Name: studying left ventricular reverse remodeling by aortic debanding in rodents
Uploaded: 2021-07-14T13:00:00
Description: Watch this Scientific Journal Video about Studying Left Ventricular Reverse Remodeling by Aortic Debanding in Rodents at JoVE.com

The authors thank Portuguese Foundation for Science and Technology (FCT), European Union, Quadro de Refer&#234;ncia Estrat&#233;gico Nacional (QREN), Fundo Europeu de Desenvolvimento Regional (FEDER) and Programa Operacional Factores de Competitividade (COMPETE) for funding UnIC (UID/IC/00051/2013) research unit. This project is supported by FEDER through COMPETE 2020 &#8211; Programa Operacional Competitividade E Internacionaliza&#231;&#227;o (POCI), the project DOCNET (NORTE-01-0145-FEDER-000003), supported by Norte Portugal regional operational programme (NORTE 2020), under the Portugal 2020 partnership agreement, through the European Regional Development Fund (ERDF), the project NETDIAMOND (POCI-01-0145-FEDER-016385), supported by European Structural And Investment Funds, Lisbon&#8217;s regional operational program 2020. Daniela Miranda-Silva and Patr&#237;cia Rodrigues are funded by Funda&#231;&#227;o para a Ci&#234;ncia e Tecnologia (FCT) by fellowship grants (SFRH/BD/87556/2012 and SFRH/BD/96026/2013 respectively). &#160;

All animal experiments comply with the Guide for the Care and Use of Laboratory Animals (NIH Publication no. 85&#8211;23, revised 2011) and the Portuguese law on animal welfare (DL 129/92, DL 197/96; P 1131/97). The competent local authorities approved this experimental protocol (018833). Seven-week-old male C57B1/J6 mice were maintained in appropriate cages, with a regular 12/12 h light-dark cycle environment, a temperature of 22 &#176;C and 60% humidity with access to water and a standard diet ad libitum.
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The model proposed herein mimics the process of LV remodeling and RR after aortic banding and debanding, respectively. Therefore, it represents an excellent experimental model to advance our knowledge on the molecular mechanisms involved in the adverse LV remodeling and to test novel therapeutic strategies able to induce myocardial recovery of these patients. This protocol details steps on how to create a rodent animal model of aortic banding and debanding with a minimally invasive and highly conservative surgical techni...

The constriction of the transverse or ascending aorta in the mouse is a widely used experimental model for pressure overload-induced cardiac hypertrophy, diastolic and systolic dysfunction and heart failure1,2,3,4. Aortic-constriction initially leads to compensated left ventricle (LV) concentric hypertrophy to normalize wall stress1. However, under certain circumstances, such as prolonged cardiac overload, this hypertrophy is insufficient to decrease the wall stress, triggering diastolic and systolic dysfunction (pathological hypertrophy)5. In parallel, changes in extracellular matrix (ECM) lead to the collagen deposition and crosslinking in a process known as fibrosis, which can be subdivided into replacement fibrosis and reactive fibrosis. Fibrosis is, in most of the cases, irreversible and compromises myocardial recovery after overload relief6,7. Nevertheless, recent cardiac magnetic resonance imaging studies revealed that reactive fibrosis is able to regress in the long term8. Altogether, fibrosis, hypertrophy and cardiac dysfunction are part of a process known as myocardial remodeling that rapidly progresses towards heart failure (HF).
Understanding the features of myocardial remodeling has become a major objective for limiting or reversing its progression, the latter known as reverse remodeling (RR). The term RR includes any myocardial alteration chronically reversed by a given intervention, such pharmacological therapy (e.g., antihypertensive medication), valve surgery (e.g., aortic stenosis) or ventricular assist devices (e.g., chronic HF). However, RR is often incomplete due to the prevailing hypertrophy or systolic/diastolic dysfunction. Thus, the clarification of RR underlying mechanisms and novel therapeutic strategies are still missing, which is mostly due to the impossibility to access and study human myocardial tissue during RR in most of these patients.
To overcome this limitation, rodent models have played a significant role in advancing our understanding of the signaling pathways involved in HF progression. Specifically, aortic debanding of mice with an aortic constriction represents a useful model to study the molecular mechanisms underlying adverse LV remodeling9 and RR10,11 as it allows the collection of myocardial samples at different time points in these two phases. Moreover, it provides an excellent experimental setting to test potential novel targets that can promote/accelerate RR. For instance, in aortic stenosis context, this model might provide information about the molecular mechanisms involved in the vast diversity of myocardial response underlying the (in)completeness of the RR6,12, as well as, the optimal timing for valve replacement, which represents a major shortcoming of the current knowledge. Indeed, the optimal timing for this intervention is a subject of debate, mainly because it is defined based on the magnitude of aortic gradients. Several studies advocate that this time point might be too late for the myocardial recovery as fibrosis and diastolic dysfunction are often already present12.
To our knowledge, this is the only animal model that recapitulates the process of both myocardial remodeling and RR taking place in conditions such as aortic stenosis or hypertension before and after valve replacement or the onset of anti-hypertensive medication, respectively.
Seeking to address the challenges summarized above, we describe a surgical animal model that can be implemented both in mice and rats, addressing the differences between these two species. We describe the main steps and details involved when carrying out these surgeries. Finally, we report the most significant changes taking place in the LV immediately before and throughout RR.

