Name: a simplified model for heterotopic heart valve transplantation in rodents
Uploaded: 2021-09-21T14:30:00
Description: Watch this Scientific Journal Video about A Simplified Model for Heterotopic Heart Valve Transplantation in Rodents at JoVE.com

Figure 1 was created with biorender.com. This work was&#160;supported in part&#160;by the AATS Foundation Surgical Investigator Program to TKR, the Children&#39;s Excellence Fund held&#160;by the Department of Pediatrics at the Medical University of South Carolina to TKR, an Emerson Rose Heart Foundation grant to TKR, Philanthropy by Senator Paul Campbell to TKR, NIH-NHLBI Institutional Postdoctoral Training Grants (T32 HL-007260) to JHK and BG, and the Medical University of South Carolina College of Medicine Pre-clerkship FLEX Research Fund to MAH.

The study was approved by the Committee of Animal Research following the National Institutes of Health Guide for Care and Use of Laboratory Animals.
1. Information on the animal model (Rats)
<ol>
	<li>Use an operating microscope (see Table of Materials) with up to 20x magnification for all surgical procedures.</li>
	<li>Use syngeneic (such as Lewis-Lewis) or allogeneic (such as Lewis-Brown Norway) strains for the transplants as needed for the experiment.</li>
	<li>Use rats o...

