Name: a rabbit model of durable transgene expression in jugular vein to common carotid artery interposition grafts
Uploaded: 2018-09-10T12:30:00.000+00:00
Duration: 13 min 58 s
Description: Watch this Scientific Journal Video about A Rabbit Model of Durable Transgene Expression in Jugular Vein to Common Carotid Artery Interposition Grafts at JoVE.com

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F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Understanding+saphenous+vein+graft+patency.">Understanding saphenous vein graft patency.</a> Circulation. 124 (3), 273-275 (2011).</li><li>Owens, C. D., Ho, K. J., Conte, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lower+extremity+vein+graft+failure:+a+translational+approach.">Lower extremity vein graft failure: a translational approach.</a> Vasc Med. 13, 63-74 (2008).</li><li>Robertson, K. E., McDonald, R. A., Oldroyd, K. G., Nicklin, S. A., Baker, A. H. <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+coronary+in-stent+restenosis+and+vein+graft+failure:+does+vascular+gene+therapy+have+a+role.">Prevention of coronary in-stent restenosis and vein graft failure: does vascular gene therapy have a role.</a> Pharmacol Ther. 136, 23-34 (2012).</li><li>Yla-Herttuala, S., Baker, A. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cardiovascular+Gene+Therapy:+Past,+Present,+and+Future.">Cardiovascular Gene Therapy: Past, Present, and Future.</a> Mol Ther. 25, 1095-1106 (2017).</li><li>Schwartz, L. 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本议定书的关键步骤包括麻醉的管理、抗凝、动脉/接枝静脉的手术操作以及接枝静脉的血流动力学测量。正确的麻醉管理在这一多项生存手术模型中是至关重要的, 包括两个相对较长的操作 (通常 3-3, 5 小时用于双侧静脉移植和 1.5-2.5 h 用于双边移植转导)。我们通过 nosecone 和气管插管进行麻醉, 发现插管改善了移植转导手术后的生存, 可能是因为正压通气预防肺不张和相关呼吸衰竭。家兔对插管有?...

动脉粥样硬化是一种慢性炎症性疾病, 其中脂质积累和炎症在血管壁导致血管腔狭窄, 心脏病发作, 中风, 四肢失去1,2。经皮介入治疗 (e., 血管成形术和支架植入术) 和医疗疗法 (g., 他汀类药物和抗血小板药物) 是治疗动脉粥样硬化的有效方法;然而, 在冠状动脉和外周循环中, 它们往往不能有效地治疗严重的梗阻性疾病。旁路移植术, 使用自体静脉段, 仍然是一个常见的程序治疗严重, 弥漫性冠状动脉和周围血管疾病3,4。然而, 在冠状动脉和外周循环中放置的静脉移植有很低的长期通畅率。在冠状动脉循环中, 大约10-20% 的静脉移植被闭塞1年, 50% 被闭塞10年5,6。在外周循环中, 静脉移植失败率为 30-50%, 5 年7。
基因治疗是一种很有吸引力的方法, 以防止静脉移植失败, 因为它可以提供一个治疗基因产品精确地在该疾病的地方。因此, 许多临床前研究已经测试了静脉移植基因治疗8,9。然而, 基本上所有这些研究已经检查的功效在早期点 (2-12 周)10,11,12,13,14,15,16,17. 我们知道没有证据表明基因治疗干预能提供持久 (多年) 保护, 防止晚期静脉移植失败, 通常是由新生内膜增生和动脉粥样硬化4引起的。我们开发了一种方法, 允许持久的转基因表达在嫁接静脉, 从而允许测试的基因治疗干预的后期以及早期的时间点。为实现持久的转基因表达, 该方法纳入 HDAd 载体和延迟转导策略。HDAd 载体提供长期的转基因表达, 因为他们缺乏病毒基因, 防止转基因细胞的识别 (和排斥) 的免疫系统18,19,20, 21. 延迟转导 (移植术后28天进行) 可防止移植22后早期发生的动脉过程中转基因细胞的丢失。
在静脉移植术中实现转基因表达的其他方法依赖于移植术10、11、12、15、16 时静脉移植的转导. ,17。在连续测量时, 使用这种方法的转基因表达在转导22、23后迅速下降。因此, 用这种方法进行的研究并没有对静脉移植后12周后的疗效进行检查, 最不评估疗效超过4周。相比之下, 我们的方法实现了静脉移植基因的表达, 稳定持续至少24周, 并根据类似的研究在动脉中进行-可能继续远长于22,24。我们知道没有其他静脉移植基因治疗干预, 达到稳定的转基因表达这一持续时间。
我们用兔子模型来发展我们的方法。另一些则使用啮齿动物、兔子或较大的动物来测试静脉移植基因治疗10、11、12、15、16、17、25、 26。与啮齿类动物模型相比, 兔子更昂贵, 并受到更严格的监管要求。然而, 与较大的动物相比(例如, 猪和狗), 兔子购买和房子的成本要低得多, 而且处理起来也更容易。此外, 兔血管与人体血管相似, 在生理上27, 它们足够大, 可以用来测试经皮介入治疗28,29, 他们提供了足够的组织,多个端点 (e., 组织学, 蛋白质, RNA) 可以使用单血管标本22,30检查。此外, 当带静脉移植的兔以高脂肪饮食喂养时, 他们发展静脉移植动脉粥样硬化31,32, 这是冠状动脉旁路移植失败的常见原因4,5.这些动脉粥样硬化兔静脉移植可以作为一个基质, 以测试的基因治疗干预提供这种方法。所提供的协议可以帮助研究者掌握在兔静脉移植中实现持久转基因表达所需的技术技能。

