The technique of orthotopic liver homotransplantation in the pig

Jourvzal of Surgical Research Clinical

and Laboratory

Volume 8 Number

THE TECHNIQUE HOMOTRANSPLANTATION J.

TERBLANCHE, J.

H.

BOWES, K.E.F.

From the Departments University of Bristol, Part of this work was the British Empire Cancer Submitted for publication

1968

ORTHOTOPIC

F.R.C.S.,

M.D.,

CH.M.,

M.B.,

F.F.A.R.C.S.,

HOBBS,

IT has become evident in recent years that a large number of patients with liver disease might possibly be treated by liver homotransplantation [4, 261. Thus research in this field is being undertaken in an increasing number of centers, following the pioneer work of Goodrich and Welch [9], Moore [ 16, 171, and Starzl [20]. A considerable body of evidence is accumulating that the dog is an unsuitable animal for experimental liver homotransplantation, because ,of the presence of hepatic vein sphincters, leading to hepatic outflow block and imts ensuing complications [ 15, 211. The pig, which appears to have no hepatic vein sphincters and therefore more closely resembles man in this respect, appears to be the best alternative. The object of this report is to present a technique of orthotopic liver homotransplantation in the pig, which was evolved as a result of a group of preliminary experiments [23] and has subsequently been used in a series of forty-one liver transplants. Some adjustments were made in the first half of the study but the ‘technique has remained rela-

ics,

4, April

IN CH.M.,

PEACOCK,

J.

OF

Investigation

of Surgery and AnesthetBristol, England. done under a grant from Campaign. May 29, 1967.

M.B.,

THE F.C.S.

LIVER PIG (%A.),

F.R.C.S., AND F.R.C.S.

tively unchanged in the later experiments. Anescthesia of the pig will be described, pointing out the particular problems related to liver homotransplantation, and the method of endotracheal intubation will be presen’ted. The operative technique used in the recipient animal, and in the preparation of the donor liver, will be presented. It is felt that this information may prove useful to other future workers contemplating the use of the pig for liver homotransplantation. Preliminary results of this series have been presented [ 12, 18, 241, and a detailed account will be published in due course [25]. Of the 41 pigs in the series, 25 survived the operative procedure, 18 of these in the last twenty-one experiments; 16 animals survived for 36 hours or longer and 1 is still alive at twenty months (operation date June 15, 1966). In none of the animals was immunosuppression employed, as the object was to study the pattern of liver rejection in pigs. The most impressive finding was that despite the fact that the pig was shown to be immunologically competent [13], the degree of liver rejection seen was much less than that reported in the dog. In addition, a significant number of animals survived for relatively long periods of time despite the lack of immunosuppressive therapy. It was concluded that the pig liver is an immuno151

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logically relatively favored organ, and it was suggested that the same might apply to man.

TECHNIQUE Blood Fresh blood in sterile, siliconized 2-litre bottles, containing 250 mg. of ampicillin sodium, and either 40 mg. of heparin or 480 ml. of acid citrate dextrose, is obtained from local abattoirs ,on the morning of each experiment. Heparinized blood is used on ‘the day of the operation only, and titrated blood is stored at 4°C. for a maximum of 2 weeks. A separate sample of clotted blood is obtained from each pig for blood grouping and cross-matching. The A and 0 blood groups of the donor and recipient animals are determined. The abattoir blood is similarly grouped and a direct compatibility test is performed between each bottle and the recipient pig; only blood of the correct group is administered to the animal. No incompatibility was detected even when the recipient animal and bottled blood were of different A and 0 groups. The one exception occurred when heparinized blood was obtained from a pig which had previously been used in a skin graft experiment.

