这项工作得到了瑞士国家科学基金会赠款150271的支持。

该试验方案已获州兽医局、弗里堡和瑞士联邦兽医局批准 (FR 2015/58). 
 注意: 使用2.8 到3.2 公斤之间的雄性 NZW 兔子。动物被安置在常规情况下 (12 h 光和黑暗的周期, 提供广告随意水和食物)。在气球剥脱之前, 动物被驯化了1周, 在这期间, 他们被喂食正常饮食。经过1周的驯化, 家兔被转换为动脉粥样硬化饮食由高脂肪 (8.6%), 和饱和脂肪酸与205毫克/千克胆固醇 (1%) 饮食的整个研究期间。在2周或4周的球囊损伤后, 左髂动脉的球囊损伤在1周后进行, 而动物则被处死. 
 1. 术前程序 <ol> <li> 在使用玻璃珠消毒器或其他合适的仪器之前对所有手术器械进行消毒。
	<li> 准备并检查球囊导管总成。
	<ol> <li> 在气囊导管的口锁部分安装一个1毫升的口锁注射器, 装上生理盐水。仔细监测没有被困的空气。通过按压注射器的柱塞, 检查泄漏并确保适当的气球膨胀. </li>
	</ol> </li> <li> 称量兔子并打开 thermopad 到37和 #176; C. </li>
	<li> 在0.3 毫克/毫升的浓度下使用丁丙诺啡溶液. 注射0.01 毫克/千克皮下的剂量. </li>
	<li> 麻醉5% 异氟醚和5升/分 O 2 在感应腔中为 10-15 min </li>
	<li> 将麻醉兔子放在手术台上的加热垫上。放置补丁和剪辑, 以监测温度, 呼吸和心电图. </li>
	<li> 将兔子的吻附着在与合适的麻醉机相连的面罩上。保持麻醉与异氟醚 (4.0% 与2.5 升/分钟 O 2 )。确认适当的麻醉 (表现为缺乏肌肉张力和失去呕吐和片反射). </li>
	将眼部软膏涂抹在眼睛上, 防止角膜干燥。用不育的手术片将兔子披上, 只有下肢外露. </li>
</ol> 2。手术协议 <ol> <li> 用动物理发钳从膝关节下方的腹侧移除毛发. </li>
	<li> 用适当的消毒剂擦拭区域以清洁皮肤并去除松散的毛发. </li>
	<li> 定位隐动脉, 用手术刀做一个长度约1.5 厘米的小皮肤切口. </li>
	<li> 在不损伤股静脉和股神经的情况下, 用小弯曲钳暴露一小部分隐动脉. </li>
	<li> 将两个松散的结扎环 (5-0 丝) 放在隐动脉下方, 并将一条结扎回路绑在动脉远端。在绷带上方放置一个微血管夹, 以阻止髂动脉的血流. </li>
	<li> 局部应用一滴罂粟碱来扩张动脉, 防止血管痉挛. </li>
	<li> 用绑结扎的帮助抬起隐动脉, 用24口径的针头做一个小的切开切口. </li>
	<li> 用细钳将切口瓣抬高, 慢慢地将静脉穿刺或引导针插入动脉腔内. </li>
	<li> 将2法国福格蒂动脉栓导管插入隐动脉。取出静脉穿刺和微血管夹. </li>
	<li> 将导管推进至第六标记 (20-25 厘米), 相应于髂分岔处大约2-5 厘米的位置. </li>
	<li> 使用1毫升的注射器或在额定压力 6 atm 上充气气球用一个被调控的手动充气, 如描述在 16 , 22 . </li>
	<li> 在旋转导管的同时, 用钳夹住气囊导管, 并通过髂 arterytoward 在插入点上向后拖6厘米. </li>
	<li> 通过拉回注射器的柱塞来使气球泄气. </li>
	<li> 重复步骤2.10 到2.13 三次, 以确保完全的内皮剥脱. </li>
	<li> 卸下导管并立即将切开站点上方的结扎环绑住以停止出血. </li>
	<li> 在伤口周围涂抹适当的防腐剂, 拭去血块。用5-0 缝线闭合皮肤切口, 用聚维酮碘溶液消毒手术部位. </li>
	<li> 重复步骤2.1 至2.16 对侧髂使用新的导管. </li>
	<li> 从眼睛拭去眼部药膏. </li>
</ol> 3。术后护理 <ol> <li> 在手术后立即管理磺胺40毫克/千克和苄8毫克/千克或任何其他合适的抗生素. </li>
	<li> 在麻醉恢复期间, 将兔子放在一个清洁的蒸压笼中. </li>
	<li> 删除监视修补程序和剪辑. </li>
	<li> 恢复后, 将兔子返回到他们的家庭笼子。注射皮下丁丙诺啡 0.05-0.1 毫克/千克术每 6-12 ĥ 48 h. 继续动脉粥样硬化饮食, 再过两个星期或四周. </li>
</ol> 4。斑块组成的组织采集与分析 <ol> <li> 在两周后 (对于早期薄斑块) 或三周的气球损伤, 麻醉用异氟醚与上述类似的方式使用了兔子. </li>
	<li> intracardial 失血打开胸腔和安乐兔. </li>
	<li> 隔离髂动脉, 如 23 所述。
	<ol> <li> 简要地打开腹部并露出后。追踪主动脉向髂分岔, 并将其系在分岔之上。仔细清除周围的组织, 以揭露和孤立髂动脉. </li>
	</ol> </li> <li> 解剖两个髂动脉并将它们浸泡在 ice-cold 磷酸盐缓冲盐水中。在镊子的帮助下取出血块。将每个髂动脉分成 4-6 段, 以描述整个动脉中斑块的厚度. </li>
	<li> 立即将动脉段嵌入到包含最佳切削温度复合物的模具中, 使用液氮对齐冻结, 并将其保持在-70 和 #176; c. 使用低温中描述的5和 #181; m 厚剖面, 如 24 . </li>
	<li> 执行组织学、免疫荧光或免疫组化染色, 如 10 、 25 中所述的形态测量学、斑块脂质和细胞含量。
	注: 简要地, 用磷酸缓冲盐 (PBS) 和 permeabilize 用0.2% 海卫一冲洗动脉部分。用 PBS 冲洗切片, 用2% 牛血清白蛋白阻断非特定部位30分钟. 在37和 #176 上孵育 1 h 的部分; 具有抗 #945 的 C; SM 肌动蛋白 (1:200) 或 RAM11 抗体 (1:200)。用 pbs 冲洗切片, 用适当的二次抗体孵育30分钟, 在37和 #176; c. 再次用 pbs 洗涤, 并添加赫斯特 (5 和 #181; g/毫升), 10 分钟以检测细胞核. </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=Experimental+atherosclerosis+in+rabbits.">Experimental atherosclerosis in rabbits.</a> Arq Bras Cardiol. 95, 272-278 (2010).