Name: a modified technique for arteriovenous fistula construction in rabbits
Uploaded: 2023-02-10T13:33:00.000+00:00
Duration: 5 min
Description: Watch this Scientific Journal Video about A Modified Technique for Arteriovenous Fistula Construction in Rabbits at JoVE.com

This study was supported by grants from the Suzhou Science and Technology Plan Project (SYS2020077), Suzhou High-tech Zone Medical and Health Science and Technology Plan Project (2020z001), Suzhou Science and Technology Development Plan Project-Medical and Health Science and Technology Innovation (SYK2021030), Nanjing Medical University Science and Technology Development Fund-General Project (NMUB20210253), Suzhou Science and Technology Bureau of the application of the basic research project (No.SYSD2019205, No.SYS2020119), Jiangsu Province Traditional Chinese Medicine Science and Technology Development Plan Project (No.MS2021098), the Ministry of Education Industry-University Cooperation Collaborative Education Project (No. 202102242003),&#160;the Sixth &#34;333 High-level Talent Cultivation&#34; Project in Jiangsu Province, Suzhou Science and Technology City Hospital 2022 Hospital-level Pre-research Fund Project (SZKJCYY2022014), and Suzhou &#34;KeJiaoXingWei&#34; Youth Science and Technology Project (KJXW2022086).

The experimental procedures using laboratory animals were approved by the Experimental Animal Welfare Ethics Committee of Nanjing Medical University. New Zealand rabbits aged 10 months (of both sexes; body weight, 3.18 &#177; 0.24 kg) were used for this study. The animals were obtained from a commercial source (see Table of Materials).
1. Animal preparation
<ol>
	<li>Anesthetize the rabbits using a mixed intravenous injection of tiletamine hydrochloride and zolazepam hydrochloride (3 mg/kg) into the marginal ear vein and an intramuscular injection of sumianxin II (0.02 mL/kg) (see Table of Materials) into the hindlimb muscle. 
	NOTE: After about 1-3 min, the anesthetic effect stabilizes. Before proceeding, the level of anesthesia should be checked by pinching the skin behind the neck and observing the corneal reflex. Tiletamine hydrochloride, zolazepam hydrochloride (0.5 mg/kg), and sumianxin II (0.01 mL/kg) could be added during the operation if needed.</li>
	<li>Lay the rabbit on a fixed table (see Table of Materials) in a supine position, and tie the limbs and incisors with binds.</li>
	<li>Shave the neck and upper chest using an electric razor, and remove the hair by animal depilatory cream (see Table of Materials).</li>
	<li>Maintain sterile conditions during the surgery by autoclaving the surgical equipment and cleaning the surgery area with povidone-iodine solution.</li>
</ol>
2. Skin incision
<ol>
	<li>Position the rabbit with its head toward the surgeon.</li>
	<li>Make a longitudinal incision of ~3 cm between the mandible and the sternoclavicular joint using surgical scissors or a&#160;scalpel blade.</li>
</ol>
3. Preparation of the external jugular vein (EJV)
<ol>
	<li>Expose the incision, and identify the right EJV. Ensure that the EJV and its perivascular tissues are clearly visible and not dissected. 
	NOTE: The EJV shows an inverted &#34;Y&#34; pattern, and the branch close to the medial neck needs to be anastomosed.</li>
	<li>Make a tunnel that a vascular clamp can be passed through (see Table of Materials) along the direction perpendicular to the EJV. Ensure that the distances between the openings on both sides of the tunnel and the EJV are &#62;1 cm.</li>
	<li>Place a vascular clamp along the tunnel.</li>
	<li>Make another tunnel (same as in step 3.2) at the distal EJV using the same method. 
	NOTE: Ensure the distance between the two tunnels is &#8805;2 cm.</li>
	<li>Apply a&#160;4-0 suture (see Table of Materials) and a vascular clamp along the tunnel to control the blood flow (Figure 1A).</li>
</ol>
4. Dissecting and preparing the common carotid artery (CCA)
<ol>
	<li>Use forceps (see Table of Materials) to explore the CCA lateral to the trachea and medial to the sternocleidomastoid muscle. 
	NOTE: The CCA has a pulsatile sensation and runs parallel to the cervical nerve.</li>
	<li>Bluntly dissect the CCA to a length of about 2 cm. 
	NOTE: Avoid injury to the vagus nerve and its branches with a deep arterial course.</li>
	<li>Place a 4-0 suture thread around the CCA to control the blood flow when needed.</li>
	<li>Apply vascular clamps (see Table of Materials) as distally and proximally as possible (Figure 1B).</li>
</ol>
5. Preparing the anastomosis
<ol>
	<li>For phlebotomy and anastomosis, use microscissors (see Table of Materials) to dissect the internal part of the EJV (4 mm long) free from the surrounding tissues.</li>
	<li>Make a 4 mm long longitudinal incision with microscissors in the middle of the vein. Rinse the vein with heparin solution (100 IU/mL) to prevent thrombosis.</li>
	<li>Make a longitudinal incision measuring approximately 4 mm in the anterior wall of the artery using a sharp blade and microscissors. Rinse the artery with 100 IU/mL heparin solution until the vessel is clear of blood.</li>
</ol>
6. Side-to-side anastomosis
<ol>
	<li>Pull the EJV and CCA closely together, as close as possible.</li>
	<li>Apply Kunlin&#39;s technique12 for the side-to-side anastomosis of the CCA and EJV using 8-0 nonabsorbable sutures (see Table of Materials). Suture the posterior wall of the vessel first (Figure 1C), followed by the anterior wall of the vessel. 
	NOTE: Since the EJV wall in a rabbit is thin, care should be taken during the surgery to prevent damage to the vessels that could later compromise the patency of the anastomosis. During the process of vascular anastomosis, heparin solution (100 IU/mL) must be used repeatedly to rinse the lumen to prevent thrombosis.</li>
</ol>
7. Vascular clamp removal and ligation of the vein
<ol>
	<li>Remove the distal vascular clamp of the CCA, the proximal vascular clamp of the EJV, and the proximal vascular clamp of the CCA in turn. Observe active blood flow through the anastomosis.</li>
	<li>Ligate the distal end of the EJV using the 4-0 suture that was placed earlier. Remove the distal vascular clamp of the EJV.</li>
	<li>Remove the suture thread that was placed around the CCA (Figure 1D).</li>
</ol>
8. Skin closure and postoperative care
<ol>
	<li>After ensuring no significant bleeding in the surgical field, close the skin of the neck using interrupted sutures (4-0).</li>
	<li>Place the rabbit in a cage until it completely recovers. Typically, this takes 30-45 min. 
	NOTE: In case of incomplete or delayed recovery, care should be taken to ensure that the rabbit does not experience hemodynamic shock due to bleeding in the surgical area. If required, administer Sumianxin II (0.01 mL/kg) post-surgery.</li>
</ol>

