Name: chronic ovine model of right ventricular failure and functional tricuspid regurgitation
Uploaded: 2023-03-17T13:40:00
Description: Watch this Scientific Journal Video about Chronic Ovine Model of Right Ventricular Failure and Functional Tricuspid Regurgitation at JoVE.com

The study was funded by an internal grant from the Meijer Heart and Vascular Institute at Spectrum Health.

The protocol was approved by the Michigan State University Institutional Animal Care and Use Committee (IACUC) (Protocol 2020-035, approved on 7/27/2020). For this study, 20 adult male sheep weighing 62 &#177; 7 kg were used.
1. Preoperative steps
<ol>
	<li>Fast the animal 12 h prior to surgery (overnight).</li>
	<li>Place the animal in a sheep chair (Figure 1), and prepare for the right jugular vein cannulation using a long 11 Fr introducer she...

<ol>
	<li>Haddad, F., Hunt, S. A., Rosenthal, D. N., Murphy, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Right+ventricular+function+in+cardiovascular+disease,+part+I:+Anatomy,+physiology,+aging,+and+functional+Assessment+of+the+right+ventricle.">Right ventricular function in cardiovascular disease, part I: Anatomy, physiology, aging, and functional Assessment of the right ventricle.</a> Circulation. 117 (11), 1436-1448 (2008).</li><li>Taramasso, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+growing+clinical+importance+of+secondary+tricuspid+regurgitation.">The growing clinical importance of secondary tricuspid regurgitation.</a> Journal of the American College of Cardiology. 59 (8), 703-710 (2012).</li><li>Mangieri, 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=Mechanism+and+implications+of+the+tricuspid+regurgitation:+From+the+pathophysiology+to+the+current+and+future+therapeutic+options.">Mechanism and implications of the tricuspid regurgitation: From the pathophysiology to the current and future therapeutic options.</a> Circulation: Cardiovascular Interventions. 10 (7), 005043 (2017).</li><li>Otto, C. 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=2020+ACC/AHA+Guideline+for+the+Management+of+Patients+With+Valvular+Heart+Disease:+Executive+summary:+A+report+of+the+American+College+of+Cardiology/American+Heart+Association+Joint+Committee+on+Clinical+Practice+Guidelines.">2020 ACC/AHA Guideline for the Management of Patients With Valvular Heart Disease: Executive summary: A report of the American College of Cardiology/American Heart Association Joint Committee on Clinical Practice Guidelines.</a> Circulation. 143 (5), 35-71 (2021).</li><li>Vonk-Noordegraaf, 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=Right+heart+adaptation+to+pulmonary+arterial+hypertension:+physiology+and+pathobiology.">Right heart adaptation to pulmonary arterial hypertension: physiology and pathobiology.</a> Journal of the American College of Cardiology. 62, 22-33 (2013).</li><li>Yoganathan, 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=Tricuspid+valve+diseases:+Interventions+on+the+forgotten+heart+valve.">Tricuspid valve diseases: Interventions on the forgotten heart valve.</a> Journal of Cardiac Surgery. 36 (1), 219-228 (2021).</li><li>Vachi&#233;ry, J. 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=Pulmonary+hypertension+due+to+left+heart+diseases.">Pulmonary hypertension due to left heart diseases.</a> Journal of the American College of Cardiology. 62, 100-108 (2013).</li><li>Chin, K. M., Coghlan, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterizing+the+right+ventricle:+Advancing+our+knowledge.">Characterizing the right ventricle: Advancing our knowledge.</a> American Journal of Cardiology. 110, 3-8 (2012).</li><li>Malinowski, 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=Large+animal+model+of+acute+right+ventricular+failure+with+functional+tricuspid+regurgitation.">Large animal model of acute right ventricular failure with functional tricuspid regurgitation.</a> International Journal of Cardiology. 264, 124-129 (2018).</li><li>Borgdorff, M. A., Dickinson, M. G., Berger, R. M., Bartelds, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Right+ventricular+failure+due+to+chronic+pressure+load:+What+have+we+learned+in+animal+models+since+the+NIH+working+group+statement.">Right ventricular failure due to chronic pressure load: What have we learned in animal models since the NIH working group statement.