We would like to thank the lab of Dr. Gordana Vunjak-Novakovic, including Julie Van Hassel, Mohamed Diane, and Panpan Chen, for allowing us to use cardiac waste tissue from their experiments. This work was supported by the Congenital Heart Defect Coalition in Butler, NJ, and the National Institutes of Health in Bethesda, MD (5T32HL007854-27).

All research was performed in compliance with institutional guidelines for the care of animals.
1. Considerations and preparations for the experiment
<ol>
	<li>Utilize a suitable pulse duplicator (PD) for the simulation of cardiac output through the AV. The PD will need to be able to accommodate biological materials and be able to be cleaned.
	<ol>
		<li>Utilize PD settings appropriate for testing the AV: 70 mL displacement volume and 70 beats per minute (5 L/min cardiac out...

Evaluation of Hydrodynamic Properties in Mounted Porcine Aortic Valves

<ol>
	<li>Aluru, J. S., Barsouk, A., Saginala, K., Rawla, P., Barsouk, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Valvular+heart+disease+epidemiology.">Valvular heart disease epidemiology.</a> Medical Science. 10 (2), 32 (2022).</li><li>Herrmann, J. L., Brown, 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=Seven+decades+of+valved+right+ventricular+outflow+tract+reconstruction:+The+most+common+heart+procedure+in+children.">Seven decades of valved right ventricular outflow tract reconstruction: The most common heart procedure in children.</a> The Journal of Thoracic and Cardiovascular Surgery. 160 (5), 1284-1288 (2020).</li><li>Rotman, O. M., Bianchi, M., Ghosh, R. P., Kovarovic, B., Bluestein, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Principles+of+TAVR+valve+design,+modelling,+and+testing.">Principles of TAVR valve design, modelling, and testing.</a> Expert Review of Medical Devices. 15 (11), 771-791 (2018).</li><li>Pibarot, 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=Imaging+for+predicting+and+assessing+prosthesis-patient+mismatch+after+aortic+valve+replacement.">Imaging for predicting and assessing prosthesis-patient mismatch after aortic valve replacement.</a> JACC Cardiovascular Imaging. 12 (1), 149-162 (2019).</li><li>Ozaki, 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=Aortic+valve+reconstruction+using+self-developed+aortic+valve+plasty+system+in+aortic+valve+disease.">Aortic valve reconstruction using self-developed aortic valve plasty system in aortic valve disease.</a> Interactive Cardiovascular and Thoracic Surgery. 12 (4), 550-553 (2011).</li><li>Krane, M., Amabile, A., Ziegelm&#252;ller, J. A., Geirsson, A., Lange, R. <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+neocuspidization+(the+Ozaki+procedure).">Aortic valve neocuspidization (the Ozaki procedure).</a> Multimedia Manual of Cardiothoracic Surgery. , (2021).</li><li>Ozaki, 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+total+of+404+cases+of+aortic+valve+reconstruction+with+glutaraldehyde-treated+autologous+pericardium.">A total of 404 cases of aortic valve reconstruction with glutaraldehyde-treated autologous pericardium.</a> The Journal of Thoracic and Cardiovascular Surgery. 147 (1), 301-306 (2014).</li><li>Vandecasteele, 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=The+pulmonary+veins+of+the+pig+as+an+anatomical+model+for+the+development+of+a+new+treatment+for+atrial+fibrillation.">The pulmonary veins of the pig as an anatomical model for the development of a new treatment for atrial fibrillation.</a> Anatomia Histollogia Embryologia. 44 (1), 1-12 (2015).</li><li>G&#243;es, A. M. 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=Comparative+angiotomographic+study+of+swine+vascular+anatomy:+contributions+to+research+and+training+models+in+vascular+and+endovascular+surgery.">Comparative angiotomographic study of swine vascular anatomy: contributions to research and training models in vascular and endovascular surgery.</a> Journal Vascular Brasilerio. 20, 20200086 (2021).</li><li>Ho&#322;da, M. K., Klimek-Piotrowska, W., Koziej, M., Pi&#261;tek, K., Ho&#322;da, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+different+fixation+protocols+on+the+preservation+and+dimensions+of+cardiac+tissue.">Influence of different fixation protocols on the preservation and dimensions of cardiac tissue.</a> Journal of Anatomy. 229 (2), 334-340 (2016).</li><li>Ho&#322;da, M. K., Klimek-Piotrowska, W., Koziej, M., Tyrak, K., Ho&#322;da, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Penetration+of+formaldehyde+based+fixatives+into+heart.">Penetration of formaldehyde based fixatives into heart.