作者要感谢来自喜悦姚明和卡伦博士Alt键的技术支持，以及资金从NHMRC和NHF。

所有涉及的动物实验均批准了阿尔弗雷德医学研究和教育专用区动物伦理委员会（E /二千○十五分之一千五百三十四/ B）。全身麻醉下进行所有的手术操作和动物没有在任何阶段经历痛苦。描述的所有实验都是非恢复。 1.准备<ol><li>切细带的滤纸（1毫米×2mm）的。 </li><li>新鲜制备2溶液的4％氯化铁（重量/体积）和6％（重量/体积）稀释到去离子水中。制备罗丹明6G溶液0.3％在PBS中，通过0.22微米的过滤。 </li><li>从切包装注射器的白色塑料小片（5毫摩×1厘米）。 </li></ol> 2.肠系膜小动脉血栓形成活体显微镜观察<ol><li>权衡10-12周大的雄性C57BL / 6野生型小鼠和用氯胺酮虽然腹腔注射的混合物（100毫克/千克）和赛拉嗪（10毫克/千克）相应麻醉。通过响应于趾，尾和/或皮肤捏，从角膜和眼睑捏反应性，并且不存在晶须运动监测麻醉深度。如果需要的话，注射第二剂量的氯胺酮（50毫克/千克），以保持动物麻醉。取适量涂抹于眼睛兽医药膏，以防止干燥，而在麻醉下。鼠标放置在一个小的培养皿放置在一个加热垫调节至37℃。 注意:虽然IP注射可导致疼痛的动物，这种不适的持续时间将是最小的（小于3秒）。与注射相关的疼痛和不适会通过使用有经验和能力的人员和适当的针头大小（25克）的最小化。下面所有的程序，使用安乐死氯胺酮和甲苯噻嗪颈椎脱位过量所有的动物。 </li><li>在皮肤进行约3厘米的腹部正中切口，并仔细切腹膜。 </li><li>将鼠标上的宠物的侧里的菜，轻轻exteriorize它的肠子小心地肠系膜与2棉花棒把合适的容器到培养皿的表面。一个微妙的雨刷干正常。 注意:要限制的肠系膜血管的移动，罂粟碱可用于抑制肠道蠕动。 </li><li>浸泡的小鼠尾巴中温水以扩张血管和注入30微升若丹明6G（0.3％）的尾静脉鼠标用29克注射器标记白细胞和血小板。 </li><li>放置在培养皿倒置显微镜下，专注于使用明场通道选择的动脉。 </li><li>浸泡的滤纸的带用6％（重量/体积）的铁（III），氯化并应用滤纸上用两个镊子动脉;第一个以保持滤纸，第二个要轻轻按下它到感兴趣的区域。以下滤纸的沉积观察血栓形成在第一10秒。 注:这是很通讯上可破坏包围微血管和滤纸的沉积和轻轻压力的精度，因此重要的是限制这个问题。 </li><li>观察血栓形成由荧光显微镜（TRITC通道:峰激发557纳米，峰值发射576纳米），通过滤纸。观察循环白细胞和血小板已经采取了罗丹明6G和他们聚集到血栓因此，很容易辨认。 </li><li>曝光1分钟至铁（III），氯化后取滤纸关闭，并继续监测血栓的形成。洗用PBS的容器中。 </li><li>观察并记录动态形成突出了血小板和白细胞与罗丹明6G的标签血栓。捕获的图像，并测量血栓的大小。本文中所呈现的图像被用倒置显微镜活体获得，虽然客观4X，在TRITC荧光通道。 </li><li> Following所有的程序，使用安乐死氯胺酮和甲苯噻嗪颈椎脱位过量的动物。 </li></ol> 3.颈动脉血栓形成评估的血流速度测定<ol><li>权衡10-12周龄雄性C57B1 / 6老鼠和氯胺酮（100毫克/千克）和赛拉嗪（10毫克/千克）虽然腹腔注射的混合物相应地麻醉。取适量涂抹于眼睛兽医药膏，以防止干燥，而在麻醉下。 </li><li>固定鼠标使用胶带上腿在手术显微镜下，在加热垫上调节至37℃。 </li><li>做一个小枕头了一个擦，并用胶带鼠标的头下稍微垫高头部。使用一个线程循环用钳子拉下来吻（使用上牙）。这将会使颈动脉的区域，便于使用。 </li><li>执行皮肤的小5毫米深的切口正下方的颚，下至胸骨。 </li><li>解剖足协SCIA和隔离左或右颈总动脉分叉以上的片段。 </li><li>精心引进之间的动脉和神经把它们分开镊子。请勿打扰运行接近动脉的神经并避免接触过多的颈动脉，因为它可能造成损害的容器中。隔离的至少5mm动脉的一个部分。 </li><li>干燥动脉的区域正确地与雨刷避免任何液体干扰的FeCl 3。 </li><li>把下颈使氯化铁的分离部分的白色小塑料片在周围组织不浸泡。为了这个目的，使用第二钳子以使片至第一个再慢慢滑动动脉下的塑料纸。 </li><li>浸泡一片滤纸，用4％（重量/体积）或6％（重量/体积）氯化铁，并把它的所有动脉周围。 </li><li> 3分钟后曝光，取下滤纸，冲洗用PBS和干燥区瓦特第i个雨刷。 </li><li>放置在容器周围的多普勒流量探针在受伤部位和开始记录的变化的流动。成年小鼠的健康颈，流程通常为约1毫升/分钟。注意！探针与氯化铁接触会损坏探针所以应避免任何接触。在本文中所呈现的数据被用一个跨音速系统公司流量计，TS420血管周围模块配备有纳米多普勒流量探针0.5的PBS获得。 注:氯化铁的浓度可以修改的血栓形成导致不同的闭塞倍的动力学。因此，暴露于6％（重量/体积）氯化铁给出更快的闭塞比暴露于4％（重量/体积）的氯化铁。 </li><li>下面所有的程序，用氯胺酮和甲苯噻嗪的颈椎脱位过量安乐死的所有动物，仔细清洁多普勒探头。 </li></ol>

