The authors have no acknowledgements.

The institutional animal care committee approved all experimental protocols and animals were treated following the &#8220;Guide for the Care and Use of Laboratory Animals&#8221; prepared by the Institute of Laboratory Animal Resources, National Research Council, 1996. Male Yorkshire pigs (40&#8211;50 kg) were used to perform the orthotopic heart transplants (animal size can vary according to the investigators discretion and experimental goals).
1. Donor Procedure
<ol>
	<li>Anesthetic inducti...

<ol>
	<li>Lund, L. H., Edwards, L. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Registry+of+the+International+Society+for+Heart+and+Lung+Transplantation:+Thirty-third+Adult+Heart+Transplantation+Report-2016;+Focus+Theme:+Primary+Diagnostic+Indications+for+Transplant.">The Registry of the International Society for Heart and Lung Transplantation: Thirty-third Adult Heart Transplantation Report-2016; Focus Theme: Primary Diagnostic Indications for Transplant.</a> The Journal of Heart and Lung Transplantation. 35 (10), 1158-1169 (2016).</li><li>Lund, L. H., Edwards, L. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Registry+of+the+International+Society+for+Heart+and+Lung+Transplantation_+Thirty-first+Official+Adult+Heart+Transplant+Report-2014;+Focus+Theme_+Retransplantation.">The Registry of the International Society for Heart and Lung Transplantation_ Thirty-first Official Adult Heart Transplant Report-2014; Focus Theme_ Retransplantation.</a> The. Journal of Heart and Lung Transplantation. 33 (10), 996-1008 (2014).</li><li>Cos&#237;o Carmena, M. D. G., G&#243;mez Bueno, 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=Primary+graft+failure+after+heart+transplantation:+characteristics+in+a+contemporary+cohort+and+performance+of+the+RADIAL+risk+score.">Primary graft failure after heart transplantation: characteristics in a contemporary cohort and performance of the RADIAL risk score.</a> The Journal of Heart and Lung Transplantation. 32 (12), 1187-1195 (2013).</li><li>Fedak, P. W. 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(102), e52907 (2015).</li><li>Fukunaga, N., Bissoondath, V., Rao, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Submandibular+Gland-preserving+Technique+for+Heterotopic+Cervical+Heart+Transplantation+in+Mice.">Submandibular Gland-preserving Technique for Heterotopic Cervical Heart Transplantation in Mice.</a> Transplantation. 1, (2018).</li><li>Gong, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+Heterotopic+Abdominal+Heart+Transplant+Model.">Mouse Heterotopic Abdominal Heart Transplant Model.</a> Rodent Transplant Medicine. , 107-118 (2014).</li><li>Robinson, N., Souslian, L., Gallegos, R. P., Rivard, A. L., Dalmasso, A. P., Bianco, R. 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This manuscript describes a large-animal pre-clinical model of orthotopic heart transplantation. Various small animal models of heterotopic heart transplantation have been successfully used to study the effects of novel treatments to improve organ preservation and decrease ischemia-reperfusion injury11,12,13. Additionally, small animal models permit survival models and long-term follow-up to investigate the development of reject...