</a> Folia Medica Cracoviensia. 57 (4), 63-70 (2017).</li><li>Spampinato, R. 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=Grading+of+aortic+regurgitation+by+cardiovascular+magnetic+resonance+and+pulsed+Doppler+of+the+left+subclavian+artery:+harmonizing+grading+scales+between+imaging+modalities.">Grading of aortic regurgitation by cardiovascular magnetic resonance and pulsed Doppler of the left subclavian artery: harmonizing grading scales between imaging modalities.</a> International Journal of Cardiovascular Imaging. 36 (8), 1517-1526 (2020).</li><li>Capps, S. B., Elkins, R. C., Fronk, D. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Body+surface+area+as+a+predictor+of+aortic+and+pulmonary+valve+diameter.">Body surface area as a predictor of aortic and pulmonary valve diameter.</a> The Journal of Thoracic and Cardiovascular Surgery. 119 (5), 975-982 (2000).</li><li>Baumgartner, 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=Recommendations+on+the+echocardiographic+assessment+of+aortic+valve+stenosis:+a+focused+update+from+the+European+Association+of+Cardiovascular+Imaging+and+the+American+Society+of+Echocardiography.">Recommendations on the echocardiographic assessment of aortic valve stenosis: a focused update from the European Association of Cardiovascular Imaging and the American Society of Echocardiography.</a> European Heart Journal - Cardiovascular Imaging. 18 (3), 254-275 (2017).</li><li>Saisho, 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=An+ex+vivo+evaluation+of+two+different+suture+techniques+for+the+Ozaki+aortic+neocuspidization+procedure.">An ex vivo evaluation of two different suture techniques for the Ozaki aortic neocuspidization procedure.</a> Interactive Cardiovascular and Thoracic Surgery. 33 (4), 518-524 (2021).</li><li>Saisho, 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=Ex+vivo+evaluation+of+the+Ozaki+procedure+in+comparison+with+the+native+aortic+valve+and+prosthetic+valves.">Ex vivo evaluation of the Ozaki procedure in comparison with the native aortic valve and prosthetic valves.</a> Interactive Cardiovascular and Thoracic Surgery. 35 (3), (2022).</li><li>Paulsen, 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=Comprehensive+ex+vivo+comparison+of+5+clinically+used+conduit+configurations+for+valve-sparing+aortic+root+replacement+using+a+3-dimensional-printed+heart+simulator.">Comprehensive ex vivo comparison of 5 clinically used conduit configurations for valve-sparing aortic root replacement using a 3-dimensional-printed heart simulator.</a> Circulation. 142 (14), 1361-1373 (2020).</li><li>Al-Atassi, 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=Impact+of+aortic+annular+geometry+on+aortic+valve+insufficiency:+Insights+from+a+preclinical,+ex+vivo,+porcine+model.">Impact of aortic annular geometry on aortic valve insufficiency: Insights from a preclinical, ex vivo, porcine model.</a> The Journal of Thoracic and Cardiovascular Surgery. 150 (3), 656-664 (2015).</li><li>Sun, 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=A+biomimetic+multilayered+polymeric+material+designed+for+heart+valve+repair+and+replacement.">A biomimetic multilayered polymeric material designed for heart valve repair and replacement.</a> Biomaterials. 288, 121756 (2022).</li><li>Waller, B. F., McKay, C., Van Tassel, J., Allen, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Catheter+balloon+valvuloplasty+of+stenotic+porcine+bioprosthetic+valves:+Part+I:+Anatomic+considerations.">Catheter balloon valvuloplasty of stenotic porcine bioprosthetic valves: Part I: Anatomic considerations.</a> Clinical Cardiology. 14 (8), 686-691 (1991).</li><li>Crick, S. J., Sheppard, M. N., Ho, S. Y., Gebstein, L., Anderson, R. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anatomy+of+the+pig+heart:+comparisons+with+normal+human+cardiac+structure.">Anatomy of the pig heart: comparisons with normal human cardiac structure.</a> Journal of Anatomy. 193, 105-119 (1998).</li></ol>

The method presented here provides a platform for hydrodynamic testing of the AV in order to examine the effect of an experimental procedure or a novel medical device. By mounting the native aortic valve on a pulse duplicator machine, we are able to determine the effect of the experimental procedure on all the hydrodynamic parameters used in the investigation and approval of novel valve prostheses (ISO 5840). This provides an opportunity to fine-tune procedures and prostheses prior to use in a large animal model.
<p ...

