Name: A Porcine Heterotopic Heart Transplantation Protocol for Delivery of Therapeutics to a Cardiac Allograft (Video) | JoVE Journal
Uploaded: 2022-02-14T14:30:00.000+00:00
Duration: 8 min 30 s
Description: Learn a porcine heterotopic heart transplantation protocol for delivering therapeutics to cardiac allografts using normothermic ex vivo perfusion. This method enables extended testing of interventions to prevent rejection and improve transplant outcomes.

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We would like to thank Duke Large Animal Surgical Core and Duke Perfusion Services for their assistance during these procedures. We would also like to thank Paul Lezberg and TransMedics, Inc. for support.

<ol><li>Virani, S. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Heart+disease+and+stroke+statistics-2021+update%3A+A+report+from+the+American+Heart+Association%5BTitle%5D)+AND+%22Circulation%22%5BJournal%5D)">Heart disease and stroke statistics-2021 update: A report from the American Heart Association.</a> Circulation. 143 (8), 254(2021).</li>
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<li>Kadner, A., Chen, R. H., Adams, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Heterotopic+heart+transplantation%3A+experimental+development+and+clinical+experience%5BTitle%5D)+AND+%22European+Journal+of+Cardiothorac+Surgery%22%5BJournal%5D)">Heterotopic heart transplantation: experimental development and clinical experience.</a> European Journal of Cardiothorac Surgery. 17 (4), 474-481 (2000).</li>
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<li>Sahoo, S., Kariya, T., Ishikawa, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Targeted+delivery+of+therapeutic+agents+to+the+heart%5BTitle%5D)+AND+%22Nature+Reviews.+Cardiology%22%5BJournal%5D)">Targeted delivery of therapeutic agents to the heart.</a> Nature Reviews. Cardiology. 18 (6), 389-399 (2021).</li>
<li>Stamp, N. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Successful+heart+transplant+after+ten+hours+out-of-body+time+using+the+TransMedics+Organ+Care+System%5BTitle%5D)+AND+%22Heart%2C+Lung+%26+Circulation%22%5BJournal%5D)">Successful heart transplant after ten hours out-of-body time using the TransMedics Organ Care System.</a> Heart, Lung & Circulation. 24 (6), 611-613 (2015).</li>
<li>Ragalie, W. S., Ardehali, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Current+status+of+normothermic+ex-vivo+perfusion+of+cardiac+allografts%5BTitle%5D)+AND+%22Current+Opinion+in+Organ+Transplantation%22%5BJournal%5D)">Current status of normothermic ex-vivo perfusion of cardiac allografts.</a> Current Opinion in Organ Transplantation. 25 (3), 237-240 (2020).</li>
<li>Koerner, M. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Normothermic+ex+vivo+allograft+blood+perfusion+in+clinical+heart+transplantation%5BTitle%5D)+AND+%22Heart+Surgery+Forum%22%5BJournal%5D)">Normothermic ex vivo allograft blood perfusion in clinical heart transplantation.</a> Heart Surgery Forum. 17 (3), 141-145 (2014).</li>
<li>Rosenbaum, D. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Perfusion+preservation+versus+static+preservation+for+cardiac+transplantation%3A+effects+on+myocardial+function+and+metabolism%5BTitle%5D)+AND+%22Journal+of+Heart+and+Lung+Transplantation%22%5BJournal%5D)">Perfusion preservation versus static preservation for cardiac transplantation: effects on myocardial function and metabolism.</a> Journal of Heart and Lung Transplantation. 27 (1), 93-99 (2008).</li>
<li>Cullen, P. P., Tsui, S. S., Caplice, N. M., Hinchion, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+state-of-the-art+review+of+the+current+role+of+cardioprotective+techniques+in+cardiac+transplantation%5BTitle%5D)+AND+%22Interactive+Cardiovascular+and+Thoracic+Surgery%22%5BJournal%5D)">A state-of-the-art review of the current role of cardioprotective techniques in cardiac transplantation.</a> Interactive Cardiovascular and Thoracic Surgery. 32 (5), 683-694 (2021).</li>
<li>Rurik, J. G., Aghajanian, H., Epstein, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Immune+cells+and+immunotherapy+for+cardiac+injury+and+repair%5BTitle%5D)+AND+%22Circulation+Research%22%5BJournal%5D)">Immune cells and immunotherapy for cardiac injury and repair.</a> Circulation Research. 128 (11), 1766-1779 (2021).</li>
<li>Rincon, M. Y., VandenDriessche, T., Chuah, M. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Gene+therapy+for+cardiovascular+disease%3A+advances+in+vector+development%2C+targeting%2C+and+delivery+for+clinical+translation%5BTitle%5D)+AND+%22Cardiovascular+Research%22%5BJournal%5D)">Gene therapy for cardiovascular disease: advances in vector development, targeting, and delivery for clinical translation.</a> Cardiovascular Research. 108 (1), 4-20 (2015).</li>
<li>Kieserman, J. M., Myers, V. D., Dubey, P., Cheung, J. Y., Feldman, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Current+landscape+of+heart+failure+gene+therapy%5BTitle%5D)+AND+%22Journal+of+the+American+Heart+Association%22%5BJournal%5D)">Current landscape of heart failure gene therapy.</a> Journal of the American Heart Association. 8 (10), 012239(2019).</li>
<li>Perin, E. C. <a target="_blank" href="http://scholar.google.com/scholar?hl=en&safe=off&q=author%3aPerin%2C+EC+%22Stem+cell+and+gene+therapy+for+cardiovascular+disease%22">Stem cell and gene therapy for cardiovascular disease</a>. Perin, E. C., Miller, L. W., Taylor, D. A., Wilkerson, J. T. , Academic Press. 279-287 (2016).</li>
<li>Bishawi, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+normothermic+ex+vivo+organ+perfusion+delivery+method+for+cardiac+transplantation+gene+therapy%5BTitle%5D)+AND+%22Scientific+Reports%22%5BJournal%5D)">A normothermic ex vivo organ perfusion delivery method for cardiac transplantation gene therapy.</a> Scientific Reports. 9 (1), 8029(2019).</li>
<li>Hulot, J. S., Ishikawa, K., Hajjar, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Gene+therapy+for+the+treatment+of+heart+failure%3A+promise+postponed%5BTitle%5D)+AND+%22European+Heart+Journal%22%5BJournal%5D)">Gene therapy for the treatment of heart failure: promise postponed.</a> European Heart Journal. 37 (21), 1651-1658 (2016).</li>
<li>Ho, C. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Molecular+characterization+of+swine+leucocyte+antigen+class+I+genes+in+outbred+pig+populations%5BTitle%5D)+AND+%22Animal+Genetics%22%5BJournal%5D)">Molecular characterization of swine leucocyte antigen class I genes in outbred pig populations.</a> Animal Genetics. 40 (4), 468-478 (2009).</li>
<li>Ho, C. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Molecular+characterization+of+swine+leucocyte+antigen+class+II+genes+in+outbred+pig+populations%5BTitle%5D)+AND+%22Animal+Genetics%22%5BJournal%5D)">Molecular characterization of swine leucocyte antigen class II genes in outbred pig populations.</a> Animal Genetics. 41 (4), 428-432 (2010).</li>
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Paul Lezberg is employed by TransMedics, Inc. Carmelo Milano received a financial gift from TransMedics, Inc. to fund heterotopic heart transplant surgeries. Michelle Mendiola Pla is supported by T32HL007101. The other authors have no conflicts of interest to declare.