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J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Plasminogen+activator+inhibitor-1+and+its+cofactor+vitronectin+stabilize+arterial+thrombi+after+vascular+injury+in+mice.">Plasminogen activator inhibitor-1 and its cofactor vitronectin stabilize arterial thrombi after vascular injury in mice.</a> Circulation. 103 (4), 576-583 (2001).</li><li>Li, W., McIntyre, T. M., Silverstein, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ferric+chloride-induced+murine+carotid+arterial+injury:+A+model+of+redox+pathology.">Ferric chloride-induced murine carotid arterial injury: A model of redox pathology.</a> Redox Biol. 1 (1), 50-55 (2013).</li></ol>

The authors have nothing to disclose.

氯化铁诱发血栓模型是一个很好的研究工具。如该研究中，这是非常容易实现，并且与活体显微镜或多普勒流量计结合使用时，它提供了血栓形成一个良好的实时监控。调整时间曝光和的FeCl 3的浓度，它也提供了，以产生非闭塞性或闭塞性血栓的可能性。 但是，这种方法也有一定的局限性。在颈动脉中，主要的缺点是，虽然阻塞时间可以有效地被修改，则该模型的再...

参与血栓的发展和抗血栓药物的有效性的评价机制的研究需要很好地建立实验动物模型。大型动物模型是第一个被使用，因为它们提供大血管更类似于人类比啮齿类1。然而，成本高，需要较大的设施和操纵他们的困难是基因主要缺点它们的使用和大型动物，现在仅限于后期临床前研究，一旦对啮齿动物的初步试验给出结论性的结果2。用转基因和基因敲除株和它们的小尺寸，最大限度地减少所需的体内测试抗血栓药物的量广泛的可用性，小鼠主要用于血栓形成研究。因此，已经开发了在小鼠中3几种型号血栓形成性疾病的。 许多老牌血栓模型扰乱intim层的血管壁，然后由子内皮细胞外基质到血流诱导血块4的形成的曝光。血栓可能导致胶原的曝光这触发血小板活化和/或从组织因子的曝光其激活凝血级联5。然后几种技术被用来实现初始血管损伤。 Pierangeli 等人开发了一种机械破碎模型上的股静脉6一个显微工具。 Kikushi 等人描述了一种模型，该模型包括在相片反应性化合物（玫瑰红），其积聚在血管内皮细胞随后用绿光（540纳米）7感兴趣的血管壁的特定激励的脂质双层的施用。这次受伤也可以通过短高强度的脉冲激光照射诱发8。另一种技术首先对大鼠的颈动脉既定包括在氯化铁的局部施用（的FeCl 3）9。在这种情况下，从通过的FeCl 3产生自由基的容器剥蚀结果导致内皮细胞10的脂质过氧化和破坏。的损伤引起几个粘附分子触发血小板粘附和聚集以及白细胞募集的表达。它已经证明，白细胞，特别是嗜中性粒细胞，在血液凝固级联导致血栓形成11的活化中发挥至关重要的作用。这种方法非常适合于重现凝血级联;研究者必须牢记的是，在该小鼠模型中，血栓形成通常诱导健康血管而血栓在人类中是主要发生在患病如 。动脉粥样硬化的血管。 因为此模型是非常简单的实现，也是有效的小鼠，它现在是最常用血栓形成模式l对于小动物体内研究。此外，这种技术提供了以诱导在各种血管血栓形成的可能性。靶血管可以是动脉或大直径（颈动脉，股腔静脉）或小直径（肠系膜，提睾肌）12-14脉。最近，也有人用在大脑中动脉近端制定行程15的模型。