Name: an immunological model for heterotopic heart and cardiac muscle cell transplantation in rats
Uploaded: 2020-05-08T13:30:00
Description: Watch this Scientific Journal Video about An Immunological Model for Heterotopic Heart and Cardiac Muscle Cell Transplantation in Rats at JoVE.com

我们要感谢布里塔·特劳特维格、科林娜·勒伯特和英格丽德·梅德的承诺。

所有动物经验均按照下萨克森州（德国奥尔登堡州拉维斯）消费者保护和食品安全区域当局当地道德动物审查委员会的准则进行，并经批准，12/0768 和 17/2472。
1. 心脏外植和灌注
注：作为移植捐赠者，使用7-22周年龄的雌性或雄性大鼠。
<ol><li>通过吸入的二氟拉纳对供体大鼠进行麻醉（感应5%，维持3%，O2流量为1升/分钟）。注射5毫克Carprofen下皮每...

<ol>
	<li>Abbott, C. 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=A+Technique+for+Heart+Transplantation+In+the+Rat.">A Technique for Heart Transplantation In the Rat.</a> Archives of Surgery. 89 (4), 645-652 (1964).</li><li>Tomita, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heart+homotransplantation+in+the+rat.">Heart homotransplantation in the rat.</a> Sapporo igaku zasshi. The Sapporo Medical Journal. 30 (4), 165-183 (1966).</li><li>Ono, K., Lindsey, E. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Improved+technique+of+heart+transplantation+in+rats.">Improved technique of heart transplantation in rats.</a> The Journal of Thoracic and Cardiovascular Surgery. 57 (2), 225-229 (1969).</li><li>Wen, 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=A+simple+technique+for+a+new+working+heterotopic+heart+transplantation+model+in+rats.">A simple technique for a new working heterotopic heart transplantation model in rats.</a> Transplantation Proceedings. 45 (6), 2522-2526 (2013).</li><li>Benke, K., 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=Heterotopic+abdominal+rat+heart+transplantation+as+a+model+to+investigate+volume+dependency+of+myocardial+remodeling.">Heterotopic abdominal rat heart transplantation as a model to investigate volume dependency of myocardial remodeling.</a> Transplantation. 101 (3), 498-505 (2017).</li><li>Kearns, M. J., 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=Rat+Heterotopic+Abdominal+Heart/Single-lung+Transplantation+in+a+Volume-loaded+Configuration.">Rat Heterotopic Abdominal Heart/Single-lung Transplantation in a Volume-loaded Configuration.</a> Journal of Visualized Experiments. (99), 52418 (2015).</li><li>Ibrahim, 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=Heterotopic+abdominal+heart+transplantation+in+rats+for+functional+studies+of+ventricular+unloading.">Heterotopic abdominal heart transplantation in rats for functional studies of ventricular unloading.</a> The Journal of Surgical Research. 179 (1), e31-e39 (2013).</li><li>Liu, F., Kang, S. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heterotopic+heart+transplantation+in+mice.">Heterotopic heart transplantation in mice.</a> Journal of Visualized Experiments. (6), 238 (2007).</li><li>Lu, W., 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+new+simplified+volume-loaded+heterotopic+rabbit+heart+transplant+model+with+improved+techniques+and+a+standard+operating+procedure.">A new simplified volume-loaded heterotopic rabbit heart transplant model with improved techniques and a standard operating procedure.</a> Journal of Thoracic Disease. 7 (4), 653-661 (2015).</li><li>Kitahara, H., 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=Heterotopic+transplantation+of+a+decellularized+and+recellularized+whole+porcine+heart.">Heterotopic transplantation of a decellularized and recellularized whole porcine heart.</a> Interactive Cardiovascular and Thoracic Surgery. 22 (5), 571-579 (2016).</li><li>Kadner, 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=Heterotopic+heart+transplantation:+experimental+development+and+clinical+experience.">Heterotopic heart transplantation: experimental development and clinical experience.</a> European Journal of Cardiothoracic Surgery. 17 (4), 474-481 (2000).</li><li>Zin&#246;cker, 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=Immune+reconstitution+and+graft-versus-host+reactions+in+rat+models+of+allogeneic+hematopoietic+cell+transplantation.">Immune reconstitution and graft-versus-host reactions in rat models of allogeneic hematopoietic cell transplantation.</a> Frontiers in Immunology. 3 (NOV), 1-12 (2012).</li><li>Klempnauer, J., 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=Genetic+control+of+rat+heart+allograft+rejection:+effect+of+different+MHC+and+non-MHC+incompatibilities.">Genetic control of rat heart allograft rejection: effect of different MHC and non-MHC incompatibilities.</a> Immunogenetics. 30, 81-88 (1989).</li><li>Huang, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+manipulations+in+the+rat:+Progress+and+prospects.">Genetic manipulations in the rat: Progress and prospects.