Ce travail est soutenu par la Fondation Leducq à travers le réseau Caerus (RJH), par le NIH R01 HL093183, HL088434, HL071763, HL080498, HL083156 et P20HL100396 (RJH). DL a été soutenu par la Fondation allemande de recherche. Nous tenons à remercier Lauren Leonardson pour son aide technique dans ce projet.

1. Infarctus du myocarde création <ol><li> Femme porcs Yorkshire (20 kg) à partir des industries animales Biotech (Doylestown, PA) sont pré-médicamenteux en utilisant télazol (tilétamine / zolazépam) (8,0 mg / kg) et la buprénorphine (0,6 mg). </li><li> Les animaux sont intubés et ventilés, puis avec de l&#39;oxygène à 100%. L&#39;anesthésie générale est maintenue avec du propofol (5-8mg/kg/hr) pendant toute la procédure. </li><li> Le site du fémur est préparé avec de l&#39;alcool isopropylique à 70% suivie par la polyvidone iodée. Une ponction percutanée permet d&#39;accéder à l&#39;artère et de placement de la gaine. Après l&#39;insertion gaine, 3000-5000 UI d&#39;héparine est administrée IV pour maintenir un temps de coagulation activé de 250-300 secondes. </li><li> Un bolus d&#39;amiodarone (75 mg) est administrée par voie intramusculaire. L&#39;injection intraveineuse d&#39;un sac de 1000 ml saline avec l&#39;amiodarone (75 mg), l&#39;atropine (2 mg), et l&#39;acétate de potassium (20 mEq) est démarré et maintenu en utilisant un taux de 150 ml / h pendant toute la procédure. </li><li> A 7 Fr crosse de hockey cathéter (Cordis; Miami, Floride) est avancé à l&#39;artère coronaire gauche et une coronarographie est réalisée. Un fil guide 0.014 pouces (Abbott, Park, IL) est avancé dans la partie antérieure artère interventriculaire (DAL). Un 8-mm de long, 4,0 mm Voyager over-the-wire ballon (Abbott, Park, IL) est avancé et positionné à l&#39;IVA proximale, et le ballon est gonflé à coronaires 3 atm. L&#39;occlusion de l&#39;artère interventriculaire antérieure est confirmée par angiographie. </li><li> Avec l&#39;électrocardiogramme en continu et la surveillance hémodynamique, l&#39;occlusion est maintenue pendant 90 minutes. Arythmies ventriculaires malignes peuvent parfois se produire dans les 40 premières minutes, dans ce cas, la livraison immédiate des chocs directs actuels à partir de 200 J à 300 J sont administrés. Habituellement, il ya deux périodes de pointe de l&#39;arythmie, l&#39;un entre 2 et 10 minutes et l&#39;autre entre 25 et 35 minutes après le gonflage du ballon. Surveillance diligente et défibrillation immédiate sont la clé de la création d&#39;infarctus du myocarde avec succès, car il ya incidence de plus de 95% d&#39;entrer au sein de la fibrillation ventriculaire les délais indiqués. Si la défibrillation immédiate et appropriée est effectuée, le taux de survie est très élevé. Si la pression artérielle diminue de moins de 80 mmHg, charge volumique est accomplie avec rapidité perfusion de solution saline par voie intraveineuse. </li><li> Le ballon est dégonflé et tous les périphériques sont supprimés. </li><li> La gaine est retirée de l&#39;artère fémorale, l&#39;anesthésie est retenu, et l&#39;animal est surveillé pendant la récupération. Salines intraveineuse avec l&#39;amiodarone, atropine et de la perfusion acétate de potassium est diminué à 50 ml / h et est donné pendant la nuit. Un des injections intramusculaires de Nitroglycerline et Furoscemide est administré. Puis l&#39;animal est autorisé à récupérer et minutieuse surveillance ECG est poursuivie pendant au moins 3 heures. </li><li> L&#39;aspirine (200 mg) PO sera donnée pendant cinq jours. </li></ol> Les complications communes durant et après la procédure de création d&#39;infarctus du myocarde sont la dissection coronarienne, hématome site de ponction, l&#39;arythmie et l&#39;insuffisance cardiaque. Un affiner la technique et la surveillance minutieuse est nécessaire pour prévenir et gérer ces complications. 2. Injection du virus <ol><li> Les animaux sont préparés comme indiqué ci-dessus dans la création infarctus du myocarde steps1-3. La même dose d&#39;héparine doit être donnée à prévenir la thrombose lors de l&#39;injection du virus. </li><li> Un cathéter bâton de hockey 5Fr (Cordis; Miami, Floride) est avancé à l&#39;artère coronaire gauche. Après l&#39;angiographie, deux des fils-guides 0,014 pouces sont avancés, l&#39;un dans la DAL et un dans l&#39;artère coronaire gauche circonflexe (LCX) pour fixer la position du cathéter. </li><li> Le sang artériel (-20 ml) est tiré du cathéter. Pour éviter l&#39;ischémie pendant la perfusion, la moitié du sang dilué est utilisé pour la solution. Le virus est mélangé avec 10 ml de sang et dilué à 20 ml avec une solution saline. Le solde de 10 ml est également dilué à 20 ml et utilisé offrir virus résiduel dans la lumière du cathéter. </li><li> Nitroglycérine intraveineuse (20 mg / min) est lancé à travers la veine de l&#39;oreille. </li><li> L&#39;extrémité du cathéter est placé à l&#39;appareil principal proximal gauche. La pression à travers le cathéter est contrôlé pour s&#39;assurer que le cathéter n&#39;est pas de calage. </li><li> Le sang du virus mixtes (15 ml) est injectée par le cathéter plus de 15 minutes en utilisant une pompe à perfusion dans l&#39;artère coronaire gauche. Après 15 minutes, la seringue est échangé à la chasse d&#39;eau, et la perfusion est poursuivie pendant 10 minutes. </li><li> Un fil et un cathéter est fixé à l&#39;artère coronaire droite la même manière que l&#39;artère coronaire gauche. </li><li> Le solde de 5 ml de sang mélangé avec le virus est injecté dans l&#39;artère coronaire droite pendant 5 minutes suivies de 10 minutes de rinçage. </li></ol> 3. Cas représentatif La figure 1 montre la ventriculographie gauche à 1 mois après la création d&#39;infarctus du myocarde. Le mouvement du ventricule gauche mur est compromise de manière significative avec une fraction d&#39;éjection (FE) de31%. La figure 2 montre les sections du ventricule gauche à 1 mois. Une grande cicatrice de la base au sommet de la paroi antérieure est vu. L&#39;EF moyenne à un mois après un infarctus du myocarde dans ce modèle est de 36% et le volume moyen de fin de diastole est de 83 ml, alors que la moyenne de ceux de l&#39;animal de même taille, sans un infarctus du myocarde est de 68% et 42 ml, respectivement. La figure 3 montre l&#39;expression de la bêta-gal dans la région de zone ventriculaire gauche frontalière un mois après l&#39;injection de virus adéno-associés (2 x 10 12 particules DNAse resitant des rAAV1 CMV Lac-Z). Une expression accrue a été trouvé dans la zone injectée le virus. <img src="/files/ftp_upload/2778/2778fig1.jpg" alt="Figure 1" /> La fraction d&#39;éjection Figure 1. Mesurée à partir ventriculographie gauche 1 mois après l&#39;infarctus du myocarde de création. Ventricule gauche est dilaté à 92,5 ml et mouvement de la paroi antérieure est significativement altérée. <img src="/files/ftp_upload/2778/2778fig2.jpg" alt="Figure 2" /> Figure 2. Les sections du ventricule gauche moins 1 mois après l&#39;infarctus du myocarde de création, taché par du chlorure de triphényl tétrazolium pour délimiter la zone cicatricielle. Une grande cicatrice de la base au sommet de la paroi antérieure est vu. <img src="/files/ftp_upload/2778/2778fig3.jpg" alt="Figure 3" /> Figure 3. Bêta-gal expression induite par le virus adéno-associés (2 x 10 12 particules DNase résistantes de rAAV1 CMV Lac-Z). Une expression accrue se trouve dans la zone de la zone frontière entre l&#39;infarctus et le tissu normal, un mois après l&#39;injection du virus intracoronaire. Photo de droite inférieure montre le contrôle négatif, où seule une solution saline normale a été injecté dans les artères coronaires utilisant le même protocole.

