Name: procedure for human saphenous veins ex vivo perfusion and external reinforcement
Uploaded: 2014-10-01T15:00:00.000+00:00
Duration: 9 min 50 s
Description: Watch this Scientific Journal Video about Procedure for Human Saphenous Veins Ex Vivo Perfusion and External Reinforcement at JoVE.com

这项工作是从SNF [31003A-138528]，该Octav和马塞拉Botnar基金会，诺华基金会和艾玛Muschamp基金会的资助。我们感谢马丁Lambelet，和让 - 克里斯托夫施特勒的出色技术援助。

洛桑大学的伦理委员会批准了试验，这是按照1975年赫尔辛基宣言提出的原则，修订于1983年为使用人体组织。 1，人类大隐静脉丰收<ol><li>从发生下肢搭桥手术缺血患者获得非曲张的大隐静脉的盈余部分。在手术室，消毒与碘溶液的整个腿和悬垂性，患者从腹股沟到脚露出的腿。 </li><li>请从腹股沟到膝盖正中切口（在离开中断的皮肤部分）。 </li><li>收获大隐静脉与周围组织（非接触式技术）的蒂。用4-0丝束静脉安全侧枝。在4℃下立即存储最小为9厘米的大隐静脉的长剩余段，具有2.5-4毫米的外径在RPMI-1640ģlutamax介质，辅以12.5％胎牛血清，并把它带到实验室。 </li></ol> 2，每股内含价值的设计<ol><li>装配在图1所示的通用设备。高压灭菌的所有设备，并保持所有组件在无菌条件下。此外，确保系统具有防水和不漏化学品到培养基中。为封面用聚甲基丙烯酸甲酯（PMMA-GS）。钢（X5铬镍18 10）和聚甲醛塑料（聚甲醛）作为静脉的支持。 </li><li>设计灌注腔至所需的几何形状，以使放置在静脉和其连接的。确保深度（或半径如果使用圆柱形结构）是至少2.5厘米所以它允许随着恒定覆盖的培养基（ 图1）最小屈曲容器和扩张。密封是一个主要问题，是长方形的PMMA-GS建设使用的原因。 </li><li>设计静脉支持所需的geometr年。避免静脉扭结或以上腹胀，允许长度调节通过推动或拉动（螺钉不能用于这一目的，因为静脉将与螺钉扭沿）。 注:完全钢杆被2连接滑动的L形件支持2静脉缸（直径5毫米，以适应容器）和静脉（ 图1B和图2）被用在这里。 </li><li>设计压力柱，使得施加到该系统的"静息压力"是:P = 0-10 = HXρXG，其p =压力（N / m 2时，Pa）的流体柱（米）为h =高度ρ =液体（千克/米3）和g =重力常数（9.81米/秒2）的密度。设计四个连接管，从顶部到底部:以施加压力，对所述流出（从静脉），所述流入（到静脉），并允许更换培养基。 </li><li>准备中。根据以往的研究5,11-14，选择的RPMI-1640，辅以Glutamax的，12.5％胎牛血清和1％抗生素 - 抗真菌剂溶液（10,000 U / ml的青霉素G，加上10毫克/毫升硫酸链霉素，外加25毫克/毫升两性霉素B，加0.5微克/毫升:庆大霉素）。剪切应力（SS）由SS = 4μQ/π* R 3 Q给定的流速（毫升/秒），R的静脉段的半径（厘米），并且μ是灌注介质的粘度。 <ol><li>调制的SS通过另外的70 kDa的葡聚糖调节粘度。测量用粘度计的粘度。这里，加8％70kDa的葡聚糖设置SS到9-15达因/厘米2。 </li></ol></li><li>设置齿轮泵诱导的60个脉冲/分钟，恒定幅度产生的150±15毫升/分钟的单向流动，独立于由计算机系统中的应用和控制的压力脉动心形信号。确保该驱动软件集成恒定采集与监控压力，流速，脉搏速率，和信号。如果需要，使用第二个泵（非同步hronized），以产生一个非层流，湍流。 </li></ol> 3 EVPS组件（图1） <ol><li>在开始之前，请确保所有的设备是无菌的。执行层流罩下无菌以下所有步骤。 </li><li>将静脉在培养皿中充满介质。使用手术刀片，分化静脉分成3等份。 </li><li>立即冲洗的PBS一段。划分段三部分，在福尔马林固定一个用于形态。冻结的其它两个用于定量转录物（RT-PCR）和蛋白质（免疫印迹）分析。考虑这些片段作为对照，非静脉灌注。 </li><li>用剩下的2段灌注。 <ol><li>轻轻地注入介质插入静脉并确定正常流动方向;在阀门存在的静脉是相反的。 </li><li>密封脉是最重要的实验取得成功。检查通过络脉泄漏。争取用6-0丝线缝合任何泄漏。 </li> &lt;/ OL&gt; </li><li>连接这两个金属圆柱体之间的静脉段，一端在一个时间（2.3， 图1）。确保与Ethibon 3-0气缸周围的凹部（ 图1A和B）。 <ol><li>将整个静脉段进入灌注腔预先填充有介质。重复同样的步骤进行的第二部分。 注意:不正确密封静脉缸将泄漏的来源，需要再次介入，并显著增加感染和实验失败的风险。 </li></ol></li><li>加强（目）的第二段，松开两个汽缸（装有静脉）从L形件（2.3和图1）。 <ol><li>要轻柔，不要触碰静脉使用任何手段。第一张幻灯片网格上的圆筒，然后到静脉。推/拉推撞就会在静脉网。 </li><li>一旦网格覆盖静脉的整个表面上固定的第jacketed静脉与Ethibon 3-0缸。 </li><li>重新组装静脉/气缸化合物为L形支撑，并将其转移到灌注腔，预先填充有介质。 </li></ol></li><li>使用过氧化处理的硅橡胶管具有3.2毫米的内径（在和流出），每个金属筒连接到一个Y型分离器。 </li><li>连接的流出分离器使用相同类型管道的第二Y型分离器。从这个Y型分线器，​​使用一管来测量通过两艘船的灌注压。连接的另一个回柱，以形成一个封闭的回路系统（ 图2）。 </li><li>内的培养箱中，使用长（一米长）管与低压塔连接到泵头部。 </li><li>完成建立连接的泵头部，以在流入Y型分光器与另一个长长度管（ 图1）。 </li></ol> 4，静脉灌注<ol><li>之后的每股内含价值组件已经合作mpleted，填充介质柱（保持低于静脉流出道，让加气站）。添加更多的介质进入塔之前，系统已满。将所有系统进入维持在37±0.1℃，pH值为孵化器保持恒定在7.40±0.01（使用基于恒基哈塞尔巴赫方程的CO 2 / pH值算法）。 </li><li>使齿轮泵头部培养箱外，并将其连接到齿轮泵的驱动。螺丝杆，以确保大会。 </li><li>交换机上的泵浦功率，确保其在驱动软件激活，允许5分钟的灭火剂能均匀地分布在每一个车厢。 </li><li>要监视的压力，用动脉在线监测。连接EVPS压力输出（它对应于动脉导管），以连接到计算机中的压力换能器。 <ol><li>确保将管完全充满介质，并且不包含任何气泡。脱泡的培养体系，通过"动脉里NE"管（ 图2）。注意显示器，寻找60个脉冲/恒定振幅的分的心搏动信号。在这点上，平均压力为0-10毫米汞柱之间。如果压力&lt;0且列逐渐清空寻找（静脉和管之间的静脉侧支或密封不足）的泄漏。 </li></ol></li><li>设置最小压力至6毫米汞柱的静脉试验或在90毫米汞柱为动脉的测试。在这些条件下，空气喷射器施加所需压力的柱和系统。 </li><li>通过使用连接到高压塔的管更换培养基，每2天。为了防止压力变化造成的破坏，先打开柱插头。 </li></ol> 5，完成灌注的<ol><li>灌注后3天或7天:乘坐每股内含价值从孵化器和下马的静脉。丢弃5毫米近端和远端静脉端连接于设备。切中央5毫米厚的RIN从剩余的部分GS和福尔马林（形态）修复。冻结剩余的碎片和降低成粉末作进一步的分子分析。 </li></ol>