Delivery of therapeutics during ex vivo perfusion in cardiac transplantation offers a strategy to modify the allograft and potentially improve transplant outcomes. The protocol presented here incorporates the state-of-the-art normothermic ex vivo sanguinous perfusion storage and offers promising potential to test isolated delivery of cell, gene, or immunotherapies to the allograft11,12,13. To date, cardiac deli...

Heart failure is a condition that affects an estimated 6 million adults in the United States and is projected to increase to 8 million adults by the year 20301. Cardiac transplantation is the gold standard treatment for end stage heart failure. However, it is not without its limitations and complications. It remains limited by the number of available donor hearts, primary graft dysfunction, rejection of the heart, and the side effects of long-term immunosuppression2. These limitations are particularly important in young recipients who may experience allograft failure and require subsequent re-transplantation to achieve normal life expectancy.
An ideal intervention to overcome these limitations would treat entire cardiac allografts with therapeutics prior to implantation into the recipient that can improve the viability of the allograft and confer &#34;cardioprotection.&#34; Such interventions would be given prophylactically to minimize the incidence of ischemic insults, allograft rejection, cardiac allograft vasculopathy, and even repair marginal allografts. Translational studies for developing these types of interventions require a large-animal model of cardiac transplantation to allow for the long-term surveillance of the cardiac graft. The porcine, heterotopic heart transplantation model in the intraabdominal position has proven ideal for this purpose. Heart transplantation in this position allows for testing the effects of novel therapies and assessing the immunopathology of graft rejection. Additionally, the heterotopic model is advantageous over the orthotopic model due to better overall survival of the recipient, no requirement for cardiopulmonary bypass, and no requirement of the graft to maintain the recipient&#39;s circulation3.
Effective delivery of therapeutic interventions to the heart, such as gene, cell, or immuno- therapy, is a significant barrier to clinical application4,5. The technology introduced by ex vivo perfusion devices allows grafts to be continually perfused, maintaining them in a nonworking but metabolically active state6,7,8,9. This offers a unique opportunity to treat a whole heart with advanced therapeutics while minimizing the potential side effects of systemic delivery10,11,12,13. Another advantage of utilizing ex vivo perfusion devices for therapeutic delivery is that they allow the administration of medications to the coronary circulation over extended periods that are not feasible using traditional cold static storage methods. This allows for more global delivery of the therapeutics to the graft14. Using the protocol presented here, we successfully delivered the firefly luciferase gene to a whole porcine cardiac graft using adenoviral vectors15. The aim of this protocol is to provide a reproducible and robust approach for achieving delivery of a therapeutic to the entire cardiac allograft prior to transplantation.