</li><li>Waksman, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=PhotoPoint+photodynamic+therapy+promotes+stabilization+of+atherosclerotic+plaques+and+inhibits+plaque+progression.">PhotoPoint photodynamic therapy promotes stabilization of atherosclerotic plaques and inhibits plaque progression.</a> J Am Coll Cardiol. 52, 1024-1032 (2008).</li><li>Fernandez-Parra, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pharmacokinetic+Study+of+Paclitaxel+Concentration+after+Drug-Eluting+Balloon+Angioplasty+in+the+Iliac+Artery+of+Healthy+and+Atherosclerotic+Rabbit+Models.">Pharmacokinetic Study of Paclitaxel Concentration after Drug-Eluting Balloon Angioplasty in the Iliac Artery of Healthy and Atherosclerotic Rabbit Models.</a> J Vasc Interv Radiol. 26, 1380-1387 (2015).</li><li>Dussault, S., Dhahri, W., Desjarlais, M., Mathieu, R., Rivard, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Elsibucol+inhibits+atherosclerosis+following+arterial+injury:+multifunctional+effects+on+cholesterol+levels,+oxidative+stress+and+inflammation.">Elsibucol inhibits atherosclerosis following arterial injury: multifunctional effects on cholesterol levels, oxidative stress and inflammation.</a> Atherosclerosis. 237, 194-199 (2014).</li><li>Manderson, J. A., Mosse, P. R., Safstrom, J. A., Young, S. B., Campbell, G. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Balloon+catheter+injury+to+rabbit+carotid+artery.+I.+Changes+in+smooth+muscle+phenotype.">Balloon catheter injury to rabbit carotid artery. I. Changes in smooth muscle phenotype.</a> Arteriosclerosis. 9, 289-298 (1989).</li><li>Miyake, 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=Prevention+of+neointimal+formation+after+angioplasty+using+nuclear+factor-kappaB+decoy+oligodeoxynucleotide-coated+balloon+catheter+in+rabbit+model.">Prevention of neointimal formation after angioplasty using nuclear factor-kappaB decoy oligodeoxynucleotide-coated balloon catheter in rabbit model.</a> Circ Cardiovasc Interv. 7, 787-796 (2014).</li><li>Fulcher, J., Patel, S., Nicholls, S. J., Bao, S., Celermajer, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optical+coherence+tomography+for+serial+in+vivo+imaging+of+aortic+plaque+in+the+rabbit:+a+preliminary+experience.">Optical coherence tomography for serial in vivo imaging of aortic plaque in the rabbit: a preliminary experience.</a> Open Heart. 2, e000314 (2015).</li><li>Abela, O. G., 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=Plaque+Rupture+and+Thrombosis:+the+Value+of+the+Atherosclerotic+Rabbit+Model+in+Defining+the+Mechanism.">Plaque Rupture and Thrombosis: the Value of the Atherosclerotic Rabbit Model in Defining the Mechanism.</a> Curr Atheroscler Rep. 18, 29 (2016).</li><li>Yamashita, A., Asada, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+rabbit+model+of+thrombosis+on+atherosclerotic+lesions.">A rabbit model of thrombosis on atherosclerotic lesions.</a> J Biomed Biotechnol. 2011, 424929 (2011).</li></ol>