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The authors have no potential conflicts of interest related to the drugs and materials used in this procedure.

Currently, several animal models are available for AVF. Among them, pigs, sheep, and dogs are mostly used as large animal models13,14,15. The small animal models used include rabbits, rats, and mice16,17,18. New Zealand rabbits were used in this study. New Zealand rabbits have abundant perivenous tissues around the EJV, which makes them conducive for the study of the MNTT method. The advantages of using New Zealand rabbits include the simple surgical operation, convenient feeding, and low cost. However, large animal models have benefits over rabbit models when studying hemodynamics.
This study proposed a unique CCA-EJV AVF procedure using MNTT without the dissection of the perivenous tissue or cutting off the artery. Functional end-to-side anastomosis19,20 for AVF creation was accomplished by side-to-side arteriovenous anastomosis followed by the ligation of the distal EJV. Compared to conventional techniques (Supplementary Figure 3), AVF creation with MNTT more adequately preserved the perivenous tissue. During arteriovenous vascular anastomosis, due to the preservation of the perivenous tissue, the venous wall could be more fully exposed by pulling the perivenous tissue, which was conducive to vascular anastomosis.
In ultrasonography, spiral laminar flow was observed in the vein and artery near the anastomosis, indicating that the MNTT may have more favorable hemodynamics, which may account for the excellent rates of patency and maturation21,22. On histological observation, significant venous intimal hyperplasia was observed at the AVF anastomosis, whereas no significant intimal hyperplasia was present at the proximal EJV of the anastomosis. This finding is probably related to the improvement of the juxta-anastomotic stenosis by this surgical technique or the spiral laminar flow.
Common problems encountered and suggestions 
Given the thin wall of the EJV, gentle operation is necessary when anastomosing blood vessels to prevent damage to the EJV. Since the tissue around the EJV is preserved, it can be pulled during anastomosis to unfold the vessel and make it more conducive for suturing. However, the preservation of the tissue surrounding the EJV is bothersome. After venotomy, given the blood outflow from the venous vessels, the blood vessels collapse and cause EJV retraction. During EJV anastomosis, microvascular tweezers must be used to pull the surrounding tissues of the EJV and fully expose the vein wall. Additionally, if the distance between arteries and veins is long, the CCA should be allowed a free sufficient length to ensure that the two are close to each other and, thus, to facilitate the anastomosis. An 8-0 sterile vascular suture was used for the vascular anastomosis to reduce damage to the vessels.
Technical limitations 
The preparation of the vein still requires tunneling and clamping along the tunnel, and this maneuver can cause venous injury. Prior to performing an arterial and venous side-to-side anastomosis, vascular injury may result from pulling the artery and vein. Since the patency rate of 38.2% was lower than that of other AVF models23,24, further improving the care and detection of rabbits after AVF surgery is necessary.
Applications of the technique 
To further study the mechanisms of the MNTT and related hemodynamics, pathological, molecular, and genomic studies are needed to validate this technique.
Conclusion 
A CCA-EJV AVF was successfully created in this study using the MNTT method. The operation was simple, with good reproducibility and a high success rate, indicating that this technique has the potential to be ideal for further studies on the application of MNTT in AVF surgery.

