B.K.M. is supported by a National Institute of Health (NIH) / National Heart Lung and Blood Institute (NHLBI) grant (R01- HL085487). M.H. is supported by the American Heart Association National (AHA) Scientist Development Grant (SDG) and the Society of Geriatric Cardiology.

1. Preparation of the Operating Area
<ol>
	<li>The operating surface is first prepared by disinfecting the area with 70% ethanol.</li>
	<li>All surgical instruments are to be sterilized with a hot bead sterilizer before surgery (and in between individual rat surgeries). These instruments include: surgical scissors (2), forceps (1), curved forceps (1), needle holder (2), and a chest rectractor.</li>
	<li>Cotton gauze and applicators are to be available in order to manage bleeding.</li>
	<li>Adjust the temperature of a homeothermic blanket system (heating pad), to be used to maintain the body temperature of the animal at 37&plusmn;1&deg;C, in order to prevent a dramatic fall in body temperature; the size of an infarct is dependent on the duration of occlusion as well body temperature (Tarnavski et al.).</li>
	<li>Prior to anesthesia, each animal will receive a dosage of buprenorphine (0.1-2.5 mg/kg SQ) and carprofen oral tablets (Rimadyl, Bioserv tablet; 1 tablet) the day before the surgery. At the time immediately prior to the surgery, each animal will again receive a dosage of buprenorphine (0.1-2.5 mg/kg SQ).</li>
<li>Note: For studies involving the assessment of cardiac function and cardiac remodeling over a period of time (7- to 28-days and beyond), a dosage of buprenorphine (0.1-2.5 mg/kg SQ) will be administered, every 6-hours after surgery for 24-48 hours post surgery and carprofen oral tablets will be provided once daily (see "Post-Operative Recovery").</li>
</ol>
2. Preparation and Intubation of Rats
<ol>
	<li>Prior to the induction of anesthesia, animals will be examined to identify any pre-existing conditions that may complicate surgical outcome; physical examination will include visual inspection and measurement of heart rate, respiratory rate, and body temperature and body weight.</li>
	<li>If the animals are determined to be healthy, 250-300 gram male rats (e.g. Sprague Dawley) will be weighed and identifying characteristics will be recorded.</li>
	<li>Rats are then anesthetized with 80 mg/kg of pentobarbital; the correct level of anesthesia is verified through the limb withdrawal response by applying pressure of the rat nail bed (toe-pinch reflex) and periodically throughout the procedure.</li>
<li>Buprenorphine (0.1-2.5 mg/kg; SQ) will be administered at the time of surgery.</li>	
<li>An electric shaver is then used to shave the fur form the neck and chest areas. Note: For studies involving the assessment of cardiac function and cardiac remodeling over a period of time (7- to 28-days and beyond), fur must be completely shaved away from the surgical area to maintain sterility.</li>
	<li>The shaved areas are to be scrubbed and disinfected with betadine solution followed by wiping the area with 70% alcohol; this betadine and alcohol scrub and disinfection procedure is to be repeated three times.</li>
	<li>The anesthetized animal is then placed in a supine position on a homeothermic blanket system (heating pad) in order to maintain body temperature.</li>
	<li>Endotracheal intubation is then performed by first carefully exposing the trachea, to enhance visualization of intubation, by making a 5 mm mid-neck incision and retraction of muscle tissue, just above the trachea. </li>
	<li>This allows better visualization for insertion of the endotracheal tube (polyethylene size 90; PE 90) where the tubing is beveled on the edge for ease of entrance through the larynx.</li>
	<li>The endotracheal tube is inserted into the trachea, with visualization aided by using a surgical stereo microscope, by carefully manipulating the tongue while viewing the trachea and the entrance of the endotracheal tube.</li>
	<li>Mechanical ventilation is then achieved by connecting the endotracheal tube to Kent Scientific ventilator (TOPO Dual Mode) cycling at 80 breaths per minute and a tidal volume of 1.2 ml per 100 gram body weight.</li>
	<li>A sterile drape is then placed over the rat, leaving only the surgical area exposed, thus minimizing potential contamination during the operation. </li>
	<li>The animal surgeon and assistants are to wear mask, gown, and gloves; with gloves being changed between individual rat surgeries.</li>
</ol>
3. Transient Left Anterior Descending (LAD) Artery Ligation
<ol>
	<li>Once steady breathing is established, the chest is opened at the left fourth intercostal space, by using hemostats or round end scissors to open the space, but without cutting the tissue so that the risk of bleeding can be reduced.</li>
	<li>A chest retractor is then positioned within the fourth intercostal space; in order to spread the ribs so that the left ventricle (LV) is exposed, taking maximal care not to damage the lung.</li>
	<li>The pericardium is then opened, but left in place if possible.</li>
	<li>The proximal left anterior descending (LAD) artery will be identified (Figure-1), with the use of a surgical stereo microscope, and the LAD is then transiently ligated (or can be tied with a slipknot) using a 6-0 prolene suture for a 30-minute ischemic period (Figure-1), but without exteriorization of the heart.</li>
	<li>To allow cardiac reperfusion, microsurgical scissors are used to cut the knot in the ligature (or by releasing the slipknot), made by the 6-0 prolene suture, 30-minutes following ligation of the LAD.</li>
	<li>In sham control rats, the procedure is identical, except the LAD is not transiently ligated.</li>
	<li>The outflow is then briefly (1-2 seconds) pinched off on the respirator to allow re-inflation of the lungs. </li>
	<li>The chest retractor is then removed and the ribs are drawn together using a 2-0 prolene suture with an interrupted suture pattern. </li>
