This work was supported by the American Heart Association Scientist Development Grant and the Foundation for Anesthesia Education and Research (MLJ). We would like to thank Talaignair N. Venkatraman PhD for his assistance with magnetic resonance imaging.

Declaración de Ética: Este protocolo ha sido aprobado por el Comité de la Universidad de Duke Institucional Cuidado de Animales y Uso y sigue todas las pautas para el uso ético de los animales. 1. Preparación de Equipo <ol><li> Autoclave las herramientas quirúrgicas antes de la cirugía. </li><li> Desinfectar el aparato estereotáctico con etanol al 70%. </li><li> Encienda el baño de agua y mantener la temperatura del agua a 42 ° C. </li><li> Disolver Tipo IV-S clostridial colagenasa en solución salina normal a una concentración de 0,075 U por 0,4 l. </li></ol> 2. Colagenasa Modelo Inyección <ol><li> Pesar el ratón. </li><li> Anestesiar el ratón en una cámara de inducción con 5% de isoflurano en el 30% O 2/70% N 2. Anestesia adecuada se indica después de aproximadamente 2 minutos, cuando la respiración de ratón han reducido a 1 por segundo. </li><li> Intubar la tráquea con una G catéter intravenoso 30 mm 20. </li><li> Conectar el catéter a unrespirador de roedores y ventilar mecánicamente los pulmones con el 1,6% de isoflurano en el 30% O 2/70% N 2, a razón de 105 respiraciones por minuto, con un volumen corriente entregado de 0,75 ml .. </li><li> Afeitar el cuero cabelludo con una máquina de afeitar electrónica. Una vez que el ratón se anestesia y intubado, moverlo a una estación de trabajo diferente para el afeitado y luego regresó a la banca quirúrgica. </li><li> Asegure la cabeza en un marco estereotáctico y nivelar la cabeza con la sutura coronal y sagital tanto como puntos de referencia. </li><li> Aplique una pomada oftálmica para los ojos. </li><li> Inserte una sonda de temperatura rectal. Mantenga la temperatura rectal a 37,0 ± 0,2 ° C por medio de una cama de agua que circula debajo de la carrocería. </li><li> Limpie el área quirúrgica con betadine seguido con etanol al 70% y repetir 3 veces. </li><li> Haga una incisión en la línea media del cuero cabelludo 1 cm y limpie periostio lateralmente con un aplicador con punta de algodón estéril para exponer bregma. </li><li> Taladro 1 mm de diámetro agujero de trépano de 2,2 mm dejó latEral al bregma con un taladro refrigerado por agua. </li><li> Gire vial colagenasa 5 veces, luego lavar una jeringa de 0,5 l con 25 G aguja (que se adjunta al marco estereotáctico) con 0,5 l solución de colagenasa 5 veces (Deja 0,5 l de solución de colagenasa en la jeringa después de la última de lavado). </li><li> Alinee la punta de la aguja con orificio de trepanación luego expulsar a 0,1 l de la jeringa y limpie bisel de la aguja con la maquinilla de afeitar de desprenderse. </li><li> El uso de un micromanipulador, avanzar la aguja de 3 mm de profundidad de la corteza y dejar inmóvil durante 30 segundos. </li><li> Inyectar 0,4 l más de 90 seg. </li><li> Disminuir isoflurano al 1% y dejar inmóvil aguja durante 5 min. </li><li> Retire la aguja lentamente. </li><li> Aplique 1-2 gotas de 0,25% por vía subcutánea bupivacaína y suturar la piel. </li><li> Apague vaporizador isoflurano y quitar ratón del marco estereotáctico. </li><li> Permita ratón para recuperar la ventilación espontánea con extubación traqueal posterior. </li><li> Ratón Regreso a una jaula limpia y permitir el libre acceso aalimentos y agua. </li></ol> 3. Autólogo Modelo Inyección de sangre <ol><li> Siga los pasos de 02.01 a 02.11 para el modelo de inyección de colagenasa. </li><li> Dibujar 50 l de solución salina normal estéril en una jeringa de 30 l G 50. </li><li> Coloque la jeringa de microlitro con un tubo PE10 70 cm. </li><li> Expulsar todo la solución salina normal de la jeringa en el tubo de microlitro a un tubo de PE10-air de completamente. </li><li> Tire de la jeringa de pistón microlitro a cabo 1 mm para hacer una burbuja de aire en la abertura distal del aparato de jeringuilla de tubo de PE10-microlitro para evitar la mezcla de solución salina y la sangre durante los procedimientos posteriores. </li><li> Limpiar la región de la arteria central de la cola distal del ratón con etanol al 70%, y cortar la arteria con una maquinilla de afeitar de 0,5 a 1 cm a la punta de la cola. </li><li> Recoger 40 l de sangre de la cola cortada en el aparato de jeringuilla de tubo de microlitro-PE10. Nota: que la heparina no se utiliza en la aguja, tubo, o el ratón. </li><li> Conecte la jeringa de microlitro a la injebomba cción. </li><li> Conectar la porción de cánula de metal de una aguja de 27 G para el extremo del tubo de PE10, y asegurar la aguja a un micromanipulador en el marco estereotáctico. </li><li> Expulsar a 2 l de sangre de 27 G de la aguja y limpie bisel de la aguja con la maquinilla de afeitar de desprenderse. </li><li> Alinee la punta de la aguja con orificio de trepanación e inserte la aguja 3 mm de profundidad de la corteza. </li><li> Inyectar 35 l de sangre autóloga a razón de 2 l por min. </li><li> Disminuir isoflurano al 1% y dejar inmóvil aguja durante 10 min. </li><li> Retire la aguja durante 30 s. </li><li> Aplique 1-2 gotas de 0,25% por vía subcutánea bupivacaína y suturar la piel. </li><li> Apague vaporizador isoflurano y quitar ratón del marco estereotáctico. </li><li> Permita ratón para recuperar la ventilación espontánea con la posterior de la extubación. </li><li> Ratón Regreso a una jaula limpia y permitir el acceso libre a comida y agua. </li></ol> 4. Sham Operación <ol><li> Siga los mismos procedimientos para injectio colagenasamodelo n, excepto que sin la inyección después de la inserción de la aguja. </li></ol> 5. Cuidados después de la cirugía <ol><li> Inyectar 0,5 ml de solución salina normal por vía subcutánea en la noche de la intervención quirúrgica en la parte posterior del cuello del animal. </li><li> Proporcionar alimentos ablandada con agua y la comida en gel en pequeñas tazas de plástico colocadas en el suelo de la jaula. Vuelva a colocar la comida al día durante 7 días. </li><li> Compruebe si hay pérdida de peso, cicatrización de heridas y signos de malestar al día durante 7 días. </li><li> Si se requieren intervalos de recuperación de más de 7 días, retirada de la sutura se puede realizar bajo anestesia ligera con inhalada (aproximadamente 1% de isoflurano en 30% de O 2/70% N 2), si es necesario. </li></ol>