<ol>
	<li>Sal Go, A., 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=Executive+summary:+heart+disease+and+stroke+statistics--2014+update:+a+report+from+the+american+heart+association.">Executive summary: heart disease and stroke statistics--2014 update: a report from the american heart association.</a> Circulation. 129, 399-410 (2014).</li><li>Sal Conte, M., 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=Results+of+PREVENT+III:+a+multicenter,+randomized+trial+of+edifoligide+for+the+prevention+of+vein+graft+failure+in+lower+extremity+bypass+surgery.">Results of PREVENT III: a multicenter, randomized trial of edifoligide for the prevention of vein graft failure in lower extremity bypass surgery.</a> Journal of Vascular Surgery. 43, 742-751 (2006).</li><li>Yu, P., Nguyen, B. T., Tao, M., Bai, Y., Ozaki, C. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+vein+graft+hemodynamic+manipulations+to+enhance+experimental+utility.">Mouse vein graft hemodynamic manipulations to enhance experimental utility.</a> The American Journal of Pathology. 178, 2910-2919 (2011).</li><li>Davies, M. G., Hagen, P. O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reprinted+article+&amp;#34;Pathophysiology+of+vein+graft+failure:+a+review&amp;#34;.">Reprinted article &amp;#34;Pathophysiology of vein graft failure: a review&amp;#34;.</a> European journal of vascular and endovascular surgery : the official journal of the European Society for Vascular Surgery. 42, S19-S29 (2011).</li><li>Berard, X., 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=Role+of+hemodynamic+forces+in+the+ex+vivo+arterialization+of+human+saphenous+veins.">Role of hemodynamic forces in the ex vivo arterialization of human saphenous veins.</a> Journal of Vascular Surgery. 57, 1371-1382 (2013).</li><li>Vijayan, V., 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=Long-term+reduction+of+medial+and+intimal+thickening+in+porcine+saphenous+vein+grafts+with+a+polyglactin+biodegradable+external+sheath.">Long-term reduction of medial and intimal thickening in porcine saphenous vein grafts with a polyglactin biodegradable external sheath.</a> Journal of Vascular Surgery. 40, 1011-1019 (2004).</li><li>Jeremy, J. Y., 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=On+the+biology+of+saphenous+vein+grafts+fitted+with+external+synthetic+sheaths+and+stents.">On the biology of saphenous vein grafts fitted with external synthetic sheaths and stents.</a> Biomaterials. 28, 895-908 (2007).</li><li>Zilla, P., 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=Constrictive+external+nitinol+meshes+inhibit+vein+graft+intimal+hyperplasia+in+nonhuman+primates.">Constrictive external nitinol meshes inhibit vein graft intimal hyperplasia in nonhuman primates.</a> The Journal of Thoracic and Cardiovascular Surgery. 136, 717-725 (2008).</li><li>Zilla, P., 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=Utilization+of+shape+memory+in+external+vein-graft+meshes+allows+extreme+diameter+constriction+for+suppressing+intimal+hyperplasia:+a+non-human+primate+study.">Utilization of shape memory in external vein-graft meshes allows extreme diameter constriction for suppressing intimal hyperplasia: a non-human primate study.</a> Journal of Vascular Surgery. 49, 1532-1542 (2009).</li><li>Yeoman, M. 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=A+constitutive+model+for+the+warp-weft+coupled+non-linear+behavior+of+knitted+biomedical+textiles.">A constitutive model for the warp-weft coupled non-linear behavior of knitted biomedical textiles.</a> Biomaterials. 31, 8484-8493 (2010).</li><li>Longchamp, A., 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=The+use+of+external+mesh+reinforcement+to+reduce+intimal+hyperplasia+and+preserve+the+structure+of+human+saphenous+veins.">The use of external mesh reinforcement to reduce intimal hyperplasia and preserve the structure of human saphenous veins.</a> Biomaterials. 35, 2588-2599 (2014).</li><li>Saucy, F., 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=Ex+vivo+pulsatile+perfusion+of+human+saphenous+veins+induces+intimal+hyperplasia+and+increased+levels+of+the+plasminogen+activator+inhibitor+1.">Ex vivo pulsatile perfusion of human saphenous veins induces intimal hyperplasia and increased levels of the plasminogen activator inhibitor 1.</a> European Surgical Research. Europaische Chirurgische Forschung. Recherches Chirurgicales Europeennes. 45, 50-59 (2010).</li><li>Dubuis, C., 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=Atorvastatin-loaded+hydrogel+affects+the+smooth+muscle+cells+of+human+veins.">Atorvastatin-loaded hydrogel affects the smooth muscle cells of human veins.</a> The Journal of pharmacology and experimental. 347, 574-581 (2013).</li><li>Deglise, 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=Increased+connexin43+expression+in+human+saphenous+veins+in+culture+is+associated+with+intimal+hyperplasia.">Increased connexin43 expression in human saphenous veins in culture is associated with intimal hyperplasia.</a> Journal of Vascular Surgery. 41, 1043-1052 (2005).</li><li>Muto, A., Model, L., Ziegler, K., Eghbalieh, S. D., Dardik, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms+of+vein+graft+adaptation+to+the+arterial+circulation:+insights+into+the+neointimal+algorithm+and+management+strategies.">Mechanisms of vein graft adaptation to the arterial circulation: insights into the neointimal algorithm and management strategies.</a> Circulation Journal : Official Journal of the Japanese Circulation Society. 74, 1501-1512 (2010).</li><li>Tao, M., 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+simplified+murine+intimal+hyperplasia+model+founded+on+a+focal+carotid+stenosis.">A simplified murine intimal hyperplasia model founded on a focal carotid stenosis.</a> The American Journal of Pathology. 182, 277-287 (2013).</li><li>Berard, X., 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=Salvage+treatment+for+venous+aneurysm+complicating+vascular+access+arteriovenous+fistula:+use+of+an+exoprosthesis+to+reinforce+the+vein+after+aneurysmorrhaphy.">Salvage treatment for venous aneurysm complicating vascular access arteriovenous fistula: use of an exoprosthesis to reinforce the vein after aneurysmorrhaphy.</a> European Journal of Vascular and Endovascular Surgery : the Official Journal of the European Society for Vascular Surgery. 40, 100-106 (2010).</li></ol>