血栓形成可以直接血小板和白细胞的荧光标记后通过活体显微镜观察或通过测量血流量减少与温度探针或多普勒探针12,16,17监测。几个参数，如阻断时间，血栓形成时间或血栓大小然后可以进行调查。容器之间的生理差异中获得的血栓显著变化调查结果。因此，调查人员根据他们想measu参数通常选择目标船只重新和/或疾病的设置，他们希望调查。通常，在颈动脉的模型是更相关的动脉粥样硬化与心肌梗死或中风而上腔静脉的研究更相关于深静脉血栓形成研究研究。不同的容器的访问也决定用来衡量血栓增长的方法。例如，肠系膜血管很容易就可以访问使得该模型适合于活体显微镜观察和血栓形成的动力学的研究。颈动脉是不太容易，但做大，使血流量的测量，并提供研究血栓闭塞一个很好的模式。 氯化铁诱发血栓模型在这病理的认识提供了巨大的进步。它已被用于在许多研究集中于血管性血友病因子在血栓形成18,19的作用。结合遗传MODIfication技术中，已经允许参与血栓形成病症的许多特定基因的鉴定。拉姆拉尼等 。例如已经表明，一敲在JAK2 V617F基因都与一个加速形成不稳定凝块20的相关联。 Zhang 等人调查了该P2Y12血小板受体的生理含义，并表明转基因小鼠过度表达特别这种受体在血小板只，显示了更快速，稳定的血栓形成的肠系膜动脉的FeCl 3 21受伤。的组织型纤溶酶原激活剂（tPA）和尿激酶型纤溶酶原激活物（uPA）的纤维蛋白降解过程中的关键作用也进行了研究在该方法22。此外，该模型还提供了测试的许多新颖的药物的血纤维蛋白溶解能力在体内的简单且精确的方式。例如，Wang 等人已经用这种模型对于第靶向激活的血小板23的新型重组纤溶酶原激活电子临床验证。这种方法也使治疗性蛋白蜱，吸血蝙蝠，和蚊子的唾液或蛇与目标24-27的特定标识的毒液中分离的验证。这些实施例说明了氯化铁模型的多功能性。在这篇文章中，我们专注于两种方法，研究在两个不同的容器类型三氯化铁诱发血栓形成;肠系膜血管和颈动脉。

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

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

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

荧光活体显微镜观察肠系膜将揭示罗丹明6G的积累标记的血小板和白细胞沿着血管壁由三氯化铁3人受伤。渐进形成局部血栓在200μm的肠系膜血管（ 图1）进行监测。血栓慢慢出现，显然是暴露的第一分钟到氯化铁后可识别（ 图1，T = 60秒）。 40秒的去除浸泡的FeCl 3滤纸之后，血栓形成迅速的进展，并且最后存在于所观察到的整个容器部分的壁（ <str...

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

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

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

三氯化铁诱发血栓形成小鼠模型上的颈动脉和肠系膜血管

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

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

Research

JoVE Journal

Medicine

17.5K 次观看.  Baker IDI Heart and Diabetes Institute. The FeCl3 induced thrombosis model in mice is described herein. A method to monitor thrombus growth by intravital microscopy observation on a mesenteric vessel and by blood flow measurement in the carotid artery is presented.