</a> Heart Failure Review. 20 (4), 475-491 (2015).</li><li>Andersen, 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=Animal+models+of+right+heart+failure.">Animal models of right heart failure.</a> Cardiovascular Diagnosis and Therapy. 10 (5), 1561-1579 (2020).</li><li>Dixon, J. A., Spinale, F. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Large+animal+models+of+heart+failure:+A+critical+link+in+the+translation+of+basic+science+to+clinical+practice.">Large animal models of heart failure: A critical link in the translation of basic science to clinical practice.</a> Circulation: Heart Failure. 2 (3), 262-271 (2009).</li><li>Miyagi, 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=Large+animal+models+of+heart+failure+with+preserved+ejection+fraction.">Large animal models of heart failure with preserved ejection fraction.</a> Heart Failure Review. 27 (2), 595-608 (2022).</li><li>Sato, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Large+animal+model+of+chronic+pulmonary+hypertension.">Large animal model of chronic pulmonary hypertension.</a> American Society for Artificial Internal Organs Journal. 54 (4), 396-400 (2008).</li><li>Bogaard, H. 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=Chronic+pulmonary+artery+pressure+elevation+is+insufficient+to+explain+right+heart+failure.">Chronic pulmonary artery pressure elevation is insufficient to explain right heart failure.</a> Circulation. 120 (20), 1951-1960 (2009).</li><li>Xie, X. 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=Tricuspid+leaflet+resection+in+an+open+beating+heart+for+the+creation+of+a+canine+tricuspid+regurgitation+model.">Tricuspid leaflet resection in an open beating heart for the creation of a canine tricuspid regurgitation model.</a> Interactive Cardiovascular and Thoracic Surgery. 22 (2), 149-154 (2016).</li><li>Hoppe, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Percutaneous+technique+for+creation+of+tricuspid+regurgitation+in+an+ovine+model.">Percutaneous technique for creation of tricuspid regurgitation in an ovine model.</a> Journal of Vascular and Interventional Radiology. 18, 133-136 (2007).</li><li>Malinowski, 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=Large+animal+model+of+functional+tricuspid+regurgitation+in+pacing+induced+end-stage+heart+failure.">Large animal model of functional tricuspid regurgitation in pacing induced end-stage heart failure.</a> Interactive Cardiovascular and Thoracic Surgery. 24 (6), 905-910 (2017).</li><li>Ukita, R., 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+large+animal+model+for+pulmonary+hypertension+and+right+ventricular+failure:+Left+pulmonary+artery+ligation+and+progressive+main+pulmonary+artery+banding+in+sheep.">A large animal model for pulmonary hypertension and right ventricular failure: Left pulmonary artery ligation and progressive main pulmonary artery banding in sheep.</a> Journal of Visualized Experiments. (173), e62694 (2021).</li><li>Dufva, 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=Pulmonary+arterial+banding+in+mice+may+be+a+suitable+model+for+studies+on+ventricular+mechanics+in+pediatric+pulmonary+arterial+hypertension.">Pulmonary arterial banding in mice may be a suitable model for studies on ventricular mechanics in pediatric pulmonary arterial hypertension.</a> Journal of Cardiovascular Magnetic Resonance. 23 (1), 66 (2021).</li><li>Verbelen, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanical+support+of+the+pressure+overloaded+right+ventricle:+An+acute+feasibility+study+comparing+low+and+high+flow+support.">Mechanical support of the pressure overloaded right ventricle: An acute feasibility study comparing low and high flow support.</a> American Journal of Physiology-Heart and Circulatory Physiology. 309 (4), 615-624 (2015).</li></ol>

In this model, 8 weeks of pulmonary artery banding resulted in a stable chronic ovine model of right ventricular dysfunction and, in most cases, significant FTR. The strengths of the presented chronic PAB model include the precise afterload adjustment during the procedure, although its influence on RV responses may differ. The model is suitable for evaluating varying degrees of RV failure or FTR, with the severity modulated by the degree of pulmonary artery constriction. Moreover, the application of fixed and stable resi...