<ol>
	<li>Asch, C. 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=Incidence,+case+fatality,+and+functional+outcome+of+intracerebral+haemorrhage+over+time,+according+to+age,+sex,+and+ethnic+origin:+a+systematic+review+and+meta-analysis.">Incidence, case fatality, and functional outcome of intracerebral haemorrhage over time, according to age, sex, and ethnic origin: a systematic review and meta-analysis.</a> Lancet Neurology. 9, 167-176 (2010).</li><li>Qureshi, A. I., Mendelow, A. D., Hanley, D. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intracerebral+haemorrhage.">Intracerebral haemorrhage.</a> Lancet. 373, 1632-1644 (2009).</li><li>Participants, N. I. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Priorities+for+clinical+research+in+intracerebral+hemorrhage:+report+from+a+National+Institute+of+Neurological+Disorders+and+Stroke+workshop.">Priorities for clinical research in intracerebral hemorrhage: report from a National Institute of Neurological Disorders and Stroke workshop.</a> Stroke. 36, (2005).</li><li>Morgenstern, L. 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=Guidelines+for+the+management+of+spontaneous+intracerebral+hemorrhage:+a+guideline+for+healthcare+professionals+from+the+American+Heart+Association/American+Stroke+Association.">Guidelines for the management of spontaneous intracerebral hemorrhage: a guideline for healthcare professionals from the American Heart Association/American Stroke Association.</a> Stroke. 41, 2108-2129 (2010).</li><li>James, M. L., Warner, D. S., Laskowitz, D. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preclinical+models+of+intracerebral+hemorrhage:+a+translational+perspective.">Preclinical models of intracerebral hemorrhage: a translational perspective.</a> Neurocrit Care. 9, 139-152 (2008).</li><li>Winkler, D. 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=Spontaneous+hemorrhagic+stroke+in+a+mouse+model+of+cerebral+amyloid+angiopathy.">Spontaneous hemorrhagic stroke in a mouse model of cerebral amyloid angiopathy.</a> J Neurosci. 21, 1619-1627 (2001).</li><li>Sinar, E. J., Mendelow, A. D., Graham, D. I., Teasdale, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+intracerebral+hemorrhage:+effects+of+a+temporary+mass+lesion.">Experimental intracerebral hemorrhage: effects of a temporary mass lesion.</a> J Neurosurg. 66, 568-576 (1987).</li><li>Mendelow, A. D., Bullock, R., Teasdale, G. M., Graham, D. I., McCulloch, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intracranial+haemorrhage+induced+at+arterial+pressure+in+the+rat.+Part+2:+Short+term+changes+in+local+cerebral+blood+flow+measured+by+autoradiography.">Intracranial haemorrhage induced at arterial pressure in the rat. Part 2: Short term changes in local cerebral blood flow measured by autoradiography.</a> Neurol Res. 6, 189-193 (1984).</li><li>Bullock, R., Mendelow, A. D., Teasdale, G. M., Graham, D. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intracranial+haemorrhage+induced+at+arterial+pressure+in+the+rat.+Part+1:+Description+of+technique,+ICP+changes+and+neuropathological+findings.">Intracranial haemorrhage induced at arterial pressure in the rat. Part 1: Description of technique, ICP changes and neuropathological findings.</a> Neurol Res. 6, 184-188 (1984).</li><li>Sang, Y. H., Su, H. X., Wu, W. T., So, K. F., Cheung, R. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Elevated+blood+pressure+aggravates+intracerebral+hemorrhage-induced+brain+injury.">Elevated blood pressure aggravates intracerebral hemorrhage-induced brain injury.</a> J Neurotrauma. 28, 2523-2534 (2011).</li><li>Krafft, P. 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=Modeling+intracerebral+hemorrhage+in+mice:+injection+of+autologous+blood+or+bacterial+collagenase.">Modeling intracerebral hemorrhage in mice: injection of autologous blood or bacterial collagenase.</a> J Vis Exp. , (2012).</li><li>Sansing, L. H., 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=Autologous+blood+injection+to+model+spontaneous+intracerebral+hemorrhage+in+mice.">Autologous blood injection to model spontaneous intracerebral hemorrhage in mice.</a> J Vis Exp. , (2011).</li><li>Rosenberg, G. A., Mun-Bryce, S., Wesley, M., Kornfeld, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Collagenase-induced+intracerebral+hemorrhage+in+rats.">Collagenase-induced intracerebral hemorrhage in rats.</a> Stroke. 21, 801-807 (1990).</li><li>Nath, F. P., Jenkins, A., Mendelow, A. D., Graham, D. I., Teasdale, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early+hemodynamic+changes+in+experimental+intracerebral+hemorrhage.">Early hemodynamic changes in experimental intracerebral hemorrhage.</a> J Neurosurg. 65, 697-703 (1986).</li><li>Anderson, C. 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=Rapid+blood-pressure+lowering+in+patients+with+acute+intracerebral+hemorrhage.">Rapid blood-pressure lowering in patients with acute intracerebral hemorrhage.</a> N Engl J Med. 368, 2355-2365 (2013).</li><li>Clark, W., Gunion-Rinker, L., Lessov, N., Hazel, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Citicoline+treatment+for+experimental+intracerebral+hemorrhage+in+mice.">Citicoline treatment for experimental intracerebral hemorrhage in mice.