	<li>Once the ribs are closed, the outflow of the ventilator is again briefly (1-2 seconds) pinched off to ensure proper breathing.</li>
	<li>The skin is closed using 6-0 prolene suture with a continuous suture pattern.</li>
</ol>
4. Confirmation of Successful Ligation of the LAD Artery
<ol>
	<li>Proper ligation of the LAD will be confirmed by observing blanching of myocardial tissue distal to the suture as well as dysfunction of the anterior wall; as observed during the transient LAD ligation. </li>
	<li>Reperfusion will be verified by the return of red color to the myocardial tissue and the demonstration of some recovery of anterior wall motion; as observed immediately following the transient LAD ligation.</li>
	<li>Note: For studies involving the assessment of cardiac function and cardiac remodeling over a period of time (7- to 28-days and beyond), an echocardiogram will be performed 24-hours post LAD ligation, in order to determine the extent of the induced myocardial infarction. All animals with a left ventricular fractional shortening (LV FS) &lt; 20%, as determined from an echocardiogram, as well as anterior wall motion dysfunction consistent with the infarct, will be identified and used for follow-up cardiac function and remodeling studies. Sham control rats are expected to have a normal LV FS between 40-45% (del Monte et al).</li>
</ol>
5. Post-Operative Recovery
<ol>
	<li>Once the surgery is complete, the sterile drape over the animal will be removed. The body temperature (by a rectal thermometer), respiratory rate (by visual inspection), heart rate (by palpitation), and abnormal signs of pain are to be monitored until the animal becomes ambulatory.</li>
	<li>The rat will be removed from the ventilator and then be placed in an animal cage and allowed to recover from anesthesia. In this cage, one end of the cage will be warmed by a heating pad to allow the rat to seek warmth. Also, water will be available to the rat during this observation period.</li>
	<li>If signs of dehydration are observed after surgery, 0.5 ml of sterile saline will be injected intraperitoneally (IP). </li>
	<li>Buprenorphine (0.1&#8208;2.5 mg/kg; SQ) will then be administered immediately after surgery, and during post-operative recovery.</li>
	<li>Note: For studies involving the assessment of cardiac function and cardiac remodeling over a period of time (7- to 28-days and beyond), a dosage of buprenorphine (0.1-2.5 mg/kg SQ) will be administered, every 6-hours after surgery for 24-48 hours post surgery. Carprofen oral tablets (Rimadyl, Bioserv tablet) will also be provided (1-tablet daily) for at least 48-hours post surgery.</li>
</ol>
6. Histological Evaluation of the Ischemia Reperfusion
<ol>
	<li>Following completion of the 4-hour reperfusion period, rats are to be anesthetized again with 80 mg/kg of pentobarbital and the LAD is ligated once again (or the slipknot can be retied) (Huang et al and Kaiser et al).</li>
	<li>1 ml of 5 % Evan's Blue (in phosphate-buffered saline) is then injected into the right ventricular chamber and allowed to perfuse for 2-minutes.</li>
	<li>After 2-minutes, 1 ml of saturated potassium chloride solution is then injected into the right ventricular chamber in order to stop the heart from beating.</li>
	<li>The heart is then rapidly excised, embedded in 2% agarose phosphate-buffered saline, and then sectioned in 1-2 mm slices.</li>
	<li>The viable myocardium is then stained with 2% triphenyl tetrazolium chloride (TTC) for 20-minutes and images of each slice is captured. The images are then quantified in order to identify the amount of myocardial area: (1) that is not-at-risk (blue sections), (2) area-at-risk (AAR; red and white sections), and (3) area that is infarcted (white "unstained" sections) (Figure-2).</li>
</ol>
7. Representative Results
For studies involving the assessment of infarct size and the area-at-risk (AAR), by Evan's blue dye and triphenyl tetrazolium chloride (TTC), following 4-hours of reperfusion, proper ligation of the LAD can be confirmed by observing blanching of myocardial tissue distal to the suture as well as dysfunction of the anterior wall; as observed during the transient LAD ligation. Reperfusion can be verified by the return of red color to the myocardial tissue and the demonstration of some recovery of anterior wall motion; observed immediately following the transient LAD ligation. For studies involving the assessment of cardiac function and cardiac remodeling over a period of time (7- to 28- days and beyond), proper ligation of the ischemia followed by reperfusion can also be confirmed by blanching of the tissue, dysfunction of the anterior wall and the return of red color with some recovery of the wall motion following reperfusion. In addition, impaired cardiac function can be determined by an echocardiogram being performed 24-hours post LAD ligation, in order to determine the extent of the induced myocardial infarction. All animals with a left ventricular fractional shortening (LV FS) &lt; 20%, as determined from an echocardiogram, as well as anterior wall motion dysfunction consistent with the infarct, will be identified and used for follow-up cardiac function and remodeling studies. Sham control rats are expected to have a normal LV FS between 40-45% (del Monte).
<img src="/files/ftp_upload/2464/2464fig1.jpg" alt="Figure 1" /> Figure 1. Schematic Diagram of a Myocardial Infarction and LAD Ligation. Schematic diagram of (1) myocardial infarction and (2) ligation of the left ventricular anterior descending (LAD) artery using a suture, used to mimic the myocardial infarction caused by plague formation.
<img src="/files/ftp_upload/2464/2464fig2.jpg" alt="Figure 2" /> Figure 2. Representative Image of a Rat Heart Following AMI. Representative image of the rat heart showing the area in blue (non-occluded coronary perfusion area), area in red (occluded coronary perfusion area indicating the area-at-risk (AAR) for infarction, and area in white/yellow (infarcted zone within the AAR).

