ISSN:0041-1337    

出版商:OVID    

年代:2000

数据来源: OVID

ISSN:0041-1337    

出版商:OVID    

年代:2000

数据来源: OVID

ISSN:0041-1337    

出版商:OVID    

年代:2000

数据来源: OVID

ISSN:0041-1337    

出版商:OVID    

年代:2000

数据来源: OVID

ISSN:0041-1337    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Background.Gene delivery through a nonviral, receptor-mediated system widely expressed in transplanted tissue would have important advantages in transplantation, where gene delivery is performed ex vivo. Integrins are widely expressed cell surface receptors and can be targeted for gene delivery.Methods.A synthetic 31 amino acid DNA vector (polylysine-molossin) comprising a 15-amino acid moiety for targeting cellular integrins (derived from the snake venom, molossin) and a 16-amino acid polylysine moiety for DNA-binding, has been evaluated. The 31-amino acid vector, as well as its separate 15-amino acid integrin-binding and (lys)16components, were individually synthesized, and a monoclonal antibody was raised to the molossin peptide for these studies. Binding to cell lines and tissue sections and capacity for gene delivery were examined.Results.Flow cytometric studies with the ECV304 cell line demonstrated that the binding of polylysine-molossin and polylysine-molossin/DNA complexes involved both electrostatic and integrin-mediated interactions with the cells, with the electrostatic binding being sufficient for maximal binding. However, binding to cellular integrins was essential for successful gene transfer. Binding studies on frozen tissue sections of the rat and pig demonstrated that the molossin peptide bound to many cell types of interest in transplantation, but not to all. Among the negative tissues were vascular endothelium and pancreatic islets. Small species differences in tissue binding were noted between the rat and pig.Conclusions.This study defines the cooperative nature of the binding of this vector system to target cells and establishes the cell types most likely to be effectively targeted for DNA transfer.

ISSN:0041-1337    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Background.Oxygen-derived free radicals play a central role in pathomechanisms of reperfusion injury after organ transplantation. Endogenous radical scavenger systems such as superoxide dismutase (SOD) degrade toxic radicals; however, SOD is degraded rapidly when given exogenously. Therefore, the hypothesis that treatment of the donor liver with an adenoviral vector encoding the Cu/Zn-SOD gene (Ad-SOD1) would lead to permanent gene expression and therefore protect the organ against injury and increase survival in a rat model of liver transplantation was tested.Methods.Some donors were infected with Ad-SOD1, whereas untreated grafts and livers infected with the indicator genelacZencoding bacterial &bgr;-galactosidase (Ad-lacZ) served as controls. After orthotopic liver transplantation, survival, serum transaminases, and histopathology were evaluated.Results.Approximately 80% of hepatocytes expressed &bgr;-galactosidase 72 hr after injection of Ad-lacZ. Moreover, SOD1 gene expression and activity were increased 3- and 10-fold in the Ad-SOD1 group, respectively. After transplantation, 20–25% of rats treated with Ad-lacZ survived. In contrast, all SOD1-treated animals survived. Transaminases measured 8 hr after transplantation in Ad-SOD1 rats were only 40% of those in controls, which increased 40-fold above normal values. Approximately 20% of hepatocytes in untreated and Ad-lacZ-infected organs were necrotic 8 hr after reperfusion, whereas necrosis was nearly undetectable in grafts from rats treated with Ad-SOD1.Conclusions.This study provides clear evidence for the first time that gene therapy with Ad-SOD1 increases survival and decreases hepatic injury after liver transplantation. Genetic modification of the liver represents a future approach to protect organs against injury where oxygen-derived free radicals are involved.