Right ventricular failure (RVF) is recognized as an important factor contributing to the morbidity and mortality of cardiac patients. The most common causes of RVF are left-sided heart disease and pulmonary hypertension1. During the progression of RVF, functional tricuspid regurgitation (FTR) may arise as a consequence of right ventricular (RV) dysfunction, annular dilatation, and subvalvular remodeling. Moderate to severe FTR is an independent predictor of mortality2,3, and it is estimated that 80%-90% of tricuspid regurgitation cases are functional in nature4. FTR itself may promote adverse ventricular remodeling by influencing either afterload or preload5. The tricuspid valve has historically been considered the forgotten valve6, and it was believed that the treatment of left-sided heart disease would resolve the associated RV pathology and FTR7. Recent data have shown this to be a faulty strategy, and current clinical guidelines advocate a much more aggressive approach to FTR4. However, the pathophysiology of severe FTR associated with right ventricular dysfunction is still poorly understood, leading to suboptimal clinical results8. The currently available large animal models of RVF are based on pressure, volume, or mixed overload. We have previously described a large animal model of RVF and TR but only in an acute setting9.
The current study focuses on a chronic ovine model of pulmonary artery banding (PAB) to increase RV afterload (pressure overload) and induce RV dysfunction and FTR. The afterload model is reliable and reproducible compared to pulmonary hypertension models, in which changes in microvasculature are less predictable and more likely10. The goal of the study was to develop a chronic large animal model of RVF and FTR that would most accurately mimic RV pressure overload in patients with left-sided heart disease and pulmonary hypertension. The establishment of such a model would permit in-depth studies on the pathophysiology of the ventricular and valvular remodeling associated with RV dysfunction and tricuspid insufficiency. The ovine model was chosen based on our prior work on the mitral valve and the published literature supporting the anatomical and physiological similarities between human and ovine hearts11,12,13.
For this study, 20 adult sheep (62 &#177; 7 kg) underwent a left thoracotomy and main pulmonary artery banding (PAB) to at least double the systolic pulmonary artery pressure (SPAP), thus inducing RV pressure overload. The animals were followed for 8 weeks, and the symptoms of heart failure were treated with diuretics when clinically apparent. Surveillance echocardiography was performed periodically to assess RV function and valvular competence. Following the completion of the experimental protocol for model development (8 weeks), the animals were taken back to the operating room for median sternotomy and the implantation of sonomicrometry crystals on the epicardial and intra-cardiac structures. This procedure was performed using cardiopulmonary bypass with the heart beating and with bicaval control. There were no problems in weaning the animals from the cardiopulmonary bypass or acquiring the sonomicrometry data in a stable steady-state hemodynamic environment without the need for inotropes for right heart support. We anticipate performing tricuspid ring annuloplasty and other right heart procedures in the near future using a right thoracotomy approach in both terminal and survival experiments. The current experience leads us to believe that it will be possible to wean the animals from the cardiopulmonary bypass without difficulty and that long-term survival is feasible. As such, we believe the model will permit the performance of clinically pertinent cardiac procedures. Below is a description of the steps (perioperative and operative) performed for carrying out the ovine experimental protocol.

