombolysis by intravital microscopy.</a> Thromb Haemost. 107 (5), 962-971 (2012).</li><li>Konstantinides, S., Sch&#228;fer, K., Thinnes, T., Loskutoff, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Plasminogen+activator+inhibitor-1+and+its+cofactor+vitronectin+stabilize+arterial+thrombi+after+vascular+injury+in+mice.">Plasminogen activator inhibitor-1 and its cofactor vitronectin stabilize arterial thrombi after vascular injury in mice.</a> Circulation. 103 (4), 576-583 (2001).</li><li>Li, W., McIntyre, T. M., Silverstein, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ferric+chloride-induced+murine+carotid+arterial+injury:+A+model+of+redox+pathology.">Ferric chloride-induced murine carotid arterial injury: A model of redox pathology.</a> Redox Biol. 1 (1), 50-55 (2013).</li></ol>

The authors have nothing to disclose.

El modelo de trombosis inducida por cloruro férrico es una excelente herramienta de investigación. Como se muestra en este estudio, es extremadamente fácil de implementar y cuando se utiliza en combinación con microscopía intravital o medidor de flujo Doppler, que proporciona un buen seguimiento en tiempo real de la formación de trombos. Ajuste del tiempo de exposición y la concentración de FeCl 3, sino que también ofrece la posibilidad de producir ya sea trombos no oclusivos o oclusivos. <p clas...

