Name: optimization of the cuff technique for murine heart transplantation
Uploaded: 2020-06-26T13:00:00
Description: Watch this Scientific Journal Video about Optimization of the Cuff Technique for Murine Heart Transplantation at JoVE.com

This work was supported by the Fujian Provincial Health Education Joint Research Project (WKJ2016-2-20), the National Natural Science Foundation of China (81771271 and 81800664), the National Key R&#38;D Program of China (2018YFA0108304) and the Education and Scientific Research Project for Young and Middle-aged Teachers in Fujian Province (JAT170714), Natural Science Foundation of Hunan Province of China (2019JJ50842) and Huxiang Young Talents of Hunan Province (2019RS2013).

Animals (BALB/c, C57BL/6, male, 8-12 weeks) are housed in a specific pathogen-free facility at the Xiamen University Laboratory Animal Center. C57BL/6 is used as recipient and BALB/c is used as donor. All the procedures are performed according to the Institutional Animal Care and Use Committee (IACUC) guidelines.
NOTE: A set of microsurgical instruments, including a micro scissor, micro straight forceps, micro curved forceps and micro needle holders, are necessary for the operation (Ta...

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Mouse heart transplants models are important tools for transplant immunology research, as tools and materials for evaluating the immune mechanisms of this model and a large number of gene-modified mice are available. However, microsurgical technical challenges, such as vessels suture and eversion, have limited its widespread use. In the present study, we have investigated certain core technical challenges of murine heart transplantation and have obtained good outcomes. A critical step of the protocol the insertion of an ...

The establishment of mouse heterotopic heart transplantation through end-to-end anastomosis within the abdomen in 1973 was a major milestone in basic transplant immunology research1. This model provided an important and valid tool for analyzing the mechanisms of ischemia reperfusion injury2, immunological rejection, and tolerance3,4. However, the complex and time-consuming nature of the surgery as well as the potential for infections can result in severe perioperative abdominal adhesions and inflammatory reactions, resulting in a low efficiency for heterotopic heart transplantation model.
The cervical heterotopic heart transplantation technique was first described by Chen in 19915. In this model, the recipient&#8217;s external jugular vein is anastomosed to the pulmonary artery of the graft and the carotid artery is anastomosed to the ascending aorta. The main advantages of this method are the convenience of monitoring and the reducing of trauma to the recipient. In the same year, Matsuura described an improved technique, in which the end of the external jugular vein and carotid artery were everted over a Teflon cuff and fixed with a circumferential silk ligature6. Some researchers also fixed the cuff to the right pulmonary artery in the donor heart before inserting the cuff into the external jugular vein of the recipient7. Thus far, the cuff technique has been widely applied in various vascular pedicle transplant models, including those for lung8, liver9, and renal10 transplantation.
To date, there are several difficulties associated with the cuff technique. For example, the carotid artery is difficult to evert over the cuff due to the additional elasticity, resulting in the tissue flipping backwards. Hence, additional practice and a microsurgical dilator may be required to complete this step. In addition, the cervical vessel preparation can take up to 25 minutes.
To resolve these issues, we introduce the inner tube technique, which is based on the cuff technique and includes fixing the cuff on the external jugular vein and carotid artery using an inner tube to helps with the eversion of the vessel wall. In addition, with simple training, the recipient preparation is reduced to 15.5 minutes. This technique reduces the complexity of the operation and does not require additional practice or the use of a vascular dilator. It can be applied in all transplantation immune research, especially for verifying third-party immune tolerance during which the recipient receives two cardiac allografts, one within the abdomen and the other in the neck11. We also recommend co-operation between two skilled surgeons to establish this model, with one surgeon preparing the recipient animal and the other harvesting and implanting the donor heart. Such collaboration can shorten the operation time to 25 minutes. Using this optimized procedure, we have established syngeneic, allogeneic12,13,14,15,16,17,18,19, and xenogeneic mouse heart transplantation models20.
The rationale for the development of the inner tube technique was to reduce the operation time for the establishment of a mouse heart transplant model with a high success rate. Optimization of the cervical heart transplantation model facilitates the acquisition of high success rates in a short period of surgery time compared to the traditional suture and cuff technique21. Further, the cooperation model can further reduce the warm ischemic time of the donor heart compared to surgeries performed with a single operator.