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halothane in a nitrous oxide and oxygen mixture (5 and 2 liters/mm., respectively) administered through a nose cone. At this stage it is important not to inflate the animal with anesthetic gases under pressure, as this distends both the stomach and the bowel, which later severely restricts the surgical field. In the past the difficulties encountered in intubating the pig have tended to limit its use to short-term nonsurvival studies. These difficulties are the distance between the incisors and the larynx, the fact that the large epiglottis obstructs the view, and the axis of the trachea is such that a curved endotracheal tube has to be twisted through 180 degrees to enable it to pass down the trachea (Fig. 1). Becently descriptions of the technique of intubation in the pig have appeared in the literature [5, 81. A long straight-bladed laryngoscope is used. Intubation is accom-

Animals The average weight of the pigs used was 40 kg. Donor and recipient animals were of different breeds [ 18, 251. They were starved for 24 hours preoperatively, as food was still present in the stomach with shorter periods of starvation. Both animals were operated upon simultaneously by two teams. Anesthesia No premeditation is necessary in pigs of this size. No long-acting sedative is given, as these agents might have an unexpectedly prolonged action due to the lack of a functioning liver during the transplant procedure. Inhalation anesthesia is used [5, 19, 271, as the pig lacks accessible veins, with the exception of small friable veins on the dorsum of the ear. In both donor and recipient animals anesthesia is induced with up to 4%

Fig. 1. The technique of endotracheal intubation in the pig. (A) Epiglottis obscuring the larynx. (B) Endotracheal tube impinging on the anterior wall of the trachea. (C) Further passage of the tube accomplished with ease after rotating it through 180 degrees.

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plished when the vocal cords are sufficiently relaxed to remain open, but while the animal is still breathing spontaneously. The long epiglottis frequently lies coiled with its tip in the esophagus, leading to a puzzling total absence of laryngeal landmarks (Fig. 1A). To view the larynx it is necessary to pass the endoctracheal tube into the esophagus, and on extracting the tube the tip of the epiglottis can be hooked forward. At this stage the endotracheal tube can be passed through the vocal cords, but it impinges in an area where the tracheal rings are weak, with the resultant possible danger of rupturing the trachea (Fig. 1B). To enable the tip to pass down the axis of the trachea the tube is rotated through 180 degrees and its further passage is usually accomplished with ease (Fig. 1C). A largebore gastric tube is passed into the stomach to keep the bowel deflated during the procedure. Anesthesia is maintained with up to 2% halothane in the same nitrous oxide and oxygen mixture. The animals are ventilated with Starling pumps (tidal volume 450 to 500 ml., and rate 18 to 20) via a circle system and soda lime absorber. The rate and tidal volume used are greater than would be thought necessary for this size of animal, This relative overventilation is to prevent atelectasis [ 71, which can be produced by constant ventilation [ 11. If the soda lime absorber in this system is fully switched onto the circuit, the pC0, falls sharply to values below 20 mm. Hg; therefore, it is adjusted so as to maintain the pCOZ between 30 and 40 mm. Hg. The donor animal usually presents no further problems. The arterial pressure is monitored and the mean pressure is expected to be between 80 and 100 mm. Hg. If it falls, the halothane in the mixture is reduced. The recipient animal requires careful observation throughout the course of the operation. The condition of the animal is judged from the electrocardiogram, arterial and central venous pressures, and half-hourly estimations of the arterial pCOa, pH, and 0, saturation. The mean femoral arterial

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blood pressure is read from a mercury manometer, while the arterial pressure tracing and lead I of the electrocardiogram are displayed son the oscilloscope of a Sanborn 4-channel recording system. The central venous pressure is read from a venous pressure manometer. Blood glucose estimations are performed at regular intervals, and approximately 25 gm. of glucose is administered during the procedure. The recipient is overtransfused with 800 to 1,000 ml. of fresh heparinized blood while the liver is being removed, as this is helpful in maintaining a satisfactory mean arterial blood pressure. In addition about 500 ml. of blood is lost when the implanted donor liver is flushed to clear acid metabolites prior to reestablishing normal circulation through this organ. There is a tendency for metabolic acidosis to develop during the bypass phase, and this is sharply accentuated when the circulation is reestablished through the implanted donor liver. As the blood pressure appears to be better maintained when the animal is alkalotic, sodium bicarbonate (60 mEq./lOO ml. in 10% glucose) is slowly introduced into the portal bypass line, where the 0ow is sufficient to ensure adequate dilution of this hypertonic solution. These animals tend to develop cardiac irregularities in the form of ectopic beats and atrioventricular block, possibly due to decreased conduction in the alkalotic heart as a result of a change in the ratio of ionized to unionized calcium [ 141. When irregularities occur they can usually be effectively reversed by the administration of calcium chloride. Calcium gluconate has been found to be less effective in the anhepatic animal. A number of problems arise in relation to anesthetizing an animal lacking a functioning liver. This exists in the recipient animal during the period when the blood bypasses the liver and immediately after implanting the donor liver. There is a very real danger of the potentiation of the action of anesthetic agents. Although it is tempting to avoid general anesthesia and use epidural anesthesia [3, 11, 281, the necessity to operate in the cervical region prevents the use of this 153