<table><tbody><tr><td>&lt;strong&gt;Disposables&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>3mL syringe with 24G needle</td><td>Becton Dickinson</td><td>309571</td><td>2x for vein graft surgery; 2x for gene transfer&amp;nbsp; surgery</td></tr><tr><td>1 mL syringe with 27G needle</td><td>Becton Dickinson</td><td>309623</td><td>2x for vein graft surgery, 5x for gene transfer surgery</td></tr><tr><td>19G needle</td><td>Becton Dickinson</td><td>305187</td><td>Gene transfer surgery</td></tr><tr><td></td><td></td><td></td><td></td></tr><tr><td>20 mL syringe, luer lock</td><td>Nipro Medical Corp</td><td>JD+20L</td><td></td></tr><tr><td>Catheters, 24Ga x 3/4&amp;rdquo;</td><td>Terumo Medical Products</td><td>SROX2419V</td><td></td></tr><tr><td>21G needle</td><td>Becton Dickinson</td><td>305165</td><td>Gene transfer surgery and for 20 mL syringe of saline</td></tr><tr><td>Gauze 4&amp;rdquo; x 4&amp;rdquo;</td><td>Dynarex</td><td>3242</td><td>~10-15 per surgery</td></tr><tr><td>3-0 silk suture</td><td>Covidien Ltd.</td><td>S-244</td><td></td></tr><tr><td>5-0 silk suture</td><td>Covidien Ltd.</td><td>S-182</td><td></td></tr><tr><td>7-0 polypropylene suture</td><td>CP Medical</td><td>8703P</td><td>Vein graft surgery</td></tr><tr><td>7-0 polypropylene suture</td><td>CP Medical</td><td>8648P</td><td>Gene transfer surgery</td></tr><tr><td>5-0 polyglycolic acid suture</td><td>CP Medical</td><td>421A</td><td></td></tr><tr><td>3-0 polyglycolic acid suture</td><td>CP Medical</td><td>398A</td><td></td></tr><tr><td>Alcohol swabs</td><td>Covidien Ltd.</td><td>6818</td><td>For the placement of I.V. line</td></tr><tr><td>Catheter plug</td><td>Vetoquinol</td><td>411498</td><td></td></tr><tr><td>Ketamine HCl, 100 mg/mL</td><td>Vedco Inc.</td><td>5098916106</td><td></td></tr><tr><td>Xylazine, 100 mg/mL</td><td>Akorn Inc.</td><td>4821</td><td></td></tr><tr><td>Lidocaine, 20 mg/mL</td><td>Pfizer</td><td>409427702</td><td></td></tr><tr><td>Marcaine 0.5%</td><td>Pfizer</td><td>409161050</td><td></td></tr><tr><td>Beuthanasia D-Special</td><td>Intervet Inc.</td><td>NDC 00061047305</td><td>Harvest surgery only</td></tr><tr><td>Buprenorphine HCl, 0.3 mg/mL</td><td>Patterson Veterinary</td><td>12496075705</td><td></td></tr><tr><td>Saline IV bag, 0.9% sodium chloride</td><td>Baxter</td><td>2B1309</td><td></td></tr><tr><td>Heparin&amp;nbsp; (5000 U/mL)</td><td>APP Pharmaceuticals</td><td>NDC 63323-047-10</td><td></td></tr><tr><td>Papaverine (3.5 mg/ml)</td><td>American Reagent Inc.</td><td>NDC 0517-4002-25</td><td>Diluted from 30mg/mL stock; Use 1 mL maximum</td></tr><tr><td>Fentanyl patch, 25 mcg/h</td><td>Apotex Corp.</td><td>NDC 60505-7006-2</td><td></td></tr><tr><td>Isoflurane</td><td>Multiple vendors</td><td>Catalog number not available</td><td></td></tr><tr><td>&amp;nbsp;Viral vector</td><td></td><td></td><td>Gene transfer surgery only</td></tr><tr><td>&lt;strong&gt;Surgical Instruments&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Metzenbaum needle holder 7&quot; straight</td><td>Roboz</td><td>RS-7900</td><td></td></tr><tr><td>Operating scissors 6.5&quot; straight