<ol>
	<li>Van Der Linde, 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=Birth+prevalence+of+congenital+heart+disease+worldwide:+A+systematic+review+and+meta-analysis.">Birth prevalence of congenital heart disease worldwide: A systematic review and meta-analysis.</a> Journal of the American College of Cardiology. 58 (21), 2241-2247 (2011).</li><li>Jacobs, J. 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=Reoperations+for+pediatric+and+congenital+heart+disease:+An+analysis+of+the+Society+of+Thoracic+Surgeons+(STS)+congenital+heart+surgery+database.">Reoperations for pediatric and congenital heart disease: An analysis of the Society of Thoracic Surgeons (STS) congenital heart surgery database.</a> Seminars in Thoracic and Cardiovascular Surgery: Pediatric Cardiac Surgery Annual. 17 (1), 2-8 (2014).</li><li>Syedain, Z. 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=Pediatric+tri-tube+valved+conduits+made+from+fibroblast-produced+extracellular+matrix+evaluated+over+52+weeks+in+growing+lambs.">Pediatric tri-tube valved conduits made from fibroblast-produced extracellular matrix evaluated over 52 weeks in growing lambs.</a> Science Translational Medicine. 13 (585), 1-16 (2021).</li><li>Khan, M. S., Samayoa, A. X., Chen, D. W., Petit, C. J., Fraser, C. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contemporary+experience+with+surgical+treatment+of+aortic+valve+disease+in+children.">Contemporary experience with surgical treatment of aortic valve disease in children.</a> Journal of Thoracic and Cardiovascular Surgery. 146 (3), 512-521 (2013).</li><li>Boyd, R., Parisi, F., Kalfa, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=State+of+the+art:+Tissue+engineering+in+congenital+heart+surgery.">State of the art: Tissue engineering in congenital heart surgery.</a> Seminars in Thoracic and Cardiovascular Surgery. 31 (4), 807-817 (2019).</li><li>Feins, E. N., Emani, 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=Expandable+valves,+annuloplasty+rings,+shunts,+and+bands+for+growing+children.">Expandable valves, annuloplasty rings, shunts, and bands for growing children.</a> Seminars in Thoracic and Cardiovascular Surgery: Pediatric Cardiac Surgery Annual. 23, 17-23 (2020).</li><li>Lintas, V., 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=TCT-795+Human+cell+derived+off-the-shelf+tissue+engineered+heart+valves+for+next+generation+transcatheter+aortic+valve+replacement:+a+proof-of-concept+study+in+adult+sheep.">TCT-795 Human cell derived off-the-shelf tissue engineered heart valves for next generation transcatheter aortic valve replacement: a proof-of-concept study in adult sheep.</a> Journal of the American College of Cardiology. 70 (18), 271 (2017).</li><li>Blum, K. M., Drews, J. D., Breuer, C. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tissue-engineered+heart+valves:+A+call+for+mechanistic+studies.">Tissue-engineered heart valves: A call for mechanistic studies.</a> Tissue Engineering Part B: Reviews. 24 (3), 240-253 (2018).</li><li>Bernstein, 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=Cardiac+growth+after+pediatric+heart+transplantation.">Cardiac growth after pediatric heart transplantation.</a> Circulation. 85 (4), 1433-1439 (1992).</li><li>Delmo Walter, E. 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=Adaptive+growth+and+remodeling+of+transplanted+hearts+in+children.">Adaptive growth and remodeling of transplanted hearts in children.</a> Europeon Journal of Cardiothoracic Surgery. 40 (6), 1374-1383 (2011).</li><li>Simon, 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=Growth+of+the+pulmonary+autograft+after+the+Ross+operation+in+childhood.">Growth of the pulmonary autograft after the Ross operation in childhood.</a> Europeon Journal of Cardiothoracic Surgery. 19 (2), 118-121 (2001).</li><li>Oei, F. B. 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=A+size-matching+heterotopic+aortic+valve+implantation+model+in+the+rat.">A size-matching heterotopic aortic valve implantation model in the rat.</a> Journal of Surgical Research. 87 (2), 239-244 (1999).</li><li>Oei, F. B. 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=Heart+valve+dysfunction+resulting+from+cellular+rejection+in+a+novel+heterotopic+transplantation+rat+model.">Heart valve dysfunction resulting from cellular rejection in a novel heterotopic transplantation rat model.</a> Transplant International. 13, (2000).</li><li>El Khatib, H., Lupinetti, F. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antigenicity+of+fresh+and+cryopreserved+rat+valve+allografts.">Antigenicity of fresh and cryopreserved rat valve allografts.</a> Transplantation. 49 (4), 765-767 (1990).</li><li>Yankah, A. C., Wottge, H. U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Allograft+conduit+wall+calcification+in+a+model+of+chronic+arterial+graft+rejection.">Allograft conduit wall calcification in a model of chronic arterial graft rejection.</a> Journal of Cardiac Surgery. 12 (2), 86-92 (1997).</li><li>Moustapha, 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=Aortic+valve+grafts+in+the+rat:+Evidence+for+rejection.">Aortic valve grafts in the rat: Evidence for rejection.</a> Journal of Thoracic and Cardiovascualr Surgery. 114 (6), 891-902 (1997).</li><li>L&#233;gar&#233;, J. F., 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=Prevention+of+allograft+heart+valve+failure+in+a+rat+model.">Prevention of allograft heart valve failure in a rat model.</a> Journal of Thoracic and Cardiovascualr Surgery. 122 (2), 310-317 (2001).</li><li>Legare, J. F., Lee, T. D. G., Creaser, K., Ross, D. B., Green, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=T+lymphocytes+mediate+leaflet+destruction+and+allograft+aortic+valve+failure+in+rats.">T lymphocytes mediate leaflet destruction and allograft aortic valve failure in rats.</a> The Annals of Thoracic Surgery. 70 (4), 1238-1245 (2000).