The construction of an arteriovenous fistula (AVF) is widely used in clinical practice for patients undergoing maintenance hemodialysis (MHD), and it has higher patency and fewer complications than an arteriovenous graft (AVG) or tunneled cuffed catheter (TCC)1,2. Although AVF is the preferred mode of vascular access, it is not perfect and has inherent limitations. The 1 year primary AVF patency rates are only 60%-65%, with many failures occurring in the near anastomotic region3,4,5.
Vessels undergo different degrees of damage during the traditional surgical approach, which ultimately affects the maturation of the AVF. New surgical modalities, such as the no-touch technique (NTT)&#160;(Supplementary Figure 1) proposed by H&#246;rer et al.6 and radial artery excursion and reimplantation (RADAR) proposed by Sadaghianloo et al.7,8 and Bai et al.9, were designed to decrease the rate of juxta-anastomotic stenosis and to improve the fistula patency by modifying the surgical technique. Although the effect of RADAR was better than that of NTT, inflow arterial stenosis was observed to be more prominent with RADAR. To further reduce injury to the veins and arteries during the operation, in 2021, a new modified no-touch technique (MNTT) was proposed to create a radio-cephalic AVF by preserving the perivenous tissue around the cephalic vein without cutting the radial artery (Supplementary Figure 1 and Supplementary Figure 2). The preliminary results showed increased primary patency, decreased juxta-anastomotic stenosis, and no arterial stenosis10,11.
Considering the current lack of animal models of AVF using MNTT, and to further explore the mechanism of MNTT in AVF surgery, this study introduces a common carotid artery (CCA)-external jugular vein (EJV) AVF procedure using MNTT.

<table><tbody><tr><td>Animal Depilatory</td><td>Fuzhou Feijing Biotechnology Co., Ltd.</td><td>PH1877</td><td></td></tr><tr><td>Curved hemostatic forceps</td><td>Xinhua Surgical Instrument Co., Ltd.</td><td>ZH131R/RN</td><td></td></tr><tr><td>Dissecting forceps</td><td>Xinhua Surgical Instrument Co., Ltd.</td><td>ZDO25R/RN</td><td></td></tr><tr><td>electrical razor</td><td>Shenbao Technology Co., Ltd</td><td>PGC-660</td><td></td></tr><tr><td>Fixed Table</td><td>Zhenhua Biomedical Instrument Co., Ltd</td><td>ZH-DSB019</td><td></td></tr><tr><td>Halsey needleholder</td><td>Xinhua Surgical Instrument Co., Ltd.</td><td>ZM208R/RN</td><td></td></tr><tr><td>Heparin Dodium Injection</td><td>Jiangsu Wanbang Biochemical Pharmaceutical Group Co., Ltd.</td><td>H32020612</td><td></td></tr><tr><td>Medical gauze dressing</td><td>Nanchang Kangjie medical hygiene products Co., Ltd</td><td>20172640135</td><td></td></tr><tr><td>Micro forceops</td><td>Xinhua Surgical Instrument Co., Ltd.</td><td>ZD275RN/T</td><td></td></tr><tr><td>Micro needle holder forceps</td><td>Xinhua Surgical Instrument Co., Ltd.</td><td>ZF2618RB/T</td><td></td></tr><tr><td>Micro scissors</td><td>Xinhua Surgical Instrument Co., Ltd.</td><td>ZF022T</td><td></td></tr><tr><td>Non-silk sutures 4-0</td><td>Kollsut Medical Instrument Co., Ltd.</td><td>NMB020RRCN26C075-1</td><td></td></tr><tr><td>Non-absorbable sutures 8-0 (double needle)</td><td>Yangzhou Yuankang Medical Instrument Co., Ltd.</td><td>10299023602</td><td></td></tr><tr><td>Povidone iodine solution</td><td>Shanghai Likang Disinfection High-tech Co., Ltd.</td><td>310512</td><td></td></tr><tr><td>Rinse needle</td><td>Jiangsu Tonghui Medical Instrument Co., Ltd</td><td>20180039</td><td></td></tr><tr><td>scalpel handle</td><td>Shanghai Medical Instrument (Group) Co., Ltd. Surgical Instruments Factory</td><td>J11030</td><td></td></tr><tr><td>Sharp blade</td><td>Suzhou Medical Products Factory Co., Ltd.</td><td>TY21232001</td><td></td></tr><tr><td>Sodium Chloride Injection&amp;nbsp; (100 mL)</td><td>Guangdong Otsuka Pharmaceutical Co., Ltd.</td><td>B21K0904</td><td></td></tr><tr><td>Sugical Scissors</td><td>Xinhua Surgical Instrument Co., Ltd.</td><td>ZC120R/RN</td><td></td></tr><tr><td>Sumianxin II</td><td>Jilin Dunhua Shengda Animal Pharmaceutical Co., Ltd.</td><td>20180801</td><td></td></tr><tr><td>Syringe with needle?5 mL)</td><td>BD medical devices (Shanghai) Co., Ltd</td><td>2006116</td><td></td></tr><tr><td>Tiletamine Hydrochloride and Zolazepam Hydrochloride for Injection</td><td>Virbac Pet Health, France</td><td>83888204</td><td></td></tr><tr><td>Triangle needle</td><td>Hangzhou Huawei medical supplies Co., Ltd</td><td>7X17</td><td></td></tr><tr><td>Vascular clamp</td><td>Xinhua Surgical Instrument Co., Ltd.</td><td>ZF220RN</td><td></td></tr><tr><td>New Zealand rabbits</td><td>Suzhou Huqiao Biological Co., Ltd.</td><td>SCXK2020-0001</td><td></td></tr></tbody></table>