Fifty-years following the first successful report, cardiac transplantation remains the gold-standard treatment for eligible patients with advanced heart failure1. Although ischemic times of up to four hours are tolerated adequately, an ischemic time of greater than six hours is associated with inferior outcomes2. Primary graft dysfunction remains the principal cause of early morbidity and mortality following transplantation2,3. The causes of primary graft dysfunction are multifactorial and include the use of marginal organs, recipient pulmonary vascular disease, hyperacute rejection, and ischemia-reperfusion injury sustained at the time of transplantation3.
Multiple studies have investigated novel methods for donor heart preservation to reduce the incidence of primary graft dysfunction4,5,6,7. It is common practice to assess new techniques and treatments in murine models of ischemia-reperfusion injury or heterotopic heart transplantation. Additionally, small animal models permit survival models and long-term follow-up to investigate the development of rejection and cardiac allograft vasculopathy11,12,13. However, most of these strategies fail initial clinical pilot trials or never reach this stage. It is of paramount importance to evaluate new therapies in a clinically relevant large-animal model for efficient and reliable translation of basic studies&#39;&#160;findings.
The porcine heart is often considered the most anatomically similar to the human heart when using large-animal models. As such, it is an ideal platform to perform cardiac surgical research. However, there are several important factors to consider when using a porcine model. First, the tissue is typically described as fragile and friable, especially in the right atrium and the pulmonary artery, being prone to tears14. Additionally, the pig heart is considered sensitive to manipulation and prone to arrhythmias, which is why one should routinely administer an anti-arrythmetic to each animal at the beginning of the experiment. An important anatomical difference between the porcine model and clinical heart transplantation is the left hemiazygous vein in the swine which drains directly into the coronary sinus. This has to be ligated during the recipient procedure to avoid continuous bleeding. Finally, the porcine model is very sensitive to ischemia, but it is still appropriate for acute studies in heart transplantation15.
This manuscript describes a pre-clinical large-animal (porcine) model of orthotopic heart transplantation that has been firmly established and utilized to investigate novel cardioprotective strategies5,6,8,9.

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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.

This pre-clinical model has been used successfully since 19945,6,8,9. Table 1 demonstrates representative results from pressure-volume relationships and echocardiographic parameters taken at baseline, and 3 h post-transplantation in a set of 5 experiments. Although we see a decrease in myocadial contractility following transplantation, this was not statistically significant.</p...

Watch this Scientific Journal Video about A Pre-Clinical Porcine Model of Orthotopic Heart Transplantation at JoVE.com

A Pre-Clinical Porcine Model of Orthotopic Heart Transplantation

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.

