摘要:

We have shown that a rare population of regulatory CD4+ T cells plays a key role in the acquisition of infectious tolerance in rat sensitized recipients of cardiac allografts pretreated with nondepleting anti-CD4 mAb. This study was designed to analyze the TCR V&bgr; expression patterns in this transplantation model. First, we used V&bgr;-specific RT-PCR to show that there was no differential usage of TCR V&bgr; genes by T cells mediating rejection or tolerance. Indeed, graft-infiltrating lymphocytes expressed most of the 22 known rat TCR V&bgr; genes in both recipient groups, suggesting unrestricted TCR V&bgr; repertoire in alloreactive T cells. Then, we applied CDR3 spectrotyping of TCR &bgr;-chain to assess the clonality of T cells at different anatomic sites. CDR3 size restriction, indicative of the presence of T cell clones, was observed in graft-infiltrating lymphocytes but not in draining lymph nodes or spleen of tolerant hosts. Consisent with the clonal expansion, T cells in tolerated grafts exhibited the memory phenotype at a much higher percentage as compared with peripheral lymphoid organs. Moreover, in tolerated graft-infiltrating lymphocytes, the CD3 size restriction occurred in limited V&bgr; gene families, with V&bgr;8.1 and V&bgr;18 most frequently detected. Hence, T cells at the graft site of tolerant recipients contain T cell clones expressing selective V&bgr; genes. This phenotypic characteristics of the tolerogenic GILs may potentially be used as a novel marker to identify operational regulatory T cells in organ allograft recipients.

ISSN:0041-1337    

出版商:OVID    

年代:2001

数据来源: OVID

摘要:

Background.The inability to diagnose early rejection of an islet allograft has previously proved to be a major impediment to progress in clinical islet transplantation. The need to detect early rejection will become even more relevant as new tolerance-inducing protocols are evaluated in the clinic. We explored three novel approaches toward development of early diagnostic markers of islet rejection after islet allotransplantation.Methods.(a) Canine islet allograft transplant recipients were immunosuppressed for 1 month, then therapy was withdrawn. Serum glutamic acid decarboxylase antigen (GAD65), an endogenous islet protein, was monitored daily with a CO2release assay. (b) Rodent islets were genetically engineered to express a unique foreign protein (&bgr;-galactosidase) by using adenoviral vectors, and after allograft transplantation, the viral-specific protein was measured in serum using optical luminescence. (c) Rodents receiving islet allografts were immunosuppressed temporarily, and daily glucose tolerance tests were followed until graft failure occurred.Results.(a) Although serum monitoring of GAD65antigen demonstrated elevated levels preceding loss of graft function in preliminary studies, the effect was not reproducible in all animals. (b) Genetically engineered rodent islets demonstrated normal insulin kinetics in vitro (insulin stimulation index 2.57±0.2 vs. 2.95±0.3 for control islets,P=ns), and purified viral protein products had a stable half-life of 8 hr in vivo. After islet allotransplantation, there were two peak elevations in serum viral proteins, confirming that an intra-islet “sentinel signal” could be detected serologically during acute rejection. There was no lead-time ahead of hyperglycemia, however. (c) Daily sequential intravenous glucose tolerance (IVGT) tests demonstrated evidence of allograft dysfunction (decline in KG) with a 2-day lead time to hyperglycemia (2.58±0.3 vs. 1.63±0.2%/min, respectively,P<0.001), with an accuracy of 89%, sensitivity of 78%, and specificity of 95%.Conclusions.Of the three diagnostic tests, metabolic assessment with an abbreviated IVGT was the most effective method of demonstrating early islet dysfunction due to rejection.

ISSN:0041-1337    

出版商:OVID    

年代:2001

数据来源: OVID

摘要:

Background.Our purpose was to investigate the occurrence and the evolution of apoptosis of enterocytes during acute and chronic rejection in an experimental model of allogeneic heterotopic small bowel transplantation (SBTx).Methods.Forty-five rats were divided in 10 experimental groups according to the dose of FK506 administration and donor bone marrow infusions (DBMI). Groups 1 and 2 did not received BMI. Groups 3 and 4 received 150×106cells at day 0, groups 5 and 6 received 75×106cells at days 0–4, groups 7 and 8 received 75×106cells at days 4 and 10, and groups 9 and 10 received 30×106cells at days 4, 10, 15, 20, and 25. Animals of groups 1, 3, 5, 7, and 9 were immunosuppressed with 0.5 mg/kg FK 506, although the remaining groups with 1 mg/kg FK 506, from day 0 to 4 after transplant. Fragment end labeling of DNA was used to detect apoptosis.Results.The number of apoptotic cells detected was highest at day 15 (184±154) and then progressively decreased thereafter (day 30=159±197; day 45=80±167; day 60=0). The number of apoptotic enterocytes was found increased during mild (151±108) and moderate (281±161) allograft rejection, although a low apoptotic rate was observed in cases without rejection (59±13) and during severe (53±131) and chronic rejection (46±136). Furthermore the number of labeled cells was found inversely correlated with fibrosis (P<0.0001). There was no correlation between apoptosis and the presence or absence of DBMI; however, at day 15 rats receiving 1 mg/day of FK 506 had a significantly lower number of apoptotic cells detected (127±103 vs. 233±174;P<0.02).Conclusions.In this study the number of apoptotic cells correlated positively with mild and moderate rejection episodes. In case of severe and chronic rejection a low apoptotic rate was found due probably to extensive necrosis and fibrosis of the mucosa. These data suggest an important role of apoptosis in acute and chronic intestinal rejection in a rat model of intestinal transplantation. Determination of apoptosis in allografts might represent an early sign of small bowel rejection and a useful marker in defining the degree of rejection and its outcome/prognosis.

ISSN:0041-1337    

出版商:OVID    

年代:2001

数据来源: OVID

摘要:

A new strategy for organ allografts that does not require recourse to immunosuppressants is established in mice. The strategy includes sublethal (7 Gy) irradiation followed by the injection of donor bone marrow cells (BMCs) via the portal vein (P.V.) and organ allografts 1 day after irradiation. Irradiation doses (≤7 Gy) are found to allow the recipients to survive without the need to reconstitute the BMCs, as the recipient hematolymphoid cells can gradually recover. One hundred percent of recipients irradiated with 7 Gy followed by either P.V. or i.v. injection of donor BMCs accept organ allografts (the skin, pancreas, and adrenal glands) for more than 1 year. However, organ allograft survival rates decrease when irradiation doses are reduced; the skin graft survival rate of mice treated with 6.5 Gy and P.V. injection of BMCs is 79%, whereas that of mice treated with 6.5 Gy and i.v. injection is 50%, indicating that the P.V. injection of BMCs induces persistent tolerance more effectively than the i.v. injection. H-2 typing reveals that almost all the hematolymphoid cells (>98%) in the peripheral blood and hematolymphoid organs are donor-derived even 1 year after the treatment (7 Gy and P.V.). The T cells are tolerant to both donor-type and host-type MHC determinants. The major mechanism underlying the persistent tolerance induced by this strategy seems to be because of clonal deletion. This simple and safe strategy would be of great advantage for human organ transplantation.

ISSN:0041-1337    

出版商:OVID    

年代:2001

数据来源: OVID

摘要:

Background.In a pig liver transplantation model, we compared the effects of Celsior solution (CS), an extracellular preservation solution, with Viaspan (University of Wisconsin solution, UW) on graft function and animal survival.Methods.Pig livers were flushed with either CS or UW solution and cold-stored for 12 hr (group 1) or for 8 to 10 hr (group 2). Grafts were transplanted orthotopically. Intrahepatic reduced and oxidized glutathione and adenine nucleotides were evaluated 1 hr after reperfusion. Liver function of transplanted animals was monitored for up to 6 days by serum transaminases, total bilirubin, purine nucleoside phosphorylase, and prothrombin levels.Results.In group 1, all animals died within 24 hr after reperfusion regardless of the preservation solution used. In group 2, no significant difference was seen in survival between the CS (72%) and the UW (67%) groups 6 days after transplantation, and there were no statistically significant differences in the biochemical data. There were no differences in histological evaluation of the livers at the time of death or killing of the animals between the CS and UW groups.Conclusion.Within the limits of this pilot study, CS is equivalent to UW in terms of graft function and animal survival.

ISSN:0041-1337    

出版商:OVID    

年代:2001

数据来源: OVID

摘要:

Background.Bone marrow transplantation is reportedly effective in preventing the progression of neurological deterioration in lysosomal storage disorders, although the mechanism underlying the therapeutic effects remains to be elucidated. Recent research on stem cell biology suggests that bone marrow cells contain nonhematopoietic stem cells, including brain precursor cells. To evaluate the contribution of bone marrow cells as carriers for cell and gene therapy of neurological disorders, we studied the fate of transplanted bone marrow cells in the adult mouse brain.Methods.Bone marrow cells were genetically marked with a retroviral vector containing the green fluorescence protein gene and then transplanted into irradiated mice by either systemic infusion or direct injection. To identify cell types, brain sections were stained with specific antibodies against neuronal cell markers—neuron specific enolase for neurons, glial fibrillary acidic protein (GFAP) for astrocytes, carbonic anhydrase II (CAII) for oligodendrocytes, and ionized calcium binding adaptor molecule 1 (Iba1) for microglia—and then examined under a confocal microscope.Results.Twenty-four weeks after systemic infusion, transplanted cells expressed Iba1 but none of the other brain cell markers. Conversely, 12 weeks after direct injection, transplanted cells were stained with antibodies against GFAP, CAII, and Iba1.Conclusions.Bone marrow contains cells capable of differentiating into oligodendrocytes, astrocytes, and microglia when exposed to the brain microenvironment. Autologous bone marrow cells may be useful as carriers for ex vivo gene therapy for lysosomal disorders with neurological symptoms.

ISSN:0041-1337    

出版商:OVID    

年代:2001

数据来源: OVID

摘要:

Background.Studies of cardiac allograft arteriosclerosis, i.e., chronic rejection, have largely focused on mononuclear inflammatory cell infiltrates in the vascular wall and periphery of the occluded vessels. The purpose of this study was to investigate the role of mast cells in the development of acute and chronic rejection in rat cardiac allografts.Methods.In the acute rejection model, transplant recipients were not treated with immunosuppressants, and the grafts were removed 5 days after transplantation at the time of severe acute rejection. In the chronic rejection model, the recipients were administered triple-drug immunosuppression, and the grafts were removed 90 days after transplantation.Results.During acute rejection, the number of mast cells was not increased, but the localization pattern differed from that of syngeneic grafts. In acute rejection, mast cells were located in the perivascular region of the allografts, but in syngeneic grafts, mast cells had a more interstitial location. In the chronic rejection model, the cardiac allografts with severe intimal thickening showed large numbers of mast cells at perivascular sites of occluded intramyocardial vessels and in the interstitium. Linear regression analysis revealed a significant correlation between the numbers of perivascular and interstitial mast cells and the intensity of intimal thickening. The majority of mast cells showed positive immunoreactivity to basic fibroblast growth factor (bFGF). Macrophage bFGF expression was not so prominent, but macrophages were more frequent in numbers. Tumor necrosis factor-&agr; expression was detected mainly in macrophages and in only a few mast cells. When the intensity of arteriosclerosis was decreased by an increase in the intensity of immunosuppression, the numbers of intragraft mast cells and other mononuclear cells, and also the production of their respective cytokines, bFGF and tumor necrosis factor-&agr;, gradually diminished.Conclusions.Taken together, our data show that the intensity of intramyocardial mast cell infiltration was associated with the intensity of chronic inflammation and allograft arteriosclerotic changes, but not with acute rejection, and that mast cells, in addition to macrophages, are a major source of myocardial bFGF. The results also demonstrate that when the T-cell activation pathway is blocked using cyclosporin, the number of mast cells is decreased. Cyclosporin may have affected the cytokine production that interfered with both the mast cell-dependent initiation and the leukocyte- and mast cell-dependent amplification and progression of the immune responses influenced by mast cell-leukocyte cytokine cascades. bFGF produced by mast cells may contribute to enhanced inflammation, neovascularization, and fibrosis during cardiac allograft arteriosclerosis.

ISSN:0041-1337    

出版商:OVID    

年代:2001

数据来源: OVID

摘要:

Background.High total plasma homocysteine (tHcy) levels are accompanied by an increased risk for premature development of atherosclerosis and atherothrombosis. Adult renal transplant recipients have elevated tHcy levels. Corresponding data in pediatric, adolescent, and young adult renal transplant recipients are scarce. We investigated whether tHcy levels were elevated in stable renal transplant recipients who received kidney grafts before age 18.Methods.This cross-sectional study was conducted during routine posttransplantation follow-up. Fasting tHcy levels, serum creatinine, and lipoprotein profile were measured in 38 clinically stable renal transplant recipients with different degrees of renal function. No patient was receiving B vitamin or folic acid supplementation. Estimated glomerular filtration rate (GFR) was assessed according to Schwartz’s formula. All patients followed a triple-drug immunosuppressive regimen, with the exception of three patients (deflazacort and azathioprine). Forty-one apparently healthy subjects constituted the control group. tHcy levels were determined by fluorescence polarization immunoassay in an IMx analyzer.Results.Mean tHcy levels in transplant recipients were significantly higher than in controls (16.8±8.7 &mgr;mol/L and 9.5±2.3 &mgr;mol/L, respectively;P<0.01). A significant positive correlation between tHcy and serum creatinine levels was observed for both transplant recipients (rS=0.70,P<0.01) and controls (rS=0.54,P<0.01). In transplant recipients, tHcy correlated negatively with estimated GFR (rS=[minus]0.47,P<0.05). Fasting tHcy levels in excess of 14.6 &mgr;mol/L (>95thpercentile in controls) were present in 19 (50%) patients; 14 of these patients had an estimated GFR<60 ml/min per 1.73 m2. When the renal transplant recipients were analyzed by renal function, mean tHcy was significantly higher in patients with an estimated GFR<60 ml/min per 1.73 m2compared with patients with an estimated GFR≥60 ml/min per 1.73 m2(20.5±9.9 vs. 13.2±5.8 &mgr;mol/L,P<0.01). Both groups were significantly different from controls (P<0.01). No relationship was found between tHcy level and either cumulative cyclosporine or cumula-tive methylprednisone doses. No differences were observed in tHcy levels or lipoprotein profile between patients who were receiving deflazacort and those on methylprednisone.Conclusions.Hyperhomocysteinemia in renal transplant recipients is a common condition. Testing for fasting tHcy level might be a useful tool to identify patients at increased risk for development of vascular disease.

ISSN:0041-1337    

出版商:OVID    

年代:2001

数据来源: OVID

摘要:

Background.Cigarette smoking contributes to a number of health-related problems, but its impact on renal transplant survival beyond accelerated patient death is unclear.Methods.We performed a cohort study of 645 adult renal allograft recipients from 1985 to 1995 to evaluate the relationship between smoking and graft outcome.Results.Twenty-four percent of recipients (156/645) were smokers at the time of transplant evaluation. Of these, 90% continued to smoke after transplantation. Pretransplant smoking was significantly associated with reduced overall graft and death-censored graft survival. Patients who were smokers at the time of pretransplant evaluation had kidney graft survival of 84%, 65%, and 48% at 1, 5, and 10 years, respectively, compared with graft survival in nonsmokers of 88%, 78%, and 62% (P=0.007). Pretransplant smoking adversely affected death-censored graft survival in recipients of cadaveric (P=0.02) and of living donor kidneys (P=0.02). Reduced graft survival in pretransplant smokers could not be accounted for by differences in rejection (64% vs. 61%,P=0.35). In a multivariate analysis, pretransplant smoking was associated with a relative risk of 2.3 for graft loss. Among patients with a smoking history before transplantation, death-censored graft survival was significantly higher for those who quit smoking before transplant evaluation.Conclusions.Cigarette smoking before kidney transplantation contributes significantly to allograft loss. The effect of smoking on graft outcome is not explained by increases in rejection or patient death. Smoking cessation before renal transplantation has beneficial effects on graft survival. These effects should be emphasized to patients with end-stage renal disease who are considering renal transplantation.

ISSN:0041-1337    

出版商:OVID    

年代:2001

数据来源: OVID

摘要:

BackgroundsThe changes in the basement membrane occurring in acutely deteriorated renal allografts (ADR) have not been extensively investigated. Our purpose is to elucidate the alteration of collagen IV, a main constituent of the basement membrane in ADR.Methods.Fifty biopsy specimens of ADR and 10 of chronic transplant nephropathy (CTN) were examined with two monoclonal antibodies specific for collagen IV. JK199 and JK132 are monoclonal antibodies that recognize triple helical collagen IV containing the &agr;1 chain. JK199 recognizes all the basement membrane containing [&agr;1 (IV)]2&agr;2(IV), although JK132 reacts only with a limited portion of it. In the normal kidney, JK199 reacts with the mesangial matrix, the basement membrane of Bowman‘s capsule (BBM), and the tubular basement membrane, as well as with the glomelular basement membrane (GBM). JK132 reacts with the mesangial matrix, BBM, and the tubular basement membrane.Results.In ADR, increased intensity of JK199 was observed in GBM, the mesangial matrix, BBM, the tubular basement membrane, and the interstitium. Increased intensity of JK132 was observed in the mesangial matrix, BBM, and the tubular basement membrane, but was not remarkable in GBM or the interstitium. In contrast, biopsy specimens of CTN showed increased intensity of JK132 in GBM, the mesangial matrix, BBM, the tubular basement membrane and the interstitium.Conclusion.These results suggest that collagen IV is up-regulated in ADR. Differential staining of collagen IV with JK199 and JK132 in GBM and the interstitium may contribute to diagnose CTN.

ISSN:0041-1337    

出版商:OVID    

年代:2001

数据来源: OVID