<ol>
	<li>Hunt, S. A., Baker, D. W., Chin, M. H., Cinquegrani, M. P., Feldmanmd, A. M., Francis, G. S., Ganiats, T. G., Goldstein, S., Gregoratos, G., Jessup, M. L., Noble, R. J., Packer, M., Silver, M. A., Stevenson, L. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=SC.ACC/AHA+Guidelines+for+the+Evaluation+and+Management+of+Chronic+Heart+Failure+in+the+Adult:+Executive+Summary+A+Report+of+the+American+College+of+Cardiology/American+Heart+Association+Task+Force+on+Practice+Guidelines+(Committee+to+Revise+the+1995+Guidelines+for+the+Evaluation+and+Management+of+Heart+Failure).">SC.ACC/AHA Guidelines for the Evaluation and Management of Chronic Heart Failure in the Adult: Executive Summary A Report of the American College of Cardiology/American Heart Association Task Force on Practice Guidelines (Committee to Revise the 1995 Guidelines for the Evaluation and Management of Heart Failure).</a> Circulation. 104, 2996-3007 (2001).</li><li>Goldman, S., Raya, T. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rat+infarct+model+of+myocardial+infarction+and+heart+failure.">Rat infarct model of myocardial infarction and heart failure.</a> J Card Fail. 1, 169-177 (1995).</li><li>Tarnavski, O., McMullen, J. R., Schinke, M., Nie, Q., Kong, S., &amp;amp, I. z. u. m. o., S, . <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+cardiac+surgery:+comprehensive+techniques+for+the+generation+of+mouse+models+of+human+diseases+and+their+application+for+genomic+studies.">Mouse cardiac surgery: comprehensive techniques for the generation of mouse models of human diseases and their application for genomic studies.</a> Physiol Genomics. 16, 349-360 (2004).</li><li>Thygesen, K., Alpert, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Universal+Definition+of+Myocardial+Infarction.">Universal Definition of Myocardial Infarction.</a> Circulation. 116, 2634-2653 (2007).</li><li>Vasilyev, N., Williams, T., Brennan, M. L., Unzek, S., Zhou, X., Heinecke, J. W., Spitz, D. R., Topol, E. J., Hazen, S. L., Penn, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Myeloperoxidase-generated+oxidants+modulate+left+ventricular+remodeling+but+not+infarct+size+after+myocardial+infarction.">Myeloperoxidase-generated oxidants modulate left ventricular remodeling but not infarct size after myocardial infarction.</a> Circulation. 112, 2812-2820 (2005).</li><li>McConnell, B. K., Popovic, Z., Mal, N., Lee, K., Bautista, J., Forudi, F., Schwartzman, R., Jin, J. P., Penn, M., Bond, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Disruption+of+protein+kinase+A+interaction+with+A-kinase-anchoring+proteins+in+the+heart+in+vivo:+effects+on+cardiac+contractility,+protein+kinase+A+phosphorylation,+and+troponin+I+proteolysis.">Disruption of protein kinase A interaction with A-kinase-anchoring proteins in the heart in vivo: effects on cardiac contractility, protein kinase A phosphorylation, and troponin I proteolysis.</a> J Biol Chem. 284, 1583-1592 (2009).</li><li>Monte, D. e. l., Butler, F., Boecker, K., Gwathmey, W., K, J., Hajjar, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Novel+technique+of+aortic+banding+followed+by+gene+transfer+during+hypertrophy+and+heart+failure.">Novel technique of aortic banding followed by gene transfer during hypertrophy and heart failure.</a> Physiol. Genomics. 9, 49-56 (2002).</li><li>Huang, M. H., Nguyen, V., Wu, Y., Rastogi, S., McConnell, B. K., Wijaya, C., Uretsky, B. F., Poh, K. K., Tan, H. C., Cheng, J., Perin, E., Fujise, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synergistic+Infarct+Size+Reduction+by+Combination+Therapy+with+Milrinone+and+Esmolol+at+Early+Reperfusion+during+Acute+Myocardial+Infarction.">Synergistic Infarct Size Reduction by Combination Therapy with Milrinone and Esmolol at Early Reperfusion during Acute Myocardial Infarction.</a> American Journal of Physiology. , .</li><li>Kaiser, R. A., Bueno, O. F., Lips, D. J., Doevendans, P. A., Jones, F., Kimball, T. F., Molkentin, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Targeted+inhibition+of+p38+mitogen-activated+protein+kinase+antagonizes+cardiac+injury+and+cell+death+following+ischemia-reperfusion+in+vivo.">Targeted inhibition of p38 mitogen-activated protein kinase antagonizes cardiac injury and cell death following ischemia-reperfusion in vivo.</a> J Biol Chem. 279, 15524-15530 (2004).</li></ol>

