The protocol would help provide a step-by-step approach to the precarious procedure of pancreas transplantation in porcine models. The technique follows nearly the same steps as observed in humans, and therefore, is the closest possible model to replicate in clinical practice. The porcine visceral structures and the vessels are extremely fragile when compared to humans.
And therefore, that provides the biggest challenge for anybody who is doing the surgery for the first time. Begin by making a midline incision from the xiphisternum to the symphysis pubis using the pure cutting mode of Bovie's electrocautery. Ensure to keep lateral to the urethra in the lower part of the incision beyond the phallus to avoid injury.
Place the self retaining abdominal wall retractor to avoid injuring the spleen and liver, to optimize exposure to the surgical field. To mobilize the head of the pancreas, provide countertraction to the pancreaticoduodenal groove with the help of the assistant. Dissect along the avascular fascial plane between the duodenal lobe of pancreatic parenchyma in the infrahepatic IVC.
Initiate the pancreatic ring mobilization by dissecting along the fascia on either aspect of the portal vein to ensure complete mobilization of the connecting lobe from the overlying portal vein. The safest way is to start dissecting along either side of the portal vein and separate the tissues in a clockwise or anti-clockwise direction. Next, identify the junction between the pancreatic and parental tissues and start dissecting the tail off the underlying splenic vein, staying close to the parenchyma.
Ligate and divide the small venous tributaries to the parenchyma, taking care not to injure the splenic vein, using Silk 3-0 ties. For the pancreatic corpus mobilization, dissect along the thin layer of tissue separating the parenchyma from the large bowel and the stomach, preserving the thin vascular arcade along the infrapyloric region. Then for the pancreaticoduodenal mobilization, separate the parenchyma from the C-loop of the duo by sharp dissection in the pancreaticoduodenal groove by traction, countertraction, preserving the thin vascular arcade along the C-loop of the duodenum.
Next, identify the pancreatic duct. Ligate with Silk 2-0 ties and divide, keeping around three to five millimeters stump of the duct on the duodenal C-loop, staying away from the duodenal vascular arcade. Complete the last part of the mobilization by dissecting the parenchyma from the junction between the connecting lobe, duodenum, and colon on either side of the portal vein.
Extract the specimen in mass by dividing the connecting lobe on the anterior aspect of the portal vein. The parenchymal division is required to extract the specimen because of its circumferential location around the portal vein. Ensure hemostasis along the areas of dissection and inspect the duodenum for any injury and vascular congestion.
Prepare the graft portal vein and proximal aortic end by trimming the edges for anastomosis. Place the prepared graft in the organ bag with UW solution on an ice bath. Take core needle biopsies from the pancreatic tail, store them in Formalin, and snap freeze.
Extract RNA at a later stage. Suture the biopsy site using Prolene 6-0 in a figure-of-eight fashion. Begin the aortocaval dissection by identifying the right ureter and dissecting along the fascia, separating the ureter from the retroperitoneal covering over the inferior vena cava.
Dissect along the lateral border of the aorta to expose it from the underlying soleus muscle. Next, dissect along the interaortocaval groove to separate the aorta from the IVC. Ensure adequate mobilization of the aorta and IVC by a trial of Satinsky's vascular clamp placement.
Switch position with the first assistant and prepare the surgical field by retracting the bowel loops on the left side to expose the major vessels for anastomosis. Place the trial clamps on IVC and aorta to reassess an adequate mobilization of both the vessels for anastomosis. After placing the vascular side-biting clamp in the IVC, make a small opening in the anterior wall of the vessel and extend it to an adequate size cranially and caudally using Potts scissors.
Flush the lumen with heparinized saline using the flexible sheath of an IV cannula. Using Prolene 6-0 double needle, make corner inside-out sutures on the IVC and graft portal vein. Secure the coddle corner suture, and run one of the needles along the posterior wall of the graft and recipient vein in a continuous fashion.