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这项研究揭示了体外静脉灌注系统（每股内含价值）在人类静脉进行大量的血流动力学研究。这个系统允许根据在没有通过体内循环细胞所释放的加重炎症和生长因子定义的血流动力学参数隐静脉灌注。因此，它提供了更好的理解涉及IH在人静脉移植物5,11,12,15的控制的基本途径。 可重复和可定量的血液动力学扰动体内限定。几个复杂的鼠显微手术操作进行了描述。通过采用来自供体小鼠腔静脉的介入旁路同系移植模式进入右颈总动脉和流出分支结扎的额外创造，中移植或颈总动脉狭窄，水流和SS剧烈下降，增强了IH 3。外向与内向重塑还可以是跨rogated使用中焦与远端的颈总动脉狭窄16。在大型动物（羊，猪，和狒狒），旁路移植物在技术上更简单，并代表一个有吸引力的方式来测试的临床前人体尺寸的装置，如在本研究8-10中使用的网格。然而，它的成本和有效的分子工具的缺乏限制了使用这些策略。最后，这些流程操作不断改变壁紧张，未能询问单一成分。此外，血流动力学和免疫系统和内分泌系统进一步之间的错综复杂的关系，限制单个演员的分析。 一些问题的出现与使用的每股内含价值的。 1）低度细菌污染往往伴随着人类静脉收获和离体情况下的循环细胞支架感染的一个重要原因。这主要是防止由手工洗涤的碎片分开，然后高压灭菌所有材料之前使用。此外该组件在少于90分钟，并在严格无菌进行。 2）密封的承受反复消毒。出于这个原因，一个长方形的PMMA-GS结构被使用时，避免了使用接头和限制形变。 3）SS和壁张力计算在定义的时间点，根据该血管腔半径（组织学），流程和粘度为常数。纵向成像（高清晰度摄像头，激光或多普勒），可持续监控静脉内径和/或流量将提供本地流量变化的详细信息，并允许循环应变计算的融合。 4）两个平行脉段可以具有在其壁顺应性和半径不受控制的差异。因此，我们只从同一静脉比较片段，并假定在相同的压力下，将流量特性是在这两个区段相似。 在这项研究中，静脉被提交到脉动层流;然而，50％氧˚F内膜增生性病变发生在静脉移植物，在层流被破坏的端 - 端吻合口周围的区域。湍流条件也可以与另外的第二泵来建模的，非同步的第一泵。将来的研究将被执行以具体评估IH层流中断和钢筋网的潜在有益效果的影响。有趣的是，该网状物的柔性结构允许周向缠绕在吻合部位，如已经进行的修复动脉瘤瘘17（ 图4）。因此，网格可以证明是有用的，以限制吻合口周围扩张，层流中断，从而降低IH在吻合部位。这可能是特别有益的远侧旁路移植，经常受静脉和胫骨或腓骨动脉之间直径的不匹配。 总之，此处所示的设置允许人类诉平行灌注一型，在相同的血流动力学条件。这些数据表明，使用外部的大孔的管状聚酯网状物是一种有效的方法，以限制IH的发展中静脉插入动脉环境11移植。该系统可实现利润研究若干领域。特别是，它的出现是一个强大的工具来执行测试的可行性和各种方法的效率，减少IH在人力物力临床前研究，是一个有价值的补充体内动物模型。其他假体支架或网格涂有药剂将使用此方法6-10进行评价。此外，人们可以设想以测试局部施加的药理学的分子，以防止IH在人体组织中，接近生理状态下。基因疗法的干预也是可以实现的，转导静脉段过度或感兴趣的沉默靶基因。 总之，我们的系统会增加我们的未derstanding对人类移植静脉疾病的血流动力学的贡献。它提供了一个创新的平台，以测试新的治疗策略和可能出现的"板凳旁边的翻译工具。"

