A dog model for acetaminophen-induced fulminant hepatic failure.

The development of a large animal model of fulminant hepatic failure produced with acetaminophen that should be useful in the development and evaluation of potential medical therapies for the important clinical problem of fulminant hepatic failure is described. Acetaminophen in dimethyl sulfoxide (600 mg/ml) given as three subcutaneous injections, with the first dose (750 mg/kg body wt) being given at noon, the second dose (200 mg/kg body wt) being given 9 h later, and the third dose (200 mg/kg body wt) being given 24 h after the initial dose consistently produces fulminant hepatic failure in dogs. The dimethyl sulfoxide vehicle, injected intramuscularly, does not influence either animal survival or hepatic function in control-treated dogs. No deaths occur within the first 36 h. By 72 h after initial drug administration, the mortality is 90%. Histopathological and biochemical investigations demonstrate a high degree of hepatocellular necrosis in nonsurviving animals without appreciable damage to the kidneys, lungs, or heart. The drug schedule and preparation outlined avoids the administration of large volumes of vehicle and results in prolonged high levels of acetaminophen in the blood sufficient to induce severe hepatic injury. Ranitidine (120 mg/kg body wt i.m.) given 30 min before each acetaminophen dose significantly reduces the mortality and hepatic necrosis produced using this model. This model satisfies all criteria established by Miller et al. for the production of a suitable large animal model of fulminant acute hepatic failure

Isolation and primary cultures of human intrahepatic bile ductular epithelium

A technique for the isolation of human intrahepatic bile ductular epithelium, and the establishment of primary cultures using a serum- and growth-factor-supplemented medium combined with a connective tissue substrata is described. Initial cell isolates and monolayer cultures display phenotypic characteristics of biliary epithelial cells (low molecular weight prekeratin positive; albumin, alphafetoprotein, and Factor VIII-related antigen negative). Ultrastructural features of the cultured cells show cell polarization with surface microvilli, numerous interepithelial junctional complexes and cytoplasmic intermediate prekeratin filaments. © 1988 Tissue Culture Association, Inc

A dog model for acetaminophen-induced fulminant hepatic failure.

The development of a large animal model of fulminant hepatic failure produced with acetaminophen that should be useful in the development and evaluation of potential medical therapies for the important clinical problem of fulminant hepatic failure is described. Acetaminophen in dimethyl sulfoxide (600 mg/ml) given as three subcutaneous injections, with the first dose (750 mg/kg body wt) being given at noon, the second dose (200 mg/kg body wt) being given 9 h later, and the third dose (200 mg/kg body wt) being given 24 h after the initial dose consistently produces fulminant hepatic failure in dogs. The dimethyl sulfoxide vehicle, injected intramuscularly, does not influence either animal survival or hepatic function in control-treated dogs. No deaths occur within the first 36 h. By 72 h after initial drug administration, the mortality is 90%. Histopathological and biochemical investigations demonstrate a high degree of hepatocellular necrosis in nonsurviving animals without appreciable damage to the kidneys, lungs, or heart. The drug schedule and preparation outlined avoids the administration of large volumes of vehicle and results in prolonged high levels of acetaminophen in the blood sufficient to induce severe hepatic injury. Ranitidine (120 mg/kg body wt i.m.) given 30 min before each acetaminophen dose significantly reduces the mortality and hepatic necrosis produced using this model. This model satisfies all criteria established by Miller et al. for the production of a suitable large animal model of fulminant acute hepatic failure

Abdominal multivisceral transplantation

Under FK506-based immunosuppression, 13 abdominal multivisceral transplantations were performed in 6 children and 7 adults. Of the 13 recipients, 7 (53.8%) are alive and well with functioning grafts after 9 to 31 months. Six recipients died: Three from PTLD, one from rejection, one from sepsis, and one from respiratory failure. In addition to rejection, postoperative complications occurring in more than isolated cases included PTLD (n=6), abdominal abscess formation (n=5), pancreatitis (n=3), and ampullary dysfunction (n=2). In addition, infection by enteric microorganisms was common during the early postoperative period. Currently, all 7 survivors are on an oral diet and have normal liver function. Two recipients (one insulin-dependent) require antidiabetes treatment, in one case following distal pancreatectomy and in the other after two episodes of pancreatic rejection. Thus, abdominal multivisceral transplantation is a difficult but feasible operation that demands complex and prolonged posttransplantation management. It is not yet ready for application and awaits a better strategy of immune modulation. © 1995 by Williams & Wilkins

The current status of hepatic transplantation at the University of Pittsburgh.

