摘要:

The shortage of human livers available for hepatocyte isolation limits its clinical application. The availability of cloned, conditionally immortalized hepatocytes that could be grown in culture but would lose their transformed phenotype and provide metabolic support upon transplantation would greatly facilitate the treatment of acute liver failure. Toward this goal, we transduced isolated Lewis rat hepatocytes using a replication-defective recombinant retrovirus capable of transferring a gene encoding a thermolabile mutant simian virus 40 T antigen (SV40ts). The cloned, immortalized hepatocytes proliferate at 33°C. At the nonpermissive temperatures (37-39°C), they stop growing and exhibit characteristics of differentiated hepatocytes. These cells did not produce tumors when transplanted in mice with severe combined immunodeficiency disease or in syngeneic rats. To induce acute liver failure, Lewis rats were subjected to 90% hepatectomy (Hpx) and given 5% oral dextrose. All rats that did not undergo hepatocyte transplantation died within 96 hr. Fifty percent of rats that received intrasplenic injection of 10×106primary Lewis rat hepatocytes (G2, n=6) or 10×106SV40ts-conditionally immortalized (SV40ts-ci) hepatocytes (G3, n=8) 1 day before 90% hepatectomy survived, whereas 80% of rats that received an intraperitoneal injection of 200×106primary Lewis rat hepatocytes (G4, n=10) or 200×106SV40ts-ci hepatocytes (G5, n=10) on the day of hepatectomy survived. Survival after intraperitoneal injection of a cellular homogenate of 200×106primary Lewis rat (G7, n=9) or SV40ts-ci hepatocytes (G8, n=10) on the day of Hpx was 33% and 40%, respectively, whereas survival after intraperitoneal injection of 200×106Lewis rat bone marrow cells (G6, n=7) was 29%. Thus, transplanted, conditionally immortalized hepatocytes can be as effective as primary hepatocytes in supporting life during acute liver insufficiency. This work represents the first step in developing an hepatocyte cell line that would partially alleviate the organ-donor shortage and could be of potential clinical value.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.Nisoldipine, a calcium antagonist, has been reported to improve the quality of grafted rat livers. We thus assessed the protective effect of two calcium antagonists, nisoldipine and nickel, during extended cold ischemia-reperfusion.Methods.Rat livers were isolated and perfused before or after 24 hr of cold ischemia in University of Wisconsin solution (4°C) with or without nisoldipine or nickel. Sinusoidal endothelial cell and hepatocyte functions were measured by hyaluronic acid and taurocholate elimination, respectively.Results.Similar alterations in hepatocyte and sinusoidal cell functions were found in all groups after cold ischemia with or without calcium antagonists. In a second set of experiments, liver transplantation was performed in two groups of rats with livers stored under identical conditions with or without nisoldipine. Seven of 12 animals (62.5%) in both groups survived for over 10 days after 24-hr preservation in University of Wisconsin solution. Survival rates were similar in both groups.Conclusions.Calcium antagonists do not appear to have a direct protective effect on sinusoidal endothelial cell and hepatocyte functions, nor on the overall liver preservation after extended cold preservation-reperfusion.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.Heterotopic guinea pig (GP) cardiac xenografts (XG) are hyperacutely rejected within minutes when transplanted into rats.Methods.In this GP to rat cardiac XG model, we studied the effect on graft survival of a short cold preservation time (1 hr at 4°C) in the presence or absence of rat anti-GP IgM preformed antibodies. The complete depletion of circulating IgM was obtained by two intraperitoneal injections of anti-rat IgM monoclonal antibody (MARM-4) on preoperative days -3 and -1.Results.When the GP cardiac XG was cold preserved for 1 hr before transplantation, the mean graft survival time (MST) was 13.5±2.8 min, whereas without previous cold preservation, the MST was significantly prolonged to 51.5±12.3 min (P<0.001). Interestingly, the complete depletion of preformed circulating IgM before grafting significantly prolonged the MST of a cold-preserved XG to 37.1±11.3 min in comparison with a nondepleted recipient of a cold-preserved XG (P<0.02), but did not prolong the graft survival of a XG that was not cold preserved (42.5±14.1 min). To assess the effect of cold preservation and/or ischemia reperfusion, we