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In addition it is felt interesting technique. that inhalation agents, with their excretion through the lungs, are safer than intramuscular opiates, which would be a necessary adjuvant to local anesthesia. In the earlier experiments, because of the fear of using a high concentration ,of halothane for a prolonged period of time, relaxant drugs were added to the anesthetic regime. Gallamine triethiodide was found to be unsatisfactory because of the accompanying tachycardia and the possible fall in the cardiac output [6], the tendency to increased bleeding, and the apparent large doses required. Tubarine chloride (0.25 mg./kg.) was used. This dose was repeated if necessary, but the last dose was always given at least an hour before blood bypassed the liver, by which time the relaxation effect appeared to have worn off. Even this cautious regimen, however, led to postoperative apnea, apparently directly related to the use of tubarine chloride. This apnea was at least partially responsible for a number of the earlier deaths, and the use of relaxants was abandoned in the second half of the study. The anesthetic agents used were therefore halothane with nitrous oxide and oxygen. Initial fears that sufficient immobilization and relaxation for major abdominal surgery would not be attained without using a concentration of halothane that a marked fall in arterial would produce pressure have been proved groundless. When the “Fluotec” setting was at 2% the mean arterial blood pressure was within normal limits for pigs of the size used in this series [6]. There is, however, evidence that the percentage of halothane is lower in a Palmer than the circle absorber system, Pump “Fluotec” setting would indicate [lo], so that the actual percentage of halothane delivered is unknown. The possibility of halothane’s being toxic to the liver has been considered, but evidence of the type of cellular damage associated with halogenated agents, was not found in postmortem and biopsy specimens in this series [ 121. To facilitate abdominal closure at the end of the procedure, the concentration of halothane is increased. Relaxants are unnecessary, 154

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although from 80 to 160 mg. of gallamine triethiodide was used in some of the earlier experiments. Donor

Liver

Preparation

The donor animal is maintained in the supine position by securing its legs to the operating table. The preparation of the abdominal wall is similar to that in the recipient animal. The left femoral artery is cannulated and connected to a mercury manometer to enable continuous arterial blood pressure measurements to be made. The abdomen is entered through a longitudinal midline or right extending from the paramedian incision, xiphisternum to below the umbilicus. In the male a ligature is applied to the urethra to prevent urine from contaminating the operative field. The edges of the incision are widely retracted. A biopsy is taken from the edge of the right central or left central lobe of the liver to act as a control for subsequent histological studies. The first part of the dissection is performed with the bowel retracted to the animal’s left (Fig. 2A). The inferior vena cava is identified where it enters the lower surface of the liver, and it is cleared of adventitia down to the level of the adrenal gland. If the dissection is carried too far down, the venous drainage of the closely adherent right adrenal gland will be damaged, with ensuing troublesome bleeding. The portal vein is identified in the lesser omentum and it is dissected free of its covering peritoneum and lymphatic tissue as far as the pancreas. All of its branches, with the exception of the splenic vein, are ligated and divided. If the splenic vein is left undisturbed until late in the procedure, the blood pressure remains more stable, and the spleen does not become congested and friable. For reasons that will be discussed later it is important to avoid occluding the lumen of the portal vein during this dissection, as this is always accompanied by a fall in blood pressure. The common bile duct is ligated and divided distal to the point of entry of the cystic duct. At times the cystic duct runs parallel to the common hepatic duct for a