a modified technique for arteriovenous fistula construction in rabbits

Juxta-anastomotic stenosis is a challenging problem that often causes non-maturation and decreases the patency of an arteriovenous fistula (AVF). Injury to the veins and arteries during the operation and hemodynamic changes can lead to intimal hyperplasia, leading to juxta-anastomotic stenosis. To reduce injury to the veins and arteries during the operation, this study proposes a new modified no-touch technique (MNTT) for AVF construction that can decrease the rate of juxta-anastomotic stenosis and improve the AVF patency. To unravel the hemodynamic changes and mechanisms of the MNTT, this study presented an AVF procedure using this technique. Although this procedure is technically challenging, 94.4% procedural success was achieved after adequate training. Ultimately, 13 out of 34 rabbits had a functional AVF 4 weeks after the surgery, leading to a 38.2% AVF patency rate. However, at 4 weeks, the survival rate was 86.1%. Ultrasonography showed active blood flow through AVF anastomosis. Furthermore, the spiral laminar flow was observed in the vein and artery near the anastomosis, suggesting that this technique may improve the hemodynamics of the AVF. On histological observation, significant venous intimal hyperplasia was observed at the AVF anastomosis, whereas no significant intimal hyperplasia was observed at the proximal external jugular vein (EJV) of the anastomosis. This technique will improve the understanding of the mechanisms underlying the use of MNTT for AVF construction and provide technical support for the further optimization of the surgical approach in AVF construction.