心血管疾病是发病率和死亡率在西方国家1的主要原因。尽管血管内治疗取得了进步，搭桥手术仍然是现代治疗的主体，从而五十多万元静脉移植在美国每年进行的。然而，尽管几十年的研究，下肢静脉移植30-60％范围内的最初几年，由于内膜增生（1H）2失败。机械力，特别是低切应力（SS）和高壁张力，关键是在这个增生反应3,4的启动和发展。为了解决这个问题，一个离体血管灌注系统（EVPS）中产生的学习，严格控制的血流动力学条件（压力和剪切应力），人隐静脉的行为下。在这项研究中，下列插入到动脉样循环，高压（平均= 100毫米汞柱）足以刺激prolif关合作和平滑肌细胞进入血管内膜层（1H）迁移。 哺乳动物的研究表明使用外部加固作为一种有效的方法来支持"静脉动脉化"，并抵消急性血液动力学改变静脉面临一次植入动脉周围环境。网状防止过 ​​度膨胀，增大的剪切应力，并降低了壁张力，因而IH 6-10。然而，潜在的机制和其适用于人类静脉改善旁路通畅性没有得到充分的表征。我们EVPS用于比较，在条件模拟的静脉面临一次插入动脉方案的改变（高剪切应力和压力），在外部大孔聚酯管状网的不存在和存在的大隐静脉的行为。防止病理性重构和IH，网格提供了其潜在的临床有效率11证据</sup&gt;。 这项研究1）引入体外控制下的压力和剪切应力2）人类隐静脉灌注模型显示，外部大孔涤纶网眼降低IH和提供了其潜在的临床应用至关重要的信息。

<table><tbody><tr><td>RPMI 1640 - Glutamax</td><td>Life Technologies</td><td>61870-010</td><td></td></tr><tr><td>Penicilline/Streptomycine/Fungizone</td><td>Bioconcept</td><td>4-02F00-H</td><td></td></tr><tr><td>Dextran from Leuconostoc spp. 500 g</td><td>Sigma-Aldrich</td><td>31390</td><td></td></tr><tr><td>Tampon PBS CHUV pH 7.1-7.3 1 L</td><td>Laboratorium und Grosse Apotheke Dr. G. Bichsel AG</td><td>100 0 324 00</td><td></td></tr><tr><td>Cryosectionning embedding medium - Tissue-Tek OCT Compound</td><td>Fisher Scientific</td><td>14-373-65</td><td></td></tr><tr><td>Silicon Tubing (Peroxide) L/S 16 (96400-16 ) - 7.5 m</td><td>Idex Health &amp;amp; Science GMBH</td><td>MF0037ST</td><td></td></tr><tr><td>Y-splitter&amp;nbsp;</td><td>Idex Health &amp;amp; Science GMBH</td><td>Y-connector</td><td></td></tr><tr><td>35 mm Culture dish</td><td>Sigma-Aldrich</td><td>CLS430165-100EA</td><td></td></tr><tr><td>15 ml Falcon tube</td><td>BD Bioscence</td><td>352096</td><td></td></tr><tr><td>50 ml Falcon tube</td><td>BD Bioscence</td><td>352098</td><td></td></tr><tr><td>Gearing pump - Reglo-Z</td><td>Idex Health &amp;amp; Science GMBH</td><td>SM 895&amp;nbsp;</td><td>&amp;nbsp;App-Nr 03736-00194</td></tr><tr><td>Pump Head</td><td>Idex Health &amp;amp; Science GMBH</td><td>MI0008&amp;nbsp;</td><td></td></tr><tr><td>Monitoring Kit TRANSPAC IV</td><td>icumedical</td><td>011-0J736-01</td><td></td></tr><tr><td>20 ml Syringes</td><td>B. Braun Medical SA</td><td>4612041-02</td><td></td></tr><tr><td>Etibon 3-0 FS-2</td><td>Ethicon- Johnson&amp;amp;Johnson</td><td>EH7346H</td><td></td></tr><tr><td>Mesh ProVena 6-8mm</td><td>B. Braun Medical SA</td><td>1105012-14</td><td></td></tr><tr><td>NaCl: Sodium chloride solution perfusion 0.9% (100 ml)</td><td>B. Braun Medical SA</td><td>534534</td><td></td></tr><tr><td>Masterflex L/S Standard Drive</td><td>Cole-Parmer Instrument Co</td><td>7521-10</td><td></td></tr><tr><td>Acquisition card</td><td>National Instruments</td><td>PCI-6024 E</td><td></td></tr><tr><td>Flowmeter module</td><td>Transonic Systems Inc.</td><td>TS410 and T402</td><td></td></tr><tr><td>Stopcock with 3-ways</td><td>BD Connexta Luerlock</td><td>394600</td><td></td></tr><tr><td>Millex Filter</td><td>Milian</td><td>SE2M229I04</td><td></td></tr></tbody></table>

procedure for human saphenous veins ex vivo perfusion and external reinforcement

现代疗法的广泛闭塞性动脉疾病的主要方法是静脉旁路移植术。然而，它的耐久性受到威胁内膜增生（1H），最终导致血管闭塞和移植失败。的机械力，特别是在低剪切应力和高壁张力，被认为是发起和维持这些细胞和分子变化，但其确切的贡献仍有待解开。以选择性地评估的压力和剪切应力IH的生物学上的作用， 离体灌注系统（EVPS）已创建灌注下动脉方案的大隐静脉（高剪切应力和高压力）的片段。进一步的技术创新，使两部分的同时灌注同出一脉，一个是外部的网状钢筋。脉中使用非接触式技术收获并立即在EVPS转移到实验室进行组装。新鲜ISOL的一个片段ATED静脉未灌注（对照组，第0天）。另外两个节段灌注长达7天，一个是完全为4毫米（直径）外部网的庇护。压力，流速，和脉冲速率连续地监测并调节以模仿血液动力学条件盛行的股动脉。在灌注完成后，静脉被拆卸，并用于组织学和分子分析。下离体条件下，高压灌注（动脉，平均= 100毫米汞柱）足以产生IH和人类静脉的重塑。这些改变都减小外部聚酯丝网的存在。

