Name: pre-clinical model of cardiac donation after circulatory death
Uploaded: 2019-08-02T13:30:00
Description: Watch this Scientific Journal Video about Pre-clinical Model of Cardiac Donation after Circulatory Death at JoVE.com

Ein Teil dieser Arbeit wurde durch einen großzügigen Beitrag der Fondation Marcel et Rolande Gosselin und der Fondation Stefane Foumy unterstützt. Nicolas Noiseux ist Gelehrter der FRQ-S.
Die Autoren bedanken sich bei Josh Zhuo Le Huang, Gabrielle Gascon, Sophia Ghiassi und Catherine Scalabrini für ihre Unterstützung bei der Datenerhebung.

Alle Tierpflege- und Versuchsprotokolle entsprachen dem Leitfaden für die Pflege und Verwendung von Labortieren und wurden vom institutionellen Tierpflege- und Einsatzausschuss des Centre Hospitalier de l'Université de Montréal Research Center genehmigt.
1. Vorvorbereitungen
<ol><li>Schalten Sie das Wasserbad ein, um das Cardioplegie-Abgabesystem (Abbildung 1A) und das Langendorff ex vivo Perfusionssystem (Abbildung 1B) zu erwär...

<ol>
	<li>Gass, A. L., 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+Transplantation+in+the+New+Era.">Cardiac Transplantation in the New Era.</a> Cardiology in Review. 23 (4), 182-188 (2015).</li><li>von Dossow, V., Costa, J., D'Ovidio, F., Marczin, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Worldwide+trends+in+heart+and+lung+transplantation:+Guarding+the+most+precious+gift+ever.">Worldwide trends in heart and lung transplantation: Guarding the most precious gift ever.</a> Best Practice &amp; Research. Clinical Anaesthesiology. 31 (2), 141-152 (2017).</li><li>Hornby, K., Ross, H., Keshavjee, S., Rao, V., Shemie, S. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Non-utilization+of+hearts+and+lungs+after+consent+for+donation:+a+Canadian+multicentre+study.">Non-utilization of hearts and lungs after consent for donation: a Canadian multicentre study.</a> Canadian Journal Of Anaesthesia. 53 (8), 831-837 (2006).</li><li>Manyalich, M., Nelson, H., Delmonico, F. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+need+and+opportunity+for+donation+after+circulatory+death+worldwide.">The need and opportunity for donation after circulatory death worldwide.</a> Current Opinion In Organ Transplantation. 23 (1), 136-141 (2018).</li><li>Shemie, S. 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=National+recommendations+for+donation+after+cardiocirculatory+death+in+Canada:+Donation+after+cardiocirculatory+death+in+Canada.">National recommendations for donation after cardiocirculatory death in Canada: Donation after cardiocirculatory death in Canada.</a> CMAJ : Canadian Medical Association Journal. 175 (8), S1 (2006).</li><li>Page, A., Messer, S., Large, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heart+transplantation+from+donation+after+circulatory+determined+death.">Heart transplantation from donation after circulatory determined death.</a> Annals of Cardiothoracic Surgery. 7 (1), 75-81 (2018).</li><li>Monteagudo Vela, M., Garcia Saez, D., Simon, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Current+approaches+in+retrieval+and+heart+preservation.">Current approaches in retrieval and heart preservation.</a> Annals of Cardiothoracic Surgery. 7 (1), 67-74 (2018).</li><li>Dhital, K. K., Chew, H. C., Macdonald, P. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Donation+after+circulatory+death+heart+transplantation.">Donation after circulatory death heart transplantation.</a> Current Opinion In Organ Transplantation. 22 (3), 189-197 (2017).</li><li>McNally, S. J., Harrison, E. M., Wigmore, S. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ethical+considerations+in+the+application+of+preconditioning+to+solid+organ+transplantation.">Ethical considerations in the application of preconditioning to solid organ transplantation.</a> Journal of Medical Ethics. 31 (11), 631-634 (2005).</li><li>Rao, V., Feindel, C. M., Weisel, R. D., Boylen, P., Cohen, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Donor+blood+perfusion+improves+myocardial+recovery+after+heart+transplantation.">Donor blood perfusion improves myocardial recovery after heart transplantation.</a> The Journal of Heart and Lung Transplantation. 16 (6), 667-673 (1997).</li><li>Ramzy, 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+allograft+preservation+using+donor-shed+blood+supplemented+with+L-arginine.">Cardiac allograft preservation using donor-shed blood supplemented with L-arginine.</a> The Journal of Heart and Lung Transplantation. 24 (10), 1665-1672 (2005).</li><li>Xin, L., 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+New+Multi-Mode+Perfusion+System+for+Ex+vivo+Heart+Perfusion+Study.">A New Multi-Mode Perfusion System for Ex vivo Heart Perfusion Study.</a> Journal of Medical Systems. 42 (2), 25 (2017).</li><li>Messer, S., Ardehali, A., Tsui, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Normothermic+donor+heart+perfusion:+current+clinical+experience+and+the+future.">Normothermic donor heart perfusion: current clinical experience and the future.</a> Transplant International. 28 (6), 634-642 (2015).</li><li>Flecknell, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Laboratory Animal Anaesthesia (Fourth Edition). , 77-108 (2016).</li><li>Kearns, 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=A+Rodent+Model+of+Cardiac+Donation+After+Circulatory+Death+and+Novel+Biomarkers+of+Cardiac+Viability+During+Ex+vivo+Heart+Perfusion.">A Rodent Model of Cardiac Donation After Circulatory Death and Novel Biomarkers of Cardiac Viability During Ex vivo Heart Perfusion.</a> Transplantation. 101 (8), e231-e239 (2017).</li><li>Sandha, J. K., 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=Steroids+Limit+Myocardial+Edema+During+Ex+vivo+Perfusion+of+Hearts+Donated+After+Circulatory+Death.">Steroids Limit Myocardial Edema During Ex vivo Perfusion of Hearts Donated After Circulatory Death.</a> The Annals of Thoracic Surgery. 105 (6), 1763-1770 (2018).</li><li>Iyer, 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=Increasing+the+tolerance+of+DCD+hearts+to+warm+ischemia+by+pharmacological+postconditioning.">Increasing the tolerance of DCD hearts to warm ischemia by pharmacological postconditioning.