The outcome of the successful application of this technique is a patent AVF in the rabbit neck. This study used the following criteria to evaluate the success: (1) when the vascular anastomosis is completed, the venous tremor of the AVF can be touched, and the vascular murmur can be heard; (2) 4 weeks after the AVF is established, the active blood flow through the internal fistula anastomosis can be measured by color Doppler ultrasound; (3) 4 weeks after the AVF is established, hematoxylin-eosin (H&#38;E) staining shows significant venous intimal hyperplasia at the AVF anastomosis.
In all, 36 healthy New Zealand rabbits were included in this study. In total, 34 rabbits had an immediately successful AVF using the MNTT. Three rabbits had significant postoperative bleeding, and one died because of blood loss. The remaining two rabbits required compression hemostasis to arrest the bleeding. Additionally, four rabbits died after surgery, with common symptoms including sneezing, coughing, runny nose, anorexia, and diarrhea. Finally, 31 rabbits survived, and 13 had a functional AVF 4 weeks after surgery. The survival rate was 86.1% (Figure 2).
The AVF was assessed using color Doppler ultrasonography (CDU) 4 weeks after surgery to confirm patency, defined as active blood flow through the AVF anastomosis (Figure 3). Moreover, the spiral laminar flow was observed at both the vein and artery near the anastomosis (Figure 3). In terms of the ultrasound parameters between the AVF and normal vessels on the contralateral neck, there were significant differences in the diameter and PSV of the EJV and the diameter of the CCA (Table 1).
The AVF was obtained at 4 weeks after surgery and made into sections. H&#38;E staining was performed on all the obtained sections. Significant venous intimal hyperplasia was observed at the site of the AVF anastomosis (Figure 4), whereas no significant intimal hyperplasia was observed at the proximal EJV of the anastomosis (Figure 4).
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/64352/64352fig01.jpg" /> 
Figure 1: The CCA-EJV AVF created in rabbits using the MNTT. (A) Two tunnels were made along the direction perpendicular to the EJV. (B) The CCA was mobilized. (C) Using Kunlin&#39;s technique, a side-to-side anastomosis of the CCA and EJV was performed. (D) The distal end of the EJV was ligated, and patent blood flow was visible through the proximal end. Abbreviations: CCA = common carotid artery; EJV = external jugular vein; AVF = arteriovenous fistula; MNTT = modified no-touch technique. <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/64352/64352fig01large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/64352/64352fig02.jpg" /> 
Figure 2: Survival curves for the rabbits. One rabbit died immediately after surgery because of blood loss. The remaining four rabbits died on the 3rd, 7th, 10th, and 26th days after surgery. Finally, 31 rabbits were alive at 4 weeks after surgery. <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/64352/64352fig02large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/64352/64352fig03.jpg" /> 
Figure 3: The CDU evaluation atlas of the AVF. (A) The CCA showed unidirectional low-resistance blood flow spectra, the loss of normal triphasic blood flow, widened systolic peaks, and abundant diastolic blood flow. (B) The EJV showed artery-like low-resistance blood flow spectra, with an increased PSV and widened spectra. (C) Active blood flow through the AVF anastomosis. (D) Spiral laminar flow was observed in the EJV outflow tracts. Abbreviations: AVF = arteriovenous fistula; CCA = common carotid artery; EJV = external jugular vein; PSV = peak systolic velocity. <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/64352/64352fig03large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/64352/64352fig04.jpg" /> 
Figure 4: Observation of the AVF morphology of rabbits at 4 weeks after surgery (H&#38;E staining). (A) No significant intimal hyperplasia of the CCA with a patent AVF was observed. (B) The elastic membrane of the EJV at the anastomosis site with a patent AVF was severely disrupted, with significant intimal hyperplasia. Thick hyperplastic fibrous tissues were clearly visible on the inner side of the elastic membranes, with reduced and fragmented elastic fibers. (C) The proximal EJV of the anastomosis had no significant intimal hyperplasia with a patent AVF. This showed intact elastic membranes and slender, wavy elastic fibers. Abbreviations: AVF = arteriovenous fistula; CCA = common carotid artery; EJV = external jugular vein. Magnification: 200x. <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/64352/64352fig04large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td rowspan="2">Group</td>
			<td colspan="2">External jugular vein</td>
			<td colspan="2">Common carotid artery</td>
		</tr>
		<tr>
			<td>Diameter (mm)</td>
			<td>PSV (cm/s)</td>
			<td>Diameter (mm)</td>
			<td>PSV (cm/s)</td>
		</tr>
		<tr>
			<td>AVF</td>
			<td>7.21 &#177; 1.55</td>
			<td>79.64 &#177; 39.31</td>
			<td>3.06 &#177; 0.32</td>
			<td>59.38 &#177; 32.25</td>
		</tr>
		<tr>
			<td>Normal vessel</td>
			<td>3.13 &#177; 0.66</td>
			<td>9.21 &#177; 2.77</td>
			<td>2.17 &#177; 0.41</td>
			<td>39.02 &#177; 11.56</td>
		</tr>
		<tr>
			<td>t</td>
			<td>5.413</td>
			<td>3.996</td>
			<td>3.779</td>
			<td>1.329</td>
		</tr>
		<tr>
			<td>P</td>
			<td>0.001</td>
			<td>0.004</td>
			<td>0.005</td>
			<td>0.22</td>
		</tr>
	</tbody>
</table>
Table 1: Comparison of ultrasound parameters between the AVF and normal vessels on the contralateral neck in rabbits (n = 5). Abbreviation: PSV = peak systolic velocity. t-test is used for data analysis. When the P value is &#60;0.05, the comparison between the two groups is statistically significant.
Supplementary Figure 1: Schematic diagram of the vascular anastomosis modes in AVF surgery. (A) Traditional AVF surgery. (B) An AVF created using the NTT. (C) An AVF created using the MNTT. Abbreviations: AVF = arteriovenous fistula; NTT = no-touch technique; MNTT = modified no-touch technique. <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/64352/Supplementary Figure 1.tif" target="_blank">Please click here to download this File.</a>
Supplementary Figure 2: Functional end-to-side anastomosis AVF with the MNTT. Abbreviations: AVF = arteriovenous fistula; MNTT = modified no-touch technique. <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/64352/Supplementary Figure 2.tif" target="_blank">Please click here to download this File.</a>
Supplementary Figure 3: Rabbit model of AVF created using the conventional technique. (A) The EJV was dissected from the perivenous tissue. (B) The EJV and CCA were pulled together. (C) Using Kunlin&#39;s technique, a side-to-side anastomosis of the CCA and EJV was conducted. (D) The distal end of the EJV was ligated, and patent blood flow visibly passed through the proximal end. Abbreviations: CCA = common carotid artery; EJV = external jugular vein; AVF = arteriovenous fistula. <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/64352/Supplementary Figure 3.tif" target="_blank">Please click here to download this File.</a>

Watch this Scientific Journal Video about A Modified Technique for Arteriovenous Fistula Construction in Rabbits at JoVE.com

A Modified Technique for Arteriovenous Fistula Construction in Rabbits

The present protocol proposes the creation of an arteriovenous fistula in rabbits using a modified no-touch technique. The technique involves the side-to-side anastomosis of the common carotid artery and external jugular vein without the dissection of the perivenous tissues or cutting off the artery.

Juxta-anastomotic stenosis is a challenging problem that often causes non-maturation and decreases the patency of an arteriovenous fistula (AVF). Injury ...

a-modified-technique-for-arteriovenous-fistula-construction-in-rabbits

Universidad San Sebastian

Research

JoVE Journal

Medicine

1.3K Views.  The First Affiliated Hospital of Soochow University. The present protocol proposes the creation of an arteriovenous fistula in rabbits using a modified no-touch technique. The technique involves the side-to-side anastomosis of the common carotid artery and external jugular vein without the dissection of the perivenous tissues or cutting off the artery.