</a> Stroke. 29, 2136-2140 (1998).</li><li>Mayer, S. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Efficacy+and+safety+of+recombinant+activated+factor+VII+for+acute+intracerebral+hemorrhage.">Efficacy and safety of recombinant activated factor VII for acute intracerebral hemorrhage.</a> N Engl J Med. 358, 2127-2137 (2008).</li><li>Mendelow, A. 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=Early+surgery+versus+initial+conservative+treatment+in+patients+with+spontaneous+supratentorial+lobar+intracerebral+haematomas+(STICH+II):+a+randomised+trial.">Early surgery versus initial conservative treatment in patients with spontaneous supratentorial lobar intracerebral haematomas (STICH II): a randomised trial.</a> Lancet. , (2013).</li><li>James, M. L., Blessing, R., Bennett, E., Laskowitz, D. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Apolipoprotein+E+modifies+neurological+outcome+by+affecting+cerebral+edema+but+not+hematoma+size+after+intracerebral+hemorrhage+in+humans.">Apolipoprotein E modifies neurological outcome by affecting cerebral edema but not hematoma size after intracerebral hemorrhage in humans.</a> J Stroke Cerebrovasc Dis. 18, 144-149 (2009).</li><li>James, M. L., Blessing, R., Phillips-Bute, B. G., Bennett, E., Laskowitz, D. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=S100B+and+brain+natriuretic+peptide+predict+functional+neurological+outcome+after+intracerebral+haemorrhage.">S100B and brain natriuretic peptide predict functional neurological outcome after intracerebral haemorrhage.</a> Biomarkers. 14, 388-394 (2009).</li><li>James, M. L., Sullivan, P. M., Lascola, C. D., Vitek, M. P., Laskowitz, D. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pharmacogenomic+effects+of+apolipoprotein+e+on+intracerebral+hemorrhage.">Pharmacogenomic effects of apolipoprotein e on intracerebral hemorrhage.</a> Stroke. 40, 632-639 (2009).</li><li>James, M. 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=Brain+natriuretic+peptide+improves+long-term+functional+recovery+after+acute+CNS+injury+in+mice.">Brain natriuretic peptide improves long-term functional recovery after acute CNS injury in mice.</a> J Neurotrauma. 27, 217-228 (2010).</li><li>Indraswari, F., 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=Statins+improve+outcome+in+murine+models+of+intracranial+hemorrhage+and+traumatic+brain+injury:+a+translational+approach.">Statins improve outcome in murine models of intracranial hemorrhage and traumatic brain injury: a translational approach.</a> J Neurotrauma. 29, 1388-1400 (2012).</li><li>Laskowitz, D. 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=The+apoE-mimetic+peptide,+COG1410,+improves+functional+recovery+in+a+murine+model+of+intracerebral+hemorrhage.">The apoE-mimetic peptide, COG1410, improves functional recovery in a murine model of intracerebral hemorrhage.</a> Neurocrit Care. 16, 316-326 (2012).</li><li>Lei, 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=Interaction+between+sex+and+apolipoprotein+E+genetic+background+in+a+murine+model+of+intracerebral+hemorrhage.">Interaction between sex and apolipoprotein E genetic background in a murine model of intracerebral hemorrhage.</a> Translational Stroke Research. 3, (2012).</li><li>Lekic, 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=Evaluation+of+the+hematoma+consequences,+neurobehavioral+profiles,+and+histopathology+in+a+rat+model+of+pontine+hemorrhage.">Evaluation of the hematoma consequences, neurobehavioral profiles, and histopathology in a rat model of pontine hemorrhage.</a> J Neurosurg. 118, 465-477 (2013).</li><li>Nakamura, 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=Intracerebral+hemorrhage+in+mice:+model+characterization+and+application+for+genetically+modified+mice.">Intracerebral hemorrhage in mice: model characterization and application for genetically modified mice.</a> J Cereb Blood Flow Metab. 24, 487-494 (2004).</li><li>Yang, 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=Statins+Protect+the+Blood+Brain+Barrier+Acutely+after+Experimental+Intracerebral+Hemorrhage.">Statins Protect the Blood Brain Barrier Acutely after Experimental Intracerebral Hemorrhage.</a> J Behav Brain Sci. 3, 100-106 (2013).</li><li>Rynkowski, M. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+mouse+model+of+intracerebral+hemorrhage+using+autologous+blood+infusion.">A mouse model of intracerebral hemorrhage using autologous blood infusion.</a> Nature Protocols. 3, 122-128 (2008).</li><li>Wang, J., Fields, J., Dore, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+development+of+an+improved+preclinical+mouse+model+of+intracerebral+hemorrhage+using+double+infusion+of+autologous+whole+blood.">The development of an improved preclinical mouse model of intracerebral hemorrhage using double infusion of autologous whole blood.</a> Brain Research. 1222, 214-221 (2008).</li><li>MacLellan, C. 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=Intracerebral+hemorrhage+models+in+rat:+comparing+collagenase+to+blood+infusion.">Intracerebral hemorrhage models in rat: comparing collagenase to blood infusion.</a> J Cereb Blood Flow Metab. 28, 516-525 (2008).</li></ol>

