Ischemic preconditioning of cadaver donor livers protects allografts following transplantation

BACKGROUND: Ischemic preconditioning (IP) has been shown in animal models to protect livers against ischemia/reperfusion injury. The aim of this clinical study is to investigate whether IP of cadaver livers prior to retrieval confers protection on the allografts. METHODS: Cadaveric donor livers were subjected to IP prior to retrieval by clamping of the hepatic pedicle for 10 min followed by reperfusion. Biopsies were obtained from the preconditioned (n=9) and control nonpreconditioned (n=14) liver transplants prior to and 2 hr following reperfusion. Cryosections were stained with antibodies against neutrophils and platelets. RESULTS: IP livers were associated with significantly lower serum levels of aspartate aminotransferase (240+/-98 IU/L vs. 382+/-163 IU/L; P>0.016) and lactate (0.81+/-0.07 mmol/L vs. 1.58+/-0.9 mmol/L; P>0.018) 24 hr following transplantation. Furthermore, recipients of IP livers spent a significantly shorter time in the intensive care unit following transplantation compared to those given nonpreconditioned allografts (1 vs. 2.8+/-1.6 days; P=0.0008). Increases in neutrophil infiltration were detected in 6/14 (43%; P=0.022) and in CD41 deposition in 5/14 (36%; P=0.042) of nonpreconditioned livers. However, none of the IP allografts showed any change in the levels of platelets or neutrophil infiltration following transplantation. CONCLUSION: IP is an effective method of protecting cadaver donor allografts from cold ischemia and subsequent reperfusion injury. IP is also associated with a reduction in the nonspecific inflammatory response.     

Utilization of Activated Carbon Hemoperfusion to Assist Recovery of Ischemically Damaged Canine Liver Allografts

The sensitivity of liver allografts to even minimal periods of ischemia currently limits the duration of hepatic preservation prior to liver transplantation. This study evaluates the role of activated carbon hemoperfusion (ACH) for assisting the recovery of canine livers ischemically damaged by a 20-min occlusion of the portal vein and hepatic artery prior to organ harvesting. Animals in Group I (n = 5) receiving damaged liver allografts without ACH survived a mean (+/- SD) of 18.0 +/- 13.5 h. One ACH treatment given to recipients immediately after liver transplantation in Group II (n = 5) resulted in improved survival to a mean of 3.8 +/- 2.16 days (p less than 0.05). The best survival was obtained after three ACH treatments in Group III (n = 6) on days 0, 1, and 2 (26.6 +/- 27.1 days) (p less than 0.05). These results indicate that ACH may be helpful in assisting the recovery of ischemically damaged liver allografts after transplantation.     

Initial hepatic metabolic function in canine liver and pancreas cluster transplantation

In this study, initial hepatic metabolic function was evaluated by determining the arterial ketone body ratio (AKBR) and plasma amino acid concentrations in an experimental orthotopic combined hepatopancreatic transplantation (OHPT), and comparing the same values in orthotopic liver transplantation (OLT). In OHPT, AKBR decreased in the anhepatic phase and recovered to the preoperative value just 1 h after reperfusion. On the other hand, in OLT, the recovery of AKBR took 3 h after reperfusion with a significant difference compared to OHPT (P<0.05). Plasma amino acid levels, especially alanine and total free plasma amino acids increased in the anhepatic phase and recovered within 1 h of reperfusion in OHPT. However, they did not recover until 3 h after reperfusion in OLT. This rapid recovery of hepatic metabolic function in OHPT should be attributed to the order of reperfusion in which the reconstruction of arterial blood flow precedes that of portal blood flow. This model is useful for assessing the best way by which the grafted liver can control the timing, order, rate, and volume of blood that should be released.     

