Name: a modified technique for transverse aortic constriction in mice
Uploaded: 2022-08-18T15:00:00
Description: Watch this Scientific Journal Video about A Modified Technique for Transverse Aortic Constriction in Mice at JoVE.com

This work is funded by the National Natural Science Foundation of China (NSFC 81822002). We thank all the members who took part in this work.

The current protocol is approved by the ethics committee of Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China. This procedure is performed on male adult C57/BL6 mice (&#62;10 weeks of age). All surgical instruments were sterilized by autoclaving before the operation.
1. Preparation of surgical instrument
<ol>
	<li>Prepare a 5 mL syringe and pinch off the tip of the needle with a needle holder to blunt it.&#160;</li>
	<li>Pre...

<ol>
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(121), e55293 (2017).</li><li>Lao, Y., 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=Operating+transverse+aortic+constriction+with+absorbable+suture+to+obtain+transient+myocardial+hypertrophy.">Operating transverse aortic constriction with absorbable suture to obtain transient myocardial hypertrophy.</a> Journal of Visualized Experiments. (163), e61686 (2020).</li><li>Veldhuizen, R. A., Slutsky, A. S., Joseph, M., McCaig, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+mechanical+ventilation+of+isolated+mouse+lungs+on+surfactant+and+inflammatory+cytokines.">Effects of mechanical ventilation of isolated mouse lungs on surfactant and inflammatory cytokines.</a> European Respiratory Journal. 17 (3), 488-494 (2001).</li><li>Withaar, C., Lam, C. S. P., Schiattarella, G. G., de Boer, R. 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The induction of sustained pressure overload can gradually cause cardiac hypertrophy and heart failure. This model has been used in numerous laboratories around the world14,15,16. The protocol provided an improved TAC method that does not need microsurgical skills or mechanical ventilation.
The most important step in this protocol is passing silk suture under the aortic arch. When the snare has hooked...