No conflicts of interest declared.

An acute myocardial infarction (AMI) in rats, also referred to as ischemia-reperfusion (IR), is commonly used to mimic human ventricular remodeling leading to heart failure. The progression of LV dysfunction and heart failure in animal models (rats and mice) is similar to that observed in humans who sustain a MI, survive and then subsequently develops heart failure (Goldman). This procedure outlined one version of this technique and also contained additional notes describing methodological variations. Additional such var...

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acute myocardial infarction in rats

With heart failure leading the cause of death in the USA (Hunt), biomedical research is fundamental to advance medical treatments for cardiovascular diseases. Animal models that mimic human cardiac disease, such as myocardial infarction (MI) and ischemia-reperfusion (IR) that induces heart failure as well as pressure-overload (transverse aortic constriction) that induces cardiac hypertrophy and heart failure (Goldman and Tarnavski), are useful models to study cardiovascular disease. In particular, myocardial ischemia (MI) is a leading cause for cardiovascular morbidity and mortality despite controlling certain risk factors such as arteriosclerosis and treatments via surgical intervention (Thygesen). Furthermore, an acute loss of the myocardium following myocardial ischemia (MI) results in increased loading conditions that induces ventricular remodeling of the infarcted border zone and the remote non-infarcted myocardium. Myocyte apoptosis, necrosis and the resultant increased hemodynamic load activate multiple biochemical intracellular signaling that initiates LV dilatation, hypertrophy, ventricular shape distortion, and collagen scar formation. This pathological remodeling and failure to normalize the increased wall stresses results in progressive dilatation, recruitment of the border zone myocardium into the scar, and eventually deterioration in myocardial contractile function (i.e. heart failure). The progression of LV dysfunction and heart failure in rats is similar to that observed in patients who sustain a large myocardial infarction, survive and subsequently develops heart failure (Goldman). The acute myocardial infarction (AMI) model in rats has been used to mimic human cardiovascular disease; specifically used to study cardiac signaling mechanisms associated with heart failure as well as to assess the contribution of therapeutic strategies for the treatment of heart failure. The method described in this report is the rat model of acute myocardial infarction (AMI). This model is also referred to as an acute ischemic cardiomyopathy or ischemia followed by reperfusion (IR); which is induced by an acute 30-minute period of ischemia by ligation of the left anterior descending artery (LAD) followed by reperfusion of the tissue by releasing the LAD ligation (Vasilyev and McConnell). This protocol will focus on assessment of the infarct size and the area-at-risk (AAR) by Evan's blue dye and triphenyl tetrazolium chloride (TTC) following 4-hours of reperfusion; additional comments toward the evaluation of cardiac function and remodeling by modifying the duration of reperfusion, is also presented. Overall, this AMI rat animal model is useful for studying the consequence of a myocardial infarction on cardiac pathophysiological and physiological function.