Metabolism during hepatic transplantation: indicators of allograft function

In an attempt to determine the initial function of hepatic allografts, several metabolic indicators of hepatic function were studied intraoperatively in 12 cases of hepatic transplantation. The operation was divided into three sampling periods: baseline, anhepatic, and reperfusion. During the baseline period plasma lactate levels rose at 2.6 mmol/L/hr and continued to rise at a similar rate during the anhepatic period. Baseline period total free plasma amino acid levels (TFPAA) rose at a moderate rate of 0.4 mmol/L/hr. During the anhepatic period TFPAA levels rose at a fivefold greater rate than during baseline (p less than 0.01). The ability of the hepatic allograft to reduce abnormal levels of TFPAA and lactate during the reperfusion period was associated with reduced morbidity in the first 48 hours after transplantation. Intraoperative clearance of accumulated TFPAA is currently the best means of assessing initial allograft function. Elevated preoperative total serum bilirubin levels were also associated with increased early morbidity in hepatic transplant recipients.     

Activation of intracellular neutrophil elastase in the transplantation of ischemic liver.

Ischemia-reperfusion injury is an important cause of primary nonfunction of transplanted organs, and neutrophil elastase has been implicated in the pathophysiology of ischemia-reperfusion injury. We assessed the kinetics of intracellular neutrophil elastase (INE) activity in canine liver transplantation. Mongrel dogs underwent orthotopic whole-liver transplantation. The animals in group I (n = 6) received fresh liver grafts, and all of the dogs survived longer than 24 h. The animals in group II (n = 5) received liver grafts injured by 30 min of warm ischemia. Only 1 animal survived longer than 24 h after reperfusion. A significant increase in the serum ALT and LDH levels was observed in group II after reperfusion of the graft. Isolated peripheral neutrophils were homogenized, and the neutrophil elastase activity in the supernatant was determined by using a spectrophotometric assay. The INE activity was expressed as the neutrophil elastase value per 1 x 10(10) peripheral neutrophils. In group I, the INE activity 10 min and 2 h after reperfusion was 7.6 +/- 2.6 and 6.1 +/- 2.4 U, respectively. In group II, this activity was 25.9 +/- 7.4 and 44.3 +/- 23.7 U, respectively. There was a significant correlation between serum LDH levels and INE activity 10 min after reperfusion (gamma = 0.70, p < 0.02). In conclusion, the INE activity increased more sharply after the reperfusion of ischemically injured liver grafts. The INE activity correlates with serum LDH levels immediately after reperfusion, suggesting that the increase in the INE activity depends on the severity of ischemic damage.     

Ischaemic preconditioning of the liver before transplantation

PURPOSE: Assessment of the effect of a short ischaemic time prior to liver transplantation on the liver graft. METHODS: White X Landrace pigs (N=10) were subjected to liver transplantation. Before being removed from the donor animal, the livers were randomised into two groups: group 1--pre-procurement ischaemia (15 minutes' temporary arrest of portal venous and hepatic arterial inflow to the liver, followed by reperfusion of these vessels for a period of 15 minutes); group 2--no prior inflow occlusion (control group). In group 1 a spleno-jugular bypass was established to prevent venous congestion, portal venous hypertension, intestinal oedema and bacterial translocation. The livers were perfused with Eurocollins solution (4 degrees C), after which they were stored on ice for a period of 3 hours' cold ischaemic time. Hepatocellular injury was assessed according to liver cell function tests (aspartate aminotransferase, AST), biochemical indicators of reperfusion injury (malondialdehyde) and histopathology. RESULTS: There was a significant rise of AST in both groups 1 hour after transplantation (from 51 +/- 27 IU/l to 357 +/- 152 IU/l in group 1 and from 29 +/- 10 IU/l to 359 +/- 198 IU/l in group 2). AST levels were marginally lower in group 1 at 2 and 4 hours after transplantation. There was also a rise in malondialdehyde levels in both groups at 5, 20, 40 and 60 minutes after transplantation. Levels of malondialdehyde were lower in the primed group at 5, 20 and 40 minutes, while the levels at 60 minutes after transplantation were comparable. Histological changes, as measured by vacuolisation, neutrophil infiltration and hepatic cell necrosis, were less in livers transplanted after ischaemic preconditioning, although the difference was not significant. CONCLUSIONS: Ischaemic preconditioning of the donor liver seems to decrease hepatocellular damage, reperfusion injury and histological changes in the liver after transplantation. Further studies with larger numbers are indicated.     

High preoperative recipient plasma 7beta-hydroxycholesterol is associated with initial poor graft function after liver transplantation.

