The study was supported by research grants from the Roche Organ Transplant Research Foundation (ROTRF) and Astellas. Markus Selzner was supported by an ASTS Career Development Award. Matthias Knaak was supported by the Astellas Research Scholarship. We thank Uwe Mummenhoff and the Birmingham family for their generous support.

所有动物人文关怀符合&#39;"实验动物管理的原则&#39;为医学研究和"指南"对实验动物的关怀"制定的国家红会"由卫生部，安大略省，加拿大国家研究院​​公布。在多伦多综合研究所的动物护理委员会批准的所有研究。 1.器官检索<ol><li>移植前，30和35公斤的研究机构1周间宅男约克夏猪，以防止应激诱导的生理反应（这可以改变灌注的结果10,11），并使其恢复动物的住房条件。快猪最少6小时前麻醉诱导。 </li><li>麻醉供体猪通过肌肉注射（IM）注射氯胺酮（25毫克/公斤），阿托品（0.04毫克/千克），和咪唑安定（0.15毫克/千克）的混合物。 </li&gt; <li>前插管，确保猪自发呼吸2升氧气的剂量用异氟烷的5％。在仰卧位，喷声带用2％利多卡因2分钟插管前，以避免声带痉挛。对于35公斤猪，使用6.5神父气管套管。堵住气管插管用3 - 5毫升室内空气。 </li><li>气管插管后，使用capnometry，确认无误插管。在监测的猪尾巴的心脏速率和氧饱和度脉搏血氧仪。降低了异氟醚蒸发到2％。 <ol><li>调整麻醉用最小肺泡浓度（MAC）值的深度;目标2 - 2.5 MAC。呼吸机设置为14 - 16呼吸/分钟和10潮气量 - 为15ml / kg体重。 </li></ol></li><li>放置一个18G的静脉内（ⅳ）导管在耳静脉中的一个，以允许输注林格氏乳酸盐溶液（200毫升/小时）。擦洗猪，它覆盖用无菌窗帘。 </li><li>在确保无菌的条件下，使中线剖腹手术弗洛由左侧横向延伸结婚。用毛巾将其移动到左侧前覆盖大大小小的肠子。 </li><li>划分镰状韧带和使用烧灼三角韧带。 </li><li>释放从右侧使用电手术刀膜片肝脏;用剪刀的静脉和膈肌的上部。解剖肝下静脉向下到右侧肾上腺静脉和左侧肾静脉的分支。 </li><li>分离肝下静脉和彼此远端主动脉;结扎主动脉分支到脊柱;隔离和黏附的组织游离肾动脉。围绕每个肾动脉以2-0平局。 </li><li>颅至左肾静脉，解剖主动脉和肠系膜上动脉。围绕肠系膜动脉以2-0平局。 </li><li>打开腹膜颅肠系膜动脉后，认真遵循对腹腔干主动脉。尾部解剖腹腔干到门户已经在;围绕脾和胃左动脉，哪个分支关闭后方腹腔干。解剖腹腔干断门静脉。 </li><li>由胰腺和门静脉之间的腹膜切开松开门静脉。摘下领带静脉从胰腺门静脉引流。 </li><li>从肝十二指肠韧带分开胆管结扎后远端把它。 </li><li>结扎肝十二指肠韧带内的淋巴管，以防止淋巴漏。分之间的关​​系胃十二指肠动脉和胃右动脉。结扎静脉较小。 </li><li>解剖心脏和腹腔干之间的隔膜背后的主动脉。周围放置主动脉颅2-0领带腹腔干。 </li><li>切除胆囊和烧灼从胆囊床出血的任何。 </li><li>打开隔膜。施用肝素1000国际单位/公斤施主体重心内或iv以达到设置的异氟烷％至5％（&gt; 2.5，MAC）更深层次的麻醉。对于循环死亡（DCD）模型后供体，通过心脏内注射40 MVAL氯化钾3分钟肝素用药后诱发心脏骤停。设置心脏停搏作为暖缺血的起点。 </li><li>摘下领带绕肾，脾，肠系膜及胃左动脉先前设定的关系。摘下领带主动脉远端肾和髂动脉之间，并用导管插入器官冲洗管线主动脉。 </li><li>摘下领带门静脉近端为尽可能与其他器官平齐线导管插入它。 </li><li>闭围绕近端主动脉预先设定的领带后，经由主动脉（压力袋）和门静脉（重力驱动）使用双灌注冲洗肝脏用2L威斯康辛（UW）溶液冷学报。 </li><li>切除肝脏，留下所有剩余的船只长。离开周围的肝上静脉慷慨的膈肌边缘。将肝脏转化为冰无菌风琴袋。 </li><li>在回台的准备，CL安培使用Satinsky钳肝上静脉，并经由下腔静脉逆行冲洗肝脏第二次用约0.5升UW液直到门静脉流出是明确的。 </li><li>打结主动脉和腹腔干的所有的动脉分支。与剩余的0.5升UW液12执行动脉背表压力灌注。 </li><li>使用冲洗UW液胆管。 </li><li>修剪膈肌修补程序以一个体面的大小。关闭所有膈静脉 - 通常为3，每1左右，而1后 - 用单丝4-0针聚丙烯。 </li><li>关闭风琴袋和肝脏储存在冰。 </li></ol> 2.收件人肝切除术<ol><li>通过IM注射氯胺酮（25毫克/公斤），阿托品（0.04毫克/千克），和咪唑安定（0.15毫克/千克）的混合物麻醉收件人猪。 </li><li>发生在仰卧位猪到手术台上的加热垫的顶部。涵盖猪无线次热循环毯。 </li><li>前插管，确保猪自发呼吸2升氧气的剂量用异氟烷的5％。插管前喷声带用2％利多卡因2分钟，以避免声带痉挛。对于35公斤猪，使用6.5神父气管套管。堵住气管插管用3 - 5毫升室内空气。 </li><li>气管插管后，使用capnometry，确认无误插管。在监测的猪尾巴的心脏速率和氧饱和度脉搏血氧仪。地点和固定的温度探头插入猪的口鼻部。降低了异氟醚蒸发到2％（目标为2 - 2.5，MAC）。呼吸机设置为14 - 16呼吸/分钟和10潮气量 - 为15ml / kg体重。 </li><li>使用对眼睛的药膏，以防止干燥，而在麻醉下。 </li><li>使用Seldinger技术13插入鞘诱导（8.5星期五）插入左外颈静脉。后来换了积极的门户腔静脉，颈静脉旁路使用该导管。 </li><li>使用塞尔丁格techniquE要插入一个全肠外营养（TPN）导管（9.5 FR）到右颈外静脉。 </li><li>在无菌条件下，解剖右颈动脉和用于侵入动脉压监测插入聚丙烯导管（18 G）的。周围动脉以2-0的丝质领带，允许紧急结扎。 </li><li>调低异氟烷蒸发器，以1％（1.5 - 2，MAC），并添加异丙酚（5 - 8毫克/公斤/小时ⅳ）以维持麻醉深度。镇痛，使用柠檬酸芬太尼的连续静脉输注（最优选的，2微克/公斤/小时）或瑞芬太尼（第二选择，15微克/公斤/小时）。 </li><li>在切开皮肤，给予头孢呋辛1000毫克和500毫克甲硝唑四。设置使用乳酸林格氏液，5％葡萄糖150毫升/小时的输液泵。 </li><li>放置在热循环毯到头部和颈部。擦洗猪，它覆盖用无菌窗帘。 </li><li>在无菌条件下，使一个中线剖腹手术。插入腹部řetractor获得足够的访问右上腹。 </li><li>划分falciforme韧带和使用烧灼三角韧带。 </li><li>在多个步骤，分肝十二指肠韧带接近关系的肝脏。确定，划分，并标注肝动脉和胆管的分支。 </li><li>解剖肝动脉逆行到胃十二指肠动脉的分工。确保牛头犬夹在周围肝总动脉近端适合于胃十二指肠动脉后夹紧。 </li><li>无组织粘连门静脉。 </li><li>动员使用电手术刀右侧腹膜后腔静脉。用剪刀的静脉和膈肌的上部。解剖肝下静脉向下到右侧肾上腺静脉和左侧肾静脉的分支。 </li><li>暴露脾门处。大约一半沿脾的长度，仔细清除f显示脾动脉和静脉从坚持腹膜层。围绕这两个脾动脉和静脉4 2-0真丝领带。 </li><li>插入一个8.5护套神父与诱导在导管的尖端2个额外的小孔进入脾静脉，指着远端朝门静脉。远端固定导管插入其用2-0的关系之一，并关闭静脉近端插入另一2-0平手。保留其他2纽带开放。 </li><li>抽取血液从导管，用10ml生理盐水冲洗，并关闭导管的夹子。 </li><li>填充盐水成旁路由离心泵头，一个颈管（3/16"），并同时从门静脉分支（3/16流入"）和一个腔静脉分支（1/4"，鲁尔锁定在近端连接器开幕）。把一个管夹上腔静脉管的近端。 </li><li>连接在入口和旁通到两个护套导引导管（ 图1）的颈静脉开口和密封连接到ction用金属软管夹圈。将离心泵头到其的泵位置。 </li><li>给予氨甲环酸1000毫克和肝素3分钟静脉交叉阻断前万IU。降低异丙酚输注速率至2mg / kg /小时的无肝期的时间。调节异氟醚浓度的动脉压和猪的反应性。 </li><li>打开两个护套导引导管和交叉夹紧门静脉的夹子。确保血液通过旁路运行被动。 </li><li>启动离心泵以约1500转/分钟。继续交叉夹紧如果一个）的猪是cardiovascularly稳定和b）该旁路在约500毫升/分钟运行。如果猪不容忍交叉夹紧，替代量（晶体液或胶体液）和正性肌力药物（去甲肾上腺素小波丽）。 </li><li>交叉夹紧肝下腔静脉颅只是用一个德Bakey骨节夹肾静脉。公司施加压力到肝脏TISSUE挤出的残血的一部分。交叉夹紧肝上下腔静脉，包括使用Satinsky夹，而收回肝尾侧横膈边缘。 </li><li>直接切肝上腔静脉在其边境的肝脏组织。接下来，切靠近肝门门静脉。 </li><li>肝下静脉夹具的约4cm颅，切孔进入的腔静脉前壁。这个洞，定位旁路"静脉开口与鲁尔锁定前方面临的连接器。 </li><li>固定连接在肝下静脉用1-0真丝领带。然后，打开旁通的静脉分支的管夹。 </li><li>最后，打开肝下静脉钳，允许在除了现有的波尔图颈旁一腔静脉，颈静脉旁路。提高泵的转速，以约2500转/分钟，使得在颈静脉侧旁通流出是900之间 - 1100毫升/分。 </li><li>切除肝脏在其剩余Çonnection颅肝下旁路连接器，以确保安全旁路连接的纽带是不切。定位旁路管小心，以避免扭曲。给500毫克的甲泼尼龙初始化免疫抑制。 </li></ol> 3.血管重建<ol><li>使用4-0聚丙烯单丝缝合，关闭了接收方在肝上静脉的光圈所有3膈静脉口。在接收方一侧肝上下静脉的两个角缝双臂4-0单丝聚丙烯缝线内外。 </li><li>拆下制冰盒的供体器官袋。打开风琴袋，取出供肝，并将其放入腹腔。 </li><li>对于肝上下静脉的端至端吻合术，修剪供体肝上下腔静脉，以适应接收者侧。使用接收方侧角线圈的内侧针，做一个内外角缝在每个suprah的侧面的epatic捐助静脉。 </li><li>拍摄权缝线的两端在一起。近似​​受援国和捐助国静脉窦口都，再配合左缝线的两个结局。 </li><li>拍摄较短的一端，做接受者静脉外，内缝背墙旁边的领带。运行在后墙，地理位置优越，外翻静脉壁。 </li><li>添加2-3前壁与拆线，一旦右侧已经达到相同的缝合，再出手这个缝合的结局。在运行使用从左上角的剩余缝合前壁。配合使用的背面和正面的墙壁都缝线在一起。配合其他两个缝合结局在右上角。 </li><li>修整供体门静脉到适当长度后，以相同的方式执行一个端至端门静脉吻合术，使用6-0单丝聚丙烯缝线。与前壁整理前不久，插管肝下静脉的内腔与另一冲洗管和冲洗出UW液用1L的盐水通过肝下cavaportal静脉在RT。 </li><li>完成吻合和领带的缝合线，留下生长因子约为0.5公分。换个德Bakey-贝克钳到捐助肝下静脉。 </li><li>打开肝上静脉夹和检查出血。然后，通过打开门钳位reperfuse肝脏。 </li><li>用6-0单丝聚丙烯缝线止血为针。降低旁路泵的转速约1500转/分钟，并关闭门户鞘管导引导管的夹子。 </li><li>重新夹紧肝下腔静脉的接收者侧，把管钳旁路的腔静脉部分。停止离心泵。 </li><li>切静脉连接的纽带，并删除它。归还剩余血液从旁路猪通过颈静脉导管。 </li><li>关闭颈静脉导管的夹子，断开旁路。得到硫酸鱼精蛋白的100毫克拮抗肝素。特别照顾的圆周率在这些步骤G的血流动力学;使用儿茶酚胺的压力支持，并以碳酸氢钠为代谢性酸中毒。 </li><li>在上述方式再次进行肝下静脉的端至端吻合，采用单丝5-0缝线的聚丙烯。通过释放两个夹子Reperfuse肝下下静脉。 </li><li>修剪周围的供体的腹腔干的主动脉补丁。摘下领带收件人胃十二指肠动脉附近的肝总动脉。把牛头犬夹到肝总动脉近端到胃十二指肠动脉的交界处。修剪小动脉修补用剪刀波茨使用周围的交界处血管组织。 </li><li>冲洗供体肝动脉用10毫升肝素生理盐水，并把另一斗牛犬钳进一步远端，以避免血液回流。吻合动脉窦口端至端在运行中的降落伞技术中，使用6-0单丝聚丙烯缝线。首先OPE Reperfuse宁远端，然后近端斗牛犬夹。 </li><li>吻合胆管端至端与使用上述运行方法2 6-0单丝聚丙烯缝线。确保胆管周围组织的大部分被包括在内，因为猪胆管是非常脆弱的，容易流泪。 </li><li>检查止血后，从脾静脉移除鞘导引导管。关闭近端和远端与其余2的关系。 </li><li>用大小为1单丝可吸收缝合线，关闭腹壁。关闭或者皮肤与皮肤缝合或任何运行2-0缝合。 </li></ol> 4.术后阶段<ol><li>使用电热垫和热循环毯，以保持温暖猪。 </li><li>样品血气小时。通过增加输液速度调脱水。 </li><li>断奶麻醉。通风猪另外2小时。 </li><li>从左颈除去护套导引导管已经。按坚定了几分钟，以防止出血。 </li><li>隧道TPN导管皮下猪的脖子的一侧。用缝线2-0保护它。 </li><li>经过2小时取出动脉行，如果猪是不支持儿茶酚胺血流动力学稳定。确保穿刺侧不出血。否则，使周围的血管孔6-0荷包缝合，无需关闭动脉。关闭切口部位。 </li><li>停止通风，一旦猪能够自主呼吸。断开气管插管通气管道。反复检查猪呼吸充分。 </li><li>放置在俯卧的猪到配备有加热灯的单个动物笔。一旦拔管猪能够独立托起他的头。不要让动物无人值守，直到它已经恢复了意识足以维持胸骨斜卧。众议院分别猪整个手术后期间。 </li><li>提供足够四止痛药手术后（例如，丁丙诺啡0.01 - 0.05毫克/公斤，每6小时）。 </li><li>如果猪不喝水独立，足以替代第四卷</li><li>继续甲泼尼龙作为免疫抑制剂（250毫克手术后一天（POD）1日上午，然后125毫克每天早晨）。开始头孢2毫克/千克PO两次从POD2上一天。 </li><li>管理500毫克的灭滴灵和1000毫克的头孢唑啉的每日两次，以及20毫克的泮托拉唑每日直至POD3一次。 </li><li>密切监测猪。不要犹豫牺牲它，如果它显示的痛苦的迹象（例如，嗜睡，回输喝，持久性酸中毒或低血糖，或出血或腹膜炎体征）。对于安乐死，抽血深下异氟烷麻醉（5％，&gt; 2.5 MAC）通过切割肝上静脉的猪。 </li></ol>