blunt/blunt</td><td>Roboz</td><td>RS-6828</td><td></td></tr><tr><td>Needle holder /w suture scissors</td><td>Miltex</td><td>8-14-IMC</td><td></td></tr><tr><td>Castroviejo scissors</td><td>Roboz</td><td>RS-5658</td><td></td></tr><tr><td>Castroviejo needle holder, 5.75&quot; straight with lock</td><td>Roboz</td><td>RS-6412</td><td></td></tr><tr><td>Stevens scissors 4.25&quot; curved blunt/blunt</td><td>Roboz</td><td>RS-5943</td><td></td></tr><tr><td>Alm retractor 4&quot; 4X4 5mm blunt prongs</td><td>Roboz</td><td>RS-6514</td><td>2x</td></tr><tr><td>Backhaus towel clamp 3.5&quot;</td><td>Roboz</td><td></td><td>4x</td></tr><tr><td>Micro clip setting forceps 4.75&quot;</td><td>Roboz</td><td>RS-6496</td><td></td></tr><tr><td>Micro vascular clips, 11 mm</td><td>Roboz</td><td></td><td></td></tr><tr><td>Surg-I-Loop</td><td>Scanlan International</td><td>1001-81M</td><td>5 cm length</td></tr><tr><td>Bonaccolto forceps, 4&amp;rdquo; (10 cm) long longitudinal serrations, cross serrated tip, 1.2mm tip width</td><td>Roboz</td><td>RS-5210</td><td></td></tr><tr><td>Dumont #3 forceps Inox tip size .17 &amp;times; .10 mm</td><td>Roboz</td><td>RS-5042</td><td></td></tr><tr><td>Graefe forceps, 4&amp;rdquo; (10 cm) long serrated straight, 0.8 mm tip</td><td>Roboz</td><td>RS-5280</td><td></td></tr><tr><td>Halstead mosquito forceps,&amp;nbsp; 5&quot; straight, 1.3 mm tips</td><td>Roboz</td><td>RS-7110</td><td>2x</td></tr><tr><td>Halstead mosquito forceps,&amp;nbsp; 5&quot; curved, 1.3mm tips</td><td>Roboz</td><td>RS-7111</td><td></td></tr><tr><td>Jacobson mosquito forceps 5&quot; curved extra delicate, 0.9 mm tips</td><td>Roboz</td><td>RS-7117</td><td></td></tr><tr><td>Kantrowitz forceps, 7.25&quot; 90 degree delicate, 1.7 mm tips</td><td>Roboz</td><td>RS-7305</td><td></td></tr><tr><td>Tissue forceps 5&quot;, 1X2 teeth, 2 mm tip width</td><td>Roboz</td><td>RS-8162</td><td></td></tr><tr><td>Allis-Baby forceps, 12 cm, 4x5 teeth, 3 mm tip width</td><td>Fine Science Tools</td><td>11092-12</td><td>2x</td></tr><tr><td>Adson forceps, 12 cm, serrated, straight</td><td>Fine Science Tools</td><td>11006-12</td><td></td></tr><tr><td>Veterinary electrosurgery handpiece and electrode</td><td>MACAN Manufacturing</td><td>HPAC-1; R-F11</td><td></td></tr><tr><td>&lt;strong&gt;Surgical Suite Equipment&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Circulating warm water blanket and pump</td><td>Multiple vendors</td><td>Catalog number not available</td><td></td></tr><tr><td>Bair hugger warming unit</td><td>3M</td><td>Model 505</td><td></td></tr><tr><td>IV infusion pump</td><td>Heska</td><td>Vet IV 2.2</td><td></td></tr><tr><td>Isoflurane vaporizer and scavenger</td><td>Multiple vendors</td><td>Catalog number not available</td><td></td></tr><tr><td>Veterinary multi-parameter monitor</td><td>Surgivet</td><td>Surgivet Advisor</td><td></td></tr><tr><td>Veterinary electrosurgery unit</td><td>MACAN Manufacturing</td><td>MV-9</td><td></td></tr><tr><td>Surgical microscope</td><td>D.F. Vasconcellos</td><td>M900</td><td>25X magnification for vein graft surgery; 16X magnification for gene transfer surgery</td></tr></tbody></table>