Lei, Sheng, Wang, Lascola, Warner, and Laskowitz have no conflicts of interest to declare. James received grant funding by American Heart Association, National Institutes of Health, and Cephalogics.

A pesar de que emerge la investigación preclínica y grandes ensayos clínicos resultantes de la promesa de la terapéutica de 15-18, no hay intervenciones farmacológicas demostradas para mejorar el resultado en el ICH, y la atención se mantiene en gran medida de apoyo. Las listas de posibles terapias pueden ser generados por tecnologías de alto rendimiento, como el trabajo transcriptómica y proteómica. Si bien estas tecnologías siguen avanzando nuestro conocimiento de posibles dianas terapéuticas, Tr...

La hemorragia intracerebral (HIC) es una forma relativamente común de enfermedad cerebrovascular con aproximadamente el 40-50% de los pacientes afectados de morir dentro de 30 días 1. Desafortunadamente, pocas mejoras se han hecho en la tasa de mortalidad en los últimos 20 años 2. Los informes de los Institutos Nacionales de Salud 3 y las directrices de la American Heart Association 4 subrayaron la importancia de desarrollar modelos clínicamente relevantes de ICH para ampliar la comprensión de la fisiopatología y desarrollar objetivos para nuevos enfoques terapéuticos. Existen varios modelos para imitar ICH humana 5. Como la comprensión de la fisiopatología de la ICH madura, se ha hecho evidente que una variedad de modelos puede ser utilizado para examinar diferentes aspectos de la enfermedad. Modelos utilizados anteriormente incluyen angiopatía amiloide murino 6, inserción microbalón intraparenquimatosa y la inflación 7, y la sangre arterial directainfiltración de 8,9. Lobar hemorragia de angiopatía amiloide se ha modelado con el uso de ratones transgénicos y representa un subtipo de la ICH distinta. Modelos microbalón imitan el efecto de masa aguda de la formación de hematomas, pero no logran captar la respuesta celular del cerebro a la presencia de sangre. Por último, la infiltración de sangre arterial directa somete el cerebro a las presiones arteriales de la arteria femoral. Por lo tanto, este modelo imita presiones arteriales y la presencia de sangre, pero no somete al cerebro a la lesión microvascular de pequeña ruptura de los vasos sanguíneos. Además, este modelo tiene inherentemente alta variabilidad. Curiosamente, las ratas espontáneamente hipertensas 10 desarrollan HIC espontánea a medida que envejecen. El estudio de estos animales después del desarrollo de la ICH puede imitar la enfermedad en presencia de uno de los principales comorbilidades que predisponen a los seres humanos de la ICH. Si bien existen estos otros modelos, la inyección de intraestriatal clostridial colagenasa 11 o inyección instrastiatal de unsangre entera utologous 12 son, en la actualidad, los dos modelos más comunes utilizados en la investigación preclínica ICH. La selección del modelo de la ICH debe hacerse con base en el objetivo de la pregunta experimental, incluyendo la selección y el método de inducir la formación de hematomas especies. Por ejemplo, los cerdos son animales grandes con volúmenes relativamente grandes de la materia blanca del cerebro en comparación con los ratones. Por lo tanto, son adecuados modelos porcinos para estudiar la materia blanca fisiopatología siguiente ICH. En contraste, los cerebros de roedores son materia principalmente de color gris, pero los sistemas transgénicos hacen roedores útil para evaluar los mecanismos moleculares de la lesión y la recuperación después de la ICH. Cada modelo tiene sus fortalezas y debilidades (Tabla 1) inherentes, que debe ser considerado cuidadosamente antes de la experimentación. Los siguientes protocolos demuestran los modelos de sangre y de inyección de colagenasa autólogas en ratones. Estos modelos han sido cada uno traducida de modelos desarrollados originalmente en ratas13,14 y permitir el uso de la tecnología transgénica ampliamente disponibles para explorar los mecanismos moleculares asociados con la muerte celular después de la ICH. Ambos representan claramente diferentes mecanismos de lesión de ICH humano, y ambos tienen muy diferentes resultados esperados en términos de medidas conductuales e histológicas. Por lo tanto, ciertas hipótesis pueden prestarse a un modelo sobre el otro, pero muchas ideas pueden requerir la validación en ambos modelos. Tabla 1. Comparación de las características de los modelos de hemorragia intracerebral inyección de sangre autóloga colagenasa-e. <table border="0" cellpadding="0" cellspacing="0"><tbody><tr><td style="width:197px;"></td><td style="width:80px;"> La inyección de colagenasa </td><td style="width:66px;"> Inyección de sangre </td></tr><tr><td style="width:197px;"> Facilidad de Uso </td><td style="width:80px;"> + + + </td><td style="width:66px;"> + + </td></tr><tr&gt; <td style="width:197px;"> Reproducibilidad </td><td style="width:80px;"> + + </td><td style="width:66px;"> + + </td></tr><tr><td style="width:197px;"> Control de la Hemorragia Tamaño </td><td style="width:80px;"> + + </td><td style="width:66px;"> + + + </td></tr><tr><td style="width:197px;"> El reflujo de sangre </td><td style="width:80px;"> + </td><td style="width:66px;"> + + </td></tr><tr><td style="width:197px;"> Simula Enfermedades Humanas </td><td style="width:80px;"> + </td><td style="width:66px;"> - </td></tr><tr><td style="width:197px;"> Sencillez </td><td style="width:80px;"> + + </td><td style="width:66px;"> + + </td></tr><tr><td style="width:197px;"> El uso en múltiples especies </td><td style="width:80px;"> + + </td><td style="width:66px;"> + + </td></tr></tbody></table>