Watch this Scientific Journal Video about Acute Myocardial Infarction in Rats at JoVE.com

Acute Myocardial Infarction in Rats

The rat model of acute myocardial infarction (AMI) is useful to study the consequence of a MI on cardiac pathophysiological and physiological function.

With heart failure leading the cause of death in the USA (Hunt), biomedical research is fundamental to advance medical treatments for cardiovascular ...

Research

JoVE Journal

Medicine

52.8K ビュー.  University of Texas Medical Branch. 急性心筋梗塞（AMI）のラットモデルでは、心臓の病態生理学的および生理学的機能に関するMIの結果を研究するのに便利です。

ビデオ: Acute Myocardial Infarction in Rats

Induction of Myocardial Infarction in Adult Zebrafish Using Cryoinjury

The mammalian heart is incapable of significant regeneration following an acute injury such as myocardial infarction1. By contrast, urodele amphibians and teleost fish retain a remarkable capacity for cardiac regeneration with little or no scarring throughout life2,3. It is not known why only some non-mammalian vertebrates can recreate a complete organ from remnant tissues4,5. To understand the molecular and cellular differences between regenerative responses in different species, we need to use similar approaches for inducing acute injuries.
In mammals, the most frequently used model to study cardiac repair has been acute ischemia after a ligation of the coronary artery or tissue destruction after cryoinjury6,7. The cardiac regeneration in newts and zebrafish has been predominantly studied after a partial resection of the ventricular apex2,3. Recently, several groups have established the cryoinjury technique in adult zebrafish8-10. This method has a great potential because it allows a comparative discussion of the results obtained from the mammalian and non-mammalian species.
Here, we present a method to induce a reproducible disc-shaped infarct of the zebrafish ventricle by cryoinjury. This injury model is based on rapid freezing-thawing tissue, which results in massive cell death of about 20% of cardiomyocytes of the ventricular wall. First, a small incision was made through the chest with iridectomy scissors to access the heart. The ventricular wall was directly frozen by applying for 23-25 seconds a stainless steel cryoprobe precooled in liquid nitrogen. To stop the freezing of the heart, fish water at room temperature was dropped on the tip of the cryoprobe. The procedure is well tolerated by animals, with a survival rate of 95%.
To characterize the regenerative process, the hearts were collected and fixed at different days after cryoinjury. Subsequently, the specimen were embedded for cryosectioning. The slides with sections were processed for histological analysis, in situ hybridization and immunofluorescence. This undertaking enhances our understanding of the factors that are required for the regenerative plasticity in the zebrafish, and provide new insights into the machinery of cardiac regeneration. A conceptual and molecular understanding of heart regeneration in zebrafish will impact both developmental biology and regenerative medicine.

Zebrafish represents a valuable model to study the mechanisms of heart regeneration in vertebrates. Here, we present a protocol for induction of a heart infarct in adult zebrafish using cryoinjury. This method results in massive cell death within 20% of the ventricular wall, similar to that observed in mammalian infarcts.

The mammalian heart is incapable of significant regeneration following an acute injury such as myocardial infarction1. By contrast, urodele amphibians and ...

