Name: normothermic ex situ heart perfusion in working mode: assessment of cardiac function and metabolism
Uploaded: 2019-01-12T12:30:00
Description: Watch this Scientific Journal Video about Normothermic Ex Situ Heart Perfusion in Working Mode: Assessment of Cardiac Function and Metabolism at JoVE.com

This work was supported by grants from the Canadian National Transplant Research Program. SH is the recipient of a Faculty of Medicine and Dentistry Motyl Graduate Studentship in Cardiac Sciences. DHF is a recipient of a Collaborative Research Projects (CHRP) grant in aid from the National Sciences and Engineering Research Council and Canadian Institutes of Health Research.

All the procedures in this manuscript were performed in compliance with the guidelines of the Canadian Council on Animal Care and the guide for the care and use of laboratory animals. The protocols were approved by the institutional animal care committee of the University of Alberta. This protocol has been applied in female juvenile Yorkshire pigs between 35&#8211;50 kg. All individuals involved in ESHP procedures had received proper biosafety training.
1. Pre-surgical Preparations
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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=Ex+vivo+perfusion+in+a+loaded+state+improves+the+preservation+of+donor+heart+function.">Ex vivo perfusion in a loaded state improves the preservation of donor heart function.</a> Canadian Journal of cardiology. 31 (10), s202 (2015).</li><li>White, 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=A+wholeblood-based+perfusate+provides+superior+preservation+of+myocardial+function+during+ex+vivo+heart+perfusion.">A wholeblood-based perfusate provides superior preservation of myocardial function during ex vivo heart perfusion.</a> Journal of Heart and Lung Transplantation. (14), (2014).</li><li>Lips, D. 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Preload recruitable stroke work, may be a possible candidate.</a> Journal of Cardiology. 71 (4), 325-326 (2018).</li><li>Hatami, 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> Canadian Society for Transplantation. , (2017).</li><li>Anthony, 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=Ex+vivo+coronary+angiographic+evaluation+of+a+beating+donor+heart.">Ex vivo coronary angiographic evaluation of a beating donor heart.</a> Circulation. 130 (25), e341-e343 (2014).</li><li>Sandha, J. 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Successful perfusion is defined according to the aims of the study; however, this should include uninterrupted ESHP for the desired amount of time and complete collection of the data on cardiac function during the perfusion. For this purpose, a few critical steps in the protocol must be followed.
The heart is an organ with high oxygen and energy demands, and minimizing the ischemic time before cannulation and perfusion is an important principle that must be followed. The process of procurement...

Clinical relevance
While most aspects of cardiac transplantation have evolved significantly since the first heart transplant in 1967, cold storage (CS) remains the standard for donor heart preservation1. CS exposes the organ to a period of cold ischemia that limits the safe preservation interval (4&#8211;6 hours) and increases the risk of primary graft dysfunction2,3,4. Due to the static nature of CS, assessments of function or therapeutic interventions are not possible in the time between the organ procurement and transplantation. This is a particular limitation in extended criteria donors including hearts donated after circulatory death (DCD), creating an obstacle for overcoming the considerable gap between demand and the current donor pool5,6. To address this limitation, ex situ heart perfusion has been proposed as a novel, semi-physiologic method of preserving donated hearts, minimizing exposure to cold ischemia by providing oxygenated, nutrient-rich perfusate to the heart during preservation time1,7,8.
Ex situ heart perfusion
One of the most frequently used methods for ex situ examination of the isolated heart is Langendorff perfusion. In this method, introduced by Oskar Langendorff in 1895, the blood flows into the coronary arteries and out the coronary sinus of the isolated heart, with the heart in an empty and beating state9,10. Clinical ESHP in a Langendorff mode with the Transmedics Organ Care System apparatus (OCS) has been shown to be non-inferior to CS in the preservation of standard-criteria donor hearts1, and has facilitated the clinical transplantation of DCD hearts11. However, there are concerns about the ability of the device to evaluate organ viability, as a number of donor hearts initially thought to be transplantable were discarded after perfusion on the OCS3. The OCS supports the heart in the Langendorff (non-working) mode, and thus possesses a limited capacity for evaluation of the pumping function of the heart3,12. A growing body of evidence suggests that functional parameters offer a better way to assess organ viability, suggesting that assessments of cardiac function may become a reliable tool for the evaluation and selection of hearts for transplantation during ESHP3,12,13,14, Furthermore, our studies on ex situ perfused porcine hearts suggest that ESHP in working mode provides enhanced functional preservation of the heart during the perfusion interval15,16.
An ESHP apparatus capable of preserving the heart in a working mode must possess a level of automation to safely and precisely maintain preload, afterload and flow rates. Also, such a system should possess the flexibility to facilitate comprehensive assessments of cardiac function to be undertaken. The ESHP apparatus used here is equipped with custom software that 1) provides and maintains desired aortic (Ao) and left atrial (LA) pressure/flow and 2) provides real-time analysis of functional parameters and visual evaluation of pressure waveforms with minimal need for supervision. Pressure data is acquired with standard fluid-filled pressure transducers, and flow data is acquired with transit-time doppler flow probes. These signals are digitized with a bridge and analog input, respectively. The heart is positioned horizontally with a slight elevation to the great vessels on a soft silicone membrane. The cannulation attachments pass through the membrane, incorporating a compliance chamber for dampening ventricular ejection. The goal of this work is to provide researchers in the field of cardiac transplantation with a protocol for ex situ perfusion and evaluation of the heart, under normothermic, semi-physiologic conditions in working mode, in a large mammal (Yorkshire pig) model.