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The authors have nothing to disclose.

实验猪OLTx是没有临床情况的重症监护资源研究设定一个具有挑战性的过程。可能的并发症包括血流动力学不稳定，出血，器官缺血，低体温，和代谢，以及呼吸，代偿失调。对于任何研究组中，外科技术5以及猪麻醉14,15的足够程序训练是强制性的，以实现具有代表性和可再现的结果。 许多技术微妙之处已经在文献中进行了描述，特别是关于血管重建阶段5。上述OLTx协议提供了一个静脉-替换模型类似于人OLTx所需的信息。所提供的结果证明可靠的动物生存和移植物复苏都HBD和DCD模式。该协议适用于移植再灌注EXPE用短期生存场景riments，例如，以及在长期生存车型如公差研究。 猪OLTx的一个很大的障碍就是静脉和门静脉交叉夹紧相对耐受性差。在无肝期内脏充血引起静脉高压和毛细血管损伤甚至器官再灌注后7可能导致重大肠缺血和血液动力学不稳定的不可逆性休克的地步。由于腔静脉完全嵌入在肝实质中，静脉保背驮式程序是不可行的。下腔静脉在静脉重建阶段完全闭塞损害猪的血流动力学稳定。虽然几个报告表明，猪OLTx可以小于25分钟16,17总静脉，门静脉闭塞期间完成，血管重建时的门-静脉-颈静脉旁路技术是更安全和更实际的选择7- 9,18。在日Ë作者的经验，一个被动的波尔图，颈静脉旁路不是最佳的在无肝期，保持猪血流动力学稳定。旁路模式，包括肝下静脉和门静脉活跃的减压，使得肝上下腔静脉和门静脉吻合甚至延长阻断时间的平静重建阶段，由于不可预见的并发症。相反，以前的报告7，脾切除术是不是强制性的，当门户旁路导管被删除。无论脾动脉和静脉被关闭大约一半沿脾的长度留一半近端充分灌注。像并发症出血或空气栓塞，由于旁路断线都是可以避免通过确保旁路小心放置和固定正常。 在长期的生存OLTx实验，胆管吻合术被认为是一个薄弱点，由于其并发症发生率高19。胆道组织是很脆弱的，NEEDS特殊照顾，当被处理。许多不同的吻合技术已经描述5,19。端至端吻合术在技术上是容易的，并以最少的并发症19相关联。连续缝合与非切割针包括胆管周围结缔组织的大段似乎要优于间断缝合。胆管下放置不必要的紧张，当单针间断缝合打结是。这可能会导致组织撕裂和连续胆汁泄漏。缝合材料 - 可吸收或不可吸收的 - 通常是不重要的，因为它的寿命有限，直到猪被终止。在过去的数个月的长期生存模型，可吸收缝合线 - 就像人类OLTx - 是可取的。 具体必须注意与术后随访。充足的营养和流体供应，有可靠的疼痛缓解协议，和一个适当的免疫抑制制度是强制性的。对于长期实验中，免疫抑制显得尤为重要。相对于其他哺乳类动物，猪表明OLTx 20,21后一个令人惊讶的低免疫排斥反应率。圆形细胞浸润是在移植后的第二周最大，减少自发即使没有免疫抑制。拒绝是很少死亡猪OLTx 22后的原因。然而，即使使用免疫抑制协议涉及这里提到给予类固醇四和钙调磷酸酶抑制剂口服，移植物排斥是由OLTx之后开始在约4天转氨酶轻度增加表示，并通过表观门户字段圆形细胞浸润证实。钙调磷酸酶抑制剂可以给任何婆23,24或25,26 IV;这两种方法都有缺点。即使口服应用助剂，实际量到达胃肠道仍然难以捉摸。另一方面，持续静脉输注猪的笔有积极的动物是困难的。因此，四应用必须进行以大丸剂，这导致在高药物浓度的峰值随着潜在的毒性作用。然而，应用这两种方法出现，以允许长期生存。 类似于临床上，手术后应激性溃疡的预防建议。手术后出血从消化性溃疡是一种常见的问题，并可能与一个肝功能损害27。经过这两个团OLTx消化道出血少数情况下，作者开始定期预防与泮托拉唑，并没有遇到任何胃肠道出血至今。 严格维持在无菌条件术，可比的条件在临床手术室，以及随之而来的预防性抗生素，降低感染并发症的风险。 总之，本品Provides实用信息在研究环境建立猪OLTx程序。足够的奉献精神，实践和团队合作是为了降低学习期间，产生可靠的结果，并降低成本，研究动物的数量重要。