<ol>
	<li>Perez de Prado, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Closed-chest+experimental+porcine+model+of+acute+myocardial+infarction-reperfusion.">Closed-chest experimental porcine model of acute myocardial infarction-reperfusion.</a> J Pharmacol Toxicol Methods. 60, 301-306 (2009).</li><li>Angeli, F. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Left+ventricular+remodeling+after+myocardial+infarction:+characterization+of+a+swine+model+on+beta-blocker+therapy.">Left ventricular remodeling after myocardial infarction: characterization of a swine model on beta-blocker therapy.</a> Comp Med. 59, 272-279 (2009).</li><li>Urdaneta, F., Lobato, E. B., Kirby, D. S. &. a. m. p. ;. a. m. p., Sidi, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Treating+myocardial+stunning+randomly,+with+either+propofol+or+isoflurane+following+transient+coronary+occlusion+and+reperfusion+in+pigs.">Treating myocardial stunning randomly, with either propofol or isoflurane following transient coronary occlusion and reperfusion in pigs.</a> Ann Card Anaesth. 12, 113-121 (2009).</li><li>Jaski, 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=Calcium+upregulation+by+percutaneous+administration+of+gene+therapy+in+cardiac+disease+(CUPID+Trial),+a+first-in-human+phase+1/2+clinical+trial.">Calcium upregulation by percutaneous administration of gene therapy in cardiac disease (CUPID Trial), a first-in-human phase 1/2 clinical trial.</a> J Card Fail. 15, 171-181 (2009).</li></ol>