</li><li>Niimi, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+technique+for+heterotopic+cardiac+transplantation+in+mice:+Experience+of+3000+operations+by+one+surgeon.">The technique for heterotopic cardiac transplantation in mice: Experience of 3000 operations by one surgeon.</a> Journal of Heart and Lung Transplantation. 20 (10), 1123-1128 (2001).</li><li>Burgin, 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=Kidney+Subcapsular+Allograft+Transplants+as+a+Model+to+Test+Virus-Derived+Chemokine-Modulating+Proteins+as+Therapeutics.">Kidney Subcapsular Allograft Transplants as a Model to Test Virus-Derived Chemokine-Modulating Proteins as Therapeutics.</a> Methods in molecular biology. 2225, 257-273 (2021).</li><li>Foglia, R. P., DiPreta, J., Donahoe, P. K., Statter, M. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fetal+allograft+survival+in+immunocompetent+recipients+is+age+dependent+and+organ+specific.">Fetal allograft survival in immunocompetent recipients is age dependent and organ specific.</a> Annals of Surgery. 204 (4), 402-410 (1986).</li><li>Cunha, G. R., Baskin, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+sub-renal+capsule+transplantation+in+developmental+biology.">Use of sub-renal capsule transplantation in developmental biology.</a> Differentiation. 91 (4-5), 4-9 (2016).</li><li>Hori, J., Joyce, N., Streilein, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epithelium-deficient+corneal+allografts+display+immune+privilege+beneath+the+kidney+capsule.">Epithelium-deficient corneal allografts display immune privilege beneath the kidney capsule.</a> Investigative Opthalmology &amp; Visual Science. 41 (2), 443-452 (2000).</li><li>Mandel, T., 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=transplantation+of+organ+cultured+fetal+pig+pancreas+in+non-obese+diabetic+(NOD)+mice+and+primates+(Macaca+fascicularis).">transplantation of organ cultured fetal pig pancreas in non-obese diabetic (NOD) mice and primates (Macaca fascicularis).</a> Xenotransplantation. 2 (3), 128-132 (1995).</li><li>Ricordi, C., Flye, M. W., Lacy, P. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Renal+subcapsular+transplantation+of+clusters+of+hepatocytes+in+conjunction+with+pancreatic+islets.">Renal subcapsular transplantation of clusters of hepatocytes in conjunction with pancreatic islets.</a> Transplantation. 45 (6), 1148-1150 (1988).</li><li>Shultz, L. 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=Subcapsular+transplantation+of+tissue+in+the+kidney.">Subcapsular transplantation of tissue in the kidney.</a> Cold Spring Harbor Protocols. 2014 (7), 737-740 (2014).</li><li>Vanden Berg, C. 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=Renal+subcapsular+transplantation+of+PSC-derived+kidney+organoids+induces+neo-vasculogenesis+and+significant+glomerular+and+tubular+maturation+in+vivo.">Renal subcapsular transplantation of PSC-derived kidney organoids induces neo-vasculogenesis and significant glomerular and tubular maturation in vivo.</a> Stem Cell Reports. 10 (3), 751-765 (2018).</li><li>Mitchell, R. N., Jonas, R. A., Schoen, F. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pathology+of+explanted+cryopreserved+allograft+heart+valves:+Comparison+with+aortic+valves+from+orthotopic+heart+transplants.">Pathology of explanted cryopreserved allograft heart valves: Comparison with aortic valves from orthotopic heart transplants.</a> Journal of Thoracic and Cardiovasular Surgery. 115 (1), 118-127 (1998).</li><li>Valante, 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=The+aortic+valve+after+heart+transplantation.">The aortic valve after heart transplantation.</a> Annals of Thoracic Surgery. 60, (1995).</li><li>O'Brien, M. F., Stafford, E. G., Gardner, M. A. H., Pohlner, P. G., McGiffin, D. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+comparison+of+aortic+valve+replacement+with+viable+cryopreserved+and+fresh+allograft+valves,+with+a+note+on+chromosomal+studies.">A comparison of aortic valve replacement with viable cryopreserved and fresh allograft valves, with a note on chromosomal studies.</a> Journal of Thoracic and Cardiovascular Surgery. 94 (6), 812-823 (1987).</li><li>Ng, T. F., Osawa, H., Hori, J., Young, M. J., Streilein, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Allogeneic+neonatal+neuronal+retina+grafts+display+partial+immune+privilege+in+the+subcapsular+space+of+the+kidney.">Allogeneic neonatal neuronal retina grafts display partial immune privilege in the subcapsular space of the kidney.</a> The Journal of Immunology. 169 (10), 5601-5606 (2002).</li><li>Heslop, B. F., Wilson, S. E., Hardy, B. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antigenicity+of+aortic+valve+allografts.">Antigenicity of aortic valve allografts.</a> Annals of Surgery. 177 (3), 301-306 (1973).</li><li>Steinmuller, D., Weiner, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evocation+and+persistence+of+transplantation+immunity+in+rats.">Evocation and persistence of transplantation immunity in rats.</a> Transplantation. 1 (1), 97-106 (1963).</li><li>Billingham, R. E., Brent, L., Brown, J. B., Medawar, P. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Time+of+onset+and+duration+of+transplantation+immunity.">Time of onset and duration of transplantation immunity.</a> Plastic and Reconstructive Surgery. 24 (1), 410-413 (1959).</li><li>Tector, A. J., Boyd, W. C., Korns, M. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Aortic+valve+allograft+rejection.">Aortic valve allograft rejection.</a> Journal of Thoracic and Cardiovascular Surgery. 62 (4), 592-601 (1971).</li><li>Sugimura, Y., Schmidt, A. K., Lichtenberg, A., Akhyari, P., Assmann, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+rat+model+for+the+in+vivo+assessment+of+biological+and+tissue-engineered+valvular+and+vascular+grafts.">A rat model for the in vivo assessment of biological and tissue-engineered valvular and vascular grafts.</a> Tissue Engineering Methods (Part C). 23 (12), 982-994 (2017).</li></ol>