<table>
	<tbody>
		<tr>
			<td>Debakey-Metzenbaum dissecting scissors</td>
			<td>Pilling</td>
			<td>342202</td>
			<td></td>
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			<td>MAYO dissecting scissors</td>
			<td>Pilling</td>
			<td>460420</td>
			<td></td>
		</tr>
		<tr>
			<td>THUMB forceps</td>
			<td>Pilling</td>
			<td>465165</td>
			<td></td>
		</tr>
		<tr>
			<td>Debakey straight vascular tissue forceps&#160;</td>
			<td>Pilling</td>
			<td>351808</td>
			<td></td>
		</tr>
		<tr>
			<td>CUSHING Gutschdressing forceps</td>
			<td>Pilling</td>
			<td>466200</td>
			<td></td>
		</tr>
		<tr>
			<td>JOHNSON needle holder</td>
			<td>Pilling</td>
			<td>510312</td>
			<td></td>
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		<tr>
			<td>DERF needle holder</td>
			<td>Pilling</td>
			<td>443120</td>
			<td></td>
		</tr>
		<tr>
			<td>Sternal saw</td>
			<td>Stryker</td>
			<td>6207</td>
			<td></td>
		</tr>
		<tr>
			<td>Sternal retractor</td>
			<td>Pilling</td>
			<td>341162</td>
			<td></td>
		</tr>
		<tr>
			<td>Vorse tubing clamp</td>
			<td>Pilling</td>
			<td>351377</td>
			<td></td>
		</tr>
		<tr>
			<td>MORRIS ascending aorta clamp</td>
			<td>Pilling</td>
			<td>353617</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical snare (tourniquet) set</td>
			<td>Medtronic</td>
			<td>CVR79013</td>
			<td></td>
		</tr>
		<tr>
			<td>2-0 SILK black 12 X 18&#34; strands</td>
			<td>ETHICON</td>
			<td>A185H</td>
			<td></td>
		</tr>
		<tr>
			<td>3-0 PROLENE blue 18&#34; PS-2 cutting</td>
			<td>ETHICON</td>
			<td>8687H</td>
			<td></td>
		</tr>
		<tr>
			<td>Biomedicus pump drive (modified)</td>
			<td>Medtronic</td>
			<td>540</td>
			<td>Modified to allow remote electronic control of pump speed</td>
		</tr>
		<tr>
			<td>Biomedicus pump</td>
			<td>Maquet</td>
			<td>BPX-80</td>
			<td></td>
		</tr>
		<tr>
			<td>Membrane oxigenator D 905</td>
			<td>SORIN GROUP</td>
			<td>50513</td>
			<td></td>
		</tr>
		<tr>
			<td>Tubing flow module&#160;&#160;</td>
			<td>Transonic</td>
			<td>Ts410</td>
			<td></td>
		</tr>
		<tr>
			<td>PXL clamp-on flow sensor</td>
			<td>Transonic</td>
			<td>ME9PXL-BL37SF</td>
			<td></td>
		</tr>
		<tr>
			<td>TruWave pressure transducer</td>
			<td>Edwards</td>
			<td>VSYPX272</td>
			<td></td>
		</tr>
		<tr>
			<td>Intercept tubing 3/8&#34; X 3/32&#34; X 6&#39;</td>
			<td>Medtronic</td>
			<td>3506</td>
			<td></td>
		</tr>
		<tr>
			<td>Intercept tubing 1/4&#34; X 1/16&#34; X 8&#39;</td>
			<td>Medtronic</td>
			<td>3108</td>
			<td></td>
		</tr>
		<tr>
			<td>Heated/Refrigerated Bath Circulator&#160;</td>
			<td>Grant</td>
			<td>TX-150</td>
			<td></td>
		</tr>
		<tr>
			<td>ABL 800 FLEX Blood Gas Analyzer</td>
			<td>Radiometer</td>
			<td>989-963</td>
			<td></td>
		</tr>
		<tr>
			<td>5F Ventriculr straight pigtail cathter</td>
			<td>CORDIS</td>
			<td>534550S</td>
			<td></td>
		</tr>
		<tr>
			<td>5F AVANTI+ Sheath Introducer</td>
			<td>CORDIS</td>
			<td>504605A</td>
			<td></td>
		</tr>
		<tr>
			<td>Emerald Amplatz Guidewire</td>
			<td>CORDIS</td>
			<td>502571A</td>
			<td></td>
		</tr>
		<tr>
			<td>Dual chamber pace maker</td>
			<td>Medtronic</td>
			<td>5388</td>
			<td></td>
		</tr>
		<tr>
			<td>Defibrilltor</td>
			<td>CodeMaster</td>
			<td>M1722B</td>
			<td></td>
		</tr>
		<tr>
			<td>Infusion pump</td>
			<td>Baxter</td>
			<td>AS50</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical electrocautery device</td>
			<td>Kls Martin</td>
			<td>ME411</td>
			<td></td>
		</tr>
		<tr>
			<td>Gas mixer</td>
			<td>SECHRIST</td>
			<td>3500 CP-G</td>
			<td></td>
		</tr>
		<tr>
			<td>Medical oxygen tank</td>
			<td>praxair</td>
			<td>2014408</td>
			<td></td>
		</tr>
		<tr>
			<td>Cabon dioxide tank</td>
			<td>praxair</td>
			<td>5823115</td>
			<td></td>
		</tr>
		<tr>
			<td>Bovine serum albumin</td>
			<td>MP biomedicals</td>
			<td>218057791</td>
			<td></td>
		</tr>
	</tbody>
</table>

