Kidney Procurement in a Preclinical Large Animal Model

Our research aims to develop transformative new tissue and organ preservation techniques, such as using nature-inspired, next-generation cryoprotective agents that allow for ice-free preservation of organs at sub-zero temperatures. Innovative microsurgical small and large animal model are utilized as translational platforms to advance organ and tissue refrigeration, immune modulation, and immune monitoring. Technological advancement are currently transforming the field of transplantation.

However, there are remaining challenges in establishing clinically relevant large animal settings to test these technologies in vivo. By extending preservation capabilities from hours to several days, this research has the potential to alleviate one of the most daunting challenges in transplantation, the extremely limited time for organ storage, while also increasing the number of organs available for transplant. To begin, perform a median laparotomy on a surgically prepped anesthetized animal, incising about 25 to 30 centimeters, to gain optimal access to both kidneys.

Insert a standard abdominal retractor. Cover the colon and small bowel with towels soaked in warm saline. Retract the bowels to the right for access to the left kidney or to the left for access to the right kidney.

Open the peritoneum overlying the kidney and dissect around the kidney to free any adhesions. Dissect the ureter until 10 to 12 centimeters of length is obtained. Next, dissect the renal vein until its origin from the inferior vena cava is exposed.

Similarly, dissect the artery until its origin from the aorta is exposed. After complete renal dissection, tie the ureter distally with a 2-0 silk ligature. Cut proximally to the tie, leaving the proximal ureter end open for urine drainage.

Administer 100 international units per kilogram of heparin intravenously and wait for two minutes to ensure adequate heparinization of the kidney. Clamp the renal artery close to the aorta using Satinsky vascular clamps. Similarly, clamp the renal vein close to the inferior vena cava.

Remove the kidney graft by cutting the renal artery and vein close to the clamps. Flush the kidney with ice-cold University of Wisconsin, or UW, solution or Custodiol histidine-tryptophan-ketoglutarate, or HTK, preservation solution. Remove the perfusion cannula and place the kidney in a sterile organ bag filled with the same ice-cold preservation solution used for flushing.

Place the bag within a second sterile organ bag. Ligate the renal artery stump with a 2-0 silk ligature and close the renal vein stump using a two-layer running suture with 6-0 polypropylene. To harvest blood for machine perfusion, identify the infrarenal abdominal aorta.

Free any large adhesions or tissue covering the vessel. Insert the blood collection bag needle directly into the aorta. Hang the bag below the animal to facilitate filling.

Kidneys were successfully retrieved for transplantation, machine perfusion, and primary cell culture experiments. Notably, successful kidney retrieval was achieved in all experimental groups, implying no complications with this surgical procedure model. The gross appearance of kidney grafts varied at different stages as described in this figure.

The native kidney appeared deep red and vascularized. The kidney graft after flushing with ice-cold preservation solution appeared pale. After machine reperfusion, the kidney graft regained a pinkish hue with visible vascular flow.

After transplantation, the kidney appears homogeneously perfused, similar to a native kidney. 64%of kidneys exhibit standard anatomy of one renal artery and one renal vein. The most common variations included one artery with two veins, one artery with three veins, two arteries with one vein, and two arteries with two veins.

Bilaterally typical anatomy was observed in 46%of pigs while 22%had one atypical kidney on the left, 14%on the right, and 19%had both kidneys atypical.