Tacrolimus is a more potent and satisfactory immunosuppressant than CyA for combination therapy with prednisone. In randomized trials comparing the 2 drugs, the ability of tacrolimus to rescue intractably rejecting grafts on the competing CyA arm allowed equalization of patient and graft survival on both arms when the intent-to-treat analytic methodology was applied. The ability of tacrolimus to systematically rescue the treatment failures of CyA suggested, as a matter of common sense, that it is the preferred baseline drug for hepatic transplantation. This conclusion was supported by analysis of secondary end points, including the ability to prevent rejection. Hepatic-intestinal, multivisceral and isolated intestinal transplantation became feasible on a practical basis only after the advent of tacrolimus. Nevertheless, better management strategies must be devised before intestinal transplantation, alone or with other abdominal viscera, will meet its potential. One such strategy is based on the discovery of the presence of previously unsuspected, low-level donor leukocyte chimerism in long-surviving allograft recipients. We believe that this chimerism is the essential explanation for the feasibility of organ transplantation and a link to the acquired neonatal tolerance demonstrated by Billingham, Brent and Medawar (32). The hematolymphopoietic chimerism in organ recipients explains why weaning to a drug-free state in selected long-term survivors is frequently feasible and particularly if the allograft is a liver. Weaning should never be attempted without a stepwise protocol and careful monitoring of graft function. Recognition of the natural chimerism that develops after whole organ transplantation has led to efforts to augment it with perioperative donor BM infusion. This procedure has been shown to be free of significant complications (including GVHD) in all kinds of whole organ recipients, including those given intestine. The prospects of clinical xenotransplantation must be evaluated in the same context of chimerism as that delineated for allotransplantation with the discovery of spontaneous chimerism. Before addressing chimerism-related questions in xenotransplantation, the additional barrier of the complement activation syndromes that cause hyperacute rejection will have to be surmounted. Although measures to effectively transplant xenografts have so far eluded us, the availability of the more potent drug, tacrolimus, and recognition of the seminal basis of allograft (or xenograft) acceptance via chimerism has inserted an element of reality into the largely wishful thinking that has been evident in discussions about the future of xenotransplantation

Humoral and cellular immunopathology of hepatic and cardiac hamster-into-rat xenograft rejection: Marked stimulation of IgM^++bright/IgD^+dull splenic B cells

Normal Lewis rat serum contains antibodies (IgM > IgG) that bind to hamster leukocytes and endothelial cells. Transplantation of either the heart or liver from hamster rat results in release of hamster hematolymphoid cells from the graft, which lodge in the recipient spleen (cell migration), where recipient T- and B-cell populations initiate DNA synthesis within one day. There is marked stimulation of splenic IgM++(bright)/IgD+(dull) B cells in the marginal zone and red pulp, which account for 48% of the total splenic blast cell population by 4 days after liver transplantation. CD4+ predominant T-cell proliferation in the splenic periarterial lymphatic sheath and paracortex of peripheral lymph nodes occurs almost simultaneously. The effector phase of rejection in cardiac recipients is dominated by complement-fixing IgM antibodies, which increase daily and result in graft destruction in 3 to 4 days, even in animals treated with FK506. In liver recipients, combined antibody and cellular rejection, associated with graft infiltration by OX8+ natural killer, and fewer W3/25+ (CD4) lymphocytes, are responsible for graft failure in untreated recipients at 6 to 7 days. FK506 inhibits the T-cell response in liver recipients and significantly prolongs graft survival, but does not prevent the rise or deposition of IgM antibodies in the graft. However, a single injection of cyclophosphamide 10 days before transplantation effectively depletes the splenic IgM++(bright)/IgD+(dull) cells and in combination with FK506, results in 100% survival of both cardiac and hepatic xenografts for more than 60 days. Although extrapolation of morphological findings to functional significance is fraught with potential problems, we propose the following mechanisms of xenograft rejection. The reaction initially appears to involve primitive host defense mechanisms, including an IgM-producing subpopulation of splenic B cells and natural killer cells. Based on the reaction and distribution of OX8+ and W3/25+ cells, antibody dependent cell cytotoxicity and delayed-type hypersensitivity responses seem worthy of further investigation as possible effector mechanisms. Effective control of xenograft rejection is likely to require a dual pharmaceutical approach, one to contain T-cell immunity and another to blunt the primitive B-cell response