Importance and potential applications 
While mechanical and bioprosthetic heart valves are routinely used in adult patients requiring valve replacement, these valves lack the potential to grow and, therefore, are suboptimal for pediatric patients. Heart valve transplantation is an experimental operation designed to deliver growing heart valve replacements for neonates and infants with congenital heart disease. However, unlike the transplant immunobiology of conventional heart transplants, the transp...

Congenital heart defects are the most common congenital disability in humans, affecting 7 in 1,000 live-born children each year1. Unlike adult patients in which various mechanical and bioprosthetic valves are routinely implanted, pediatric patients currently have no good options for valve replacement. These conventional implants do not have the potential to grow in recipient children. As a result, morbid re-operations are required to exchange the heart valve implants for successively larger versions as the children grow, with affected kids often requiring up to five or more open-heart surgeries in their lifetime2,3. Studies have shown that freedom from intervention or death is significantly poor for infants than older children, with 60% of infants with prosthetic heart valves facing re-operation or death within 3 years of their initial operation4. Therefore, there is an urgent need to deliver a heart valve that can grow and maintain function in pediatric patients.
For decades, attempts to deliver growing heart valve replacements have been centered on tissue engineering and stem cells. However, attempts to translate these valves to the clinic have been unsuccessful thus far5,6,7,8. For addressing this, a heart valve transplantation is proposed as a more creative operation for delivering growing heart valve replacements having the ability to self-repair and avoid thrombogenesis. Instead of transplanting the whole heart, only the heart valve is transplanted and will then grow with the recipient child, similar to conventional heart transplants or a Ross pulmonary autograph9,10,11. Post-operatively, recipient children will receive immunosuppression until the transplanted valve can be exchanged for an adult-sized mechanical prosthetic when the growth of the valve is no longer required. However, the transplant biology of heart valve transplant grafts remains unexplored. Therefore, animal models are needed to study this new type of transplant.
Several rat models have been previously described for heterotopic transplantation of the aortic valve into the abdominal aorta12,13,14,15,16,17,18. However, these models are prohibitively tricky, often requiring trained surgeons to operate successfully. Additionally, they are costly and time-consuming19. A novel rat model was developed to create a simpler animal model for studying the immunobiology of heart valve transplants. Single aortic valve leaflets are excised and inserted into the renal subcapsular space. The kidney is especially suited to study transplant rejection as it is highly vascularized with access to circulating immune cells20,21. While several others have utilized a renal subcapsular model to study the transplant biology of other allograft transplants such as pancreas, liver, kidney, and cornea22,23,24,25,26,27, this is the first description of transplantation of cardiac tissue in this position. Here, the transplantation technique is described, providing a significant step forward in studying the transplant immunology of heart valve transplantation.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>0.9% Sodium Chlordie, USP</td>
			<td>Baxter</td>
			<td>NDC 0338-0048-04</td>
