Hepatic Allograft Research Articles

The apoptosis protector, bcl-2 protein, is downregulated in bile duct epithelial cells of human liver allografts

Background/Aims: Apoptosis of bile duct cells occurs in hepatic allografts and is correlated with acute rejection. bcl-2 protein counteracts apoptosis and prolongs cell survival. We therefore tested the expression of bcl-2 protein in bile ducts of liver grafts in comparison with those of liver cirrhosis. Methods: 115 biopsies from 17 liver allografts and 47 biopsies of liver cirrhosis were analyzed and compared with 22 normal controls and with biopsies from patients with primary sclerosing cholangitis or primary biliary cirrhosis. bcl-2 protein and PCNA (proliferating cell nuclear antigen) expression was assessed using immunohistochemistry, and apoptosis was analyzed employing in situ DNA end-labeling. Results: A high apoptotic rate was detected in bile duct cells of allograft biopsies. In contrast to controls, bile duct cells of allografts and liver cirrhosis had high proliferative activity (mean PCNA labeling index: 1.0% vs. 40.7% and 18.6%, respectively). In liver grafts, bcl-2 protein positivity of bile duct and ductular cells was found in 3.6% and 4.4% of sections, respectively, and in cirrhosis in 44% and 79%, respectively (allografts vs. cirrhosis p<0.01). In controls, only one biopsy was bcl-2 positive. Conclusions: Whereas increased proliferative activity of small bile ducts and ductules in cirrhosis is associated with a high degree of bcl-2 expression, bile duct and ductular cells in liver grafts have a very low bcl-2 protein reactivity, even though their proliferative activity is high. These findings suggest that downregulation of bcl-2 expression in allograft bile duct cells might play a role in the increased apoptosis of these cells in acute rejection.

Split liver transplantation benefits the recipient of the ‘leftover liver’

The division of a single hepatic allograft to create two reduced-size grafts has been reported with decreased graft survival (50%) resulting in decreased enthusiasm for this approach. The authors reviewed their experience with 12 recipients of this procedure to evaluate the outcome of the children electively undergoing transplant with the “leftover liver.” A retrospective review of six pairs of children receiving part of one hepatic allograft included donor anatomy, recipient operation, and allograft and patient outcomes. Recipient pairs were selected according to blood type compatibility, medical priority, and size restrictions of the larger right lobe and the smaller left lateral segment. Patient and graft survival were compared with elective and urgent patients undergoing whole or reduced-size transplants. Six donors weighed 71.8 ± 17.4 kg and were 22.6 ± 11.0 years of age. Recipients of the right lobe were 11.8 ± 4.2 years of age and weighed 41.9 ± 14 kg. Recipients of the left lateral segment were 1.81 ± 1.1 years of age and weighed 9.85 ± 1.82 kg. Six patients were initially offered the donor allograft because of their hospitalization, critical illness or waiting time. Six additional patients electively underwent transplantation with the leftover liver. Donor organs were screened for normal arterial anatomy. Division of the allograft was performed on the back table in the falciform groove. Generally the left lateral segment graft received the major portion of the hepatic artery and the right lobe the major portion of the portal vein. Five of six (83%) elective patients, two receiving the right lobe and three receiving the left lateral segment had prompt recovery and left the hospital without surgical complication. One recipient of a right lobe transplant died from primary allograft nonfunction. These results are not different from the outcomes of all elective patients who underwent transplantation with whole or reduced-sized transplants in the same program. The authors conclude that split liver transplantation benefits the stable patient who electively receives the liver leftover after reducing the size of a large donor liver for a critically ill child.