Tous les auteurs ne sont pas des associations commerciales qui pourraient poser comme un conflit d&#39;intérêts en rapport avec le manuscrit.

Parmi les différentes causes de l&#39;insuffisance cardiaque, l&#39;ischémie est de loin la plus fréquente dans l&#39;étiologie. Pour surmonter cette grave maladie, différentes approches thérapeutiques sont en cours de développement. Évaluations soigneuses des thérapies sont nécessaires avant que ces thérapies prometteuses sont peut être appliquée à la clinique. Par conséquent, l&#39;établissement d&#39;un modèle d&#39;insuffisance cardiaque ischémique, qui représente les conditions cliniques des pati...

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		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Telazol</td>
<td>&nbsp;</td>
<td>Fort Dodge Animal Health</td>
<td>0856-9050-49</td>
<td>&nbsp;</td>
<tr>
<td>Propofol</td>
<td>&nbsp;</td>
<td>APP Pharmaceuticals</td>
<td>63323-270-20</td>
<td>&nbsp;</td>
<tr>
<td>Heparin</td>
<td>&nbsp;</td>
<td>APP Pharmaceuticals</td>
<td>63323-540-11</td>
<td>&nbsp;</td>
<tr>
<td>Amiodarone</td>
<td>&nbsp;</td>
<td>APP Pharmaceuticals</td>
<td>63323-616-03</td>
<td>&nbsp;</td>
<tr>
<td>Atropin Sulfate</td>
<td>&nbsp;</td>
<td>Med-Pharmex</td>
<td>54925-063-10</td>
<td>&nbsp;</td>
<tr>
<td>Potassium acetate</td>
<td>&nbsp;</td>
<td>American Regent, Inc.</td>
<td>2053-25</td>
<td>&nbsp;</td>
<tr>
<td>Nitroglycerine</td>
<td>&nbsp;</td>
<td>American Regent, Inc.</td>
<td>4810-25 </td>
<td>&nbsp;</td>
<tr>
<td>Salix (Furosemide)</td>
<td>&nbsp;</td>
<td>Intervet Inc.</td>
<td>710461</td>
<td>&nbsp;</td>
<tr>
<td>5 Fr Hockey stick catheter</td>
<td>&nbsp;</td>
<td>Cordis</td>
<td>556-278-0L</td>
<td>&nbsp;</td>
<tr>
<td>7 Fr Hockey stick catheter</td>
<td>&nbsp;</td>
<td>Cordis</td>
<td>778-278-00</td>
<td>&nbsp;</td>
<tr>
<td>Coronary wire</td>
<td>&nbsp;</td>
<td>Abbott Laboratories</td>
<td>1044588</td>
<td>&nbsp;</td>
<tr>
<td>Coronary balloon</td>
<td>&nbsp;</td>
<td>Abbott Laboratories</td>
<td>1009447-08</td>
<td>&nbsp;</td>
<tr>
<td>8Fr sheath</td>
<td>&nbsp;</td>
<td>TERUMO</td>
<td>RSS801</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