Oxidative stress is implicated in the pathogenesis of hepatic ischemia-reperfusion injury, a major determinant of initial poor graft function (IPGF) after orthotopic liver transplantation (OLT). We prospectively investigated the association between the recipient plasma preoperative oxidative stress and the occurrence of IPGF after deceased-donor OLT and indirectly studied the source-hepatic or extra-hepatic-of systemic oxidative stress in vivo in cirrhosis. We used a recently developed specific and sensitive mass spectrometry assay to measure 7beta-hydroxycholesterol and 7-ketocholesterol (oxysterols), markers of oxidative stress, in biological matrices. At univariate analysis, preoperative recipient 7beta-hydroxycholesterol plasma concentration was significantly higher in transplants with subsequent IPGF (n = 9) compared with those with initial good graft function (IGGF; n = 23) [mean +/- SD: 30.63 +/- 26.42 and 11.57 +/- 15.76 ng/mL, respectively] (P = 0.017). In a logistic regression model, which included also the Model for End-Stage Liver Disease (MELD) score, 7beta-hydroxycholesterol plasma concentration was an independent predictor of IPGF with an odds ratio of 1.17 (95% CI, 1.02-1.33, P = 0.028). Patients with cirrhosis (n = 32) had increased oxysterol plasma levels compared with healthy controls (n = 49); livers with cirrhosis (n = 21), however, had oxysterol content comparable with normal livers obtained from organ donors (n = 19). Oxysterols persisted elevated in plasma 1 month after OLT (n = 23). In conclusion, cirrhosis presents upregulated systemic oxidative stress likely of extrahepatic source that is associated with graft failure after OLT.     

Extension of ischemic tolerance of porcine livers by cold preservation including postconditioning with gaseous oxygen

BACKGROUND: Transplantation of organs from nonheartbeating donors was recommended to reduce organ shortage. In vitro experiments with rat livers have shown that the warm ischemic tolerance of the liver may be extended by persufflation with gaseous oxygen during cold storage. The qualification of this method for procurement of livers harvested after cardiac arrest was tested in an in vivo approach with pigs. METHODS: Livers from 15 donor pigs were explanted, heparinized, flushed with and stored in University of Wisconsin solution for 4 hr at 4 degrees C, and then implanted into 15 recipients. The organs were dissected immediately after cardiac arrest (group 1) or after 60 min of warm ischemia (groups 2 and 3). Group 2 livers received 75,000 IU of superoxide dismutase together with the flush solution and were persufflated with gaseous oxygen via the venous vascular system during cold storage. Main end point was survival after 5 days. Additionally, metabolic, functional and inflammatory criteria were measured in the blood. RESULTS: All animals of the groups 1 and 2 survived, all animals of group 3 died within 3 hr after reperfusion. In all groups the blood parameters reflected significant damage of the livers. However, the ischemic damage was comparable in the groups 1 and 2 whereas the livers of group 3 exhibited significantly higher levels of aspartate alanine aminotransferase and lactate dehydrogenase, and a significantly elongated partial thromboplastin time 1 hr after reperfusion (P=0.016). CONCLUSIONS: Venous systemic oxygen persufflation in combination with antioxidative medication is a promising new method of resuscitating ischemically altered livers from nonheartbeating donors for successful transplantation.     

The impact of arterialization on hepatic microcirculation and leukocyte accumulation after liver transplantation in the rat.