Video: A Modified Technique for Arteriovenous Fistula Construction in Rabbits

Microsurgical Venous Pouch Arterial-Bifurcation Aneurysms in the Rabbit Model: Technical Aspects

For ruptured human cerebral aneurysms endovascular embolization has become an equivalent alternative to aneurysm clipping.1 However, large clinical trials have shown disappointing long-term results with unacceptable high rates of aneurysm recanalization and delayed aneurysm rupture.2 To overcome these problems, animal experimental studies are crucial for the development of better endovascular devices.3-5
Several animal models in rats, rabbits, canines and swine are available.6-8 Comparisons of the different animal models showed the superiority of the rabbit model with regard to hemodynamics and comparability of the coagulation system and cost-effectiveness.9-11 
The venous pouch arterial bifurcation model in rabbits is formed by a venous pouch sutured into an artificially created true bifurcation of both common carotid arteries (CCA). The main advantage of this model are true bifurcational hemodynamics.12 The major drawbacks are the sofar high microsurgical technical demands and high morbidity and mortality rates of up to 50%.13 These limitations have resulted in less frequent use of this aneurysm model in the recent years. These shortcomings could be overcome with improved surgical procedures and modified peri- and postoperative analgetic management and anticoagulation.14-16 Our techniques reported in this paper demonstrate this optimized technique for microsurgical creation of arterial bifurcation aneurysms.

An optimized technique for the microsurgical creation of arterial bifurcation aneurysms mimicking bifurcation human cerebral aneurysms is described. A venous pouch is sutured into an artificially created true bifurcation of both common carotid arteries. Facilitated microsurgical techniques and aggressive postoperative anticoagulation and analgesia lead to minimized morbidity rates and high aneurysm patency rates.

For ruptured human cerebral aneurysms endovascular embolization has become an equivalent alternative to aneurysm clipping.1 However, large clinical trials ...

Technical Aspects of the Mouse Aortocaval Fistula

Technical aspects of creating an arteriovenous fistula in the mouse are discussed. Under general anesthesia, an abdominal incision is made, and the aorta and inferior vena cava (IVC) are exposed. The proximal infrarenal aorta and the distal aorta are dissected for clamp placement and needle puncture, respectively. Special attention is paid to avoid dissection between the aorta and the IVC. After clamping the aorta, a 25 G needle is used to puncture both walls of the aorta into the IVC. The surrounding connective tissue is used for hemostatic compression. Successful creation of the AVF will show pulsatile arterial blood flow in the IVC. Further confirmation of successful AVF can be achieved by post-operative Doppler ultrasound.

The goal is to produce an arteriovenous fistula that is simple and reproducible. This method does not use sutures or glue adhesive. Therefore the samples can be used with the least amount of foreign materials for analysis.

Technical aspects of creating an arteriovenous fistula in the mouse are discussed. Under general anesthesia, an abdominal incision is made, and the aorta ...

The Arteriovenous (AV) Loop in a Small Animal Model to Study Angiogenesis and Vascularized Tissue Engineering

A functional blood vessel network is a prerequisite for the survival and growth of almost all tissues and organs in the human body. Moreover, in pathological situations such as cancer, vascularization plays a leading role in disease progression. Consequently, there is a strong need for a standardized and well-characterized in vivo model in order to elucidate the mechanisms of neovascularization and develop different vascularization approaches for tissue engineering and regenerative medicine.
We describe a microsurgical approach for a small animal model for induction of a vascular axis consisting of a vein and artery that are anastomosed to an arteriovenous (AV) loop. The AV loop is transferred to an enclosed implantation chamber to create an isolated microenvironment in vivo, which is connected to the living organism only by means of the vascular axis. Using 3D imaging (MRI, micro-CT) and immunohistology, the growing vasculature can be visualized over time. By implanting different cells, growth factors and matrices, their function in blood vessel network formation can be analyzed without any disturbing influences from the surroundings in a well controllable environment. 
In addition to angiogenesis and antiangiogenesis studies, the AV loop model is also perfectly suited for engineering vascularized tissues. After a certain prevascularization time, the generated tissues can be transplanted into the defect site and microsurgically connected to the local vessels, thereby ensuring immediate blood supply and integration of the engineered tissue. By varying the matrices, cells, growth factors and chamber architecture, it is possible to generate various tissues, which can then be tailored to the individual patient's needs.

The Arteriovenous AV Loop in a Small Animal Model to Study Angiogenesis and Vascularized Tissue Engineering

We describe a microsurgical approach for the generation of an arteriovenous (AV) loop as a model for analyzing vascularization in vivo in an isolated and well-characterized environment. This model is not only useful for the investigation of angiogenesis, but is also optimally suited for engineering axially vascularized and transplantable tissues.

A functional blood vessel network is a prerequisite for the survival and growth of almost all tissues and organs in the human body. Moreover, in ...

Bioengineering

A Novel Murine Model of Arteriovenous Fistula Failure: The Surgical Procedure in Detail

The arteriovenous fistula (AVF) still suffers from a high number of failures caused by insufficient remodeling and intimal hyperplasia from which the exact pathophysiology remains unknown. In order to unravel the pathophysiology a murine model of AVF-failure was developed in which the configuration of the anastomosis resembles the preferred situation in the clinical setting. A model was described in which an AVF is created by connecting the venous end of the branch of the external jugular vein to the side of the common carotid artery using interrupted sutures. At a histological level, we observed progressive stenotic intimal lesions in the venous outflow tract that is also seen in failed human AVFs. Although this procedure can be technically challenging due to the small dimensions of the animal, we were able to achieve a surgical success rate of 97% after sufficient training. The key advantage of a murine model is the availability of transgenic animals. In view of the different proposed mechanisms that are responsible for AVF failure, disabling genes that might play a role in vascular remodeling can help us to unravel the complex pathophysiology of AVF failure.