的每股内含价值提供了一个有价值的工具，以独立评估血流动力学的力量对人类隐静脉移植重建和IH。 图1示出了灌注腔和静脉支撑。在图1A和B，前静脉载体（ 图1A）和之后（ 图1B）组件，分别被描绘。它是由（从顶部至底部）的1平纹不锈钢管尺寸为9厘米，这可作为2为L形件，可以容易地滑动（从左至右）的载体，并提供了可靠的技术，以调节支承尺寸的静脉。每个这些片段的持有POM盘，以适应钢筒（静脉连接器）固定在适当位置以集成螺钉（箭头）。 图1C-D示出了单独使用（C）和插入静脉载体（D）的后灌注腔。在灌注腔，凹部被设计成保持在适当位置（顶部）的静脉的支持，并避免在连接管的扭结进出从静脉（底部）。 图2示出的实时图像（ 图2A）和EVPS的示意图（ 图2B）。灌注腔，静脉和它的支撑，以及在高压塔，被保持在受控的环境（温度，CO 2和O 2），而泵，压力喷射器，和控制装置全部留在培养箱外。该图中示出了齿轮泵（1），其生成由计算机（2），其监视流速（3），压力控制的脉冲信号（4），并且控制最小舒张压（5）;相同的隐静脉的两段并联连接到内部分离灌注腔室的灌注泵（6a和6b）放置在细胞培养孵育箱。 在图3中，组织形态学分析表明，外部强化防止内膜增生和病理媒体改造后的高压下7天（动脉方案，平均为100毫米汞柱），否则灌注观察。在图3A中 ，染色的苏木精-曙红代表性组织切片（HE）揭示了内腔的内皮细胞的细胞核和平滑肌细胞的细胞核中的各种情况下，介质层衬里。 图3B-C示出了代表性的范Gieson染色弹力膜（ VGEL）染色切片。在图3B中 ，内膜增厚（IH）在静脉灌注在高压（平均= 100毫米汞柱），连续7天进行比较，以控制非灌注静脉，这种现象在很大程度上降低在外部网孔的存在。 图3C示出病理向外重塑和媒体变薄的血管受到7天动脉压。这在很大程度上防止了由外部加固。 FurthermorE，在图3D中，Masson三色染色（蓝色=结缔组织，红色=肌肉）关联这个病理重塑仅为三个肌肉层和平滑肌细胞的积累，内层（内膜）中的一个持久性。外部加固保留校董会和媒体结构的分布。 图4示出的外部网孔的当前临床应用。一个说明性的例子是由动脉瘤动静脉瘘（血液透析通路）的外部加强件提供。 图4A示出了静脉加强件的时间进程表示（从顶部至底部）。首先，网格周围放置一个刚性管，而静脉的末端被固定到心轴（上图）。然后，静脉穿过管由于芯棒被拉扯。一旦静脉已经到位，将管缓慢缩回，留下周围的静脉网状（UPPER和中图）。在这种特定情况下，该步骤被重复的静脉的两侧，并在静脉段和网状增强件是由一个端-端吻合术（下面板）组装而成。 图4B中提供的动静脉端的放大视图 - 至侧吻合，这表明网状缠绕在吻合沿动脉后壁捆扎它。 <img alt="图1" fo:content-width="5in" src="/files/ftp_upload/52079/52079fig1highres.jpg" width="500" /> 图1中的静脉支撑和灌注腔。 A.静脉支撑是由1滑动管，2为L形件，光盘和气缸（形成顶部至底部）B.静脉支撑一旦组装。C.视图灌注腔。D.灌注室被设计到位，以保持静脉支撑，并允许其连接离子的静脉和连接管。 <img alt="图2" fo:content-width="5in" src="/files/ftp_upload/52079/52079fig2highres.jpg" width="500" /> 图2中的离体灌注系统。答完全组装EVPS在细胞培养箱中。B.示意图。 1）泵产生搏动心波; 2）在计算机控制的压力，流量（类型，速度和幅度）; 3）流量计; 4）中的压力换能器 - 动脉管路; 5）高压注射器; 6）一个非常相同的大隐静脉的两段无灌注（6A）或与外部钢筋网（6B）。 <img alt="图3" fo:content-width="5in" src="/files/ftp_upload/52079/52079fig3highres.jpg" width="500" /> 图3:外部加固防止内膜增生。 á代表性的组织切片用苏木精-伊红（HE）。律师表示染色弹力（VGEL）50微米。 公元前代表组织切片。 L =流明M =媒体，1H =内膜增生。律师表示沾上Masson染色50微米。D.代表组织切片。律师代表50微米。 <img alt="图4" fo:content-width="5in" src="/files/ftp_upload/52079/52079fig4highres.jpg" width="500" /> 图中的动脉瘤瘘4外部加固。 A.时间过程的照片静脉增强剂（从上到下）。动脉-静脉端-侧吻合术B.大视图。

Watch this Scientific Journal Video about Procedure for Human Saphenous Veins Ex Vivo Perfusion and External Reinforcement at JoVE.com

Procedure for Human Saphenous Veins Ex Vivo Perfusion and External Reinforcement

导致内膜增生（IH）和静脉移植物衰竭的发展机制仍然知之甚少。这项研究描述了一种体外系统灌注在控制流量和压力的人脉。另外外部钢筋网的效率限制IH的发展进行了评价。

步骤人类隐静脉<em&gt;前体内</em&gt;灌注和外部强化

The mainstay of contemporary therapies for extensive occlusive arterial disease is venous bypass graft. However, its durability is threatened by intimal ...

procedure-for-human-saphenous-veins-ex-vivo-perfusion-external

Nanyang Technological University

Research

JoVE Journal

Medicine

Procedure for Human Saphenous Veins Ex Vivo Perfusion and External Reinforcement

11.2K 次观看.  Brigham and Women's Hospital/Harvard Medical School. 导致内膜增生（IH）和静脉移植物衰竭的发展机制仍然知之甚少。这项研究描述了一种体外系统灌注在控制流量和压力的人脉。另外外部钢筋网的效率限制IH的发展进行了评价。 

视频: Procedure for Human Saphenous Veins Ex Vivo Perfusion and External Reinforcement

Tissue Engineering by Intrinsic Vascularization in an In Vivo Tissue Engineering Chamber

In reconstructive surgery, there is a clinical need for an alternative to the current methods of autologous reconstruction which are complex, costly and trade one defect for another. Tissue engineering holds the promise to address this increasing demand. However, most tissue engineering strategies fail to generate stable and functional tissue substitutes because of poor vascularization. This paper focuses on an in vivo tissue engineering chamber model of intrinsic vascularization where a perfused artery and a vein either as an arteriovenous loop or a flow-through pedicle configuration is directed inside a protected hollow chamber. In this chamber-based system angiogenic sprouting occurs from the arteriovenous vessels and this system attracts ischemic and inflammatory driven endogenous cell migration which gradually fills the chamber space with fibro-vascular tissue. Exogenous cell/matrix implantation at the time of chamber construction enhances cell survival and determines specificity of the engineered tissues which develop. Our studies have shown that this chamber model can successfully generate different tissues such as fat, cardiac muscle, liver and others. However, modifications and refinements are required to ensure target tissue formation is consistent and reproducible. This article describes a standardized protocol for the fabrication of two different vascularized tissue engineering chamber models in vivo.