<table border="0" cellpadding="0" cellspacing="0" style="width:965px;" width="964">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Absorption Spears</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">18105-03</td>
			<td style="width:393px;">To absorb fluids during the surgery</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Blades</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">10011-00</td>
			<td>To perform the skin incision</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Buprenorphine</td>
			<td style="width:145px;">Buprelieve</td>
			<td style="width:161px;"></td>
			<td>Analgesia drug</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Catutery</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">18010-00</td>
			<td style="width:393px;">To prevent exsanguination</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Catutery tips</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">18010-01</td>
			<td style="width:393px;">To prevent exsanguination</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">cotton swab</td>
			<td style="width:145px;">Johnson&#39;s</td>
			<td style="width:161px;"></td>
			<td style="width:393px;">To absorb fluids during the surgery</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Depilatory cream</td>
			<td style="width:145px;">Veet</td>
			<td style="width:161px;"></td>
			<td>To delipate the animal</td>
		</tr>
		<tr height="17">
			<td height="17" style="height:18px;">Disposable operating room table cover</td>
			<td style="width:145px;">MEDKINE</td>
			<td style="width:161px;">DYND4030SB</td>
			<td>To cover the surgical area</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Echo probe</td>
			<td style="width:145px;">Siemens</td>
			<td>Sequoia 15L8W</td>
			<td>Ultrasound signal aquisition</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Echocardiograph</td>
			<td style="width:145px;">Siemens</td>
			<td>Acuson Sequoia C512</td>
			<td>Ultrasound signal aquisition</td>
		</tr>
		<tr height="25">
			<td height="25" style="height:25px;width:265px;">End-tidal CO2 monitor</td>
			<td style="width:145px;">Kent Scientific</td>
			<td style="width:161px;">CapnoStat</td>
			<td>To control expiration gas saturation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Forcep/Tweezers</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">11255-20</td>
			<td>To dissect the tissues and aorta</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Forcep/Tweezers</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">11272-30</td>
			<td>To dissect the tissues and aorta</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Forcep/Tweezers</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">11151-10</td>
			<td>To dissect the tissues and aorta</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Forcep/Tweezers</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">11152-10</td>
			<td>To dissect the tissues and aorta</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Gas system</td>
			<td>Penlon Sigma Delta</td>
			<td style="width:161px;"></td>
			<td>To anesthesia and mechanical ventilation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Hemostats</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">13010-12</td>
			<td>To hold the suture before tight the aorta</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Hemostats</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">13011-12</td>
			<td>To hold the suture before tight the aorta</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Ligation aids</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">18062-12</td>
			<td>To place a suture around the aorta</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Magnetic retractor</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">18200-20</td>
			<td>To help keep the animal in a proper position</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Needle holder</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">12503-15</td>
			<td>To suture the animal</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Needle 26G</td>
			<td style="width:145px;">B-BRAUN</td>
			<td style="width:161px;">4665457</td>
			<td>To serve as a molde of aortic constriction diameter</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Oxygen</td>
			<td style="width:145px;">Air Liquide</td>
			<td style="width:161px;"></td>
			<td>To anesthesia and mechanical ventilation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Polipropilene suture</td>
			<td style="width:145px;">Vycril</td>
			<td style="width:161px;">W8304/W8597</td>
			<td>To suture the animal and to do the constriction</td>
		</tr>
		<tr height="24">
			<td height="24" style="height:24px;">Povidone-iodine solution</td>
			<td style="width:145px;">Betadine&#174;</td>
			<td style="width:161px;"></td>
			<td>Skin antiseptic</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">PowerLab</td>
			<td>Millar instruments</td>
			<td>ML880 PowerLab 16/30</td>
			<td>PV loop Signal Aquisition</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Pulse oximeter</td>
			<td style="width:145px;">Kent Scientific</td>
			<td style="width:161px;">MouseStat</td>
			<td>To control heart rate and blood saturation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">PVAN software</td>
			<td style="width:145px;">Millar Instruments</td>
			<td style="width:161px;"></td>
			<td>To analyse the haemodynamic data</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">PV loop cathether</td>
			<td>Millar instruments</td>
			<td>SPR-1035. 1.4 F</td>
			<td>PV loop Signal Aquisition</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Retractor</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">17000-01</td>
			<td>To provide a better overview of the aorta</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Scalpet handle</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">10003-12</td>
			<td>To perform the skin incision</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Scissors</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">15070-08</td>
			<td>To cut the suture in debanding surgery</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Scissors</td>
			<td style="width:145px;">F.S.T</td>
			<td style="width:161px;">14084-09</td>
			<td>To cut other material during the surgery e.g. suture, papper</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Sevoflurane</td>
			<td style="width:145px;">Baxter</td>
			<td style="width:161px;">533-CA2L9117</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Temperature control module</td>
			<td style="width:145px;">Kent Scientific</td>
			<td style="width:161px;">RightTemp</td>
			<td>To control animal corporal temperature</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:265px;">Ventilator</td>
			<td style="width:145px;">Kent Scientific</td>
			<td style="width:161px;">PhysioSuite</td>
			<td>To ventilate the animal</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Water-bath</td>
			<td style="width:145px;">Thermo Scientific&#8482;</td>
			<td style="width:161px;">TSGP02</td>
			<td>To maintain water temperature adequate to heat the P-V loop catethers</td>
		</tr>
	</tbody>
</table>