<table><tbody><tr><td>0 Looped Maxon suture</td><td>Covidien</td><td>GMM-341L</td><td>Used to close fascia of the laparotomy incision</td></tr><tr><td>0 Silk ties</td><td>Medtronic, Inc</td><td>S346</td><td></td></tr><tr><td>18 G Angiocath</td><td>BD</td><td>381144</td><td>Used to de-air the left ventricle of the donor heart after implantation</td></tr><tr><td>20 Fr LV vent</td><td>Medtronic, Inc</td><td>12002</td><td></td></tr><tr><td>2-0 Silk sutures</td><td>Ethicon, Inc.</td><td>SA11G</td><td></td></tr><tr><td>2-0 Silk ties</td><td>Ethicon, Inc.</td><td>SA65H</td><td></td></tr><tr><td>2-0 Vicryl suture</td><td>Ethicon, Inc.</td><td>J259H</td><td></td></tr><tr><td>24 Fr venous cannula</td><td>Medtronic, Inc</td><td>68124</td><td></td></tr><tr><td>3-0 Prolene sutures</td><td>Ethicon, Inc.</td><td>8522</td><td></td></tr><tr><td>4-0 Monocryl suture</td><td>Ethicon, Inc.</td><td>Y469G</td><td></td></tr><tr><td>4-0 Prolene sutures</td><td>Ethicon, Inc.</td><td>8521</td><td></td></tr><tr><td>Animal hair cutting clipper</td><td>Wahl</td><td>8786-452</td><td></td></tr><tr><td>Aortic clamp</td><td>V. Mueller</td><td>CH6201</td><td></td></tr><tr><td>Army Navy retractor</td><td>V. Mueller</td><td>SU3660</td><td></td></tr><tr><td>ATF 40, Cell saver disposable set</td><td>Fresenius Kabi</td><td>9108494</td><td>Cell saver device insert</td></tr><tr><td>Balfour retractor</td><td>V. Mueller</td><td>SU3042</td><td>Used as an abdominal wall retractor</td></tr><tr><td>C.A.T.S cell saver</td><td>Fresenius Kabi</td><td>ES0019</td><td>Cell saver device used to wash donor blood</td></tr><tr><td>Cardiac defibrillator</td><td>Zoll</td><td>M Series</td><td>Cardiac defibrillator</td></tr><tr><td>Castro needle holder</td><td>V. Mueller</td><td>CH8589</td><td></td></tr><tr><td>CG4 iStat cartridges</td><td>Abbott</td><td>03P85-25</td><td>POC testing</td></tr><tr><td>CG8 iStat cartridges</td><td>Abbott</td><td>03P88-25</td><td>POC testing</td></tr><tr><td>DeBakey forceps</td><td>V. Mueller</td><td>CH5902</td><td></td></tr><tr><td>Electrocautery disposable pencil</td><td>Covidien</td><td>E2450H</td><td></td></tr><tr><td>Gerald forceps</td><td>V. Mueller</td><td>NL1451</td><td></td></tr><tr><td>Hemotherm 400CE Dual Reservoir Cooler/Heater</td><td>Cincinnati Sub-Zero</td><td>86022</td><td>Heater cooler used to regulate perfusion temperature on the ex vivo perfusion device</td></tr><tr><td>iSTAT 1</td><td>Abbott</td><td>04P75-03</td><td>POC testing device</td></tr><tr><td>Kocher clamp</td><td>V. Mueller</td><td>SU2790</td><td></td></tr><tr><td>Large clip applier</td><td>Sklar</td><td>50-4300</td><td></td></tr><tr><td>Large clips</td><td>Teleflex</td><td>4200</td><td></td></tr><tr><td>Large soft pledgets</td><td>Covidien</td><td>8886867901</td><td></td></tr><tr><td>Medium clip applier</td><td>Sklar</td><td>50-4335</td><td></td></tr><tr><td>Medium clips</td><td>Teleflex</td><td>2200</td><td></td></tr><tr><td>Metzenbaum scissor</td><td>V. Mueller</td><td>CH2006-001</td><td></td></tr><tr><td>No. 10 scalpel blade</td><td>Swann-Mortan</td><td>301</td><td>Used for skin incision</td></tr><tr><td>No. 11 scalpel blade</td><td>Kiato Plus</td><td>18111</td><td>Used for vascular incision</td></tr><tr><td>OCS device with base</td><td>TransMedics, Inc.</td><td></td><td>Ex vivo perfusion device</td></tr><tr><td>OCS disposable</td><td>TransMedics, Inc.</td><td></td><td>Ex vivo perfusion device insert with perfusion kits</td></tr><tr><td>Pacing cable</td><td>Remington Medical</td><td>FL-601-97</td><td></td></tr><tr><td>Pediatric cardioplegia catheter (4Fr)</td><td>Medtronic, Inc</td><td>10218</td><td>Used to deliver cardioplegia to the donor aortic root</td></tr><tr><td>Pediatric Foley catheter</td><td>Teleflex</td><td>RSH170003080</td><td>Placed pre-op to decompress the recipient&#039;s bladder</td></tr><tr><td>Potts scissors</td><td>V. Mueller</td><td>CH13038</td><td></td></tr><tr><td>Pressure bag x2 (1,000 mL)</td><td>Novaplus</td><td>V4010H</td><td>Used to deliver cardioplegia at a set pressure</td></tr><tr><td>Satinsky clamp</td><td>V. Mueller</td><td>CH7305</td><td>Vascular clamp used for creating anastomoses between donor heart and recipient vessels</td></tr><tr><td>Scissors</td><td>Felco</td><td>FELCO 200A-50</td><td>Used to perform sternotomy</td></tr><tr><td>Small hard pledgets</td><td>Covidien</td><td>8886867701</td><td></td></tr><tr><td>Sternal retractor</td><td>V. Mueller</td><td>CH6950-007</td><td></td></tr><tr><td>Temporary cardiac pacing wires</td><td>Ethicon, Inc.</td><td>TPW32</td><td></td></tr><tr><td>Temporary dual chamber pacemaker</td><td>Medtronic, Inc</td><td>5388</td><td>Cardiac pacing device</td></tr><tr><td>Tourniquet kit</td><td>Medtronic, Inc</td><td>79005</td><td>Rummel tourniquets</td></tr><tr><td>Umbilical tape</td><td>Covidien</td><td>8886861903</td><td></td></tr><tr><td>Vessel loops</td><td>Covidien</td><td>31145686</td><td></td></tr></tbody></table>