Secure the anterior wall in an outside-in continuous fashion, and secure the knot in the center of the anterior wall after flushing the lumen of the anastomosis with heparinized saline. Then place a Bulldog's vascular clamp on the graft portal vein away from the anastomosis, and slowly release the side-biting clamp from the recipient IVC. Check for venous refilling, and any major bleed from the anastomosis.
Next, place the side-biting clamp on the aorta without injuring the lumbar branches along the posteromedial wall of the vessel. Instruct the anesthetist to inject heparin and methylprednisolone through the central venous catheter. Make an opening on the the anterior wall of the vessel, and extend it cranially and caudally, taking care not to dissect along the vessel wall.
Flush the lumen with heparinized saline. And using Prolene 6-0 double needle, suture the proximal end of the graft aorta to the recipient aortic opening continuously by the parachute technique. Flush the lumen with heparinized saline before tying the final knot on the anterior wall of the vessel.
Before initiating reperfusion, check the hemodynamic alterations for any drastic fall in the mean BP.Monitor the invasive BP minute-to-minute, and perform a dose titration of the vasopressor and fluid preload to keep the target mean BP between 45 to 50 millimeters of mercury. Then maintain the hemostasis after removing the Bulldog clamp from the vein by assessing for any major bleed from the parenchyma, paraaortic region, and periportal area. Release the aortic clamp and check for bleeding from the arterial anastomotic site.
Secure the bleeding points with ligatures and hemostatic suturing. Ask the anesthetist to inject a vial of tranexamic acid through the central venous catheter. It is essential to minimize aggressive handling of the pancreas during this phase of reperfusion to avoid graft edema and hematoma.
After ensuring no mesenteric torsion, isolate a loop of jejunum 40 to 50 centimeters away from the duodenojejunal junction, and bring it close to the graft. Anastomose the graft duodenum to the recipient jejunum in a side-to-side continuous fashion using polydioxanone 4-0 suture. Confirm hemostasis around the graft and at the anastomosis sites.
Then take three core needle biopsies from the pancreatic tail one hour after reperfusion. Store in Formalin, snap freeze, and extract RNA later. Suture the biopsy site with Prolene 6-0 in a figure-of-eight fashion.
After assessing for retained mops and securing hemostasis, suture the rectus abdominis in a continuous fashion using polydioxanone 0 needle, keeping the lower midline away from the urethra. Close the skin using continuous Silk 0 sutures. Create a subcutaneous tunnel on the neck, and secure the central venous catheter by burying it under the tunnel and suturing the overlying skin with a Silk 0-0 cutting needle.
After ensuring a stable mean BP and an improving trend of pH and lactate levels on the blood gas analysis, remove the arterial catheter and ligate the vessel to secure hemostasis in the neck. Close the overlying skin with Silk 0 suture in a continuous fashion. The mean heart rate one hour after reperfusion was 135 beats per minute on average, and the mean blood pressure was approximately 37 milliliters of mercury.
These levels were maintained under a high dose of norepinephrine infusion that could be tapered and discontinued in all five cases before shifting the animal back to its pen. The lactate levels were monitored in the operating room for three hours after reperfusion. All five animals were clinically alert, evident by active movement of limbs and breathing patterns within five to six hours of reperfusion.
The postoperative amylase, serum lipase, and serum lactate dehydrogenase levels were monitored for the first three days. A 120 minute for glucose tolerance test was performed on the third postoperative day. Blood samples were subsequently withdrawn and processed for glucose levels at two minutes, five minutes, 10 minutes, 20 minutes 30 minutes, 60 minutes, 90 minutes, and 120 minutes.
Meticulous handling of the vascular arcade around the pancreas during pancreatectomy is crucial. And performing this step carefully ensures a good vascular supply to the duodenum at the end of the surgery, which in turn is a key determinant of success after pancreatectomy. This technique has opened up the option of machine perfusion with transplantation in porcine models, and establishing survival models.