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	<tbody>
		<tr>
			<td>Anesthesia Machine Drager Narkomed MRI-2</td>
			<td>Drager</td>
			<td>4116091-001</td>
			<td></td>
		</tr>
		<tr>
			<td>angiocatheter</td>
			<td>BD</td>
			<td>BD382268</td>
			<td>14GAx8.25cm</td>
		</tr>
		<tr>
			<td>BD ChloraPrep Scrub Teal 26 ml applicator with a sterile solution</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Blade #11</td>
			<td>Bard-Parker</td>
			<td>371111</td>
			<td></td>
		</tr>
		<tr>
			<td>Buprenorphine&#160;</td>
			<td>HIKMA</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>cefazolin 1.0g</td>
			<td>Hikma</td>
			<td>0143-9924-90</td>
			<td></td>
		</tr>
		<tr>
			<td>Diprivan 200mg/20ml</td>
			<td>63323-0269-29</td>
			<td>FRESENIUS KABI</td>
			<td></td>
		</tr>
		<tr>
			<td>Electrosurgical generator Valleylab Force FX</td>
			<td>Valleylab</td>
			<td>CF5L44233A</td>
			<td></td>
		</tr>
		<tr>
			<td>Gentamicin Sulfate 40 mg / mL</td>
			<td>Fresenius</td>
			<td>406365</td>
			<td></td>
		</tr>
		<tr>
			<td>i-Stat Blood analyzer MN 300</td>
			<td>Abbott</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Lidocaine HCl 1%</td>
			<td>Pfizer</td>
			<td>243243</td>
			<td></td>
		</tr>
		<tr>
			<td>Open ligating clip appliers Horizon Medium</td>
			<td>Teleflex</td>
			<td>237061</td>
			<td></td>
		</tr>
		<tr>
			<td>PERMAHAND Silk Suture</td>
			<td>PERMA HAND</td>
			<td>SA 63H</td>
			<td></td>
		</tr>
		<tr>
			<td>Pinnacle Introducer sheath</td>
			<td>Terrumo</td>
			<td>RSS102</td>
			<td>sheath length 10cm</td>
		</tr>
		<tr>
			<td>Prolene 3-0</td>
			<td>ETHICON</td>
			<td>8684H</td>
			<td></td>
		</tr>
		<tr>
			<td>Titanium Clips Medium</td>
			<td>Teleflex</td>
			<td>2200</td>
			<td></td>
		</tr>
		<tr>
			<td>Umbilical tape</td>
			<td>Ethicon</td>
			<td>EFA 1165</td>
			<td></td>
		</tr>
		<tr>
			<td>VICRYL 2 coated undyed 1X54&#34; TP-1</td>
			<td>ETHICON</td>
			<td>J 880T</td>
			<td></td>
		</tr>
		<tr>
			<td>Vicryl 2-0</td>
			<td>ETHICON</td>
			<td>J269H</td>
			<td></td>
		</tr>
	</tbody>
</table>

chronic ovine model of right ventricular failure and functional tricuspid regurgitation

The pathophysiology of severe functional tricuspid regurgitation (FTR) associated with right ventricular dysfunction is poorly understood, leading to suboptimal clinical results. We set out to establish a chronic ovine model of FTR and right heart failure to investigate the mechanisms of FTR. Twenty adult male sheep (6-12 months old, 62 &#177; 7 kg) underwent a left thoracotomy and baseline echocardiography. A pulmonary artery band (PAB) was placed and cinched around the main pulmonary artery (PA) to at least double the systolic pulmonary artery pressure (SPAP), inducing right ventricular (RV) pressure overload and signs of RV dilatation. PAB acutely increased the SPAP from 21 &#177; 2 mmHg to 62 &#177; 2 mmHg. The animals were followed for 8 weeks, symptoms of heart failure were treated with diuretics, and surveillance echocardiography was used to assess for pleural and abdominal fluid collection. Three animals died during the follow-up period due to stroke, hemorrhage, and acute heart failure. After 2 months, a median sternotomy and epicardial echocardiography were performed. Of the surviving 17 animals, 3 developed mild tricuspid regurgitation, 3 developed moderate tricuspid regurgitation, and 11 developed severe tricuspid regurgitation. Eight weeks of pulmonary artery banding resulted in a stable chronic ovine model of right ventricular dysfunction and significant FTR. This large animal platform can be used to further investigate the structural and molecular basis of RV failure and functional tricuspid regurgitation.

Following the completion of the experimental protocol for model development (nearly 8 weeks), the animals were taken back to the operating room for median sternotomy and the implantation of sonomicrometry crystals on the epicardial and intra-cardiac structures. This procedure was performed using cardiopulmonary bypass with the heart beating and with bicaval control, as&#160;described by our group in detail previously9. There were no problems in weaning the animals from the cardiopulmonary bypass o...

Watch this Scientific Journal Video about Chronic Ovine Model of Right Ventricular Failure and Functional Tricuspid Regurgitation at JoVE.com

Chronic Ovine Model of Right Ventricular Failure and Functional Tricuspid Regurgitation

Right ventricular failure and functional tricuspid regurgitation are associated with left-sided heart disease and pulmonary hypertension, which contribute significantly to morbidity and mortality in patients. Establishing a chronic ovine model to study right ventricular failure and functional tricuspid regurgitation will help in understanding their mechanisms, progression, and possible treatments.

The pathophysiology of severe functional tricuspid regurgitation (FTR) associated with right ventricular dysfunction is poorly understood, leading to ...

