Name: the inverted heart model for interstitial transudate collection from the isolated rat heart
Uploaded: 2017-06-20T15:00:00
Description: Watch this Scientific Journal Video about The Inverted Heart Model for Interstitial Transudate Collection from the Isolated Rat Heart at JoVE.com

This study was funded by NSFC 81570244, FoKo 23/2013, and SFB 1116/B01 and by the Cardiovascular Research Institute D&#252;sseldorf (CARID).

All experiments were approved by the local regulatory agency (LANUV of Nordrhein-Westfalen, Germany) and were performed according to the guidelines of animal use. Animals were fed with a standard chow diet and received tap water ad libitum. All equipment and chemicals necessary to each step of the experiment are available in the Table of Materials.
1. Preparation of the Latex Cap and Intraventricular Balloon
<ol>
	<li>Make an aluminum mold using a milling m...

<ol>
	<li>Henkel, D. M., Redfield, M. M., Weston, S. A., Gerber, Y., Roger, V. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Death+in+heart+failure:+a+community+perspective.">Death in heart failure: a community perspective.</a> Circ Heart Fail. 1 (2), 91-97 (2008).</li><li>Limana, F., 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=Myocardial+infarction+induces+embryonic+reprogramming+of+epicardial+c-kit(+)+cells:+role+of+the+pericardial+fluid.">Myocardial infarction induces embryonic reprogramming of epicardial c-kit(+) cells: role of the pericardial fluid.</a> J Mol Cell Cardiol. 48 (4), 609-618 (2010).</li><li>Brunner, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cardiac+tissue+endothelin-1+levels+under+basal,+stimulated,+and+ischemic+conditions.">Cardiac tissue endothelin-1 levels under basal, stimulated, and ischemic conditions.</a> J Cardiovasc Pharmacol. 26, S44-S46 (1995).</li><li>de Lannoy, L. 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Local production of angiotensin I.</a> Hypertension. 29 (6), 1240-1251 (1997).</li><li>Strupp, M., Kammermeier, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interstitial+Lactate+And+Glucose-Concentrations+Of+the+Isolated-Perfused+Rat-Heart+before,+during+And+after+Anoxia.">Interstitial Lactate And Glucose-Concentrations Of the Isolated-Perfused Rat-Heart before, during And after Anoxia.</a> Pflugers Arch. 423 (3-4), 232-237 (1993).</li><li>Wienen, W., Jungling, E., Kammermeier, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enzyme-Release+into+the+Interstitial+Space+of+the+Isolated+Rat-Heart+Induced+by+Changes+in+Contractile+Performance.">Enzyme-Release into the Interstitial Space of the Isolated Rat-Heart Induced by Changes in Contractile Performance.</a> Cardiovasc Res. 28 (8), 1292-1298 (1994).</li><li>De Deckere, E. A., Ten Hoor, ., 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+modified+Langendorff+technique+for+metabolic+investigations.">A modified Langendorff technique for metabolic investigations.</a> Pflugers Arch. 370 (1), 103-105 (1977).</li><li>Tschubar, F., Rose, H., Kammermeier, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fatty+acid+transfer+across+the+myocardial+capillary+wall.">Fatty acid transfer across the myocardial capillary wall.</a> J Mol Cell Cardiol. 25 (4), 355-366 (1993).</li><li>Wienen, W., Kammermeier, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intra-+and+extracellular+markers+in+interstitial+transudate+of+perfused+rat+hearts.">Intra- and extracellular markers in interstitial transudate of perfused rat hearts.</a> Am J Physiol. 254 (4 Pt 2), H785-H794 (1988).</li><li>Sasse, A., Ding, Z. P., Wallich, M., Godecke, A., Schrader, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Vascular+transfer+of+adenovirus+is+augmented+by+nitric+oxide+in+the+rat+heart.">Vascular transfer of adenovirus is augmented by nitric oxide in the rat heart.</a> Am J Physiol Heart Circ Physiol. 287 (3), H1362-H1368 (2004).</li><li>Gnecchi, M., Zhang, Z., Ni, A., Dzau, V. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Paracrine+mechanisms+in+adult+stem+cell+signaling+and+therapy.">Paracrine mechanisms in adult stem cell signaling and therapy.</a> Circ Res. 103 (11), 1204-1219 (2008).</li><li>Herr, D. J., Aune, S. E., Menick, D. 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(101), e52908 (2015).</li><li>Ding, Z., 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=Epicardium-Derived+Cells+Formed+After+Myocardial+Injury+Display+Phagocytic+Activity+Permitting+In+Vivo+Labeling+and+Tracking.">Epicardium-Derived Cells Formed After Myocardial Injury Display Phagocytic Activity Permitting In Vivo Labeling and Tracking.</a> Stem Cells Transl Med. 5 (5), 639-650 (2016).</li><li>Hartwig, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Secretome+profiling+of+primary+human+skeletal+muscle+cells.">Secretome profiling of primary human skeletal muscle cells.</a> Biochim Biophys Acta. 1844 (5), 1011-1017 (2014).</li><li>Smolenski, R. T., Lachno, D. R., Ledingham, S. J. M., Yacoub, M. 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K., Juengling, E., Kammermeier, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Interstitial+transudate+concentration+of+adenosine+and+inosine+in+rat+and+guinea+pig+hearts.">Interstitial transudate concentration of adenosine and inosine in rat and guinea pig hearts.</a> Am J Physiol. 254 (6 Pt 2), H1125-H1132 (1988).</li><li>Heller, L. J., Mohrman, D. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Estimates+of+interstitial+adenosine+from+surface+exudates+of+isolated+rat+hearts.">Estimates of interstitial adenosine from surface exudates of isolated rat hearts.</a> J Mol Cell Cardiol. 20 (6), 509-523 (1988).</li></ol>