Retransplantation for failed hepatic allografts in children

Retransplantation for failed hepatic allografts in children

Liver transplantation for jejunoileal bypass-associated cirrhosis: Allograft histology in the setting of an intact bypassed limb

Jejunoileal bypass (JIB) is a well known cause of steatohepatitis, which may, on occasion, progress to cirrhosis and require liver transplantation. We report 3 patients who underwent orthotopic liver transplantation (OLT) for steatohepatitic cirrhosis secondary to JIB in which the JIB was left intact. All 3 patients have demonstrated recurrent steatosis in the graft after liver transplantation. In two of the cases, the changes are moderately severe, whereas in one case the changes are mild. All 3 patients have essentially normal liver function tests and are clinically asymptomatic; 1 of the patients has undergone removal of the JIB 2.5 years after transplantation. Control hepatic allografts in patients with primary biliary cirrhosis and primary sclerosing cholangitis show negligible fatty change, and in patients who receive transplants for alcoholic steatohepatitis, they rarely (2 of 20 patients) contain greater than 10% fat. We conclude that transplantation alone is not associated with subsequent steatosis. Presence of JIB is, therefore, a continuing risk factor for steatosis in patients who have undergone OLT. Reversal of JIB after OLT may be considered if fatty changes are severe or associated with significant fibrosis.

Immunological study of unresponsive state in rat hepatic transplant model. 1. Phenotypic and functional analyses of infiltrating cells.

In order to elucidate the immunological characteristics of rat liver transplantation, graft-infiltrating cells (GIC) isolated from rat hepatic allografts were analyzed phenotypically and functionally. GIC from long-surviving recipients (Brown Norway livers into Lewis hosts) and acutely rejecting recipients (DA livers into Lewis hosts) were compared. The relative proportions of all T cells and activated T cells determined by flow cytometry were significantly higher in acutely rejecting Lewis recipients than in long-surviving recipients on day 6 after grafting. Phenotypic kinesis of GIC on days 6, 14, and 45 after transplantation from long-surviving Lewis hosts was analyzed. Each proportion of all T cells, OX8-positive cells (cytotoxic T and natural killer cells), and OX39-positive cells (IL-2 receptor), was greatest on day 6 and decreased by day 45. Cytotoxic activity of GIC toward donor lymphocytes on day 6 was greater in acutely rejecting versus long-surviving recipients. These results demonstrate that an immunosuppressive mechanism is already present on day 6 posttransplantation, and that infiltration or activation of cytotoxic T cells is inhibited in the long-surviving rat hepatic allografts.

Immunologic barriers to hepatic adenoviral gene therapy for transplantation.

Adenoviral gene transfer has potential use to attenuate the immunogenicity of hepatic allografts. However, the clinical application of adenoviral gene therapy is currently impeded by the potent host immune response to the virus that limits the duration of its effects. In these studies, we identify the cellular and humoral immune responses to recombinant adenovirus in the liver of mice and define the immunologic barriers to the successful application of this technology to transplantation. The immunobiology of recombinant adenovirus was studied in mouse liver using vectors containing the lacZ and alkaline phosphatase marker genes. The duration of transgene expression was studied in various immunodeficient mice to determine the mechanism of viral clearance. Adoptive transfer of serum to B lymphocyte deficient mice and neutralizing antibody assays were used to define the antiviral humoral response. Hepatic adenoviral transgene expression was prolonged in animals deficient in CD4+ or CD8+ T cells indicating their importance in viral clearance. Unexpectedly, mice lacking B lymphocytes also had delayed elimination of virus suggesting that B cells play a role in the primary immune response. Effective repeat gene transfer was blocked by adenoviral-specific neutralizing antibody. Therefore, a T lymphocyte response results in viral elimination after a primary intravenous inoculation of recombinant adenovirus and a potent humoral response inhibits effective repeat adenoviral gene transfer. The immunogenicity of the vector must be overcome for adenoviral gene therapy to have therapeutic application for hepatic transplantation.

Prostaglandins in liver transplantation.

This review summarizes experimental studies and clinical experiences with prostaglandins in liver transplantation emphasizing two randomized, double blinded placebo controlled clinical trials of prostaglandin E1 (PGE1) involving nearly 300 liver transplant recipients. Resource utilization and pharmacoeconomic aspects are also discussed. In the randomized trials, PGE1 did not affect patient and graft survival. Acute cellular rejection and primary allograft non-function were not reduced by PGE1. Postoperative renal failure was significantly less frequent among PGE1 patients in both trials and in one study perioperative blood product utilization was significantly lower. PGE1 treated patients had reduced intensive care unit length of stay, shorter hospitalization, and significantly lower total health care charges. Despite its failure to show improvements in patient and graft survival, rejection, or primary non-function, PGE1 use in hepatic allograft recipients reduces morbidity and results in notable reduction in the cost of liver transplantation.