This study investigated the influence of hepatic arterialization on early graft function, microcirculation, and leukocyte-endothelial interaction after syngeneic orthotopic liver transplantation in Lewis rats. Livers were preserved for 17 hr in UW solution and transplanted without rearterialization (group 1: n = 10) or with immediate arterial reconstruction (group 2: n = 10). Graft function was analyzed by bile flow; microcirculation was assessed by laser Doppler flowmetry (LDF) and intravital microscopy (IVM). In addition, flow behavior of leukocytes was quantified by IVM after i.v. injection of the WBC marker acridine orange. Improved graft function in group 2 was indicated by increased bile production during the observation period of 90 min after reperfusion (7.18 +/- 0.62 vs. 3.63 +/- 0.63 ml/100 g liver [mean +/- SEM] P < 0.001). In arterialized grafts LDF values increased by 22.9 +/- 3.8% upon reperfusion of the hepatic artery (P = 0.004). Arterialization increased WBC velocities in sinusoids (group 1: 0.29 +/- 0.02 mm/sec, group 2: 0.34 +/- 0.01 mm/sec, P < 0.001) and postsinusoidal venules (0.43 +/- 0.05 vs. 0.64 +/- 0.05 mm/sec, P = 0.029). In addition, the number of nonperfused midzonal sinusoids decreased significantly (8.5 +/- 2.2% of all sinusoids analyzed vs. 4.2 +/- 1.3%, P = 0.048). However, the marked sinusoidal and venular WBC adherence observed 1 hr after reperfusion was not altered by arterialization. It is concluded that arterial reconstruction in rat liver transplantation improves microvascular perfusion and graft function but this improvement does not relate to WBC accumulation within the graft. We propose that studies on hepatic preservation and postischemic reperfusion in the rat should be based on the physiological model of dual vascularization.     

Amino acid clearance in cirrhosis. A predictor of postoperative morbidity and mortality

The central plasma clearance rate of amino acids (CPCR-AA), the ratio of peripheral amino acid entry rate into blood plasma to arterial amino acid concentration, was measured preoperatively in 149 noninfected cirrhotic patients. In 50 survivors of shunting or general surgical procedures, the mean (+/- SEM) CPCR-AA was 201 +/- 17 mL/m2/min; in 39 subsequent deaths, the mean ratio was 87 +/- 14 mL/m2/min. Comparing Child's classification with CPCR-AA reveals the following values: class A (mortality, two of ten patients) survivors, 152 +/- 23 mL/m2/min; class A deaths, 96 +/- 54 mL/m2/min; class C (mortality, 13 of 19 patients) survivors, 214 +/- 47 mL/m2/min; class C deaths, 101 +/- 13 mL/m2/min. The preoperative CPCR-AA of 46 patients receiving liver transplants was 91 +/- 9 mL/m2/min; 69% of these patients survived. Preoperative CPCR-AA values correlated significantly with rates of hepatic protein synthesis in incubated liver slices obtained by biopsy at operation in 22 patients. Thus, CPCR-AA determination is a true liver function test, valuable in predicting surgical mortality and selecting transplantation or other operations for cirrhotic patients.     

Cobalt-protoporphyrin induced heme oxygenase overexpression and its impact on liver regeneration

Cobalt-protoporphyrin (CoPP)-dependent induction of heme oxygenase (HO)-1 has been shown to protect from ischemia-reperfusion injury, which remains a major source of graft loss after liver transplantation. The impact of HO-1 on liver regeneration, especially in reduced-size grafts, has not yet been evaluated. Using an experimental model, we investigated HO-1 induction by CoPP treatment on postoperative recovery of ischemically injured livers following partial (70%) hepatectomy. Wistar rats underwent partial hepatectomy under temporary inflow occlusion (30 minutes). One group of animals received CoPP (5 mg/kg body weight i.p.) 24 hours prior to surgery to induce high levels of HO-1 at the time of surgery, and the second group served as nontreated controls. At postoperative days 1, 4, 7, and 10, animals were exsanguinated, and blood and liver samples were stored for enzymatic (serum AST and ALT levels) and histologic (mitotic index) analyses (n = 5 each day). Additionally, postoperative body weight and weight of the remnant liver were measured. Although serum AST and ALT levels as well as remnant liver weight were comparable between both groups, CoPP-treated animals recovered from surgery more quickly as indicated by postoperative body weight. Moreover, the number of mitotic cells was significantly increased in this group at day 1 (33 +/- 5 versus 20 +/- 5 per 2000 hepatocytes) as compared with nontreated animals. Liver regeneration of ischemically injured livers following partial hepatectomy was improved by HO-1 overexpression following preoperative CoPP administration. Thus, it is conceivable that prevention of ischemia-reperfusion injury by HO-1 overexpression also might be beneficial for reduced-size liver grafts without affecting their proliferative capacity.     