a porcine heterotopic heart transplantation protocol for delivery of therapeutics to a cardiac allograft

A Porcine Heterotopic Heart Transplantation Protocol for Delivery of Therapeutics to a Cardiac Allograft

Cardiac transplantation is the gold standard treatment for end-stage heart failure. However, it remains limited by the number of available donor hearts and complications such as primary graft dysfunction and graft rejection. The recent clinical use of an ex vivo perfusion device in cardiac transplantation introduces a unique opportunity for treating cardiac allografts with therapeutic interventions to improve function and avoid deleterious recipient responses. Establishing a translational, large-animal model for therapeutic delivery to the entire allograft is essential for testing novel therapeutic approaches in cardiac transplantation. The porcine, heterotopic heart transplantation model in the intraabdominal position serves as an excellent model for assessing the effects of novel interventions and the immunopathology of graft rejection. This model additionally offers long-term survival for the pig, given that the graft is not required to maintain the recipient's circulation. The aim of this protocol is to provide a reproducible and robust approach for achieving ex vivo delivery of a therapeutic to the entire cardiac allograft prior to transplantation and provide technical details to perform a survival heterotopic transplant of the ex vivo perfused heart.

This group has successfully survived 9 pigs between 5 and 35 days following the protocol as presented here, depending on the study design. Out of 10 pigs that have undergone this protocol, only 1 died prematurely from surgical complications, yielding a 90% survival rate. Demonstrated in Figure 2 is a diagram of the configuration of a heterotopic heart transplanted in the intraabdominal position in a pig. When determining the site for anastomosis of the allograft, select a site that minimizes...

We present a protocol for utilizing a normothermic ex vivo sanguinous perfusion system for the delivery of therapeutics to an entire cardiac allograft in a porcine heterotopic heart transplant model.

Cardiac transplantation is the gold standard treatment for end-stage heart failure. However, it remains limited by the number of available donor hearts ...

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Learn a porcine heterotopic heart transplantation protocol for delivering therapeutics to cardiac allografts using normothermic ex vivo perfusion. This method enables extended testing of interventions to prevent rejection and improve transplant outcomes.

A Porcine Heterotopic Heart Transplantation Protocol for Delivery of Therapeutics to a Cardiac Allograft

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