作者声明没有竞争的金融利益。

兔髂动脉粥样硬化模型广泛应用于动脉粥样硬化的研究。与这个协议兔子迅速地开发了更加严厉和先进的匾与自发损伤开发与仅胆固醇膳食。重要的是, 动物从手术中迅速恢复。
动脉粥样硬化的主要刺激是由气囊导管造成的机械损伤, 它会伤害内皮细胞, 扩张血管壁26。这一过程诱发重塑反应的特点是炎症与巨噬细胞的招募和脂质积累时, 与 hypercholestorolemic 饮...

易损性动脉粥样硬化斑块破裂是工业化国家的主要死因之一1。尽管过去几十年的研究已经展现了一些分子和细胞机制参与了斑块的进展, 但仍然需要继续努力, 不仅要解开疾病进展的复杂机制, 还要测试新的治疗方法.已经提出了几种动物模型来研究动脉粥样硬化。基因操作、胆固醇喂养或机械内皮损伤是多数动物模型的共同的标准策略包括小鼠、兔子或者小型。在这些, NZW 兔子是敏感的胆固醇饮食, 而正常的大鼠和小鼠不显着吸收膳食胆固醇2,3,4。家兔自发地发展出富含巨噬细胞的主动脉病变, 当喂食富含胆固醇的饮食时5,6。然而, 4-8 月的长期准备时间诱导动脉粥样硬化 plaquesby 喂养胆固醇饮食单独6,7是大多数实验设置的主要缺点。为了在较短的时间内诱导病变, Baumgarter 和施图德博土8开发了高胆固醇饮食和球囊损伤的结合。这项技术的总目标是诱导2周内高家兔的动脉粥样硬化斑块 (类似于人的脂肪条纹)。本技术介绍了基于 Baumgarter 方法的动脉壁损伤过程, 采用气囊导管进入 NZW 高兔髂动脉。
加上胆固醇丰富的饮食, 由气球引起的损伤导致损伤将导致动脉粥样硬化。球囊损伤加速动脉粥样硬化病变的形成, 并产生均匀大小和分布的斑块。内膜增厚超过一段时间, 内中膜细胞浸润在损伤后数日内开始增加。脂肪条纹与大量巨噬细胞开始出现后 7-10 天的气球损伤, 并表示为 II 型病变根据分类的美国心脏协会。兔的球囊损伤常在主动脉中进行, 以研究斑块的组成。膜内皮表达高水平的细胞间黏附分子。斑块与内侧解剖和外的变化有关。动脉粥样硬化病变由脂质、增殖性平滑肌细胞 (校董)、胶原纤维和在再生内皮下积聚的炎症细胞组成, 主要为 II. 型。兔斑块的拓扑分布类似于人类脉的报告9,10原则上, 与髂动脉相比, 大动脉的大小更大, 并且会产生更大长度的斑块。然而, 利用髂动脉作为动脉粥样硬化部位的主要优势是其可获得性, 其肌肉含量与人冠状动脉的相似性11, 均匀病变的发展为12, 高组织因子活动13和一致的血管尺寸, 可与人冠状动脉相媲美, 允许对商业制造的设备进行评估, 以进行形态学和血管造影终点。对活体动物的兔髂动脉斑块进行了侵入性和无创的分析研究。以前的报告描述了使用磁共振成像 (MRI) 的帮助下, 2.35-特斯拉先生系统14此外, 血管内超声 (IVUS) 或光学相干断层扫描 (OCT) 导管可适当应用于图像兔髂动脉动脉粥样硬化斑块。髂动脉可用于超声成像时, 使用高分辨率声像和主动脉也可以探索与该技术。
在过去的十年中, 这种兔气球损伤模型有助于进一步了解斑块进展的机制15和斑块回归16。此外, 该模型已被用来研究新的治疗剂, 如他汀类药物, 标准抗血小板药物, 抗氧化剂17,18和药物洗脱支架, 如司或zotarolimus 洗脱支架19,20膜增厚。该模型还用于研究近红外荧光成像导管的血管内成像21。