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Fie. 2. Donor liver nrenaration. (A) \ , Bowel retracted to the animal’s left for the first nart of the di&ection. (B) MobiizaAon of the thoracic aorta. (C) The liver after its removal from the donor animal. I

distance before joining it to form the common bile duct. Immediately after this the gallbladder is opened at its apex (the site of the subsequent cholecystoduodenostomy ). It is emptied of bile and the lumen is washed out with saline to prevent autodigestion by bile. It is allowed to drain freely throughout the rest of the operation. The lesser omenturn is divided close to its attachment to the stomach and duodenum, thereby dividing the branches of the celiac and hepatic arteries to these viscera. Great care is taken to avoid damage to the main hepatic artery or to any of the small hepatic branches which arise some distance from the liver. If one of the branches is inadvertently divided, infarction of an area of liver tissue ensues. The proximal end of the main hepatic artery is freed from the ensheathing nerve plexus and dissected down to its origin from the celiac artery. As it is difficult to complete the dissection from this approach, further dissection is performed from the animal’s left side. The upper end of the abdominal incision is continued across the costal margin and down the left side of the thorax in the eighth intercostal space. The diaphragm is divided radially from this incision down to the aorta. The lung and abdominal contents are re-

tracted to the animal’s right (Fig. 2B ). The aorta is mobilized from the arch to just below the celiac axis and its intercostal branches are ligated and divided. This is facilitated by the fact that they usually arise as a single posterior midline trunk, before bifurcating. The dissection of the celiac axis and hepatic artery is completed from this side by ligating and dividing the remaining branches. In all of the experiments except one in this series, the hepatic artery was left attached to a segment of thoracic aorta, similar to the segment of abdominal aorta used by Starzl

1201. The final stage of the preparation of the donor liver is performed from the animal’s right side. The abdominal contents are retracted to the left. The splenic vein is ligated and divided. To prevent thrombosis in the hepatic artery, 50 mg. of heparin is administered. A flask containing 4 liters of sterile Ringer lactate solution at about 4°C. is set up and connected to g-inch bore polyvinylchloride (P.V.C. ) tubing,* which is carefully filled to exclude all air bubbles. The portal vein is ligated close to the pancreas and the P.V.C. tubing is inserted into the portal vein through a transverse venot*Velbex Beverage Ltd.,

Grantham

Works,

Hose. Supplied Manningtree,

by B. X. Plastics Essex, England.

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omy and secured with a double ligature. It is advantageous to leave the portal vein attached at this stage, as this helps to hold the tubing in place during the subsequent dissection. The hepatic artery is clamped and the Ringer lactate solution is allowed to run into the liver at a rate of 1 to 1% liters per minute. This provides rapid cooling and flushes the liver of its contained blood. The liver soon becomes a pallid color. After cross-clamping, the aorta is divided distal to the arch and is ligated and divided distal to the origin of the celiac trunk, which leaves the hepatic artery free and attached to a length of thoracic aorta ( Fig. 2B ) . To ensure that no small branches have been left unligated, the arterial segment is tested with heparinized saline under pressure, The lower inferior vena cava is transected immediately above the right adrenal gland, and the are divided. The portal hepatic ligaments vein is freed by completing its division at the site of entry of the P.V.C. tubing. Finally the liver is removed by transecting the proximal end of the inferior vena cava flush with the undersurface of the diaphragm. The liver is placed in a sterile receptacle and perfused with another 4 liters of chilled Ringer lactate solution through the portal vein (Fig. 2C). The effluent from the vena cava fills the receptacle and covers the liver. The aim at this stage is to have the temperature of the liver below 12°C. to help protect it prior to its insertion into the recipient animal. Recipient

Operation

The recipient animal is prepared to receive the donor liver, Hepatectomy is performed, which necessitates occlusion of portal vein and inferior vena caval blood flow. In the pig it has been shown that occlusion of the portal vein for periods exceeding four minutes leads to a profound fall in blood pressure [23]. To overcome this, portal-to-superiorvena-cava and inferior-vena-cava-to-superiorvena-cava bypass systems are instituted (Fig. 4A) [17, 221. After induction of anesthesia the recipient animal is placed in the supine position and 156