The reversed-heart model is based on the well-established Langendorff heart perfusion technique12 and is performed by simply inverting the heart into an upside-down position and holding this position using a rigid intra-ventricular balloon catheter. In such a way, cardiac interstitial transudate can be physically separated from coronary venous effluent perfusate, dripping by gravity from the base of the heart9. The CT can be continuously collected by means of a thin and fle...

Heart failure (HF) is the leading cause of death in humans worldwide1. HF often occurs because of myocarditis, ischemic insults to the myocardium, and left ventricular remodeling, leading to the progressive deterioration of cardiac contractile function and patients&#39; quality of life. Although advances in cardiology and cardiac surgery have remarkably lowered HF mortality, they merely serve as transient &#34;delayers&#34; of an inevitably progressive disease process that carries signi&#64257;cant morbidity. Therefore, the current lack of effective treatment underscores the need to identify novel molecular targets that can prevent or even reverse HF. This includes changes in the extracellular matrix, uncontrolled cardiac immune response, and interactions between cardiac and non-cardiac cells2.
It is important to recognize that the microenvironment that cardiac cells are exposed to directly shapes the immune and regenerative response of the injured heart. In the isolated, saline-perfused heart, CT is generated on the heart surface in the form of small droplets that are derived from the interstitial fluid space (i.e., microenvironment), both under physiological and pathophysiological conditions3,4,5. Therefore, analysis of the CT (i.e., interstitial fluid) may help to identify factors that regulate cardiac metabolism and contractile function6 or influence immune cell functions after migration into the injured heart. Potentially, this may lead to the development of novel therapeutic strategies for the treatment of HF.
The collection of CT from murine hearts is technically challenging. In regular Langendorff-perfused hearts, the exclusive collection of CT is difficult because the mixture of the CT with coronary venous effluent perfusate unpredictably dilutes any concentration of metabolites/enzymes released from the interstitial space. One possible strategy to overcome this limitation is to exclude the venous effluent by cannulating the pulmonary and simultaneously ligating the pulmonary vein7. However, this method faces difficulties associated with the cannulation and ligation of the pulmonary artery and vein, causing potential leakage of venous effluent into the cardiac transudate. The concept of using a reverse heart model was first introduced by the group of Kammermeier, who inverted the isolated, perfused heart into an upside-down position and placed a thin latex cap on the epicardial surface to continuously sample CT without the contamination of venous effluent8,9. Using this procedure, CT was shown to provide a very sensitive measure of the metabolites released from the heart9, the capillary transfer of fatty acids8, and viral particles10.
More recently, paracrine factors that may regulate the local immune response and augment cardiac angiogenesis11 have been implicated in the beneficial effects of stem cell-based therapy for heart disease. The analysis of CT in the reversed heart may help to chemically identify these individual paracrine factors. In addition, CT may help to identify the factors involved in the in vivo activation of immune cells in the heart.
The detailed description of CT collection from the heart surface, provided here, is experimentally useful for researchers studying the interplay of immune cells, fibroblasts, endothelial cells, and cardiomyocytes in relation to overall cardiac function. As mentioned above, the interstitial fluid carries the information for cell-to-cell communication within the heart, which can conveniently be assessed by the collection of CT. The detailed technical description, including a video protocol of how to collect CT from the reversed heart, should facilitate the future application of this unique technique.

