The overall goal of this procedure is to establish an accurate model and easily feasible method to investigate the development of transplant vasculopathy. This is accomplished by using the PVG to a CI strain combination. The second step of the procedure is to harvest the donor aorta.
The third step of the procedure is to prepare the recipient. The final step of the procedure is to anastomose the donor aorta with the recipient's aorta. Ultimately, results can be obtained that show neointima formation using patho histology.
Hi, I'm Sonya Schwefel, professor in transplant immunology in Thet Eye Lab at the University Heart Center in Hamburg, Germany. And I am Manish Duboff. Also from the TS Eye Lab.
My laboratory is interested in transplant immunology. In this video we'll introduce you to the Orthotopic aortic transplantation model, an easy and valid model to study the development of transplant vasculopathy In Wes. We use this procedure in our laboratory to investigate chronic rejection in cardiovascular questionings.
In the Orthotopic aortic transplant model, the aorta can easily be evaluated histologically. In addition, the PVG to a CI model is especially useful for transplant vasculopathy studies. Since acute vascular rejection is not a major confounding factor and the development of vasculopathy is similar to the findings in the clinic.
So let's get started. The donor used in this ORTHOTOPIC aortic transplantation is A PVG rat weighing approximately 250 to 300 grams. After the animal is anesthetized shave its abdominal and thoracic hair.
Place the rat on its back and place a face mask over its nose and mouth. To maintain the anesthesia, disinfect the abdominal and thoracic area widely using Provo iodine, followed by 80%ethanol. Next, check the reflexes of the animal by pinching its hind feet.
To confirm that it is sufficiently anesthetized. Exsanguinate the rat by opening the abdomen and cutting a hole into the abdo aorta. Carefully open the thorax to find the descending thoracic aorta.
Dissect the thoracic aorta from surrounding tissues, such as fat nerves and the esophagus. Withdraw a 1.5 centimeter piece of the aorta using acuta without tissue damage of the aorta. Perfuse the aortic graft thoroughly with cold saline and store the graft at four degrees Celsius.
The recipient for this orthotopic aortic transplantation is an A CI rat weighing approximately 250 to 300 grams. To begin this procedure, shave the abdominal area of the anesthetized rat and apply eye ointment to prevent the eyes from drying during anesthesia. Place the rat on its back and place a face mask over its nose and mouth to maintain the anesthesia.
Next, disinfect the abdominal area using provo iodine, followed by 80%ethanol. After pinching the hind feet to confirm that the rat is sufficiently anesthetized, open the abdomen by performing a midline abdominal incision and separating the skin and muscle. Two steps.
Once the abdomen is open, place the intestines in the saline ized glove. Fold the glove around the intestines to prevent loss of moisture. Remove the fatty tissue covering the abdominal aorta.
Dissect the aorta from the infrarenal region to the bifurcation, being careful not to cause damage to the branches of the vessels. If necessary, ligate branches of the aorta after dissecting the aorta. First, use a microsurgical clamp on the infrarenal part of the aorta to stop the blood flow.
Next place a second clamp close to the bifurcation of the aorta. Now that the blood flow has been stopped, remove a short segment of the aorta. Then take the donor aortic graft that has been kept in 0.9%saline on ice.
Shorten it to the appropriate length and position it in the gap in the aorta. Connect the donor aorta to the recipient aorta by performing running eight zero prolene sutures, starting with the cranial end-to-end anastomosis. When the distal end-to-end anastomosis is finished, carefully open the distal clamp.
Next, carefully open the cranial clamp. There should be a visible pulse at the distal end of the aorta. Now move the intestines back into the abdomen and flush the abdomen with prewarm sterile saline.
Close the muscle layer of the abdominal wall using running six zero proline sutures. Then use running five zero proline sutures to close the skin while the rat is still under an anesthesia. Inject four to five milligrams per kilogram of carprofen subcutaneously.
Return the animal to its cage. Add methimazole to the drinking water for pain medication for three days. Post transplantation.
The histopathology results after 120 days will be demonstrated on our histology microscope. The pictures will clearly show the development of transplant vasculopathy. The developed luminal narrowing will be highlighted by computer animation.
We have just shown you how to perform an auto topic aortic transplantation in rats. We've also demonstrated the successful development of transplant vasculopathy after 120 days using histopathology. This method enables the investigation of initial effects of drugs on chronic allograft vasculopathy.