<table><tbody><tr><td>New Zealand White rabbits</td><td>Charles River laboratories,France</td><td>Cre:KBL(NZW)</td><td></td></tr><tr><td>Cholesterol rich diet</td><td>Ssniff spezialdi&amp;auml;ten</td><td>Ssniff EF K High Fat and Cholesterol</td><td></td></tr><tr><td>Glass bead sterilizer-Germinator 500</td><td>VWR, Leicestershire, UK</td><td>101326-488</td><td></td></tr><tr><td>Fogarty balloon embolectomy catheters, 2 French</td><td>Edwards Lifesciences, Switzerland</td><td>120602F</td><td>For single use only</td></tr><tr><td>Luer Lock Syringe</td><td>Becton, Dickinson and Company, USA</td><td>309628</td><td></td></tr><tr><td>Thermopad Type 226</td><td>Solis, Switzerland AG</td><td>397387</td><td></td></tr><tr><td>Buprenorphine- Temgesic</td><td>Reckitt Benckiser AG, Switzerland</td><td>7.68042E+12</td><td></td></tr><tr><td>Isoflurane</td><td>Piramal Critical Care, Inc, Bethlehem, PA 18017</td><td>2667-46-7</td><td></td></tr><tr><td>Anaesthesia machine-combi-vet Base Anesthesia System</td><td>Rothacher Medical GmbH, Switzerland</td><td>CV 30-301-A</td><td></td></tr><tr><td>Cardell touch veterinary vital signs monitor</td><td>Midmark, Ohio, USA</td><td>8013-001</td><td></td></tr><tr><td>Ophthalmic ointment-Humigel</td><td>Virbac, France</td><td></td><td></td></tr><tr><td>Animal hair clippers</td><td>Aesculap AG, Germany</td><td>GT420</td><td></td></tr><tr><td>Disinfectant-Betadine solution</td><td>MundipharmaMedicalCompany, Switzerland</td><td>14671-1203</td><td></td></tr><tr><td>Dumont #7 Forceps</td><td>FST Germany</td><td>11274-20</td><td></td></tr><tr><td>Medium and small microscissors</td><td>Medline International Switzerland S&amp;agrave;rl</td><td>UC4337</td><td></td></tr><tr><td>Microvascular clamps</td><td>FST, Germany</td><td>18051-28</td><td></td></tr><tr><td>Papaverine</td><td>ESCA chemicals, Switzerland</td><td>RE 356 803</td><td></td></tr><tr><td>Vein Pick</td><td>Harvard Apparatus, Cambridge, UK</td><td>72-4169</td><td>For single use only</td></tr><tr><td>Saline</td><td>Laboratorium Dr. G. Bichsel AG, , Switzerland</td><td>1330055</td><td></td></tr><tr><td>Polysorb 5-0 suture</td><td>Covidien AG, Switzerland</td><td>UL 202</td><td>Monofilament</td></tr><tr><td>Sulfadoxine and Trimethoprim-Trimethazol</td><td>Werner Stricker AG, Switzerland</td><td>Swissmedic Nr. 50&#039;361</td><td></td></tr><tr><td>Antiseptic- Octenisept</td><td>Sch&amp;uuml;lke &amp;amp; Mayr AG, Switzerland</td><td>GTIN: 4032651214068</td><td></td></tr><tr><td>Phosphate Buffered Saline</td><td>Roth</td><td>1058.1</td><td></td></tr><tr><td>Isobutanol-2-Methylbutane</td><td>Sigma-Aldrich, Switzerland</td><td>M32631-1L</td><td></td></tr><tr><td>Optimum Cutting Temperature compound-Tissue-Tek</td><td>VWR Chemicals, Belgium</td><td>25608-930</td><td></td></tr><tr><td>Cryostat</td><td>Leica, Glattbrugg, Switzerland</td><td>Leica CM1860 UV</td><td></td></tr><tr><td>Glass slide- Superfrost Plus</td><td>Thermo Scientific</td><td>4951PLUS4</td><td></td></tr><tr><td>Mayer&#039;s Haematoxylin</td><td>Sigma-Aldrich, Switzerland</td><td>MHS32-1L</td><td></td></tr><tr><td>Eosin 0.5% aq.</td><td>Sigma-Aldrich, Switzerland</td><td>HT110232-1L</td><td></td></tr><tr><td>Oil Red O</td><td>Sigma-Aldrich, Switzerland</td><td>O0625-25G</td><td></td></tr><tr><td>&amp;alpha;-smooth muscle actin antibody</td><td>Abcam, UK.</td><td>ab7817</td><td></td></tr><tr><td>Macrophage Clone RAM11 antibody</td><td>DAKO, Switzerland</td><td>M063301</td><td></td></tr><tr><td>Hoechst</td><td>Abcam, UK.</td><td>ab145596</td><td></td></tr><tr><td>Goat polyclonal Secondary Antibody (Chromeo 546)</td><td>Abcam, UK.</td><td>ab60316</td><td></td></tr><tr><td>Alexa Fluor 488/547</td><td>Abcam, UK.</td><td></td><td></td></tr><tr><td>Glycergel Mounting Medium, Aqueous</td><td>DAKO, Switzerland</td><td>C056330</td><td></td></tr><tr><td>Hematoxylin for Movat pentachrome staining</td><td>Sigma-Aldrich, Switzerland</td><td>H3136-25G</td><td></td></tr><tr><td>Ferric chloride for Movat pentachrome staining</td><td>Sigma-Aldrich, Switzerland</td><td>157740-100G</td><td></td></tr><tr><td>Iodine for Movat staining</td><td>Sigma-Aldrich, Switzerland</td><td>207772-100G</td><td></td></tr><tr><td>Potassium iodide for Movat pentachrome staining</td><td>Sigma-Aldrich, Switzerland</td><td>60400-100G-F</td><td></td></tr><tr><td>Alcian blue for Movat staining</td><td>Sigma-Aldrich, Switzerland</td><td>A5268-10G</td><td></td></tr><tr><td>Strong Ammonia for Movat pentachrome staining</td><td>Sigma-Aldrich, Switzerland</td><td>320145-500ML</td><td></td></tr><tr><td>Brilliant crocein MOO for Movat pentachrome staining</td><td>Sigma-Aldrich, Switzerland</td><td>210757-50G</td><td></td></tr><tr><td>Acid Fuchsin for Movat pentachrome staining</td><td>Sigma-Aldrich, Switzerland</td><td>F8129-50G</td><td></td></tr><tr><td>Sodium Thiosulfate for Movat pentachrome staining</td><td>Sigma-Aldrich, Switzerland</td><td>72049-250G</td><td></td></tr><tr><td>Phosphotungstic acid for Movat pentachrome staining</td><td>Sigma-Aldrich, Switzerland</td><td>79690-100G</td><td></td></tr><tr><td>Crocin for Movat pentachrome staining</td><td>Sigma-Aldrich, Switzerland</td><td>17304-5G</td><td></td></tr><tr><td>EUKITT for Movat pentachrome staining</td><td>Sigma-Aldrich, Switzerland</td><td>03989-100ML</td><td></td></tr></tbody></table>