</a> Current Opinion in Nephrology and Hypertension. 20 (4), 391-399 (2011).</li><li>Gordon, C. 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R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+new+modified+technique+for+heterotopic+femoral+heart+transplantation+in+rats.">A new modified technique for heterotopic femoral heart transplantation in rats.</a> The Journal of Surgical Research. 139 (2), 157-163 (2007).</li><li>Ma, Y., Wang, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+2+heterotopic+heart+transplant+techniques+in+rats:+cervical+and+abdominal+heart.">Comparison of 2 heterotopic heart transplant techniques in rats: cervical and abdominal heart.</a> Experimental and Clinical Transplantation. 9 (2), 128-133 (2011).</li><li>Bektas, H., 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=Differential+effect+of+donor-specific+blood+transfusions+after+kidney,+heart,+pancreas,+and+skin+transplantation+in+major+histocompatibility+complex-incompatible+rats.">Differential effect of donor-specific blood transfusions after kidney, heart, pancreas, and skin transplantation in major histocompatibility complex-incompatible rats.</a> Transfusion. 37 (2), 226-230 (1997).</li><li>Saiho, K. O., 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+allograft+acceptance+induced+by+single+dose+anti-leukocyte+common+antigen+(RT7)+antibody+in+the+rat.">Long-term allograft acceptance induced by single dose anti-leukocyte common antigen (RT7) antibody in the rat.</a> Transplantation. 71 (8), 1124-1131 (2001).</li><li>Bektas, H., 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=Blood+transfers+infectious+immunologic+tolerance+in+MHC-incompatible+heart+transplantation+in+rats.">Blood transfers infectious immunologic tolerance in MHC-incompatible heart transplantation in rats.</a> Journal of Heart and Lung Transplantation. 24 (5), 614-617 (2005).</li><li>J&#228;ger, M. D., 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=Sirolimus+promotes+tolerance+for+donor+and+recipient+antigens+after+MHC+class+II+disparate+bone+marrow+transplantation+in+rats.">Sirolimus promotes tolerance for donor and recipient antigens after MHC class II disparate bone marrow transplantation in rats.</a> Experimental Hematology. 35 (1), 164-170 (2007).</li><li>Timrott, K., 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=Application+of+allogeneic+bone+marrow+cells+in+view+of+residual+alloreactivity:+Sirolimus+but+not+cyclosporine+evolves+tolerogenic+properties.">Application of allogeneic bone marrow cells in view of residual alloreactivity: Sirolimus but not cyclosporine evolves tolerogenic properties.</a> PLoS ONE. 10 (4), 1-16 (2015).</li><li>Hadamitzky, 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=Memory-updating+abrogates+extinction+of+learned+immunosuppression.">Memory-updating abrogates extinction of learned immunosuppression.</a> Brain, Behavior, and Immunity. 52, 40-48 (2016).</li><li>Beetz, O., 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=Recipient+natural+killer+cells+alter+the+course+of+rejection+of+allogeneic+heart+grafts+in+rats.">Recipient natural killer cells alter the course of rejection of allogeneic heart grafts in rats.</a> Plos One. 14 (8), e0220546 (2019).</li><li>Hirschburger, 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=Nicotine+Attenuates+Macrophage+Infiltration+in+Rat+Lung+Allografts.">Nicotine Attenuates Macrophage Infiltration in Rat Lung Allografts.</a> The Journal of Heart and Lung Transplantation. 28 (5), 493-500 (2009).</li><li>Ruzza, 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=Heterotopic+heart+transplantation+in+rats:+improved+anesthetic+and+surgical+technique.">Heterotopic heart transplantation in rats: improved anesthetic and surgical technique.</a> Transplantation Proceedings. 42 (9), 3828-3832 (2010).</li><li>Wang, D., 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+technique+for+heart+transplantation+in+rats:+abdominal+vessel+branch-sparing+and+modified+venotomy.">A simplified technique for heart transplantation in rats: abdominal vessel branch-sparing and modified venotomy.</a> Microsurgery. 26 (6), 470-472 (2006).</li><li>Wang, 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=A+modified+method+for+heterotopic+mouse+heart+transplantion.">A modified method for heterotopic mouse heart transplantion.</a> Journal of Visualized Experiments. 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先前描述的大鼠异位心脏移植方法主要基于1969年对小野和林赛的描述。自那时以来，对各种物种进行了若干修改，导致该模型的广泛多样性。结合其中几个修改，并介绍我们自己的经验，在实验室进行了超过30年的异位心脏移植，我们创造了一个可行的手术方法，不需要长时间的训练或手术背景。在下面，我们将讨论此模型的一般限制，并强调协议的关键步骤。此外，我们将强?...