a rabbit model of durable transgene expression in jugular vein to common carotid artery interposition grafts

静脉搭桥术是治疗闭塞性动脉疾病的常见方法;然而, 由于血栓形成、内膜增生和动脉粥样硬化的移植失败, 长期的成功是有限的。本文的目的是演示一种将双侧静脉介入移植在兔身上的方法, 然后用基因转移载体传感移植物, 达到持久的转基因表达。该方法可用于研究基因及其蛋白产物在正常静脉移植稳态中的生物学作用。它还允许测试转基因的活动, 以防止静脉移植失败, e., 转基因的表达是否能防止新生内膜的生长, 减少血管炎症, 或降低家兔动脉粥样硬化高脂肪饮食。在最初的生存手术中, 左颈外静脉的部分被切除, 并放置在双边上, 作为反向端对侧颈总动脉介入移植。在第二次生存手术中, 28 天后进行, 每一个移植从循环与血管片段分离, 流明是填补 (通过 arteriotomy) 与解决方案, 其中包含帮助者依赖腺病毒 (HDAd) 向量。经过20分钟的孵化, 矢量解吸气, arteriotomy 修复, 流恢复。这些静脉是在个别实验规程所规定的时间点上收获的。移植术与转导之间的28天延迟是确保移植静脉适应动脉循环的必要条件。这种适应避免了快速丢失的转基因表达, 发生在静脉移植转基因之前或紧接移植后。该方法具有在嫁接静脉中获得持久、稳定的转基因表达能力的独特性。与其他大型动物静脉移植模型相比, 家兔具有成本低、处理方便等优点。与啮齿动物静脉移植模型相比, 家兔具有更大、更易于操作的血管, 为分析提供了丰富的组织。

在操作员可以使用此方法生成实验数据之前, 必须验证新操作员的技术熟练程度。新的操作者必须达到的第一个里程碑是, 在初始静脉移植术和随后的延迟转导手术后, 保持静脉移植通畅。每次手术后90% 多的通畅是可取的和可实现的。可通过经皮超声评估无创通畅, 我们通常在术后5-7 天进行。在具有专利静脉的家兔中, 超声检查将显示一大血管, 在前颈部两侧有轻快的骶管?...