</a> American Journal of Transplantation. 14 (8), 1744-1752 (2014).</li><li>Sanz, M. N., 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=Cardioprotective+reperfusion+strategies+differentially+affect+mitochondria:studies+in+an+isolated+rat+heart+model+of+donation+after+circulatory+death+(DCD).">Cardioprotective reperfusion strategies differentially affect mitochondria:studies in an isolated rat heart model of donation after circulatory death (DCD).</a> American Journal of Transplantation. , (2018).</li><li>Van de Wauwer, 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=The+mode+of+death+in+the+non-heart-beating+donor+has+an+impact+on+lung+graft+quality.">The mode of death in the non-heart-beating donor has an impact on lung graft quality.</a> European Journal of Cardio-Thoracic Surgery. 36 (5), 919-926 (2009).</li><li>Quader, 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=Determination+of+Optimal+Coronary+Flow+for+the+Preservation+of+&amp;#34;Donation+after+Circulatory+Death&amp;#34;+in+Murine+Heart+Model.">Determination of Optimal Coronary Flow for the Preservation of &amp;#34;Donation after Circulatory Death&amp;#34; in Murine Heart Model.</a> ASAIO journal (American Society for Artificial Internal Organs : 1992). 64 (2), 225-231 (2018).</li><li>Priebe, H. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+acute+open-chest+model.">The acute open-chest model.</a> British Journal Of Anaesthesia. 60 (8 Suppl 1), 38-41 (1988).</li><li>Narita, 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=Cardiac+effects+of+vecuronium+and+its+interaction+with+autonomic+nervous+system+in+isolated+perfused+canine+hearts.">Cardiac effects of vecuronium and its interaction with autonomic nervous system in isolated perfused canine hearts.</a> Journal of Cardiovascular Pharmacology. 19 (6), 1000-1008 (1992).</li><li>Dhital, K. K., 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=Adult+heart+transplantation+with+distant+procurement+and+ex-vivo+preservation+of+donor+hearts+after+circulatory+death:+a+case+series.">Adult heart transplantation with distant procurement and ex-vivo preservation of donor hearts after circulatory death: a case series.</a> Lancet (London, England). 385 (9987), 2585-2591 (2015).</li><li>Messer, S. 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=Functional+assessment+and+transplantation+of+the+donor+heart+after+circulatory+death.">Functional assessment and transplantation of the donor heart after circulatory death.</a> The Journal of Heart and Lung Transplantation. 35 (12), 1443-1452 (2016).</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=Assessment+of+donor+heart+viability+during+ex+vivo+heart+perfusion.">Assessment of donor heart viability during ex vivo heart perfusion.</a> Canadian Journal of Physiology and Pharmacology. 93 (10), 893-901 (2015).</li><li>Mayr, 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=Cardiac+troponin+T+and+creatine+kinase+predict+mid-term+infarct+size+and+left+ventricular+function+after+acute+myocardial+infarction:+a+cardiac+MR+study.">Cardiac troponin T and creatine kinase predict mid-term infarct size and left ventricular function after acute myocardial infarction: a cardiac MR study.</a> Journal of Magnetic Resonance Imaging. 33 (4), 847-854 (2011).</li><li>Remppis, 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=Intracellular+compartmentation+of+troponin+T:+release+kinetics+after+global+ischemia+and+calcium+paradox+in+the+isolated+perfused+rat+heart.">Intracellular compartmentation of troponin T: release kinetics after global ischemia and calcium paradox in the isolated perfused rat heart.</a> Journal of Molecular and Cellular Cardiology. 27 (2), 793-803 (1995).</li><li>Rossello, X., Hall, A. R., Bell, R. M., Yellon, 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=Characterization+of+the+Langendorff+Perfused+Isolated+Mouse+Heart+Model+of+Global+Ischemia-Reperfusion+Injury:+Impact+of+Ischemia+and+Reperfusion+Length+on+Infarct+Size+and+LDH+Release.">Characterization of the Langendorff Perfused Isolated Mouse Heart Model of Global Ischemia-Reperfusion Injury: Impact of Ischemia and Reperfusion Length on Infarct Size and LDH Release.</a> Journal of Cardiovascular Pharmacology and Therapeutics. 21 (3), 286-295 (2016).</li><li>Dornbierer, 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=Early+reperfusion+hemodynamics+predict+recovery+in+rat+hearts:+a+potential+approach+towards+evaluating+cardiac+grafts+from+non-heart-beating+donors.">Early reperfusion hemodynamics predict recovery in rat hearts: a potential approach towards evaluating cardiac grafts from non-heart-beating donors.</a> PloS One. 7 (8), e43642 (2012).</li><li>Henry, P. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Positive+staircase+effect+in+the+rat+heart.">Positive staircase effect in the rat heart.</a> The American Journal of Physiology. 228 (2), 360-364 (1975).</li><li>Markert, 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=Evaluation+of+a+method+to+correct+the+contractility+index+LVdP/dt(max)+for+changes+in+heart+rate.">Evaluation of a method to correct the contractility index LVdP/dt(max) for changes in heart rate.</a> Journal of Pharmacological and Toxicological Methods. 66 (2), 98-105 (2012).</li><li>Azar, T., Sharp, J., Lawson, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heart+rates+of+male+and+female+Sprague-Dawley+and+spontaneously+hypertensive+rats+housed+singly+or+in+groups.">Heart rates of male and female Sprague-Dawley and spontaneously hypertensive rats housed singly or in groups.</a> Journal of the American Association for Laboratory Animal Science. 50 (2), 175-184 (2011).</li><li>Bonney, S., Hughes, K., Eckle, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anesthetic+cardioprotection:+the+role+of+adenosine.">Anesthetic cardioprotection: the role of adenosine.</a> Current Pharmaceutical Design. 20 (36), 5690-5695 (2014).</li><li>Ali, A. 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=Rat+model+of+veno-arterial+extracorporeal+membrane+oxygenation.">Rat model of veno-arterial extracorporeal membrane oxygenation.</a> Journal of Translational Medicine. 12, 37 (2014).</li></ol>