gene transfer for ischemic heart failure in a preclinical model

Diverses nouvelles technologies sont développées pour les patients atteints d&#39;insuffisance cardiaque. Bien établies des évaluations précliniques sont nécessaires pour déterminer leur efficacité et leur sécurité. La thérapie génique utilisant des vecteurs viraux est l&#39;une des approches les plus prometteuses pour le traitement des maladies cardiaques. La livraison de divers gènes viraux différents en changeant le gène transporteur a incommensurable potentiel thérapeutique. Dans cette vidéo, le processus complet d&#39;un modèle animal de la création d&#39;une insuffisance cardiaque suivie par transfert de gènes est présentée en utilisant un modèle porcin. Tout d&#39;abord, l&#39;infarctus du myocarde est créé par l&#39;occlusion proximale antérieure descendante gauche l&#39;artère coronaire. Résultats de remodelage cardiaque dans l&#39;insuffisance cardiaque chronique. Unique à notre modèle est une cicatrice assez importante qui reflète véritablement les patients atteints d&#39;insuffisance cardiaque grave et qui nécessitent un traitement agressif pour des résultats positifs. Après la création du myocarde infarctus et le développement de tissu cicatriciel, une injection intracoronaire de virus est démontrée avec une perfusion de nitroglycérine simultanée. Notre méthode d&#39;injection permet le transfert de gène simple et efficace avec l&#39;expression génétique. Cette combinaison d&#39;un modèle d&#39;infarctus du myocarde porcine avec la livraison intracoronarienne virus s&#39;est avéré être une méthodologie cohérente et reproductible, ce qui permet non seulement de tester l&#39;effet des gènes individuels, mais aussi de comparer l&#39;efficacité de nombreux gènes comme des candidats thérapeutiques.

Watch this Scientific Journal Video about Gene Transfer for Ischemic Heart Failure in a Preclinical Model at JoVE.com

Gene Transfer for Ischemic Heart Failure in a Preclinical Model

Une méthode de transfert de gènes pour le traitement de l&#39;insuffisance cardiaque ischémique est décrit en utilisant un modèle porcin d&#39;infarctus du myocarde. Notre méthode simple et reproductible nous permet de facilement évaluer l&#39;efficacité des transferts de gènes différents avec une manière très simple et reproductible.

Transfert de gènes pour insuffisance cardiaque ischémique dans un modèle préclinique

Various emerging technologies are being developed for patients with heart failure. Well-established preclinical evaluations are necessary to determine ...

gene-transfer-for-ischemic-heart-failure-in-a-preclinical-model

Research

JoVE Journal

Medicine

12.8K vues.  Mount Sinai School of Medicine. Une méthode de transfert de gènes pour le traitement de l'insuffisance cardiaque ischémique est décrit en utilisant un modèle porcin d'infarctus du myocarde. Notre méthode simple et reproductible nous permet de facilement évaluer l'efficacité des transferts de gènes différents avec une manière très simple et reproductible.

Vidéo: Gene Transfer for Ischemic Heart Failure in a Preclinical Model

A Simplified Technique for Producing an Ischemic Wound Model

One major obstacle in current diabetic wound research is a lack of an ischemic wound model that can be safely used in diabetic animals. Drugs that work well in non-ischemic wounds may not work in human diabetic wounds because vasculopathy is one major factor that hinders healing of these wounds. We published an article in 2007 describing a rabbit ear ischemic wound model created by a minimally invasive surgical technique. Since then, we have further simplified the procedure for easier operation. On one ear, three small skin incisions were made on the vascular pedicles, 1-2 cm from the ear base. The central artery was ligated and cut along with the nerve. The whole cranial bundle was cut and ligated, leaving only the caudal branch intact. A circumferential subcutaneous tunnel was made through the incisions, to cut subcutaneous tissues, muscles, nerves, and small vessels. The other ear was used as a non-ischemic control. Four wounds were made on the ventral side of each ear. This technique produces 4 ischemic wounds and 4 non-ischemic wounds in one animal for paired comparisons. After surgery, the ischemic ear was cool and cyanotic, and showed reduced movement and a lack of pulse in the ear artery. Skin temperature of the ischemic ear was 1-10 &deg;C lower than that on the normal ear and this difference was maintained for more than one month. Ear tissue high-energy phosphate contents were lower in the ischemic ear than the control ear. Wound healing times were longer in the ischemic ear than in the non-ischemic ear when the same treatment was used. The technique has now been used on more than 80 rabbits in which 23 were diabetic (diabetes time ranging from 2 weeks to 2 years). No single rabbit has developed any surgical complications such as bleeding, infection, or rupture in the skin incisions. The model has many advantages, such as little skin disruption, longer ischemic time, and higher success rate, when compared to many other models. It can be safely used in animals with reduced resistance, and can also be modified to meet different testing requirements.

We have developed a minimally invasive technique to create a rabbit ischemic ear wound model by dividing the central artery and nerve and the cranial neurovascular bundle. A subcutaneous tunnel then cuts all subcutaneous tissues. This procedure causes minimal skin disruption and can be safely used in diabetic animals.