The model is similar to those previously described in the literature. Although our specific goal was to induce significant functional tricuspid regurgitation rather than just isolated right heart dysfunction. The technique may be used in the future to study mechanisms of functional tricuspid regurgitation with its associated right ventricular annular and subular remodeling, as well as tissue changes.The model may be helpful in identifying molecular pathways responsible for functional tricuspid regurgitation and establishing more efficient and complex surgical techniques for its treatment. Begin by placing the animal in a sheep chair. Shave the right anterior aspect of the neck around 10 to 15 centimeters lateral from the midline for the right jugular vein using clippers.After disinfecting the surgical area, turn the animal's head to the left to expose the right anterior and lateral aspects of the neck. Then localize the jugular vein course by compressing the bottom of the neck to distend the vein. Now, anesthitize the insertion site with 1%lidocaine.Using a number 11 blade, cut the skin above and perpendicular to the vein and cannulate with a 14 gauge angio catheter. Once the angio catheter is in place, remove the needle, pass the guide wire, followed by removing the catheter. Then, place the 11 french sheath and remove the guide wire before securing the sheath.Ensure the patency and proper placement of the cannula by withdrawing dark red blood and performing a saline flush to confirm the flow and absence of swelling at the insertion site. After inducing propofol intravenously, intubate with an endotracheal tube using a laryngoscope with a number five blade as described in the manuscript. Confirm proper placement by the bilateral breath sounds and condensation on the endotracheal tube.After preparing the operative field, shave the left anterior thorax. Then, apply sterile drapes around the disinfected surgical area. Make a 10 centimeter long skin and subcutaneous incision at the level of the fourth intercostal space.Confirm the correct intercostal space is incised by identifying the thoracic inlet and counting the intercostal spaces downward. Subsequently, continue the incision in the center and along the fourth intercostal space. Now, divide the intercostal muscles, open the chest cavity and spread the ribs with a mini thoracotomy finochietto style retractor.Avoid injuring the left internal mammary artery at the sternal border of the incision and the lung at the superior border. Then, perform baseline epicardial echocardiography to assess the biventricular function and valvular competence. Identify the left internal mammary artery at the sternal border of the incision, remove the adjacent tissues around it, and prepare to establish an arterial line for pressure monitoring.Now, place two 4-0 silk sutures around the artery with one proximal and one distal to the cannulation site. Use titanium clips with a clip applier to clip the left internal mammary artery distal to the planned cannulation site to prevent backflow bleeding during cannulation. Make a perpendicular incision that is half of the circumference of the catheter in the left internal mammary artery with a number 11 blade.Insert an 18 gauge angio catheter and attach it to the arterial line module. When proper placement of the angio catheter is confirmed by arterial pressure line reading, secure the catheter using two 4-0 silk sutures placed earlier. Next, perform pericardiotomy starting at the level of pulmonary artery sinuses and going four to five centimeters laterally along the main pulmonary artery taking care not to injure the left phrenic nerve.Apply four to five retraction stitches to the opened pericardium to create a pericardial well which facilitates exposure and dissection between the pulmonary trunk and aorta. Using blunt right angle forceps, dissect the main pulmonary artery from ascending aorta around two to three centimeters from its origin as described in the manuscript. To separate the main pulmonary artery from the ascending aorta, use electrocautery or scissors to remove the connective tissue between the two structures.Pass an umbilical tape around the main pulmonary artery with a blunt right angle clamp. Then establish a main pulmonary artery pressure line by placing a 5-0 monofilament purse string suture one centimeter distal from the main pulmonary artery sinuses. Insert a 20 gauge angio catheter and connect it to a monitoring line.Before cinching the umbilical tape, ensure correct main pulmonary artery and arterial line readings are achieved. After holding both ends of the umbilical tape, clip them together to reduce the lumen of the main pulmonary artery. Then, tighten the band with the successive application of a clip applier as described in the manuscript.Perform post banding echocardiography to assess the biventricular function and valvular competence as demonstrated earlier. Remove the main pulmonary artery pressure line. When maximal cinching and stable hemodynamic conditions are achieved, secure the umbilical tape to the adventitia of the main pulmonary artery using a 5-0 monofilament suture to avoid distal migration.Then, remove the arterial line and ensure good hemostasis by checking for any bleeding from the area where the band and arterial lines were placed. Place a chest tube in the left thorax with the entry site one intercostal space below the initial incision. Close the ribs with two vicryl 2-0 sutures and close the wound with three layer continuous sutures.Vicryl 2-0 for the muscle and subcutaneous tissues and proline 3-0 for the skin. Once good hemostasis is confirmed, remove the chest tube and extubate the animal. After the follow-up period, the mean TR grade increased from 0.4 to 3.2, demonstrating signs of evolving right ventricular failure and the development of significant TR after eight weeks of pulmonary banding.Consistent with the echocardiographic examination. Over or under tightening of the band may result in the early animal demise or not inducing redicular failure and functional tricuspid regurgitation. Additional methods inducing heart failure may be performed after initial made pulmonary artery banding, like ventricle art pacing or ascending aorta banding, or band may be released to elevate right heart failure.