Das hier vorgestellte Protokoll stellt ein einfaches, bequemes und vielseitiges Modell von Herz-DCD vor, das die Möglichkeit bietet, die herzfunktionelle Erholung, Gewebeschäden und den Einsatz von kardioprotektive naktiven Nachhilfemitteln zu bewerten, um die Genesung des Spenders zu verbessern. Herzen, die sonst zur Transplantation verworfen werden. Ex-vivo-Herzperfusionssysteme (EVHP) wurden optimiert, um eine Plattform zur Beurteilung der Herzfunktion zu bieten und eine einzigartige Möglichkeit zu bieten, modifizi...

Feste Organtransplantationen im Allgemeinen und Herztransplantationen im Besonderen sind weltweit auf dem Vormarsch1,2. Die Standardmethode der Organbeschaffung ist die Spende nach dem Hirntod (DBD). Angesichts der strengen Inklusionskriterien der DBD werden weniger als 40% der angebotenen Herzenakzeptiert 3, wodurch das Angebot angesichts der steigenden Nachfrage begrenzt und die Orgelwarteliste erweitert wird. Um dieses Problem anzugehen, wird die Verwendung von Organen, die nach dem Kreislauftod gespendet werden (DCD), als mögliche Lösung betrachtet4.
Bei DCD-Spendern sind jedoch eine agonale Phase nach Entzug der Pflege und eine Periode ungeschützter warmer Ischämie vor Reanimation unvermeidlich5. Die mögliche Organverletzung nach dem Kreislauftod kann zu Organfunktionsstörungen führen, was die Abneigung gegen routinemäßige DCD-Herztransplantationen erklärt. Es wird berichtet, dass nur 4 Zentren DCD-Herzen klinisch verwenden, mit strengen Kriterien, die sehr kurze warme Ischämie-Zeiten und junge Spender ohne chronische Pathologienumfasst 6,7. Aus ethischen und rechtlichen Gründen können bei Spendern vor dem Kreislauftod5,8,9begrenzte oder keine kardioprotektive Eingriffe angewendet werden. Daher ist jede Abschwächung zur Linderung der Ischämie-Reperfusion (IR)-Verletzung auf kardioprotektive Therapien beschränkt, die während der frühen Reperfusion mit kardiopdiogischen Lösungen eingeleitet werden, und lässt keine ordnungsgemäße funktionelle Bewertung zu. Ex-vivo-Herzperfusion (EVHP) und Rekonditionierung des DCD-Herzes mit speziellen Plattformen wurde als alternative Lösung vorgeschlagen und von verschiedenen Wissenschaftlern untersucht10,11,12,13 . EVHP bietet eine einzigartige Gelegenheit, Post-Konditionierungs-Agenten an DCD-Herzen zu liefern, um die funktionelle Erholung zu verbessern. Für eine effiziente klinische Übersetzung müssen jedoch noch viele technische und praktische Fragen angegangen werden, und dies wird noch verstärkt durch einen Mangel an Konsens über eine Reihe von Perfusions- und funktionellen Kriterien zur Bestimmung der Transplantierbarkeit6, 8.
Hierin berichten wir über die Entwicklung eines reproduzierbaren präklinischen DCD-Protokolls für Kleintiere in Kombination mit einem Ex-vivo-Herzperfusionssystem, das zur Untersuchung der zum Zeitpunkt der Beschaffung eingeleiteten Organpostkonditionierung während der ersten Reperfusion verwendet werden kann, und /oder in ganz EVHP.