intravenously injected a superoxide-dismutase mimetic (EUK-134) just before transplantation of a cold-preserved XG. This antioxidant regimen improved the MST from 13.5±2.8 min to 35.3±7.3 min (P<0.001). These results clearly suggested that either preservation lesions or preformed IgM are capable of accelerating the loss of the cardiac graft function, but also that the presence of preformed IgM seems to be especially deleterious when the cardiac XG has previously been ischemically injured. Analyzing the histological data, we also observed that the prompt cessation of cardiac function seen in cold-preserved grafts was uniformly associated with massive interstitial hemorrhage, thereby suggesting a particular susceptibility of the GP cardiac XG to cold preservation. To assess the effect of preservation on the GP cardiac function in a nonimmunological model, we performed syngeneic GP cardiac grafts and found that 1 hr of cold preservation provoked massive interstitial hemorrhage capable of promptly inducing the cessation of the heartbeat.Conclusions.Overall, this study demonstrated that both ischemic lesions and immunological processes might induce the cessation of cardiac graft function in the GP to rat model and this cessation of graft function is probably often misinterpreted as a XG rejection only.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.Cholestasis is a complication that occurs during the rejection of liver transplants. The aim of this study was to investigate the association of activated Kupffer cells (KCs) and Na+,K+-ATPase activity for taurocholate cotransport and bile canalicular (BC) Mg++-ATPase activity for hepatobiliary excretion in rat liver allograft.Methods.Quantitative analyses of KC number and size in relationship to enzyme activity of Na+,K+-ATPase and of BC Mg++-ATPase were conducted in rejected liver after allogenic transplantation and after prevention of rejection using cyclosporine.Results.The animals were examined on the 10th postoperative day. In the rejection group, the number of KCs significantly increased more than fourfold in comparison with the number of KCs in the control livers. Some KCs were found in the sinusoids, but the majority were located in the space of Disse. Na+,K+-ATPase activity vanished from the basolateral plasma membrane, whereas BC Mg++-ATPase activity was restored in the apical domain. With immunosuppression, KCs showed the same behavior as in the control group, and activity of both ATPases was observed as strong electron-dense precipitates in basolateral and apical plasma membrane domains.Conclusions.In this study, we demonstrate that activated KCs migrate into the donor liver and release cytokines, which leads to the loss of Na+,K+-ATPase activity in the rejection group. BC Mg++-ATPase activity was not influenced by these mediators of activated macrophages. Since Na+,K+-ATPase is the cotransporter for hepatocyte taurocholate uptake, these data may contribute to understanding the mechanisms for cholestasis during hepatic allograft rejection.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.We have reproducibly induced specific tolerance to multiple minor histocompatibility antigens with nondepleting anti-CD4 and -CD8 monoclonal antibodies. The tolerance induced is effective for the lifetime of the host. We have tested this therapy in a number of mouse strain combinations to further understand the mechanisms.Methods.Various mouse strains were grafted with allogeneic tail skin with and without nondepleting CD4- and CD8-specific monoclonal antibody therapy. The grafts were monitored daily for signs of rejection.Results.Whereas the CBA/Ca (H2k) strain can be made tolerant to skin grafts that are mismatched at multiple minor histocompatibility antigens indefinitely, using the same protocol, long-term survival of similarly mismatched grafts on the HW80 (B6 congenic for BALB H1) mouse strain is limited to around 8 weeks. Interestingly, the B10.BR strain, which is also of the H2khaplotype, is also not readily tolerized. In addition, an F1 between the CBA/Ca and the resistant B10.BR strains is B10.BR-like in its susceptibility to tolerance induction. Susceptibility to such antibody-dependent tolerance induction is not related to immunogenicity because grafts mismatched at only a single minor antigen also do not reproducibly survive beyond 8 weeks when grafted onto HW80 mice in the presence of the antibody therapy.Conclusions.The data strongly suggest that the B6/B10 genetic background confers a level of resistance to CD4- and CD8-specific monoclonal antibody-dependent tolerance induction.