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Research

JoVE Journal

Medicine

Assessment of Right Ventricular Structure and Function in Mouse Model of Pulmonary Artery Constriction by Transthoracic Echocardiography

Emerging clinical data support the notion that RV dysfunction is critical to the pathogenesis of cardiovascular disease and heart failure1-3. Moreover, the RV is significantly affected in pulmonary diseases such as pulmonary artery hypertension (PAH). In addition, the RV is remarkably sensitive to cardiac pathologies, including left ventricular (LV) dysfunction, valvular disease or RV infarction4. To understand the role of RV in the pathogenesis of cardiac diseases, a reliable and noninvasive method to access the RV structurally and functionally is essential.
A noninvasive trans-thoracic echocardiography (TTE) based methodology was established and validated for monitoring dynamic changes in RV structure and function in adult mice. To impose RV stress, we employed a surgical model of pulmonary artery constriction (PAC) and measured the RV response over a 7-day period using a high-frequency ultrasound microimaging system. Sham operated mice were used as controls. Images were acquired in lightly anesthetized mice at baseline (before surgery), day 0 (immediately post-surgery), day 3, and day 7 (post-surgery). Data was analyzed offline using software.
Several acoustic windows (B, M, and Color Doppler modes), which can be consistently obtained in mice, allowed for reliable and reproducible measurement of RV structure (including RV wall thickness, end-diastolic&#160;and end-systolic dimensions), and function (fractional area change, fractional shortening, PA peak velocity, and peak pressure gradient) in normal mice and following PAC.
Using this method, the pressure-gradient resulting from PAC was accurately measured in real-time using Color Doppler mode and was comparable to direct pressure measurements performed with a Millar high-fidelity microtip catheter. Taken together, these data demonstrate that RV measurements obtained from various complimentary views using echocardiography are reliable, reproducible and can provide insights regarding RV structure and function. This method will enable a better understanding of the role of RV cardiac dysfunction.

Right ventricle (RV) dysfunction is critical to the pathogenesis of cardiovascular disease, yet limited methodologies are available for its evaluation. Recent advances in ultrasound imaging provide a noninvasive and accurate option for longitudinal RV study. Herein, we detail a step-by-step echocardiographic method using a murine model of RV pressure overload.

Emerging clinical data support the notion that RV dysfunction is critical to the pathogenesis of cardiovascular disease and heart failure1-3. Moreover, ...

Isolation and Functional Characterization of Human Ventricular Cardiomyocytes from Fresh Surgical Samples

Cardiomyocytes from diseased hearts are subjected to complex remodeling processes involving changes in cell structure, excitation contraction coupling and membrane ion currents. Those changes are likely&#160;to be responsible for the increased arrhythmogenic risk and the contractile alterations leading to systolic and diastolic dysfunction in cardiac patients. However, most information on the alterations of myocyte function in cardiac diseases has come from animal models.
Here we describe and validate a protocol to isolate viable myocytes from small surgical samples of ventricular myocardium from patients undergoing cardiac surgery operations. The protocol is described in detail. Electrophysiological and intracellular calcium measurements are reported to demonstrate the feasibility of a number of single cell measurements in human ventricular cardiomyocytes obtained with this method.
The protocol reported here can be useful for future investigations of the cellular and molecular basis of functional alterations of the human heart in the presence of different cardiac diseases. Further, this method can be used to identify novel therapeutic targets at cellular level and to test the effectiveness of new compounds on human cardiomyocytes, with direct translational value.

