我们要感谢俄亥俄州立大学认知和行为脑成像中心的财政和科学支持, 以发展和执行犬磁共振成像。

所述调查符合《国家卫生研究院护理和使用实验动物指南》, 并获得俄亥俄州立大学机构动物保育和使用委员会 (#2015A00000029) 的批准。所有手术操作都是在深部麻醉下进行的, 在手术过程中, 动物在任何阶段都没有疼痛。所描述的所有实验都是不恢复的。
1. 准备
<ol><li>新鲜地准备50毫升氯化铁 (FeCl3) 在 50% (瓦特/v) 稀释在去离子水。</li>
	<li>将脐带切成5.5 厘米片, 浸泡在新配制的 50% FeCl3溶液中, 在手术前1周。 注: 使用的脐胶带厚, 需要一些时间来吸收 50% FeCl3溶液。</li>
	<li>每只狗在手术前一夜禁食。</li>
	<li>标签 cryovials 为血浆和尿收集, 50 毫升无菌离心机管为所有器官存贮, 并且10% 个福尔马林容器为脑和颈动脉固定为 H & E 染色与动物辨认、出生日期、手术日期和治疗。</li>
	<li>为每条警犬的手术准备100毫升: 4% 多聚甲醛磷酸盐缓冲盐水 (ph 7.4), 10% 中性缓冲福尔马林, 2% 23, 5-triphenyltetrazolium 氯化物 [在 PBS (pH 7.4), 存储在黑暗中]。</li>
	<li>获取充足的液氮为所有 cryovials 和50毫升组织管为闪光冷冻在过程的天。</li>
	<li>获得两个 ADP/胶原蛋白盒为 PFA-100 (血小板功能分析仪) 血小板反应性为每只犬每一次的兴趣点。</li>
	<li>获得4.5 毫升枸橼酸钠采血管, 为每一个血液绘制前牺牲, 为血浆分离和储存。在牺牲, 绘制16血液收集管为血浆存贮。每次采血管满后, 三十年代旋转 4000 x g, 隔离血小板丰富的血浆, 将透明层转移到 cryovials。在贮存前-80 摄氏度的液态氮气中的闪光冻结。</li>
	<li>获得一个4.5 毫升的锂肝素采血管, 为每个 PFA-100 时间点记录血小板反应性。在每次采血时, 将800µL 的全血从一根锂肝素采血管引入到每一个 ADP/胶原蛋白盒中, 而没有气泡来跟随血小板反应。</li>
	<li>获得两个 edta K3 采血管 (1.8 毫克无水 edta 每1毫升血液), 为完整的血液计数 (CBC) 在基线和在牺牲的时候。 注: 本卷对于大多数核心设施来说都是足够的, 在机械错误的情况下每两次运行一次。虽然在受伤前的基线和牺牲时都运行了 CBC, 但调查人员可以在动物协议中增加更多的血液, 以进行额外的分析。检查设备的容量和交货要求。</li>
	<li>准备一个2毫升血气注射器的填充和涂层注射器的日期的过程中的血液气体分析在牺牲的时候。 注: 检查设备的容量和交货要求。</li>
</ol>2. 犬颈动脉闭塞
<ol><li>称成人 (> 18 月) 比格犬和肌内乙酰丙嗪 (0.025-2 毫克/千克) 前用药, 其次是最初腹腔注射氯胺酮 (5-20 毫克/千克) 和咪达唑仑 (0.01-25 毫克/千克)。根据心率、呼吸和颚音, 诱导1-5% 异氟醚深层麻醉。 注: 确切的麻醉是由重量和批准的动物协议决定的。虽然注射可能会导致动物疼痛, 但这种不适的持续时间将是最小的 (少于3秒)。</li>
	<li>在把动物带到手术桌前, 通过修剪来去除毛发。将润滑眼膏应用于麻醉动物的眼睛, 以防止干燥。</li>
	<li>使用定位系统将动物置于仰卧位, 并附上心电图 (ECG) 导致脚垫。插管通过气管与6.5 毫米袖口气管插管, 机械通气在14呼吸/分钟, 并保持每只狗持续吸入1-5% 异氟醚 (取决于批准的动物协议)。</li>
	<li>为了确定颈动脉的疏导程度和闭塞血栓的长度, 在受伤前进行血管造影, MRI 前, 并在牺牲时 (见下文以了解其他说明)。</li>
	<li>插入一个股动脉鞘和一个塑料导管, 并确保与松散的0丝缝合。在运输和影像学研究中暂时关闭鞘和导管。请参阅下面的犬血管造影附加说明)。</li>
	<li>对于血液绘制, 插入一个 16G x 45 毫米静脉导管通过完整的皮肤远端的股骨动脉鞘切断现场。将导管插入手术暴露中, 并将其可视化为与动脉鞘相邻的外露股静脉。一旦完全插入静脉, 取出针, 用3路旋塞阀将导管盖上, 用2毫升0.9% 无菌生理盐水冲洗。用0丝缝线将导管固定在皮肤上。</li>
	<li>在右颈总动脉区的顶端直接做一个8-10 厘米的皮肤切口, 解剖筋膜, 将血管与周围组织隔离。</li>
	<li>小心地引入钳颈动脉与迷走神经之间的分离。避免接触颈动脉超过必要的, 因为它可能导致血管收缩。</li>
	<li>用无菌纱布干燥动脉外露区域, 避免 FeCl3溶液稀释。</li>
	<li>在颈动脉周围放置多普勒血流探针, 允许缓行空间在颈动脉周围包裹脐带, 而不接触流探针, 开始记录血液流速并继续贯穿整个过程 (图 1 顶部)。在应用脐胶带之前, 记录基线流量为 ~ 5 分钟。 注: FeCl3脐放置前的基线流量通常平均约250毫升/分钟。</li>
	<li>将准备好的 FeCl3脐带绕颈动脉, 使用止血直接低于流探针, 不接触它15分钟, 删除和丢弃。 注: 遮挡被指定为零流量毫升/分钟。</li>