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.Previous studies of cyclosporine-induced cholestasis were flawed by confounders encountered in human studies and discrepancies in acute animal experiments. Even the cyclosporine vehicle, polyoxyethylated castor oil (Cremophor EL), had been implicated in cholestasis. The purpose of this study was to investigate how cyclosporine affects bile salt kinetics and biliary lipid secretion in a rat model under steady state conditions.Methods.Three groups of male Lewis rats (n=10) were given daily subcutaneous injections of either cyclosporine (CsA; 10 mg/kg body weight), Cremophor, or NaCl (control) for 1 week. Twenty-four-hour bile collection was performed 18 hr after the last injection. The first hour's output measured bile flow and organic bile solute secretion rates. Bile salt pool size and basal synthesis were determined with the washout technique.Results.CsA significantly reduced basal bile flow and bile salt secretion by 25%. Bile salt synthesis was suppressed 45% (CsA: 3.50±0.8 μmol/g liver/24 hr vs. control: 6.31±1.17 μmol/g liver/24 hr;P<0.05), which resulted in a 28% reduction in the bile salt pool size (CsA: 16.9±1.9 μmol/g liver vs. control: 23.6±2.0 μmol/g liver;P<0.05). Bile salt-independent flow was significantly suppressed (29%), whereas bile salt-dependent flow was only modestly reduced. Biliary phospholipid output decreased 23% (CsA: 11.7±0.8 nmol/min/g liver vs. control 15.2±1.1 nmol/min/g liver;P<0.05), but cholesterol secretion was unaltered, resulting in a 29% increase in the cholesterol saturation index (CsA: 0.40±0.03 vs. control 0.31±0.02;P<0.05). Cremophor had no significant effects on bile secretion or bile salt kinetics.Conclusions.CsA induces cholestasis by decreasing both bile flow and bile salt secretion. Its suppression of bile salt synthesis reduces the bile salt pool size. The drug inhibits bile salt and phospholipid secretion without a corresponding change in cholesterol secretion and thus elevates cholesterol saturation in bile, a potential risk for gallstone formation.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.Tissue samples for the diagnosis of pancreatic allograft rejection are now obtained routinely through the application of the percutaneous needle biopsy technique. The availability of biopsy material (89% adequate for diagnosis in our setting) presents a challenge for pathologists who are asked to provide a fast and accurate diagnosis of rejection and its severity, while at the same time being able to differentiate rejection from other causes of graft dysfunction.Methods.To differentiate rejection from other pathologic processes, 26 histologic features were assessed in 92 biopsies performed for confirmation of clinical diagnosis of rejection and the results were compared with 31 protocol biopsies, 12 allograft pancreatectomies with non-rejection pathology, and 30 native pancreas resections with various disease processes.Results.Based on these comparisons, a constellation of findings relating to the vascular, septal, and acinar inflammation was identified for the diagnosis of rejection. Application of these features led us to revise our scheme for grading rejection (ranging from 0-normal to V-severe rejection) to include the categories of “inflammation of undetermined significance” and “minimal rejection.” The scheme was used by five pathologist to grade 20 biopsies independently of any clinical data and the interobserver level of agreement was highly significant (κ=0.83,P<0.0001). This grading scheme was applied blindly to all (183) biopsies from 77 patients with 6-52 months of follow-up. The correlation of the highest degree of rejection on each patient and ultimate graft loss (0% for grades 0-I, 11.5% for grade II, 17.3% for grade III, 37.5% for grade IV, and 100% for grade V) was highly statistically significant (P<0.002). The fraction of grafts lost due to pure immunologic causes increased proportionally to the grade of rejection (0, 50, 66, and 100% for grades II, III, IV, and V, respectively).Conclusions.This study provides strong support for the proposed pancreas rejection grading scheme and confirms its potential for practical use.