<table border="0" cellpadding="0" cellspacing="0" width="416">
	<tbody>
		<tr height="43">
			<td height="43" style="height:43px;width:152px;">Name</td>
			<td style="width:88px;">Company</td>
			<td style="width:88px;">Catalog Number</td>
			<td style="width:88px;">Comments</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:152px;">Stereotactic frame</td>
			<td style="width:88px;">Stoelting Co.</td>
			<td style="width:88px;">51603</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:152px;">Probe holder with corner clamp</td>
			<td style="width:88px;">Stoelting Co.</td>
			<td style="width:88px;">51631</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:152px;">Mini grinder</td>
			<td style="width:88px;">Power Glide</td>
			<td style="width:88px;">Model 60100002</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:152px;">0.5 &#181;l Hamilton syringe</td>
			<td style="width:88px;">Hamilton Co.</td>
			<td style="width:88px;">86259</td>
			<td style="width:88px;">25 gauge needle</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:152px;">50 &#181;l Hamilton syringe</td>
			<td style="width:88px;">Hamilton Co</td>
			<td style="width:88px;">7637-01</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:152px;">26G Hamilton needle</td>
			<td style="width:88px;">Hamilton Co</td>
			<td style="width:88px;">7804-03</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:152px;">Syringe pump</td>
			<td style="width:88px;">KD Scientific</td>
			<td style="width:88px;">Model 100</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:152px;">Heat therapy water pump</td>
			<td style="width:88px;">Gaymar Industries, Inc.</td>
			<td style="width:88px;">Model# TP650</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:152px;">Circulating waterbed</td>
			<td style="width:88px;">CMS Tool &#38; Die, Inc.</td>
			<td style="width:88px;"></td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:152px;">Rodent ventilator</td>
			<td style="width:88px;">Harvard Apparatus</td>
			<td style="width:88px;">Model 683</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:152px;">Isoflurane vaporizer</td>
			<td style="width:88px;">Drager</td>
			<td style="width:88px;">Vapor 19.1</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:152px;">Air flowmeter</td>
			<td style="width:88px;">Cole Parmer</td>
			<td style="width:88px;">Model PMR1-010295</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:152px;">Induction chamber</td>
			<td style="width:88px;"></td>
			<td style="width:88px;"></td>
			<td style="width:88px;">Self made</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:152px;">Otoscope</td>
			<td style="width:88px;">Welch Allyn</td>
			<td style="width:88px;">22820</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:152px;">intravenous catheter</td>
			<td style="width:88px;">Becton-Dickinson</td>
			<td style="width:88px;">381534</td>
			<td style="width:88px;">20-gauge, 1.16 inch Insyte-W</td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:152px;">Isoflurane</td>
			<td style="width:88px;">Baxter Healthcare Corporation</td>
			<td style="width:88px;">NDC10019-360-69</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:152px;">Collagenase Type IV-S</td>
			<td style="width:88px;">Sigma</td>
			<td style="width:88px;">C1889</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:152px;">Polyethylene tubing PE20</td>
			<td style="width:88px;">Becton-Dickinson</td>
			<td style="width:88px;">427406</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:152px;">Polyethylene tubing PE10</td>
			<td style="width:88px;">Becton-Dickinson</td>
			<td style="width:88px;">427401</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:152px;">30G 1 inch needle</td>
			<td style="width:88px;">Becton-Dickinson</td>
			<td style="width:88px;">305128</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:152px;">27G 1 1/4 inch needle</td>
			<td style="width:88px;">Becton-Dickinson</td>
			<td style="width:88px;">305136</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:152px;">Surgical scissors</td>
			<td style="width:88px;">Miltex</td>
			<td style="width:88px;">21-539</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:152px;">Forceps</td>
			<td style="width:88px;">Miltex</td>
			<td style="width:88px;">17-307</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:152px;">Needle holder</td>
			<td style="width:88px;">Boboz</td>
			<td style="width:88px;">RS-7840</td>
			<td style="width:88px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:152px;">Monofilament suture</td>
			<td style="width:88px;">Ethicon</td>
			<td style="width:88px;">8698</td>
			<td style="width:88px;">Size 5-0</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:152px;">Indicating controller</td>
			<td style="width:88px;">YSI</td>
			<td style="width:88px;">73ATD</td>
			<td style="width:88px;"></td>
		</tr>
	</tbody>
</table>

intrastriatal injection of autologous blood or clostridial collagenase as murine models of intracerebral hemorrhage

La hemorragia intracerebral (ICH) es una forma común de enfermedad cerebrovascular y se asocia con morbilidad y mortalidad significativas. La falta de tratamiento y el fracaso efectivo de grandes ensayos clínicos dirigidos a la hemostasia y la eliminación de coágulos demostrar la necesidad de una mayor investigación mecanismo impulsado de ICH. Esta investigación se puede realizar a través del marco establecido por los modelos preclínicos. Dos modelos murinos en uso popular incluyen (ganglios basales) inyección intraestriatal de sangre entera autóloga o colagenasa clostridial. Dado que, cada modelo representa claramente diferentes características fisiopatológicas relacionadas con la ICH, el uso de un modelo en particular se puede seleccionar sobre la base de qué aspecto de la enfermedad es a ser estudiado. Por ejemplo, la inyección de sangre autóloga representa con mayor precisión la respuesta del cerebro a la presencia de sangre intraparenquimatosa, y puede reproducir más de cerca posible hemorragia lobar. Inyección de colagenasa clostridial representa con mayor precisión las sruptura de los vasos centro comercial y hematoma evolución característico de hemorragias profundas. Por lo tanto, cada uno de los resultados del modelo en diferente formación de hematomas, neuroinflammatory respuesta, el desarrollo de edema cerebral, y los resultados neuroconductuales. La robustez de una intervención terapéutica pretendida puede ser mejor evaluada usando ambos modelos. En este protocolo, la inducción de ICH con modelos de demostración, post-operatorio inmediato de la lesión, y las técnicas de atención postoperatoria temprana se demuestran. Ambos modelos resultan en lesiones reproducibles, volúmenes hematoma, y ​​déficits neuroconductuales. Debido a la heterogeneidad de ICH humana, se necesitan varios modelos preclínicos para explorar a fondo los mecanismos fisiopatológicos y probar estrategias terapéuticas potenciales.

Debido a las diferencias en la formación de un hematoma (Figura 1), giro ipsilateral se muestra inmediatamente después de despertar para los ratones inyectados de sangre autóloga y dentro de 2 a 4 horas después de la inyección de colagenasa, como la expansión del hematoma se produce (Figura 2). Ausencia de giro ipsilateral debe plantear la preocupación por la ausencia de lesiones significativas. En el primer día después de la lesión, los ratones en ambos modelos deben demostra...

Watch this Scientific Journal Video about Intrastriatal Injection of Autologous Blood or Clostridial Collagenase as Murine Models of Intracerebral Hemorrhage at JoVE.com

Intrastriatal Injection of Autologous Blood or Clostridial Collagenase as Murine Models of Intracerebral Hemorrhage

Preclinical models of intracerebral hemorrhage are utilized to mimic certain aspects of clinical disease. Thus, mechanisms of injury and potential therapeutic strategies may be explored. In this protocol, two models of intracerebral hemorrhage are described, intrastriatal (basal ganglia) injections of autologous blood or collagenase.

Inyección intraestriatal de sangre autóloga o clostridial colagenasa como modelos murinos de la hemorragia intracerebral

Intracerebral hemorrhage (ICH) is a common form of cerebrovascular disease and is associated with significant morbidity and mortality. Lack of effective ...

intrastriatal-injection-autologous-blood-or-clostridial-collagenase

Research

JoVE Journal

Medicine

14.6K vistas.  Duke University. Preclinical models of intracerebral hemorrhage are utilized to mimic certain aspects of clinical disease. Thus, mechanisms of injury and potential therapeutic strategies may be explored. In this protocol, two models of intracerebral hemorrhage are described, intrastriatal (basal ganglia) injections of autologous blood or collagenase. 