	<li>稳定闭塞血栓45分钟. 在探头中添加额外的盐水以保持信号。 注: 根据年龄、性别和犬类的不同, 在重新应用前, 可能需要额外放置准备好的 FeCl3脐胶带, 使30分钟的闭塞发生。在猎犬中, 氯化铁的应用与性别在血栓形成中没有发现差异。没有发现有必要用这个模型多次重新应用新鲜的 FeCl3脐磁带, 从来没有偏离 FeCl3脐准备协议。如果实验室使用的是不同品种或年龄的犬, FeCl3脐胶带应适用于最初的主题, 以确保血栓形成协议不偏离, 或一组犬可能需要建立 FeCl3该品种或年龄在其具体研究中的应用时间。在这个实验中, 遮挡时间从 FeCl3脐胶带应用后的8-30 分钟不等。</li>
	<li>为了确定中风或出血量, 除了可能导致药物干预的下游血栓事件, 运输犬与氯胺酮注射液的磁共振成像机 (见下文附加处理)。</li>
	<li>虽然仍然在一个从最后的血管造影/MRI (见下文为成像协议) 的麻醉深度手术飞机, 表明对有害刺激的反应, 收集全血所需的后期分析, 然后打开胸部切开通过肋骨从胸骨开始, 通过切割血管 (放血) 来切除心脏。 注: 在本实验中, 在基线、5分钟后用药后, 从股鞘灌注60分钟后, 全血 (< 10 毫升) 提取, 对溶栓过程无影响。在牺牲时, 如果在动物协议中得到批准, 可以为额外的等离子存储绘制无限量的体积。在30分钟的牺牲中分析、处理和储存全部血液。确保血液不超过所批准的动物体重的核定比例超过批准的动物议定书。所描述的所有实验都是不恢复的。</li>
	<li>如果需要的话, 在液氮中用0.9% 盐水和闪光冷冻样品冲洗心脏。</li>
	<li>在移除颈动脉之前, 通过在它旁边放置一个标尺并记录来测量血块的长度。将组织标记放在血栓中央, 并放置在与侧颈动脉相同的位置, 以便于组织学修整。用同一长度标记对侧颈动脉。 注: 这个模型的平均血栓长度为1.75 厘米。</li>
	<li>清除两个血管内的血栓的整个长度, 并修复10% 福尔马林。在组织标记标记的长度内移除对侧颈动脉。将对侧颈动脉在血栓中央的一半处切开。闪光冷冻半侧颈动脉在液氮后分析和固定的另一半10% 福尔马林嵌入在受伤的颈动脉下面的组织学分析。</li>
	<li>取出器官进行任何需要的分析, 用0.9% 盐水冲洗, 并在液氮中快速冷冻。</li>
	<li>用圆骨锯取出头骨。慢慢移除头骨而不损害大脑下方。</li>
	<li>大脑从头骨中取出后, 用冰冷的0.9% 盐水冲洗, 从大脑中间切开两个4毫米的切片, 用锋利的手术刀横向切割。将其中的4毫米部分放入2% 个 TTC 为缺血性分界 (参见其他处理为脑子 ttc), 而另一片放置10% 福尔马林7天为 H & e 染色 (参见另外的处理为脑子 h & e)。</li>
</ol>3. 犬血管造影
注意: 这如图 2所示, 是在手术期间的时间点进行的。
<ol><li>感觉在右腹股沟区域的脉搏, 并作出一个3-5 厘米腹侧矢状切口。</li>
	<li>用钝夹层暴露股动脉并将其与周围组织分离。</li>
	<li>使用直角止血在暴露的动脉远端周围放置0丝缝线。</li>
	<li>用止血剂将松散的0丝缝合端到皮肤上, 配合远端0丝缝合, 并对动脉施加轻微的张力。</li>
	<li>在外露段的近端处放置第二0丝缝线。</li>
	<li>用18G 介绍人针将导丝插入动脉, 在近端和远端0丝缝合线之间的动脉穿刺。</li>
	<li>将组装的6Fr 护套和扩张装入导丝上, 当它从组件中退出时, 抓住导线。</li>
	<li>在导丝上推进扩张和鞘。在推进扩张鞘总成时, 一定要保持对导线的牢固的把握, 因为它从扩张中凸出。</li>
	<li>充分插入后, 将近0丝缝线缝合在鞘周围, 以维持动脉穿刺部位的止血。</li>
	<li>同时取下扩张和导丝。</li>
	<li>打开单向阀, 以验证是否有脉动血流和冲洗3-5 毫升生理盐水。</li>
	<li>将接入套单向阀连接到压力传感器上的流体填充延伸线, 以监测和记录侵入性血压测量。</li>
	<li>预加载 0.35 "导丝到 4Fr. 血管造影导管, 并连接到与杜希 y 适配器设置的对比注射歧管。</li>
	<li>将导丝尖端拉出导管的远端, 用生理盐水冲洗整个组件。</li>
	<li>将血管造影导管插入鞘内的止血阀, 然后将导丝在远端导管末端的5厘米外向前推进。</li>
	<li>在透视指导下, 使用逆行的反式主动脉方法, 慢慢地将导管插入主弓。</li>
	<li>注射2-3 毫升的造影剂稀释1:1 与正常生理盐水, 以确定起飞的头臂大动脉树干。</li>
	<li>使用导丝引导导管进入头臂大动脉躯干, 然后选择性地进入右颈总动脉。</li>
	<li>取出导丝并注入2-3 毫升稀释的对比度, 以验证导管是否放在颈动脉内。</li>
	<li>注射3-4 毫升未稀释的对比, 同时记录造影运行使用数字减法和标准血管造影技术。</li>
	<li>重复步骤 3.18-3. 21 为重复血管造影在额外的时间点。</li>
</ol>4. 磁共振扩散加权成像 (DWI) 和 T2 加权成像 (T2WI) 的犬脑
注意: 这如图 3A-3B所示。
<ol><li>确保狗在深度麻醉下。</li>
	<li>确保犬的生理参数在整个成像包括心电图和心率中得到监控。</li>
	<li>确保狗趴在仰卧位, 头部位于大脑线圈的双边开口处。</li>
	<li>启用3特斯拉 (3T) 磁铁。</li>
	<li>执行 T2 加权成像 (T2WI), 一个基本的脉搏序列在 MRI。 注意: 序列使用不同病理条件下组织 T2 松弛时间的差异。</li>