studying left ventricular reverse remodeling by aortic debanding in rodents

To better understand the left ventricular (LV) reverse remodeling (RR), we describe a rodent model wherein, after aortic banding-induced LV remodeling, mice undergo RR upon removal of the aortic constriction. In this paper, we describe a step-by-step procedure to perform a minimally invasive surgical aortic debanding in mice. Echocardiography was subsequently used to assess the degree of cardiac hypertrophy and dysfunction during LV remodeling and RR and to determine the best timing for aortic debanding. At the end of the protocol, terminal hemodynamic evaluation of the cardiac function was conducted, and samples were collected for histological studies. We showed that debanding is associated with surgical survival rates of 70-80%. Moreover, two weeks after debanding, the significant reduction of ventricular afterload triggers the regression of ventricular hypertrophy (~20%) and fibrosis (~26%), recovery of diastolic dysfunction as assessed by the normalization of left ventricular filling and end-diastolic pressures (E/e&#39;&#160;and LVEDP). Aortic debanding is a useful experimental model to study LV RR in rodents. The extent of myocardial recovery is variable between subjects, therefore, mimicking the diversity of RR that occurs in the clinical context, such as aortic valve replacement. We conclude that the aortic banding/debanding model represents a valuable tool to unravel novel insights into the mechanisms of RR, namely the regression of cardiac hypertrophy and the recovery of diastolic dysfunction.

Post-operative and late survival 
The perioperative survival of the banding procedure is 80% and the mortality during the first month is typically &#60;20%. As previously mentioned, the success of the debanding surgery is highly dependent on how invasive the previous surgery was. After a learning curve, the mortality rate during the debanding procedures is around 25%. For this mortality accounts mostly deaths during the surgery procedure, including aorta or left atrium rupture (in rats, the survival...

Watch this Scientific Journal Video about Studying Left Ventricular Reverse Remodeling by Aortic Debanding in Rodents at JoVE.com

Studying Left Ventricular Reverse Remodeling by Aortic Debanding in Rodents

Here we describe a step-by-step protocol of surgical aorta debanding in the well-established mice model of aortic-constriction. This procedure not only allows studying the mechanisms underlying the left ventricular reverse remodeling and regression of hypertrophy but also to test novel therapeutic options that might accelerate myocardial recovery.

To better understand the left ventricular (LV) reverse remodeling (RR), we describe a rodent model wherein, after aortic banding-induced LV remodeling, ...

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