the rabbit model of accelerated atherosclerosis: a methodological perspective of the iliac artery balloon injury

在动脉粥样硬化斑块发育和破裂后冠状动脉闭塞引起的急性冠状动脉综合征是工业化国家死亡的主要原因。新西兰白 (NZW) 兔被广泛用作动脉粥样硬化研究的动物模型。当动脉粥样硬化饮食时, 它们会产生自发的损伤;然而, 这需要很长的时间 4-8 月。为了进一步加强和加速动脉粥样硬化, 动脉粥样硬化饮食和机械内皮损伤的结合经常被使用。提出了一种诱导家兔动脉粥样硬化斑块的方法, 用气囊导管阻断动脉粥样硬化饮食喂养的 NZW 兔左髂动脉内皮细胞。这种由气囊导管引起的机械损伤诱发了一连串的炎症反应, 引发了膜的脂质堆积。球囊损伤后动脉粥样硬化斑块显示膜增厚, 脂质浸润广泛, 平滑肌细胞含量高, 巨噬细胞衍生泡沫细胞存在。该技术简单, 重现性好, 在髂动脉内产生受控长度的斑块。整个程序在 20-30 分钟内完成。该手术是安全的低死亡率, 也提供了很大的成功获得大量内膜病变。气囊导管诱发动脉损伤的方法在两周内导致动脉粥样硬化。该模型可用于疾病病理学、诊断影像学的研究和新的治疗策略的评价。

髂动脉球囊损伤无并发症 (图 1) 成功完成。总手术时间范围从20至30分钟的伤害只在一个髂动脉, 35 至45分钟的伤害, 在两个动脉。兔在球囊损伤后1小时内恢复。所有的动物都健康, 没有明显的体重减轻。无感染、水肿或动脉血栓形成。缝合部位除轻度纤维化外, 伤口面积正常。在4周的动脉粥样硬化饮食喂养后, 家兔高胆固醇血症的44±18毫米/升。
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Watch this Scientific Journal Video about The Rabbit Model of Accelerated Atherosclerosis: A Methodological Perspective of the Iliac Artery Balloon Injury at JoVE.com