Tissue Engineering by Intrinsic Vascularization in an In Vivo Tissue Engineering Chamber

This is a guideline for constructing in vivo vascularized tissue using a microsurgical arteriovenous loop or a flow-through pedicle configuration inside a tissue engineering chamber. The vascularized tissues generated can be employed for organ regeneration and replacement of tissue defects, as well as for drug testing and disease modeling.

在组织工程由内在血管化<em&gt;在体内</em&gt;组织工程商会

In reconstructive surgery, there is a clinical need for an alternative to the current methods of autologous reconstruction which are complex, costly and ...

科研

生物工程

The Arteriovenous (AV) Loop in a Small Animal Model to Study Angiogenesis and Vascularized Tissue Engineering

A functional blood vessel network is a prerequisite for the survival and growth of almost all tissues and organs in the human body. Moreover, in pathological situations such as cancer, vascularization plays a leading role in disease progression. Consequently, there is a strong need for a standardized and well-characterized in vivo model in order to elucidate the mechanisms of neovascularization and develop different vascularization approaches for tissue engineering and regenerative medicine.
We describe a microsurgical approach for a small animal model for induction of a vascular axis consisting of a vein and artery that are anastomosed to an arteriovenous (AV) loop. The AV loop is transferred to an enclosed implantation chamber to create an isolated microenvironment in vivo, which is connected to the living organism only by means of the vascular axis. Using 3D imaging (MRI, micro-CT) and immunohistology, the growing vasculature can be visualized over time. By implanting different cells, growth factors and matrices, their function in blood vessel network formation can be analyzed without any disturbing influences from the surroundings in a well controllable environment. 
In addition to angiogenesis and antiangiogenesis studies, the AV loop model is also perfectly suited for engineering vascularized tissues. After a certain prevascularization time, the generated tissues can be transplanted into the defect site and microsurgically connected to the local vessels, thereby ensuring immediate blood supply and integration of the engineered tissue. By varying the matrices, cells, growth factors and chamber architecture, it is possible to generate various tissues, which can then be tailored to the individual patient's needs.

The Arteriovenous AV Loop in a Small Animal Model to Study Angiogenesis and Vascularized Tissue Engineering

We describe a microsurgical approach for the generation of an arteriovenous (AV) loop as a model for analyzing vascularization in vivo in an isolated and well-characterized environment. This model is not only useful for the investigation of angiogenesis, but is also optimally suited for engineering axially vascularized and transplantable tissues.

在小型动物模型的动静脉（AV）循环研究血管生成和血管组织工程

A functional blood vessel network is a prerequisite for the survival and growth of almost all tissues and organs in the human body. Moreover, in ...

Decellularization and Recellularization Methodology for Human Saphenous Veins

Vascular conduits used during most vascular surgeries are allogeneic or synthetic grafts that often lead to complications caused by immunosuppression and poor patency. Tissue engineering offers a novel solution to generate personalized grafts with a natural extracellular matrix containing the recipient&#39;s cells using the method of decellularization and recellularization. We show a detailed method for performing decellularization of the human saphenous vein and recellularization by perfusion of peripheral blood. The vein was decellularized by perfusing 1% Triton X-100, 1% tri-n-butyl-phosphate (TnBP) and 2,000 Kunitz units of deoxyribonuclease (DNase). Triton X-100 and TnBP were perfused at 35 mL/min for 4 h while DNase was perfused at 10 mL/min at 37 &#176;C for 4 h. The vein was washed in ultrapure water and PBS and then sterilized in 0.1% peracetic acid. It was washed again in PBS and preconditioned in endothelial medium. The vein was connected to a bioreactor and perfused with endothelial medium containing 50 IU/mL heparin for 1 h. Recellularization was performed by filling the bioreactor with fresh blood, diluted 1:1 in Steen solution, and adding endocrine gland-derived vascular endothelial growth factors (80 ng/mL), basic fibroblast growth factors (4 &#181;L/mL), and acetyl salicylic acid (5 &#181;g/mL). The bioreactor was then moved into an incubator and perfused for 48 h at 2 mL/min while maintaining glucose between 3 - 9 mmol/L. Later, the vein was washed with PBS, filled with endothelial medium and perfused for 96 h in the incubator. Treatment with Triton X-100, TnBP and DNase decellularized the saphenous vein in 5 cycles. The decellularized vein looked white in contrast to normal and recellularized veins (light red). The hematoxylin &#38; eosin (H&#38;E) staining showed the presence of nuclei only in normal but not in decellularized veins. In the recellularized vein, H&#38;E-staining showed the presence of cells on the luminal surface of the vein.

Here, we describe a protocol for the saphenous vein decellularization using detergents and recellularization by perfusion of the peripheral blood and the endothelial medium.

人隐静脉的去细胞和 Recellularization 方法学研究

Vascular conduits used during most vascular surgeries are allogeneic or synthetic grafts that often lead to complications caused by immunosuppression and ...

Vein Interposition Model: A Suitable Model to Study Bypass Graft Patency

Bypass grafting is an established treatment method for coronary artery disease. Graft patency continues to be the Achilles heel of saphenous vein grafts. Research models for bypass graft failure are essential for a better understanding of pathobiological and pathophysiological processes during graft patency loss. Large animal models, such as pigs or sheep, resemble human anatomical structures but require special facilities and equipment. This video describes a rat vein interposition model to investigate vein graft patency loss. Rats are inexpensive and easy to handle. Compared to mouse models, the convenient size of rats permits better operability and enables a sufficient amount of material to be obtained for further diverse analysis. In brief, the inferior epigastric vein of a donor rat is harvested and used to replace a segment of the femoral artery. Anastomosis is conducted via single stitches and sealed with fibrin glue. Graft patency can be monitored non-invasively using duplex sonography. Myointimal hyperplasia, which is the main cause for graft patency loss, develops progressively over time and can be calculated from histological cross sections.

