摘要:

In the mid-1980s, methods for making defined mutations to the genome of living cells were shown to be applicable in mammalian cells. These methods became known as gene targeting and were soon combined with techniques for manipulating totipotent mouse embryonic stem (ES*) cells, allowing researchers to generate mice homozygous for genetic changes that had been engineered in the test tube. Such targeted mice can display phenotypes that provide valuable information on gene function or can serve as models of human genetic disease. This powerful technology has already had a huge impact on a wide spectrum of biological disciplines. To date, most targeting experiments have involved disruption of the target gene, but methods for the creation of subtle, conditional, or tissue-specific mutations have been developed and are likely to become increasingly useful. Further in the future, gene targeting may well be used for the genetic modification of human somatic stem cells and ES cells from animals other than mice (e.g., pig or sheep) with major implications for autologous transplantation and xenotransplantaion, respectively. Our purpose in this brief overview is to survey established and emerging gene targeting methods, their use in understanding graft rejection, and potential applications in clinical transplantation. For a wider discussion of the techniques and their applications, the reader is referred to other reviews(1-5).

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.L-Arginine (L-Arg), the substrate for nitric oxide (NO) synthase producing NO, and the NO synthase inhibitor, N-nitro-L-arginine-methyl ester(L-NAME), have both seen shown to modify acute cyclosporine (CsA)-induced intrarenal vasoconstriction. However, the mechanism of chronic CsA nephrotoxicity characterized by progressive tubulointerstitial fibrosis (TIF) remains unclear. Thus, we examined the pathogenetic role of NO in a rat model of chronic CsA nephropathy.Methods.Rats were given vehicle, CsA (7.5 mg/kg), CsA + L-Arg (1.7 g/kg), CsA + D-arginine (1.7 g/kg), and CsA + L-NAME (3.5 mg/kg) for 28 days on a low-salt diet. NO production, glomerular filtration rate (GFR), blood and urine chemistry, and histology were assessed.Results.L-Arg treatment significantly enhanced NO biosynthesis and protected animals from impaired GFR and development of TIF induced by CsA, whereas D-arginine did not. In contrast, L-NAME strikingly reduced urinary NO and worsened both GFR and TIF compared to the CsA alone group, whereas L-NAME did not change renal function and histology in the vehicle group.Conclusions.Chronic CsA nephrotoxicity can be aggravated by NO blockade and ameliorated by NO enhancement, suggesting that NO has an important role in the mechanism of chronic CsA nephropathy.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.In contrast to adult hepatocytes, fetal hepatocytes (FH) are thought to be highly proliferative, less immunogenic, and resistant to cryopreservation and ischemic injury. These qualities could enhance FH engraftment, proliferation, and gene transfer requiring active DNA synthesis.Methods.Rat FH were obtained using the nonperfusion collagenase/DNase digestion method. Free and cultured cells were studied using electron microscopy, fluorescence-activated cell sorting, and Northern analysis usingα-fetoprotein and albumin as markers of hepatocyte lineage. DNA synthetic activity was measured in quiescent and mitogen-stimulated fetal and adult hepatocytes by [3H]thymidine incorporation. Susceptibility of cultured FH to retrovirally mediated gene transfer was studied using an amphotropic retroviral vector carrying theEscherichia colilac-Z gene. Nagase analbuminemic rats were used as recipients to study the effects of intraportal FH transplantation. Analysis of serum albumin was carried out by enzyme-linked immunosorbent assay.Results.In fetal liver, 87±2% of the cells showed morphological and molecular features of hepatocytes. DNA synthetic activity in nonstimulated cultured FH was 10 times greater than the maximal hepatocyte growth factor-driven response in adult rat hepatocytes. A total of 5-15% FH stained positive for X-gal; results of transduction in adult hepatocyte cultures were negative. In Nagase analbuminemic rat recipients, FH produced significant amounts of albumin only when a hepatic regenerative stimulus was applied. Immunohistochemistry confirmed presence of albumin-positive hepatocytes.Conclusions.Fetal rat liver from the late gestation period is highly enriched with hepatocyte progenitors. They are highly proliferative and susceptible to retroviral transduction and can engraft and function in the adult rat liver if transplanted under a hepatic regenerative stimulus.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Although agents that inhibit complement activation may be beneficial in discordant xenotransplantation, it is not known whether local complement activation occurs and is deleterious after isogeneic lung transplantation. Lungs were harvested from Lewis rats subjected to 4 °C 6-hr preservation followed by transplantation into strain-, gender-, and weight-matched recipients. Transplanted lungs demonstrated increased immunostaining for C5b-9 compared with nontransplanted controls, confirming local complement activation in this isograft model. To investigate the physiologic relevance of complement activation in the transplanted lung, the native pulmonary artery was ligated after transplantation, and pulmonary vascular resistance (mmHg/ml/min), arterial oxygenation (mmHg), graft neutrophil infiltration (myeloperoxidase activity, ΔAbs 460 nm/min), and recipient survival were measured at 30 min. Animals received either saline (control; n=22) or soluble complement receptor type-1 (sCR1, 15 mg/kg; n=19) 2 min before reperfusion. Animals treated with sCR1 showed a marked reduction in serum complement hemolytic activity (CH50; 90% lower than that of control animals,P<0.001). Compared with controls, sCR1-treated animals showed reduced pulmonary vascular resistance (2.9±1.1 vs. 8.5±1.5 mmHg/ml/min,P<0.05), improved arterial oxygenation (194±34 vs. 91±17 mmHg,P<0.05), decreased neutrophil infiltration (35% decrease,P<0.005), and improved recipient survival(74% vs. 23%,P<0.005). In parallel with the reduction in complement hemolytic activity in sCR1-treated animals, immunohistology of the transplanted lung revealed decreased C5b-9 deposition compared with controls. Taken together, these data indicate that complement activation occurs after lung preservation and transplantation in an isograft model, and that inhibiting complement activation improves outcome after transplantation.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