Une technique simplifiée pour produire un modèle de plaie ischémique

One major obstacle in current diabetic wound research is a lack of an ischemic wound model that can be safely used in diabetic animals. Drugs that work ...

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Médecine

Permanent Ligation of the Left Anterior Descending Coronary Artery in Mice: A Model of Post-myocardial Infarction Remodelling and Heart Failure

Heart failure is a syndrome in which the heart fails to pump blood at a rate commensurate with cellular oxygen requirements at rest or during stress. It is characterized by fluid retention, shortness of breath, and fatigue, in particular on exertion. Heart failure is a growing public health problem, the leading cause of hospitalization, and a major cause of mortality. Ischemic heart disease is the main cause of heart failure.
Ventricular remodelling refers to changes in structure, size, and shape of the left ventricle. This architectural remodelling of the left ventricle is induced by injury (e.g., myocardial infarction), by pressure overload (e.g., systemic arterial hypertension or aortic stenosis), or by volume overload. Since ventricular remodelling affects wall stress, it has a profound impact on cardiac function and on the development of heart failure. A model of permanent ligation of the left anterior descending coronary artery in mice is used to investigate ventricular remodelling and cardiac function post-myocardial infarction. This model is fundamentally different in terms of objectives and pathophysiological relevance compared to the model of transient ligation of the left anterior descending coronary artery. In this latter model of ischemia/reperfusion injury, the initial extent of the infarct may be modulated by factors that affect myocardial salvage following reperfusion. In contrast, the infarct area at 24 hr after permanent ligation of the left anterior descending coronary artery is fixed. Cardiac function in this model will be affected by 1) the process of infarct expansion, infarct healing, and scar formation; and 2) the concomitant development of left ventricular dilatation, cardiac hypertrophy, and ventricular remodelling.
Besides the model of permanent ligation of the left anterior descending coronary artery, the technique of invasive hemodynamic measurements in mice is presented in detail.

Heart failure is the leading cause of hospitalization and a major cause of mortality. A model of permanent ligation of the left anterior descending coronary artery in mice is applied to investigate ventricular remodelling and cardiac dysfunction post-myocardial infarction. The technique of invasive hemodynamic measurements in mice is presented.

Ligature permanente de l&#39;artère interventriculaire antérieure coronaire chez la souris: Un modèle de post-infarctus du myocarde et de remodelage de l&#39;insuffisance cardiaque

Heart failure is a syndrome in which the heart fails to pump blood at a rate commensurate with cellular oxygen requirements at rest or during stress. It ...

In Vivo Gene Transfer to the Rabbit Common Carotid Artery Endothelium

The goal of this method is to introduce a transgene into the endothelium of isolated segments of both rabbit common carotid arteries. The method achieves focal endothelial-selective transgenesis, thereby allowing an investigator to determine the biological roles of endothelial-expressed transgenes and to quantify the in vivo transcriptional activity of DNA sequences in large artery endothelial cells. The method uses surgical isolation of rabbit common carotid arteries and an arteriotomy to deliver a transgene-expressing viral vector into the arterial lumen. A short incubation period of the vector in the lumen, with subsequent aspiration of the lumen contents, is sufficient to achieve efficient and durable expression of the transgene in the endothelium, with no detectable transduction or expression outside of the isolated arterial segment. The method allows assessment of the biological activities of transgene products both in normal arteries and in models of human vascular disease, while avoiding systemic effects that could be caused either by targeting gene delivery to other sites (e.g. the liver) or by the alternative approach of delivering genetic constructs to the endothelium by germ line transgenesis. Application of the method is limited by the need for a skilled surgeon and anesthetist, a well-equipped operating room, the costs of purchasing and housing rabbits, and the need for expertise in gene-transfer vector construction and use. Results obtained with this method include: transgene-related alterations in arterial structure, cellularity, extracellular matrix, or vasomotor function; increases or reductions in arterial inflammation; alterations in vascular cell apoptosis; and progression, retardation, or regression of diseases such as intimal hyperplasia or atherosclerosis. The method also allows measurement of the ability of native and synthetic DNA regulatory sequences to alter transgene expression in endothelial cells, providing results that include: levels of transgene mRNA, levels of transgene protein, and levels of transgene enzymatic activity.

In Vivo Gene Transfer to the Rabbit Common Carotid Artery Endothelium

This method is to introduce a transgene into the endothelium of rabbit carotid arteries. Introduction of the transgene allows the assessment of the biological role of the transgene product either in normal arteries or disease models. The method is also useful for measuring activity of DNA regulatory sequences.