	<li>使用 T2-weighted 梯度回声成像序列, 在解剖成像前采集狗脑的先导图像, 并确定视场 (FOV)、切片数和切片厚度以完全覆盖大脑。</li>
	<li>使用以下 T2-parameters: 切片厚度 = 3 毫米, TR = 4000 毫秒, TE = 75 ms, ETL = 7, 采集矩阵 = 320 x 256, FA = 180ᵒ, FOV = 320 x 320 像素, 图像分辨率 = 2.4615 像素每毫米。</li>
	<li>在 MCA 闭塞后立即进行扩散加权成像 (DWI), 使用4小时的回声刨床序列。</li>
	<li>使用下面的 DWI 参数: b = 1500 秒/毫米2, 切片厚度 = 3 毫米, TR = 4600 毫秒, ET = 86 毫秒, ETL = 55, 采集矩阵 = 140 x 140, FA = 90ᵒ, FOV = 231x257, 图像分辨率 = 0.9333 像素/毫米 (表 1)。</li>
</ol>5. 苏木精和伊红 (H & E) 犬脑染色
注意: 这如图 3D所示。
<ol><li>7天后, 10% 福尔马林, 嵌入4毫米脑切片石蜡。</li>
	<li>修剪石蜡块, 直到它们是水平与切片, 清除任何额外的石蜡从顶部的脑组织, 然后削减脑组织在4µm 和放置一个大脑部分在每 2 "x 3" 英寸幻灯片。 注意: 染色前, 将所有解决方案放在单独的容器中, 这样幻灯片就可以从一个解决方案移到另一个溶液, 而不干燥。</li>
	<li>paraffinize 每一个大脑滑动和水合物与自来水。</li>
	<li>把每个大脑幻灯片的苏木精560为8分钟。</li>
	<li>在自来水中冲洗每个脑滑动。</li>
	<li>用1% 酸酒精将每个脑部的幻灯片区分为 1 s 三倍。</li>
	<li>在自来水中冲洗每个脑滑动。</li>
	<li>蓝色的每一个大脑幻灯片与1% 氢氧化铵为1秒。</li>
	<li>在自来水中冲洗每个脑滑动2分钟。</li>
	<li>在70% 乙醇 (乙醇) 中, 每一个脑都有 1 s 十二倍的脱水。</li>
	<li>Counterstain 每个大脑在1分钟内都能滑动。</li>
	<li>在95% 乙醇中, 每一个脑都有 1 s 十二倍的脱水。</li>
	<li>在100% 乙醇中, 每个脑都有脱水。</li>
	<li>清除每个脑滑动在二甲苯, 然后应用一个2英寸 x 3 寸盖玻片在大脑的幻灯片上, 消除气泡与树脂安装媒体。 注意: 除水和乙醇外, 所有解决方案都被重复使用, 多天的染色和多张幻灯片。所有的处理后, 脑部被放置在幻灯片是在玻璃染色罐子, 但塑料染色容器也商业上可用。更换任何解决方案, 当它变色, 并保持紧密覆盖。</li>
</ol>6. 犬颈动脉的苏木精和伊红 (H & E) 染色
注意: 这如图 1所示 (左)。
<ol><li>在10% 福尔马林24-72 小时后, 将受伤的颈动脉切开一半在血栓中部, 并在同一石蜡盒内嵌入1厘米受伤的颈动脉和1厘米的对侧控制。 注: 颈动脉内的血块是脆弱的。等待, 直到该船只被固定在10% 福尔马林前切割, 以使血栓不受干扰。将受伤和控制的颈动脉嵌入同一石蜡块中, 可同时在血管的同一位置切割。</li>
	<li>修剪石蜡块直到水平与切片, 除去任何额外的石蜡。然后切切片在4µm 到25毫米 x 75 毫米 x 1 毫米幻灯片不旋转石蜡块, 使对侧颈动脉部分放置在幻灯片的顶端。 注: 三颈动脉切片放置在每张幻灯片, 但你可以添加取决于污点的兴趣。</li>
	<li>paraffinize 每个颈动脉滑动和水合水中的水合物。 注: 与应单独处理的脑滑块不同, 颈动脉滑块可以通过放置在每次适合几张幻灯片而不相互接触的罐子中进行批量处理。</li>
	<li>用苏木精染色每张颈动脉滑动8分钟。</li>
	<li>在自来水中冲洗每张颈动脉。</li>
	<li>将每张颈动脉滑动与1% 酸酒精区分为 1 s 三倍。</li>
	<li>在自来水中冲洗每张颈动脉。</li>
	<li>蓝色每个颈动脉滑动与1% 氢氧化铵为 1 s。</li>
	<li>冲洗每张颈动脉在自来水中的滑动2分钟。</li>
	<li>将每张颈动脉的70% 乙醇脱水1秒十二次。</li>
	<li>Counterstain 每张颈动脉滑动1分钟。</li>
	<li>将每张颈动脉的95% 乙醇, 1 x 12。</li>
	<li>每张颈动脉100% 乙醇的脱水。</li>
	<li>清除二甲苯中的每个颈动脉滑动, 并添加一个24毫米 x 50 毫米盖玻片与树脂安装介质, 以消除气泡。</li>
</ol>7. 23, 5-苯基-2 四氮唑氯 (TTC) 染色的犬脑
注意: 这如图 3C所示。
<ol><li>将其他4毫米的部分从大脑中央立即移除至 H & E 部分, 并将其置于先前准备的 2% TTC (颈动脉闭塞) 中。在黑暗中孵化37摄氏度至少20分钟, 翻转大脑部分, 甚至染色每5分钟。 注:20 分钟后, 该部分应为樱桃红色两侧。根据 TTC 的新鲜度和组织切片的厚度, 可能需要额外的时间。</li>
	<li>删除的 TTC 解决方案, 并取代它与4% 多聚甲醛在 PBS, pH 7.4, 以优化颜色对比度。</li>
	<li>当白色和红色染色的对比度最佳时, 将 TTC 脑切片放在透明的塑料片之间, 使多余的液体干燥, 并在高分辨率下扫描以追踪缺血性区域。 注意: 节可以无限期地存储在多聚甲醛解决方案中, 但染色会褪色。</li>
</ol>