原位肝移植（OLTx）是治疗终末期肝病或晚期肝癌的唯一的治疗选择。在过去的25年中，考生在轮候名单上的人数逐渐增加，现在远远超过可移植的数量。在大多数移植的地区，病人轮候肝脏移植20〜30％死亡，而不会收到器官移植或摘牌是由于疾病的进展。策略，以增加供体库，因此，可移植的数量，都迫切需要。扩展标准器官分配，长时间保存移植和诱导免疫耐受仍然代表主要的临床挑战1-3。因此，实验OLTx研究是枢转，以优化临床OLTx实践。 猪OLTx是类似于人类OLTx在许多方面，包括升行之有效的实验模型艾弗大小，解剖学，生理学4-6。因此，它已成为一个标准的实验方法在研究领域，如外科手术技术，生理学，免疫学，保存，和缺血再灌注损伤。许多接枝采购，收件人肝切除，和，特别是，血管重建，技术在文献5已经描述。合适的技术的选择，根据研究人员的偏好和技术能力而变化。 与此相反的人的情况下，在无肝期内脏拥塞表示在猪OLTx一个重要问题。随后肠缺血和充血性血管损伤可引起严重的血液动力学不稳定，危及猪存活，因此，在实验7-9中的成功。因此，足够的肠减压是强制性的，尤其是在缺乏技术精制而成的实验设置。 我们荷兰国际集团积极波尔图腔静脉，颈静脉分流术无肝期的持续时间是一个可靠的选择，以避免肠道堵塞。该系统可用于早期再灌注的实验以及长期生存的情景。下面的协议中包含的所有信息建立一个稳定和可重复的肝移植模型的猪，包括供肝采购，受体手术切除包括和终端到终端的血管重建技术，和术后护理。