In Vivo Transfert de gènes à l’endothélium de l’artère carotide commune lapin

The goal of this method is to introduce a transgene into the endothelium of isolated segments of both rabbit common carotid arteries. The method achieves ...

Tachycardia-Induced Cardiomyopathy As a Chronic Heart Failure Model in Swine

A stable and reliable model of chronic heart failure is required for many experiments to understand hemodynamics or to test effects of new treatment methods. Here, we present such a model by tachycardia-induced cardiomyopathy, which can be produced by rapid cardiac pacing in swine.
A single pacing lead is introduced transvenously into fully anaesthetized healthy swine, to the apex of the right ventricle, and fixated. Its other end is then tunneled dorsally to the paravertebral region. There, it is connected to an in-house modified heart pacemaker unit that is then implanted in a subcutaneous pocket. 
After 4 - 8 weeks of rapid ventricular pacing at rates of 200 - 240 beats/min, physical examination revealed signs of severe heart failure - tachypnea, spontaneous sinus tachycardia, and fatigue. Echocardiography and X-ray showed dilation of all heart chambers, effusions, and severe systolic dysfunction. These findings correspond well to decompensated dilated cardiomyopathy and are also preserved after the cessation of pacing.
This model of tachycardia-induced cardiomyopathy can be used for studying the pathophysiology of progressive chronic heart failure, especially hemodynamic changes caused by new treatment modalities like mechanical circulatory supports. This methodology is easy to perform and the results are robust and reproducible.

Here, we present a protocol to produce tachycardia-induced cardiomyopathy in swine. This model represents a potent way to study the hemodynamics of progressive chronic heart failure and the effects of applied treatment.

Cardiomyopathie induite par la tachycardie comme un modèle de l’insuffisance cardiaque chronique chez le porc

A stable and reliable model of chronic heart failure is required for many experiments to understand hemodynamics or to test effects of new treatment ...

Isolation of Atrial Cardiomyocytes from a Rat Model of Metabolic Syndrome-related Heart Failure with Preserved Ejection Fraction

In this article, we describe an optimized, Langendorff-based procedure for the isolation of single-cell atrial cardiomyocytes (ACMs) from a rat model of metabolic syndrome (MetS)-related heart failure with preserved ejection fraction (HFpEF). The prevalence of MetS-related HFpEF is rising, and atrial cardiomyopathies associated with atrial remodeling and atrial fibrillation are clinically highly relevant as atrial remodeling is an independent predictor of mortality. Studies with isolated single-cell cardiomyocytes are frequently used to corroborate and complement in vivo findings. Circulatory vessel rarefication and interstitial tissue fibrosis pose a potentially limiting factor for the successful single-cell isolation of ACMs from animal models of this disease.
We have addressed this issue by employing a device capable of manually regulating the intraluminal pressure of cardiac cavities during the isolation procedure, substantially increasing the yield of morphologically and functionally intact ACMs. The acquired cells can be used in a variety of different experiments, such as cell culture and functional Calcium imaging (i.e., excitation-contraction-coupling).
We provide the researcher with a step-by-step protocol, a list of optimized solutions, thorough instructions to prepare the necessary equipment, and a comprehensive troubleshooting guide. While the initial implementation of the procedure might be rather difficult, a successful adaptation will allow the reader to perform state-of-the-art ACM isolations in a rat model of MetS-related HFpEF for a broad spectrum of experiments.

Here, we describe an optimized, Langendorff-based procedure for the isolation of single-cell atrial cardiomyocytes from a rat model of metabolic syndrome-related heart failure with preserved ejection fraction. A manual regulation of intraluminal pressure of cardiac cavities is implemented to yield functionally intact myocytes suitable for excitation-contraction-coupling studies.

Isolement des Cardiomyocytes auriculaires d’un modèle de Rat de métaboliques liées Syndrome une insuffisance cardiaque avec Fraction d’éjection préservée

In this article, we describe an optimized, Langendorff-based procedure for the isolation of single-cell atrial cardiomyocytes (ACMs) from a rat model of ...