Watch this Scientific Journal Video about A Rabbit Model of Durable Transgene Expression in Jugular Vein to Common Carotid Artery Interposition Grafts at JoVE.com

A Rabbit Model of Durable Transgene Expression in Jugular Vein to Common Carotid Artery Interposition Grafts

颈动脉介入移植中颈静脉持久性转基因表达的家兔模型

该方法描述了介入静脉移植在家兔中的位置、移植物的转导以及持久性转基因表达的实现。这允许调查转基因及其蛋白产品在接枝静脉中的生理和病理作用, 以及对静脉移植病基因治疗的检测。

Vein graft bypass surgery is a common treatment for occlusive arterial disease; however, long-term success is limited by graft failure due to thrombosis, ...

a-rabbit-model-durable-transgene-expression-jugular-vein-to-common

Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

10.3K 次观看.  University of Washington School of Medicine. 该方法描述了介入静脉移植在家兔中的位置、移植物的转导以及持久性转基因表达的实现。这允许调查转基因及其蛋白产品在接枝静脉中的生理和病理作用, 以及对静脉移植病基因治疗的检测。

视频: A Rabbit Model of Durable Transgene Expression in Jugular Vein to Common Carotid Artery Interposition Grafts

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

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

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

培养一个精密的显微外科技术在大鼠颈总动脉，颈外静脉移植：介入

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

科研

医学

Murine Cervical Heart Transplantation Model Using a Modified Cuff Technique

Mouse models are of special interest in research since a wide variety of monoclonal antibodies and commercially defined inbred and knockout strains are available to perform mechanistic in vivo studies. While heart transplantation models using a suture technique were first successfully developed in rats, the translation into an equally widespread used murine equivalent was never achieved due the technical complexity of the microsurgical procedure. In contrast, non-suture cuff techniques, also developed initially in rats, were successfully adapted for use in mice1-3. This technique for revascularization involves two major steps I) everting the recipient vessel over a polyethylene cuff; II) pulling the donor vessel over the formerly everted recipient vessel and holding it in place with a circumferential tie. This ensures a continuity of the endothelial layer, short operating time and very high patency rates4.
Using this technique for vascular anastomosis we performed more than 1,000 cervical heart transplants with an overall success rate of 95%. For arterial inflow the common carotid artery and the proximal aortic arch were anastomosed resulting in a retrograde perfusion of the transplanted heart. For venous drainage the pulmonary artery of the graft was anastomosed with the external jugular vein of the recipient5.
Herein, we provide additional details of this technique to supplement the video.

The murine cervical heart transplantation model is well suited for immunological as well as ischemia reperfusion injury studies. We modified the procedure using a non-suture cuff technique and performed more than 1,000 successful transplants with this approach.
Herein, we provide additional details of this technique to supplement the video.

用改进的袖套法小鼠颈部心脏移植模型

Mouse models are of special interest in research since a wide variety of monoclonal antibodies and commercially defined inbred and knockout strains are ...

Heterotopic Cervical Heart Transplantation in Mice

The heterotopic cervical heart transplantation in mice is a valuable tool in transplant and cardiovascular research. The cuff technique greatly simplifies this model by avoiding challenging suture anastomoses of small vessels thereby reducing warm ischemia time. In comparison to abdominal graft implantation the cervical model is less invasive and the implanted graft is easily accessible for further follow-up examinations. Anastomoses are performed by pulling the ascending aorta of the graft over the cuff with the recipient&#8217;s common carotid artery and by pulling the main pulmonary artery over the cuff with the external jugular vein. Selection of appropriate cuff size and complete mobilization of the vessels are important for successful revascularization. Ischemia-reperfusion (I/R) injury can be minimized by perfusing the graft with a cardioplegic solution and by hypothermia. In this article, we provide technical details for a simplified and improved cuff technique, which should allow surgeons with basic microsurgical skills to perform the procedure with a high success rate.

The cuff technique shortens and facilitates the model of heterotopic cervical heart transplantation in mice by avoiding technically challenging suture anastomoses of small vessels. The technical details shown in this video paper should allow researches to establish this model in their laboratories.