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

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

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

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

一个型号的不安流致动脉粥样硬化小鼠颈动脉部分结扎术和颈动脉内皮细胞的RNA分离的简单方法

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

科研

医学

Mouse Complete Stasis Model of Inferior Vena Cava Thrombosis

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

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

鼠标完成血瘀模型的下腔静脉血栓

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

Mouse Model of Middle Cerebral Artery Occlusion

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

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

大脑中动脉闭塞的小鼠模型

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

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

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

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

颈动脉支架植入再狭窄的研究的小鼠模型

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

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

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

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

成年小鼠静脉高压型号:颈总动脉，以颈外静脉吻合。

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

Carotid Artery Infusions for Pharmacokinetic and Pharmacodynamic Analysis of Taxanes in Mice

When proposing the use of a drug, drug combination, or drug delivery into a novel system, one must assess the pharmacokinetics of the drug in the study model. As the use of mouse models are often a vital step in preclinical drug discovery and drug development1-8, it is necessary to design a system to introduce drugs into mice in a uniform, reproducible manner. Ideally, the system should permit the collection of blood samples at regular intervals over a set time course. The ability to measure drug concentrations by mass-spectrometry, has allowed investigators to follow the changes in plasma drug levels over time in individual mice1, 9, 10. In this study, paclitaxel was introduced into transgenic mice as a continuous arterial infusion over three hours, while blood samples were simultaneously taken by retro-orbital bleeds at set time points. Carotid artery infusions are a potential alternative to jugular vein infusions, when factors such as mammary tumors or other obstructions make jugular infusions impractical. Using this technique, paclitaxel concentrations in plasma and tissue achieved similar levels as compared to jugular infusion. In this tutorial, we will demonstrate how to successfully catheterize the carotid artery by preparing an optimized catheter for the individual mouse model, then show how to insert and secure the catheter into the mouse carotid artery, thread the end of the catheter out through the back of the mouse&#8217;s neck, and hook the mouse to a pump to deliver a controlled rate of drug influx. Multiple low volume retro-orbital bleeds allow for analysis of plasma drug concentrations over time.

This method was developed with the goal of delivering a steady drug solution via the carotid artery, to assess the pharmacokinetics of novel drugs in mouse models.

颈动脉输液的紫杉烷类化合物在小鼠体内的药代动力学和药效学分析

When proposing the use of a drug, drug combination, or drug delivery into a novel system, one must assess the pharmacokinetics of the drug in the study ...

Vascular Balloon Injury and Intraluminal Administration in Rat Carotid Artery

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

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

血管球囊损伤和腔内管理在大鼠颈动脉

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

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.

三氯化铁诱导小鼠模型的血栓形成

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

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

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

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

氯化铁诱导的犬颈动脉血栓形成: 一种大的血管损伤动物模型

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

Ferric Chloride-Induced Arterial Thrombosis and Sample Collection for 3D Electron Microscopy Analysis

Cardiovascular diseases are a leading cause of mortality and morbidity worldwide. Aberrant thrombosis is a common feature of systemic conditions like diabetes and obesity, and chronic inflammatory diseases like atherosclerosis, cancer, and autoimmune diseases. Upon vascular injury, usually the coagulation system, platelets, and endothelium act in an orchestrated manner to prevent bleeding by forming a clot at the site of the injury. Abnormalities in this process lead to either excessive bleeding or uncontrolled thrombosis/insufficient antithrombotic activity, which translates into vessel occlusion and its sequelae. The FeCl3-induced carotid injury model is a valuable tool in probing how thrombosis initiates and progresses in vivo. This model involves endothelial damage/denudation and subsequent clot formation at the injured site. It provides a highly sensitive, quantitative assay to monitor vascular damage and clot formation in response to different degrees of vascular damage. Once optimized, this standard technique can be used to study the molecular mechanisms underlying thrombosis, as well as the ultrastructural changes in platelets in a growing thrombus. This assay is also useful to study the efficacy of antithrombotic and antiplatelet agents. This article explains how to initiate and monitor FeCl3-induced arterial thrombosis and how to collect samples for analysis by electron microscopy.

The present protocol describes how to use a FeCl3-mediated injury to induce arterial thrombosis, and how to collect and prepare arterial injury samples at various stages of thrombosis for electron microscopy analysis.

氯化铁诱导的动脉血栓形成和用于3D电子显微镜分析的样本采集

Cardiovascular diseases are a leading cause of mortality and morbidity worldwide. Aberrant thrombosis is a common feature of systemic conditions like ...