<ol>
	<li>Adams, H. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stroke:+a+vascular+pathology+with+inadequate+management.">Stroke: a vascular pathology with inadequate management.</a> Journal of Hypertension Supplement. 21 (5), S3-S7 (2003).</li><li>Lansberg, M. G., Bluhmki, E., Thijs, V. N. <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+tissue+plasminogen+activator+3+to+4.5+hours+after+acute+ischemic+stroke:+a+metaanalysis.">Efficacy and safety of tissue plasminogen activator 3 to 4.5 hours after acute ischemic stroke: a metaanalysis.</a> Stroke. 40 (7), 2438-2441 (2009).</li><li>Nagel, 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=Therapy+of+acute+basilar+artery+occlusion:+intraarterial+thrombolysis+alone+vs+bridging+therapy.">Therapy of acute basilar artery occlusion: intraarterial thrombolysis alone vs bridging therapy.</a> Stroke. 40 (1), 140-146 (2009).</li><li>Ciccone, A., Motto, C., Abraha, I., Cozzolino, F., Santilli, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glycoprotein+IIb-IIIa+inhibitors+for+acute+ischaemic+stroke.">Glycoprotein IIb-IIIa inhibitors for acute ischaemic stroke.</a> The Cochrane database of systematic reviews. 3 (3), (2014).</li><li>The National Institute of Neurological Disorders and Stroke rt-PA Stroke Study Group. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tissue+Plasminogen+Activator+for+Acute+Ischemic+Stroke.">Tissue Plasminogen Activator for Acute Ischemic Stroke.</a> New England Journal of Medicine. 333 (24), 1581-1588 (1995).</li><li>Leadley, R., Chia, L., Rebellob, S., Gagnon, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contribution+of+in+vivo+models+of+thrombosis+to+the+discovery+and+development+of+novel+antithrombotic+agents.">Contribution of in vivo models of thrombosis to the discovery and development of novel antithrombotic agents.</a> Journal of Pharmacological and Toxicological Methods. 43 (2), 101-116 (2000).</li><li>Bodary, P. F., Eitzman, 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=Animal+Models+of+Thrombosis.">Animal Models of Thrombosis.</a> Current Opinion In Hematology. 16 (5), 342-346 (2009).</li><li>Sachs, U. J. H., Nieswandt, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+thrombus+formation+in+murine+models.">In vivo thrombus formation in murine models.</a> Circulation Research. 100 (7), 979-991 (2007).</li><li>Bonnard, T., Hagemeyer, C. E. <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+Thrombosis+Mouse+Model+on+Carotid+Artery+and+Mesentery+Vessel.">Ferric Chloride-induced Thrombosis Mouse Model on Carotid Artery and Mesentery Vessel.</a> Journal of Visualized Experiments. (100), e52838 (2015).</li><li>Kurz, K. D., Main, B. W., Sandusky, G. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rat+model+of+arterial+thrombosis+induced+by+ferric+chloride.">Rat model of arterial thrombosis induced by ferric chloride.</a> Thrombosis Research. 60 (4), 269-280 (1990).</li><li>Karatas, H., Erdener, S. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thrombotic+distal+middle+cerebral+artery+occlusion+produced+by+topical+FeCl(3)+application:+a+novel+model+suitable+for+intravital+microscopy+and+thrombolysis+studies.">Thrombotic distal middle cerebral artery occlusion produced by topical FeCl(3) application: a novel model suitable for intravital microscopy and thrombolysis studies.</a> Journal of Cerebral Blood Flow and Metabolism. 31 (6), 1452-1460 (2011).</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 Biology. 1 (1), 50-55 (2013).</li><li>Vilahur, G., Padro, T., Badimon, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Atherosclerosis+and+Thrombosis:+Insights+from+Large+Animal+Models.">Atherosclerosis and Thrombosis: Insights from Large Animal Models.</a> Journal of Biomedicine and Biotechnology. 2011, 1-12 (2011).</li><li>Coller, B. S., Folts, J. D., Smith, S. R., Scudder, L. E., Jordan, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Abolition+of+in+vivo+Platelet+Thrombus+Formation+in+Primates+with+Monoclonal+Antibodies+to+the+Platelet+GPIIb/IIIa+Receptor.+Correlation+with+Bleeding+Time,+Platelet+Aggregation,+and+Blockade+of+GPIIb/IIIa+Receptors.">Abolition of in vivo Platelet Thrombus Formation in Primates with Monoclonal Antibodies to the Platelet GPIIb/IIIa Receptor. Correlation with Bleeding Time, Platelet Aggregation, and Blockade of GPIIb/IIIa Receptors.</a> Circulation. 80 (6), 1766-1774 (1989).</li><li>Folts, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+in+vivo+Model+of+Experimental+Arterial+Stenosis,+Intimal+Damage,+and+Periodic+Thrombosis.">An in vivo Model of Experimental Arterial Stenosis, Intimal Damage, and Periodic Thrombosis.</a> Circulation. 83 (6 Suppl), (1991).</li><li>Yasuda, 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=A+canine+model+of+coronary+artery+thrombosis+with+superimposed+high+grade+stenosis+for+the+investigation+of+rethrombosis+after+thrombolysis.">A canine model of coronary artery thrombosis with superimposed high grade stenosis for the investigation of rethrombosis after thrombolysis.</a> Journal of the American College of Cardiology. 13 (6), 1409-1414 (1989).</li><li>Schob, 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=Correlation+Between+Aquaporin+4+Expression+and+Different+DWI+Parameters+in+Grade+I+Meningioma.">Correlation Between Aquaporin 4 Expression and Different DWI Parameters in Grade I Meningioma.</a> Molecular Imaging and Biology : MIB : the Official Publication of the Academy of Molecular Imaging. 19 (1), 138-142 (2017).</li><li>Schob, 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=Diffusion-Weighted+Imaging+Using+a+Readout-Segmented,+Multishot+EPI+Sequence+at+3+T+Distinguishes+between+Morphologically+Differentiated+and+Undifferentiated+Subtypes+of+Thyroid+Carcinoma-A+Preliminary+Study.">Diffusion-Weighted Imaging Using a Readout-Segmented, Multishot EPI Sequence at 3 T Distinguishes between Morphologically Differentiated and Undifferentiated Subtypes of Thyroid Carcinoma-A Preliminary Study.</a> Translational Oncology. 9 (5), 403-410 (2016).</li><li>Schob, 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=Diffusion-Weighted+MRI+Reflects+Proliferative+Activity+in+Primary+CNS+Lymphoma.">Diffusion-Weighted MRI Reflects Proliferative Activity in Primary CNS Lymphoma.</a> Public Library of Science One. 11 (8), e0161386 (2016).</li></ol>