The Rabbit Model of Accelerated Atherosclerosis: A Methodological Perspective of the Iliac Artery Balloon Injury

动物模型的动脉粥样硬化是必不可少的了解机制和研究新的方法, 以防止斑块的发展或破裂, 在工业化世界的主要死因。该方案采用球囊损伤和胆固醇丰富的饮食组合, 诱导兔髂动脉动脉粥样硬化斑块。

家兔加速动脉粥样硬化模型的研究--髂动脉球囊损伤的方法学观察

Acute coronary syndrome resulting from coronary occlusion following atherosclerotic plaque development and rupture is the leading cause of death in the ...

the-rabbit-model-accelerated-atherosclerosis-methodological

Research

JoVE Journal

Medicine

12.0K 次观看.  University of Fribourg. 动物模型的动脉粥样硬化是必不可少的了解机制和研究新的方法, 以防止斑块的发展或破裂, 在工业化世界的主要死因。该方案采用球囊损伤和胆固醇丰富的饮食组合, 诱导兔髂动脉动脉粥样硬化斑块。

视频: The Rabbit Model of Accelerated Atherosclerosis: A Methodological Perspective of the Iliac Artery Balloon Injury

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

In vivo Near Infrared Fluorescence (NIRF) Intravascular Molecular Imaging of Inflammatory Plaque, a Multimodal Approach to Imaging of Atherosclerosis

The vascular response to injury is a well-orchestrated inflammatory response triggered by the accumulation of macrophages within the vessel wall leading to an accumulation of lipid-laden intra-luminal plaque, smooth muscle cell proliferation and progressive narrowing of the vessel lumen. The formation of such vulnerable plaques prone to rupture underlies the majority of cases of acute myocardial infarction. The complex molecular and cellular inflammatory cascade is orchestrated by the recruitment of T lymphocytes and macrophages and their paracrine effects on endothelial and smooth muscle cells.1
Molecular imaging in atherosclerosis has evolved into an important clinical and research tool that allows in vivo visualization of inflammation and other biological processes. Several recent examples demonstrate the ability to detect high-risk plaques in patients, and assess the effects of pharmacotherapeutics in atherosclerosis.4 While a number of molecular imaging approaches (in particular MRI and PET) can image biological aspects of large vessels such as the carotid arteries, scant options exist for imaging of coronary arteries.2 The advent of high-resolution optical imaging strategies, in particular near-infrared fluorescence (NIRF), coupled with activatable fluorescent probes, have enhanced sensitivity and led to the development of new intravascular strategies to improve biological imaging of human coronary atherosclerosis.
Near infrared fluorescence (NIRF) molecular imaging utilizes excitation light with a defined band width (650-900 nm) as a source of photons that, when delivered to an optical contrast agent or fluorescent probe, emits fluorescence in the NIR window that can be detected using an appropriate emission filter and a high sensitivity charge-coupled camera. As opposed to visible light, NIR light penetrates deeply into tissue, is markedly less attenuated by endogenous photon absorbers such as hemoglobin, lipid and water, and enables high target-to-background ratios due to reduced autofluorescence in the NIR window. Imaging within the NIR 'window' can substantially improve the potential for in vivo imaging.2,5
Inflammatory cysteine proteases have been well studied using activatable NIRF probes10, and play important roles in atherogenesis. Via degradation of the extracellular matrix, cysteine proteases contribute importantly to the progression and complications of atherosclerosis8. In particular, the cysteine protease, cathepsin B, is highly expressed and colocalizes with macrophages in experimental murine, rabbit, and human atheromata.3,6,7 In addition, cathepsin B activity in plaques can be sensed in vivo utilizing a previously described 1-D intravascular near-infrared fluorescence technology6, in conjunction with an injectable nanosensor agent that consists of a poly-lysine polymer backbone derivatized with multiple NIR fluorochromes (VM110/Prosense750, ex/em 750/780nm, VisEn Medical, Woburn, MA) that results in strong intramolecular quenching at baseline.10 Following targeted enzymatic cleavage by cysteine proteases such as cathepsin B (known to colocalize with plaque macrophages), the fluorochromes separate, resulting in substantial amplification of the NIRF signal. Intravascular detection of NIR fluorescence signal by the utilized novel 2D intravascular NIRF catheter now enables high-resolution, geometrically accurate in vivo detection of cathepsin B activity in inflamed plaque.
In vivo molecular imaging of atherosclerosis using catheter-based 2D NIRF imaging, as opposed to a prior 1-D spectroscopic approach,6 is a novel and promising tool that utilizes augmented protease activity in macrophage-rich plaque to detect vascular inflammation.11,12 The following research protocol describes the use of an intravascular 2-dimensional NIRF catheter to image and characterize plaque structure utilizing key aspects of plaque biology. It is a translatable platform that when integrated with existing clinical imaging technologies including angiography and intravascular ultrasound (IVUS), offers a unique and novel integrated multimodal molecular imaging technique that distinguishes inflammatory atheromata, and allows detection of intravascular NIRF signals in human-sized coronary arteries.