Aortic valve disease represents a significant public health burden, particularly aortic stenosis, which affects 9 million people worldwide1. Strategies to address this disease are currently evolving and include aortic valve repair and aortic valve replacement. In the pediatric population especially, there is a significant incentive to repair rather than replace the valve since currently available prostheses are prone to structural valve degeneration (SVD) and are not growth tolerant, requiring reoperation for re-replacement as the patient grows. Even the Ross procedure, which replaces the diseased aortic valve (AV) with the native pulmonary valve (PV), requires a prosthesis or graft in the pulmonary position that is also subject to SVD and often limited growth tolerance2. New approaches to aortic valve disease are being developed, and there is a need for testing in a biologically relevant context prior to application in a large animal model.
We have developed a method for testing a porcine AV that can provide insights into the function of the valve before and after an investigational procedure or application of a novel medical device. By mounting the porcine AV on a commercially available pulse duplicator machine, we are able to compare the hydrodynamic characteristics that are commonly used in the investigation and ultimately approval of valve prostheses, including regurgitation fraction (RF), effective orifice area (EOA), and mean positive pressure difference (PPD)3,4. The intervention can then be fine-tuned in a biologically relevant context before use in a large animal model, thus limiting the number of animals needed to produce a procedure or prosthesis that can be used in humans. The hearts used for this experiment can be obtained from the local abattoir or waste tissue from other experiments, so it is not necessary to sacrifice an animal solely for the purposes of this experiment.
In our work, we used this method to develop a novel patch material for valve repair and replacement. We tested the hydrodynamic function of a variety of patch materials by performing an aortic valve neocuspidization procedure (Ozaki procedure5,6,7) on porcine AVs and testing them in the pulse duplicator before and after the procedure. This enabled us to fine-tune the material based on its hydrodynamic performance. Thus, this method provides a platform for hydrodynamic testing of experimental procedures and novel medical devices for use on the AV prior to application&#160;in a large animal model.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>3D Printer</td>
			<td>Ultimaker</td>
			<td>Ultimaker S5</td>
			<td>Used for printing custom fixtures for hydrodynamic testing</td>
		</tr>
		<tr>
			<td>Crile-Wood Needle Driver</td>
			<td>Emerald Instruments</td>
			<td>2.0638.15</td>
			<td>Used for suturing ventricle</td>
		</tr>
		<tr>
			<td>Debakey Forceps</td>
			<td>Jarit</td>
			<td>320-110</td>
			<td>Used for dissection and sample preparation (can use multiple if working with an assistant)</td>
		</tr>
		<tr>
			<td>Ethanol 200 proof</td>
			<td>Decon Labs Inc.</td>
			<td>DSP-MD.43</td>
			<td>Used for fixed tissue storage</td>
		</tr>
		<tr>
			<td>Formalin 10%</td>
			<td>Epredia</td>
			<td>5701</td>
			<td>Used for tissue fixation</td>
		</tr>
		<tr>
			<td>Gerald Forceps</td>
			<td>Jarit</td>
			<td>285-126</td>
			<td>Used for dissection and sample preparation</td>
		</tr>
		<tr>
			<td>Glass jars</td>
			<td>QAPPDA</td>
			<td>B07QCP54Z3</td>
			<td>Used for tissue storage</td>
		</tr>
		<tr>
			<td>Glutaraldehyde 25%</td>
			<td>Electron Microscopy Sciences</td>
			<td>16400</td>
			<td>Used for tissue fixation</td>
		</tr>
		<tr>
			<td>HEPES 1 M buffer solution</td>
			<td>Fisher</td>
			<td>BP299-100</td>
			<td>Used to make glutaraldehyde 0.6%</td>
		</tr>
		<tr>
			<td>Mayo Scissors</td>
			<td>Jarit</td>
			<td>099-200</td>
			<td>Used for cutting suture</td>
		</tr>
		<tr>
			<td>Metzenbaum Scissors</td>
			<td>Jarit</td>
			<td>099-262</td>
			<td>Used for dissection and sample preparation</td>
		</tr>
		<tr>
			<td>O-ring</td>
			<td>Sterling Seal &#38; Supply Inc.</td>
			<td>AS568-117</td>
			<td>Used as a gasket on the end of the 3D printed fixtures</td>
		</tr>
		<tr>
			<td>Polylactic acid resin</td>
			<td>Ultimaker</td>
			<td>1609</td>
			<td>Used for 3D printing fixtures</td>
		</tr>
		<tr>
			<td>Polyproplene suture</td>
			<td>Covidien</td>
			<td>VP-762-X</td>
			<td>Used for suturing ventricle, tapered needle</td>
		</tr>
		<tr>
			<td>Pulse Duplicator</td>
			<td>BDC Laboratories</td>
			<td>HDTi-6000</td>
			<td>Used for hydrodynamic testing</td>
		</tr>
		<tr>
			<td>Silk ties</td>
			<td>Covidien</td>
			<td>S-193</td>
			<td>Used for ligating coronary arteries</td>
		</tr>
		<tr>
			<td>Tonsil Clamp</td>
			<td>Aesculap</td>
			<td>BH957R</td>
			<td>Used for coronary artery dissection</td>
		</tr>
		<tr>
			<td>Zip ties (6 inch)</td>
			<td>Advanced Cable Ties, Inc.</td>
			<td>AL-06-18-9-C</td>
			<td>Used for securing sample to fixtures, 157.14 mm long (6 inches), 2.5 mm wide</td>
		</tr>
		<tr>
			<td>Zip ties (8 inch)</td>
			<td>GTSE</td>
			<td>GTSE-20025B.1000</td>
			<td>Used for securing sample to fixtures, 203 mm long (8 inches), 2.5 mm wide</td>
		</tr>
	</tbody>
</table>