Video: Intrastriatal Injection of Autologous Blood or Clostridial Collagenase as Murine Models of Intracerebral Hemorrhage

Fixed Volume or Fixed Pressure: A Murine Model of Hemorrhagic Shock

It is common knowledge that severe blood loss and traumatic injury can lead to a cascade of detrimental signaling events often resulting in mortality. 1, 2, 3, 4, 5 These signaling events can also lead to sepsis and/or multiple organ dysfunction (MOD). 6, 7, 8, 9 It is critical then to investigate the causes of suppressed immune function and detrimental signaling cascades in order to develop more effective ways to help patients who suffer from traumatic injuries. 10 This fixed pressure Hemorrhagic Shock (HS) procedure, although technically challenging, is an excellent resource for investigation of these pathophysiologic conditions. 11, 12, 13 Advances in the assessment of biological systems, i.e. Systems Biology have enabled the scientific community to further understand complex physiologic networks and cellular communication patterns. 14 Hemorrhagic Shock has proven to be a vital tool for unveiling these cellular communication patterns as they relate to immune function. 15, 16, 17, 18 This procedure can be mastered! This procedure can also be used as either a fixed volume or fixed pressure approach. We adapted this technique in the murine model to enhance research in innate and adaptive immune function. 19, 20, 21 Due to their small size HS in mice presents unique challenges. However due to the many available mouse strains, this species represents an unparalleled resource for the study of the biologic responses. The HS model is an important model for studying cellular communication patterns and the responses of systems such as hormonal and inflammatory mediator systems, and danger signals, i.e. DAMP and PAMP upregulation as it elicits distinct responses that differ from other forms of shock. 22, 23, 24, 25 The development of transgenic murine strains and the induction of biologic agents to inhibit specific signaling have presented valuable opportunities to further elucidate our understanding of the up and down regulation of signal transduction after severe blood loss, i.e. HS and trauma 26, 27, 28, 29, 30. 
There are numerous resuscitation methods (R) in association with HS and trauma. 31, 32, 33, 34 A fixed volume resuscitation method of solely lactated ringer solution (LR), equal to three times the shed blood volume, is used in this model to study endogenous mechanisms such as remote organ injury and systemic inflammation. 35, 36, 38 This method of resuscitation is proven to be effective in evaluating the effects of HS and trauma 38, 39.

The Hemorrhagic Shock model has been a reliable and reproducible resource facilitating the identification and understanding of signaling cascades associated with inflammation and end-organ damage after trauma. This article provides a step-by-step description of surgical and mechanical aspects associated with the Hemorrhagic Shock experimental procedure in mice.

Volumen fijo o fijado a presión: un modelo murino de shock hemorrágico

It is common knowledge that severe blood loss and traumatic injury can lead to a cascade of detrimental signaling events often resulting in mortality. 1, ...

Investigación

Medicina

Coronary Artery Ligation and Intramyocardial Injection in a Murine Model of Infarction

Mouse models are a valuable tool for studying acute injury and chronic remodeling of the myocardium in vivo. With the advent of genetic modifications to the whole organism or the myocardium and an array of biological and/or synthetic materials, there is great potential for any combination of these to assuage the extent of acute ischemic injury and impede the onset of heart failure pursuant to myocardial remodeling. 
Here we present the methods and materials used to reliably perform this microsurgery and the modifications involved for temporary (with reperfusion) or permanent coronary artery occlusion studies as well as intramyocardial injections. The effects on the heart that can be seen during the procedure and at the termination of the experiment in addition to histological evaluation will verify efficacy. 
Briefly, surgical preparation involves anesthetizing the mice, removing the fur on the chest, and then disinfecting the surgical area. Intratracheal intubation is achieved by transesophageal illumination using a fiber optic light. The tubing is then connected to a ventilator. An incision made on the chest exposes the pectoral muscles which will be cut to view the ribs. For ischemia/reperfusion studies, a 1 cm piece of PE tubing placed over the heart is used to tie the ligature to so that occlusion/reperfusion can be customized. For intramyocardial injections, a Hamilton syringe with sterile 30gauge beveled needle is used. When the myocardial manipulations are complete, the rib cage, the pectoral muscles, and the skin are closed sequentially. Line block analgesia is effected by 0.25% marcaine in sterile saline which is applied to muscle layer prior to closure of the skin. The mice are given a subcutaneous injection of saline and placed in a warming chamber until they are sternally recumbent. They are then returned to the vivarium and housed under standard conditions until the time of tissue collection. At the time of sacrifice, the mice are anesthetized, the heart is arrested in diastole with KCl or BDM, rinsed with saline, and immersed in fixative. Subsequently, routine procedures for processing, embedding, sectioning, and histological staining are performed. 
Nonsurgical intubation of a mouse and the microsurgical manipulations described make this a technically challenging model to learn and achieve reproducibility. These procedures, combined with the difficulty in performing consistent manipulations of the ligature for timed occlusion(s) and reperfusion or intramyocardial injections, can also affect the survival rate so optimization and consistency are critical.

Numerous genetic manipulations and/or intramyocardial injections of genes, proteins, cells, and/or biomaterials are superimposed upon the dimension of time in studies of acute ischemia/ reperfusion injury and chronic remodeling in mice. This video illustrates the microsurgical procedures for ischemia/reperfusion, permanent coronary artery ligation, and intramyocardial injection studies.

Ligadura de las arterias coronarias y la inyección intramiocárdica en un modelo murino de infarto

Mouse models are a valuable tool for studying acute injury and chronic remodeling of the myocardium in vivo.  With the advent of genetic modifications to ...