异位心脏移植是一种常用的实验模型，用于不同研究移植耐受性、急性和慢性异体移植排斥、缺血再灌注损伤、机器灌注或心脏重塑等。除其他优点外，移植功能可以通过触觉进行非侵入性监测，移植失败不会导致接受者与其他器官（如肾脏或肝脏）相比的重要损伤。
1964年，Abbott等人最初描述了大鼠1的异位腹心移植。后来，在1966年，原子塔等人对阿纳斯托莫的端到端技术作了描述。1969年，奥诺和林赛报告了目前使用的模型的基础。在过去的几十年中，已经发表了一些修改，以创建不同类型的卸载，部分加载或加载左心室心脏移植，包括联合异位心肺移植44，5，6。5,6对于免疫分析，最常见的是非体积加载心脏移植移植。在这种情况下，血流逆行进入供体上升主动脉，随后进入冠状动脉。静脉排水沿冠状窦进入右中庭和心室（图1A-B）。因此，除了来自Thebesian静脉的少量血液外，左心室被排除在血流之外。这也使它成为研究左心室辅助装置治疗7期间的病理生理机制的有用模型。
异位心脏移植已在各种物种中进行，包括小鼠、兔子、猪，甚至被用作,人类8、9、10、119的单体或双心形辅助8装置。10,11大鼠仍然是移植模型的流行实验动物，特别是因为不同大鼠菌株组合的移植存活时间在过去已经明确，大量的免疫试剂是可访问的12，13。12,与老鼠不同，大鼠做手术和获得淋巴组织进行免疫分析更可行12。此外，近年来在大鼠身上引入商业性克隆技术，很可能导致对实验鼠模型14的反复关注。
一般来说，异位心脏移植可以通过进行宫颈或腹部麻醉附着在受体血管上。然而，一些研究表明，由于更好的获得手动触觉或跨体波他心动图，因此可以更精确地检测移植失败15，16，因此，股骨性麻醉有助于改善监测。15,
结果表明，两种阿生虫技术在操作时间、并发症率、结果和移植生存时间上没有差异。显然，必须提及有足够的排空淋巴结，作为宫颈麻醉的好处;但是，需要较长的培训期。相反，腹部麻醉对于免疫学研究来说不那么复杂，同样有价值，特别是当结合一种新疗法的异源性心肌细胞和随后的宫颈淋巴切除术的结果时。两种模型的组合提供了广泛的干预后免疫分析。
以下协议是指对外科医生进行手术，以减少缺血时间。但是，所有实验都可以由一个人执行。图2A-B显示了心脏外植和植入仪器和材料的设置。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Anesthesia device (including isoflurane vaporizer)</td>
			<td>Summit Anesthesia Solutions</td>
			<td>No Catalog Number available</td>
			<td></td>
		</tr>
		<tr>
			<td>Cannula (27 G)</td>
			<td>BD Microlance</td>
			<td>302200</td>
			<td></td>
		</tr>
		<tr>
			<td>Carprofen</td>
			<td>Pfizer</td>
			<td>Rimadyl 50 mg/mL</td>
			<td></td>
		</tr>
		<tr>
			<td>Cellstar Tubes (15 mL)</td>
			<td>GreinerBioOne</td>
			<td>188271</td>
			<td></td>
		</tr>
		<tr>
			<td>Cell strainer (40 &#181;m)</td>
			<td>BD Falcon</td>
			<td>2271680</td>
			<td></td>
		</tr>
		<tr>
			<td>Collagenase Type CLSII</td>
			<td>Biochrome</td>
			<td>C2-22</td>
			<td></td>
		</tr>
		<tr>
			<td>Compresses 5x5 cm</td>
			<td>Fuhrmann</td>
			<td>31501</td>
			<td></td>
		</tr>
		<tr>
			<td>Compresses 7.5x7.5 cm</td>
			<td>Fuhrmann</td>
			<td>31505</td>
			<td></td>
		</tr>
		<tr>
			<td>Cotton swabs</td>
			<td>Heinz Herenz Medizinalbedarf</td>
			<td>1032128</td>
			<td></td>
		</tr>
		<tr>
			<td>Dexpathenol (5 %)</td>
			<td>Bayer</td>
			<td>&#34;Bepanthen&#34;</td>
			<td></td>
		</tr>
		<tr>
			<td>DPBS BioWhittaker</td>
			<td>Lonza</td>
			<td>17-512F</td>
			<td></td>
		</tr>
		<tr>
			<td>Forceps</td>
			<td>B. Braun</td>
			<td>Aesculap BD557R</td>
			<td></td>
		</tr>
		<tr>
			<td>Forceps</td>
			<td>B. Braun</td>
			<td>Aesculap BD313R</td>
			<td></td>
		</tr>
		<tr>
			<td>Forceps</td>
			<td>B. Braun</td>
			<td>Aesculap BD35</td>
			<td></td>
		</tr>
		<tr>
			<td>Heating mat</td>
			<td>Gaymar Industries</td>
			<td>&#34;T/Pump&#34;</td>
			<td></td>
		</tr>
		<tr>
			<td>Hemostatic gauze</td>
			<td>Ethicon</td>
			<td>Tabotamp</td>
			<td></td>
		</tr>
		<tr>
			<td>Heparin-Natrium 25 000 I.E.</td>
			<td>Ratiopharm</td>
			<td>No Catalog Number available</td>
			<td></td>
		</tr>
		<tr>
			<td>Isofluran CP</td>
			<td>CP-Pharma</td>
			<td>No Catalog Number available</td>
			<td></td>
		</tr>
		<tr>
			<td>Large-pored sieve (stainless steel)</td>