Induction and Phenotyping of Acute Right Heart Failure in a Large Animal Model of Chronic Thromboembolic Pulmonary Hypertension

The development of acute right heart failure (ARHF) in the context of chronic pulmonary hypertension (PH) is associated with poor short-term outcomes. The morphological and functional phenotyping of the right ventricle is of particular importance in the context of hemodynamic compromise in patients with ARHF. Here, we describe a method to induce ARHF in a previously described large animal model of chronic PH, and to phenotype, dynamically, right ventricular function using the gold standard method (i.e., pressure-volume PV loops) and with a non-invasive clinically available method (i.e., echocardiography). Chronic PH is first induced in pigs by left pulmonary artery ligation and right lower lobe embolism with biological glue once a week for 5 weeks. After 16 weeks, ARHF is induced by successive volume loading using saline followed by iterative pulmonary embolism until the ratio of the systolic pulmonary pressure over systemic pressure reaches 0.9 or until the systolic systemic pressure decreases below 90 mmHg. Hemodynamics are restored with dobutamine infusion (from 2.5 &#181;g/kg/min to 7.5 &#181;g/kg/min). PV-loops and echocardiography are performed during each condition. Each condition requires around 40 minutes for induction, hemodynamic stabilization and data acquisition. Out of 9 animals, 2 died immediately after pulmonary embolism and 7 completed the protocol, which illustrates the learning curve of the model. The model induced a 3-fold increase in mean pulmonary artery pressure. The PV-loop analysis showed that ventriculo-arterial coupling was preserved after volume loading, decreased after acute pulmonary embolism and was restored with dobutamine. Echocardiographic acquisitions allowed to quantify right ventricular parameters of morphology and function with good quality. We identified right ventricular ischemic lesions in the model. The model can be used to compare different treatments or to validate non-invasive parameters of right ventricular morphology and function in the context of ARHF.

We present a protocol to induce and phenotype an acute right heart failure in a large animal model with chronic pulmonary hypertension. This model can be used to test therapeutic interventions, to develop right heart metrics&#160;or to improve the understanding of acute right heart failure pathophysiology.

Induction et phénotypage de l’insuffisance cardiaque droite aiguë dans un grand modèle animal d’hypertension pulmonaire thromboembolique chronique

The development of acute right heart failure (ARHF) in the context of chronic pulmonary hypertension (PH) is associated with poor short-term outcomes. The ...

Preclinical Model of Hind Limb Ischemia in Diabetic Rabbits

Peripheral vascular disease is a widespread clinical problem that affects millions of patients worldwide. A major consequence of peripheral vascular disease is the development of ischemia. In severe cases, patients can develop critical limb ischemia in which they experience constant pain and an increased risk of limb amputation. Current therapies for peripheral ischemia include bypass surgery or percutaneous interventions such as angioplasty with stenting or atherectomy to restore blood flow. However, these treatments often fail to the continued progression of vascular disease or restenosis or are contraindicated due to the overall poor health of the patient. A promising potential approach to treat peripheral ischemia involves the induction of therapeutic neovascularization to allow the patient to develop collateral vasculature. This newly formed network alleviates peripheral ischemia by restoring perfusion to the affected area. The most frequently employed preclinical model for peripheral ischemia utilizes the creation of hind limb ischemia in healthy rabbits through femoral artery ligation. In the past, however, there has been a strong disconnect between the success of preclinical studies and the failure of clinical trials regarding treatments for peripheral ischemia. Healthy animals typically have robust vascular regeneration in response to surgically induced ischemia, in contrast to the reduced vascularity and regeneration in patients with chronic peripheral ischemia. Here, we describe an optimized animal model for peripheral ischemia in rabbits that includes hyperlipidemia and diabetes. This model has reduced collateral formation and blood pressure recovery in comparison to a model with a higher cholesterol diet. Thus, the model may provide better correlation with human patients with compromised angiogenesis from the common co-morbidities that accompany peripheral vascular disease.

We describe a surgical procedure used to induce peripheral ischemia in rabbits with hyperlipidemia and diabetes. This surgery acts as a preclinical model for conditions experienced in peripheral artery disease in patients. Angiography is also described as a means to measure the extent of introduced ischemia and recovery of perfusion.

Modèle préclinique d’ischémie des membres postérieurs chez les lapins diabétiques

Peripheral vascular disease is a widespread clinical problem that affects millions of patients worldwide. A major consequence of peripheral vascular ...

Establishing a Swine Model of Post-myocardial Infarction Heart Failure for Stem Cell Treatment

Although advances have been achieved in the treatment of heart failure (HF) following myocardial infarction (MI), HF following MI remains one of the major causes of mortality and morbidity around the world. Cell-based therapies for cardiac repair and improvement of left ventricular function after MI have attracted considerable attention. Accordingly, the safety and efficacy of these cell transplantations should be tested in a preclinical large animal model of HF prior to clinical use. Pigs are widely used for cardiovascular disease research due to their similarity to humans in terms of heart size and coronary anatomy. Therefore, we sought to present an effective protocol for the establishment of a porcine chronic HF model using closed-chest coronary balloon occlusion of the left circumflex artery (LCX), followed by rapid ventricular pacing induced with pacemaker implantation. Eight weeks later, the stem cells were administered by intramyocardial injection in the peri-infarct area. Then the infarct size, cell survival, and left ventricular function (including echocardiography, hemodynamic parameters, and electrophysiology) were evaluated. This study helps establish a stable preclinical large animal HF model for stem cell treatment.