This video demonstrates a model to study the development of myointimal hyperplasia after venous interposition surgery in rats.

静脉介入型号:研究旁路移植通畅一个合适的模型

Bypass grafting is an established treatment method for coronary artery disease. Graft patency continues to be the Achilles heel of saphenous vein grafts. ...

医学

A Reliable Porcine Fascio-Cutaneous Flap Model for Vascularized Composite Allografts Bioengineering Studies

Vascularized Composite Allografts (VCA) such as hand, face, or penile transplant represents the cutting-edge treatment for devastating skin defects, failed by the first steps of the reconstructive ladder. Despite promising aesthetic and functional outcomes, the main limiting factor remains the need for a drastically applied lifelong immunosuppression and its well-known medical risks, preventing broader indications. Therefore, lifting the immune barrier in VCA is essential to tip the ethical scale and improve patients&#39; quality of life using the most advanced surgical techniques. De novo creation of a patient-specific graft is the upcoming breakthrough in reconstructive transplantation. Using tissue engineering techniques, VCAs can be freed of donor cells and customized for the recipient through perfusion-decellularization-recellularization. To develop these new technologies, a large-scale animal VCA model is necessary. Hence, swine fascio-cutaneous flaps, composed of skin, fat, fascia, and vessels, represent an ideal model for preliminary studies in VCA. Nevertheless, most VCA models described in the literature include muscle and bone. This work reports&#160;a reliable and reproducible technique for saphenous fascio-cutaneous flap harvest in swine, a practical tool for various research fields, especially vascularized composite tissue engineering.

The present protocol describes the porcine fascio-cutaneous flap model and its potential use in vascularized composite tissue research.

一种用于血管化复合同种异体移植物生物工程研究的可靠猪筋膜-皮肤皮瓣模型

Vascularized Composite Allografts (VCA) such as hand, face, or penile transplant represents the cutting-edge treatment for devastating skin defects, ...

Synergizing Antegrade Endoscopic with Bridging Vein Harvesting for Improvement of Great Saphenous Vein Graft Quality from the Lower Leg

Antegrade endoscopic harvesting of autografts for bypass grafting may be an optimal strategy addressing excellent graft quality and reduced post-operative wound complications. This standardized protocol for antegrade endoscopic vein harvesting (EVH) from the lower leg has the potential to be introduced to routine coronary artery bypass grafting (CABG). Patients undergoing CABG surgery are positioned on a surgical table with two additional foam rollers below the extended legs, enabling antegrade EVH from the lower leg. Following minimally invasive surgical access through a bridging vein harvest technique, an endoscopic optical dissector is inserted antegrade into the wound. The main vessel and side branches are dissected under continuous optical control of vein quality status and the working channel. After, an endoscopic optical retractor is inserted with an internal bipolar electrocoagulation device for precise, safe, and tissue-protective interruption of side branches. After release of the vein, the vessel is cut off at the proximal and distal ends under optical control, retrieved from the wound, then cannulated and flushed with heparinized saline. Finally, all side branches of the vein graft are double-clipped. Vascular histology is analyzed in a randomized selection of vein samples. After applying this standardized EVH protocol, the learning curve was shown to be steep, and graft quality was sufficient for coronary artery bypass grafting in every case. There was no conversion to surgical harvesting and low risks for tissue damage and bleeding. Leg positioning and synergizing EVH with bridging vein harvesting improved procedural success and vein graft quality. In our hands, antegrade EVH from the lower leg was feasible, demonstrating straightforward graft dissection as well as adequate macroscopic and microscopic graft quality with preserved endothelial integrity. In conclusion, the introduced technique is safe, shows excellent vein autograft quality, and illustrates feasibility for elective and urgent isolated CABG and combined CABG scenarios.

Presented here is a protocol for antegrade endoscopic vein harvesting from the lower leg, which can safely be introduced in routine coronary artery bypass grafting. Vein grafts present excellent graft quality following this standardized protocol with positioning of the legs, minimally invasive access to the vein, and antegrade endoscopic vein harvesting.

协同前体内窥镜与桥接静脉收获，提高从下腿大萨普努斯静脉移植质量

Antegrade endoscopic harvesting of autografts for bypass grafting may be an optimal strategy addressing excellent graft quality and reduced post-operative ...

A Training and Testing System for Performing Vascular Reconstruction In Vitro

Manual vascular reconstruction training is essential for a beginner surgeon. However, an optimal training system for vascular reconstruction in vitro has yet to be developed. In this study, we introduce an in vitro training and testing system using a magnetic anchoring technique with which a trainee can practice manual vascular reconstruction individually. Additionally, this system can also be used to test the quality of the reconstruction. The described system includes a vascular reconstruction training machine, magnetic tractors, and a magnetic suture puller. In this manuscript, we detail an end-to-end vein anastomosis using porcine right and left iliac veins. To identify the potential damage caused by a magnetic suture puller on the suture, we created three groups with six segments of 4-0 polypropylene sutures each: a control group with no intervention on the polypropylene suture, a group in which the polypropylene suture is manually pulled with sterile gloves 20x, and a magnetic puller group in which the magnetic puller pulled the polypropylene suture 20x. These groups were tested by light microscopy and breaking strength tests, and the effect of reconstruction was assessed. In the light microscopy test, the control group was less likely to be damaged (p &#60; 0.05) and the number of damaged points of the manual group and magnetic puller group were similar (p &#62; 0.05). The results of the breaking strength test were compared across groups and no significant difference was observed (p &#62; 0.05). The end-to-end anastomosis of the porcine iliac veins was successfully performed using this training system, and the reconstructed veins could undergo 2.0 kPa perfusion pressure. Using this training and testing system the trainee can practice manual vascular reconstruction in vitro individually with the aid of magnetic tractors and a magnetic suture puller, and the quality of the reconstruction can be tested.

Here we present a training and testing system where a trainee can complete manual vascular reconstruction in vitro individually using a magnetic anchoring technique. The system can also be used to test the quality of reconstruction.