El estudio de los mecanismos implicados en el desarrollo de la trombosis y la evaluación de la eficacia de los fármacos anti-trombóticos requiere bien establecido modelos animales experimentales. Modelos animales grandes fueron los primeros en ser utilizados ya que proporcionan grandes vasos más similar a los humanos que los roedores 1. Sin embargo, el alto costo, las grandes instalaciones requeridas y la dificultad para manipularlos genéticamente son los principales inconvenientes para su uso y animales grandes ahora se limitan a estudios preclínicos finales una vez que las pruebas preliminares sobre roedores han dado resultados concluyentes 2. Con amplia disponibilidad de cepas transgénicas y knockout y su pequeño tamaño que minimiza la cantidad de fármacos antitrombóticos requeridos para las pruebas in vivo, los ratones se utilizan principalmente para la investigación trombosis. Por lo tanto, se han desarrollado varios modelos de trastornos trombóticos en ratones 3. Muchos modelos de trombosis establecidos interrumpen las intimuna capa de la pared del vaso, seguido por la exposición de la matriz extracelular sub endotelial para el flujo de sangre induciendo la formación de coágulos de sangre 4. El trombos puede resultar de la exposición del colágeno que desencadena la activación de las plaquetas y / o de la exposición de factor tisular que activa la cascada de coagulación 5. Se emplean a continuación varias técnicas para lograr la lesión inicial buque. Pierangeli et al. Desarrolló un modelo de rotura mecánica con una herramienta de microcirugía en la vena femoral 6. Kikushi et al. Describe un modelo que consiste en la administración de un compuesto reactivo foto (Rosa de Bengala) que se acumula en la bicapa lipídica de las células endoteliales, seguido de la excitación específica de la pared del vaso de interés con luz verde (540 nm) 7. La lesión también puede ser inducida por un corto de alta intensidad del láser de impulsos de iluminación 8. Otra técnica estableció en primer lugar en la arteria carótida de las ratasconsiste en la aplicación tópica de cloruro férrico (FeCl3) 9. En este caso, los resultados de denudación recipiente de radicales libres generados por FeCl 3 que hace que la peroxidación de lípidos y la destrucción de las células endoteliales 10. La lesión induce la expresión de varias moléculas de adhesión de activación adhesión y agregación plaquetaria, así como el reclutamiento de leucocitos. Se ha demostrado que los leucocitos, especialmente neutrófilos, juegan un papel crucial en la activación de la cascada de la coagulación de la sangre que lleva a la trombosis 11. Este método es muy adecuado para reproducir la cascada de la coagulación; los investigadores deben tener en cuenta que, en este modelo de ratón, la trombosis es típicamente inducida en vasos sanos mientras que la trombosis en los seres humanos ocurre principalmente en por ejemplo enfermo. vasos ateroscleróticos. Como este modelo es muy simple de implementar y también es eficaz en ratones, ahora es el modo de trombosis utilizado principalmentel para pequeños animales en estudios in vivo. Además, esta técnica ofrece la posibilidad de inducir la formación de trombos en una variedad de vasos. Vasos de destino pueden ser arterias o venas de gran diámetro (carótida, femoral, la vena cava) o pequeño diámetro (mesenterio, cremaster) 12-14. Más recientemente, también se utiliza en la arteria cerebral media proximal para desarrollar un modelo de accidente cerebrovascular 15. La formación de trombosis puede ser observada directamente por microscopía intravital después del marcaje fluorescente de plaquetas y leucocitos o monitoreada midiendo la disminución del flujo sanguíneo con una sonda de temperatura o una sonda Doppler 12,16,17. Varios parámetros tales como el tiempo de oclusión, el tiempo de la formación de trombos o tamaño del trombo pueden entonces ser investigadas. Las diferencias fisiológicas entre los vasos investigaron resultado en variaciones significativas en los trombos obtenido. Por lo tanto, los investigadores suelen seleccionar el vaso diana de acuerdo con los parámetros que desean Measure y / o el ajuste que quieren investigar la enfermedad. Típicamente, el modelo de la arteria carótida es más relevante para la investigación en aterotrombosis relacionada con el infarto de miocardio o un accidente cerebrovascular mientras que los estudios sobre la vena cava son más relevantes para la investigación de la trombosis venosa profunda. La accesibilidad de los diferentes vasos también determina el método utilizado para medir el crecimiento de trombos. Por ejemplo, los vasos mesentéricos son de fácil acceso haciendo que este modelo muy adecuado para la observación microscópica intravital y el estudio de la dinámica de la formación de trombos. La arteria carótida es menos accesible pero más grande que permitan a las mediciones de flujo de sangre y proporcionan un excelente modelo para estudiar la trombosis oclusiva. El modelo de trombosis inducida por cloruro férrico ha proporcionado enormes progresos en la comprensión de esta patología. Se ha utilizado en muchos estudios centrados en el papel del factor von Willebrand en la formación de trombosis 18,19. Combinado con modificaciones genéticastécnicas ficación, que ha permitido la identificación de muchos genes específicos implicados en los trastornos trombóticos. Lamrani et al. por ejemplo han demostrado que un knock-in del gen JAK2 V617F se asocia con la formación de un coágulo de aceleración de la inestable 20. Zhang et al. Han investigado la implicación fisiológica del receptor plaquetario P2Y12 y demostrado que los ratones transgénicos que sobreexpresan específicamente este receptor en plaquetas solamente, muestran una formación de trombos más rápido y estable en la arteria mesentérica lesionado con FeCl 3 21. El papel crucial de activador de plasminógeno de tipo tisular (tPA) y del activador del plasminógeno de tipo uroquinasa (uPA) en el proceso de degradación de la fibrina también ha sido investigado en este método 22. Además, este modelo también ofrece una manera sencilla y exacta de probar las capacidades de muchos fármacos fibrinolíticos novedosas in vivo. Por ejemplo, Wang et al., Han utilizado este modelo para THcorreo de validación preclínica de una novela del activador del plasminógeno recombinante dirigido contra las plaquetas activadas 23. Este método también permitió la validación de proteínas terapéuticas aisladas a partir de la saliva de garrapatas, murciélagos vampiro, y mosquitos o a partir del veneno de serpientes con la identificación específica del objetivo 24-27. Estos ejemplos demuestran la versatilidad del modelo de cloruro férrico. En este artículo, nos centramos en dos métodos y estudiamos cloruro férrico trombosis inducida en dos tipo de buque diferentes; buque mesentérica y la arteria carótida.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>Whatman chromatography paper</td>
			<td>GE Healthcare</td>
			<td>3030917</td>
			<td></td>
		</tr>
		<tr>
			<td>Iron (III) chloride 40 % (w/v)</td>
			<td>VWR</td>
			<td>24212.298</td>
			<td></td>
		</tr>
		<tr>
			<td>Rhodamine 6G</td>
			<td>Sigma</td>
			<td>R4127</td>
			<td></td>
		</tr>
		<tr>
			<td>Inverted microscope&#160;</td>
			<td>Olympus</td>
			<td>IX81</td>
			<td></td>
		</tr>
		<tr>
			<td>Digital black-and-white camera&#160;</td>
			<td>Olympus</td>
			<td>XM10</td>
			<td></td>
		</tr>
		<tr>
			<td>Doppler flowmeter</td>
			<td>Transonic</td>
			<td>TS420</td>
			<td></td>
		</tr>
		<tr>
			<td>Nano-doppler flow probe</td>
			<td>Transonic</td>
			<td>0.5 PBS</td>
			<td></td>
		</tr>
		<tr>
			<td>Ketamine</td>
			<td>Hospira&#160;</td>
			<td>0409-2051-05</td>
			<td></td>
		</tr>
		<tr>
			<td>Xylazine (Rampun)</td>
			<td>Bayer</td>
			<td>75313&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Petri dish</td>
			<td>Sarstedt</td>
			<td>82.1472</td>
			<td></td>
		</tr>
		<tr>
			<td>Insulin syringe (29 G)</td>
			<td>BD Ultra-Fine</td>
			<td>326103</td>
			<td></td>
		</tr>
		<tr>
			<td>Cotton tipped applicators</td>
			<td>BSN medical</td>
			<td>211827A</td>
			<td></td>
		</tr>
		<tr>
			<td>Dynek dysilk sutures</td>
			<td>Dynek Pty Ltd</td>
			<td>CS30100</td>
			<td></td>
		</tr>
		<tr>
			<td>Dulbecco&#39;s phosphate buffer saline (PBS)</td>
			<td>Gibco life technologies</td>
			<td>21600-069</td>
			<td></td>
		</tr>
		<tr>
			<td>Heating pad</td>
			<td>Kirchner</td>
			<td>T60</td>
			<td></td>
		</tr>
	</tbody>
</table>