MRI and PET in Mouse Models of Myocardial Infarction

Myocardial infarction is one of the leading causes of death in the Western world. The similarity of the mouse heart to the human heart has made it an ideal model for testing novel therapeutic strategies.
In vivo magnetic resonance imaging (MRI) gives excellent views of the heart noninvasively with clear anatomical detail, which can be used for accurate functional assessment. Contrast agents can provide basic measures of tissue viability but these are nonspecific. Positron emission tomography (PET) is a complementary technique that is highly specific for molecular imaging, but lacks the anatomical detail of MRI. Used together, these techniques offer a sensitive, specific and quantitative tool for the assessment of the heart in disease and recovery following treatment.
In this paper we explain how these methods are carried out in mouse models of acute myocardial infarction. The procedures described here were designed for the assessment of putative protective drug treatments. We used MRI to measure systolic function and infarct size with late gadolinium enhancement, and PET with fluorodeoxyglucose (FDG) to assess metabolic function in the infarcted region. The paper focuses on practical aspects such as slice planning, accurate gating, drug delivery, segmentation of images, and multimodal coregistration. The methods presented here achieve good repeatability and accuracy maintaining a high throughput.

We describe how to perform MRI and PET imaging of the mouse heart. The protocol is tailored to assess treatment efficacy in models of myocardial infarction and heart failure.

Myocardial infarction is one of the leading causes of death in the Western world. The similarity of the mouse heart to the human heart has made it an ...

Myocardial Infarction and Functional Outcome Assessment in Pigs

Introduction of newly discovered cardiovascular therapeutics into first-in-man trials depends on a strictly regulated ethical and legal roadmap. One important prerequisite is a good understanding of all safety and efficacy aspects obtained in a large animal model that validly reflect the human scenario of myocardial infarction (MI). Pigs are widely used in this regard since their cardiac size, hemodynamics, and coronary anatomy are close to that of humans. Here, we present an effective protocol for using the porcine MI model using a closed-chest coronary balloon occlusion of the left anterior descending artery (LAD), followed by reperfusion. This approach is based on 90 min of myocardial ischemia, inducing large left ventricle infarction of the anterior, septal and inferoseptal walls. Furthermore, we present protocols for various measures of outcome that provide a wide range of information on the heart, such as cardiac systolic and diastolic function, hemodynamics, coronary flow velocity, microvascular resistance, and infarct size. This protocol can be easily tailored to meet study specific requirements for the validation of novel cardioregenerative biologics at different stages (i.e. directly after the acute ischemic insult, in the subacute setting or even in the chronic MI once scar formation has been completed). This model therefore provides a useful translational tool to study MI, subsequent adverse remodeling, and the potential of novel cardioregenerative agents.

This protocol describes the porcine myocardial infarction (MI) model using a 90 min closed-chest coronary balloon occlusion of the left anterior descending artery (LAD), followed by reperfusion. Furthermore, the protocol for several outcome parameters, such as cardiac function, hemodynamics, microvascular resistance, and infarct size, are also presented.

Introduction of newly discovered cardiovascular therapeutics into first-in-man trials depends on a strictly regulated ethical and legal roadmap. One ...

Minimal Invasive Surgical Procedure of Inducing Myocardial Infarction in Mice

Myocardial infarction still remains the main cause of death in western countries, despite considerable progress in the stent development area in the last decades. For clarification of the underlying mechanisms and the development of new therapeutic strategies, the availability of valid animal models are mandatory. Since we need new insights into pathomechanisms of cardiovascular diseases under in vivo conditions to combat myocardial infarction, the validity of the animal model is a crucial aspect. However, protection of animals are highly relevant in this context. Therefore, we establish a minimally invasive and simple model of myocardial infarction in mice, which assures a high reproducibility and survival rate of animals. Thus, this models fulfils the requirements of the 3R principle (Replacement, Refinement and Reduction) for animal experiments and assure the scientific information needed for further developing of therapeutical strategies for cardiovascular diseases.

A highly reproducible model for myocardial infarction in mice with minimal invasive manipulations is described. The model can be easily performed, resulting in a high reproducibility and survival rate. Thus, the described model will reduce the number of required animals as requested by the 3R principle (Replacement, Refinement and Reduction).

Myocardial infarction still remains the main cause of death in western countries, despite considerable progress in the stent development area in the last ...

Primary Outcome Assessment in a Pig Model of Acute Myocardial Infarction

Mortality after acute myocardial infarction remains substantial and is associated with significant morbidity, like heart failure. Novel therapeutics are therefore required to confine cardiac damage, promote survival and reduce the disease burden of heart failure. Large animal experiments are an essential part in the translational process from experimental to clinical therapies. To optimize clinical translation, robust and representative outcome measures are mandatory. The present manuscript aims to address this need by describing the assessment of three clinically relevant outcome modalities in a pig acute myocardial infarction (AMI) model: infarct size in relation to area at risk (IS/AAR) staining, 3-dimensional transesophageal echocardiography (TEE) and admittance-based pressure-volume (PV) loops. Infarct size is the main determinant driving the transition from AMI to heart failure and can be quantified by IS/AAR staining. Echocardiography is a reliable and robust tool in the assessment of global and regional cardiac function in clinical cardiology. Here, a method for three-dimensional transesophageal echocardiography (3D-TEE) in pigs is provided. Extensive insight into cardiac performance can be obtained by admittance-based pressure-volume (PV) loops, including intrinsic parameters of myocardial function that are pre- and afterload independent. Combined with a clinically feasible experimental study protocol, these outcome measures provide researchers with essential information to determine whether novel therapeutic strategies could yield promising targets for future testing in clinical studies.