Impact of endothelin-1 on microcirculatory disturbance after partial hepatectomy under ischemia/reperfusion in thioacetamide-induced cirrhotic rats

BACKGROUND: Endothelin (ET)-1 contributes to hepatic ischemia and reperfusion (HIR) injury in normal liver. This study was conducted to clarify the role of ET-1 in HIR injury in cirrhotic state. MATERIALS AND METHODS: Using thioacetamide-induced cirrhotic rats with spontaneous portosystemic shunt, we determined the changes in plasma aspartate aminotransferase (AST) levels, plasma and hepatic ET-1 values, 7-day survival rates, and hepatic oxygen saturation (SO(2)) by time-resolved spectroscopy as an indicator of hepatic microcirculation under intermittent or continuous total hepatic ischemia with subsequent partial hepatectomy. RESULTS: Hepatic ET-1 levels in cirrhotic rats were significantly higher than those in noncirrhotic rats. Plasma and hepatic ET-1 levels at 1, 3 and 6 h of reperfusion after intermittent hepatic ischemia were significantly lower than those after continuous hepatic ischemia. In cirrhotic animals subjected to intermittent hepatic ischemia, the elevation of plasma AST levels at 1, 3 and 6 h of reperfusion and the decline in hepatic SO(2) at the end of 60-min hepatic ischemia and after reperfusion were significantly suppressed when compared with those subjected to continuous hepatic ischemia. Pretreatment with a nonselective endothelin receptor antagonist in continuous hepatic ischemia significantly ameliorated plasma AST levels and hepatic SO(2) values with less hepatic sinusoidal congestion, resulting in an improvement in the 7-day survival rate. CONCLUSIONS: Continuous hepatic ischemia in the cirrhotic liver has disadvantages relating to microcirculatory derangement with more ET-1 production in partial hepatectomy. In liver surgery, pharmacological regulation of ET-1 production may lead to attenuation of reperfusion injuries for ischemically damaged cirrhotic liver.     

The effects of fasting on the quality of liver preservation by simple cold storage

Although livers can be successfully preserved for 24 hr or more, often the transplanted livers have poor or no (primary nonfunction) function. The quality of the liver does not appear dependent upon the time of preservation but may be dependent upon the condition of the donor. In this study we have investigated the effects of fasting on the quality of livers for transplantation. Rabbits were fasted (48 hr) and livers preserved in the UW solution for 6-8 hr. Functions of the liver were analyzed by isolated perfusion for 2 hr. Also, pigs were fasted for 72 hr, livers preserved for 12 hr, and viability determined by orthotopic transplantation. Fasting depleted the liver glycogen by 85% but had no effect on ATP or glutathione concentrations. Rabbit livers from fasted animals produced similar amounts of bile, released similar concentrations of lactate dehydrogenase (LDH) and aspartate amino transaminase (AST) into the perfusate, maintained similar concentrations of ATP and glutathione in the tissue, and had a similar intracellular K:Na ratio after 24-hr preservation when compared to livers from fed animals. After 48-hr preservation, livers from fasted animals were less viable than livers from fed animals, including: reduced bile production (2.0 +/- 0.3 vs. 5.0 +/- 0.9 ml/2 hr, 100 g), greater release of LDH (3701 +/- 562 units vs. 1123 +/- 98 units) and AST, less ATP (0.326 +/- 74 vs. 0.802 +/- 160 nmol/g), less glutathione (0.303 +/- 13 vs. 0.933 +/- 137 nmol/g), and a lower K:Na ratio (1.5 +/- 0.9 vs. 7.4 +/- 0.6). Pigs receiving livers from fed animals preserved for 12 hr had better survival (5/6, 83%) than livers from fasted animals (3/6, 50%). The results show that the nutritional status of the donor can affect the outcome of liver preservation and transplantation. Increased injury in livers from fasted animals may be due to the loss of glycogen that may be an essential source of energy in the initial posttransplant period. In clinical liver transplantation the nutritional status of the donor may be an important factor in the initial function of the liver, and methods to increase the nutritional status of the donor may be important in increasing the quality of livers.     