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			<td style="width:257px;">Hospira Healthcare Corporation</td>
			<td align="right" style="width:201px;">37869</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Propofol 1000mg/100mL</td>
			<td style="width:257px;">Pharmascience Inc.</td>
			<td align="right" style="width:201px;">2244379</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Protamine Sulfate 50mg/5mL</td>
			<td style="width:257px;">Pharmaceutical Partners of Canada Inc.</td>
			<td align="right" style="width:201px;">2139537</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Saline 0.9%, 1L</td>
			<td style="width:257px;">Baxter Corporation</td>
			<td align="right" style="width:201px;">60208</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Sodium Bicarbonate 50 mEq/50mL</td>
			<td style="width:257px;">Hospira Healthcare Corporation</td>
			<td align="right" style="width:201px;">261998</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Solu-Medrol 500mg</td>
			<td style="width:257px;">Pfizer Canada Inc.</td>
			<td align="right" style="width:201px;">2367963</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Tranexamic Acid 1000mg/10mL</td>
			<td style="width:257px;">Pfizer Canada Inc.</td>
			<td align="right" style="width:201px;">2064413</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">University of Wisconsin Solution, SPS-1</td>
			<td style="width:257px;">Organ Recovery Systms</td>
			<td style="width:201px;">SPS-1</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Xylocaine Endotracheal 10mg/50mL</td>
			<td style="width:257px;">AstraZeneca</td>
			<td align="right" style="width:201px;">2003767</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;"></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Appose ULC 35W skin stapler</td>
			<td style="width:257px;">Covidien Canada</td>
			<td align="right" style="width:201px;">803712</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Maxon, 1</td>
			<td style="width:257px;">Covidien Canada</td>
			<td align="right" style="width:201px;">606173</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Sofsilk, 0</td>
			<td style="width:257px;">Covidien Canada</td>
			<td style="width:201px;">S606</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Sofsilk, 2-0</td>
			<td style="width:257px;">Covidien Canada</td>
			<td style="width:201px;">S405</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Sofsilk, 3-0</td>
			<td style="width:257px;">Covidien Canada</td>
			<td style="width:201px;">S404</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Surgipro II, 4-0</td>
			<td style="width:257px;">Covidien Canada</td>
			<td style="width:201px;">VP581X</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Surgipro II, 5-0</td>
			<td style="width:257px;">Covidien Canada</td>
			<td style="width:201px;">VP725X</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Surgipro II, 6-0</td>
			<td style="width:257px;">Covidien Canada</td>
			<td style="width:201px;">VP733X</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;"></td>
			<td style="width:257px;"></td>
			<td style="width:201px;"></td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Catheter i.v, 18 G</td>
			<td style="width:257px;">BD Canada</td>
			<td align="right" style="width:201px;">381147</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Cook TPN catheter, 9.5Fr</td>
			<td style="width:257px;">Cook Medical Company</td>
			<td style="width:201px;">C-TPNS-9.5-90</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">PSI Kit for sheath catheter, 8.5Fr</td>
			<td style="width:257px;">Arrow International</td>
			<td style="width:201px;">ASK-09803-UHN</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;"></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Infusion Pump Line</td>
			<td style="width:257px;">Smith Medical ASD Inc.</td>
			<td style="width:201px;">21-0442-25</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Liver Admin Set (flush line)</td>
			<td style="width:257px;">CardioMed Supplies Inc</td>
			<td align="right" style="width:201px;">17175</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Mallinckrodt, Tracheal Tube, 6.5mm</td>
			<td style="width:257px;">Covidien Canada</td>
			<td align="right" style="width:201px;">86449</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Med-Rx Suction Connecting Tube</td>
			<td style="width:257px;">Benlan Inc.</td>
			<td style="width:201px;">70-8120</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Organ Bag</td>
			<td style="width:257px;">CardioMed Supplies Inc</td>
			<td align="right" style="width:201px;">2990</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Suction Tip</td>
			<td style="width:257px;">Tyco Healthcare Group LP</td>
			<td align="right" style="width:201px;">8888501023</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Valleylab, Cautery Pencil</td>
			<td style="width:257px;">Covidien Canada</td>
			<td style="width:201px;">E2515H</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Valleylab, Patient Return Electrode</td>
			<td style="width:257px;">Covidien Canada</td>
			<td style="width:201px;">E7507</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;"></td>
			<td style="width:257px;"></td>
			<td style="width:201px;"></td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Bypass Connector 3/8&#8221; x 1/4&#8220;</td>
			<td style="width:257px;">Raumedic AG</td>
			<td style="width:201px;">955083-001</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Bypass Connector 3/8&#8221; x 3/8&#8221; Luer Lock</td>
			<td style="width:257px;">Raumedic AG</td>
			<td style="width:201px;">955163-001</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Bypass Connector Y 3/8&#8221; x 3/8&#8221; x 1/4&#8221;</td>
			<td style="width:257px;">Raumedic AG</td>
			<td style="width:201px;">961360-002</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Bypass Tubing 1/4&#8221; x 1/16&#8221;</td>
			<td style="width:257px;">Raumedic AG</td>
			<td style="width:201px;">039505-010</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Bypass Tubing 3/8&#8221; x 3/32&#8221;</td>
			<td style="width:257px;">Raumedic AG</td>
			<td style="width:201px;">039535-005</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Rotaflow Cenrtifugal Pump</td>
			<td style="width:257px;">Maquet-Dynamed</td>
			<td style="width:201px;">HC 2821</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Stainless Steel Hose Clamp Ring, 5mm</td>
			<td style="width:257px;">Oetiker</td>
			<td align="right" style="width:201px;">16700007</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;"></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Abdominal Retractor</td>
			<td style="width:257px;">Medite GmbH</td>
			<td style="width:201px;">N/A</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">De Bakey &#8211; Beck, Infrahepatic Cava Clamp</td>
			<td style="width:257px;">Aesculap Inc.</td>
			<td style="width:201px;">FB519R</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Diethrich, Atraumaitc Clamp (Portal Vein)</td>
			<td style="width:257px;">Aesculap Inc.</td>
			<td style="width:201px;">FB525R</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Gregory Bull Dog Clamp, curved</td>
			<td style="width:257px;">Aesculap Inc.</td>
			<td style="width:201px;">FB382R</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Gregory Bull Dog Clamp, straight</td>
			<td style="width:257px;">Aesculap Inc.</td>
			<td style="width:201px;">FB381R</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Potts &#8211; De Martel, Scissors</td>
			<td style="width:257px;">Aesculap Inc.</td>
			<td style="width:201px;">BC648R</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Satinsky, Suprahepatic Cava Clamp</td>
			<td style="width:257px;">Aesculap Inc.</td>
			<td style="width:201px;">FB605R</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:276px;">Symetrical Tubing Clamp</td>
			<td style="width:257px;">Codman Instruments</td>
			<td align="right" style="width:201px;">198010</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;"></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Anesthesia Machine, Optimax</td>
			<td style="width:257px;">Moduflex Anesthesia Equipment</td>
			<td style="width:201px;">SN5180</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Bypass Flow meter, HT 110</td>
			<td style="width:257px;">Transonic Systems Inc.</td>
			<td style="width:201px;">HT110B11106</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Flow meter probe, H6XL</td>
			<td style="width:257px;">Transonic Systems Inc.</td>
			<td style="width:201px;">H6Xl689</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Heat Therapy Pump, T/Pump</td>
			<td style="width:257px;">Gaymar Industries Inc</td>
			<td style="width:201px;">TP500-G89D19</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Infusion Pump 3000</td>
			<td style="width:257px;">SIMS Graseby LTD.</td>
			<td style="width:201px;">SN300050447</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Isoflurane Vapor 19.1</td>
			<td style="width:257px;">Draeger Medical Canada Inc.</td>
			<td style="width:201px;">N/A</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Monitor, Datex AS 3</td>
			<td style="width:257px;">Instrumentarium Corp./ Hitachi</td>
			<td style="width:201px;">D-VHC14-23-02</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Rotaflow Centrifugal Drive Unit</td>
			<td style="width:257px;">Marquet-Dynamed</td>
			<td align="right" style="width:201px;">952301</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Rotaflow Console</td>
			<td style="width:257px;">Marquet-Dynamed</td>
			<td align="right" style="width:201px;">706035</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Temperature Therapy Pad</td>
			<td style="width:257px;">Gaymar Industries Inc</td>
			<td style="width:201px;">TP26E</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Valleylab Force Tx</td>
			<td style="width:257px;">Valleylab Inc.</td>
			<td align="right" style="width:201px;">216151480</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Ventilator, AV 800</td>
			<td style="width:257px;">DRE Medical Equipment</td>
			<td style="width:201px;">40800AVV</td>
			<td style="width:240px;"></td>
		</tr>
		<tr height="19">
			<td height="19" style="height:19px;width:276px;">Warm Touch, Patient Warming System</td>
			<td style="width:257px;">Nellcor/ Covidien Canada</td>
			<td style="width:201px;">5015300A</td>
			<td style="width:240px;"></td>
		</tr>
	</tbody>
</table>

technique of porcine liver procurement and orthotopic transplantation using an active porto-caval shunt

The success of liver transplantation has resulted in a dramatic organ shortage. Each year, a considerable number of patients on the liver transplantation waiting list die without receiving an organ transplant or are delisted due to disease progression. Even after a successful transplantation, rejection and side effects of immunosuppression remain major concerns for graft survival and patient morbidity. 
Experimental animal research has been essential to the success of liver transplantation and still plays a pivotal role in the development of clinical transplantation practice. In particular, the porcine orthotopic liver transplantation model (OLTx) is optimal for clinically oriented research for its close resemblance to human size, anatomy, and physiology. 
Decompression of intestinal congestion during the anhepatic phase of porcine OLTx is important to guarantee reliable animal survival. The use of an active porto-caval-jugular shunt achieves excellent intestinal decompression. The system can be used for short-term as well as long-term survival experiments. The following protocol contains all technical information for a stable and reproducible liver transplantation model in pigs including post-operative animal care.