Modeling Intracerebral Hemorrhage in Mice: Injection of Autologous Blood or Bacterial Collagenase

Spontaneous intracerebral hemorrhage (ICH) defines a potentially life-threatening neurological malady that accounts for 10-15% of all stroke-related hospitalizations and for which no effective treatments are available to date1,2. Because of the heterogeneity of ICH in humans, various preclinical models are needed to thoroughly explore prospective therapeutic strategies3. Experimental ICH is commonly induced in rodents by intraparenchymal injection of either autologous blood or bacterial collagenase4. The appropriate model is selected based on the pathophysiology of hemorrhage induction and injury progression. The blood injection model mimics a rapidly progressing hemorrhage. Alternatively, bacterial collagenase enzymatically disrupts the basal lamina of brain capillaries, causing an active bleed that generally evolves over several hours5. Resultant perihematomal edema and neurofunctional deficits can be quantified from both models. In this study, we described and evaluated a modified double injection model of autologous whole blood6 as well as an ICH injection model of bacterial collagenase7, both of which target the basal ganglia (corpus striatum) of male CD-1 mice. We assessed neurofunctional deficits and brain edema at 24 and 72 hr after ICH induction. Intrastriatal injection of autologous blood (30 &mu;l) or bacterial collagenase (0.075U) caused reproducible neurofunctional deficits in mice and significantly increased brain edema at 24 and 72 hr after surgery (p&lt;0.05). In conclusion, both models yield consistent hemorrhagic infarcts and represent basic methods for preclinical ICH research.

Clinically relevant animal models of intracerebral hemorrhage (ICH) are needed to extend our knowledge of hemorrhagic stroke and to examine novel therapeutic strategies. In this study, we describe and evaluate two ICH models that implement unilateral injections of either autologous whole blood or bacterial collagenase into the basal ganglia (corpus striatum) of mice.

Modelado Hemorragia intracerebral en ratones: La inyección de sangre autóloga o colagenasa bacteriana

Spontaneous intracerebral hemorrhage (ICH) defines a potentially life-threatening neurological malady that accounts for 10-15% of all stroke-related ...

A Murine Model of Subarachnoid Hemorrhage

In this video publication a standardized mouse model of subarachnoid hemorrhage (SAH) is presented. Bleeding is induced by endovascular Circle of Willis perforation (CWp) and proven by intracranial pressure (ICP) monitoring. Thereby a homogenous blood distribution in subarachnoid spaces surrounding the arterial circulation and cerebellar fissures is achieved. Animal physiology is maintained by intubation, mechanical ventilation, and continuous on-line monitoring of various physiological and cardiovascular parameters: body temperature, systemic blood pressure, heart rate, and hemoglobin saturation. Thereby the cerebral perfusion pressure can be tightly monitored resulting in a less variable volume of extravasated blood. This allows a better standardization of endovascular filament perforation in mice and makes the whole model highly reproducible. Thus it is readily available for pharmacological and pathophysiological studies in wild type and genetically altered mice.

A standardized mouse model of subarachnoid hemorrhage by intraluminal Circle of Willis perforation is described. Vessel perforation and subarachnoid bleeding are monitored by intracranial pressure monitoring. In addition various vital parameters are recorded and controlled to maintain physiologic conditions.

Un modelo murino de hemorragia subaracnoidea

In this video publication a standardized mouse model of subarachnoid  hemorrhage (SAH) is presented. Bleeding is induced by endovascular Circle of  Willis ...

Stereological and Flow Cytometry Characterization of Leukocyte Subpopulations in Models of Transient or Permanent Cerebral Ischemia

Microglia activation, as well as extravasation of haematogenous macrophages and neutrophils, is believed to play a pivotal role in brain injury after stroke. These myeloid cell subpopulations can display different phenotypes and functions and need to be distinguished and characterized to study their regulation and contribution to tissue damage. This protocol provides two different methodologies for brain immune cell characterization: a precise stereological approach and a flow cytometric analysis. The stereological approach is based on the optical fractionator method, which calculates the total number of cells in an area of interest (infarcted brain) estimated by a systematic random sampling. The second characterization approach provides a simple way to isolate brain leukocyte suspensions and to characterize them by flow cytometry, allowing for the characterization of microglia, infiltrated monocytes and neutrophils of the ischemic tissue. In addition, it also details a cerebral ischemia model in mice that exclusively affects brain cortex, generating highly reproducible infarcts with a low rate of mortality, and the procedure for histological brain processing to characterize infarct volume by the Cavalieri method.

Brain myeloid cells characterization following stroke can be performed by stereology using the optical fractionator method, or by a flow cytometric analysis of brain leukocytes suspensions. Both are useful techniques to perform an accurate phenotypical distinction of the main myeloid cell subsets found in the ischemic brain.

Estereológico y Citometría de Flujo Caracterización de leucocitos subpoblaciones en Modelos de transitorios o isquemia cerebral permanente

Microglia activation, as well as extravasation of haematogenous macrophages and neutrophils, is believed to play a pivotal role in brain injury after ...

The Rabbit Blood-shunt Model for the Study of Acute and Late Sequelae of Subarachnoid Hemorrhage: Technical Aspects

Early brain injury and delayed cerebral vasospasm both contribute to unfavorable outcomes after subarachnoid hemorrhage (SAH). Reproducible and controllable animal models that simulate both conditions are presently uncommon. Therefore, new models are needed in order to mimic human pathophysiological conditions resulting from SAH.
This report describes the technical nuances of a rabbit blood-shunt SAH model that enables control of intracerebral pressure (ICP). An extracorporeal shunt is placed between the arterial system and the subarachnoid space, which enables examiner-independent SAH in a closed cranium. Step-by-step procedural instructions and necessary equipment are described, as well as technical considerations to produce the model with minimal mortality and morbidity. Important details required for successful surgical creation of this robust, simple and consistent ICP-controlled SAH rabbit model are described.

The experimental intracranial pressure-controlled blood shunt subarachnoid hemorrhage (SAH) model in the rabbit combines the standard procedures &#8212; subclavian artery cannulation and transcutaneous cisterna magna puncture, which enables close mimicking of human pathophysiological conditions after SAH. We present step-by-step instructions and discuss key surgical points for successful experimental SAH creation.