Effects of exogenous endothelin-1 application on liver perfusion in native and transplanted porcine livers

PURPOSE: This study was designed to assess and differentiate the impact of progressivly increasing portal venous endothelin-1 (ET) plasma concentrations on hepatic micro- and macroperfusion of native porcine livers (Group A) and liver grafts after experimental transplantation (Group B). METHODS: A standardized gradual increment in systemic ET plasma concentration (0-58 pg/ml) was induced by continuous ET-1 infusion into the portal vein in both groups (A: n = 10, B: n = 10). Control animals received only saline (n = 5, each group). Hepatic microcirculation (HMC) was quantified by thermodiffusion electrodes, hepatic artery flow (HAF), and portal venous flow (PVF) by Doppler flowmetry. RESULTS: No changes in ET or perfusion parameters were observed in controls. The mean ET level after orthotopic liver transplantation (OLT) in Group B was elevated (baseline: 3.8 +/- 2.4 pg/ml) compared with Group A (2.8 +/- 1.9 pg/ml). With rising ET levels HAF decreased progressively in Group A from 205 +/- 97 (baseline) to 160 +/- 72 ml/min, and in Group B from 161 +/- 87 to 146 +/- 68 ml/min. PVF decreased in Group A from 722 +/- 253 to 370 +/- 198 ml/min, and in Group B from 846 +/- 263 to 417 +/- 203 ml/min. Baseline HMC in Group A was 86 +/- 15 and decreased significantly to 29 +/- 9 ml/100 g/min, and baseline MC in Group B was 90 +/- 22 and decreased to 44 +/- 32 ml/100 g/min. No significant alteration in systemic circulation was noted at the ET concentrations investigated. CONCLUSIONS: Significant impairment of hepatic micro- and macrocirculation was detected after induction of systemic ET levels above 9.4 pg/ml both in native and in transplanted livers. Disturbance of HMC was caused predominantly by reduction of portal venous flow, while the effect of ET on HAF was less pronounced. Characteristics of flow impairment in transplanted and native livers were analogous after short cold ischemic graft storage (6 h).     

Basal and meat extract plasma gastrin before and after parietal cell vagotomy and selective gastric vagotomy with drainage in patients with duodenal ulcer.

Basal and meat extract stimulated plasma gastrin (PG) levels and basal and stimulated gastric acid secretion were evaluated pre and postoperatively in duodenal ulcer patients who underwent parietal cell vagotomy without antral drainage (normal duodena) (PC, n=32) or selective vagotomy with drainage (pyloric stenosis) (SV +P, n=11). Before operation, both groups had comparable basal PG values of 52+/-13 pg/ml (PCVP) AND 51+/-18 PG/ML (SV+P), while the peak gastrin level to meat extract stimulation was 173+/-40 pg/ml for the total group of patients. After both operations basal PG levels increased (107+/-18 pg/ml (PCV) and 152+/-45 pg/ml (SV+P) and the gastrin response to meat extract stimulation was augmented after PCV, while the response after SV+P was the same as before operation. Patients with PCV often demonstrated an acid response following meat extract stimulation (3.6+/-0.9 mEq HC1/hr), and pentapeptide stimulation (18.8+/-2.0 mEq/hr) while patients with SV/P showed a minimal response (1.3+/-1.2 mEq HC1/hr meat extract), and 10.7+/-1.8 mEq/hr pentapeptide stimulation. The comparatively intact acid response in the PCV patients may augur a high ultimate recurrence rate.     

Protective effects of trifluoperazine on the microcirculation of cold-stored livers