在体外进行血管重建的训练和测试系统

Manual vascular reconstruction training is essential for a beginner surgeon. However, an optimal training system for vascular reconstruction in vitro has ...

A Rabbit Venous Interposition Model Mimicking Revascularization Surgery using Vein Grafts to Assess Intimal Hyperplasia under Arterial Blood Pressure

Although vein grafts have been commonly used as autologous grafts in revascularization surgeries for ischemic diseases, the long-term patency remains poor because of the acceleration of intimal hyperplasia due to the exposure to arterial blood pressure. The present protocol is designed for the establishment of experimental venous intimal hyperplasia by interposing rabbit jugular veins to the ipsilateral carotid arteries. The protocol does not require surgical procedures deep in the body trunk and the extent of the incision is limited, which is less invasive for the animals, allowing long-term observation after implantation. This simple procedure enables researchers to investigate strategies to attenuate the progression of intimal hyperplasia of the implanted vein grafts. Using this protocol, we reported the effects transduction of microRNA-145 (miR-145), which is known to control the phenotype of vascular smooth muscle cells (VSMCs) from the proliferative to the contractile state, into harvested vein grafts. We confirmed the attenuation of intimal hyperplasia of vein grafts by transducing miR-145 before implantation surgery through the phenotype change of the VSMCs. Here we report a less invasive experimental platform to investigate the strategies that can be used to attenuate intimal hyperplasia of vein grafts in revascularization surgeries.

The present protocol aims to experimentally create venous intimal hyperplasia by subjecting veins to arterial blood pressure for developing strategies to attenuate venous intimal hyperplasia following revascularization surgery using vein grafts.

兔子静脉介词模型模仿再血管化手术使用静脉移植评估动脉血压下的影响增生

Although vein grafts have been commonly used as autologous grafts in revascularization surgeries for ischemic diseases, the long-term patency remains poor ...

Occlusion of the Great and Small Saphenous Vein Using Copolymeric Glue Based on N-Butyl Cyanoacrylate and Methacryloxy Sulfolane

We present the preliminary results of a longitudinal observational study aimed at evaluating the effectiveness and safety at different short- and long-term follow-ups of vascular occlusion of the great saphenous vein (GSV) and small saphenous vein (SSV) affected by severe pathological reflux, using an innovative modified cyanoacrylate surgical glue composed of N-butyl cyanoacrylate and methacryloxy sulfolane (NBCA+MS). Ninety patients, prospectively recruited for 1 year, underwent the study with EcoColor-Doppler (ECD) to evaluate the maximum diameters of the GSV and SSV in the orthostatic position and the reflux time (RT). An RT greater than 0.5 s was considered pathologic. Clinical, etiology, anatomy, and pathophysiology (CEAP) assessment was used for the complete evaluation of each patient in the study. All the patients were treated by NBCA+MS glue to obtain vein occlusion and observed before treatment (baseline; T0), within 6 h after treatment (T1), 1 month after treatment (T2), 3 months after treatment (T3), 6 months after treatment (T4), and 1 year after treatment (T5). Chi-square (&#967;) analysis was performed to evaluate the effectiveness and safety of the treatment. All the patients participated in the entire duration of the study. Complete occlusion was maintained in 100% of patients at T1, 98.9% at T2 and T3, and 97.8% at T4 and T5 (p &lt; 0.001). None of the patients suffered from post-surgical thrombosis. No blue hyperpigmentation, or paresthesia was observed during the entire observation period. Immediately after treatment, 7.7% of patients needed painkillers; 1 week after treatment, 100% of patients returned to normal life. Vascular occlusion of the great or small saphenous vein using NBCA+MS glue is a safe procedure with persistent benefits after a 1 year follow-up. This procedure can be performed with local anesthesia, allowing a quick return to normal life. Thanks to its low invasiveness, the treatment is not painful.

Here, we present a protocol to treat the great saphenous vein (GSV) and the small saphenous vein (SSV) affected by severe pathological reflux, using an original sclerosing and embolizing cyanoacrylate-based glue composed of N-butyl cyanoacrylate and methacryloxy sulfolane (NBCA+MS).

使用基于氰基丙烯酸正丁酯和甲基丙烯酰氧基环丁基磺烷的共聚物胶咬合大隐脉和小隐脉

We present the preliminary results of a longitudinal observational study aimed at evaluating the effectiveness and safety at different short- and ...

Human Saphenous Vein Endothelial Cell Isolation and Exposure to Controlled Levels of Shear Stress and Stretch

Coronary artery bypass graft (CABG) surgery is a procedure to revascularize ischemic myocardium. Saphenous vein remains used as a CABG conduit despite the reduced long-term patency compared to arterial conduits. The abrupt increase of hemodynamic stress associated with the graft arterialization results in vascular damage, especially the endothelium, that may influence the low patency of the saphenous vein graft (SVG). Here, we describe the isolation, characterization, and expansion of human saphenous vein endothelial cells (hSVECs). Cells isolated by collagenase digestion display the typical cobblestone morphology and express endothelial cell markers CD31 and VE-cadherin. To assess the mechanical stress influence, protocols were used in this study to investigate the two main physical stimuli, shear stress and stretch, on arterialized SVGs. hSVECs are cultured in a parallel plate flow chamber to produce shear stress, showing alignment in the direction of the flow and increased expression of KLF2, KLF4, and NOS3. hSVECs can also be cultured in a silicon membrane that allows controlled cellular stretch mimicking venous (low) and arterial (high) stretch. Endothelial cells' F-actin pattern and nitric oxide (NO) secretion are modulated accordingly by the arterial stretch. In summary, we present a detailed method to isolate hSVECs to study the influence of hemodynamic mechanical stress on an endothelial phenotype.

We describe a protocol to isolate and culture human saphenous vein endothelial cells (hSVECs). We also provide detailed methods to produce shear stress and stretch to study mechanical stress in hSVECs.

人隐静脉内皮细胞分离和暴露于受控水平的剪切应力和拉伸

Coronary artery bypass graft (CABG) surgery is a procedure to revascularize ischemic myocardium. Saphenous vein remains used as a CABG conduit despite the ...

步骤人类隐静脉

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