颈椎异位心脏移植小鼠

The heterotopic cervical heart transplantation in mice is a valuable tool in transplant and cardiovascular research. The cuff technique greatly simplifies ...

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. ...

In Vivo Gene Transfer to the Rabbit Common Carotid Artery Endothelium

The goal of this method is to introduce a transgene into the endothelium of isolated segments of both rabbit common carotid arteries. The method achieves focal endothelial-selective transgenesis, thereby allowing an investigator to determine the biological roles of endothelial-expressed transgenes and to quantify the in vivo transcriptional activity of DNA sequences in large artery endothelial cells. The method uses surgical isolation of rabbit common carotid arteries and an arteriotomy to deliver a transgene-expressing viral vector into the arterial lumen. A short incubation period of the vector in the lumen, with subsequent aspiration of the lumen contents, is sufficient to achieve efficient and durable expression of the transgene in the endothelium, with no detectable transduction or expression outside of the isolated arterial segment. The method allows assessment of the biological activities of transgene products both in normal arteries and in models of human vascular disease, while avoiding systemic effects that could be caused either by targeting gene delivery to other sites (e.g. the liver) or by the alternative approach of delivering genetic constructs to the endothelium by germ line transgenesis. Application of the method is limited by the need for a skilled surgeon and anesthetist, a well-equipped operating room, the costs of purchasing and housing rabbits, and the need for expertise in gene-transfer vector construction and use. Results obtained with this method include: transgene-related alterations in arterial structure, cellularity, extracellular matrix, or vasomotor function; increases or reductions in arterial inflammation; alterations in vascular cell apoptosis; and progression, retardation, or regression of diseases such as intimal hyperplasia or atherosclerosis. The method also allows measurement of the ability of native and synthetic DNA regulatory sequences to alter transgene expression in endothelial cells, providing results that include: levels of transgene mRNA, levels of transgene protein, and levels of transgene enzymatic activity.

In Vivo Gene Transfer to the Rabbit Common Carotid Artery Endothelium

This method is to introduce a transgene into the endothelium of rabbit carotid arteries. Introduction of the transgene allows the assessment of the biological role of the transgene product either in normal arteries or disease models. The method is also useful for measuring activity of DNA regulatory sequences.

体内兔颈总动脉内皮的基因转移

The goal of this method is to introduce a transgene into the endothelium of isolated segments of both rabbit common carotid arteries. The method achieves ...

Murine Cervical Aortic Transplantation Model using a Modified Non-Suture Cuff Technique

With the introduction of powerful immunosuppressive protocols, distinct advances are possible in the prevention and therapy of acute rejection episodes. However, only minor improvement in the long-term results of transplanted solid organs could be observed over the past decades. In this context, chronic allograft vasculopathy (CAV) still represents the leading cause of late organ failure in cardiac, renal and pulmonary transplantation.
Thus far, the underlying pathogenesis of CAV development remains unclear, explaining why effective treatment strategies are presently missing and emphasizing a need for relevant experimental models in order to study the underlying pathophysiology leading to CAV formation. The following protocol describes a murine heterotopic cervical aortic transplantation model using a modified non-suture cuff technique. In this technique, a segment of the thoracic aorta is interpositioned in the right common carotid artery. With the use of the non-suture cuff technique, an easy to learn and reproducible model can be established, minimizing the possible heterogeneity of sutured vascular micro anastomoses.

Here, we present a protocol of heterotopic aortic transplantation in mice using the non-suture cuff technique in a cervical murine model. This model can be used to study the underlying pathology of chronic allograft vasculopathy (CAV) and can help evaluate new therapeutic agents in order to prevent its formation.

使用改性非缝合铐技术的宫颈主动脉移植模型

With the introduction of powerful immunosuppressive protocols, distinct advances are possible in the prevention and therapy of acute rejection episodes. ...

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 ...