Current knowledge on the cellular basis of cardiac diseases mostly relies on studies on animal models. Here we describe and validate a novel method to obtain single viable cardiomyocytes from small surgical samples of human ventricular myocardium. Human ventricular myocytes can be used for electrophysiological studies and drug testing.

Cardiomyocytes from diseased hearts are subjected to complex remodeling processes involving changes in cell structure, excitation contraction coupling and ...

Urinary Bladder Distention Evoked Visceromotor Responses as a Model for Bladder Pain in Mice

Approximately 3-8 million people in the United States suffer from interstitial cystitis/bladder pain syndrome (IC/BPS), a debilitating condition characterized by increased urgency and frequency of urination, as well as nocturia and general pelvic pain, especially upon bladder filling or voiding. Despite years of research, the cause of IC/BPS remains elusive and treatment strategies are unable to provide complete relief to patients. In order to study nervous system contributions to the condition, many animal models have been developed to mimic the pain and symptoms associated with IC/BPS. One such murine model is urinary bladder distension (UBD). In this model, compressed air of a specific pressure is delivered to the bladder of a lightly anesthetized animal over a set period of time. Throughout the procedure, wires in the superior oblique abdominal muscles record electrical activity from the muscle. This activity is known as the visceromotor response (VMR) and is a reliable and reproducible measure of nociception. Here, we describe the steps necessary to perform this technique in mice including surgical manipulations, physiological recording, and data analysis. With the use of this model, the coordination between primary sensory neurons, spinal cord secondary afferents, and higher central nervous system areas involved in bladder pain can be unraveled. This basic science knowledge can then be clinically translated to treat patients suffering from IC/BPS.

Approximately 3-8 million people in the United States suffer from interstitial cystitis/bladder pain syndrome (IC/BPS), a debilitating condition characterized in part by pelvic pain. In order to study nervous system contributions to the condition, a physiological model of urinary bladder pain is used in mice and rats.

Approximately 3-8 million people in the United States suffer from interstitial cystitis/bladder pain syndrome (IC/BPS), a debilitating condition ...

Chronic Thromboembolic Pulmonary Hypertension and Assessment of Right Ventricular Function in the Piglet

An original piglet model of Chronic Thromboembolic Pulmonary Hypertension (CTEPH) associated with chronic Right Ventricular (RV) dysfunction is described. Pulmonary Hypertension (PH) was induced in 3-week-old piglets by a progressive obstruction of the pulmonary vascular bed. A ligation of the left Pulmonary Artery (PA) was performed first through a mini-thoracotomy. Second, weekly embolizations of the right lower pulmonary lobe were done under fluoroscopic guidance with n-butyl-2-cyanoacrylate during 5 weeks. Mean Pulmonary Arterial Pressure (mPAP) measured by ritght heart catheterism, increased progressively, as well as Right Atrial pressure and Pulmonary Vascular Resistances (PVR) after 5 weeks compared to sham animals. Right Ventricular (RV) structural and functional remodeling were assessed by transthoracic echocardiography (RV diameters, RV wall thickness, RV systolic function). RV elastance and RV-pulmonary coupling were assessed by Pressure-Volume Loops (PVL) analysis with conductance method. Histologic study of the lung and the right ventricle were also performed. Molecular analyses on RV fresh tissues could be performed through repeated transcutaneous endomyocardial biopsies. Pulmonary microvascular disease in obstructed and unobstructed territories was studied from lung biopsies using molecular analyses and pathology. Furthermore, the reliability and the reproducibility was associated with a range of PH severity in animals. Most aspects of the human CTEPH disease were reproduced in this model, which allows new perspectives for the understanding of the underlying mechanisms (mitochondria, inflammation) and new therapeutic approaches (targeted, cellular or gene therapies) of the overloaded right ventricle but also pulmonary microvascular disease.

Chronic Thromboembolic Pulmonary Hypertension (CTEPH) and Right Ventricular (RV) dysfunction were induced in piglets by progressive obstruction of the pulmonary arteries. Consequences were remarkably similar to those observed in CTEPH patients. This animal model would be a very useful tool for pathophysiology and therapeutic experiments on CTEPH and RV failure.

