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 os procedimentos e manipulações de animais foram aprovados pelo Institutional Animal Care e Uso Comissões (IACUC) da The Cleveland Clinic, em conformidade com a política dos Estados Unidos Serviço de Saúde Pública sobre o cuidado humano e uso de animais, e o Guia NIH para o Cuidado e uso de Animais de laboratório. 1. Preparações: <ol><li> Corante fluorescente para a rotulagem Plaquetas <ol><li> Preparar solução de rodamina 6G, 0,5 mg / ml, ...

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O FeCl3 induzida pelo modelo é um dos modelos mais amplamente utilizados de trombose, que não só pode fornecer informações valiosas sobre as modificações genéticas na função das plaquetas e trombose 7,8,16,19,31-33, mas também pode ser uma ferramenta valiosa para a avaliação de compostos terapêuticos e estratégias para o tratamento e prevenção das doenças aterotrombóticas 11,17,34-37. Aqui temos mostrado nossas modificações e aperfeiçoamentos do presente modelo e mo...

trombose arterial (coágulo de sangue) é uma complicação comum de muitas doenças sistêmicas associadas à inflamação crônica, incluindo a aterosclerose, diabetes, obesidade, câncer e doenças reumatológicas auto-imunes crônicas. Trombos que ocorrem no tronco circulação arterial a partir inadequado ativação plaquetária, agregação e mecanismos coagulante subsequentes e estão implicados em ataques cardíacos, derrames e perda de extremidade. A parede do vaso é um sistema complexo que inclui vários tipos de células e é influenciada por uma multitude de factores extrínsecos, incluindo tensão de corte, as células sanguíneas, hormonas e citocinas, bem como a expressão de proteínas antioxidantes que circula na parede do vaso. Experiências in vitro não pode replicar neste ambiente complexo e, por conseguinte, em estudos in vivo utilizando modelos animais são críticos para permitir uma melhor compreensão dos mecanismos envolvidos em doenças trombóticas. Os ratos têm mostrado ter SIMmecanismos lhantes aos seres humanos em termos de trombose, aterosclerose, inflamação e diabetes 1,2. Além disso, camundongos transgênicos e knockout podem ser criados para testar a função de produtos de genes específicos em uma fisiológica complexa ou ambiente patológico. Tais estudos imitar patologia humana e pode fornecer informações mecanicista importante relacionado à descoberta de novos caminhos e terapias, bem como fornecer detalhes importantes na caracterização de efeitos de drogas sobre a trombose. Trombos arteriais patológicas ocorrer devido à lesão endotelial camada ou disfunção e a exposição do fluxo de sangue para a matriz subendotelial 3,4. Vários modelos de trombose foram desenvolvidos para induzir esse dano endotelial tais como dano mecânico, dano oxidativo baseado em Bengal composto fotorreativa Rose e ferimentos de laser 5. Neste espectro, cloreto férrico (FeCl3) induzida por lesão vascular é um modelo amplamente usado de trombose. Este reagente quandoaplicada ao aspecto exterior dos navios induz danos oxidativos para células vasculares 6-8, com a perda da protecção de células endoteliais a partir de plaquetas e componentes da cascata de coagulação circulantes. O modelo FeCl3 é simples e sensível para ambos os anticoagulantes e anti-plaquetas drogas, e foi realizada em artérias carótida e femoral, veias jugulares e mesentérica e arteríolas cremastérico e vênulas em ratinhos, ratos, cobaias e coelhos 6-15. Um parâmetro mensurável neste modelo é o tempo decorrido de uma lesão para completar oclusão do vaso, medida como a cessação do fluxo sanguíneo com um medidor de fluxo de Doppler ou sob observação directa com 6,7,9 microscopia intravital. Uma gama de tempos de entre 5 a 30 min tem sido relatada em diferentes estudos em ratinhos C57BL6 7-10,16, sugerindo que as concentrações de FeCl 3, tipos de anestesia, técnicas cirúrgicas, idade rato, fundo genómico, método de medição bfluxo lood, e outras variáveis ​​ambientais têm efeitos significativos neste modelo. Esta ampla variabilidade faz com que seja difícil comparar estudos de diferentes grupos de pesquisa e pode fazer a detecção de diferenças sutis difícil. Com uma visão para minimizar tais variabilidades e estabelecer um uniforme reprodutível em sistema de modelo vivo, temos refinado o modelo de artéria carótida induzida por FeCl3 que faz com que os dados altamente reproduzíveis com o mínimo de variação 6-10,16-19. Neste artigo descrevemos e compartilhar as habilidades e relatar vários exemplos experimentais representativos que podem beneficiar a partir deste modelo.

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			<td style="width:325px;">cut and hold skin</td>
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			<td style="width:159px;">Fine Science Tools&#160;</td>
			<td>11018-12</td>
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			<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.

Carótida Thrombosis Modelo Em ratos com C57BL6 fundo, recomendamos a utilização de 7,5% FeCl3 para tratar o navio durante 1 min como ponto de partida. Sob o tratamento de 7,5% FeCl3, fronteiras da área lesada e parede normal do vaso são facilmente identificados sob o microscópio (Veja o vídeo on-line 1), sugerindo que a camada endotelial foi significativamente danificada. Os trombos formados imediatamente após Fe...

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

Ferric Chloride-induced Murine Thrombosis Models

Nós relatamos um procedimento refinada do cloreto férrico (FeCl3) modelos de trombose induzidas na carótida e mesentérica artéria, bem como veia, caracterizado de forma eficiente através de microscopia intravital para monitorar tempo para oclusiva formação de trombos.

Férricos induzida pelo cloreto modelos murinos Trombose

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

Eletrolítico Veia Cava Inferior Model (EIM) de trombose 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 ...

Medicina

Mouse Complete Stasis Model of Inferior Vena Cava Thrombosis

Modelo do mouse completa de Stasis Inferior Trombose da veia cava

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

Segmentação e medição de volumes de gordura em modelos de obesidade em roedores usando raios-X Tomografia Computadorizada

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

Injection intraestriatal de sangue autólogo ou Clostridial colagenase como modelos murinos de 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.

Injeção Orthotopic de células cancerosas da mama na mamária Fat Pad de Ratos para o Estudo de crescimento tumoral.

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

Adaptação Humanos videofluoroscópicos Andorinha Métodos de Estudo de detectar e caracterizar Disfagia em modelos de doenças Murino

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

De alta frequência Ecografia de mouse linfonodos cervicais

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

A Multicenter MRI Protocolo para a Avaliação e Quantificação de Trombose 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

Férrico Thrombosis Modelo rato na artéria carótida e Mesentery Vessel induzida por Chloride

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