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maintained by securing all four limbs to the operating table with stout tapes, the limbs being protected from damage by foam rubber pads. The neck, abdomen, and groin are shaved. The electrocardiogram leads are attached and thermometer leads are inserted into the rectum and midesophagus (Fig. 4AI). The operative field is prepared with Hibitane solution and the animal is draped with sterile disposable paper towels. The first step in the preparation of the recipient animal is the exposure of groin and neck vessels. A longitudinal incision is made in the left groin and the femoral artery exposed. A cannula is inserted to record arterial blood pressure (Fig. 4AI). The external jugular veins are exposed through bilateral longitudinal cervical incisions and the adventitia removed for 3 inches above the subclavian veins. Side branches are ligated and divided. The veins are exposed at this early stage to ensure that they are large enough to accommodate the bypass cannulas. On four occasions it was necessary to reject a pig because one of the external jugular veins was abnormally small, To record central venous pressure, a cannula is inserted into the vena cava via a right subclavian vein (Fig. 4AI). The next step is the preparation of the iliac and splenic veins, the preparation of the lower aorta, and the early mobilization of the animal’s liver prior to its removal. A longitudinal midline abdominal incision is made from the xiphisternum to the pubis. In males the inferior half of the incision is a right paramedian to avoid the penis and urethra. The wound edges are widely retracted. The abdominal dissection is commenced in the lower abdomen and continued up to the liver, and excessive bowel handling or evisceration is avoided to prevent a fall in blood pressure. The right external iliac vein is identified and cleared of adventitia. When its caliber is excessively small, the dissection is continued proximally to expose the common iliac vein, and the internal iliac vein is then ligated and divided. The lower abdominal aorta is identified and an inchlong segment is exposed just above the bi-

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furcation to prepare for the arterial anasOne or two lumbar arteries tomosis. occasionally require ligation but this can often be avoided. The abdominal aorta of the pig has a tough adventitia and the dissection is carried out in an extraadventitial plane. Care is taken to avoid damage to the large channels. The mobile adherent lymphatic spleen is delivered over the left-hand edge of the wound, and the main splenic vein is dissected to the pancreas, dividing and ligating some of the splenic branches. The bowel is retracted to the animal’s left side and the inferior vena cava is cleared below the liver as in the donor animal. Only a short segment of this vessel is available above the renal veins, as it is ensheathed in a downward prolongation of liver tissue. The dissection of the porta hepatis is commenced by clearing a short segment of portal vein about half an inch below the liver. The common and the bile duct is ligated and divided anterior peritoneum of the lesser omentum is incised. The animal is given 50 mg. of heparin. This, together with the heparin used in the transfused blood, raises the total heparin dosage to about 2 mg. per kilogram In the earlier transplants of body weight. no heparin was given because of the fear of hemorrhage. However, the bypass lines tended to thrombose, and when this occurred in the vital portal bypass, it frequently led to the death of the animal. Protamine sulphate was used at the end of some experiments to reverse the heparin, but this has been discontinued, as it was found to be unnecessary [2], and it was our impression that it increased capillary hemorrhage. During the preliminary dissection the vena caval and portal bypass lines are prepared. A simple veno-venous bypass without ( Fig. 4AI). Flexible a pump is adequate P.V.C. tubing (internal diameter g in. ) is used for the bypass lines and to each end are attached Portex cannulas* (7/32 in. or H in. internal diameter) by means of stainlesssteel reducing connectors. Each connector used at the cervical end has a side arm which *Right arterial cannulas. Supplied Plastics Ltd., Hythe, Kent, England.

hy Portland

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Fig. 3. Recipient animal preparation. (A) Insertion of the bypass cannula into the external jugular vein. (B) I nsertion of the bypass cannula into the splenic vein. (C) The liver bed. is attached to a venous pressure manometer and a bottle of heparinized normal saline (containing 5 mg. of heparin per liter). The bypass lines are carefully filled with saline to remove all air bubbles. The cervical cannula of the vena caval bypass is inserted into the right external jugular vein, and that of the portal bypass into the left external jugular vein (Fig. 3A). The lower end of the vein is occluded with a vascular clamp, and the cephalad end is ligated, The lubricated cannula is inserted through a longitudinal venotomy, maintaining a flow ‘of saline to prevent air embolism. The clamp is removed and the cannula is advanced into the anterior cava and secured with a double ligature. To prevent thrombi from forming at the tip of the cannula, a slow infusion of saline is continued. The jugular vein is friable and easily damaged; therefore, if any difficulty is experienced in passing the larger cannula, it is immediately replaced by a smaller one. The other end of the portal bypass is inserted into the splenic vein using a similar technique ( Fig. 3B ) . The tip of the cannula is carefully positioned behind the tail of the pancreas, but it is not advanced too far; otherwise it partially obstructs the portal flow. The lower end of the vena caval bypass is similarly inserted into the right 157