ferric chloride-induced thrombosis mouse model on carotid artery and mesentery vessel

Severe thrombosis and its ischemic consequences such as myocardial infarction, pulmonary embolism and stroke are major worldwide health issues. The ferric chloride injury is now a well-established technique to rapidly and accurately induce the formation of thrombi in exposed veins or artery of small and large diameter. This model has played a key role in the study of the pathophysiology of thrombosis, in the discovery and validation of novel antithrombotic drugs and in the understanding of the mechanism of action of these new agents. Here, the implementation of this technique on a mesenteric vessel and carotid artery in mice is presented. The method describes how to label circulating leukocytes and platelets with a fluorescent dye and to observe, by intravital microscopy on the exposed mesentery, their accumulation at the injured vessel wall which leads to the formation of a thrombus. On the carotid artery, the occlusion caused by the clot formation is measured by monitoring the blood flow with a Doppler probe.

El fluorescente observación microscopía intravital del mesenterio revelará la acumulación de Rodamina 6G etiquetado plaquetas y leucocitos lo largo de la pared de los vasos lesionados por FeCl3. La formación progresiva de un trombo parcial se supervisa en un recipiente mesenterio 200 micras (Figura 1). Un trombo aparece lentamente y es claramente identificable después de la primera minutos de exposición a FeCl 3 (Figura 1, t = 60 seg). 40 seg después de la ...

Watch this Scientific Journal Video about Ferric Chloride-induced Thrombosis Mouse Model on Carotid Artery and Mesentery Vessel at JoVE.com

Ferric Chloride-induced Thrombosis Mouse Model on Carotid Artery and Mesentery Vessel

The FeCl3 induced thrombosis model in mice is described herein. A method to monitor thrombus growth by intravital microscopy observation on a mesenteric vessel and by blood flow measurement in the carotid artery is presented.

Trombosis modelo de ratón de la arteria carótida y Mesenterio Embarcaciones inducida por cloruro férrico-

Severe thrombosis and its ischemic consequences such as myocardial infarction, pulmonary embolism and stroke are major worldwide health issues. The ferric ...

ferric-chloride-induced-thrombosis-mouse-model-on-carotid-artery

Research

JoVE Journal

Medicine

17.3K vistas.  Baker IDI Heart and Diabetes Institute. The FeCl3 induced thrombosis model in mice is described herein. A method to monitor thrombus growth by intravital microscopy observation on a mesenteric vessel and by blood flow measurement in the carotid artery is presented.

