A Rodent Model of The Ross Operation: Syngeneic Pulmonary Artery Graft Implantation in A Systemic Position

Congenital aortic stenosis is a subgroup of congenital heart disease, characterized by an obstruction of the left ventricular tract in which the lesion is located at the valvular level. The options available for the correction of a congenital aortic stenosis are many. Each with their own and disadvantages.

An interesting therapeutic option in the pediatric population is the transfer of the pulmonary autograph to the aortic position named Ross operation. In this case, the pulmonary valve is then replaced with the homograft. The pulmonary autograph is characterised by growth potential, and does not carry the risks of lifelong anticoagulant therapy.

Despite this, its use remains extremely limited due to the possible dilatation of the pulmonary autograph and subsequent aortic regurgitation. In this study, we sought to establish a rodent model of pulmonary valve graft implantation in a systemic position, in order to evaluate the adaptation of the pulmonary graft. A checklist of all materials required was performed before each operation.

After the induction anesthesia, with 4%sevoflurane in one liter per minute of oxygen, the sedated animal was placed on a cork tray and its abdomen was shaved with a razor. The skin was then sterilized using an iodine solution. In order to protect the animal from getting wet and to prevent heat dispersion during the surgery, the animal was covered with a transparent plastic wrap.

The level of anesthesia was evaluated before performing the Pfannenstiel pubic incision by assessing the absence of response to obnoxious stimulus. Pericardiectomy and atherectomy were carried out in order to obtain a complete view of the aortic arch. The remaining fatty tissues surrounding the aorta was also removed.

A micro plier was inserted under posterior wall of the vessel to isolate the pulmonary artery. To maximize the link to the graft, the latter was cut as close as possible to its bifurcation using micro scissors. The pulmonary artery was then gently held with a ring tipped micro forceps, and separated from the right ventricular using the micro spring scissors.

The pulmonary graft was therefore harvested including some right ventricular muscle. The pulmonary artery is placed on a gauze moistened with cold saline on the operating table and the pulmonary root was inspected under the operating microscope. Any abundance surrounding tissue was cut off.

Only one millimeter of ventricular muscle was left while the link to the vessel was set at five millimeters. A median longitudinal abdominal incision was performed and two mini retractors were used to keep the abdomen open. The intestines were extracted, allowing the visualization and exposure of the infrarenal abdominal aorta.

A 2-0 silk suture was used to create a loop around the abdominal aorta in order to lift the vessel and separate the abdominal aorta from inferior vena cava. Two Yasargil clips were used to clamp the infrarenal abdominal aorta and placed at a distance of 1.5 centimeters from each other. The abdominal aorta was transected at the midpoint between the two clips.

The pulmonary artery was placed between the two ends in the ventricular end towards the cranial portion of the animal. A 10-0 Prolene suture was used to perform two landmark single stitches connecting the pulmonary artery to the abdominal aorta. These sutures were placed on opposite sides of the proximal vessel circumference.

The same procedure was performed on the distal end of the graft. An end to end anastomosis between pulmonary artery and abdominal aorta was then performed beginning with the distal end. The end located more distally to the surgeon was used for the posterior anastomosis, using the recipient to graft out-in, in-out sequence in order to create a running suture of about six stitches.

Once the suture reached the proximal landmark, a double half stitch completed by making a square knot using the suture and one of the two ends of the proximal landmark suture. Protected mosquitoes were then applied to the sutures to provide traction. The same anastomosis was performed on the anterior wall.

The entire procedure was then carried out on the proximal end of the pulmonary artery and particular attention was taken in order to avoid including any leaflet in the suture line. The distal clip was released first to let the pulmonary graft fill up with retrograde blood, that is a low pressure flow in order to check the anastomosis. Once the distal anastomosis was evaluated, the same procedure was performed at the proximal end.

At the end of the operation, the rat was then placed under a heating lamp and visually monitored until wakening. During the follow up, the animals underwent serial ultrasound studies at one week, one month and two months. These studies allow the measurement of vessel diameter, peaks systolic velocity and end-diastolic velocity.

These parameters were measured inside the graft and at the level of proximal and distal abdominal aorta. We utilized a total of 39 adult Lewis rats. 17 animals were used as pulmonary graft donors, 17 animals as recipients, and five animals as sham operated and considered the control group.

No fatal event occurred at the time of the operation. We had two death after the operation with an overall survival rate of 91%at two months. The median body weight of the animals was a little lower for the donor group when compared to the recipients, 328 grams versus 387 grams.

We have experimented at 6%degrees in body weight after one week, and then all rats regained their weight reaching a median value of 397 grams after two months. The result of the study showed a rapid significantly increase of the pulmonary graft diameter within the first week of implantation followed by a plateau during the following two months. The median value of the pulmonary graft in genetic position was 3.2 millimeter, and it increased immediately to 4.0 centimeter after one week and it maintained to 4.07 millimeter after one month and 4.27 millimeter at two months.

The other result were able to demonstrate that there was a rapid significantly decrease in the peak systolic velocity measure at the graft level with 2D echo after a one week of implantation, follow by an increase during the last two months, reaching the value that we are comparable to the one obtained in sham operated rats. When considering the astrological analysis of the graph and the time of explantation, we didn’t see any significantly sign of endothelia thrombosis or multi-cification. This experimental model of syngeneic pulmonary valve draft implantation in systemic position in a rodent model reveal safe, effective, and reproducible.

It gave us the possibility of characterizing the modification on the pulmonary artery after exposure to systemic pressure and this could represent the basis for understanding the causes of pulmonary autograft failure. The small size of the animal used simplified the surgical and the postoperative management. This allowed us to obtain a useful model with limited bacterias and animal expenses.

Conclusion, the current study showed that the systemically place in syngeneic pulmonary graft in a rodent model represent the simple and physical platform for the development and evaluation of novel surgical techniques, such as the root reinforcement and drug therapies aimed at reducing this dilatation of the graft, maybe with the aim of further improving the outcomes of the Ross operation.