normothermic ex situ heart perfusion in working mode: assessment of cardiac function and metabolism

The current standard method for organ preservation (cold storage, CS), exposes the heart to a period of cold ischemia that limits the safe preservation time and increases the risk of adverse post-transplantation outcomes. Moreover, the static nature of CS does not allow for organ evaluation or intervention during the preservation interval. Normothermic ex situ heart perfusion (ESHP) is a novel method for preservation of the donated heart that minimizes cold ischemia by providing oxygenated, nutrient-rich perfusate to the heart. ESHP has been shown to be non-inferior to CS in the preservation of standard-criteria donor hearts and has also facilitated the clinical transplantation of the hearts donated after the circulatory determination of death. Currently, the only available clinical ESHP device perfuses the heart in an unloaded, non-working state, limiting assessments of myocardial performance. Conversely, ESHP in working mode provides the opportunity for comprehensive evaluation of cardiac performance by assessment of functional and metabolic parameters under physiologic conditions. Moreover, earlier experimental studies have suggested that ESHP in working mode may result in improved functional preservation. Here, we describe the protocol for ex situ perfusion of the heart in a large mammal (porcine) model, which is reproducible for different animal models and heart sizes. The software program in this ESHP apparatus allows for real-time and automated control of the pump speed to maintain desired aortic and left atrial pressure and evaluates a variety of functional and electrophysiological parameters with minimal need for supervision/manipulation.

At the start of the perfusion (in non-working mode), the heart will normally resume a sinus rhythm when the temperature of the system and perfusate approaches normothermia. When entering working mode, as the LA pressures are approaching the desired values, ejection on the Ao pressure tracing should be observed and the LA flow (a reflection of cardiac output) should increase gradually. In a Yorkshire pig model (35&#8211;50 kg) and a starting heart weight of 180&#8211;220 grams, the initial...

Watch this Scientific Journal Video about Normothermic Ex Situ Heart Perfusion in Working Mode: Assessment of Cardiac Function and Metabolism at JoVE.com

Normothermic Ex Situ Heart Perfusion in Working Mode: Assessment of Cardiac Function and Metabolism

Normothermic ex situ heart perfusion (ESHP), preserves the heart in a beating, semi-physiologic state. When performed in a working mode, ESHP provides the opportunity to perform sophisticated assessments of donor heart function and organ viability. Here, we describe our method for myocardial performance evaluation during ESHP.

The current standard method for organ preservation (cold storage, CS), exposes the heart to a period of cold ischemia that limits the safe preservation ...

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