Heart failure is a complex clinical symptom that results from impaired structure and function of ventricular filling or ejecting blood1. The disease stage is mainly defined via the New York Heart Association function classification based on the severity of symptoms and physical activity2. For those patients with an ejection fraction of over 50%, structural and/or functional abnormalities raised natriuretic peptides to support the diagnosis of heart failure with preserved ejection fraction (HFpEF)2. Ischemic heart disease is a leading cause among multiple etiologies of heart failure. Thus, the myocardial infarction model (such as permanent coronary ligation) is often used to study pathophysiology after cardiac hypoperfusion or ischemia-reperfusion injury3,4. Besides acute myocardial injury, other risk factors such as hypertension, diabetes, obesity, and a family history of cardiomyopathy also contribute to the development of heart failure. After patients pass Stage A (at risk for heart failure) and enter Stage B (pre-heart failure), structural modification occurs1. For example, hypertensive patients first go through adaptive left ventricle hypertrophy, and then gradually develop into maladaptive cardiac hypertrophy and transit to heart failure through pathological remodeling5.
As the terminal stage of various cardiovascular diseases, chronic heart failure has been studied for decades6. Multiple mouse models have been widely used in heart failure research, including drug infusion (angiotensin II), metabolic disorders (diabetes or high caloric diet), and aortic constriction7. Among these models, angiotensin II perfusion is accompanied by various organ side effects, such as kidney7. Inducing metabolic disorders usually require a rather long period of time. Ascending aortic constriction has been considered to have limited relevance to human disease7.
TAC is a reliable model that increases afterload and induces cardiac hypertrophy as well as heart failure8. Open-chest TAC model was first described by Rockman et al. and was used in numerous laboratories around the world9. However, this classical TAC procedure causes a rather large trauma to mice and changes their normal behavior, which may take a long recovery time and disturb further treatment10. Other modified closed-chest TAC procedures did reduce some invasive steps but required microsurgical skills or mechanical ventilation10,11.
The present protocol details a step-by-step method with a minimally invasive approach to the aortic arch using a self-made retractor via a 3 mm midline incision of the upper edge of the sternum. This model does not need microsurgical skill, mechanical ventilation, or cutting through the ribs, thereby providing a rapid, surgical trauma-limited, uncomplicated, inexpensive way to perform TAC surgery.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>4-0 nonabsorbable suture</td>
			<td>Jinhuan</td>
			<td>HM403</td>
			<td>Used for suturing the skin</td>
		</tr>
		<tr>
			<td>5 mL syringe</td>
			<td>Haifuda Technology Co., Ltd.</td>
			<td>BD-309628</td>
			<td>Used for making snare containing retractor</td>
		</tr>
		<tr>
			<td>7-0 nonabsorbable suture</td>
			<td>Jinhuan</td>
			<td>HM701</td>
			<td>Used for aorta ligation</td>
		</tr>
		<tr>
			<td>Animal temperature monitor</td>
			<td>Kaerwen</td>
			<td>FT3400</td>
			<td>Used for monitoring body temperature</td>
		</tr>
		<tr>
			<td>Buprenorphine&#160;</td>
			<td>Sigma</td>
			<td>B-044</td>
			<td>Used for post-surgical pain treatment</td>
		</tr>
		<tr>
			<td>Depilatory cream&#160;</td>
			<td>Veet</td>
			<td>N/A</td>
			<td>Used for remove body hair from the surgical area</td>
		</tr>
		<tr>
			<td>Heating Pad</td>
			<td>Xiaochuangxin</td>
			<td>N/A</td>
			<td>Used for maintaining body temperature</td>
		</tr>
		<tr>
			<td>Ibuprofen</td>
			<td>MCE</td>
			<td>HY-78131</td>
			<td>Used for post-surgical pain treatment</td>
		</tr>
		<tr>
			<td>Iron wire (0.5 mm)</td>
			<td>Qing Yuan</td>
			<td>Iron wire #26</td>
			<td>Used for making snare containing retractor</td>
		</tr>
		<tr>
			<td>Microscopic tweezers</td>
			<td>RWD</td>
			<td>F12006-10</td>
			<td>Used for penetrating and separating the tissue to open operation space</td>
		</tr>
		<tr>
			<td>Needle holder</td>
			<td>RWD</td>
			<td>F12005-10</td>
			<td>Used for pinching off the tip of gauge needle and blunting it</td>
		</tr>
		<tr>
			<td>Ophthalmic forceps</td>
			<td>RWD</td>
			<td>F14012-10&#160;</td>
			<td>Used for holding skin and other tissues</td>
		</tr>
		<tr>
			<td>Ophthalmic scissors</td>
			<td>RWD</td>
			<td>S11001-08</td>
			<td>Used for making sking incision of mouse</td>
		</tr>
		<tr>
			<td>Pentobarbital sodium</td>
			<td>Sigma</td>
			<td>P3761</td>
			<td>Used for mouse anesthesia</td>
		</tr>
		<tr>
			<td>Sterile operating mat</td>
			<td>Hale &#38; hearty</td>
			<td>211002</td>
			<td>Used for placing animal during surgery</td>
		</tr>
		<tr>
			<td>Ultra-sound imaging system</td>
			<td>Fujifilm visualsonics</td>
			<td>vevo1100</td>
			<td>Used for measure the blood flow velocity, left ventricular wall thickness and ejection fraction, https://www.visualsonics.com/product/imaging-systems/vevo-1100</td>
		</tr>
	</tbody>
</table>

a modified technique for transverse aortic constriction in mice

Transverse aortic constriction (TAC) is a frequently used surgery in research regarding heart failure and cardiac hypertrophy based on the formation of pressure overload in mouse models. The main challenge of this procedure is to clearly visualize the transverse aortic arch and precisely band the target vessel. Classical approaches perform a partial thoracotomy to expose the transverse aortic arch. However, it is an open-chest model that causes a rather large surgical trauma and requires a ventilator during the surgery. To prevent unnecessary trauma and simplify the operating procedure, the aortic arch is approached via the proximal proportion of the sternum, reaching and binding the target vessel using a small self-made retractor that contains a snare. This procedure can be conducted without entering the pleura cavity and does not need a ventilator or microsurgical operation, which leaves the mice with physiological breathing patterns, simplifies the procedure, and significantly reduces operation time. Due to the less invasive approach and less operation time, mice can undergo fewer stress reactions and recover rapidly.