			<td>Forschungswerkst&#228;tten Hannover Medical School</td>
			<td>No Catalog Number available</td>
			<td></td>
		</tr>
		<tr>
			<td>Lidocaine</td>
			<td>Astra Zeneca</td>
			<td>2 % Xylocain</td>
			<td></td>
		</tr>
		<tr>
			<td>Metamizol-Natrium</td>
			<td>Ratiopharma</td>
			<td>Novaminsulfon 500 mg/mL</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro forceps</td>
			<td>B. Braun</td>
			<td>Aesculap BD3361</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro needle holder</td>
			<td>Codman, Johnson &#38; Johnson Medical</td>
			<td>Codmann 80-2003</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro needle holder</td>
			<td>B. Braun</td>
			<td>Aesculap BD336R</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro needle holder</td>
			<td>B. Braun</td>
			<td>Aesculap FD241R</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro scissors</td>
			<td>B. Braun</td>
			<td>Aesculap FD101R</td>
			<td></td>
		</tr>
		<tr>
			<td>Micro scissors</td>
			<td>B. Braun</td>
			<td>Aesculap FM471R</td>
			<td></td>
		</tr>
		<tr>
			<td>Needle holder</td>
			<td>B. Braun</td>
			<td>Aesculap BM221R</td>
			<td></td>
		</tr>
		<tr>
			<td>Penicillin/Streptomycin/Glutamine (100x)</td>
			<td>PAA</td>
			<td>P11-010</td>
			<td></td>
		</tr>
		<tr>
			<td>Peripheral venous catheter (18 G)</td>
			<td>B. Braun</td>
			<td>4268334B</td>
			<td></td>
		</tr>
		<tr>
			<td>Peripheral venous catheter (22 G)</td>
			<td>B. Braun</td>
			<td>4268091B</td>
			<td></td>
		</tr>
		<tr>
			<td>Probe pointed scissors</td>
			<td>B. Braun</td>
			<td>Aesculap BC030R</td>
			<td></td>
		</tr>
		<tr>
			<td>Retractors</td>
			<td>Forschungswerkst&#228;tten Hannover Medical School</td>
			<td>No Catalog Number available</td>
			<td></td>
		</tr>
		<tr>
			<td>RPMI culture medium</td>
			<td>Lonza</td>
			<td>BE12-702F</td>
			<td></td>
		</tr>
		<tr>
			<td>Saline solution (NaCl 0.9 %)</td>
			<td>Baxter</td>
			<td>No Catalog Number available</td>
			<td></td>
		</tr>
		<tr>
			<td>Scissors</td>
			<td>B. Braun</td>
			<td>Aesculap BC414</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical microscope</td>
			<td>Carl-Zeiss</td>
			<td>OPMI-MDM</td>
			<td></td>
		</tr>
		<tr>
			<td>Sutures (anastomoses)</td>
			<td>Catgut</td>
			<td>Mariderm 8-0 monofil</td>
			<td></td>
		</tr>
		<tr>
			<td>Sutures (ligature)</td>
			<td>Resorba</td>
			<td>Silk 5-0 polyfil</td>
			<td></td>
		</tr>
		<tr>
			<td>Sutures (skin, fascia)</td>
			<td>Ethicon</td>
			<td>Mersilene 3-0</td>
			<td></td>
		</tr>
		<tr>
			<td>Syringe (1 mL)</td>
			<td>B. Braun</td>
			<td>9166017V</td>
			<td></td>
		</tr>
		<tr>
			<td>Syringe (10 mL)</td>
			<td>B. Braun</td>
			<td>4606108V</td>
			<td></td>
		</tr>
		<tr>
			<td>Syringe (20 mL)</td>
			<td>B. Braun</td>
			<td>4606205V</td>
			<td></td>
		</tr>
		<tr>
			<td>Vascular clamp</td>
			<td>B. Braun</td>
			<td>Aesculap FB708R</td>
			<td></td>
		</tr>
	</tbody>
</table>