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The recipient animal. (AI) The donor liver in the orthotopic position. (AII) Vascular anascomplete. Portal and vena caval bypass lines in situ. (B) Cholecystoduodenostomy.

iliac vein (Fig. 4AII). Portal and vena caval bypass flows are commenced, and the portal vein and vena cava are occluded below the liver with vascular clamps. The adequacy of the important portal bypass is assessedby the line pressure reading and the color of the bowel. The liver is rapidly removed, The lesser omentum is ligated and divided close to the liver, thus preserving the arterial supply to the lesser curvature of the stomach as far as possible. The inferior vena cava is occluded above the liver with two clamps.* They are applied so as to include a portion of the diaphragm, thereby leaving a segment of vena cava for subsequent anastomoses (Fig. 3C). The vena cava is divided below these clamps, The portal vein and lower inferior vena cava are transected and the remaining peritoneal attachments of the liver divided. The liver is finally removed. The cooled donor liver is placed in the orthotopic position. The vascular anastomoses are rapidly performed using a continuous over-and-over technique with 0000 atrau-

matic black silk or Tavdekf sutures (Fig. 4AII). To help secure the liver in position and prevent subsequent kinking, the vein lengths are kept as short as possible. The upper vena caval anastomosis is performed first. No clamp is employed on the donor liver side to enable the anastomosis to be performed flush with the undersurface of the diaphragm. The posterior layer is anastomosed from within the lumen, and a second posterior layer is sometimes inserted to ensure hemostasis in this site which is inaccessible later in the procedure. Prior to completing the anterior layer of this anastomosis, the lumen is filled with heparinized saline to exclude air bubbles. The portal vein is anastomosed next. Interrupted sutures are employed for one-third of the anterior layer to allow for subsequent growth. Then the posterior layer of the inferior vena cava below the liver is anastomosed, leaving the anterior layer open. The recipient animal’s lower abdominal aorta is occluded with vascular clamps and an anterior longitudinal arteriotomy performed. Hepatic artery cir-

*Potts-Satinsky Vena Cava Clamps. Supplied Down Brothers and Meyer and Phelps Ltd., Cavendish Street, London W.l, England.

plied by England.

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fManufactured Charles

by Deknatel, Thackray Ltd.,

New York City. SupPark

Street,

Leeds,

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culation is reconstituted by an end-to-side donor-aorta to recipient-aorta anastomosis. This form of arterial reconstruction was used in all but one of the experiments, when a direct hepatic-artery to hepatic-artery anastomosis was performed. The portal vein and aortic clamps are removed, and about 500 ml. of blood is allowed to flush through the liver and discharge through the opening in the anterior wall of the lower inferior vena caval anastomosis. The aim is to clear the liver of acid metabolites that accumulate during the ischemic period, and since this step has been included, operative survival has improved. The donor vena cava below the liver is occluded. The Potts-Satinsky clamps are removed from the upper vena cava, thereby allowing portal and arterial blood to flow through the liver and return to the heart. The lower vena caval anastomosis is rapidly completed, and normal vena caval flow is recommenced. The portal and vena caval bypass lines are removed, and all the vessels are ligated. Finally biliary drainage is established by a cholecystoduodenostomy using a continuous single layer of 00 chromic catgut (Fig. 4B ) . No problems have arisen in relation to this anastomosis. Careful attention is paid to hemostasis. The abdomen is closed in layers with continuous braided nylon. The cannula in the right subclavian vein is brought out behind the ear through a subcutaneous tunnel to enable intravenous fluids to be administered and postoperative blood samples to be obtained. The neck incisions are closed. The arterial pressure cannula is removed from the left femoral artery, which is ligated, and the wound is closed.