FeCl3诱导的血管损伤模型广泛用于研究小动物血栓形成, 易于转化为大型动物, 具有多种优势的临床前模型。将该协议调整为犬类的轻微修改, 允许利用磁共振成像对中风和出血量进行药理干预和血管造影评估, 评估血管疏导之前、期间和治疗后。其他血栓性大动物模型未研究损伤部位的稳定闭塞血栓, 因此不能利用颈动脉血管造影和组织学评估每种预防性或溶栓疏导的程度。治疗。除了大?...

中风治疗主要是在冠状动脉疾病治疗后进行模拟的, 因为心血管疾病的干预对药物治疗和血管内干预措施有很好的反应1。然而, 这些治疗并没有成功地转化为脑梗塞。目前中风治疗的难点是重组组织纤溶酶原激活剂 (rTPA) 不能逆转, 而且该管理具有6.4% 的出血转化率2,3的风险,4. 由此产生的发病率和死亡率限制了它对一个小的、常常无法实现的窗口5的使用。同时, 再狭窄和闭塞往往发生在最初的溶栓后, 逆转最初的神经系统改善。总之, 有一个狭窄的时间窗口管理 rTPA, 排除大多数 (90%) 的患者谁患有缺血性脑血管的侮辱。
静脉抗血小板治疗在改善血管再通、生存和结局2方面显示了治疗缺血性中风的前景。不幸的是, 这些药物有一个可预测的副作用的颅内和颅内出血, 主要是因为没有办法来充分逆转或控制他们的活动2。虽然有效的预防血小板聚集, 出血的风险和无法逆转他们的活动已排除在日常护理中风患者使用。因此, 有必要采取有效的抗血栓药物, 单独或组合, 以防止和溶解血栓还有一个安全的配置文件, 将允许使用在封闭的, 低体积的空间, 如大脑, 出血是不耐受的。
了解动脉血栓形成和再狭窄的机制, 评估预防再狭窄的栓和药物, 需要小动物模型作为临床前药物开发的一部分。氯化铁诱导的血管损伤是快速、准确诱导小鼠、大鼠 、豚鼠和家兔6、7、8等暴露血管血栓形成的一种广泛应用的技术,9,10,11,12. 这些较小的物种提供了一些好处, 包括易于遗传操作、廉价的动物购买和低的每日津贴住房费用。不幸的是, 小动物实验在手术中否定了多项血液绘制, 以获得血小板反应性、血气分析和炎症反应。更重要的是, 大型动物更紧密地模仿人类血小板生理学6,13。FeCl3型颈动脉损伤模型在研究血栓的病理生理学、新的抗血小板和抗凝血药物的有效性以及发现潜在栓6中发挥了主导作用。,7,8,9,10,11,12. 以前在小鼠、大鼠、豚鼠和兔子身上的模型为基因操作提供了方便和灵活性, 但可翻译的临床前模型对药物治疗药物的剂量和毒性研究有重要意义6 ,13。虽然在小鼠中已经形成了几种血栓紊乱模型, 但适用于周围血管疾病、中风和心肌梗塞的大动物血栓形成模型却很少。第一个血栓模型在猴子, 狗和猪集中于狭窄, 应用止血剂和以后圆筒对容器, 通常导致循环流减少14,15,16。而不是在内皮损伤部位的闭塞血栓, 如在氯化铁模型中, 这些模型中的血栓导致循环血栓形成, 远端栓塞, 恢复正常血流。相比之下, 在大动物这里修改的氯化铁模型, 导致损伤部位闭塞血栓, 并在溶栓治疗前通过血管造影进行稳定和验证。如果调查人员有足够的资金用于每日津贴和购买犬齿以及足够的外科专业技能, 我们这里详细介绍一大犬血管损伤模型, 让实验室通过手术、影像学和组织学研究血栓形成。技术。