Previous studies have shown a protective effect of trifluoperazine (TFP), a calmodulin inhibitor, upon the microcirculation of cold-stored kidneys. The present study points to similar beneficial effects of TFP on the microcirculation of cold-stored livers; 25 canine livers were preserved for 24 hr with Euro-Collins' solution (EC) (n = 8), University of Wisconsin solution (UW) (n = 7), or UW + TFP (n = 10). The stored livers underwent heterotopic transplantation (HLTX); hepatic-artery and portal-vein pressure and flow were monitored; oxygen consumption and extraction were measured before HLTX and at 15-min intervals after reperfusion, for 1 hr. Mean hepatic-artery and portal-vein flow (HAF & PVF) prior to donor hepatectomy were 172 and 530 cc/min, respectively. Poor HAF and PVF occurred in EC-HLTX (mean 35, 175 cc/min, respectively). The damaged EC-flushed livers could not compensate to the decreased hepatic blood flow by increased oxygen extraction (oxygen consumption and extraction, 8.7 vol.% and 48%, respectively). Light and electron microscopy showed severe liver necrosis and periportal hemorrhages. Improved hepatic-artery and portal-vein flows were seen in UW HLTX (105 and 254 cc/min), and oxygen consumption and extraction were 16.4 vol.% and 66%, respectively. Liver biopsy taken just before reperfusion revealed well-preserved liver architecture. Liver biopsy obtained 1 hr after reperfusion revealed marked edema of the portal triad, sinusoid congestion, and hemorrhage. Electron-microscopy biopsies obtained during reperfusion at 15-min intervals revealed severe vasospasm of the terminal hepatic arterioles and progressive damage to the liver microcirculation. The addition of TFP to the UW-flush solution resulted in excellent protection of the liver microcirculation. Marked increase in hepatic-artery and portal-vein blood flow was noted after reperfusion (mean 167 and 421 cc/min, respectively (P 0.02 vs. UW: P 0.001 vs. EC). The recovery of metabolic activity was evident by the high oxygen consumption and extraction (25.8 vol.% and 80%, respectively). And serial liver biopsies obtained after reperfusion have shown excellent protection of liver architecture and the absence of hepatic arteriolar vasospasm. Taken together, these data suggest that the addition of TFP to the UW solution protects the liver microcirculation by rendering the hepatic microcirculation insensitive to vasospastic stimuli during reperfusion, thus permitting better metabolic recovery after transplantation.     

Hemodynamic and biochemical changes during normothermic and hypothermic sanguinous perfusion of the porcine hepatic graft

Using an ex vivo liver sanguinous perfusion system, hemodynamic and biochemical changes of the porcine livers were studied, which were preserved cold (4 degrees C) for 24 hr in University of Wisconsin solution and reperfused with normothermic (37 degrees C) (n = 8) or hypothermic (32 degrees C) (n = 8) blood for 3 hr. Six more livers were reperfused with normothermic blood (37 degrees C) immediately after procurement as controls. The total hepatic blood flow was adjusted to 1 ml/min/g liver weight, in which hepatic artery and portal vein flows were administered at a 1:2 ratio. In livers stored cold for 24 hr in UW solution and perfused normothermically, a statistically higher hepatic artery resistance was exhibited at 30 and 60 min after reperfusion (P less than 0.05), and there was lower bile output (P less than 0.05) at 90 and 120 min as compared to the controls. In livers stored cold for 24 hr in UW solution and perfused hypothermically, as compared to ones perfused normothermically, statistically higher hepatic-artery and portal-vein resistances (P less than 0.05) were observed throughout the perfusion period and 60 min after reperfusion, respectively. In addition, bile output and oxygen consumption of these livers were statistically lower than those of ones perfused normothermically (P less than 0.05). In contrast, chemistries of the perfusate of livers perfused hypothermically were comparable to ones perfused normothermically. Histologic examination of the liver perfused hypothermically demonstrated hepatic arterial and/or portal venous congestion and mild-to-moderate hemorrhage in the portal triads. This study suggests that livers preserved for a prolonged period of time demonstrate a high hepatic arterial resistance shortly after revascularization, and that recipient hypothermia after revascularization may be a risk factor for the development of hepatic arterial thrombosis following liver transplantation.     

Bile acids in xenogeneic ex-vivo liver perfusion: function of xenoperfused livers and compatibility with human bile salts and porcine livers