<table>
	<tbody>
		<tr>
			<td>0,9% Sodium Chloride. 1L bag</td>
			<td>Baxter</td>
			<td></td>
			<td>Electrolyte solution for flushing in the modified Langendorff system.</td>
		</tr>
		<tr>
			<td>14G 2&#34; I.V catheter</td>
			<td>Jelco</td>
			<td>4098</td>
			<td>To act as endotracheal tube.</td>
		</tr>
		<tr>
			<td>2,3,5-Triphenyltetrazolium chloride</td>
			<td>Milipore-Sigma</td>
			<td>T8877</td>
			<td>Vital coloration</td>
		</tr>
		<tr>
			<td>22G 1&#34; I.V catheter</td>
			<td>BD</td>
			<td>383532</td>
			<td>I.V catheter with extension tube that facilitates manipulation for carotid catheterization</td>
		</tr>
		<tr>
			<td>Adson Dressing Fcp, 4 3/4&#34;, Serr</td>
			<td>Skalar</td>
			<td>50-3147</td>
			<td>Additional forceps for tissue manipulation</td>
		</tr>
		<tr>
			<td>Alm Self-retaining retractor 4x4 Teeth Blunt 2-3/4&#34;</td>
			<td>Skalar</td>
			<td>22-9027</td>
			<td>Tissue retractor used to maintain the chest open.</td>
		</tr>
		<tr>
			<td>Bridge amp</td>
			<td>ADinstruments</td>
			<td>FE221</td>
			<td>Bridge amp for intracarotid blood pressure measurement</td>
		</tr>
		<tr>
			<td>Calcium chloride</td>
			<td>Milipore-Sigma</td>
			<td>C1016</td>
			<td>CaCl2 anhydrous, granular, &#8804;7.0 mm, &#8805;93.0% Part of the Krebs solution</td>
		</tr>
		<tr>
			<td>D-(+)-Glucose</td>
			<td>Milipore-Sigma</td>
			<td>G8270</td>
			<td>D-Glucose &#8805;99.5% Part of the Krebs solution</td>
		</tr>
		<tr>
			<td>DIN(8) to Disposable BP Transducer</td>
			<td>ADinstruments</td>
			<td>MLAC06</td>
			<td>Adapter cable for link between bridge amp and pressure transducer</td>
		</tr>
		<tr>
			<td>Disposable BP Transducer (stopcock)</td>
			<td>ADinstruments</td>
			<td>MLT0670</td>
			<td>Pressure transducer for intracarotid blood pressure measurement</td>
		</tr>
		<tr>
			<td>dPBS</td>
			<td>Gibco</td>
			<td>14190-144</td>
			<td>Electrolyte solution without calcium or magnesium.</td>
		</tr>
		<tr>
			<td>Eye Dressing Fcp, Str, Serr, 4&#34;</td>
			<td>Skalar</td>
			<td>66-2740</td>
			<td>Additional forceps for tissue manipulation</td>
		</tr>
		<tr>
			<td>Formalin solution, neutral buffered, 10%</td>
			<td>Milipore-Sigma</td>
			<td>HT501128</td>
			<td>Fixative solution</td>
		</tr>
		<tr>
			<td>Heating Pad</td>
			<td>Sunbean</td>
			<td>756-CN</td>
			<td></td>
		</tr>
		<tr>
			<td>Heparin sodium 1000 UI/mL</td>
			<td>Sandoz</td>
			<td></td>
			<td>For systemic anticoagulation</td>
		</tr>
		<tr>
			<td>Hydrochloric Acid 36,5 to 38,0%</td>
			<td>Fisher scientific</td>
			<td>A144-500</td>
			<td>Diluted 1:1 for pH correction</td>
		</tr>
		<tr>
			<td>Ketamine</td>
			<td>Bimeda</td>
			<td></td>
			<td>Anesthetic. 100 mg/ml</td>
		</tr>
		<tr>
			<td>LabChart</td>
			<td>ADinstruments</td>
			<td></td>
			<td>Control software for the Powerlab polygraph, allowing off-line analyses. Version 7, with blood pressure and PV loop modules enabled</td>
		</tr>
		<tr>
			<td>Left ventricle pressure balloon</td>
			<td>Radnoti</td>
			<td>170404</td>
			<td>In latex. Size 4.</td>
		</tr>
		<tr>
			<td>Lidocaine HCl 2% solution</td>
			<td>AstraZeneca</td>
			<td></td>
			<td>Antiarrhythmic for the cardioplegic solution</td>
		</tr>
		<tr>
			<td>Magnesium Chloride ACS</td>
			<td>ACP Chemicals</td>
			<td>M-0460</td>
			<td>MgCl2+6H2O &#8805;99.0% Part of the Krebs solution</td>
		</tr>
		<tr>
			<td>Micro pressure sensor</td>
			<td>Radnoti</td>
			<td>159905</td>
			<td>Micro pressure sensor and amplifier connected to the intraventricular balloon</td>
		</tr>
		<tr>
			<td>Pacemaker</td>
			<td>Biotronik</td>
			<td>Reliaty</td>
			<td>Set to generate a pulse each 200 ms for a heart rate of 300 bpm.</td>
		</tr>
		<tr>
			<td>pH bench top meter</td>
			<td>Fisher scientific</td>
			<td>AE150</td>
			<td></td>
		</tr>
		<tr>
			<td>Physiological monitor</td>
			<td>Kent Scientific</td>
			<td>Physiosuite</td>
			<td>For continuous monitoring of rodent temperature and saturation during the procedure</td>
		</tr>
		<tr>
			<td>Plasma-Lyte A</td>
			<td>Baxter</td>
			<td></td>
			<td>Electrolyte solution used as base to prepare cardioplegia</td>
		</tr>
		<tr>
			<td>Potassium Chloride</td>
			<td>Milipore-Sigma</td>
			<td>P4504</td>
			<td>KCl &#8805;99.0% Part of the Krebs solution</td>
		</tr>
		<tr>
			<td>Potassium Chloride 2 meq/ml</td>
			<td>Hospira</td>
			<td></td>
			<td>Part of the cardioplegic solution</td>
		</tr>
		<tr>
			<td>PowerLab 8/30 Polygraph</td>
			<td>ADinstruments</td>
			<td></td>
			<td>Electronic polygraph</td>
		</tr>
		<tr>
			<td>Silk 2-0</td>
			<td>Ethicon</td>
			<td>A305H</td>
			<td>Suture material for Langendorff apparatus</td>
		</tr>
		<tr>
			<td>Silk 5-0</td>
			<td>Ethicon</td>
			<td>A302H</td>
			<td>Suture material for carotid</td>
		</tr>
		<tr>
			<td>Small animal anesthesia workstation</td>
			<td>Hallowell EMC</td>
			<td>000A2770</td>
			<td>Small animal ventilator</td>
		</tr>
		<tr>
			<td>Sodium bicarbonate</td>
			<td>Milipore-Sigma</td>
			<td>S5761</td>
			<td>NaHCO3 &#8805;99,5% Part of the Krebs solution</td>
		</tr>
		<tr>
			<td>Sodium Chloride</td>
			<td>Milipore-Sigma</td>
			<td>S7653</td>
			<td>NaCl &#8805;99.5% Part of the Krebs solution</td>
		</tr>
		<tr>
			<td>Sodium Hydroxide pellets</td>
			<td>ACP chemicals</td>
			<td>S3700</td>
			<td>Diluted to 5N (10 g in 50 ml) for pH correction</td>
		</tr>
		<tr>
			<td>Sodium phosphate monobasic</td>
			<td>Milipore-Sigma</td>
			<td>S0751</td>
			<td>NaH2PO4 &#8805;99.0% Part of the Krebs solution</td>
		</tr>
		<tr>
			<td>Stevens Tenotomy Sciss, Str, Delicate, SH/SH, 4 1/2&#34;</td>
			<td>Skalar</td>
			<td>22-1240</td>
			<td>Small scisors for atria and cava vein opening</td>
		</tr>
		<tr>
			<td>Tissue slicer blades</td>
			<td>Thomas scientific</td>
			<td>6727C18</td>
			<td>Straight carbon steel blades for tissue slicing at the end of the protocol</td>
		</tr>
		<tr>
			<td>Tuberculin safety syringe with needle 25G 5/8&#34;</td>
			<td>CardinalHealth</td>
			<td>8881511235</td>
			<td>For heparin injection</td>
		</tr>
		<tr>
			<td>Veterinary General Surgery Set</td>
			<td>Skalar</td>
			<td>98-1275</td>
			<td>Surgery instruments including disection scisors and mosquito clamps</td>
		</tr>
		<tr>
			<td>Veterinary Micro Set</td>
			<td>Skalar</td>
			<td>98-1311</td>
			<td>Surgery instruments with microscisors used for carotid artery opening</td>
		</tr>
		<tr>
			<td>Working Heart Rat/Guinea Pig/Rabbit system</td>
			<td>Radnoti</td>
			<td>120101BEZ</td>
			<td>Modular working heart system modified for the needs of the protocol. Includes all the necesary tubbing, water jacketed reservoirs and valves, including 2 and 3 way stop cock</td>
		</tr>
		<tr>
			<td>Xylazine</td>
			<td>Bayer</td>
			<td></td>
			<td>Sedative. 20 mg/ml</td>
		</tr>
	</tbody>
</table>