Video: Ferric Chloride-induced Thrombosis Mouse Model on Carotid Artery and Mesentery Vessel

A Model of Disturbed Flow-Induced Atherosclerosis in Mouse Carotid Artery by Partial Ligation and a Simple Method of RNA Isolation from Carotid Endothelium

Despite the well-known close association, direct evidence linking disturbed flow to atherogenesis has been lacking. We have recently used a modified version of carotid partial ligation methods [1,2] to show that it acutely induces low and oscillatory flow conditions, two key characteristics of disturbed flow, in the mouse common carotid artery. Using this model, we have provided direct evidence that disturbed flow indeed leads to rapid and robust atherosclerosis development in Apolipoprotein E knockout mouse [3]. We also developed a method of endothelial RNA preparation with high purity from the mouse carotid intima [3]. Using this mouse model and method, we found that partial ligation causes endothelial dysfunction in a week, followed by robust and rapid atheroma formation in two weeks in a hyperlipidemic mouse model along with features of complex lesion formation such as intraplaque neovascularization by four weeks. This rapid in vivo model and the endothelial RNA preparation method could be used to determine molecular mechanisms underlying flow-dependent regulation of vascular biology and diseases. Also, it could be used to test various therapeutic interventions targeting endothelial dysfunction and atherosclerosis in considerably reduced study duration.

This describes a partial carotid ligation surgery, which causes disturbed flow conditions and subsequent atherosclerosis development (in two weeks) with intraplaque neo-vascularization (in four weeks) in the mouse common carotid artery. We also describe a novel method of RNA isolation from the carotid intima, providing high purity endothelial RNA.

Un modelo de Disturbed inducida por el flujo de Aterosclerosis en la arteria carótida del ratón por la ligadura parcial y un método sencillo para el aislamiento de ARN del endotelio carótida

Despite the well-known close association, direct evidence linking disturbed flow to atherogenesis has been lacking. We have recently used a modified ...

Investigación

Medicina

Mouse Complete Stasis Model of Inferior Vena Cava Thrombosis

Venous thromboembolism (VTE) includes both deep vein thrombosis (DVT) and pulmonary embolism (PE). In the United States (U.S.), the high morbidity and mortality rates make VTE a serious health concern 1-2. After heart disease and stroke, VTE is the third most common vascular disease 3. In the U.S. alone, there is an estimated 900,000 people affected each year, with 300,000 deaths occurring annually 3. A reliable in vivo animal model to study the mechanisms of this disease is necessary.
The advantages of using the mouse complete stasis model of inferior vena cava thrombosis are several. The mouse model allows for the administration of very small volumes of limited availability test agents, reducing costs dramatically. Most promising is the potential for mice with gene knockouts that allow specific inflammatory and coagulation factor functions to be delineated. Current molecular assays allow for the quantitation of vein wall, thrombus, whole blood, and plasma for assays. However, a major concern involving this model is the operative size constraints and the friability of the vessels. Also, due to the small IVC sample weight (mean 0.005 grams) it is necessary to increase animal numbers for accurate statistical analysis for tissue, thrombus, and blood assays such as real-time polymerase chain reaction (RT-PCR), western blot, enzyme-linked immunosorbent (ELISA), zymography, vein wall and thrombus cellular analysis, and whole blood and plasma assays 4-8.
The major disadvantage with the stasis model is that the lack of blood flow inhibits the maximal effect of administered systemic therapeutic agents on the thrombus and vein wall.

The mouse complete stasis model of inferior vena cava thrombosis yields quantifiable amounts of vein wall tissue and thrombus. It has proven useful for evaluating interactions between the vein wall and the occlusive thrombus and in assessing the progression from acute to chronic inflammation.

Ratón completo modelo de estasis Inferior Vena Cava trombosis

Venous thromboembolism (VTE) includes both deep vein thrombosis (DVT) and pulmonary embolism (PE).  In the United States (U.S.), the high morbidity and ...