ALLOREACTIVE T LYMPHOCYTES CULTURED FROM LIVER TRANSPLANT BIOPSIES: ASSOCIATIONS OF HLA SPECIFICITY WITH CLINICOPATHOLOGICAL FINDINGS.

Lymphocyte cultures grown from liver allograft biopsies were shown to exhibit alloreactivity towards donor cells as measured by primed lymphocyte testing (PLT). The PLT specificity was determined in assays using HLA typed panel cells and/or by inhibition testing with HLA specific monoclonal antibodies. Certain cultures exhibited PLT specificity towards class I HLA antigens of the donor, whereas others were specific for class II HLA antigens or recognized mixtures of class I and II antigens. These PLT specificity patterns were compared with clinical, histological and laboratory findings on the liver transplant patients at the time of the biopsy. Biopsies yielding class I specific PLT cells were taken generally during the earlier posttransplant period, whereas class II specific cells were grown from later biopsies. There was no significant correlation of the PLT specificity towards class I vs II antigens with the levels of total or direct bilirubin, serum glutamate oxaloacetic transaminase (SGOT), and serum glutamate pyruvate transaminase (SGPT), although a trend towards higher values was noted for biopsies presenting with a class II specific infiltrate. However, the levels of gamma glutamyl transpeptidase (GGTP) and alkaline phosphatase (AP) were significantly increased when biopsies yielded class II specific rather than class I specific PLT cells. Biopsy histology showed more damage to bile duct epithelium in association with class II PLT specificity whereas intense but often reversible infiltrates were found in biopsies yielding class I specific cells. The elevated GGTP and AP levels are probably related to the interaction of class II specific T cells with bile duct epithelium, which has been shown to express induced class II HLA antigens on their cell surface

Three years clinical experience with intestinal transplantation

BACKGROUND: After the successful evolution of hepatic transplantation during the last decade, small bowel and multivisceral transplantation remains the sole elusive achievement for the next era of transplant surgeons. Until recently, and for the last thirty years, the results of the sporadic attempts of intestinal transplantation worldwide were discouraging because of unsatisfactory graft and patient survival. The experimental and clinical demonstration of the superior therapeutic efficacy of FK 506, a new immunosuppressive drug, ushered in the current era of small bowel and multivisceral transplantation with initial promising results. STUDY DESIGN: Forty-three consecutive patients with short bowel syndrome, intestinal insufficiency, or malignant tumors with or without associated liver disease, were given intestinal (n=15), hepatic and intestinal (n=21), or multivisceral allografts that contained four or more organs (n=7). Treatment was with FK 506 based immunosuppression. The ascending and right transverse colon were included with the small intestine in 13 of the 43 grafts, almost evenly distributed between the three groups. RESULTS: After six to 39 months, 30 of the 43 patients are alive, 29 bearing grafts. The most rapid convalescence and resumption of diet, as well as the highest three month patient survival (100 percent) and graft survival (88 percent) were with the isolated intestinal procedure. However, this advantage was slowly eroded during the first two postoperative years, in part because the isolated intestine was more prone to rejection. By the end of this time, the best survival rate (86 percent) was with the multivisceral procedure. With all three operations, most of the patients were able to resume diet and discontinue parenteral alimentation, and in the best instances, the quality of life approached normal. However, the surveillance and intensity of care required for these patients for the first year, and in most instances thereafter, was very high, being far more than required for patients having transplants of the liver, kidney or heart. CONCLUSIONS: Although intestinal transplantation has gone through the feasibility phase, strategies will be required to increase its practicality. One possibility is to combine intestinal transplantation with contemporaneous autologous bone marrow transplantation