pre-clinical model of cardiac donation after circulatory death

Die Nachfrage nach Herztransplantationen steigt; Dennoch ist die Verfügbarkeit von Organen aufgrund eines Mangels an geeigneten Spendern begrenzt. Organspende nach Kreislauftod (DCD) ist eine Lösung, um diese begrenzte Verfügbarkeit zu adressieren, aber aufgrund einer Zeit der längeren warmen Ischämie und des Risikos von Gewebeverletzungen ist ihre routinemäßige Anwendung bei Herztransplantationen selten zu sehen. In diesem Manuskript stellen wir ein detailliertes Protokoll zur Verfügung, das aktuelle klinische Praktiken im Kontext von DCD mit kontinuierlicher Überwachung der Herzfunktion nachahmt, so dass die Bewertung neuartiger kardioprotektive Strategien und Interventionen Ischämie-Reperfusionsverletzung.
In diesem Modell wird das DCD-Protokoll bei anästhesierten Lewis-Ratten initiiert, indem die Beatmung gestoppt wird, um den Kreislauftod auszulösen. Wenn der systolische Blutdruck unter 30 mmHg sinkt, wird die warme ischämische Zeit eingeleitet. Nach einer voreingestellten warmen ischämischen Periode werden die Herzen mit einer normothermen kardioplegischen Lösung gespült, beschafft und auf ein Langendorff ex vivo Herzperfusionssystem montiert. Nach 10 min anfänglicher Reperfusion und Stabilisierung wird die Herzrekonditionierung 60 min lang kontinuierlich mit hilfe der intraventrikulären Drucküberwachung evaluiert. Eine Herzverletzung wird durch Messung des Herztroponins T beurteilt und die Infarktgröße wird durch histologische Färbung quantifiziert. Die warme ischämische Zeit kann moduliert und auf die gewünschte Menge an strukturellen und funktionellen Schäden zugeschnitten werden. Dieses einfache Protokoll ermöglicht die Bewertung verschiedener kardioprotektive Konditionierungsstrategien, die zum Zeitpunkt der Kardioplegie, der anfänglichen Reperfusion und/oder während der Ex-vivo-Perfusion eingeführt wurden. Die aus diesem Protokoll gewonnenen Erkenntnisse können in großen Modellen reproduziert werden, was die klinische Übersetzung erleichtert.

Nach der Extubation sinkt der Blutdruck in einem vorhersagbaren Muster rapide (Abbildung 3). Die erwartete Todeszeit beträgt weniger als 5 min.
Abbildung 4 zeigt eine durchschnittliche Druck-/Zeitkurve zu Beginn der Instandsetzung nach 0, 10 und 15 min WIT. Die Kontraktile-Funktion wird sich im Laufe der Zeit verbessern. Die Verwendung kurzer WIT-Zeiträume wird es ermöglichen, dass sich die Kontraktilität wieder normalisiert, und ...

Watch this Scientific Journal Video about Pre-clinical Model of Cardiac Donation after Circulatory Death at JoVE.com

Pre-clinical Model of Cardiac Donation after Circulatory Death

Dieses Protokoll zeigt einen einfachen und flexiblen Ansatz für die Bewertung neuer Konditionierungsmittel oder Strategien zur Erhöhung der Durchführbarkeit von Herzspenden nach Kreislauftod.

Präklinisches Modell der Herzspende nach Kreislauftod

Cardiac transplantation demand is on the rise; nevertheless, organ availability is limited due to a paucity of suitable donors. Organ donation after ...

