Name: ferric chloride-induced murine thrombosis models
Uploaded: 2016-09-05T13:00:00
Description: Watch this Scientific Journal Video about Ferric Chloride-induced Murine Thrombosis Models at JoVE.com

This work was supported by the National Heart Lung and Blood Institute (NHLBI) of the National Institutes of Health under award numbers R01 HL121212 (PI: Sen Gupta), R01 HL129179 (PI: Sen Gupta, Co-I: Li) and R01 HL098217 (PI: Nieman). The content of this publication is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Todos los procedimientos y manipulaciones de los animales han sido aprobados por Institucional Cuidado de Animales y el empleo Comités (IACUC) de la Clínica de Cleveland, en conformidad con la política de Estados Unidos Servicio de Salud Pública en el cuidado humano y Uso de Animales, y la Guía de los NIH para el Cuidado y uso de Animales de laboratorio. 1. Preparación: <ol><li> Tinte fluorescente para el etiquetado de plaquetas <ol><li> Preparar la so...

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El FeCl3 inducida modelo es uno de los modelos de trombosis más ampliamente usados, que no sólo pueden proporcionar información valiosa acerca de las modificaciones genéticas sobre la función plaquetaria y trombosis 7,8,16,19,31-33, pero también puede ser una herramienta valiosa para la evaluación de compuestos terapéuticos y estrategias para el tratamiento y prevención de enfermedades aterotrombóticos 11,17,34-37. Aquí hemos demostrado que nuestros modificaciones y refinamien...

La trombosis arterial (coágulos de sangre) es una complicación común de muchas enfermedades sistémicas asociadas con la inflamación crónica, incluyendo la aterosclerosis, la diabetes, la obesidad, el cáncer y los trastornos autoinmunes reumatológicas crónicas. Los trombos que se presentan en el tronco circulación arterial de la activación plaquetaria inadecuada, agregación y mecanismos procoagulantes posteriores, y están implicados en los ataques cardíacos, accidentes cerebrovasculares y la pérdida de la extremidad. La pared del vaso es un sistema complejo que incluye múltiples tipos de células y está influenciada por una multitud de factores extrínsecos como el estrés de cizallamiento, las células de la sangre, hormonas y citoquinas, así como la expresión de proteínas antioxidantes que circula en la pared del vaso. Experimentos in vitro no pueden replicar este entorno complejo y por lo tanto en estudios in vivo utilizando modelos animales son críticos para permitir una mejor comprensión de los mecanismos implicados en los trastornos trombóticos. Los ratones han demostrado tener simmecanismos ILAR a los seres humanos en términos de la trombosis, aterosclerosis, inflamación y diabetes 1,2. Además, los ratones transgénicos y knockout pueden ser creados para probar la función de los productos génicos específicos en un complejo fisiológico o patológico ambiente. Tales estudios imitan la patología humana y pueden proporcionar información sobre el mecanismo importante relacionada con el descubrimiento de nuevas vías y terapias, así como proporcionar detalles importantes en la caracterización de los efectos de la droga sobre la trombosis. Trombos arteriales patológicas se producen debido a la lesión endotelial capa o disfunción y la exposición de la corriente de la sangre a la matriz subendotelial 3,4. Varios modelos de trombosis se han desarrollado para inducir este daño endotelial tales como la lesión mecánica, lesión oxidativa a base de Bengala compuesto fotorreactivo Rose y lesiones láser 5. En este espectro, cloruro férrico (FeCl3) inducida por la lesión vascular es un modelo ampliamente utilizado de trombosis. Este reactivo cuandose aplica a la cara externa de los vasos induce daño oxidativo a las células vasculares 6-8, con pérdida de la protección de las células endoteliales a partir de plaquetas y componentes de la cascada de coagulación circulantes. El modelo de FeCl 3 es simple y sensible tanto a anticoagulante y anti-plaquetas medicamentos, y se ha realizado en arterias carótida y femoral, vena yugular, y mesentérica y arteriolas y vénulas cremastéricos en ratones, ratas, cobayas y conejos 6-15. Un parámetro medible en este modelo es el tiempo transcurrido desde la lesión hasta completar la oclusión del vaso, medida como cese el flujo de sangre con un medidor de flujo Doppler o bajo la observación directa con 6,7,9 microscopía intravital. Una gama de tiempos entre 5 a 30 min se ha informado en diferentes estudios en ratones C57Bl6 7-10,16, lo que sugiere que FeCl 3 concentraciones, tipos de anestesia, las técnicas quirúrgicas, la edad del ratón, fondo genómico, método de medición de bLood flujo, y otras variables ambientales tienen efectos significativos en este modelo. Esta amplia variabilidad hace que sea difícil comparar los estudios de diferentes grupos de investigación y puede hacer la detección de diferencias sutiles difícil. Con una visión para minimizar tales variabilidades y establecer un reproducible uniformemente en el sistema de modelo in vivo, hemos refinado el modelo de arteria carótida inducida por FeCl3 que hace que los datos altamente reproducibles con una variación mínima 6-10,16-19. En este trabajo se describen y compartir las habilidades y presenta varios ejemplos experimentales representativos que pueden beneficiarse de este modelo.

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			<td height="40" style="height:40px;width:351px;">Metzenbaum Fino Scissors - Tungsten Carbide/Curved/Blunt-Blunt/14.5cm</td>
			<td style="width:159px;">Fine Science Tools&#160;</td>
			<td style="width:119px;">14519-14&#160;&#160;</td>
			<td>to dissect and separate soft tissue</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:351px;">Ultra Fine Hemostat - Smooth/Curved/12.5cm</td>
			<td style="width:159px;">Fine Science Tools&#160;</td>
			<td>13021-12</td>
			<td>to dissect and separate soft tissue</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Graefe Forceps - Serrated/Straight/10cm</td>
			<td style="width:159px;">Fine Science Tools&#160;</td>
			<td style="width:119px;">11050-10</td>
			<td>to dissect and separate soft tissue</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:351px;">Dumont #5 Fine Forceps - Biology Tips/Straight/Inox/11cm</td>
			<td style="width:159px;">Fine Science Tools&#160;</td>
			<td>11254-20&#160;</td>
			<td>Isolate vessel from surounding tissue</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:351px;">Dumont #5XL Forceps - Standard Tips/Straight/Inox/15cm</td>
			<td style="width:159px;">Fine Science Tools&#160;</td>
			<td>11253-10</td>
			<td>Isolate vessel from surounding tissue</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:351px;">Blunt Hook- 12cm/0.3mm Tip Diameter</td>
			<td style="width:159px;">Fine Science Tools&#160;</td>
			<td>10062-12</td>
			<td>Isolate vessel from surounding tissue</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:351px;">Castroviejo Micro Needle Holders</td>
			<td style="width:159px;">Fine Science Tools&#160;</td>
			<td>12061-02</td>
			<td>Needle holders</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:351px;">Suture Thread 4-0</td>
			<td style="width:159px;">Fine Science Tools&#160;</td>
			<td>18020-40</td>
			<td>For fix the incisors to the plate</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:351px;">Suture Thread 6-0</td>
			<td style="width:159px;">Fine Science Tools&#160;</td>
			<td>18020-60</td>
			<td>For all surgery and ligation</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:351px;">Kalt Suture Needles</td>
			<td style="width:159px;">Fine Science Tools&#160;</td>
			<td>12050-03</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">rhodamine 6G&#160;</td>
			<td style="width:159px;">Sigma</td>
			<td>83697-1G</td>
			<td>To lebel platelets</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;">FeCl3 (Anhydrous)</td>
			<td style="width:159px;">Sigma</td>
			<td style="width:119px;">12321</td>
			<td>To induce vessel injury</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Papaverine hydrochloride</td>
			<td style="width:159px;">Sigma</td>
			<td>P3510</td>
			<td>To inhibit gut peristalsis.</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:351px;">Medline Surgical Instrument Sterilization Steam Autoclave Tapes</td>
			<td style="width:159px;">Medline</td>
			<td>111625</td>
			<td>To fix the mouse to the plate</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:351px;">Fisherbrand&#8482;&#160;Syringe Filters - Sterile 0.22&#181;m</td>
			<td style="width:159px;">Fisher</td>
			<td>09-720-004</td>
			<td>For sterlization of solutions injected to mice</td>
		</tr>
		<tr height="26">
			<td height="26" style="height:26px;width:351px;">Fisherbrand&#8482;&#160;Syringe Filters - Sterile 0.45&#181;m</td>
			<td style="width:159px;">Fisher</td>
			<td>09-719D</td>
			<td>To filter the FeCl3 solution</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:351px;">Sterile Alcohol Prep Pad</td>
			<td style="width:159px;">Fisher</td>
			<td>06-669-62</td>
			<td>To sterilize the surgical site</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:351px;">Agarose&#160;</td>
			<td style="width:159px;">BioExpress</td>
			<td>E-3120-500</td>
			<td>To make gel stage</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:351px;">Leica DMLFS fluorescent microscope</td>
			<td style="width:159px;">Leica</td>
			<td style="width:119px;"></td>
			<td>Intravital microscope</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:351px;">GIBRALTAR Platform and X-Y Stage System</td>
			<td style="width:159px;">npi electronic GmbH</td>
			<td style="width:119px;"></td>
			<td style="width:325px;">http://www.npielectronic.de/products/micropositioners/burleigh/gibraltar.html</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:351px;">Streampix version 3.17.2 software</td>
			<td style="width:159px;">NorPix</td>
			<td style="width:119px;"></td>
			<td>https://www.norpix.com/</td>
		</tr>
	</tbody>
</table>

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.

La trombosis de la arteria carótida Modelo En ratones C57BL6 con el fondo, se recomienda utilizar 7,5% FeCl3 para el tratamiento de la embarcación durante 1 min como punto de partida. Bajo tratamiento de 7,5% FeCl3, las fronteras de la zona lesionada y la pared vascular normal se identifican fácilmente bajo el microscopio (Ver video en línea 1), lo que sugiere que la capa endotelial fue dañado de manera significativa. El...

Watch this Scientific Journal Video about Ferric Chloride-induced Murine Thrombosis Models at JoVE.com

Ferric Chloride-induced Murine Thrombosis Models

Se presenta un procedimiento de refinado del cloruro férrico (FeCl3) modelos de trombosis inducidas en la carótida y la arteria mesentérica, así como la vena, que se caracteriza de manera eficiente mediante microscopía intravital para monitorear el tiempo de oclusiva formación de trombos.

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, ...

Research

JoVE Journal

Medicine

Electrolytic Inferior Vena Cava Model (EIM) of Venous Thrombosis

Electrolytic Inferior Vena Cava Model EIM of Venous Thrombosis

Electrolítico de la vena cava inferior del modelo (EIM) de la Trombosis Venosa

Animal models serve a vital role in deep venous thrombosis (DVT) research in order to study thrombus formation, thrombus resolution and to test potential ...

Investigación

Medicina

Mouse Complete Stasis Model of Inferior Vena Cava Thrombosis

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 ...

Segmentation and Measurement of Fat Volumes in Murine Obesity Models Using X-ray Computed Tomography

La segmentación y la medición de los volúmenes de grasa en modelos murinos de obesidad Usando rayos X de tomografía computarizada

Obesity is associated with increased morbidity and mortality as well as reduced metrics in quality of life.1 Both environmental and genetic factors are ...

Quantitative Analysis and Characterization of Atherosclerotic Lesions in the Murine Aortic Sinus

Atherosclerosis is a disease of the large arteries and a major underlying cause of myocardial infarction and stroke. Several different mouse models have ...

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

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 ...

Orthotopic Injection of Breast Cancer Cells into the Mammary Fat Pad of Mice to Study Tumor Growth.

Inyección ortotópico de células de cáncer de mama en el Mamaria Fat Pad de ratones para estudiar el crecimiento del tumor.

Breast cancer growth can be studied in mice using a plethora of models. Genetic manipulation of breast cancer cells may provide insights into the ...

Adapting Human Videofluoroscopic Swallow Study Methods to Detect and Characterize Dysphagia in Murine Disease Models

Adaptación videofluoroscópicos Swallow Métodos de Estudio Humanos para detectar y caracterizar disfagia en modelos murinos de enfermedades

This study adapted human videofluoroscopic swallowing study (VFSS) methods for use with murine disease models for the purpose of facilitating ...

High-frequency Ultrasound Imaging of Mouse Cervical Lymph Nodes

Alta frecuencia de imágenes por ultrasonido del mouse ganglios linfáticos cervicales

High-frequency ultrasound (HFUS) is widely employed as a non-invasive method for imaging internal anatomic structures in experimental small animal ...

A Multicenter MRI Protocol for the Evaluation and Quantification of Deep Vein Thrombosis

Un Protocolo MRI Multicéntrico para la Evaluación y cuantificación de la trombosis venosa profunda

We evaluated a magnetic resonance venography (MRV) approach with gadofosveset to quantify total thrombus volume changes as the principal criterion for ...

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

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 ...