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the inverted heart model for interstitial transudate collection from the isolated rat heart

The present protocol describes a unique approach that enables the collection of cardiac transudate (CT) from the isolated, saline-perfused rat heart. After isolation and retrograde perfusion of the heart according to the Langendorff technique, the heart is inverted into an upside-down position and is mechanically stabilized by a balloon catheter inserted into the left ventricle. Then, a thin latex cap &#8211; previously cast to match the average size of the rat heart &#8211; is placed over the epicardial surface. The outlet of the latex cap is connected to silicon tubing, with the distal opening 10 cm below the base level of the heart, creating slight suction. CT continuously produced on the epicardial surface is collected in ice-cooled vials for further analysis. The rate of CT formation ranged from 17 to 147 &#181;L/min (n = 14) in control and infarcted hearts, which represents 0.1-1% of the coronary venous effluent perfusate. Proteomic analysis and high performance liquid chromatography (HPLC) revealed that the collected CT contains a wide spectrum of proteins and purinergic metabolites.

The reversed-heart model enables the collection of cardiac interstitial transudate in an isolated, retro-perfused rat heart (Figure 1A-C). When perfused at a constant pressure of 100 cmH2O, the rate of interstitial fluid formation ranged between 17 and 147 &#181;L/min, amounting to 0.1-1% of the coronary venous effluent in the isolated heart.
The protein content of the CT...

Watch this Scientific Journal Video about The Inverted Heart Model for Interstitial Transudate Collection from the Isolated Rat Heart at JoVE.com 

The Inverted Heart Model for Interstitial Transudate Collection from the Isolated Rat Heart (Video) | JoVE 

This protocol describes a method to collect cardiac interstitial fluid from the isolated, perfused rat heart. To physically separate interstitial transudate from coronary venous effluent perfusate, the Langendorff perfused heart is inverted, and the transudate (interstitial fluid) formed on the cardiac surface is collected using a soft latex cap.

The Inverted Heart Model for Interstitial Transudate Collection from the Isolated Rat Heart

The present protocol describes a unique approach that enables the collection of cardiac transudate (CT) from the isolated, saline-perfused rat heart. ...

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Isolation of Atrial Cardiomyocytes from a Rat Model of Metabolic Syndrome-related Heart Failure with Preserved Ejection Fraction (Video) | JoVE 

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

A Cryoinjury Model to Study Myocardial Infarction in the Mouse

A Cryoinjury Model to Study Myocardial Infarction in the Mouse (Video) | JoVE 

The use of animal models is essential for developing new therapeutic strategies for acute coronary syndrome and its complications. In this article, we ...

An Image Guided Transapical Mitral Valve Leaflet Puncture Model of Controlled Volume Overload from Mitral Regurgitation in the Rat

An Image Guided Transapical Mitral Valve Leaflet Puncture Model of Controlled Volume Overload from Mitral Regurgitation in the Rat (Video) | JoVE 

Mitral regurgitation (MR) is a widely prevalent heart valve lesion, which causes cardiac remodeling and leads to congestive heart failure. Though the ...