<table border="0" cellpadding="0" cellspacing="0" width="1136">
	<tbody>
		<tr height="40">
			<td height="40" style="height:40px;width:397px;">1/8&#8221; umbilical tape&#160;</td>
			<td style="width:195px;">Jorgensen Laboratories Inc.,&#160;</td>
			<td style="width:344px;">#J0025UA&#160;</td>
			<td style="width:201px;">for ferric chloride application</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">4% paraformaldehyde in PBS</td>
			<td>Alfa Aesar</td>
			<td>AAJ61899AP</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">10% neutral buffered formalin&#160;</td>
			<td>Richard-Allan Scientific</td>
			<td>5701</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">&#160;2% 2,3,5-triphenyltetrazolium chloride (TTC in PBS, pH 7.4)&#160;</td>
			<td>Sigma Aldrich</td>
			<td>T8877</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">ADP/Collagen cartridges</td>
			<td>Siemens Diagnostics</td>
			<td>B417021A</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">4.5 ml 3.2% sodium citrate blood vacutainer&#160;</td>
			<td>Becton Dickinson</td>
			<td>BD 369714</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">4.5 ml lithium heparin vacutainer&#160;</td>
			<td>Becton Dickinson</td>
			<td>BD 368056</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">EDTA K3 vacutainers&#160;</td>
			<td>Becton Dickinson</td>
			<td>BD455036</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Doppler flow probe</td>
			<td>Transonic Systems Inc</td>
			<td>MA2.5PSL</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Hematoxylin 560&#160;</td>
			<td>Surgipath</td>
			<td>3801570</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Eosin</td>
			<td>Surgipath</td>
			<td>3801602</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">LabChart Software</td>
			<td>ADInstruments Inc.</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Prisma Fit 3 tesla (3T) magnet</td>
			<td>Siemen&#39;s Diagnostics</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Sodium heparin for injection (to coat blood gas syringe)</td>
			<td>NovaPlus</td>
			<td>402525D</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">HUG-U-VAC positioning system &#160;</td>
			<td>DRE Veterinary</td>
			<td>1320</td>
			<td></td>
		</tr>
	</tbody>
</table>

ferric chloride-induced canine carotid artery thrombosis: a large animal model of vascular injury

