Pancreas transplantation is a curative treatment for people with diabetes. However, organs are scarce, and strategies to improve the donor pool are necessary. Normothermic Ex Vivo Perfusion has already been successful for other organs that could potentially allow for assessment and repair of pancreas allografts.
Pancreas is the most often organ discarded for transplantation. This technique has a potential to increase the organ pool for people who live with complicated diabetes. This method will also allow us to better understand ischemia-reperfusion injury, and identify biomarkers of pancreas damage.
The setup can be a little complicated, and considering that the pancreas is a very sensitive organ, practice for its retrieval is necessary. To begin, disinfect and cover the surgical field. Perform a midline incision from the xiphoid to the pubic symphysis, and extend the surgical field with a left lateral incision for better exposure.
Dissect the inferior vena cava from the abdominal aorta. Further free the aorta from surrounding tissue, and ligate the small lumbar aortic branches. Identify and place ligatures around both renal arteries.
Once the back of the aorta is free, pass two ligatures around it where the lower ligature will eventually be tied just above the iliac artery bifurcation, and the upper ligature will be tied five centimeters above the previous tie. After dissecting the hepatic hilum, tie all arteries as close to the liver as possible. Then identify the common bile duct.
Place two ligatures close to the liver, and divide the structure. Next, identify and dissect around the supra hepatic portion of the aorta and place a tie around it. Next, open the lesser sack to allow ice to cool the pancreas and mobilize the pancreas as little as possible before flush.
Then administer 500 international units of heparin per kilogram of donor weight through the central line, and start blood collection in citrate, phosphate, dextrose, saline, adenine, glucose and mannitol bags using the jugular catheter. After tying the inferior aortic ligature, cannulate the aorta with a flush line above the iliac bifurcation tie. Secure the cannula with the upper tie, and ligate both renal arteries.
Once enough blood has been collected, tie the supraceliac aorta, and place a clamp above the tie. Next, initiate a flush with University of Wisconsin Preservation Solution. Then cut an opening in the portal vein and cava for venting.
After flushing one liter of University of Wisconsin Solution assess the pancreas tail and duodenum C-loop. And once adequately flushed, begin dissection by identifying and clamping mesenteric vessels. Retrieve the pancreatic graft, and a segment of the cava or iliac vein, for extension of the portal vein.
And place the organ inside an organ bag that is placed inside a basin filled with ice. After removing the flush line from the distal part of the aorta, close it with a tie, and fill the organ bag with the remaining UW Solution. Next, perform portal vein extension using previously recovered cava or iliac vein with 6-0 Prolene, and cannulate the portal vein and proximal aorta with one fourth times 3/8 Reducer.
Then cannulate the distal part of the duodenum with a Malecot catheter and tie. Next clamp the end of the catheter to avoid spilling duodenal content, and oversaw mesenteric vessels with 4-0 Prolene. After registering the weight of the graft, keep the graft in static cold storage until the start of the Ex Vivo Pancreas Perfusion.
Place the pancreas chamber on a Mayo table, and introduce the arterial and venous tubing through the holes intended for this purpose. Connect and turn on the external heater unit. After placing the suction tubing inside the roller pump, connect one end to the tubing that comes out from the chamber to collect the fluids, and the other end to the venous reservoir to collect all organ loss of perfusate.
Then connect the oxygen tubing to the gas tank containing the carbon mixture and the oxygenator. Then connect the heater pump unit tubing to the oxygenator. Clamp arterius and venous outflow lines as well as the outflow of the venous reservoir.
After filling the venous reservoir with the perfusate, using one syringe pump continuously administer the vasodilator at eight milliliters per hour into the arterial line. Then using a second syringe pump, continuously administer the enzyme inhibitor directly into the venous reservoir. After turning on the heart-lung machine, start up the pressure, temperature, and timer panels.
Then turn on the heater pump to warm the perfusion solution to 38 degrees Celsius, and open the oxygen carbon dioxide supply. Then remove the tubing clamp placed on the outflow of the venous reservoir, and start the centrifugal pump, taking it up to 1, 500 RPM. Next, clamp the tubing bypassing the arterial filter.
And release air from the arterial filter. Afterward, zero the arterial and venous pressure lines. After opening the organ bag where the pancreas is stored, flush with 200 milliliters of albumin through the arterial cannula.
Then remove the pancreas from the ice, and position it inside the organ chamber, ensuring that the arterial and venous tubing are air free. Next, after releasing the clamp from the arterial side, clamp the shortcut between the arterial and venous tubing. Once blood starts coming outta the arterial tubing, connect the line to the arterial cannula.
Then set arterial pressure to 20 to 25 millimeters of mercury by regulating the speed of the centrifugal pump, and connect venous tubing once blood starts coming out from the venous cannula. Next, administer one vial of verapamil directly to the arterial side when the pancreas is completely connected and no measure bleeds are observed. After recording the various parameters as described in the manuscript.
take the samples for analysis. Disconnect arterial and venous tubing when perfusion is over. after removing the graft from the organ chamber, flush with cold University of Wisconsin Solution, and weigh.
Then store the graft on ice in a sterile organ bag until the moment of transplantation. The pressure and flow were measured through the whole perfusion, and the results demonstrated that it remains stable. The metabolic activity was estimated by calculating the mean oxygen consumption of the graft.
The results show that pH, sodium, calcium, and bicarbonate measurements were within physiological values during the whole perfusion. It was observed that the lactate and potassium levels decreased during the perfusion and achieved close to normal values at three hours. Since the circuit is a closed system, amylase and lipase levels were expected to increase during the perfusion.
However, the increase in levels does not seem to correlate with the damage to the graft. It is very important to remember that the pancreas must be manipulated as little as possible before flush and during the extraction. We hope that in the future, this model will allow for therapeutic interventions to repair organs not suitable for transplant.
This technique will allow researchers to further study pancreas ischemia-reperfusion injury, preservation, assessment, and eventually repair.