<table>
	<tbody>
		<tr>
			<td>Artery cuff</td>
			<td>Self-made</td>
			<td></td>
			<td>Polyamide tube. diameter: 0.55 mm,length: 1.0 mm</td>
		</tr>
		<tr>
			<td>Artery inner tube</td>
			<td>Self-made</td>
			<td></td>
			<td>Polyamide tube. Diameter: 0.28mm</td>
		</tr>
		<tr>
			<td>Micro curved forceps</td>
			<td>Shanghai Medical Instruments (Group) Ltd., Corp. Surgical Instruments Factory</td>
			<td>WA3050</td>
			<td>1/8 arc, 0.3-mm tip without a hook</td>
		</tr>
		<tr>
			<td>Micro needle holders</td>
			<td>Shanghai Medical Instruments (Group) Ltd., Corp. Surgical Instruments Factory</td>
			<td>WA2050</td>
			<td>0.2-mm tip</td>
		</tr>
		<tr>
			<td>Micro scissors</td>
			<td>Shanghai Medical Instruments (Group) Ltd., Corp. Surgical Instruments Factory</td>
			<td>WA1050</td>
			<td>Straight, blade length: 10 mm</td>
		</tr>
		<tr>
			<td>Micro straight forceps</td>
			<td>Shanghai Medical Instruments (Group) Ltd., Corp. Surgical Instruments Factory</td>
			<td>WA3060</td>
			<td>0.15-mm tip without a hook</td>
		</tr>
		<tr>
			<td>Scanlan Vascu-Statt Bulldog Clamps</td>
			<td>Scanlan International Inc</td>
			<td>1001-531</td>
			<td>Clamping pressure 20&#8211;25 grams</td>
		</tr>
		<tr>
			<td>Vein cuff</td>
			<td>Self-made</td>
			<td></td>
			<td>Polyamide tube. diameter: 0.9 mm,length: 1.2 mm</td>
		</tr>
		<tr>
			<td>Vein inner tube</td>
			<td>Self-made</td>
			<td></td>
			<td>Polyamide tube. Diameter: 0.6 mm</td>
		</tr>
	</tbody>
</table>

optimization of the cuff technique for murine heart transplantation

Murine cardiac transplantation has been performed for more than 40 years. With advancements in microsurgery, certain new techniques have been used to improve surgical efficiency. In our lab, we have optimized the cuff technique with two major steps. First, we used the inner tube technique to insert a temporary inner tube into the external jugular vein and carotid artery blood vessel to facilitate eversion of the vessel over the cuff. Second, we performed complete heterotopic cardiac transplantation through the collaboration of two experienced surgeons. These modifications effectively reduced the operation time to 25 minutes, with a success rate of 95%. In this report, we describe these procedures in detail and provide a supplemental video. We believe that this report on the improved cuff technique will offer practical guidance for murine heterotopic heart transplantation and will enhance the utility of this mouse model for basic research.

Surgical Operation Time
After training, a skilled surgeon can successfully perform the operation within 35 minutes using the inner tube technique, wherein approximately 15.5 minutes are required for recipient preparation, 10.9 minutes are required for donor preparation, and 4.4 minutes are required for donor heart anastomoses. The cold and warm ischemia time (from donor preparation to heart implantation) is reduced to 15.3 minutes compared to the operation using the cuff techn...

Watch this Scientific Journal Video about Optimization of the Cuff Technique for Murine Heart Transplantation at JoVE.com

Optimization of the Cuff Technique for Murine Heart Transplantation

We introduce an inner tube approach to the cuff technique for mouse cervical heterotopic heart transplantation to help evert the vessel over the cuff. We found that cooperation between two experienced surgeons markedly shortens the operation time.

Murine cardiac transplantation has been performed for more than 40 years. With advancements in microsurgery, certain new techniques have been used to ...

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