an ex vivo porcine model for hydrodynamic testing of experimental aortic valve procedures and novel medical devices

The options for testing new cardiac procedures and investigative medical devices prior to use in an animal model are limited. In this study, we present a method for mounting a porcine aortic valve in a pulse duplicator to evaluate its hydrodynamic properties. These properties can then be evaluated before and after the procedure under investigation is performed and/or the investigative medical device is applied. Securing the inflow segment presents some difficulty owing to the lack of circumferential myocardium in the left ventricular outflow tract. This method addresses that issue by securing the inflow segment using the anterior leaflet of the mitral valve and then suturing the left ventricular free wall around the inflow fixture. The outflow segment is secured simply by inserting the fixture into an incision in the superior aspect of the aortic arch. We found that specimens had significantly different hydrodynamic properties before and after tissue fixation. This finding induced us to use fresh specimens in our testing and should be considered when using this method. In our work, we used this method to test novel intracardiac patch materials for use in the valvular position by performing an aortic valve neocuspidization procedure (Ozaki procedure) on the mounted porcine aortic valves. These valves were tested before and after the procedure to assess the change in hydrodynamic properties in comparison to the native valve. Herein, we report a platform for hydrodynamic testing of experimental aortic valve procedures that enables comparison with the native valve and between different devices and techniques used for the procedure under investigation.

Author Spotlight: Advancing Cardiac Procedure Testing Prior to Embarking on Large Animal Studies 

An Ex Vivo Porcine Model for Hydrodynamic Testing of Experimental Aortic Valve Procedures and Novel Medical Devices

Limited options exist for testing new cardiac procedures and investigative medical devices before use in a large animal model. Our protocol provides the opportunity to fine-tune such procedures and devices in a biologically and anatomically relevant context prior to embarking on large animal studies. Other techniques for mounting an aortic valve on a pulse duplicator require inserting or fixing a rigid structure to the aortic valve annulus, thus distorting the valve's hydrodynamic properties.Our protocol offers an innovative way to cannulate the LVOT so that all measured hydrodynamic properties are valid. This method is designed to assess the viability of leaflet substitution materials for use as a valve repair patch or prosthesis. It is a pivotal tool for our future research in which we are developing biomimetic polymeric materials for use as durable valve substitutives with native tissue, like mechanical and hydrodynamic properties.

The representative data collected from the pulse duplicator includes regurgitation fraction (RF), effective orifice area (EOA), and mean positive pressure difference (PPD). The RF and EOA, in particular, are used in the ISO standards for prosthetic valves (ISO 5840) and will be important to collect if prosthetic valve products are under investigation. The PPD offers information regarding how much pressure is required to open the valve and is commonly referenced when discussing prosthetic valve replacement<sup class="xref...

Watch this Scientific Journal Video about Advancing Cardiac Procedure Testing Prior to Embarking on Large Animal Studies at JoVE.com

We present a method for mounting a porcine aortic valve on a pulse duplicator to test its hydrodynamic properties. This method can be used to determine the change in hydrodynamics after the application of an experimental procedure or novel medical device prior to use in a large animal model.

The options for testing new cardiac procedures and investigative medical devices prior to use in an animal model are limited. In this study, we present a ...