An original piglet model of Chronic Thromboembolic Pulmonary Hypertension (CTEPH) associated with chronic Right Ventricular (RV) dysfunction is described. ...

Transvaginal Mesh Insertion in the Ovine Model

This protocol describes mesh insertion into the rectovaginal septum in sheep using a single vaginal incision technique, with and without the trocar-guided insertion of anchoring arms. Parous sheep underwent the dissection of the rectovaginal septum, followed by the insertion of an implant with or without four anchoring arms, both designed to fit the ovine anatomy. The anchoring arms were put in place using a trocar and an "outside-in" technique. The cranial arms were passed through the obturator, gracilis, and adductor magnus muscles. The caudal arms were fixed near the sacrotuberous ligament, through the coccygeus muscles. This technique allows for the mimicking of surgical procedures performed in women suffering from pelvic organ prolapse. The anatomical spaces and elements are easily identified. The most critical part of the procedure is the insertion of the cranial trocar, which can easily penetrate the peritoneal cavity or the surrounding pelvic organs. This can be avoided by a more extensive retroperitoneal dissection and by guiding the trocar more laterally. This approach is designed only for experimental testing of novel implants in large animal models, as trocar-guided insertion is currently not used clinically.

This protocol describes mesh implantation in the ovine rectovaginal septum using a single vaginal incision technique, with and without the trocar-guided insertion of anchoring arms.

This protocol describes mesh insertion into the rectovaginal septum in sheep using a single vaginal incision technique, with and without the trocar-guided ...

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.

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

Echocardiographic Measurement of Right Ventricular Diastolic Parameters in Mouse

Diastolic dysfunction is a prominent feature of right ventricular (RV) remodeling associated with conditions of pressure overload. However, the RV diastolic function is rarely quantified in experimental studies. This might be due to technical difficulties in the visualization of the RV in the apical four-chamber view in rodents. Here we describe two positions facilitating the visualization of the apical four-chamber view in mice to assess the RV diastolic function.
The apical four-chamber view is enabled by tilting the mouse fixation platform to the left and caudally (LeCa) or to the right and cranially (RiCr). Both positions provide images of comparable quality. The results of the RV diastolic function obtained from two positions are not significantly different. Both positions are comparably easy to perform. This protocol can be incorporated into published protocols and enables detailed investigations of the RV function.

Here we describe and compare two positions for obtaining the apical four-chamber view in mice. These positions enable the quantification of the right ventricular function, provide comparable results, and can be used interchangeably.

Diastolic dysfunction is a prominent feature of right ventricular (RV) remodeling associated with conditions of pressure overload. However, the RV ...

A Pulmonary Trunk Banding Model of Pressure Overload Induced Right Ventricular Hypertrophy and Failure

Right ventricular (RV) failure induced by sustained pressure overload is a major contributor to morbidity and mortality in several cardiopulmonary disorders. Reliable and reproducible animal models of RV failure are therefore warranted in order to investigate disease mechanisms and effects of potential therapeutic strategies. Banding of the pulmonary trunk is a common method to induce isolated RV hypertrophy but in general, previously described models have not succeeded in creating a stable model of RV hypertrophy and failure.
We present a rat model of pressure overload induced RV hypertrophy caused by pulmonary trunk banding (PTB) that enables different phenotypes of RV hypertrophy with and without RV failure. We use a modified ligating clip applier to compress a titanium clip around the pulmonary trunk to a pre-set inner diameter. We use different clip diameters to induce different stages of disease progression from mild RV hypertrophy to decompensated RV failure.
RV hypertrophy develops consistently in rats subjected to the PTB procedure and depending on the diameter of the applied banding clip, we can accurately reproduce different disease severities ranging from compensated hypertrophy to severe decompensated RV failure with extra-cardiac manifestations.
The presented PTB model is a valid and robust model of pressure overload induced RV hypertrophy and failure that has several advantages to other banding models including high reproducibility and the possibility of inducing severe and decompensated RV failure.

We present a surgical method to induce right ventricular hypertrophy and failure in rats.

Right ventricular (RV) failure induced by sustained pressure overload is a major contributor to morbidity and mortality in several cardiopulmonary ...

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.

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

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.

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