Here we present a murine model of arteriovenous fistula (AVF) failure in which a clinically relevant anastomotic configuration is incorporated. This model can be used to study the pathophysiology and to test possible therapeutic interventions.

The arteriovenous fistula (AVF) still suffers from a high number of failures caused by insufficient remodeling and intimal hyperplasia from which the ...

A Rabbit Model of Durable Transgene Expression in Jugular Vein to Common Carotid Artery Interposition Grafts

Vein graft bypass surgery is a common treatment for occlusive arterial disease; however, long-term success is limited by graft failure due to thrombosis, intimal hyperplasia, and atherosclerosis. The goal of this article is to demonstrate a method for placing bilateral venous interposition grafts in a rabbit, then transducing the grafts with a gene transfer vector that achieves durable transgene expression. The method allows the investigation of the biological roles of genes and their protein products in normal vein graft homeostasis. It also allows the testing of transgenes for the activities that could prevent vein graft failure, e.g., whether the expression of a transgene prevents the neointimal growth, reduces the vascular inflammation, or reduces atherosclerosis in rabbits fed with a high-fat diet. During an initial survival surgery, the segments of right and left external jugular vein are excised and placed bilaterally as reversed end-to-side common carotid artery interposition grafts. During a second survival surgery, performed 28 days later, each of the grafts is isolated from the circulation with vascular clips and the lumens are filled (via an arteriotomy) with a solution containing a helper-dependent adenoviral (HDAd) vector. After a 20-min incubation, the vector solution is aspirated, the arteriotomy is repaired, and flow is restored. The veins are harvested at time points dictated by individual experimental protocols. The 28-day delay between the graft placement and the transduction is necessary to ensure the adaptation of the vein graft to the arterial circulation. This adaptation avoids rapid loss of transgene expression that occurs in vein grafts transduced before or immediately after grafting. The method is unique in its ability to achieve durable, stable transgene expression in grafted veins. Compared to other large animal vein graft models, rabbits have advantages of low cost and easy handling. Compared to rodent vein graft models, rabbits have larger and easier-to-manipulate blood vessels that provide abundant tissue for analysis.

This method describes the placement of interposition vein grafts in rabbits, the transduction of the grafts, and the achievement of durable transgene expression. This allows the investigation of physiological and pathological roles of transgenes and their protein products in grafted veins, and testing of gene therapies for vein graft disease.

Vein graft bypass surgery is a common treatment for occlusive arterial disease; however, long-term success is limited by graft failure due to thrombosis, ...

Arterial Pouch Microsurgical Bifurcation Aneurysm Model in the Rabbit

Endovascular treatment for intracranial aneurysms gained importance over the past decades, consequently there is an increased need of testing endovascular devices. Animal models respecting rheological, hemodynamic and aneurysm wall conditions are highly warranted. Therefore, the aim of the present study was to design a novel standardized and reproducible surgical technique to create autologous arterial pouch bifurcation aneurysms with non-modified and modified wall conditions in rabbits.
Bifurcation aneurysms were created by end-to-side anastomosis of the right on the left common carotid artery, both serving as parent arteries for the arterial pouch, which was microsurgically sewn on. Grafts were taken from the proximal right common carotid artery, either for the control (n = 7, immediate autologous re-implantation) or modified (n = 7, incubated with 100 international units elastase for 20 minutes before autologous re-implantation) group. Pouch and parent artery patency were controlled by fluorescence angiography immediately after creation. At follow-up (28 days), all rabbits underwent contrast enhanced magnetic resonance angiography and fluorescence angiography followed by aneurysm harvesting, macroscopic and histological evaluation.
A total of 16 female New Zealand White rabbits were operated upon. Two animals died prematurely. At follow-up, 85.72% of all aneurysms remained patent. Both groups revealed an increase in aneurysm size over time; this was more pronounced in the control group (6.48 &#177; 1.81 mm3 at time of creation vs. 19.85 &#177; 6.40 mm3 at follow-up, p = 0.037) than in the modified group (8.03 &#177; 1.08 mm3 at time of creation vs. 20.29 &#177; 6.16 mm3 at follow-up, p = 0.054).
Our findings demonstrate the adequacy of this new rabbit model which allows for the creation of bifurcation aneurysms with different wall conditions in a microsurgical approach. Given the excellent long-term patency and the property of aneurysm growth over time, this model may serve as an important tool for preclinical evaluation of novel endovascular therapies.

Developing and testing endovascular devices for intracranial aneurysm treatment is still of great importance. Most aneurysm models used today miss either the important characteristics of an arterial degenerated wall or the hemodynamics of a true bifurcation. Therefore, we aimed to design a novel arterial pouch bifurcation model in rabbits.

Endovascular treatment for intracranial aneurysms gained importance over the past decades, consequently there is an increased need of testing endovascular ...