BackgroundThis study sought to (i) investigate the efficacy of cobra venom factor (CVF) in preventing hyperacute rejection (HAR) after pig-to-baboon heart transplantation, (ii) examine the effect of additional splenectomy (Spx) and pharmacologic immunosuppression (IS), and (iii) study delayed graft rejection when HAR is avoided by complement depletion.MethodsEleven recipient baboons received heterotopic pig heart transplants. Three received either no therapy or IS (cyclosporine + methylprednisolone± cyclophosphamide ± methotrexate) at clinically well-tolerated doses, with graft survival for only 40, 32, and 15 min, respectively. Two received CVF±Spx, which extended survival to 5 and 6 days, respectively. Six underwent Spx + CVF therapy + IS; graft survival was 3 hr(technical complication), 6 days (death from sepsis), 10, 12, and 22 days(vascular rejection), and <25 days (euthanized for viral pneumonia with a functioning graft that showed histopathologic features of vascular rejection).ResultsDense deposition of IgM and, to a lesser extent, IgG and IgA were seen on the endothelial cells within 1 hr of transplantation, but only trace levels of complement deposition were present in CVF-treated recipients. Within approximately 5-12 days, cardiac xenografts showed progressive infiltration by mononuclear cells, consisting primarily of activated macrophages producing tumor necrosis factor-α and small numbers of natural killer cells; T and B cells were absent.ConclusionsWe conclude that (i) CVF prevents HAR, (ii) the addition of Spx + IS delays rejection, but (iii) the early deposition of antibody leads to progressive graft injury, resulting in (iv) delayed vascular rejection. Our findings indicate that the features of delayed xenograft rejection described in small animal models also occur in the pig-to-baboon model, and that rejection may occur in a complement-independent manner from the effects of antibody and/or host macrophages.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.Despite numerous reports published since the early 1970s, it is frequently asserted that quality of life (QOL) outcomes of transplantation have seldom been investigated and/or that little is known about QOL. This view may have persisted due to lack of adequate cumulation and synthesis of existing data. We performed an exhaustive, quantitative literature review to determine the nature and degree of any QOL benefits associated with transplantation in adults.Methods.All independent, peer-reviewed empirical, English-language QOL studies were retrieved for six areas of transplantation: kidney, pancreas/combined kidney-pancreas, heart, lung/combined heart-lung, liver, and bone marrow. Studies' findings were analyzed to determine whether the weight of evidence suggested that (a) QOL improved from pre- to post-transplant, (b) transplant recipient QOL was better than that of patient comparison groups, and (c) recipient QOL equaled that of healthy nonpatient samples.Results.A total of 218 independent studies, evaluating a total of approximately 14,750 patients, were identified. The majority of studies demonstrated statistically significant (P<0.05) pre- to posttransplant improvements in physical functional QOL, mental health/cognitive status, social functioning, and overall QOL perceptions. The majority documented physical functional and global QOL advantages for transplant recipients relative to ill comparison groups. The studies did not indicate that recipient QOL in specific functional areas equaled that of healthy, nonpatient cohorts, although global QOL perceptions were often high.Conclusions.Although transplantation may not restore to the patient the“normal” life he/she may once have had, convergent evidence from six areas of transplantation, a variety of study designs, and demographically diverse study cohorts suggests that there are distinct QOL benefits of transplantation. Future work is required to identify background and personal factors that influence the degree of QOL benefits that any individual patient realizes from transplantation.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.Patients must wait increasingly longer periods on the kidney waiting list (WL) before receiving a transplant. Although patients can be maintained on dialysis, many deaths occur while waiting. To determine whether the risk of mortality on the WL is different from that related to the transplant procedure, data from the Organ Procurement and Transplantation Network and Scientific Registry were used to analyze all adult patients entered on the United Network for Organ Sharing (UNOS) kidney WL for a primary transplant between April 1, 1994, and December 31, 1994 (n=9925).Methods.To account for the time spent on the WL before transplant, a time dependent, nonproportional hazards model was used to assess the risk of mortality after transplant for both well-matched (zero to two HLA mismatches) and poorly-matched (three to six HLA mismatches) transplants compared with the mortality risk of remaining on the WL. This model incorporated an exponential decay component to account for the transient