We sought to establish a swine model of heart failure induced by left circumflex artery blockage and rapid pacing to test the effect and safety of intramyocardial administration of stem cells for cell-based therapies.

Établissement d’un modèle porcin d’insuffisance cardiaque d’infarctus post-myocardique pour le traitement des cellules souches

Although advances have been achieved in the treatment of heart failure (HF) following myocardial infarction (MI), HF following MI remains one of the major ...

A Preclinical Model of Exertional Heat Stroke in Mice

Heat stroke is the most severe manifestation of heat-related illnesses. Classic heat stroke (CHS), also known as passive heat stroke, occurs at rest, whereas exertional heat stroke (EHS) occurs during physical activity. EHS differs from CHS in etiology, clinical presentation, and sequelae of multi-organ dysfunction. Until recently, only models of CHS have been well established. This protocol aims to provide guidelines for a refined preclinical mouse model of EHS that is free from major limiting factors such as the use of anesthesia, restraint, rectal probes, or electric shock. Male and female C57Bl/6 mice, instrumented with core temperature (Tc) telemetric probes were utilized in this model. For familiarization with the running mode, mice undergo 3 weeks of training using both voluntary and forced running wheels. Thereafter, mice run on a forced wheel inside a climatic chamber set at 37.5 &#176;C and 40%-50% relative humidity (RH) until displaying symptom limitation (e.g., loss of consciousness) at Tc of 42.1-42.5 &#176;C, although suitable results can be obtained at chamber temperatures between 34.5-39.5 &#176;C and humidity between 30%-90%. Depending on the desired severity, mice are removed from the chamber immediately for recovery in ambient temperature or remain in the heated chamber for a longer duration, inducing a more severe exposure and a higher incidence of mortality. Results are compared with sham-matched exercise controls (EXC) and/or na&#239;ve controls (NC). The model mirrors many of the pathophysiological outcomes observed in human EHS, including loss of consciousness, severe hyperthermia, multi-organ damage as well as inflammatory cytokine release, and acute phase responses of the immune system. This model is ideal for hypothesis-driven research to test preventative and therapeutic strategies that may delay the onset of EHS or reduce the multi-organ damage that characterizes this manifestation.

The protocol describes the development of a standardized, repeatable, preclinical model of exertional heat stroke (EHS) in mice free from adverse external stimuli such as electric shock. The model provides a platform for mechanistic, preventative, and therapeutic studies.

Un modèle préclinique de coup de chaleur à l’effort chez la souris

Heat stroke is the most severe manifestation of heat-related illnesses. Classic heat stroke (CHS), also known as passive heat stroke, occurs at rest, ...

Large Animal Model for Evaluating the Efficacy of the Gene Therapy in Ischemic Heart

Coronary artery disease is one of the significant causes of mortality and morbidity worldwide. Despite the progression of current therapeutics, a considerable proportion of coronary artery disease patients remain symptomatic. Gene therapy-mediated therapeutic angiogenesis offers a novel therapeutic method for improving myocardial perfusion and relieving symptoms. Gene therapy with different angiogenic factors has been studied in few clinical trials. Due to the novelty of the method, the progress of myocardial gene therapy is a continuous path from bench to bedside. Therefore, large animal models are needed for evaluating the safety and efficacy. The more the large animal model identifies the original disease and the endpoints used in clinics, the more predictable outcomes are from clinical trials. Here, we introduce a large animal model for evaluating the efficacy of the gene therapy in the ischemic porcine heart. We use clinically relevant imaging methods such as ultrasound imaging and 15H2O-PET. For targeting the gene transfers into the desired area, electroanatomical mapping is used. The aim of this method is: (1) to mimic chronic coronary artery disease, (2) to induce therapeutic angiogenesis at hypoxic areas of the heart, and (3) to evaluate the safety and efficacy of the gene therapy by using relevant endpoints.

Myocardial gene therapy for ischemic heart disease holds great promise for future therapeutics. Here, we introduce a large animal model for evaluating the efficacy of gene therapy in the ischemic heart.