El Conejo Modelo Blood-shunt para el Estudio de la aguda y secuelas tardías de Hemorragia subaracnoidea: Aspectos Técnicos

Early brain injury and delayed cerebral vasospasm both contribute to unfavorable outcomes after subarachnoid hemorrhage (SAH). Reproducible and ...

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, ...

Ultrasound-guided Intracardiac Injection of Human Mesenchymal Stem Cells to Increase Homing to the Intestine for Use in Murine Models of Experimental Inflammatory Bowel Diseases

Crohn&#39;s disease (CD) is a common chronic inflammatory disease of the small and large intestines. Murine and human mesenchymal stem cells (MSCs) have immunosuppressive potential and have been shown to suppress inflammation in mouse models of intestinal inflammation, even though the route of administration can limit their homing and effectiveness 1,3,4,5. Local application of MSCs to colonic injury models has shown greater efficacy at ameliorating inflammation in the colon. However, there is paucity of data on techniques to enhance the localization of human bone marrow-derived MSCs (hMSCs) to the small intestine, the site of inflammation in the SAMP-1/YitFc (SAMP) model of experimental Crohn&#39;s disease. This work describes a novel technique for the ultrasound-guided intracardiac injection of hMSCs in SAMP mice, a well-characterized spontaneous model of chronic intestinal inflammation. Sex- and age-matched, inflammation-free AKR/J (AKR) mice were used as controls. To analyze the biodistribution and the localization, hMSCs were transduced with a lentivirus containing a triple reporter. The triple reporter consisted of firefly luciferase (fl), for bioluminescent imaging; monomeric red fluorescent protein (mrfp), for cell sorting; and truncated herpes simplex virus thymidine kinase (ttk), for positron emission tomography (PET) imaging. The results of this study show that 24 h after the intracardiac administration, hMSCs localize in the small intestine of SAMP mice as opposed to inflammation-free AKR mice. This novel, ultrasound-guided injection of hMSCs in the left ventricle of SAMP mice ensures a high success rate of cell delivery, allowing for the rapid recovery of mice with minimal morbidity and mortality. This technique could be a useful method for the enhanced localization of MSCs in other models of small-intestinal inflammation, such as TNF&#916;RE6. Future studies will determine if the increased localization of hMSCs by intra-arterial delivery can lead to increased therapeutic efficacy.

Murine studies in models of colonic inflammation have demonstrated that a small percentage (1 - 5%) of mesenchymal stem cells (MSC) injected intravenously or intraperitoneally home to the inflamed colon1,2. This study shows that ultrasound-guided intracardiac injections of MSCs result in increased localization to the intestine.

Guiado por ecografía intracardiaca inyección de células madre mesenquimales humanas para aumentar autoguiado hacia el blanco en el intestino para su uso en modelos murinos de enfermedades de intestino inflamatorias experimentales

Crohn&#39;s disease (CD) is a common chronic inflammatory disease of the small and large intestines. Murine and human mesenchymal stem cells (MSCs) have ...

Nerve Stimulator-guided Injection of Autologous Stem Cells Near the Equine Left Recurrent Laryngeal Nerve

Recurrent laryngeal neuropathy (RLN) commonly affects horses and is characterized by abnormal respiratory sounds and exercise intolerance. The recurrent laryngeal nerve shows lesions of demyelination. The benefit of applying stem cells to demyelinated nerves has been demonstrated in various animal models. The aim of the study was to test the feasibility and safety of a peri-neuronal injection of autologous muscle-derived mesenchymal stem cells to the left recurrent laryngeal nerve in healthy horses by using an electrical nerve stimulator.
Muscle-derived stems cell are obtained from five healthy Standardbred horses by sampling 20 mg of muscle tissue with a semi-automatic 14 G biopsy needle from the triceps muscle. Movements of the larynx are monitored via upper-airway video endoscopy. The left recurrent laryngeal nerve is approached with an insulated nerve block needle. Nerve stimulation is applied, starting at 2 mA, and the successful abduction of the left arytenoid is monitored. The stimulation intensity is reduced progressively. When a loss of the motor response is observed at 0.5 mA, 107 autologous muscle-derived stem cells are injected. Two examiners, who are blinded to the time point, score the laryngeal function of the horses prior to the treatment and at day 1, day 7, and day 28 after the injection of the cells. In a sixth horse, 1 mL of 2% lidocaine is injected to further confirm the correct positioning of the needle. This leads to a temporary paralysis of the left arytenoid cartilage.
This study proves that the recurrent laryngeal nerve can be approached with the help of an electrical nerve stimulator and that the electrical stimulation of the nerve is well tolerated by the horses. No modification of the laryngeal function was observed in any of the horses after the injection of the stem cells. Further studies should be conducted to describe the effects of a peri-neuronal injection of autologous muscle-derived mesenchymal stem cells to horses suffering from RLN.

Here, we present a protocol to inject autologous muscle-derived stem cells in the proximity to the recurrent laryngeal nerve with the help of an electrical nerve stimulator. This novel technique may become useful for the treatment of equine recurrent laryngeal neuropathy.

Nervio estimulador guiado por la inyección de células madre autólogas cerca de nervio laríngeo recurrente izquierdo equino

Recurrent laryngeal neuropathy (RLN) commonly affects horses and is characterized by abnormal respiratory sounds and exercise intolerance. The recurrent ...

Subclavian Vein Puncture As an Alternative Method of Blood Sample Collection in Rats

Blood collection with enough blood volume is essential in animal experiments. Blood collection from the tail vein of rats is popular and less stressful compared to other more aggressive methods such as retro-orbital plexus sample collection. However, this blood collection method is sometimes limited by an unsatisfactory success rate. Here, we introduce a method for blood collection through the subclavian vein puncture. The subclavian vein is located just under the clavicle and this vein is large enough to fulfill the volume requirements of blood collection. Our results show that this method is safe and applicable for blood collection sampling with the required blood volume. Blood collection through the subclavian vein puncture could serve as an alternative blood collection method in case of failed tail vein blood sampling in rats.

Here, we present a protocol to collect blood samples from the rat subclavian vein.

Punción de la vena subclavia como método alternativo de recogida de muestras de sangre en las ratas

Blood collection with enough blood volume is essential in animal experiments. Blood collection from the tail vein of rats is popular and less stressful ...