an immunological model for heterotopic heart and cardiac muscle cell transplantation in rats

50多年来，大鼠异位心脏移植一直是各种免疫学研究的常用模型。自1964年第一次描述以来，已经报告了若干修改。在大鼠进行异位心脏移植30年后，我们开发了一种简化的手术方法，无需进一步手术训练或背景，即可轻松教授和进行。
在解剖上升主动脉和肺动脉和上等类腔和肺静脉结扎后，供体心脏被收获，然后注入冰冷盐水溶液补充肝素。在夹紧和切开受体腹血管后，捐赠者提升主动脉和肺动脉分别被麻醉到受体腹部主动脉和劣质的vena卡瓦，使用连续的导联。
根据不同的供体-受体组合，此模型允许分析异体移植的急性或慢性排斥。该模型的免疫意义进一步增强，通过一种新的注射重要心肌细胞的耳内注射方法，以及随后对排空颈淋巴组织的分析， 进一步增强了该模型的免疫意义。

过去，不同的免疫问题都根据该模型得到解决，在工作小组中，500多例移植进行了验证，存活率超过95%，13、18、19、20、21、22、23、24。23,2413,18,19,20,21,22,总手术时间（包?...

Watch this Scientific Journal Video about An Immunological Model for Heterotopic Heart and Cardiac Muscle Cell Transplantation in Rats at JoVE.com

An Immunological Model for Heterotopic Heart and Cardiac Muscle Cell Transplantation in Rats

我们描述了大鼠异位腹心移植的模型，这意味着对当前策略的修改，从而导致简化的手术方法。此外，我们还描述了一种新的排斥模型，通过注射重要的心肌细胞的耳朵内，允许进一步的移植免疫学分析在大鼠。

大鼠异位心脏和心肌细胞移植的免疫模型

Heterotopic heart transplantation in rats has been a commonly used model for diverse immunological studies for more than 50 years. Several modifications ...