Liver transplantation for jejunoileal bypass–associated cirrhosis: Allograft histology in the setting of an intact bypassed limb

Jejunoileal bypass (JIB) is a well known cause of steatohepatitis, which may, on occasion, progress to cirrhosis and require liver transplantation. We report 3 patients who underwent orthotopic liver transplantation (OLT) for steatohepatitic cirrhosis secondary to JIB in which the JIB was left intact. All 3 patients have demonstrated recurrent steatosis in the graft after liver transplantation. In two of the cases, the changes are moderately severe, whereas in one case the changes are mild. All 3 patients have essentially normal liver function tests and are clinically asymptomatic; 1 of the patients has undergone removal of the JIB 2.5 years after transplantation. Control hepatic allografts in patients with primary biliary cirrhosis and primary sclerosing cholangitis show negligible fatty change, and in patients who receive transplants for alcoholic steatohepatitis, they rarely (2 of 20 patients) contain greater than 10% fat. We conclude that transplantation alone is not associated with subsequent steatosis. Presence of JIB is, therefore, a continuing risk factor for steatosis in patients who have undergone OLT. Reversal of JIB after OLT may be considered if fatty changes are severe or associated with significant fibrosis. (Liver Transpl Surg 1997 Jan;3(1):23-7)

Intrahepatic Hematoma following Needle Biopsy of Liver Graft: Incidence and Management

Percutaneous liver biopsy is a frequently used technique to diagnose hepatic allograft dysfunction after liver transplantation. One hundred and twenty-four grafts were biopsied under ultrasound contro

Prostaglandin E1 administration following orthotopic liver transplantation: a randomized prospective multicenter trial

Klein, A S; Cofer, J B; Pruett, T L; Thuluvath, P J; McGory, R; Uber, L; Stevenson, W C; Baliga, P; Burdick, J F

Bile secretory function of the arterialized versus nonarterialized rat liver allograft.

Arterialized and nonarterialized rat models of orthotopic liver transplantation are widely used for studying various aspects of graft function. Although bile duct damage has been implicated in graft nonarterialization, any impact on graft secretory function is unknown. This study sought to investigate whether nonarterialized orthotopic liver transplantation (NOLT) is a valid model for studying bile formation after the animal has fully recovered from the perioperative period. Twenty-four-hour bile collections were performed on eight arterialized and eight nonarterialized rats 4 weeks after transplantation to avoid the confounding effects of preservation-reperfusion injury and perioperative stress. Eight unoperated rats were used as control. There was no difference in mortality or biliary complications between the two transplant groups. The nonarterialized rats exhibited a higher serum aminotransferase level, but serum bilirubin was normal. NOLT resulted in more portal lymphocytic infiltration and bile ductular proliferation. Despite these histologic changes, bile duct epithelial cells remained intact, and spontaneous graft rearterialization was evident in the NOLT group. Bile salt secretion, pool size, and synthesis in both transplant groups did not differ from unoperated rats. NOLT did not adversely affect either bile acid-dependent or bile acid-independent flow. Biliary cholesterol secretion was markedly reduced in both transplant groups, resulting in a more favorable cholesterol saturation index. In conclusion, hepatic allograft secretory function is well maintained at 4 weeks even in the absence of hepatic arterial reanastomosis. Compensatory mechanisms possibly prevent irreversible hepatobiliary damage in NOLT. The NOLT model is quite reasonable to study bile formation after transplantation.

Interferon-α therapy for chronic hepatitis C in special patient populations

Interferon-alpha therapy for chronic hepatitis C in special patient populations raises a number of issues. Patients with hemophilia, kidney disease requiring hemodialysis, mixed cryoglobulinemia, HIV infection, and those receiving an allograft share some characteristics that complicate the treatment of hepatitis C virus infections. These patients generally have some degree of immune deficiency, higher levels of hepatitis C virus replication, and are infected with genotypes 1a or 1b. Each of these characteristics is often associated with a poor response to interferon therapy. Clinical research in this area also has been limited. Current data and clinical experience demonstrate that interferon-alpha therapy should be considered in patients with hemophilia who have concurrent hepatitis C viral infection. Other hepatitis C virus-infected patient populations in which interferon-alpha therapy may be beneficial include those undergoing hemodialysis, mixed cryoglobulinemia, or HIV infection. Further, the high incidence of relapse following treatment cessation in these patients warrants prolonged administration of interferon-alpha. Patients undergoing renal or hepatic allograft transplantation who develop hepatitis C virus infections are not as likely to benefit from interferon-alpha therapy. These patients may be at risk for allograft rejection during interferon treatment.

Restoration of tolerance to rat hepatic allografts by spleen-derived passenger leukocytes.

The tolerance induced by orthotopic liver transplantation (OLT) in certain combinations of rat strains can be prevented by total body irradiation (TBI) of the donor. We demonstrate here that the intravenous inoculation of splenic leukocytes into irradiated donors before OLT could re-establish tolerance in association with a state of microchimerism detected in the recipients. When donor DA (RT1a) strain rats were irradiated with 1000 rad 24 h before liver harvesting and subsequent liver implantation into PVG recipients, five out of six rats died from rejection in this normally tolerogenic OLT (DA-PVG) combination. Injection of 1.5 x 10(8) splenic leukocytes from naive DA rats into the irradiated DA donor rats 24 h before OLT restored the tolerogenic potential of the liver allografts. Immunofluorescence assay revealed an increased number of donor (DA) type cells in the PVG recipient bearing a repopulated DA liver, compared to the PVG recipient of an irradiated liver. These results suggest that passenger leukocytes reconstituted by splenic leukocytes have the capacity to protect liver allografts.

Restoration of tolerance to rat hepatic allografts by spleen-derived passenger leukocytes

Abstract The tolerance induced by orthotopic liver transplantation (OLT) in certain combinations of rat strains can be prevented by total body irradiation (TBI) of the donor. We demonstrate here that the intravenous inoculation of splenic leukocytes into irradiated donors before OLT could re-establish tolerance in association with a state of microchi-merism detected in the recipients. When donor DA (RT1a) strain rats were irradiated with 1000 rad 24 h before liver harvesting and subsequent liver implantation into PVG recipients, five out of six rats died from rejection in this normally tolerogenic OLT (DA-PVG) combination. Injection of 1.5 times 108 splenic leukocytes from naive DA rats into the irradiated DA donor rats 24 h before OLT restored the tolerogenic potential of the liver allografts. Immunofluorescence assay revealed an increased number of donor (DA) type cells in the PVG recipient bearing a repopulated DA liver, compared to the PVG recipient of an irradiated liver. These results suggest that passenger leukocytes reconstituted by splenic leukocytes have the capacity to protect liver allografts.

Immunogenicity of biliary epithelium: Investigation of antigen presentation to CD4+ T cells

The intrahepatic biliary epithelium is susceptible to extensive T-cell- mediated damage during primary biliary cirrhosis, primary sclerosing cholangitis, and hepatic allograft rejection. During these processes, human intrahepatic biliary epithelial cells (HIBEC) become activated and express high levels of the lymphocyte adhesion molecules, intercellular adhesion molecule-1 (ICAM-1) and lymphocyte-associated antigen (LFA)-3, and of class II MHC antigens. It follows that activated HIBEC may also play a direct role in the activation of antigen-specific CD4+ T lymphocytes. The capacity of class II MHC antigen-expressing HIBEC to present antigen and induce specific proliferation of CD4+ T cells was examined in this study. Lines of purified HIBEC were activated by culture with the proinflammatory cytokines interferon gamma (IFN-gamma) and tumor necrosis factor a and were mixed in coculture with allogeneic CD4+ T cells. The result of interaction between these cells was assessed by measurement of lymphoproliferation and IL-2 production. Class II MHC antigen-expressing HIBEC failed to induce either lymphoproliferation or IL-2 production. However, both of these parameters of T-cell activation were positive in cocultures when a costimulation signal was delivered to T cells by adding bivalent anti-CD28 antibodies. The antigen-specific activation of these T cells was further enhanced by the addition of a cross-linking secondary antibody that caused CD28 receptor aggregation. The failure of cytokine-stimulated HIBEC to induce T-cell activation is consistent with the observation that HIBEC do not express the costimulatory CD28 ligands B7-1 or B7-2 at either mRNA or protein levels. It may be concluded that HIBEC are unlikely to play a direct role in activation of antigen-specific CD4+ T lymphocytes within the inflamed liver. (Hepatology 1996 Sep;24(3):561-7)

Prostaglandin E1 administration following orthotopic liver transplantation: A randomized prospective multicenter trial

BACKGROUND & AIMS: Prostaglandin E1 (PGE1) has been used after orthotopic liver transplantation (OLT) based on limited clinical data suggesting PGE1 infusion improves immediate hepatic allograft function. The aim of this study was to conduct a randomized double-blinded multicenter trial to evaluate the effect of PGE1 on early hepatic and renal function in patients undergoing OLT. METHODS: One hundred eighteen patients were randomized to receive either PGE1 or crystalloid placebo intravenously after allograft revascularization. Primary end points were incidence of primary allograft nonfunction (PNF) or severe renal dysfunction. RESULTS: The incidence of PNF was 6.7% (4 of 60) and 6.9% (4 of 58) in the control and PGE1 groups, respectively. PGE1 infusion was, however, associated with improved early renal function (mean peak creatinine level of 1.4 +/- 1.0 and 2.0 +/- 1.0 in patients treated with PGE1 and placebo, respectively; P < 0.001). Severe renal dysfunction occurred more frequently in the placebo group (26.7%) than in the PGE1 group (13.8%; P = 0.65). Additionally, dialysis treatments were more frequent in the placebo group (0.7 +/- 2.0 per patient) than in the PGE1 group (0.2 +/- 1.0 per patient; P = 0.10). Initial intensive care unit stay was shorter in patients treated with PGE1 (4.0 +/- 3.6 days) compared with controls (10.5 +/- 17.1 days) (P < 0.01). CONCLUSIONS: PGE1 administration after OLT resulted in improved renal function and decreased initial postoperative intensive care unit stay but did not affect the incidence of PNF. (Gastroenterology 1996 Sep;111(3):710-5)

Immunogenicity of biliary epithelium: investigation of antigen presentation to CD4+ T cells.

The intrahepatic biliary epithelium is susceptible to extensive T-cell-mediated damage during primary biliary cirrhosis, primary sclerosing cholangitis, and hepatic allograft rejection. During these processes, human intrahepatic biliary epithelial cells (HIBEC) become activated and express high levels of the lymphocyte adhesion molecules, intercellular adhesion molecule-1 (ICAM-1) and lymphocyte-associated antigen (LFA)-3, and of class II MHC antigens. It follows that activated HIBEC may also play a direct role in the activation of antigen-specific CD4+ T lymphocytes. The capacity of class II MHC antigen-expressing HIBEC to present antigen and induce specific proliferation of CD4+ T cells was examined in this study. Lines of purified HIBEC were activated by culture with the proinflammatory cytokines interferon gamma (IFN-gamma) and tumor necrosis factor a and were mixed in coculture with allogeneic CD4+ T cells. The result of interaction between these cells was assessed by measurement of lymphoproliferation and IL-2 production. Class II MHC antigen-expressing HIBEC failed to induce either lymphoproliferation or IL-2 production. However, both of these parameters of T-cell activation were positive in cocultures when a costimulation signal was delivered to T cells by adding bivalent anti-CD28 antibodies. The antigen-specific activation of these T cells was further enhanced by the addition of a cross-linking secondary antibody that caused CD28 receptor aggregation. The failure of cytokine-stimulated HIBEC to induce T-cell activation is consistent with the observation that HIBEC do not express the costimulatory CD28 ligands B7-1 or B7-2 at either mRNA or protein levels. It may be concluded that HIBEC are unlikely to play a direct role in activation of antigen-specific CD4+ T lymphocytes within the inflamed liver.