Postoperatiue

Care

Anesthesia is discontinued and the pig is returned to a warmed recovery cage with the endotracheal tube still in place. This is removed later when movements commence. The animal is nursed on its side and turned at intervals. In the immediate postoperative period regular venous blood samples are obtained for acid base and glucose determinations, and sodium bicarbonate or glucose

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is administered if necessary. Further samples are taken during the first 12 hours, and subsequently at daily intervals for coagulation and liver function studies. No fluid is administered for the first 12 hours; then a liter of intravenous 4.8% dextrose in l/S normal saline is administered daily for the succeeding two days and banked titrated blood is administered if the pig becomes anemic, This was necessary in a number of cases because of the high incidence of bleeding esophagogastric ulcers [ 18, 251. Eating usually commences by the second or third postoperative day. Intramuscular penicillin and streptomycin are administered daily for a week. SUMMARY A technique for orthotopic liver homotransplantation in the pig is presented, describing the method of obtaining blood for transfusion; the anesthetic technique, with special reference to endotracheal intubation; and the preparation of the donor liver. The operation on the recipient animal is presented in detail and some aspects of postoperative care are mentioned, It is felt that this may be of value to future workers in this field contemplating the use of the pig. ACKNOWLEDGMENTS We wish to thank the many people who have assisted us, particularly Dr. E. J, Tierris, British Council Fellow in Surgery, Mr. McCrea, B.V.Sc., M.R.C.V.S., Senior Lecturer in Animal Husbandry, and the technicians of the University of Bristol Medical School and Department of Surgery. We are grateful to the Department of Medical Illustration, University of Bristol, for the illustrations. We acknowledge some of the suture material from Ethicon Ltd. REFERENCES 1.

Bendixen, H. H., Headley-White, J., and Laver, M. B. Impaired oxygenation in surgical patients during general anesthesia with controlled ventilation. New Eng. J. Med. 269:991, 1963.

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Blecher, T. E., Terblanche, J., and Peacock, J. H. Coagulation and haematological changes following orthotopic liver homotransplantation in the pig. Arch. Surg. In Press. Brook, G. B. Spinal (epidural) anaesthesia in domestic animals. Vet. Rec. 15:631, 1935. Couch, N. P. Supply and demand in kidney and liver transplantation: A statistical survey. Transplantation 4 : 587, 1966. Dawson, J. B. Anaesthesia for the experimental pig. Brit. J. Anaesth. 35:736, 1963. Engelhardt, W. V. Swine Cardiovascular Physiology: A Review. In L. K. Bustard and R. 0. McClellan (Eds.), Swine in Biochemical Research. Washington: United States Atomic Energy Commission, 1966. P. 307. Finley, T. N., Lenfant, C., Haab, P., Piiper, J., and Rahn, H. Venous admixture in the pulcirculation of anaesthetized dogs. J. monary Appl. Physiol. 15:418, 1960. Fowler, J., Hill, D. W., Morgan, R. L., Nunn, J. F., Weaver, B., and Woolmer, R. F. Anaesthesia for the irradiated pig: A study in remote control. Brit. J. Anaesth. 34:327, 1962. Goodrich, E. O., Welch, H. F., Nelson, J. A., Beecher, T. S., and Welch, C. S. Homotransplantation of the canine liver. Surgery 39:244, 1956. Hallen, B., Westermark, L., and Wahlin, A. The concentration of halothane with the combined use of Fluotec vaporizer and Palmer ventilator for animal experimentation. Acta Anaesth. Scud. 9:65, 1965. Hopcroft, S. C. Extradural anaesthesia in the pig. Brit. J. Anaesth. 37:982, 1965. Hunt, A. C. Pathology of liver transplantation in the pig. In A. E. Read (Ed.), Co&on Papers. Vol. XIX. The Liver. London: Butterworth, 1967. P. 337. Jaffe, W. P., Symes, M. O., and Terblanche, J. Observations on the immunological reactions of pigs. In A. E. Read (Ed.), Colston Papers, Vol. XIX. The Liver. London: Butterworth, 1967. P. 351. Loken, H. F., Havel, R. J., Gordon, G. S., and Whittington, S. L. Ultracentrifugal analysis of protein-bound and free calcium in human serum. J. Biol. Chem. 235:3654, 1960. Moore, F. D., Birtch, A. G., Dagher, F., Veith, F., Krisher, J. A., Order, S. E., Shucart, W. A., Dammin, G. J., and Couch, N. P. Immunosuppression and vascular insufficiency in liver transplantation. Ann. N.Y. Acad. Sci. 120:729, 1964: Moore, F. D., Smith, L. L., Burnap, T. K., Dal-

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