Neuroscience

A Rabbit Venous Interposition Model Mimicking Revascularization Surgery using Vein Grafts to Assess Intimal Hyperplasia under Arterial Blood Pressure

Although vein grafts have been commonly used as autologous grafts in revascularization surgeries for ischemic diseases, the long-term patency remains poor because of the acceleration of intimal hyperplasia due to the exposure to arterial blood pressure. The present protocol is designed for the establishment of experimental venous intimal hyperplasia by interposing rabbit jugular veins to the ipsilateral carotid arteries. The protocol does not require surgical procedures deep in the body trunk and the extent of the incision is limited, which is less invasive for the animals, allowing long-term observation after implantation. This simple procedure enables researchers to investigate strategies to attenuate the progression of intimal hyperplasia of the implanted vein grafts. Using this protocol, we reported the effects transduction of microRNA-145 (miR-145), which is known to control the phenotype of vascular smooth muscle cells (VSMCs) from the proliferative to the contractile state, into harvested vein grafts. We confirmed the attenuation of intimal hyperplasia of vein grafts by transducing miR-145 before implantation surgery through the phenotype change of the VSMCs. Here we report a less invasive experimental platform to investigate the strategies that can be used to attenuate intimal hyperplasia of vein grafts in revascularization surgeries.

The present protocol aims to experimentally create venous intimal hyperplasia by subjecting veins to arterial blood pressure for developing strategies to attenuate venous intimal hyperplasia following revascularization surgery using vein grafts.

Although vein grafts have been commonly used as autologous grafts in revascularization surgeries for ischemic diseases, the long-term patency remains poor ...

Creating Radio-cephalic Arteriovenous Fistula in the Forearm with a Modified No-Touch Technique

Autologous arteriovenous fistula (AVF) is the primary and best option to obtain vascular access for hemodialysis treatment; other options are arteriovenous graft (AVG) and central venous catheterization (CVC). The implementation of radio-cephalic autologous arteriovenous fistula (RC-AVF) in the forearm was preferred among patients with superior vascular conditions. However, there is a high rate of early fistula failure. The chosen surgical method is understood to have an effect on the maturation of the fistula. New surgical procedures such as radial artery deviation and reimplantation (RADAR) have been significantly improved for juxta-anastomotic stenosis. Nevertheless, new problems such as stenosis of arteries and narrowing of surgical indication were also found. In this report, we presented a modified no-touch technique (MNTT) to create an RC-AVF, in which the venous and arterial wall avoid devascularization and the radial artery does not sever.

We present a modified no-touch technique (MNTT) to create a radio-cephalic arteriovenous fistula (RC-AVF) in which the venous and arterial wall avoid devascularization and the radial artery does not sever.

Autologous arteriovenous fistula (AVF) is the primary and best option to obtain vascular access for hemodialysis treatment; other options are ...

Microsurgical Creation of Giant Bifurcation Aneurysms in Rabbits for the Evaluation of Endovascular Devices

Giant aneurysms are dangerous lesions requiring endovascular treatment, with high rates of aneurysm recanalization and re-rupture. Reliable in vivo models are rare but are required for testing new endovascular devices.&#160;We demonstrate the technical aspects of the creation of giant bifurcation aneurysms in New Zealand white rabbits (2.5-5.5 kg). A 25-30 mm long venous pouch is taken from the external jugular vein, and a bifurcation between both carotid arteries is created microsurgically. The pouch is sutured in the bifurcation to mimic a giant aneurysm. This protocol summarizes our previously published standard technique for venous pouch true arterial bifurcation aneurysms and highlights its essential modification steps for giant aneurysms. Using this modified technique, we were able to create an animal model for giant aneurysms with high comparability to humans regarding the hemodynamics and coagulation systems. Furthermore, low morbidity and high aneurysm patency rates were achieved. The proposed giant aneurysm model offers an excellent possibility for testing new endovascular devices.

Here, we describe the technique for the microsurgical creation of giant bifurcation aneurysms in rabbits for the evaluation of endovascular devices.

Giant aneurysms are dangerous lesions requiring endovascular treatment, with high rates of aneurysm recanalization and re-rupture. Reliable in vivo models ...

Carotid Artery and Jugular Vein Interposition: A Technique to Induce Venous Intimal Hyperplasia in Rabbit Model

Source: Nishio, H., et al. <a href="https://www-jove-com.bdigitaluss.remotexs.co/v/60931">A Rabbit Venous Interposition Model Mimicking Revascularization Surgery using Vein Grafts to Assess Intimal Hyperplasia under Arterial Blood Pressure</a> J. Vis. Exp. (2020).
This video demonstrates the interposition of the rabbit carotid artery with the jugular vein to induce venous intimal hyperplasia by subjecting the veins to arterial pressure. This model will help investigate molecular and genetic interventions to attenuate the progression of intimal hyperplasia after revascularization surgeries.

Interpozycja tętnicy szyjnej i żyły szyjnej: technika wywoływania przerostu żylnej błony wewnętrznej w modelu królika

Source: Nishio, H., et al. A Rabbit Venous Interposition Model Mimicking Revascularization Surgery using Vein Grafts to Assess Intimal Hyperplasia under ...