在第一个移植研究中，一个心脏跳动施主模型（HBD中，n = 5）用一个DCD模型（10）暴露于原位热缺血45分钟进行比较。在两组中，移植物被保存在冰上采购后10小时。 在HBD组，收件人猪的100％存活，直到第5天的后续移植后结束。在DCD组，只有50％的受体猪的存活由于凝结问题或代谢失调5天，作为减少术后肝功能的结果。 所有血液样品从中心静脉导管收集。离心后，将血清样品获得和分析对肝细胞损伤（天冬氨酸转氨酶，AST），胆道功能（总胆红素和碱性磷酸酶），和肝功能（INR）。的时间过程的每个标记的示于图2 - 5。 <p class="jove_content"&gt; AST水平在24小时后（1414±538 U / L的HBD组和2296±1313 U / L的DCD组，P = 0.13）达到高峰，5天后恢复到接近正常的值。同样，碱性磷酸酶值后36小时对DCD组（224±111 U / L）相比，HBD组（162±54 U / L，p值= 0.27）显着增加。而总胆红素是稳定的HBD组（≤10微摩尔遍及/ L）时，它逐渐在DCD组中直到第5天（23±31微摩尔/升，p值= 0.43）增加。在DCD组中的胆红素值大的标准偏差显示在这组奇特的异质性胆道损伤。 INR为肝功能的标志物显示出相似的AST值的趋势。值达到高峰，24小时，5天后恢复到接近正常的值。所述HBD组1.47±0.34比较给DCD组与一个峰值低的值（峰值1.70±0.36，p值= 0.32）。 图波尔图腔静脉，颈静脉分流1算法。旁路是充满了乳酸林格氏液。然后将腔静脉部分被夹紧用管夹，颈静脉和脾部分连接到预先设定的导管，旁通打开，并且离心泵门脉夹持后开始。肝切除后，旁路的腔静脉部分被插入并固定在肝下腔静脉残端，颅到肾静脉。管钳被释放，以允许静脉减压，除了门户减压。 <img alt="图2" src="/files/ftp_upload/52055/52055fig2.jpg" /> 图2.谷草转氨酶（AST）（HBD N = 5，DCD N = 10）。AST是肝细胞损伤的敏感指标。峰值船尾尔24小时是低级在HBD比对DCD组中，提示少肝再灌注损伤;较小的标准差表示HBD组中更均匀的效果。 <img alt="图3" src="/files/ftp_upload/52055/52055fig3.jpg" /> 图3.总胆红素（HBD每组5，DCD 10例），总胆红素，胆间隙和胆管完整性的标志物，显示了与HBD组在低于10微摩尔/ L的值的稳定而均匀的趋势。在DCD组中的胆红素曲线随时间逐渐增加，并且显示出高的标准偏差，在实验组的仅一部分暗示胆管损伤。 <img alt="图4" src="/files/ftp_upload/52055/52055fig4.jpg" /> 图4.碱性磷酸酶（HBD N = 5，DCD N = 10）。碱性磷酸酶我š胆管损伤的指标。对于HBD组的值是比该DCD组，这意味着较少胆管损伤的降低。 <img alt="图5" src="/files/ftp_upload/52055/52055fig5.jpg" /> 图5. INR（HBD N = 5，DCD N = 10）。高INR值表明减弱肝细胞功能，由于降低凝血因子的释放。在这两种HBD和DCD群体，INR值返回到正常值移植后第5天，这表明肝功能的恢复。对于HBD组值出现下降。

Watch this Scientific Journal Video about Technique of Porcine Liver Procurement and Orthotopic Transplantation using an Active Porto-Caval Shunt at JoVE.com

Technique of Porcine Liver Procurement and Orthotopic Transplantation using an Active Porto-Caval Shunt

Experimental animal research plays a pivotal role in the development of clinical transplantation practice. The porcine orthotopic liver transplantation model (OLTx) closely resembles human conditions and is frequently used in clinically oriented research. The following protocol contains all information for a reliable porcine OLTx model using an active porto-caval-jugular shunt.

使用Active波尔图腔静脉分流术猪肝脏采购和原位移植技术

The success of liver transplantation has resulted in a dramatic organ shortage. Each year, a considerable number of patients on the liver transplantation ...

technique-porcine-liver-procurement-orthotopic-transplantation-using

Nanyang Technological University

Research

JoVE Journal

Medicine

16.3K 次观看.  Toronto General Hospital. Experimental animal research plays a pivotal role in the development of clinical transplantation practice. The porcine orthotopic liver transplantation model (OLTx) closely resembles human conditions and is frequently used in clinically oriented research. The following protocol contains all information for a reliable porcine OLTx model using an active porto-caval-jugular shunt.

视频: Technique of Porcine Liver Procurement and Orthotopic Transplantation using an Active Porto-Caval Shunt

Orthotopic Small Bowel Transplantation in Rats

Small bowel transplantation has become an accepted clinical option for patients with short gut syndrome and failure of parenteral nutrition (irreversible intestinal failure). In specialized centers improved operative and managing strategies have led to excellent short- and intermediate term patient and graft survival while providing high quality of life 1,3. Unlike in the more common transplantation of other solid organs (i.e. heart, liver) many underlying mechanisms of graft function and immunologic alterations induced by intestinal transplantation are not entirely known6,7. Episodes of acute rejection, sepsis and chronic graft failure are the main obstacles still contributing to less favorable long term outcome and hindering a more widespread employment of the procedure despite a growing number of patients on home parenteral nutrition who would potentially benefit from such a transplant. The small intestine contains a large number of passenger leucocytes commonly referred to as part of the gut associated lymphoid system (GALT) this being part of the reason for the high immunogenity of the intestinal graft. The presence and close proximity of many commensals and pathogens in the gut explains the severity of sepsis episodes once graft mucosal integrity is compromised (for example by rejection). To advance the field of intestinal- and multiorgan transplantation more data generated from reliable and feasible animal models is needed. The model provided herein combines both reliability and feasibility once established in a standardized manner and can provide valuable insight in the underlying complex molecular, cellular and functional mechanisms that are triggered by intestinal transplantation. We have successfully used and refined the described procedure over more than 5 years in our laboratory 8-11. The JoVE video-based format is especially useful to demonstrate the complex procedure and avoid initial pitfalls for groups planning to establish an orthotopic rodent model investigating intestinal transplantation.

Small bowel transplantation has become an accepted treatment option for patients with irreversible intestinal failure. Our experimental model of orthotopic small bowel transplantation in rats serves as a reliable tool to address underlying immunologic and inflammatory processes that complicate intestinal transplantation.

大鼠原位小肠移植

Small  bowel transplantation has become an accepted clinical option for patients with  short gut syndrome and failure of parenteral nutrition ...

科研

医学

Orthotopic Liver Transplantation in Rats

Clinical progress in the field of liver transplantation has been largely supported by animal models1,2. Since the publication of the first orthotopic rat liver transplantation in 1979 by Kamada et al.3, this model has remained the gold standard despite various proposed alternative techniques4. Nevertheless, its broader use is limited by its steep learning curve5.
In this video paper, we show a simple and easy-to-establish revision of Kamada's two-cuff technique. The suprahepatic vena cava anastomosis is performed manually with a running suture, and the vena porta and infrahepatic vena cava anastomoses are performed utilizing a quick-linker cuff system6. Manufacturing the quick-linker kit is shown in a separate video paper.

We present an easy-to-establish revision of the classical two-cuff technique for orthotopic liver transplantation in rat.

大鼠原位肝移植

Clinical progress in the field of liver transplantation has been largely supported by animal models1,2. Since the publication of the first orthotopic rat ...