In vivo Near Infrared Fluorescence NIRF Intravascular Molecular Imaging of Inflammatory Plaque, a Multimodal Approach to Imaging of Atherosclerosis

We detail a new near-infrared fluorescence (NIRF) catheter for 2-dimensional intravascular molecular imaging of plaque biology in vivo. The NIRF catheter can visualize key biological processes such as inflammation by reporting on the presence of plaque-avid activatable and targeted NIR fluorochromes. The catheter utilizes clinical engineering and power requirements and is targeted for application in human coronary arteries. The following research study describes a multimodal imaging strategy that utilizes a novel in vivo intravascular NIRF catheter to image and quantify inflammatory plaque in proteolytically active inflamed rabbit atheromata.

体内近红外荧光（NIRF）血管内分子成像炎性斑块，多式联运方针成像动脉粥样硬化

The vascular response to injury is a well-orchestrated inflammatory response triggered by the accumulation of macrophages within the vessel wall leading ...

Inducing Myointimal Hyperplasia Versus Atherosclerosis in Mice: An Introduction of Two Valid Models

Various in vivo laboratory rodent models for the induction of artery stenosis have been established to mimic diseases that include arterial plaque formation and stenosis, as observed for example in ischemic heart disease. Two highly reproducible mouse models&#160;&#8211;&#160;both resulting in artery stenosis but each underlying a different pathway of development&#160;&#8211; are introduced here. The models represent the two most common causes of artery stenosis; namely one mouse model for each myointimal hyperplasia, and atherosclerosis are shown. To induce myointimal hyperplasia, a balloon catheter injury of the abdominal aorta is performed. For the development of atherosclerotic plaque, the ApoE -/- mouse model in combination with western fatty diet is used. Different model-adapted options for the measurement and evaluation of the results are named and described in this manuscript. The introduction and comparison of these two models provides information for scientists to choose the appropriate artery stenosis model in accordance to the scientific question asked.

This video shows two models of intimal plaque development in murine arteries and emphasizes the differences in myointimal hyperplasia and atherosclerosis.

诱导肌内膜增生对战动脉粥样硬化小鼠:两种有效的模型介绍

Various in vivo laboratory rodent models for the induction of artery stenosis have been established to mimic diseases that include arterial plaque ...

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 ...

A Murine Model of Arterial Restenosis: Technical Aspects of Femoral Wire Injury

Cardiovascular disease caused by atherosclerosis is the leading cause of death in the developed world. Narrowing of the vessel lumen, due to atherosclerotic plaque development or the rupturing of established plaques, interrupts normal blood flow leading to various morbidities such as myocardial infarction and stroke. In the clinic endovascular procedures such as angioplasty are commonly performed to reopen the lumen. However, these treatments inevitably damage the vessel wall as well as the vascular endothelium, triggering an excessive healing response and the development of a neointimal plaque that extends into the lumen causing vessel restenosis (re-narrowing). Restenosis remains a major cause of failure of endovascular treatments for atherosclerosis. Thus, preclinical animal models of restenosis are vitally important for investigating the pathophysiological mechanisms as well as translational approaches to vascular interventions. Among several murine experimental models, femoral artery wire injury is widely accepted as the most suitable for studies of post-angioplasty restenosis because it closely resembles the angioplasty procedure that injures both endothelium and vessel wall. However, many researchers have difficulty utilizing this model due to its high degree of technical difficulty. This is primarily because a metal wire needs to be inserted into the femoral artery, which is approximately three times thinner than the wire, to generate sufficient injury to induce prominent neointima. Here, we describe the essential surgical details to effectively overcome the major technical difficulties of this model. By following the presented procedures, performing the mouse femoral artery wire injury becomes easier. Once familiarized, the whole procedure can be completed within 20 min.

The mouse femoral artery wire injury model of restenosis is technically challenging. In this protocol we show the key technical details essential for successfully performing wire injury to induce consistent&#160;neointima for studies of restenosis.