导致脑缺血性中风和心肌梗死的闭塞性动脉血栓每年导致1300万人死亡。在这里, 我们已经把一个小动物的血管损伤模型转化成一个大动物 (犬), 轻微的修改, 可用于临床前筛查预防和溶栓剂。除了手术方法外, 修改后的协议还描述了通过血管造影评估颈动脉疏导的分步方法, 详细说明了用大脑和颈动脉进行组织学分析以证实颈动脉疏导和脑出血, 以及具体的参数, 以完成评估下游血栓发生事件, 利用磁共振成像 (MRI)。此外, 还对以前建立良好的小动物模型进行了具体的程序性改变, 以此来转化为大动物 (犬) 血管损伤。

下面的详细程序将导致开发一个模型, 可用于预防或溶栓评估闭塞动脉干预。图 1A显示了在商业软件记录的治疗之前、期间和之后的基线流速和所产生的血流速度。该记录的数据可用于确定该犬模型中颈动脉损伤和治疗后再灌注的百分比。图 1B提供了一个例子, 对侧 (顶部) 和受伤 (底部) 犬颈动脉部分染色的 H & E, 以验证在牺牲?...

Watch this Scientific Journal Video about Ferric Chloride-induced Canine Carotid Artery Thrombosis: A Large Animal Model of Vascular Injury at JoVE.com

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

在此, 我们提出了必要的修改, 以良好的特点和常用的小动物氯化铁诱导 (FeCl3) 颈动脉损伤模型, 用于大动物血管损伤模型。结果模型可用于临床前试验评估预防和溶栓的药理和机械干预。

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-canine-carotid-artery-thrombosis-large-animal

Research

JoVE Journal

Medicine

9.0K 次观看.  Ohio State University. 在此, 我们提出了必要的修改, 以良好的特点和常用的小动物氯化铁诱导 (FeCl3) 颈动脉损伤模型, 用于大动物血管损伤模型。结果模型可用于临床前试验评估预防和溶栓的药理和机械干预。

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

Electrolytic Inferior Vena Cava Model (EIM) of Venous Thrombosis

Animal models serve a vital role in deep venous thrombosis (DVT) research in order to study thrombus formation, thrombus resolution and to test potential therapeutic compounds (1). New compounds to be utilized in the treatment and prevention of DVT are currently being developed. The delivery of potential therapeutic antagonist compounds to an affected thrombosed vein has been problematic. In the context of therapeutic applications, a model that uses partial stasis and consistently generates thrombi within a major vein has been recently established. The Electrolytic Inferior vena cava Model (EIM) is mouse model of DVT that permits thrombus formation in the presence of continuous blood flow. This model allows therapeutic agents to be in contact with the thrombus in a dynamic fashion, and is more sensitive than other models of DVT (1). In addition, this thrombosis model closely simulates clinical situations of thrombus formation and is ideal to study venous endothelial cell activation, leukocyte migration, venous thrombogenesis, and to test therapeutic applications (1). The EIM model is technically simple, easily reproducible, creates consistent thrombi sizes and allows for a large sample (i.e. thrombus and vein wall) which is required for analytical purposes.

Electrolytic Inferior Vena Cava Model EIM of Venous Thrombosis

The electrolytic induction of endothelial activation to the internal surface of the Inferior Vena Cava results in venous type thrombus formation due to endothelial activation and partial blood stasis, two components of Virchow's triad.

电解下腔静脉静脉血栓形成的模型（EIM）

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

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 ...

Training a Sophisticated Microsurgical Technique: Interposition of External Jugular Vein Graft in the Common Carotid Artery in Rats

Neointimal hyperplasia is one the primary causes of stenosis in arterialized veins that are of great importance in arterial coronary bypass surgery, in peripheral arterial bypass surgery as well as in arteriovenous fistulas.1-5 The experimental procedure of vein graft interposition in the common carotid artery by using the cuff-technique has been applied in several research projects to examine the aetiology of neointimal hyperplasia and therapeutic options to address it. 6-8 The cuff prevents vessel anastomotic remodeling and induces turbulence within the graft and thereby the development of neointimal hyperplasia.
Using the superior caval vein graft is an established small-animal model for venous arterialization experiment.9-11 This current protocol refers to an established jugular vein graft interposition technique first described by Zou et al., 9 as well as others.12-14 Nevertheless, these cited small animal protocols are complicated.
To simplify the procedure and to minimize the number of experimental animals needed, a detailed operation protocol by video training is presented. This video should help the novice surgeon to learn both the cuff-technique and the vein graft interposition. Hereby, the right external jugular vein was grafted in cuff-technique in the common carotid artery of 21 female Sprague Dawley rats categorized in three equal groups that were sacrificed on day 21, 42 and 84, respectively. Notably, no donor animals were needed, because auto-transplantations were performed. The survival rate was 100 % at the time point of sacrifice. In addition, the graft patency rate was 60 % for the first 10 operated animals and 82 % for the remaining 11 animals. The blood flow at the time of sacrifice was 8&plusmn;3 ml/min. In conclusion, this surgical protocol considerably simplifies, optimizes and standardizes this complicated procedure. It gives novice surgeons easy, step-by-step instruction, explaining possible pitfalls, thereby helping them to gain expertise fast and avoid useless sacrifice of experimental animals.