Surgical Procedures for a Rat Model of Partial Orthotopic Liver Transplantation with Hepatic Arterial Reconstruction

Orthotopic liver transplantation (OLT) in rats using a whole or partial graft is an indispensable experimental model for transplantation research, such as studies on graft preservation and ischemia-reperfusion injury 1,2, immunological responses 3,4, hemodynamics 5,6, and small-for-size syndrome 7. The rat OLT is among the most difficult animal models in experimental surgery and demands advanced microsurgical skills that take a long time to learn. Consequently, the use of this model has been limited. Since the reliability and reproducibility of results are key components of the experiments in which such complex animal models are used, it is essential for surgeons who are involved in rat OLT to be trained in well-standardized and sophisticated procedures for this model.
While various techniques and modifications of OLT in rats have been reported 8 since the first model was described by Lee et al. 9 in 1973, the elimination of the hepatic arterial reconstruction 10 and the introduction of the cuff anastomosis technique by Kamada et al. 11 were a major advancement in this model, because they simplified the reconstruction procedures to a great degree. In the model by Kamada et al., the hepatic rearterialization was also eliminated. Since rats could survive without hepatic arterial flow after liver transplantation, there was considerable controversy over the value of hepatic arterialization. However, the physiological superiority of the arterialized model has been increasingly acknowledged, especially in terms of preserving the bile duct system 8,12 and the liver integrity 8,13,14.
In this article, we present detailed surgical procedures for a rat model of OLT with hepatic arterial reconstruction using a 50% partial graft after ex vivo liver resection. The reconstruction procedures for each vessel and the bile duct are performed by the following methods: a 7-0 polypropylene continuous suture for the supra- and infrahepatic vena cava; a cuff technique for the portal vein; and a stent technique for the hepatic artery and the bile duct.

Orthotopic liver transplantation in rats is an indispensable experimental model for biomedical research. Here we present our surgical procedures for orthotopic rat liver transplantation with hepatic arterial reconstruction using a 50% partial graft.

部分原位肝移植大鼠模型的肝动脉重建的外科手术

Orthotopic liver transplantation (OLT) in rats using a whole or partial graft is an indispensable experimental model for transplantation research, such as ...

Using Quantitative Real-time PCR to Determine Donor Cell Engraftment in a Competitive Murine Bone Marrow Transplantation Model

Murine bone marrow transplantation models provide an important tool in measuring hematopoietic stem cell (HSC) functions and determining genes/molecules that regulate HSCs. In these transplant model systems, the function of HSCs is determined by the ability of these cells to engraft and reconstitute lethally irradiated recipient mice. Commonly, the donor cell contribution/engraftment is measured by antibodies to donor- specific cell surface proteins using flow cytometry. However, this method heavily depends on the specificity and the ability of the cell surface marker to differentiate donor-derived cells from recipient-originated cells, which may not be available for all mouse strains. Considering the various backgrounds of genetically modified mouse strains in the market, this cell surface/ flow cytometry-based method has significant limitations especially in mouse strains that lack well-defined surface markers to separate donor cells from congenic recipient cells. Here, we reported a PCR-based technique to determine donor cell engraftment/contribution in transplant recipient mice. We transplanted male donor bone marrow HSCs to lethally irradiated congenic female mice. Peripheral blood samples were collected at different time points post transplantation. Bone marrow samples were obtained at the end of the experiments. Genomic DNA was isolated and the Y chromosome specific gene, Zfy1, was amplified using quantitative Real time PCR. The engraftment of male donor-derived cells in the female recipient mice was calculated against standard curve with known percentage of male vs. female DNAs. Bcl2 was used as a reference gene to normalize the total DNA amount. Our data suggested that this approach reliably determines donor cell engraftment and provides a useful, yet simple method in measuring hematopoietic cell reconstitution in murine bone marrow transplantation models. Our method can be routinely performed in most laboratories because no costly equipment such as flow cytometry is required.

Determining donor cell engraftment presents a challenge in mouse bone marrow transplant models that lack well-defined phenotypical markers. We described a methodology to quantify male donor cell engraftment in female transplant recipient mice. This method can be used in all mouse strains for the study of HSC functions.

采用实时定量PCR检测供体细胞植入一个具有竞争力的小鼠骨髓移植模型

Murine bone marrow transplantation models provide an important tool in measuring hematopoietic stem cell (HSC) functions and determining genes/molecules ...

Orthotopic Hind Limb Transplantation in the Mouse

In vivo animal model systems, and in particular mouse models, have evolved into powerful and versatile scientific tools indispensable to basic and translational research in the field of transplantation medicine. A vast array of reagents is available exclusively in this setting, including mono- and polyclonal antibodies for both diagnostic and interventional applications. In addition, a vast number of genotyped, inbred, transgenic, and knock out strains allow detailed investigation of the individual contributions of humoral and cellular components to the complex interplay of an immune response and make the mouse the gold standard for immunological research. 
Vascularized Composite Allotransplantation (VCA) delineates a novel field of transplantation using allografts to replace "like with like" in patients suffering traumatic or congenital tissue loss. This surgical methodological protocol shows the use of a non-suture cuff technique for super-microvascular anastomosis in an orthotopic mouse hind limb transplantation model. The model specifically allows for comparison between established paradigms in solid organ transplantation with a novel form of transplants consisting of various different tissue components. Uniquely, this model allows for the transplantation of a viable vascularized bone marrow compartment and niche that have the potential to exert a beneficial effect on the balance of immune acceptance and rejection. This technique provides a tool to investigate alloantigen recognition and allograft rejection and acceptance, as well as enables the pursuit of functional nerve regeneration studies to further advance this novel field of transplantation.

This novel model for orthotopic hind limb transplantation in the mouse, applying a non-suture cuff technique for super-microvascular anastomosis, provides a powerful tool for in&#160;vivo mechanistic immunological research related to vascularized composite allotransplantation (VCA).

原位后肢移植中的鼠标

In vivo animal model systems, and in particular mouse models, have evolved into powerful and versatile scientific tools indispensable to basic and ...

Using the Sleeve Technique in a Mouse Model of Aortic Transplantation - An Instructional Video

Orthotopic aortic transplantation using the sleeve technique reduces injury to the aorta with failure rate of only 10-20%. The time to anastomose the aorta in mice using the sleeve method was short and easy averaging 20 min, permitting studies of iso/allo grafts. The following article describes the aortic transplantation procedure used in our laboratory. The mice were anesthetized with a mixture of 1.5% volume isoflurane and 100% oxygen through a face mask. At this point, the segment of the aorta between the renal arteries and its bifurcation was separated from the vena cava, freely prepared and clampedat the proximal and distal segments with a single silk suture. Prior to the removal of the aorta, a saline solution containing heparin was injected into the inferior vena cava. Then the aorta was cut between the clamps and a saline heparin solution was used to flush the lumen. The sleeve technique with monofilament sutures was used in order to transplant the abdominal aorta in the orthotopic position.