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

It is important to evaluate new heart preservation strategies using clinically relevant large-animal model for a reliable clinical translation. The porcine heart model for research in cardiac surgery is a very useful one because human and porcine hearts share lots of similarities in terms of anatomy, physiology, even size, so it's an almost ideal platform for research. After opening the pericardium of an anesthetized donor pig, have an assistant carefully retract the right ventricular outflow tract inferiorly and the pulmonary artery to the left and carefully retract the aorta to the right to avoid direct lesions to the pulmonary artery.Use the cautery to dissect the aortal pulmonary space and clear the anterior aspect of the ascending aorta from the connective tissue. After carefully retracting the right ventricular outflow tract use a 4.0 PROLENE suture to place a purse string stitch on the proximal ascending aorta adventitia and secure the suture with a tourniquet. Insert the cardioplegia delivery cannula into the ascending aorta within the purse string suture and tighten the tourniquet to secure the cannula.Next open the left inferior pulmonary vein and the superior and inferior vena cavae to ensure adequate cardiac venting and place the aortic cross-clamp on the distal ascending aorta. Initiate cardioplegia infusion targeting an aortic root pressure of 80 to 100 millimeters of mercury. Apply a 0.9%sodium chloride ice slush to the thoracic cavity and over the organ.When the cardioplegia infusion is finished, proceed with the cardiectomy in a traditional manner, sectioning the aorta after the innominate artery and the pulmonary artery and at the bifurcation to ensure a sufficient length for the implant. To initiate a cardiopulmonary bypass in the recipient, first use Metzenbaum and right angle forceps to dissect between the superior vena cava and the innominate artery and the inferior vena cava and the pericardium of the anesthetized recipient animal. Individually encircle the superior and inferior vena cavae with a 0-silk tie and secure each tape with a tourniquet.Use a 4-0 PROLENE suture to place two concentric purse string sutures on the distal ascending aorta adventitia and place purse string sutures on the inferior and superior vena cavae at the level of the pericardial reflection. Secure each suture with a tourniquet and use the Seldinger technique to insert a 17-21 French arterial cannula into the aorta. Use a 3/8-3/8 connector to connect the cannula to the arterial line of the bypass circuit.Make a five-millimeter incision at the center of the purse string sutures placed on the vena cavae and dilate the incision with a small angled instrument. Using a 24-28 French right-angled, single-stage venous cannula, cannulate the superior and inferior vena cavae, directing the angle superiorly in the superior vena cava and inferiorly into the inferior vena cava, away from the heart. Tighten the tourniquet holding the purse string suture to secure the cannula, and use a 3/8-3/8 1/2-Y connector to connect the cannulas to the venous line of the bypass circuit.Then initiate the cardiopulmonary bypass. After cardiopulmonary bypass initiation, use a sharp dissection instrument and a right angle to encircle the left hemiazygos vein and ligate the vessel distally with an O-silk tie. Cross-clamp the recipient aorta proximally to the arterial cannula and snare both vena cavae with the previously placed O-silk ties.Compare the left atrial cuff sizes of the donor and recipient and modify each as necessary. Deliver the first cardioplegia dose to the donor heart using the previously placed cardioplegia cannula. Next use the standard biatrial anastomotic technique and an SH needle to place one 4-0 PROLENE suture at the junction between the left atrium and the right inferior margin of the inferior vena cava.Place another suture at 180 degrees from the first suture to connect the donor and recipient cuffs, then complete the posterior and anterior wall anastomoses from the superior suture to the inferior suture. For the right atrium, open the donor right atrium from the appendage towards the inferior vena cava, creating a donor cuff that matches the recipient cuff size, then, starting at the junction between the inferior vena cava and the right atrium, complete the interior wall anastomosis and then the lateral wall. For the pulmonary artery, trim the edges of both the recipient and donor pulmonary arteries to create matching sizes.Use a BB needle to place a 5-0 PROLENE suture at the left lateral wall, uniting the donor and recipient vessels, and another at the right lateral edge. Then complete the inferior wall anastomosis and the anterior wall anastomosis. For the aorta, trim the edges as just demonstrated and place a suture on the left lateral wall connecting the donor and recipient vessels.Then complete the inferior wall and the anterior wall anastomosis before initiating a 60-minute reperfusion on the cardiopulmonary bypass. For a functional assessment of the graft, place a 0-silk tie around the inferior vena cava and insert a pressure-volume conductance catheter into the left ventricle through a small apical ventriculotomy to permit continuous measurements of the left ventricular pressure-volume relations. Here representative results from pressure-volume relationships and echocardiographic parameters taken at baseline, and at three hours post-transplantation in a set of five experiments are shown.During steady-state assessments, volume-dependent parameters are recorded such as the maximum and minimum rate of developed pressure. Volume-independent parameters are obtained by intermittent occlusion of the inferior vena cava during which the volume of the left ventricle progressively decreases and the end-systolic and end-diastolic pressure-volume relationships are recorded. The pre-load recruitable stroke work data can be used to determine the relationship between the stroke work and the corresponding end-diastolic volume.Various other metabolic parameters can also be measured with this model. This procedure requires at least one trained surgeon and approximately five practice experiments to optimize the research protocol for each research group. The porcine heart is fragile, friable, and especially sensitive to manipulation and ischemia, thus being prone to tears.It is important to revise all anastomoses to ensure minimal to no bleeding. This protocol can help in the development of new myocardial preservation strategies to decrease ischemia-perfusion injury and to prevent primary graft failure.

a-pre-clinical-porcine-model-of-orthotopic-heart-transplantation

Research

JoVE Journal

Medicine

9.4K visualizações.  University Health Network. É importante avaliar novas estratégias de preservação do coração utilizando um modelo de animais de grande porte clinicamente relevante para uma tradução clínica confiável. O modelo de coração suíno para pesquisa em cirurgia cardíaca é muito útil porque os corações humanos e suínos compartilham muitas semelhanças em termos de anatomia, fisiologia, até mesmo tamanho, por isso é uma plataforma quase ideal para a pesquisa. Após abrir o pericárdio de um porco doador aneste...