动脉再狭窄小鼠模型:股导线损伤的技术方面

Cardiovascular disease caused by atherosclerosis is the leading cause of death in the developed world. Narrowing of the vessel lumen, due to ...

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 ...

Induction of Accelerated Atherosclerosis in Mice: The "Wire-Injury" Model

Atherosclerosis is a proliferative fibro-inflammatory disease developing in the arterial wall, inducing a deficient blood flow or a lack of blood flow. Moreover, by rupture of the defective vascular wall, atherosclerosis induces occlusive thrombus formation, which represents the main cause of myocardial infarction or stroke and the most frequent cause of death. Despite the advances in the cardiovascular field, many questions remain unanswered, and additional basic research is essential to improve our understanding of the molecular mechanisms during atherosclerosis and its effects. Due to limited clinical studies, there is a need for representative animal models recreating atherosclerotic conditions such as neointima formation after stent implantation, balloon angioplasty, or endarterectomy. Since the mouse presents many advantages and is the most frequently used model for studying molecular processes, the current study proposes an invasive procedure of endothelial denudation, also known as the wire-injury model, which is representative of the human condition of neointima formation in arteries after revascularization procedures.

This study describes an invasive procedure for the induction of accelerated atherosclerosis in mice. In comparison to other methods using electric- or cryo-induced injury, mechanical-induced injury mimics the human condition of restenosis after revascularization therapies and is ideal for the study of the molecular mechanisms involved.

小鼠加速动脉硬化的诱导:"电线损伤"模型

Atherosclerosis is a proliferative fibro-inflammatory disease developing in the arterial wall, inducing a deficient blood flow or a lack of blood flow. ...

Reproducible Arterial Denudation Injury by Infrarenal Abdominal Aortic Clamping in a Murine Model

Percutaneous vascular interventions uniformly result in arterial denudation injuries that subsequently lead to thrombosis and restenosis. These complications can be attributed to impairments in re-endothelialization within the wound margins. Yet, the cellular and molecular mechanisms of re-endothelialization remain to be defined. While several animal models to study re-endothelialization after arterial denudation are available, few are performed in the mouse because of surgical limitations. This undermines the opportunity to exploit transgenic mouse lines and investigate the contribution of specific genes to the process of re-endothelialization. Here, we present a step-by-step protocol for creating a highly reproducible murine model of arterial denudation injury in the infrarenal abdominal aorta using external vascular clamping. Immunocytochemical staining of injured aortas for fibrinogen and &#946;-catenin demonstrate the exposure of a pro-thrombotic surface and the border of intact endothelium, respectively. The method presented here has the advantages of speed, excellent overall survival rate, and relative technical ease, creating a uniquely practical tool for imposing arterial denudation injury in transgenic mouse models. Using this method, investigators may elucidate the mechanisms of re-endothelialization under normal or pathological conditions.

Understanding the cellular and molecular mechanisms of re-endothelialization following arterial denudation injury is of paramount importance in preventing thrombosis and restenosis of arteries. Here we describe a protocol for reproducible arterial denudation injury of the infrarenal abdominal aorta. The procedure was developed to investigate the underlying mechanisms that regulate endothelial regeneration using mouse models.

重复性的动脉剥脱损伤腹主动脉阻断在小鼠模型

Percutaneous vascular interventions uniformly result in arterial denudation injuries that subsequently lead to thrombosis and restenosis. These ...

Balloon-based Injury to Induce Myointimal Hyperplasia in the Mouse Abdominal Aorta

The use of animal models is essential for a better understanding of MH, one major cause for arterial stenosis.In this article, we demonstrate a murine balloon denudation model, which is comparable with established vessel injury models in large animals. The aorta denudation model with balloon catheters mimics the clinical setting and leads to comparable pathobiological and physiological changes. Briefly, after performing a horizontal incision in the aorta abdominalis, a balloon catheter will be inserted into the vessel, inflated, and introduced retrogradely. Inflation of the balloon will lead to intima injury and overdistension of the vessel. After removing the catheter, the aortic incision will be closed with single stiches. The model shown in this article is reproducible, easy to perform, and can be established quickly and reliably. It is especially suitable for evaluating expensive experimental therapeutic agents, which can be applied in an economical fashion. By using different knockout-mouse strains, the impact of different genes on MH development can be assessed.

This article demonstrates a murine model to study the development of myointimal hyperplasia (MH) after aortic balloon injury.

球囊损伤诱发小鼠腹主动脉 Myointimal 增生的研究

The use of animal models is essential for a better understanding of MH, one major cause for arterial stenosis.In this article, we demonstrate a murine ...

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 ...