Mouse Model of Middle Cerebral Artery Occlusion

Stroke is the most common fatal neurological disease in the United States 1. The majority of strokes (88%) result from blockage of blood vessels in the brain (ischemic stroke) 2. Since most ischemic strokes (~80%) occur in the territory of middle cerebral artery (MCA) 3, many animal stroke models that have been developed have focused on this artery. The intraluminal monofilament model of middle cerebral artery occlusion (MCAO) involves the insertion of a surgical filament into the external carotid artery and threading it forward into the internal carotid artery (ICA) until the tip occludes the origin of the MCA, resulting in a cessation of blood flow and subsequent brain infarction in the MCA territory 4. The technique can be used to model permanent or transient occlusion 5. If the suture is removed after a certain interval (30 min, 1 h, or 2 h), reperfusion is achieved (transient MCAO); if the filament is left in place (24 h) the procedure is suitable as a model of permanent MCAO. This technique does not require craniectomy, a neurosurgical procedure to remove a portion of skull, which may affect intracranial pressure and temperature 6. It has become the most frequently used method to mimic permanent and transient focal cerebral ischemia in rats and mice 7,8. To evaluate the extent of cerebral infarction, we stain brain slices with 2,3,5-triphenyltetrazolium chloride (TTC) to identify ischemic brain tissue 9. In this video, we demonstrate the MCAO method and the determination of infarct size by TTC staining.

We demonstrate in the video a method for producing a middle cerebral artery occlusion in adult mice using an intraluminal monofilament. We also show how to evaluate the extent of cerebral infarction by 2,3,5-triphenyltetrazolium chloride (TTC) staining.

Modelo de ratón de la oclusión arteria cerebral media

Stroke is the most common fatal neurological disease in the United States 1. The majority of strokes (88%) result from blockage of blood vessels in the ...

A Murine Model of Stent Implantation in the Carotid Artery for the Study of Restenosis

Despite the considerable progress made in the stent development in the last decades, cardiovascular diseases remain the main cause of death in western countries. Beside the benefits offered by the development of different drug-eluting stents, the coronary revascularization bears also the life-threatening risks of in-stent thrombosis and restenosis. Research on new therapeutic strategies is impaired by the lack of appropriate methods to study stent implantation and restenosis processes. Here, we describe a rapid and accessible procedure of stent implantation in mouse carotid artery, which offers the possibility to study in a convenient way the molecular mechanisms of vessel remodeling and the effects of different drug coatings.

A model of stent implantation in mouse carotid artery is described. Compared to other similar methods, this procedure is very rapid, simple and accessible, offering the possibility to study in a convenient way the vascular wall reaction to different drug-eluting stents and the molecular mechanisms of restenosis.

Un modelo murino de la implantación de un stent en la arteria carótida para el Estudio de la reestenosis

Despite  the considerable progress made in the stent development in the last decades,  cardiovascular diseases remain the main cause of death in western ...

Adult Mouse Venous Hypertension Model: Common Carotid Artery to External Jugular Vein Anastomosis.

The understanding of the pathophysiology of brain arteriovenous malformations and arteriovenous fistulas has improved thanks to animal models. A rat model creating an artificial fistula between the common carotid artery (CCA) and the external jugular vein (EJV) has been widely described and proved technically feasible. This construct provokes a consistent cerebral venous hypertension (CVH), and therefore has helped studying the contribution of venous hypertension to formation, clinical symptoms, and prognosis of brain AVMs and dural AVFs. Equivalent mice models have been only scarcely described and have shown trouble with stenosis of the fistula. An established murine model would allow the study of not only pathophysiology but also potential genetic therapies for these cerebrovascular diseases.
We present a model of arteriovenous fistula that produces a durable intracranial venous hypertension in the mouse. Microsurgical anastomosis of the murine CCA and EJV can be difficult due to diminutive anatomy and frequently result in a non-patent fistula. In this step-by-step protocol we address all the important challenges encountered during this procedure. Avoiding excessive retraction of the vein during the exposure, using 11-0 sutures instead of 10-0, and making a carefully planned end-to-side anastomosis are some of the critical steps. Although this method requires advanced microsurgical skills and a longer learning curve that the equivalent in the rat, it can be consistently developed.
This novel model has been designed to integrate transgenic mouse techniques with a previously well-established experimental system that has proved useful to study brain AVMs and dural AVFs. By opening the possibility of using transgenic mice, a broader spectrum of valid models can be achieved and genetic treatments can also be tested. The experimental construct could also be further adapted to the study of other cerebrovascular diseases related with venous hypertension such as migraine, transient global amnesia, transient monocular blindness, etc.

