Creation of Two Saccular Elastase-Digested Aneurysms with Different Hemodynamics in One Rabbit

Our protocol describes a reproducible technique to create a rabbit aneurysms model with both stump and bifurcation hemodynamics and degenerated wall condition in a single animal and during a single surgical approach with low morbidity and mortality rates and high patency rates. The model allows a direct comparison of the natural course of saccular aneurysms with respect to the role of hemodynamics, as well as the testing of endovascular devices in two distinct aneurysm configuration and flow condition. The main drawback of this method remains the same as for the creation of the classical bifurcation and stump aneurysms model, the need for sophisticated laboratory equipment and specific microsurgical skills.

Use a scalpel to make a median skin incision from the hyoid bone until 1.5 centimeters caudal to the manubrium sterni. Prepare the subcutaneous and fat tissue from the medial incision while performing meticulous hemostasis. Free the sternocephalicus muscle from the adherent connective tissue, and apply lidocaine topically to avoid myoclonus.

Expose the right common carotid artery, or CCA, medially of the sternocephalicus muscle, and keep it wet with wet swabs. Prepare the lateral and proximal parts of the sternocephalicus muscle, and retract it medially with a vessel loop to expose the CCA. Identify the external jugular vein, and protect it with a wet micro swab.

Dissect the connective tissue carefully along the CCA until the bifurcation of the brachiocephalic trunk to expose the artery. If small branches are emerging from the artery, coagulate them with the cauterizer. Apply two temporary clips, with the first at the origin of the CCA and the second two centimeter distal from it.

Place a rubber pad under the vessel, and rinse with papaverine hydrochloride for vasodilation. Remove the adventitia carefully using microscissors. Perform an arteriotomy below the distal clip with a 22-gauge four catheter, and insert the catheter caudally up to the proximal clip.

Flush the segment intraluminally with heparinized sodium chloride until there is no blood visible. Then fix the catheter with a 4-0 ligature. Through the catheter, inject 0.1 to 0.2 milliliters of elastase into the artery segment, and incubate for 20 minutes.

After 20 minutes of incubation time with elastase, clear the elastase solution, and change the syringe to rinse the artery segment about 10 times with 0.9%saline. Apply two ligatures, with the first at five millimeters distal of the proximal clip and the second just proximally under the arteriotomy. Cut the vessel at about three millimeters above the first ligature and one more time between the second ligature and the distal clip.

Keep this autologous graft in a heparinized solution until the creation of the bifurcation aneurysm. Carefully open the first proximal clip, and measure the aneurysm. To create a bifurcation aneurysm, remove the adventitia carefully, and make about a two-millimeter longitudinal incision laterally in the stump of the right CCA.

Apply two temporary clips on the left CCA to delimit the segment of about one centimeter, and remove the adventitia in between. Perform an arteriotomy with a 23-gauge needle. Then, flush the segment with heparinized sodium chloride.

Enlarge the arteriotomy using microscissors to about four to five millimeters to allow the suturing of the right CCA and the aneurysm pouch. To perform the anastomosis, suture the proximal back wall of the right carotid blunt with five stitches, starting at the proximal edge of the arteriotomy on the left CCA. Then, suture the backside of the aneurysm pouch with four to five stitches, starting at the distal edge of the arteriotomy on the left CCA.

Continue with the distal backside at the level of the fish mouth incision to suture with the vertical backside of the aneurysm graft with three stitches. Suture the front side of the fish mouth incision with three stitches, starting upwards and moving downwards. Finish with the front suture between the left CCA and front side of the aneurysm graft and right CCA with about six stitches.

Before finishing the anastomosis, rinse the vessels with heparinized 0.9%saline solution intraluminally. Remove the clip on the right CCA while putting some pressure on the anastomosis with the micro swabs for hemostasis. Then, continue by removing the distal clip from the left CCA.

If there is no major bleeding, continue with taking out the proximal clip on the left CCA to allow blood flow. At follow-up, aneurysm patency was confirmed by magnetic resonance angiography. A three-dimensional TOF sequence acquired with a three-tesla MRI focused on the neck arteries is shown.

The stump aneurysm is observed on the right subclavian artery, and the bifurcation aneurysm is observed on the bifurcation created by anastomosing the right CCA on the left subclavian artery. Aneurysm patency was also confirmed by macroscopic inspection after tissue extraction. Major grooves on the clip indicate one millimeter, and minor grooves indicate 0.5 millimeters.

Microscopic analysis of the stump and bifurcation aneurysms was performed after staining the samples with hematoxylin-eosin. Two steps are critical during surgery, first, the exposure and dissection of the right carotid artery until brachiocephalic trunk, and second, the protection of vital structures like trachea, jugular vein, and laryngeal nerve.