Dieses Protokoll zielt auf die frage der begrenzten Transplantationsorganversorgung ab, indem es die Bewertung verschiedener kardioprotektiver Konditionierungsstrategien erleichtert, um die Verwendung von Herzen zu ermöglichen, die nach dem Kreislauftod gespendet wurden. Legen Sie das Tier auf ein Heizkissen, das auf Medium eingestellt ist. Nachdem Sie eine fehlende Reaktion auf Zehenstiche bei einer anästhesierten erwachsenen Ratte bestätigt haben, legen Sie eine rektale Temperatursonde ein.Befestigen Sie einen transdermalen Pulsoximetersensor an einer Hinterpfote und bedecken Sie die Ratte mit einem saugfähigen Pad, um die Körpertemperatur aufrechtzuerhalten. Machen Sie einen drei- bis vier Zentimeter großen Mittellinien-Hautschnitt im Hals und verwenden Sie eine stumpfe, gekrümmte Schere, um das unterkutane Gewebe stumpf zu sezieren, um den rechten Sternohyoidmuskel freizulegen. Mit nicht-traumatischen Zangen, bewegen Sie den Muskel seitlich, bis die rechte Halsschlagader, Jugular vene, und Vagusnerv identifiziert werden.Verwenden Sie eine Schere, um den Vagusnerv sorgfältig von der Halsschlagader zu trennen. Injizieren Sie 2.000 internationale Einheiten pro Kilogramm Heparin in die rechte Jugularvene und üben Sie Druck auf die Injektionsstelle nach dem Nadelrückzug aus, um Blutlecks zu vermeiden. Verwenden Sie gekrümmte Zangen, um zwei 5-0 Seidennähte um die Halsschlagader zu übergeben.Befestigen Sie die distale Naht fest, um die Halsschlagader am überlegenen Aspekt der exponierten Arterie zu verschließen, die proximale Naht ungebunden zu halten und die Ratte unter ein Stereomikroskop zu stellen. Verwenden Sie dann eine Mikrochirurgie-Schere, um vorsichtig einen Ein-Millimeter-Schnitt über die vordere Wand der Halsschlagader zu machen, und legen Sie einen 22-Spur, ein Zoll geschlossenen intravenösen Katheter in Richtung des Aortenbogens ein. Die Verwendung von Vergrößerungs- und Mikrodissektionswerkzeugen ist für diese Schritte sehr zu empfehlen.Ein sauberes Feld kann durch Anspannung der proximalen Naht erreicht werden, um Blutungen zu vermeiden. Herzspende ist notwendig, um die ex vivo Cardioplegia Apparat zu montieren. Um das DCD-Protokoll zu initiieren, extubieren Sie zuerst das Tier, dann verwenden Sie Mückenzange, um die Luftröhre zu klemmen, beginnend mit der Agonalphase.Beginnen Sie mit der Zählung der funktionellen WIT, wenn der systolische Spitzenblutdruck unter 30 Millimeter Quecksilber oder Asystole oder Kammerflimmern fällt, je nachdem, was zuerst eintritt. Führen Sie am Ende von WIT eine mediale Sternotomie durch, wobei Sie den Thorax bei Bedarf mit einem Alm-Retraktor offen halten. Trennen Sie das Herz von den Lungenvenen und anderen thorakalen Strukturen, um die Kardiektomie zu vervollständigen.Tauchen Sie das Herz sofort in eiskalte Krebs-Lösung für einen schnellen Transport zum ex vivo-System ein. Verwenden Sie eine 2-0 Seidennaht, um die Aorta fest am Langendorff-Apparat zu fixieren, und öffnen Sie dann den Hahn. Öffnen Sie nun den Fluss zur Kanüle vollständig und beginnen Sie die anfängliche Reperfusions- und Stabilisierungszeit.Drehen Sie das Herz so, dass die Vorhöfe dem Drucksensor zugewandt sind, und sezieren Sie die Lungenvenen, um die linke ventrikuläre Vorhoföffnung bei Bedarf zu erweitern. Legen Sie als Nächstes einen Latexballon ein, der mit einem Drucksensor verbunden ist, um sicherzustellen, dass der Ballon vollständig im Ventrikel positioniert ist. Füllen Sie den Ballon langsam mit Kochsaline, bis der Diastolische Druck am Ende 15 Millimeter Quecksilber erreicht.Legen Sie die Schrittelektrode in das vordere Gesicht des Herzens ein, ohne die Herzkranzgefäße zu durchkreuzen. Sobald spontanes Schlagen beobachtet wird, stellen Sie das Tempo auf 300 Schläge pro Minute. Nach 10 Minuten Stabilisierung, initiieren Sie die kontinuierliche intraventrikuläre Druckmessung Aufzeichnung, um die Rekonditionierung sanieren und Assessment-Phase zu beginnen.Nach einer Stunde entfernen Sie das Herz aus dem Langendorff-Gerät und verwenden Sie eine gerade hochkohlenstoffhaltige Stahlklinge, um die Vorhöfe zu entfernen. Mit dem rechten Ventrikel nach unten schneiden Sie ein bis zwei Millimeter dicke querventrikuläre Scheiben. Verbrauchen Sie den rechten Ventrikel aus der dritten Scheibe des Gewebes, und fangen Sie den linken Ventrikel für nachgelagerte biochemische Analysen ein.Die restlichen Abschnitte in frisch zubereitete 5%TTC-Lösung in PBS für 10 Minuten bei 37 Grad Celsius untertauchen. Am nächsten Morgen die Proben zweimal in frisches PBS waschen und die Proben zur Bildgebung in frische PBS untertauchen. Lebensfähige Gewebe erscheinen ziegelrot in der Farbe.Nach der Extubation sinkt der Blutdruck in einem vorhersagbaren Muster rapide und die erwartete Todeszeit beträgt weniger als fünf Minuten. Hier werden durchschnittliche Druck- und Zeitkurven zu Beginn der Rekonditionierung nach Null, 10 und 15 Minuten WIT angezeigt. Die kontraktile Funktion wird sich im Laufe der Zeit verbessern, die Verwendung von kurzen Zeiträumen von WIT wird es ermöglichen, die Kontraktilität wieder normal zu machen, und morphologische Schäden werden nicht nachweisbar sein.Die Verwendung eines mit der Kardioplegie und in der Stabilisierungsphase hinzugefügten Konditionierungsmittels zeigte, dass die durch 15 Minuten WIT in diesem Modell verursachten Schäden durch kardioprotektive Mittel modulationsfähig sind. Diese Technik ermöglicht die vollständige Kontrolle aller relevanten Variablen und pharmakologischen und nichtpharmakologischen Interventionstests und kann in größeren Modellen reproduziert werden, was die klinische Übersetzung erleichtert.

pre-clinical-model-of-cardiac-donation-after-circulatory-death

Research

JoVE Journal

Medicine

Pre-clinical Evaluation of Tyrosine Kinase Inhibitors for Treatment of Acute Leukemia

Receptor tyrosine kinases have been implicated in the development and progression of many cancers, including both leukemia and solid tumors, and are attractive druggable therapeutic targets. Here we describe an efficient four-step strategy for pre-clinical evaluation of tyrosine kinase inhibitors (TKIs) in the treatment of acute leukemia. Initially, western blot analysis is used to confirm target inhibition in cultured leukemia cells. Functional activity is then evaluated using clonogenic assays in methylcellulose or soft agar cultures. Experimental compounds that demonstrate activity in cell culture assays are evaluated in vivo using NOD-SCID-gamma (NSG) mice transplanted orthotopically with human leukemia cell lines. Initial in vivo pharmacodynamic studies evaluate target inhibition in leukemic blasts isolated from the bone marrow. This approach is used to determine the dose and schedule of administration required for effective target inhibition. Subsequent studies evaluate the efficacy of the TKIs in vivo using luciferase expressing leukemia cells, thereby allowing for non-invasive bioluminescent monitoring of leukemia burden and assessment of therapeutic response using an in vivo bioluminescence imaging system. This strategy has been effective for evaluation of TKIs in vitro and in vivo and can be applied for identification of molecularly-targeted agents with therapeutic potential or for direct comparison and prioritization of multiple compounds.