We present an orthotopic aortic transplantation model using the sleeve technique in mice. It is a very rapid anastomosis method, which can be employed in studies of vascular disease.

套筒技术在主动脉移植小鼠模型中的应用-教学视频

Orthotopic aortic transplantation using the sleeve technique reduces injury to the aorta with failure rate of only 10-20%. The time to anastomose the ...

A Rat Model of Orthotopic Liver Transplantation Using a Novel Magnetic Anastomosis Technique for Suprahepatic Vena Cava Reconstruction

The rat model of orthotopic liver transplantation (OLT) is essential for transplant research. It is a very sophisticated animal model and requires a steep learning curve. The introduction of the cuff technique for anastomosis of the portal vein (PV) and infrahepatic vena cava (IHVC) has significantly simplified the transplant procedure in rats. However, due to the short anterior wall of the recipients' suprahepatic vena cava (SHVC), the cuff technique is very difficult to use for the reconstruction of the SHVC. Most researchers in this field still use the hand-suture technique for SHVC reconstruction, which makes it the bottleneck step in rat orthotopic liver transplantation. The magnetic anastomosis technique (i.e., magnamosis) is a method of connecting two vessels using the attractive force between two magnets. Our recent study has shown that the magnetic anastomosis technique is superior to the hand-suture technique for SHVC reconstruction in rats. In this article, we show a step-by-step protocol for SHVC reconstruction in rats using the novel magnetic anastomosis technique. In this model, the reconstruction of the PV and IHVC was performed by the standard cuff technique, while the reconstruction of the bile duct (BD) was performed by a stent technique. The hepatic re-arterialization was not performed. The magnetic anastomosis technique made SHVC reconstruction much easier and significantly shortened the anphepatic phase. After a reasonable learning curve, even researchers without advanced microsurgical skills can produce reliable and reproducible results using this rat model of OLT.

The reconstruction of the suprahepatic vena cava (SHVC) remains a difficult step in rat orthotopic liver transplantation. In this article, we show a step-by-step protocol for SHVC reconstruction in rats using a novel magnetic anastomosis technique.

应用新型磁吻合技术 Suprahepatic 腔静脉重建大鼠原位肝移植模型的研究

The rat model of orthotopic liver transplantation (OLT) is essential for transplant research. It is a very sophisticated animal model and requires a steep ...

A Pre-Clinical Porcine Model of Orthotopic Heart Transplantation

Fifty-years following the first successful report, cardiac transplantation remains the gold-standard treatment for eligible patients with advanced heart failure. Multiple small-animal models of heart transplantation have been used to study the acute and long-term effects of novel therapies. However, few are tested and demonstrated success in clinical trials. It is of critical importance to evaluate new therapies in a clinically relevant large-animal model for efficient and reliable translation of basic studies&#39;&#160;findings. Here, we describe a pre-clinical large-animal (porcine) model of orthotopic heart transplantation that has been firmly established and previously used to investigate novel cardioprotective strategies. This procedure focuses on acute ischemia-reperfusion injury and is a reliable method to investigate novel interventions which have been tested and validated in smaller experimental models, such as the murine model. We demonstrate its usefulness in assessing cardiac performance during the early post-transplantation period and other potential possibilities enabled by the model.

Here, we describe a pre-clinical large-animal (porcine) model of orthotopic heart transplantation that has been firmly established and utilized to investigate novel cardioprotective strategies.

正交心脏移植的临床前猪模型

Fifty-years following the first successful report, cardiac transplantation remains the gold-standard treatment for eligible patients with advanced heart ...

Reduced Complications after Arterial Reconnection in a Rat Model of Orthotopic Liver Transplantation

The rat orthotopic liver transplantation (OLT) model is a powerful tool to study acute and chronic rejection. However, it is not a complete representation of human liver transplantation due to the absence of arterial reconnection. Described here is a modified transplantation procedure that includes the incorporation of hepatic artery (HA) reconnection, leading to a marked improvement in transplant outcomes. With a mean anhepatic time of 12 min and 14 s, HA reconnection results in improved perfusion of the transplanted liver and an increase in long-term recipient survival from 37.5% to 88.2%. This protocol includes the use of 3D-printed cuffs and holders to connect the portal vein and infrahepatic inferior vena cava. It can be implemented for studying multiple aspects of liver transplantation, from immune response and infection to technical aspects of the procedure. By incorporating a simple and practical method for arterial reconnection using a microvascular technique, this modified rat OLT protocol closely mimics aspects of human liver transplantation and will serve as a valuable and clinically relevant research model.

The goal of this study is to modify the rat orthotopic liver transplant model to better represent human liver transplantation and improve recipient survival. The presented method reestablishes hepatic arterial inflow by connecting the donor liver&#39;s common hepatic artery to the recipient liver&#39;s proper hepatic artery.

矫形肝移植大鼠模型中动脉重联后并发症减少

The rat orthotopic liver transplantation (OLT) model is a powerful tool to study acute and chronic rejection. However, it is not a complete representation ...

Orthotopic Kidney Auto-Transplantation in a Porcine Model Using 24 Hours Organ Preservation And Continuous Telemetry

In the present era of organ transplantation with critical organ shortage, various strategies are employed to expand the pool of available allografts for kidney transplantation (KT). Even though, the use of allografts from extended criteria donors (ECD) could partially ease the shortage of organ donors, ECD organs carry a potentially higher risk for inferior outcomes and postoperative complications. Dynamic organ preservation techniques, modulation of ischemia-reperfusion and preservation injury, and allograft therapies are in the spotlight of scientific interest in an effort to improve allograft utilization and patient outcomes in KT.
Preclinical animal experiments are playing an essential role in translational research, especially in the medical device and drug development. The major advantage of the porcine orthotopic auto-transplantation model over ex vivo or small animal studies lies within the surgical-anatomical and physiological similarities to the clinical setting. This allows the investigation of new therapeutic methods and techniques and ensures a facilitated clinical translation of the findings. This protocol provides a comprehensive and problem-oriented description of the porcine orthotopic kidney auto-transplantation model, using a preservation time of 24 hours and telemetry monitoring. The combination of sophisticated surgical techniques with highly standardized and state-of-the-art methods of anesthesia, animal housing, perioperative follow up, and monitoring ensure the reproducibility and success of this model.

Large animal models play an essential role in preclinical transplantation research. Due to its similarities to the clinical setup, the porcine model of orthotopic kidney auto-transplantation described in this article provides an excellent in vivo setting for the testing of organ preservation techniques and therapeutic interventions.

使用24小时器官保存和连续遥测的猪模型中正交肾自动移植

In the present era of organ transplantation with critical organ shortage, various strategies are employed to expand the pool of available allografts for ...