ISSN:0041-1337    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Background.Our purpose was to develop and evaluate protocols for selective immunosuppression after liver transplantation using the monoclonal antibodies (mAbs) NDS-61, directed against the interleukin-2 receptor (CD25), and 1A29, directed against the intercellular adhesion molecule-1 (CD54), in combination with subtherapeutic cyclosporine (CsA).Methods.Orthotopic rat liver transplantation (ORLT) was performed in a DA-to-LEW strain combination. Immunosuppression was administered from day 0 to +13. Functional parameters such as survival time, body weight, and serum bilirubin levels were measured and the liver grafts were evaluated histologically.Results.A stepwise tapering of CsA from 3 to 0.25 mg/kg/day reduced the long-term survival rate. All animals died at a CsA dosage of 0.25 mg/kg/day, which was therefore defined as subtherapeutic. Monotherapy with the anti-CD25 mAb was performed at dosages of 600 and 1800 &mgr;g/kg/day. The lower mAb dosage resulted in a long-term survival rate of 12% and was defined as subtherapeutic. The combination therapy of CsA (0.25 mg/kg/day) and anti-CD25 mAb (600 &mgr;g/kg/day) produced a synergistic effect and led to a long-term survival rate of 84%. This survival rate was significantly higher than those after either CsA (P<0.005) or anti-CD25 mAb (P<0.001) monotherapy. Both dosages (10 and 30 &mgr;g/kg/day) of anti-CD54 mAb monotherapy as well as anti-CD54 mAb combined with a subtherapeutic dosage of CsA were ineffective in preventing acute allograft rejection. The addition of anti-CD54 mAb (30 &mgr;g/kg/day) to combined CsA plus anti-CD25 mAb therapy (triple therapy), however, increased the long-term survival rate to 100%. In the triple therapy group there was no rejection process in the liver allografts at any time, and donor-specific tolerance could be shown by donor-specific and third-party heterotopic heart transplantation.Conclusions.The synergistic action of subtherapeutic CsA plus anti-CD25 mAb NDS-60 could be demonstrated, whereas anti-CD54 mAb only had a positive effect in a triple therapy group. Triple therapy prevented both acute and chronic rejection and induced donor-specific tolerance.

ISSN:0041-1337    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Background.Our purpose was to optimize hyperbaric pressure as a vector forex vivotransfection of antisense oligodeoxynucleotides (AS-ODN) to intercellular adhesion molecule-1 to limit reperfusion injury (RI) in cardiac allografts.We investigated the effects of increased pressure, incubation time, and AS-ODN concentrations on transfection efficiency and toxicity.Methods and results.PVG (RT1c) donor hearts were heterotopically transplanted to ACI (RT1a) recipients. Donor hearts were harvested and the various groups were treated at: (1) different pressure (1–9 atm) for 45 min with 80 &mgr;mol/liter AS-ODN; (2) different incubation times (15 min to 6 hr) at 5 atm with 80 &mgr;mol/liter AS-ODN; 3) different AS-ODN concentrations (80–240 &mgr;mol/liter) at 5 atm for 45 min. Hearts were procured 24 or 72 hr after transplantation. Transfection efficiency was determined with flourescein-labeled AS-ODN. The degree of RI was determined with biochemical and histological analysis. Increasing pressure from ambient (1 atm) pressure to pressures as high as 9 atm leads to a increase in transfection efficiency from 1.7±.5 to 62±3.9% and a reduction in RI. Increased incubation time up to 45 min increased transfection efficiency and reduced RI, but longer incubation times induced significant toxicity to the allograft. Increased AS-ODN concentrations improved transfection and reduced RI.Conclusions.Hyperbaric pressure is a safe and effective vector for the ex vivo delivery of AS-ICAM-1-ODN to rodent cardiac allografts and results in a reduction in reperfusion injury.

ISSN:0041-1337    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Introduction.Immune privilege provides a natural paradigm for potentially down-regulating allogeneic and xenogeneic inflammatory immune responses. Fas ligand has been suggested as a general underlying mechanism of immune privilege; the human Fas ligand has been shown to ligate murine Fas in vitro.Methods.In this study, we examined whether the human testicular xenograft, a presumed immune-privileged tissue would have prolonged survival in mice. In addition, in vitro and in vivo murine xenogeneic immune responses to the human testicular xenografts were characterized using MHC class I, MHC class II, CD4, CD8, CD4/8 knockout mice.Results.Unlike in rodent testis, Fas ligand mRNA is not expressed and Fas is highly expressed in human testis. Human testicular xenografts are immunogenic, and do not induce any preferential pattern of recipient systemic Th1 or Th2 cytokine bias. Interestingly, an indefinite survival of the human testicular xenografts is observed in murine MHC class II knockoutmice, whereas the human skin xenografts were rejected without a delay. In vivo murine immune responses to human testicular xenografts require a recipient MHC class II-dependent CD4 T cell-mediated process that appears to depend on B7–1/B7–2 costimulatory signals.Conclusions.Our results demonstrate that the concept of immune privilege, as defined by the expression of Fas ligand and prolonged survival after transplantation, cannot be extended to human testis. The stringent restriction of murine xenogeneic immune responses to discordant human testicular xenografts to the indirect MHC class II-dependent CD4 T cell-mediated pathway suggests a potential venue for immune modulation to induce tolerance across a discordant species barrier.

ISSN:0041-1337    

出版商:OVID    

年代:2000

数据来源: OVID