Reliable and accurate outcome assessment is the key for translation of preclinical therapies into clinical treatment. The current paper describes how to assess three clinically relevant primary outcome parameters of cardiac performance and damage in a pig acute myocardial infarction model.

Mortality after acute myocardial infarction remains substantial and is associated with significant morbidity, like heart failure. Novel therapeutics are ...

Myocardial Infarction in Neonatal Mice, A Model of Cardiac Regeneration

Myocardial infarction induced by coronary artery ligation has been used in many animal models as a tool to study the mechanisms of cardiac repair and regeneration, and to define new targets for therapeutics. For decades, models of complete heart regeneration existed in amphibians and fish, but a mammalian counterpart was not available. The recent discovery of a postnatal window during which mice possess regenerative capabilities has led to the establishment of a mammalian model of cardiac regeneration. A surgical model of mammalian cardiac regeneration in the neonatal mouse is presented herein. Briefly, postnatal day 1 (P1) mice are anesthetized by isoflurane and placed on an ice pad to induce hypothermia. After the chest is opened, and the left anterior descending coronary artery (LAD) is visualized, a suture is placed around the LAD to inflict myocardial ischemia in the left ventricle. The surgical procedure takes 10-15 min. Visualizing the coronary artery is crucial for accurate suture placement and reproducibility. Myocardial infarction and cardiac dysfunction are confirmed by triphenyl-tetrazolium chloride (TTC) staining and echocardiography, respectively. Complete regeneration 21 days post myocardial infarction is verified by histology. This protocol can be used to as a tool to elucidate mechanisms of mammalian cardiac regeneration after myocardial infarction.

This protocol describes a highly reproducible model of cardiac regeneration by surgical induction of myocardial infarction in the left ventricle of postnatal day 1 mice. The method involves induction of hypothermic anesthesia and ligation of the left anterior descending coronary artery.

Myocardial infarction induced by coronary artery ligation has been used in many animal models as a tool to study the mechanisms of cardiac repair and ...

Real-time Pressure-volume Analysis of Acute Myocardial Infarction in Mice

Acute myocardial infarction can lead to acute heart failure and cardiogenic shock. The evaluation of hemodynamics is critical for the evaluation of any potential therapeutic approach directed against acute left ventricular (LV) dysfunction. Current imaging modalities (e.g., echocardiography and magnetic resonance imaging) have several limitations since data on LV pressure cannot directly be measured. LV catheterization in mice undergoing coronary artery occlusion could serve as a novel method for a real-time evaluation of LV function.
At the beginning of the procedure, mice were anesthetized followed by endotracheal intubation. For LV catheterization, the right carotid artery was exposed via middle-neck incision. The catheter was introduced and placed into the LV cavity. Left thoracotomy was conducted and the left main coronary artery (LCA) was ligated. To induce reperfusion, the suture was released after 45 min. Pressure-volume data was recorded at all times.
Ligation of the LCA caused a decrease in LV systolic function as evidenced by a 30% reduction in stroke volume, LV ejection fraction (EF), and cardiac output. Maximum dP/dt as a parameter for LV contractility was also significantly reduced and diastolic function was severely impaired (minimum dP/dt -40%). Reperfusion over a period of 20 min did not lead to a complete recovery of LV function.
Real-time pressure-volume analysis served as a valid procedure for monitoring cardiac function during acute myocardial infarction in mice. Maintaining stable anesthesia and a standardized surgical approach was crucial to ensure valid results. As the early phase of acute myocardial infarction is critical for morbidity and mortality, the delineated method could be beneficial for preclinical evaluation of new strategies for cardioprotection.

Acute myocardial infarction in mice induces acute but incompletely characterized changes in left ventricular (LV) function. LV catheterization in mice undergoing coronary artery occlusion serves as a novel method for a real-time evaluation of LV function.