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Blood transfusions are common in patients with end-stage liver disease (ESLD), and their effects on sensitization, rejection, and liver graft survival are not well known. These effects were examined in 121 recipients of primary liver grafts, surviving ≥30 days.Ninety-six (79%) patients received transfusions before transplantation. Transfusion recipients had significantly fewer severe or recurrent rejection episodes (18%), compared with patients who did not receive transfusions (42%,P=0.006), if the first transfusion was ≥90 days before the transplant. Patients with alcoholic ESLD (n=49) had significantly fewer severe rejection episodes when compared with the nonalcoholic (n=72) patients (12% vs. 35%,P=0.004). The transfusion benefit was, however, more apparent and significant in the nonalcoholic (26% vs. 56% in nontransfused,P=0.02) than among the alcoholic recipients (6% vs. 25%,P=0.1). This finding is, most likely, due to a combination of a higher rate of severe rejection and the statistical power of the larger number of recipients in the nonalcoholic group. This finding is further corroborated by a multivariate analysis in which blood transfusions retained their benefit (P<0.05) independent of recipient's age and diagnosis. Graft and patient survival were not significantly different in the transfused versus nontransfused groups.Transfusion recipients had a higher panel antibody (11.4±23.4 vs. 2.7±8.1,P<0.02) but no increased risk of a positive crossmatch. In liver recipients, blood transfusions diminish the risk of rejection independent of recipient's age and the cause of ESLD.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.Prophylactic treatment with ursodeoxycholic acid (UDCA) has been reported to reduce the incidence of acute rejection after liver transplantation compared with historical controls. We investigated this in a prospective, randomized, placebo-controlled multicenter study.Methods.Fifty-four liver transplant patients were allocated to the UDCA treatment group (15 mg/kg/day), and 48 patients were allocated to the placebo group. Trial medicine was started on the first postoperative day and was given for 3 months. Follow-up was for 12 months. Treatment was stratified for adults with chronic liver disease (n=77), adults with acute liver failure (n=10), and children (n=15).Results.The frequency of patients with acute rejection was 65% in the UDCA treatment group and 68% in the placebo group. The frequency of steroid-resistant rejection was similar in both groups. The probability of acute rejection, analyzed according to the intention-to-treat policy with Kaplan-Meier analysis, was similar in both treatment groups. No significant differences were found in patient survival and graft survival probabilities. For the biochemical markers of cholestasis, only γ-glutamyltransferase was significantly improved after 2 months of UDCA treatment.Conclusions.The initial optimistic report of a beneficial effect of prophylactic treatment with UDCA on acute rejection after liver transplantation was not confirmed in this controlled study.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Cytomegalovirus (CMV) is a cause of considerable morbidity and mortality among orthotopic liver transplant (OLT) recipients. To study the impact of CMV on cost and hospital length of stay in this population, we undertook an analysis of a cohort of OLT recipients from four transplant centers in Boston who participated in a CMV prophylaxis trial. First posttransplant year hospital length of stay (including the hospital stay after transplantation and readmissions within 1 year after transplantation) was available for all 141 patients included in the study. In a multiple linear regression model, bacteremia (P=0.0001), CMV disease (P=0.0007), abdominal reexploration (excluding retransplantation) (P=0.0070), recipient age ≤16 years (P=0.0352), and the number of units of blood products (red blood cells, platelets, or fresh frozen plasma) administered during transplantation (P=0.0523) were shown to be independently associated with longer first posttransplant year hospital length of stay. Cost data for in-hospital care for the year beginning with admission for liver transplantation were available for 66 OLT recipients. Using a multiple linear regression model, development of CMV disease (P=0.0001), the number of units of blood products administered during transplantation (P=0.0001), bacteremia (P=0.0002), decreased pretransplant renal function (estimated by creatinine clearance) (P=0.0109), and need for retransplantation (P=0.0619) were shown to be independently associated with higher cost. These data strongly suggest that CMV disease has a direct impact on cost and hospital length of stay in liver transplantation.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID