The majority of patients with acute liver failure (ALF) die waiting for orthotopic liver transplantation (OLT). No other treatment modality is shown to improve survival. This study was conducted to assess the safety and feasibility of hepatocyte transplantation (HT) and subsequent engraftment and function of donor cells. Functional and structural integrity of cryopreserved and thawed human hepatocytes were assessed by their morphological characteristics, induction of P-4501A1 transcription, and survival in vivo by xenotransplantation into rats. Five patients with severe ALF underwent intrasplenic (4 patients) and/or intrahepatic (2 patients) HT through angiography under cyclosporine immunosuppression. All patients had grade III to IV encephalopathy and factor V levels less than 0.5 U/mL, were ventilator and dialysis dependent, and were not OLT candidates. Three of the 5 patients who survived 48 hours after HT had substantial improvement in encephalopathy scores, arterial ammonia levels, and prothrombin times. Clinical improvement was paralleled by an increase in aminopyrine and caffeine clearances. All 3 patients lived substantially longer than expected based on clinical experience after HT (12, 28, and 52 days) but eventually died. Postmortem examination showed the presence of transplanted hepatocytes in liver and spleen by light microscopy and fluorescent in situ hybridization (FISH). Cryopreserved and thawed human hepatocytes can be transplanted into recipients with ALF with some acceptable but definite complications. Engraftment of donor hepatocytes was proven by histological examination and FISH by both transjugular biopsy and at autopsy. Improvement in brain edema, encephalopathy grade, and clearance of antipyrine and caffeine suggested function, albeit with a 24- to 72-hour delay posttransplantation.

Hepatocyte transplantation is a conceptually attractive alternative to whole organ grafting for some inborn metabolic errors and for fulminant liver failure. However, studies of the immunogenicity of transplanted allogeneic hepatocytes have yielded contradictory results. In these experiments, the effect of purification and cryopreservation of the hepatocytes on the ability of these cells to engraft in the mouse allogeneic recipients without immunosuppression was studied. BALB/ cByJ mouse crude (unpurified), modified (purified or cryopreserved), or dead (irradiated) hepatocyte preparations labeled with fluorescein dye CFSE were infused either into the portal vein or into the spleen parenchyma of the recipient CBA mice. A histological examination revealed normal appearance of engrafted modified hepatocytes with no signs of acute rejection up to 21 days posttransplant. Many of the intrasplenically implanted hepatocytes migrated into the hepatic sinusoids. The modified hepatocytes showed intact ultrastructural appearance 7 days after transplantation. The numbers of inoculated crude hepatocytes rapidly declined with signs of dense infiltration of mononuclear cells in the graft indicating destructive response. The fluorescence of dead hepatocytes was undetectable. These results suggest that reduced immunogenicity may be responsible for the longer survival time of inoculated, purified or cryopreserved hepatocytes with no adverse morphological effects.

Hepatic venoocclusive disease (VOD) is an often fatal disease which can occur during the first 100 days following bone marrow transplantation (BMT).To date multiple factors have been implicated; however, there is no clear cut study to identify the patients at increased risk before and after BMT.Thus, to identify the potential high risk groups and factors predisposing to VOD, we retrospectively reviewed the charts of patients who had BMT between 1990-96 at MD Anderson Cancer Center.Methods: In this case-control study patients' previous chemotherapy, medical and surgical history as well as hospital course, biochemical and histological data 100 days after BMT were extensively evaluated.Baltimore Clinical Diagnostic Criteria were applied to diagnose VOD within 21 days after BMT (Bilirubin _> 2 mg/dL, new onset ascites, 5% gain above admission weight, hepatomegaly and right upper quadrant pain) In addition to hyperbilirubinemia, presence of 2 out of 3 events was considered as diagnostic for VOD.Multiple stepwise logistic regression analysis was done and p<0.05 was considered as statistically significant.Results: There were 19 VOD patients and 99 controls who were age, gender and race matched.Use of pain medications, cyclosporin-A and methotrexate and presence of renal failure were the only significant independent risk factors for VOD (Table 1).