Acute myocardial infarction can lead to acute heart failure and cardiogenic shock. The evaluation of hemodynamics is critical for the evaluation of any ...

Digital PCR for Quantifying Circulating MicroRNAs in Acute Myocardial Infarction and Cardiovascular Disease

Circulating serum microRNAs (miRNAs) have shown promise as biomarkers for the cardiovascular disease and acute myocardial infarction (AMI), being released from the cardiovascular cells into the circulation. Circulating miRNAs are highly stable and can be quantified. The quantitative expression of specific miRNAs can be linked to the pathology, and some miRNAs show high tissue and disease specificity. Finding novel biomarkers for cardiovascular diseases is of importance for medical research. Quite recently, digital polymerase chain reaction (dPCR) has been invented. dPCR, combined with fluorescent hydrolysis probes, enables specific direct absolute quantification. dPCR exhibits superior technical qualities, including a low variability, high linearity, and high sensitivity compared to the quantitative polymerase chain reaction (qPCR). Thus, dPCR is a more accurate and reproducible method for directly quantifying miRNAs, particularly for the use in large multi-center cardiovascular clinical trials. In this publication, we describe how to effectively perform digital PCR in order to assess the absolute copy number in serum samples.

Circulating microRNAs have shown promise as biomarkers for cardiovascular diseases and acute myocardial infarctions. In this study, we describe a protocol for miRNA extraction, reverse transcription, and digital PCR for the absolute quantification of miRNAs in the serum of patients with cardiovascular disease.

Circulating serum microRNAs (miRNAs) have shown promise as biomarkers for the cardiovascular disease and acute myocardial infarction (AMI), being released ...

A Cryoinjury Model to Study Myocardial Infarction in the Mouse

The use of animal models is essential for developing new therapeutic strategies for acute coronary syndrome and its complications. In this article, we demonstrate a murine cryoinjury infarct model that generates precise infarct sizes with high reproducibility and replicability. In brief, after intubation and sternotomy of the animal, the heart is lifted from the thorax. The probe of a handheld liquid nitrogen delivery system is applied onto the myocardial wall to induce cryoinjury. Impaired ventricular function and electrical conduction can be monitored with echocardiography or optical mapping. Transmural myocardial remodeling of the infarcted area is characterized by collagen deposition and loss of cardiomyocytes. Compared to other models (e.g., LAD-ligation), this model utilizes a handheld liquid nitrogen delivery system to generate more uniform infarct sizes.

This article demonstrates a model to study cardiac remodeling after myocardial cryoinjury in mice.

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

Semi-Minimal Invasive Method to Induce Myocardial Infarction in Rats and the Assessment of Cardiac Function by an Isolated Working Heart System

Myocardial infarction (MI) remains the main contributor to morbidity and mortality worldwide. Therefore, research on this topic is mandatory. An easily and highly reproducible MI induction procedure is required to obtain further insight and better understanding of the underlying pathological changes. This procedure can also be used to evaluate the effects or potency of new and promising treatments (as drugs or interventions) in acute MI, subsequent remodeling and heart failure (HF). After intubation and pre-operative preparation of the animal, an anesthetic protocol with isoflurane was performed, and the surgical procedure was conducted&#160;quickly. Using a minimally invasive approach, the left anterior descending artery (LAD) was located and occluded by a ligature. The occlusion can be performed acutely for subsequent reperfusion (ischemia/reperfusion injury). Alternatively, the vessel can be ligated permanently to investigate the development of chronic MI, remodeling or HF. Despite common pitfalls, the drop-out rates are minimal. Various treatments such as remote ischemic conditioning can be examined for their cardioprotective potential pre-, peri- and post-operatively. The post-operative recovery was quick as the anesthesia was precisely controlled and the duration of the operation was short. Post-operative analgesia was administered for three days. The minimally invasive procedure reduces the risk of infection and inflammation. Furthermore, it facilitates rapid recovery. The &#8220;working heart&#8221; measurements were performed ex vivo and enabled precise control of preload, afterload and flow. This procedure requires specific equipment and training for adequate performance. This manuscript provides a detailed step-by-step introduction for conducting these measurements.

This article presents an efficient method to perform myocardial ischemia and subsequent chronic reperfusion in rats using a minimally invasive approach. In addition, left ventricular hemodynamic function of rats is assessed by echocardiography and isolated working heart methods.&#160;

Myocardial infarction (MI) remains the main contributor to morbidity and mortality worldwide. Therefore, research on this topic is mandatory. An easily ...

Acute Myocardial Infarction in Rats

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