We describe a method for creating a reliable model of cerebral venous hypertension in the adult mouse. This model has been widely described and tested in the rat. This new counterpart in the mice opens the possibility of using genetic modified animals and thereby broadens the applications of the model.

Adultos Mouse hipertensión venosa: Modelo: arteria carótida común a la vena yugular externa anastomosis.

The understanding of the pathophysiology of brain arteriovenous malformations and arteriovenous fistulas has improved thanks to animal models. A rat model ...

Vascular Balloon Injury and Intraluminal Administration in Rat Carotid Artery

The carotid artery balloon injury model in rats has been well established for over two decades. It remains an important method to study the molecular and cellular mechanisms involved in vascular smooth muscle dedifferentiation, neointima formation and vascular remodeling. Male Sprague-Dawley rats are the most frequently employed animals for this model. Female rats are not preferred as female hormones are protective against vascular diseases and thus introduce a variation into this procedure. The left carotid is typically injured with the right carotid serving as a negative control. Left carotid injury is caused by the inflated balloon that denudes the endothelium and distends the vessel wall. Following injury, potential therapeutic strategies such as the use of pharmacological compounds and either gene or shRNA transfer can be evaluated. Typically for gene or shRNA transfer, the injured section of the vessel lumen is locally transduced for 30 min with viral particles encoding either a protein or shRNA for delivery and expression in the injured vessel wall. Neointimal thickening representing proliferative vascular smooth muscle cells usually peaks at 2 weeks after injury. Vessels are mostly harvested at this time point for cellular and molecular analysis of cell signaling pathways as well as gene and protein expression. Vessels can also be harvested at earlier time points to determine the onset of expression and/or activation of a specific protein or pathway, depending on the experimental aims intended. Vessels can be characterized and evaluated using histological staining, immunohistochemistry, protein/mRNA assays, and activity assays. The intact right carotid artery from the same animal is an ideal internal control. Injury-induced changes in molecular and cellular parameters can be evaluated by comparing the injured artery to the internal right control artery. Likewise, therapeutic modalities can be evaluated by comparing the injured and treated artery to the control injured only artery.

This protocol uses a balloon catheter to cause an intraluminal injury on the rat carotid artery and henceforth elicit neointimal hyperplasia. This is a well-established model for studying the mechanisms of vascular remodeling in response to injury. It is also widely used to determine the validity of potential therapeutic approaches.

Lesión Vascular Globo y Administración intraluminal en rata arteria carótida

The carotid artery balloon injury model in rats has been well established for over two decades. It remains an important method to study the molecular and ...

A Rat Carotid Balloon Injury Model to Test Anti-vascular Remodeling Therapeutics

The rat carotid balloon injury is a well-established surgical model that has been used to study arterial remodeling and vascular cell proliferation. It is also a valuable model system to test, and to evaluate therapeutics and drugs that negate maladaptive remodeling in the vessel. The injury, or barotrauma, in the vessel lumen caused by an inflated balloon via an inserted catheter induces subsequent neointimal growth, often leading to hyperplasia or thickening of the vessel wall that narrows, or obstructs the lumen. The method described here is sufficiently sensitive, and the results can be obtained in relatively short time (2 weeks after the surgery). The efficacy of the drug or therapeutic against the induced-remodeling can be evaluated either by the post-mortem pathological and histomorphological analysis, or by ultrasound sonography in live animals. In addition, this model system has also been used to determine the therapeutic window or the time course of the administered drug. These studies can leadto the development of a better administrative strategy and a better therapeutic outcome. The procedure described here provides a tool for translational studies that bring drug and therapeutic candidates from bench research to clinical applications.

The rat carotid balloon injury model described below allows researchers to evaluate drugs or therapeutics that negate injury-induced arterial hyperplasia. Detailed pre-surgical preparation, surgical procedure, and post-surgical cares of the animal are described.

Una rata carótida lesión con el balón modelo para probar Terapéutica Remodelación Anti-vasculares

The rat carotid balloon injury is a well-established surgical model that has been used to study arterial remodeling and vascular cell proliferation. It is ...