Islet allografts transplanted into Type I diabetic recipients may be destroyed by allorejection or recurrent autoimmune diabetes. We studied islet transplantation in three murine models in order to determine the relative sensitivity of autoimmunity and alloimmunity to two immunosuppressive agents that may be useful in clinical islet transplantation: 15-deoxyspergualin (DSG) and anti-CD4 antibody (GK 1.5). In the model in which only allorejection occurs (BALB/c islets transplanted into streptozotocin-induced diabetic CBA or streptozotocin-induced diabetic NOD recipients), both DSG and anti-CD4 antibody treatment led to indefinite survival of allogeneic islets (>100 days in both treatments). In the second model in which only recurrent autoimmunity can destroy islet grafts (islets from NOD donors transplanted into spontaneously diabetic NOD recipients), only anti-CD4 treatment caused prolonged graft survival [MST 36.7 ± 6.8 days vs 9.8 ± 1.8 days (controls),P< 0.0002]. Treatment with DSG did not cause any increase in graft survival (MST 12.6 ± 5.4 days, NS). Finally, using a model in which both autoimmunity and allorejection may occur (BALB/c to spontaneously diabetic NOD mice), treatment with anti-CD4 caused marked graft prolongation [42.0 ± 14.5 days vs 7.2 ± 0.8 days (control),P< 0.002] while DSG again did not prolong graft survival with respect to untreated recipients (9.8 ± 3.0, NS). We conclude that recurrent autoimmunity in the NOD mouse involves a CD4+T cell that is not sensitive to DSG. Anti-CD4 antibody may be useful in human clinical islet transplantation trials because it seems to prevent both allorejection and recurrent autoimmunity.

Background. the purpose of this study was to determine whether an antibody to vascular cell adhesion molecule 1 (VCAM1) prolongs the survival of neovascularized pancreatic islet allografts.Methods. We treated CBA (H-2(k)) recipients of BALB/c (H-2(d)) islet allografts with anti-VCAM1 antibody (400 mu g/day for 20 days). Sensitized recipients of islet grafts also were treated with anti-VCAM1. To study mechanism we performed mixed lymphocyte reactions (MLRs) with anti-VCAM1 and studied the graft infiltrate in treated recipients.Results. Anti-VCAM1-treated CBA recipients showed prolonged graft survival with indefinite survival in five of nine cases. Anti-VCAM1 prevented proliferation in an MLR but not when added 36 hours after the beginning of the MLR. Anti-VCAM1 did not prolong allograft survival in sensitized recipients and did not prevent lymphocytic infiltration of the graft at 7 days.Conclusions. Anti-VCAM1 prolongs allograft survival in neovascularized islets in which the donor vascular endothelium plays little or no role in immunogenicity. VCAM1 appears to the important in the afferent phase (lymphocyte activation) of the allograft response. Once activated, either late in an MLR or in sensitized recipients, lymphocytes are not dependent on VCAM1 for function. Finally, anti-VCAM1 does not appear to affect the homing of lymphocytes to the allograft.

The population of T-cells that develops in any individual can be divided into families based on sequence differences in the beta-chain variable region of the T-cell receptor heterodimer. Major histocompatibility complex products and endogenous retroviral gene products have both been shown to exert powerful influences on the frequency distribution of T-cell receptor beta-chain variable region families in the mouse. In most mouse strains, these repertoire modifiers appear to be fully functional early in mouse development and shape a repertoire of antigen specificities that remains essentially unchanged from the first weeks of life until old age. In NOD mice, an inbred mouse model of type I diabetes, puberty in males coincides with a beta-chain variable region-specific T-cell expansion that mimics the results of exposure to exogenous superantigens in immunologically mature animals. The subsequent behavior of this subset indicates that it may play a role in the relative protection of male NOD mice from complete pancreatic beta-cell destruction and overt diabetes.