移植模型是免疫学研究的宝贵工具，往往需要外科专业知识。在这里，我们提出了一个可行的方法，在大鼠的异位心和心脏肌肉细胞移植。通过突出和简化我们模型的所有关键步骤，没有手术背景的研究人员应该能够在合理的时间内掌握这个模型。这两种移植模型都提供了对耐受和排斥机制的宝贵见解，并可应用于各种物种。我们有很多经验，教这个模型给个人没有事先的手术培训。在学习了关键步骤后，我们建议对死去的动物保持耐心和足够的练习。对于供体心脏外植，在确认对麻醉7至22周大大鼠供体动物踏板反射缺乏反应后，应用眼部润滑剂，使用机械剪子去除腹部和胸毛。将大鼠放在加热基座上的上点位置，用弹性带固定操作台底部的四肢。用70%乙醇对裸露的皮肤进行消毒，并进行中位腹腔切除术。将缩回器插入切口，并使用灭菌棉签将肠子调动到捐赠者左侧，以暴露劣质的维纳卡瓦。接下来，穿刺劣质的维纳卡瓦，允许静脉注射500国际单位的肝素溶解在一毫升的冰冷同位素盐水溶液中。使用棉签用轻压缩针缩回针头后，停止被刺穿地点的出血。将隔膜化，对捐赠者的两侧进行侧胸切除术，并固定胸壁。使用两个微针架钝制备，去除心绞痛和虚神经。对于排泄，将腹部血管转开，将探针的钝枝插入横向心缘鼻窦中。使用压缩解剖上升的主动脉和肺动脉尽可能在心脏的光冠牵引下。将单个 5-0 连结字围绕上优和劣质的静脉和肺静脉，并尽可能用多处拧紧连结步骤。将组织后向连结分离并提取心脏。要使被移植的心脏被移植，使用静脉导管中的18个量表管冲洗30毫升的冰冷，同位素盐水溶液，辅以1000国际单位的肝素通过上升主动脉和肺动脉，然后将心脏放在15毫升的盐水溶液在冰上。在准备了10至14周大的接受大鼠刚刚演示后，将肠道动员到接收者左上角，进行温暖、湿润的压缩。并调整手术显微镜或足够的替代方案。在调动十二指肠和近角后，分别使用五至七个全放大倍率，使用棉签和钝解剖来暴露腹主动脉和劣质的维纳卡瓦。使用两个微型针架提升腹部血管而不损伤腰椎静脉，将 Cooley 血管夹放在血管上。使用 30 至 45 度拱形 27 量表管刺穿腹部血管。并使用波茨剪刀将穿刺部位放大成与捐赠血管流明大小相匹配的纵向切口。为了去除血块和防止术后血栓形成，用盐水将血管渗透，并放入坐点。使用两个简单的中断 8 - 0单丝不可松吸的缝合线，将供体提升主动脉固定到纵向切口的颅角和骨角的接受者腹主动脉。用接受者的腹部主动脉对捐赠者的上主动脉进行原子化，在以8-0运行进行前半部分的阿托莫病时，将移植物放在接受者血管的右侧单丝缝合线。然后将移植物放在接受者血管的左侧，并进行下半部分的阿托莫病。以类似的方式将供体肺动脉固定到劣质的静脉道，在颅角和颅角有两个角结。在每个角落固定有结的捐赠血管后，从容器的外侧到容器内侧缝一针，以运行动脉瘤的上半部分的宫内结。在绑下缝合线之前，先从内部到外面再缝一针。为了防止在完成下半部分的结后外周栓塞，在紧固下半部分的结之前用盐水冲洗动脉瘤。在两个麻醉剂周围放置止血纱布，并小心释放 Cooley 血管夹，以便进行移植再灌注。用轻压缩和消毒棉签管理任何出血。大约60秒后，心脏应该开始跳动。小心地更换肠道，用单独的连续 3-0 多丝线线缝合线关闭腹部筋膜和皮肤。在适当的实验时间点，收获一个捐赠的老鼠心脏，如证明，并使用无菌手术刀或无菌剪刀粉碎心脏成三由三毫米碎片。在37摄氏度下，每毫升胶原酶补充0.5毫克，在37摄氏度下消化20毫升培养基的片段，30分钟。在消化结束时，将组织添加到大孔筛中，同时去除培养培养。加入5至10毫升的同位素盐水溶液，通过筛子对组织浆料进行应变。离心两次细胞悬浮液后，通过40微米细胞滤株过滤。用5至10毫升的同位素盐水溶液冲洗过滤器，收集任何剩余的细胞。离心后，以每毫升5倍10的浓度将分离的大鼠心脏细胞重新在盐水溶液中，再增。用细胞溶液加载一毫升注射器，将麻醉受援动物放在侧向位置，目标耳朵朝上。用手指和双面胶带固定耳朵，并使用 27 量表管皮下将 20 微升的心脏肌肉细胞溶液注射到视觉毛细血管附近到接受者的耳朵中。经过适当的实验观察期，提取排空的颈椎淋巴结，进行适当的下游兴趣分析。合成移植存活长达100天，没有移植失败的迹象。特定的供体-接受者组合可能导致异位心脏移植排斥速度的不同。被排斥心脏的低温分段显示免疫效应细胞的渗透增加，而合成移植物基本上没有免疫细胞。子宫颈淋巴切除术和口服心肌细胞注射后排出淋巴结细胞的再刺激测定揭示了异位性心脏组织在异位移植接受者中独特的菌株特异性免疫反应，值得进一步免疫学分析，如细胞因子分析。插入和提取针头时，请小心处理容器。不要太注意缝合线，以防止狭窄。不要为每次动脉瘤缝针太多。