Neointimal hyperplasia is the primary cause of stenosis in arterialized veins. We propose a new protocol whereby the right external jugular vein is grafted using the cuff technique in the common carotid artery of Sprague Dawley rats. The survival rate was 100 % at the time point of sacrifice.

培养一个精密的显微外科技术在大鼠颈总动脉，颈外静脉移植:介入

Neointimal hyperplasia is one the primary causes of stenosis in arterialized veins that are of great importance in arterial coronary bypass surgery, in ...

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 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.

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 ...

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

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

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

大鼠颈动脉球囊损伤模型试验抗血管重建治疗

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

Ferric Chloride-induced Murine Thrombosis Models

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

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

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, ...

Invasive Hemodynamic Monitoring of Aortic and Pulmonary Artery Hemodynamics in a Large Animal Model of ARDS

One of the leading causes of morbidity and mortality in patients with heart failure is right ventricular (RV) dysfunction, especially if it is due to pulmonary hypertension. For a better understanding and treatment of this disease, precise hemodynamic monitoring of left and right ventricular parameters is important. For this reason, it is essential to establish experimental pig models of cardiac hemodynamics and measurements for research purpose. 
This article shows the induction of ARDS by using oleic acid (OA) and consequent right ventricular dysfunction, as well as the instrumentation of the pigs and the data acquisition process that is needed to assess hemodynamic parameters. To achieve right ventricular dysfunction, we used oleic acid (OA) to cause ARDS and accompanied this with pulmonary artery hypertension (PAH). With this model of PAH and consecutive right ventricular dysfunction, many hemodynamic parameters can be measured, and right ventricular volume load can be detected. 
All vital parameters, including respiratory rate (RR), heart rate (HR) and body temperature were recorded throughout the whole experiment. Hemodynamic parameters including femoral artery pressure (FAP), aortic pressure (AP), right ventricular pressure (peak systolic, end systolic and end diastolic right ventricular pressure), central venous pressure (CVP), pulmonary artery pressure (PAP) and left arterial pressure (LAP) were measured as well as perfusion parameters including ascending aortic flow (AAF) and pulmonary artery flow (PAF). Hemodynamic measurements were performed using transcardiopulmonary thermodilution to provide cardiac output (CO). Furthermore, the PiCCO2 system (Pulse Contour Cardiac Output System 2) was used to receive parameters such as stroke volume variance (SVV), pulse pressure variance (PPV), as well as extravascular lung water (EVLW) and global end-diastolic volume (GEDV). Our monitoring procedure is suitable for detecting right ventricular dysfunction and monitoring hemodynamic findings before and after volume administration.

We present a protocol of creating right ventricular dysfunction in a pig model by inducing ARDS. We demonstrate invasive monitoring of left and right ventricular cardiac output using flow probes around the aorta and the pulmonary artery, as well as blood pressure measurements in the aorta and pulmonary artery.

ards 大型动物模型主动脉和肺动脉血流动力学的侵袭血流动力学监测

One of the leading causes of morbidity and mortality in patients with heart failure is right ventricular (RV) dysfunction, especially if it is due to ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

用线 Myography 测定小鼠肠系膜动脉的等轴收缩力

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

A Rat Carotid Artery Pressure-Controlled Segmental Balloon Injury with Periadventitial Therapeutic Application

Cardiovascular disease remains the leading cause of death and disability worldwide, in part due to atherosclerosis. Atherosclerotic plaque narrows the luminal surface area in arteries thereby reducing adequate blood flow to organs and distal tissues. Clinically, revascularization procedures such as balloon angioplasty with or without stent placement aim to restore blood flow. Although these procedures reestablish blood flow by reducing plaque burden, they damage the vessel wall, which initiates the arterial healing response. The prolonged healing response causes arterial restenosis, or re-narrowing, ultimately limiting the long-term success of these revascularization procedures. Therefore, preclinical animal models are integral for analyzing the pathophysiological mechanisms driving restenosis, and provide the opportunity to test novel therapeutic strategies. Murine models are cheaper and easier to operate on than large animal models. Balloon or wire injury are the two commonly accepted injury modalities used in murine models. Balloon injury models in particular mimic the clinical angioplasty procedure and cause adequate damage to the artery for the development of restenosis. Herein we describe the surgical details for performing and histologically analyzing the modified, pressure-controlled rat carotid artery balloon injury model. Additionally, this protocol highlights how local periadventitial application of therapeutics can be used to inhibit neointimal hyperplasia. Lastly, we present light sheet fluorescence microscopy as a novel approach for imaging and visualizing the arterial injury in three-dimensions.

The rat carotid artery balloon injury mimics the clinical angioplasty procedure performed to restore blood flow in atherosclerotic vessels. This model induces the arterial injury response by distending the arterial wall, and denuding the intimal layer of endothelial cells, ultimately causing remodeling and an intimal hyperplastic response.

大鼠胡萝卜动脉压力控制段气球损伤与腹腔ventit 治疗应用

Cardiovascular disease remains the leading cause of death and disability worldwide, in part due to atherosclerosis. Atherosclerotic plaque narrows the ...