			<td></td>
		</tr>
		<tr>
			<td>4-0 Polyglactin 910</td>
			<td>Ethicon</td>
			<td>J415H</td>
			<td></td>
		</tr>
		<tr>
			<td>7.5% Povidone-Iodine</td>
			<td>CareFusion</td>
			<td>29904-004</td>
			<td></td>
		</tr>
		<tr>
			<td>70% ETOH</td>
			<td>Fisher Scientific</td>
			<td>BP82031GAL</td>
			<td></td>
		</tr>
		<tr>
			<td>Anesthesia induction chamber</td>
			<td>Harvard Apparatus</td>
			<td>75-2030</td>
			<td>Air-tight inducton chamber for rats</td>
		</tr>
		<tr>
			<td>Anesthesia machine</td>
			<td>Harvard Apparatus</td>
			<td>75-0238</td>
			<td>Mobile Anesthesia System with Passive Scavenging</td>
		</tr>
		<tr>
			<td>Anesthesia Mask</td>
			<td>Harvard Apparatus</td>
			<td>59-8255</td>
			<td>Rat anesthesia mask</td>
		</tr>
		<tr>
			<td>Brown Norway Rats (BN/Crl)</td>
			<td>Charles River</td>
			<td>Strain Code 091</td>
			<td>Male, 5-7 weeks, 100-200 g</td>
		</tr>
		<tr>
			<td>Buprenorphine Hydrochloride, 0.3 mg/mL</td>
			<td>PAR Pharmaceutical</td>
			<td>NDC 42023-179-05</td>
			<td>0.03 mg/kg, administered subcutaneously</td>
		</tr>
		<tr>
			<td>Electric hair clippers</td>
			<td>WAHL</td>
			<td>79434</td>
			<td></td>
		</tr>
		<tr>
			<td>Electric Heating Pad</td>
			<td>Harvard Apparatus</td>
			<td>72-0492</td>
			<td>Maintained at 36-38 &#176;C</td>
		</tr>
		<tr>
			<td>Heparin</td>
			<td>Sagent Pharmaceuticals</td>
			<td>NDC 25021-400-10</td>
			<td>100U/100g injection into the left atrium</td>
		</tr>
		<tr>
			<td>Insulin Syringe, 1 mL</td>
			<td>Fisher Scientific</td>
			<td>14-841-33</td>
			<td></td>
		</tr>
		<tr>
			<td>Iris forceps curved</td>
			<td>World Precision Instruments</td>
			<td>15917</td>
			<td></td>
		</tr>
		<tr>
			<td>Iris forceps straight</td>
			<td>World Precision Instruments</td>
			<td>15916</td>
			<td></td>
		</tr>
		<tr>
			<td>Isoflurane, USP</td>
			<td>Piramal Critical Care</td>
			<td>NDC 66794-017-25</td>
			<td>Induced at 5% isoflurance in oxygen and maintained with 3.5% isoflurane in oxygen</td>
		</tr>
		<tr>
			<td>Lewis Rats (LEW/ Crl)</td>
			<td>Charles River</td>
			<td>Strain Code 004</td>
			<td>Male, 5-7 weeks, 100-200 g</td>
		</tr>
		<tr>
			<td>Micro forceps</td>
			<td>World Precision Instruments</td>
			<td>500233</td>
			<td>Dumont #5</td>
		</tr>
		<tr>
			<td>Micro scissors</td>
			<td>World Precision Instruments</td>
			<td>501930</td>
			<td>Spring-loaded Vannas Scissors</td>
		</tr>
		<tr>
			<td>Needle Driver</td>
			<td>World Precision Instruments</td>
			<td>500226</td>
			<td>Ryder Needle Driver</td>
		</tr>
		<tr>
			<td>Operating microscope</td>
			<td>AmScope</td>
			<td>SM-3BZ-80S</td>
			<td>3.5x - 90x Stereo Microscope</td>
		</tr>
		<tr>
			<td>Petri Dish</td>
			<td>Fisher Scientific</td>
			<td>FB0875714</td>
			<td></td>
		</tr>
		<tr>
			<td>Petrolatum ophthalmic ointment</td>
			<td>Dechra</td>
			<td>NDC 17033-211-38</td>
			<td></td>
		</tr>
		<tr>
			<td>Skin staples</td>
			<td>Ethicon</td>
			<td>PXR35</td>
			<td>Proximate 35</td>
		</tr>
		<tr>
			<td>Sterile cotton swabs</td>
			<td>Puritan</td>
			<td>25-806 1WC</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile gauze sponges</td>
			<td>Fisher Scientific</td>
			<td>22-037-902</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical Scissors</td>
			<td>World Precision Instruments</td>
			<td>1962C</td>
			<td>Metzenbaum Scissors</td>
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		<tr>
			<td>University of Wisconsin Buffer (Servator B)</td>
			<td>S.A.L.F S.p.A.</td>
			<td>6484A1</td>
			<td>Stored at 4 &#176;C</td>
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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.

A graphical depiction of the experimental design is provided for the rat model (Figure 1). Additionally, an aortic root dissected from the donor&#39;s heart and an individual aortic valve leaflet prepared for implantation is also shown in Figure 2. Next, a representative image of the position of the aortic valve leaflet under the renal capsule for implantation is shown in Figure 3A and after 3, 7, and 28 days within the recipient ra...

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A Simplified Model for Heterotopic Heart Valve Transplantation in Rodents

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.

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 ...

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. This technique is practical and more straightforward than previous methods for studying heart valve transplant immunobiology. After anesthetizing the rat, use the dissecting scissors to incise the skin from the xiphoid to the sternal notch.And perform a sternectomy by cutting the ribs on each side, lateral to the sternum until optimal access to the heart is achieved. Inject Heparin in the left atrium. Post sacrification, excise the thymus to improve the visualization of the great vessels.Then remove the heart on block with the ascending aorta till the level of the innominate artery. After placing the donor heart in a sterile Petri dish immediately following the cardiectomy, in an ice-cold, cold storage buffer, use the forceps and Vannas Spring Scissors to dissect the heart until only the aortic root remains with a one millimeter ventricular cuff proximal to the aortic valve. For opening the aortic valve, make a longitudinal cut to open the sinus of Valsalva between the left and noncoronary sinuses to visualize all three leaflets.To excise each aortic valve leaflet individually, use the blunt forceps to grasp the edge of the leaflet and use the Vannas Spring Scissors to excise the leaflet by cutting from one commissure down to the annulus then toward the next commissure. Store the excised leaflets in ice-cold storage buffer solution until they are ready to be implanted in the recipient rat, but implant all the leaflets within four hours of cold storage. Once the recipient rat is completely anesthetized, maintain the body temperature at 36 to 38 degrees celsius throughout the surgery with the heating pad.Next, place the rat in a right lateral recumbent position to access the left kidney. Administer Buprenorphine to recipient rat subcutaneously before surgery. And use the scissors to incise the skin over the flank longitudinally for one inch.Similarly, incise the underlying abdominal wall. Using the thumb and forefinger, apply a light pressure dorsally and ventrally while using the curved forceps. Lift the caudal pole of the kidney through the abdominal and skin incision.Externalize the cranial end of the kidney similarly. Alternatively, the kidney may be externalized by grasping the perirenal fat and pulling upward with light tension. Once externalized, keep the kidney moist with warm saline swabbed onto the kidney.In order to create a subcapsular pocket, begin by applying light pressure to the renal capsule using one set of blunt forceps so that the renal capsule can be clearly distinguished from the underlying parenchyma. Simultaneously using another set of blunt forceps, carefully grasp the capsule and gently pull upward to create a hole in the capsule. With blunt forceps, extend the incision until a space of about two millimeters has been created to accommodate the aortic valve leaflet.Develop a shallow subcapsular pocket that is slightly larger than the valve leaflet while lifting the edge of the incision with one set of forceps and advancing a blunt probe under the renal capsule. Initiate transplantation of the retrieved aortic valve by lifting the edge fibrous capsule advancing the aortic leaflet into the subcapsular pocket with blunt forceps. The incision and the renal capsule can be left open.Push the kidney gently back into its anatomical position using countertraction applied to the incision edges. Close the abdominal incision with a running sterile surgical suture and close the skin with staples. Following the operation, place the rat in a clean cage on a heating pad with access to food and water.If required, administer Buprenorphine to the recipient rat every six to twelve hours to alleviate the pain. Monitor the animal daily to assess for routine wound healing and signs of pain or distress. And remove the staples after seven to ten days of the surgery.A representative image shows the position of the aortic valve leaflet under the renal capsule for implementation. And after three, seven, and twenty eight days within the recipient rat, locating and recovering the transplanted tissue became easy. The aortic valve leaflets retained their native architecture after heterotopic transplantation in syngeneic animals.Demonstrating the utility of this model as a baseline to compare the immune response in allergenic transplants. The Histology with hematoxylin and eosin staining revealed that valve leaflets in syngeneic transplants after seven days were structurally intact with no signs of edematous swelling. The structural integrity of the valve leaflet after transplantation was further confirmed by Immunohistochemistry for smooth muscle actin and CD31.The aortic valve leaflets are very delicate. It is important to handle them very carefully without damaging the valves. Renal subcapsular, heart valve transplantation simplifies heterotopic heart valve transplantation.This will allow researchers with limited microsurgical capabilities to explore the immunobiology of heart valve transplantation.

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