Receptor tyrosine kinases are ectopically expressed in many cancers and have been identified as therapeutic targets in acute leukemia. This manuscript describes an efficient strategy for pre-clinical evaluation of tyrosine kinase inhibitors for the treatment of acute leukemia.

Präklinische Evaluierung von Tyrosinkinase-Inhibitoren für die Behandlung von akuter Leukämie

Receptor tyrosine kinases have been implicated in the development and progression of many cancers, including both leukemia and solid tumors, and are ...

Forschung

Medizin

Handling of the Cotton Rat in Studies for the Pre-clinical Evaluation of Oncolytic Viruses

Oncolytic viruses are a novel anticancer therapy with the ability to target tumor cells, while leaving healthy cells intact. For this strategy to be successful, recent studies have shown that involvement of the host immune system is essential. Therefore, oncolytic virotherapy should be evaluated within the context of an immunocompetent model. Furthermore, the study of antitumor therapies in tolerized animal models may better recapitulate results seen in clinical trials. Cotton rats, commonly used to study respiratory viruses, are an attractive model to study oncolytic virotherapy as syngeneic models of mammary carcinoma and osteosarcoma are well established. However, there is a lack of published information on the proper handling procedure for these highly excitable rodents. The handling and capture approach outlined minimizes animal stress to facilitate experimentation. This technique hinges upon the ability of the researcher to keep calm during handling and perform procedures in a timely fashion. Finally, we describe how to prepare cotton rat mammary tumor cells for consistent subcutaneous tumor formation, and how to perform intratumoral and intraperitoneal injections. These methods can be applied to a wide range of studies furthering the development of the cotton rat as a relevant pre-clinical model to study antitumor therapy.

Cotton rats are extremely excitable and have a strong flight-or-fight response. A handling method optimized to reduce the stress of the animals is described which will make cotton rats more accessible as a preclinical model.

Die Handhabung der Baumwolle Ratte in Studien für die präklinische Bewertung onkolytischer Viren

Oncolytic viruses are a novel anticancer therapy with the ability to target tumor cells, while leaving healthy cells intact.  For this strategy to be ...

Pre-clinical Orthotopic Murine Model of Human Prostate Cancer

To study the multifaceted biology of prostate cancer, pre-clinical in vivo models offer a range of options to uncover critical biological information about this disease. The human orthotopic prostate cancer xenograft mouse model provides a useful alternative approach for understanding the specific interactions between genetically and molecularly altered tumor cells, their organ microenvironment, and for evaluation of efficacy of therapeutic regimens. This is a well characterized model designed to study the molecular events of primary tumor development and it recapitulates the early events in the metastatic cascade prior to embolism and entry of tumor cells into the circulation. Thus it allows elucidation of molecular mechanisms underlying the initial phase of metastatic disease. In addition, this model can annotate drug targets of clinical relevance and is a valuable tool to study prostate cancer progression. In this manuscript we describe a detailed procedure to establish a human orthotopic prostate cancer xenograft mouse model.

Prostate cancer is the second most common cause of cancer-related deaths in the United States. An orthotopic cancer model provides a useful approach to understand the biology of prostate cancer and to evaluate the efficacy of therapeutic regimens. This protocol describes detailed steps necessary to establish an orthotopic prostate cancer mouse model.

Präklinische Orthotopische Mausmodell der menschlichen Prostata-Krebs

To study the multifaceted biology of prostate cancer, pre-clinical in vivo models offer a range of options to uncover critical biological information ...

Use of a Low-flow Digital Anesthesia System for Mice and Rats

A traditional vaporizer depends on flowing gas and atmospheric pressure for passive anesthetic vaporization. Newly developed direct injection vaporizers utilize a syringe pump to directly administer volatile anesthetics into a gas stream. Unlike a traditional vaporizer, it can be used at very low flow rates, making it ideal for use on mice and rats. 
The equipment's capability to use low flow rates could result in a substantial cost savings due to the reduced need for anesthetic agents, compressed gas, and charcoal scavenging filters1. A lower flow rate means less waste of anesthetic gas and likely reduces the risk of anesthetic exposure to laboratory personnel. Thus, the high levels of precision and safety associated with direct injection vaporizers, along with a reduced need for anesthetic agents, compressed gas, and charcoal filters are beneficial for research requiring small animal anesthesia. 
The goal of this protocol is to demonstrate the use of a syringe-driven direct injection vaporizer as part of a digital, low-flow anesthesia system. The direct injection vaporizer is capable of accurately delivering anesthesia at very low flow rates compared to a traditional vaporizer, making it a promising alternative for controlled gas anesthetic delivery to rodents.

Here, we present a protocol to more safely and efficiently administer anesthetic gas to mice using a digital, low flow anesthesia system utilizing a syringe-driven direct injection vaporizer.