Research

JoVE Journal

Immunology and Infection

Murine Model of CD40-activation of B cells

激活的B细胞CD40的小鼠模型

Research on B cells has shown that CD40 activation improves their antigen presentation capacity. When stimulated with interleukin-4 and CD40 ligand ...

科研

免疫与感染

Orthotopic Hind-Limb Transplantation in Rats

原位大鼠后肢移植

Composite tissue allotransplantation (CTA) now represents a valid therapeutic option after the loss of a hand, forearm or digits and has become a novel ...

An Ex vivo Model of an Oligodendrocyte-directed T-Cell Attack in Acute Brain Slices

An Ex vivo Model of an Oligodendrocyte-directed T-Cell Attack in Acute Brain Slices

一个<em&gt;体外</em&gt;急性脑片的少突胶质细胞定向T细胞攻击模式

Death of oligodendrocytes accompanied by destruction of neurons and axons are typical histopathological findings in cortical and subcortical grey matter ...

Assessing Bacterial Invasion of Cardiac Cells in Culture and Heart Colonization in Infected Mice Using Listeria monocytogenes

Assessing Bacterial Invasion of Cardiac Cells in Culture and Heart Colonization in Infected Mice Using Listeria monocytogenes

使用李斯特菌单细胞基因评估受感染小鼠培养和心脏殖民化中心脏细胞的细菌入侵

Listeria monocytogenes is a Gram-positive facultative intracellular pathogen that is capable of causing serious invasive infections in immunocompromised ...

Normal and Malignant Muscle Cell Transplantation into Immune Compromised Adult Zebrafish

正常和恶性肌细胞移植到免疫受损成年斑马鱼

Zebrafish have become a powerful tool for assessing development, regeneration, and cancer. More recently, allograft cell transplantation protocols have ...

An Endothelial Planar Cell Model for Imaging Immunological Synapse Dynamics

成像免疫突触动力学内皮平面细胞模型

Adaptive immunity is regulated by dynamic interactions between T cells and antigen presenting cells (&#39;APCs&#39;) referred to as &#39;immunological ...

Establishment of a High-throughput Setup for Screening Small Molecules That Modulate c-di-GMP Signaling in Pseudomonas aeruginosa

Establishment of a High-throughput Setup for Screening Small Molecules That Modulate c-di-GMP Signaling in Pseudomonas aeruginosa

高通量设置的建立筛选小分子能调节C-二GMP信令<em&gt;绿脓杆菌</em

Bacterial resistance to traditional antibiotics has driven research attempts to identify new drug targets in recently discovered regulatory pathways. ...

Isolation and Identification of Extravascular Immune Cells of the Heart

心脏外免疫细胞的分离与鉴定

The immune system is an essential component of a healthy heart. The myocardium is home to a rich population of different immune cell subsets with ...

Optimization of the Cuff Technique for Murine Heart Transplantation

穆林心脏移植袖口技术的优化

Murine cardiac transplantation has been performed for more than 40 years. With advancements in microsurgery, certain new techniques have been used to ...

Visualization of SARS-CoV-2 using Immuno RNA-Fluorescence In Situ Hybridization

使用免疫RNA-荧光原位杂交对SARS-CoV-2进行可视化

This manuscript provides a protocol for in situ hybridization chain reaction (HCR) coupled with immunofluorescence to visualize severe acute respiratory ...