After successful TAC surgery, pressure overload was detected using an ultrasound imaging system. Four weeks after surgery, mice develop decreased heart function. In the present study, the efficacy of TAC surgery was validated via ejection fraction (EF), fractional shortening (FS), left ventricular mass (LV mass) and left ventricular internal diameter (LVID) of mice who underwent TAC surgery after 4 weeks. EF was significantly reduced in TAC mice after 4 weeks compared to sham mice (47% &#177; 10% vs. 78% &#177; ...

Watch this Scientific Journal Video about A Modified Technique for Transverse Aortic Constriction in Mice at JoVE.com

A Modified Technique for Transverse Aortic Constriction in Mice

The present protocol describes a modified and simplified technique with a minimally invasive transverse aortic constriction (TAC) procedure using a self-made retractor. This procedure can be conducted without a ventilator or microscope and introduces pressure overload, eventually leading to cardiac hypertrophy or heart failure.

Transverse aortic constriction (TAC) is a frequently used surgery in research regarding heart failure and cardiac hypertrophy based on the formation of ...

This protocol provides a rapid surgical trauma limited, uncomplicated, and inexpensive masses to perform TAC surgery. This model approaches the aortic arch using a self-made retractor and does not need microsurgical scale or mechanical ventilation, thereby significant limiting surgical trauma. To begin, prepare a five milliliter syringe.Then make the needle blunt by pinching off its tip with a needle holder. Prepare a 27 gauge needle blunted with a needle holder and bend its tip at 90 degrees. Use a commercially obtained iron wire and twine it around the needle.Form a small snare or loop at the blunted tip of the needle. Sterilize all the surgical tools before surgery by autoclaving. Confirm the depth of anesthesia with a toe pinch to prevent dryness while under anesthesia.Apply ophthalmic ointment to both eyes. Then place the mouse in the supine position on a heating pad covered with a sterile drape with its head pointing to the operator. Fix its extremities with adhesive tape and incisors with a 4-0 suture.Now, insert a rectal probe to monitor body temperature during the operation. Using sterilized ophthalmic scissors, cut a hole in a new, disposable sterile drape. Place the sterile drape over the mouse and ensure the un-haired operating field is visible through the hole.After making an incision separate the skin and the superficial fascia and expose the trachea and proximal sternal. Then carefully make a three to four millimeter longitudinal midline incision to the sternum from the upper edge to the sternal angle and gently lift the sternum with forceps. Using straight forceps bluntly dissect the target vessel and tissues around it.Now, hold the previously self-made retractor with a wire snare in the right hand. Carefully pass the snare of the retractor under the aorta between the left common carotid artery and the origin of the right denominate artery. Pass the retractor to the left hand.Hold the microscopic tweezers in the right hand and pass a 7-0 silk suture through the snare loop at the end. Pull out the retractor. Place a 27 gauge needle parallel to the aorta and tie the suture around the needle and the aorta.Then hold the 27 gauge needle with ophthalmic forceps and the knot tied to the 27 gauge needle and aorta with another forceps. Quickly but gently pull the 27 gauge needle from the ligation. Suture the skin with 4-0 synthetic monofilament suture in an interrupted suture pattern.Place the mouse on a warm pad and wait for it to revive. Generally, the mouse revives in about 60 minutes. After four weeks of surgery.Ejection fraction and fractional shortening were significantly reduced in transverse aortic constriction mice compared to sham mice. However, left ventricular mass and left ventricular internal diameter were significantly elevated in the transverse aortic constriction mice. The most important step in this particle is passing suture under the aortic arch.All movies must be gentle to reduce unnecessary traction to the artery. Following this procedure researchers can apply medical treatment to the animal model in order to investigate potential therapeutic targets to patients with cardio hypertrophy or heart failure.

Research

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