BACKGROUND: In recent years, hepatic support systems using xenogeneic cells have been developed to support patients in fulminant hepatic failure. The extent to which xenogeneic hepatocytes metabolize and excrete human organic anions is unclear. In these studies we examined the ability of the ex vivo porcine liver to clear human bile acids during extracorporeal liver perfusion (ELP). METHODS: Four patients with fulminant hepatic failure underwent extracorporeal liver perfusion with 9 porcine livers. The venovenous circuit was designed as previously described (NEJM,1994,331:234) as were the immunologic features (Transplantation 1994,58:1162). Bile from the porcine liver and serum samples were collected hourly during perfusion. Three bile acids (glycocholic, glycodeoxycholic, taurodeoxycholic acid) were selected as markers for human bile and three (glycohyocholic, glycohyodeoxycholic, and glyco-3alpha-hydroxy-6-oxo-5beta-cholanoic acid) for markers of pig bile. Bile acids from both serum and bile were processed and analyzed through high performance liquid chromatography. The Students' t test was used for statistical analysis. RESULTS: The mean duration of perfusions was 4.1+/-1.5 hr. The mean total bile acid clearance from serum (243+/-44 micromol/h) was similar to the total bile acid biliary excretion (286+/-84 micromol/hr, P = 0.06). After 1 hr of perfusion, bile samples demonstrated a predominance of pig bile salts (65%). After 3 hr of perfusion, human bile acids made up 85% of total biliary bile acids. Pig bile acids appeared in patients' sera after 1 hr of perfusion, and after 3 hr, 35% of serum bile salts were pig-specific. CONCLUSIONS: Porcine livers perfused with human blood can clear the serum of potentially toxic human bile acids and excrete them into bile. Simultaneously, the percentage of pig-specific bile acids in patient serum increases during xenogeneic perfusion for unknown reasons. The relative hepatic uptake of bile acid from serum is similar to bile acid excretion in bile. Further development of systems using porcine livers or hepatocytes is warranted.     

Quantitation of transplanted hepatic mass necessary to cure the Gunn rat model of hyperbilirubinemia.

The minimal hepatic mass necessary to reverse the metabolic defect of unconjugated hyperbilirubinemia in the rat model of Crigler-Najjar type I deficiency was determined using heterotopic (auxiliary) partial liver transplantation (HLT) and orthotopic liver transplantation (OLT). In HLT, the donor graft consisted of the right upper and/or right lower hepatic lobe(s) depending on the final mass of liver tissue desired for transplantation. The mass of the donor graft ranged from 12% to 23% of the whole organ (n = 12). The serum unconjugated bilirubin levels decreased quickly after HLT from a preoperative value of 8.98 +/- 0.34 mg/dL to 0.63 +/- 0.11 mg/dL in 24 hours, which was similar to OLT in which the levels decreased from a preoperative value of 8.20 +/- 0.44 mg/dL to 0.24 +/- 0.07 mg/dL in 24 hours. Conjugated bilirubin was excreted from the graft liver shortly after OLT and also from both the host and graft livers after HLT. This study demonstrates that using as little as 12% of the whole liver mass in HLT reduces serum bilirubin significantly in 24 hours in a fashion similar to whole-organ OLT. The clinical application of alternative therapies to whole-organ OLT such as HLT or hepatocyte transplantation may provide sufficient replacement therapy in metabolic disease.     

N-acetylcysteine in pig liver transplantation from non-heart-beating donors.

Lipid peroxidation due to oxygen free radicals (OFR) seems to play a major role in loss of liver graft viability after warm ischemia, preservation, and transplantation. N-acetylcysteine (NAC) is an antioxidant that has a direct effect on OFR, and is also a glutathione precursor, another antioxidant. This study was designed to evaluate the efficacy of NAC in preventing ischemia-reperfusion damage of liver grafts harvested from non-heart-beating donors. Liver transplantation was performed on pigs divided into five groups: group 1 (control group; n=5) received livers from heart-beating donors; livers were subjected to 30 min of warm ischemia in groups 2 (n=3, no NAC) and group 3 (n=3; NAC treatment); warm ischemia time lasted 60 min in groups 4 (n=4; no NAC) and 5 (n=5; NAC treatment). Studied parameters included graft survival for more than 3 days, aspartate aminotransferase plasma levels, liver histology, and hepatic total glutathione concentrations. Graft survival was 100% in groups 1, 2, and 3, 0% in group 4, and 20% in group 5. NAC treatment did not influence initial mean aspartate aminotransferase release which was greater in warm ischemic livers than in controls. NAC treatment had no effect on liver hepatic total glutathione after reperfusion of animals receiving warm ischemic grants. Finally, no effect on liver histology was observed with NAC treatment. Our study suggests that in liver transplantation from non-heart-beating donors, NAC has no effect in both graft viability and lipid peroxidation. The role of OFR in primary dysfunction of transplanted warm ischemic livers remains controversial.