Ferric Chloride-induced Murine Thrombosis Models

Arterial thrombosis (blood clot) is a common complication of many systemic diseases associated with chronic inflammation, including atherosclerosis, diabetes, obesity, cancer and chronic autoimmune rheumatologic disorders. Thrombi are the cause of most heart attacks, strokes and extremity loss, making thrombosis an extremely important public health problem. Since these thrombi stem from inappropriate platelet activation and subsequent coagulation, targeting these systems therapeutically has important clinical significance for developing safer treatments. Due to the complexities of the hemostatic system, in vitro experiments cannot replicate the blood-to-vessel wall interactions; therefore, in vivo studies are critical to understand pathological mechanisms of thrombus formation. To this end, various thrombosis models have been developed in mice. Among them, ferric chloride (FeCl3) induced vascular injury is a widely used model of occlusive thrombosis that reports platelet activation and aggregation in the context of an aseptic closed vascular system. This model is based on redox-induced endothelial cell injury, which is simple and sensitive to both anticoagulant and anti-platelets drugs. The time required for the development of a thrombus that occludes blood flow gives a quantitative measure of vascular injury, platelet activation and aggregation that is relevant to thrombotic diseases. We have significantly refined this FeCl3-induced vascular thrombosis model, which makes the data highly reproducible with minimal variation. Here we describe the model and present representative data from several experimental set-ups that demonstrate the utility of this model in thrombosis research.

We report a refined procedure of the ferric chloride (FeCl3)-induced thrombosis models on carotid and mesenteric artery as well as vein, characterized efficiently using intravital microscopy to monitor time to occlusive thrombi formation.

Férricos modelos murinos trombosis inducida por cloruro

Arterial thrombosis (blood clot) is a common complication of many systemic diseases associated with chronic inflammation, including atherosclerosis, ...

Phase Contrast Magnetic Resonance Imaging in the Rat Common Carotid Artery

Phase contrast magnetic resonance imaging (PC-MRI) is a noninvasive approach that can quantify flow-related parameters such as blood flow. Previous studies have shown that abnormal blood flow may be associated with systemic vascular risk. Thus, PC-MRI can facilitate the translation of data obtained from animal models of cardiovascular diseases to pertinent clinical investigations. In this report, we describe the procedure for measuring blood flow in the common carotid artery (CCA) of rats using cine-gated PC-MRI and discuss relevant analysis methods. This procedure can be performed in a live, anesthetized animal and does not require euthanasia after the procedure. The proposed scanning parameters yield repeatable measurements for blood flow, indicating excellent reproducibility of the results. The PC-MRI procedure described in this article can be used for pharmacological testing, pathophysiological assessment, and cerebral hemodynamics evaluation.

The overall goal of this procedure is to measure the blood flow in the rat common carotid artery by using noninvasive phase contrast magnetic resonance imaging.

Fase contraste resonancia magnética en la arteria carótida común de rata

Phase contrast magnetic resonance imaging (PC-MRI) is a noninvasive approach that can quantify flow-related parameters such as blood flow. Previous ...

Ferric Chloride-induced Canine Carotid Artery Thrombosis: A Large Animal Model of Vascular Injury

Occlusive arterial thrombosis leading to cerebral ischemic stroke and myocardial infarction contributes to ~13 million deaths every year globally. Here, we have translated a vascular injury model from a small animal into a large animal (canine), with slight modifications that can be used for pre-clinical screening of prophylactic and thrombolytic agents. In addition to the surgical methods, the modified protocol describes the step-by-step methods to assess carotid artery canalization by angiography, detailed instructions to process both the brain and carotid artery for histological analysis to verify carotid canalization and cerebral hemorrhage, and specific parameters to complete an assessment of downstream thromboembolic events by utilizing magnetic resonance imaging (MRI). In addition, specific procedural changes from the previously well-established small animal model necessary to translate into a large animal (canine) vascular injury are discussed.

Here, we present the modifications necessary to a well characterized and commonly used small animal ferric chloride-induced (FeCl3) carotid artery injury model for use in a large animal vascular injury model. The resulting model can be utilized for pre-clinical trial assessment of both prophylactic and thrombolytic pharmacological and mechanical interventions.

Trombosis de la arteria carótida canina cloruro férrico-inducida: Un modelo Animal grande de lesión Vascular

Occlusive arterial thrombosis leading to cerebral ischemic stroke and myocardial infarction contributes to ~13 million deaths every year globally. Here, ...