increased risk after kidney transplantation. Patients were stratified by age, race, creatinine level, panel-reactive antibody at listing, and blood group.Results.Although there was an increased risk of mortality in the initial posttransplant period, the risk of mortality at 1 year for transplanted patients was 59% (three to six mismatches) to 67% (zero to two mismatches) less than that of patients who remained on the waiting list for an additional year.Conclusions.Kidney transplantation is more beneficial than remaining on the waiting list. Even poorlymatched kidneys provided a significant reduction in the risk of mortality by 6 months as compared with the mortality risk of continuing to wait. Patients receive the maximum benefit when transplanted with wellmatched kidneys.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.Numerous studies have demonstrated that renal allograft survival is reduced in African-Americans (AAs). This posthoc racial subgroup analysis (AAs vs. non-AAs) tested whether mycophenolate mofetil (MMF) might have favorable implications for the treatment of AA renal allograft recipients.Methods.Patients received a triple therapy regimen of corticosteroids, cyclosporine, and azathioprine (AZA) 1-2 mg/kg/day, MMF 2 g/day (MMF 2 g), or MMF 3 g/day (MMF 3 g).Results.AAs in the AZA group had the highest biopsyproven rejection/treatment failure (BPR/TF) rate (57.5% vs. 43.5% for non-AAs). AAs in the MMF 3 g group showed a significant reduction in BPR/TF (57.5% vs. 24.2%,P=0.0008). BPRs were more frequent for AAs in either the AZA (47.5%) or MMF 2 g group (31.8%) than in the MMF 3 g group(12.1%), whereas rejections were reduced for non-AAs receiving either MMF dosage (AZA, 35.5%; MMF 2 g, 15.7%; MMF 3 g, 18.8%). AAs in the AZA group experienced BPR/TF earlier than AAs in the MMF 3 g group (median onset at 64 days vs. > 183 days,P=0.0012). But AAs in the MMF 3 g experienced BPR/TF the latest among the six subgroups of treatment and race. AAs had more severe rejection episodes and higher serum creatinine levels at 6 months after transplant, regardless of treatment group.Conclusions.Dose-dependent prevention of acute rejection in AAs is best afforded by a dosage of MMF at 3 g/day, whereas 2 g/day provides a superior benefit/risk ratio for non-AAs. MMF at 3 g/day thus provides an improvement over conventional immunosuppressive strategies in reducing the frequency of acute rejections in this immunologically high-risk group.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.Previous studies comparing renal function in diabetic subjects receiving either a kidney or kidney-pancreas transplant generally have indicated no differences; however, these studies have been limited by inclusion of either a small number of patients or selected patients followed for relatively short periods of time.Methods.To compare long-term renal function and factors affecting renal function in type I diabetic patients receiving either kidney or kidney-pancreas transplants, the slopes of regression lines generated by plotting the reciprocal of serum creatinine (1/Cr) versus time were measured in 109 consecutive patients followed for at least 12 and up to 102 months after transplantation. Multivariate analyses included linear regression using the slope of 1/Cr versus time as the dependent variable and logistic regression using a positive or negative slope as the dependent variable.Results.Significant differences between kidney-pancreas (n=64) and kidney recipients (n=45) included a smaller proportion of African-Americans, lower rates of HLA matching, lower levels of panel-reactive antibodies, shorter cold ischemia times, a lower incidence of delayed graft function, and a higher incidence of acute renal allograft rejection episodes in the kidney-pancreas group. Trough cyclosporine blood levels were significantly higher in the kidney-pancreas group for the first 12 posttransplant months. The slopes of 1/Cr versus time were negative in each group with a trend toward a more negative slope in the kidney-pancreas group. Multivariate analyses indicated that a concomitant pancreas allograft did not influence long-term renal function. The total number of renal rejection episodes was the best independent predictor of a negative slope of 1/Cr versus time. However, use of OKT3 for the treatment of rejection within the first 3 months of transplantation exerted a surprisingly beneficial effect on long-term renal function, a phenomenon that was most apparent in the kidney-alone group.Conclusions.The frequency and timing of acute rejection episodes are more important than the influence of a simultaneously transplanted pancreatic allograft in determining long-term function of the transplanted kidney. A concerning trend toward late deterioration of renal function in kidney-pancreas recipients suggests that the benefits of sustained euglycemia, shorter cold ischemia times, lower rates of sensitization, and early use of OKT3 ultimately may be outweighed by the negative effects of more frequent renal rejection episodes.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID

摘要:

Background.Previous studies have identified more morbidity in simultaneous pancreas-kidney (SPK) transplant recipients compared with kidney alone (KA) recipients. With the development of novel immunosuppressive drugs, studies are needed to determine optimal treatment regimens in specific patient populations.Methods.We retrospectively compared short-term outcome in diabetic patients receiving either SPK or KA transplantation from December 10, 1991, to July 31, 1996. The SPK recipients received either cyclosporine (CsA) + azathioprine(AZA), FK506+AZA, or FK506 + mycophenolate mofetil (MM). KA group patients received either CsA+AZA or CsA+MM.Results.Recipients of SPK instead of KA transplants were younger, had a longer mean length of stay, had a decreased incidence of delayed graft function, and had more readmissions. There were no significant differences in serum creatinine at 1, 2, and 3 years after transplantation, number of rejection episodes and infections, incidence of kidney graft loss and patient death, and 1- and 3-year actuarial patient and kidney graft survival rates between the two groups.Diabetic SPK patients receiving FK506+MM had a higher mean 3-month creatinine clearance (calculated), compared with recipients of CsA+AZA or FK506+AZA. Diabetic patients after KA transplantation who received CsA+MM demonstrated fewer rejection episodes and graft losses, although differences did not reach statistical significance.Conclusions.(1) Diabetic SPK recipients have decreased rates of delayed graft function and more readmissions compared with diabetic KA recipients. (2) There is no difference in: serum creatinine levels up to 3 years after transplantation, number of rejection episodes or infections, and 1- and 3-year patient and graft survival rates between SPK and KA recipients. (3) Short-term outcome is improved in diabetic recipients of SPK and KA transplants receiving MM instead of AZA.

ISSN:0041-1337    

出版商:OVID    

年代:1997

数据来源: OVID