Implantation of Tissue-Engineered Vascular Graft in Mouse Carotid Artery via Cuff Technique

The development of small-diameter vascular grafts has been a global endeavor, with numerous research groups contributing to this field. Animal experimentation plays a pivotal role in assessing the efficacy and safety of vascular grafts, particularly in the absence of clinical applications. Compared to alternative animal models, the mouse implantation model offers several advantages, including a well-defined genetic background, a mature method for disease model construction, and a straightforward surgical procedure. Based on these advantages, the present study devised a simple cuff technique for the implantation of tissue-engineered vascular grafts in the mouse carotid artery. This technique began with the fabrication of polycaprolactone (PCL) small-diameter vascular grafts via electrostatic spinning, followed by the seeding of macrophages onto the grafts through perfusion adsorption. Subsequently,&#160;the cellularized tissue-engineered vascular grafts were transplanted into the mouse carotid artery using the cuff technique to evaluate patency and regenerative capability. After 30 days of in vivo implantation, vascular patency was found to be satisfactory, with evidence of neo-tissue regeneration and the formation of an endothelial layer within the lumen of the grafts. All data were analyzed using statistical and graphing software. This study successfully established a mouse carotid artery implantation model that can be used to explore the cellular sources of vascular regeneration and the mechanisms of action of active substances. Furthermore, it provides theoretical support for the development of novel small-diameter vascular grafts.

Here, a protocol is presented for the implantation of a tissue-engineered vascular graft into the mouse carotid artery using the cuff technique, providing a suitable animal model for investigating vascular tissue regeneration mechanisms.

通过 Cuff 技术将组织工程血管移植物植入小鼠颈动脉

The development of small-diameter vascular grafts has been a global endeavor, with numerous research groups contributing to this field. Animal ...

生物工程

A Simplified Model for Heterotopic Heart Valve Transplantation in Rodents

There is an urgent clinical need for heart valve replacements that can grow in children. Heart valve transplantation is proposed as a new type of transplant with the potential to deliver durable heart valves capable of somatic growth with no requirement for anticoagulation. However, the immunobiology of heart valve transplants remains unexplored, highlighting the need for animal models to study this new type of transplant. Previous rat models for heterotopic aortic valve transplantation into the abdominal aorta have been described, though they are technically challenging and costly. For addressing this challenge, a renal subcapsular transplant model was developed in rodents as a practical and more straightforward method for studying heart valve transplant immunobiology. In this model, a single aortic valve leaflet is harvested and inserted into the renal subcapsular space. The kidney is easily accessible, and the transplanted tissue is securely contained in a subcapsular space that is well vascularized and can accommodate a variety of tissue sizes. Furthermore, because a single rat can provide three donor aortic leaflets and a single kidney can provide multiple sites for transplanted tissue, fewer rats are required for a given study. Here, the transplantation technique is described, providing a significant step forward in studying the transplant immunology of heart valve transplantation.

This protocol describes a simple and efficient method for the transplantation of aortic valve leaflets under the renal capsule to allow for the study of alloreactivity of heart valves.

啮齿动物异位心脏瓣膜移植的简化模型

There is an urgent clinical need for heart valve replacements that can grow in children. Heart valve transplantation is proposed as a new type of ...

Jugular Vein Graft Harvest From a Rabbit Model: A Surgical Procedure to Excise a Segment of Rabbit Jugular Vein for Ex Vivo Studies

Source: Nishio, H., et al. <a href="https://jove.unicartagenaproxy.elogim.com/v/60931">A Rabbit Venous Interposition Model Mimicking Revascularization Surgery using Vein Grafts to Assess Intimal Hyperplasia under Arterial Blood Pressure</a> J. Vis. Exp. (2020).
In this video, we demonstrate a procedure for surgically harvesting a segment of the jugular vein from a rabbit model. The resected segment can be used as an autologous vein graft to study intimal hyperplasia that develops due to arterial blood pressure.

从兔模型中收获的颈静脉移植物:切除一段兔颈静脉用于离体研究的外科手术

Source: Nishio, H., et al. A Rabbit Venous Interposition Model Mimicking Revascularization Surgery using Vein Grafts to Assess Intimal Hyperplasia under ...