Die Verwendung eines Low-Flow-Digital-Anästhesiesystem für Mäuse und Ratten

A traditional vaporizer depends on flowing gas and atmospheric pressure for passive anesthetic vaporization. Newly developed direct injection vaporizers ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

Messung der isometrischen Kontraktilität der Maus mesenterialen Arterie mit Draht Myographie

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

Implantation of hiPSC-derived Cardiac-muscle Patches after Myocardial Injury in a Guinea Pig Model

Due to the limited regeneration capacity of the heart in adult mammals, myocardial infarction results in an irreversible loss of cardiomyocytes. This loss of relevant amounts of heart muscle mass can lead to the heart failure. Besides heart transplantation, there is no curative treatment option for the end-stage heart failure. In times of organ donor shortage, organ independent treatment modalities are needed. Left-ventricular assist devices are a promising therapy option, however, especially as destination therapy, limited by its side-effects like stroke, infections and bleedings. In recent years, several cardiac repair strategies including stem cell injection, cardiac progenitors or myocardial tissue engineering have been investigated. Recent improvements in cell biology allow for the differentiation of large amounts of cardiomyocytes derived from human induced pluripotent stem cells (iPSC). One of the cardiac repair strategies currently under evaluation is to transplant artificial heart tissue. Engineered heart tissue (EHT) is a three-dimensional in vitro created cardiomyocyte network, with functional properties of native heart tissue. We have created EHT-patches from hiPSC derived cardiomyocytes. Here we present a protocol for the induction of left ventricular myocardial cryoinjury in a guinea pig, followed by implantation of hiPSC derived EHT on the left ventricular wall.

Here we present a protocol for the induction of left ventricular cryoinjury followed by the implantation of a cardiac muscle patch, derived from human iPS-cell cardiomyocytes in a guinea pig model.

Implantation von HiPSC abgeleitete Herzmuskel Patches nach Myokard-Schädigung in einem Meerschweinchen-Modell

Due to the limited regeneration capacity of the heart in adult mammals, myocardial infarction results in an irreversible loss of cardiomyocytes. This loss ...

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.

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.

Ein präklinisches Porcine-Modell der orthotopischen Herztransplantation

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

Invasive Hemodynamic Assessment for the Right Ventricular System and Hypoxia-Induced Pulmonary Arterial Hypertension in Mice

Pulmonary arterial hypertension (PAH) is a chronic and severe cardiopulmonary disorder. Mice are a popular animal model used to mimic this disease. However, the evaluation of right ventricular pressure (RVP) and pulmonary artery pressure (PAP) remains technically challenging in mice. RVP and PAP are more difficult to measure than left ventricular pressure because of the anatomical differences between the left and right heart systems. In this paper, we describe a stable right heart hemodynamic measurement method and its validation using healthy and PAH mice. This method is based on open-chest surgery and mechanical ventilation support. It is a complicated procedure compared to closed chest procedures. While a well-trained surgeon is required for this surgery, the advantage of this procedure is that it can generate both RVP and PAP parameters at the same time, so it is a preferable procedure for the evaluation of PAH models.

Here, we present a protocol to perform an invasive hemodynamic assessment of the right ventricle and pulmonary artery in mice using an open-chest surgery approach.

Invasive hämodynamische Beurteilung des rechten ventrikulären Systems und Hypoxie-induzierte pulmonale arterielle Hypertonie bei Mäusen

Pulmonary arterial hypertension (PAH) is a chronic and severe cardiopulmonary disorder. Mice are a popular animal model used to mimic this disease. ...

A Pre-clinical Rat Model for the Study of Ischemia-reperfusion Injury in Reconstructive Microsurgery

Ischemia-reperfusion injury is the main cause of flap failure in reconstructive microsurgery. The rat is the preferred preclinical animal model in many areas of biomedical research due to its cost-effectiveness and its translation to humans. This protocol describes a method to create a preclinical free skin flap model in rats with ischemia-reperfusion injury. The described 3 cm x 6 cm rat free skin flap model is easily obtained after the placement of several vascular ligatures and the section of the vascular pedicle. Then, 8 h after the ischemic insult and completion of the microsurgical anastomosis, the free skin flap develops the tissue damage. These ischemia-reperfusion injury-related damages can be studied in this model, making it a suitable model for evaluating therapeutic agents to address this pathophysiological process. Furthermore, two main monitoring techniques are described in the protocol for the assessment of this animal model: transit-time ultrasound technology and laser speckle contrast analysis.

Here, we describe a pre-clinical animal model for studying the pathophysiology of ischemia-reperfusion injury in reconstructive microsurgery. This free skin flap model based on the superficial caudal epigastric vessels in the rat may also allow for the evaluation of different therapies and compounds to counteract ischemia-reperfusion injury-related damage.

Ein präklinisches Rattenmodell zur Untersuchung von Ischämie-Reperfusionsverletzungen in der rekonstruktiven Mikrochirurgie

Ischemia-reperfusion injury is the main cause of flap failure in reconstructive microsurgery. The rat is the preferred preclinical animal model in many ...

Use of an Integrated Low-Flow Anesthetic Vaporizer, Ventilator, and Physiological Monitoring System for Rodents

Low-flow digital vaporizers commonly utilize a syringe pump to directly administer volatile anesthetics into a stream of carrier gas. Per animal welfare recommendations, animals are warmed and monitored during procedures requiring anesthesia. Common anesthesia and physiological monitoring equipment include gas tanks, anesthetic vaporizers and stands, warming controllers and pads, mechanical ventilators, and pulse oximeters. A computer is also necessary for data collection and to run equipment software. In smaller spaces or when performing field work, it can be challenging to configure all this equipment in limited space.
The goal of this protocol is to demonstrate best practices for use of a low-flow digital vaporizer using both compressed oxygen and room air, along with an integrated mechanical ventilator, pulse oximeter, and far infrared warming as an all-inclusive anesthesia and physiological monitoring suite ideal for rodents.

Here, we present a protocol to safely and effectively administer anesthetic gas to mice using a digital, low flow anesthesia system with integrated ventilator and physiological monitoring modules.

Einsatz eines integrierten Low-Flow Anesthetic Vaporizer, Ventilator und Physiologisches Überwachungssystem für Nagetiere

Low-flow digital vaporizers commonly utilize a syringe pump to directly administer volatile anesthetics into a stream of carrier gas. Per animal welfare ...