FK506 inhibits human lymphocyte migration and the production of lymphocyte chemotactic factors in liver allograft recipients.

The macroglide immunosuppressant FK506 is effective at preventing and reversing hepatic allograft rejection. The establishment of graft rejection is dependent upon an influx of lymphocytes from the circulation into the graft in response to locally secreted chemotactic factors. Thus, inhibition of lymphocyte migration might be an additional mode of action of FK506 that could block lymphocyte recruitment to rejecting liver allografts. In the present study, we provide evidence to support this hypothesis because we have demonstrated, using in vitro migration assays, that FK506 can inhibit the migration of lymphocytes, including CD4+ and CD8+ T-cells, to structurally diverse chemotactic factors that are present during human liver allograft rejection. In addition, FK506 acts on lymphocytes in patients with graft rejection to inhibit migration and to block the secretion of chemotactic factors in vitro. Thus, FK506 might reverse established graft rejection by inhibiting lymphocyte recruitment to the graft in vivo.

Plasma prostaglandin E1 concentrations and hemodynamics during intravenous infusions of prostaglandin E1 in humans and swine.

Prostaglandin (PG) E1 administered intravenously has been used for the treatment of primary nonfunction of hepatic allografts and fulminant hepatic failure. It has been proposed that this therapy may improve hepatic blood flow via the vasodilating properties of PGE1. However, PGE1 undergoes extensive metabolic inactivation by the lung and the concentration of PGE1 reaching the liver during intravenous administration has not been determined. Thus, we measured plasma PGE1 concentrations in patients with hepatic dysfunction being treated with PGE1 and in a swine model of PGE1 infusion. We also determined the hemodynamic effects of PGE1 infusion in swine. Blood was sampled from the pulmonary artery, carotid artery, portal vein, and hepatic vein in swine infused with PGE1 (range, 0.67-4.9 microg/kg/hr) demonstrating: (1) a pulmonary extraction ratio of PGE1 of 0.78 +/- 0.12, (2) a splanchnic extraction ratio of PGE1 of 0.54 +/- 0.23, and (3) levels of PGE1 in the systemic circulation of </= 78 pg/ml, even at the highest infusion rates. Despite significant increases in body temperature and pulse rate, hepatic hemodynamics were not affected by the PGE1 infusions in healthy swine. Seven patients receiving intravenous PGE1 for hepatic dysfunction (0.11-1.30 microg/kg/hr) had a pulmonary extraction ratio of 0.69 +/- 0.17. Systemic arterial concentrations of PGE1 were </= 62 pg/ml. These results suggest that due to clearance of PGE2 in the pulmonary and splanchnic circulations, current clinical protocols for intravenous administration of PGE1 are not likely to affect perihepatic hemodynamics.

FK506 inhibits human lymphocyte migration and the production of lymphocyte chemotactic factors in liver allograft recipients

The macroglide immunosuppressant FK506 is effective at preventing and reversing hepatic allograft rejection. The establishment of graft rejection is dependent upon an influx of lymphocytes from the circulation into the graft in response to locally secreted chemotactic factors. Thus, inhibition of lymphocyte migration might be an additional mode of action of FK506 that could block lymphocyte recruitment to rejecting liver allografts. In the present study, we provide evidence to support this hypothesis because we have demonstrated, using in vitro migration assays, that FK506 can inhibit the migration of lymphocytes, including CD4+ and CD8+ T-cells, to structurally diverse chemotactic factors that are present during human liver allograft rejection. In addition, FK506 acts on lymphocytes in patients with graft rejection to inhibit migration and to block the secretion of chemotactic factors in vitro. Thus, FK506 might reverse established graft rejection by inhibiting lymphocyte recruitment to the graft in vivo. (Hepatology 1996 Jun;23(6):1476-83)