Biomedical subjects
Publications and source records attributed to D Talbot.
This report describes the evolution of the Birmingham, UK experience with pediatric liver transplantation from 1983 to present. Two hundred liver grafts were placed in 168 children less than 17 years of age. The current survival rate exceeds 80%.
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IFN-gamma increases the potential immunogenicity of vascular endothelial cells by up-regulation of intercellular adhesion molecule-1 (ICAM-1) and class I MHC antigen expression and by induction of class II MHC antigens and certain chemokines. In this study the mechanism by which the glycosaminoglycan (GAG) heparin antagonizes the activation of a model endothelium by IFN-gamma was investigated. Radioligand binding assays demonstrated that total binding of 125I-IFN-gamma to the EAhy.926 endothelial hybridoma cell line was reduced in the presence of heparin or heparan sulphate (HS); the structurally dissimilar GAG chondroitin sulphate had no effect. Treatment of the cells with chlorate, a metabolic inhibitor of GAG sulphation, was found to reduce both the subsequent binding of IFN-gamma and its ability to induce expression of class II MHC antigens. Treatment with heparinase II dramatically reduced the binding of IFN-gamma, while chondroitin ABC lyase had no effect. A cationic peptide from the C-terminal region of IFN-gamma was also found to reduce binding of intact IFN-gamma to the cells. These results appear to demonstrate that IFN-gamma is sequestered at the surface of endothelial cells by electrostatic interaction between specific basic amino acid residues and sulphated domains on HS, the most abundant endothelial GAG. This interaction is competitively inhibited by heparin, which is structurally related to HS. These observations are consistent with the model that IFN-gamma is bound by membrane-associated HS before engagement with the high-affinity receptor and signal transduction. Inhibition of the interaction between proinflammatory cytokines and membrane-associated GAG molecules may provide a mechanism for inducing clinically useful immunosuppression.
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One factor that is considered in the search for viable organs is the age of the potential donor. Retrospective data analysis on older donors (those aged 65 years and above) was performed to examine graft outcome. Data were collected on all older donors retrieved in one region from 1990 to 1995. Of those recipients transplanted locally (mean age, 57.6 years), creatinine values were recorded at 3 and 12 months. A recipient control group was selected from younger donors over the same period. Grafts were considered functional if the recipient was dialysis free. After 1 year, 24 recipients (82.3%) remained dialysis free, whereas 28 in the control group (90.3%) remained dialysis free during the same period. No significant differences were found in 1-year graft survival between kidneys retrieved from older donors versus those retrieved from younger donors. It therefore can be concluded that recovery from older donors is worthwhile.
Previous attempts to express functional DNA cytosine methyltransferase (EC 2.1.1.37) in cells transfected with the available Dnmt cDNAs have met with little or no success. We show that the published Dnmt sequence encodes an amino terminal-truncated protein that is tolerated only at very low levels when stably expressed in embryonic stem cells. Normal expression levels were, however, obtained with constructs containing a continuation of an ORF with a coding capacity of up to 171 amino acids upstream of the previously defined start site. The protein encoded by these constructs comigrated in SDS/PAGE with the endogenous enzyme and restored methylation activity in transfected cells. This was shown by functional rescue of Dnmt mutant embryonic stem cells that contain highly demethylated genomic DNA and fail to differentiate normally. When transfected with the minigene construct, the genomic DNA became remethylated and the cells regained the capacity to form teratomas that displayed a wide variety of differentiated cell types. Our results define an amino-terminal domain of the mammalian MTase that is crucial for stable expression and function in vivo.
In order to prevent cyclosporine nephrotoxicity in the ischemic kidney, pentoxifylline was used in a rat model. Seventy-two rats were divided into six groups according to treatment after right nephrectomy: Group I was the control, group II was treated with 25 mg/kg cyclosporine, group III underwent renal ischemia for 45 min, group IV was given 25 mg/kg cyclosporine and subjected to renal ischemia, and group V was subjected to renal ischemia and given 45 mg/kg pentoxifylline (repeated at 12, 36, and 48 hr), group VI underwent renal ischemia and was then given both cyclosporine and pentoxifylline. BUN, creatinine, and potassium levels were significantly elevated 24 hr after cyclosporine (group II), ischemia (group III), and cyclosporine and ischemia (group IV). Sodium levels remained unaffected. BUN levels normalized in all but groups III and IV after 48 hr. Creatinine levels normalized in all but group IV after 48 hr. Creatinine clearance fell in all groups and remained low even after 48 hr. Pentoxifylline prevented dramatic rises in BUN and creatinine and levels nearly normalized after 48 hr. It also histologically prevented extensive tissue damage seen after ischemia. In conclusion, pentoxifylline has a protective effect upon the kidney when subjected to cyclosporine in the presence of ischemia.
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The association of a positive flow cytometric crossmatch between recipient IgG directed against donor T lymphocytes and poor outcome is well described in renal transplantation. Until now, no long-term follow-up on such patients has been available. A total of 117 renal transplant patients were followed up for a period of 5 years. Of these, 21 were known to have donor T cell-directed IgG and 5 had B lymphocyte-directed IgG. Both groups of patients with these antibodies had a significantly poorer outcome at 5 years than did the group of patients without IgG (P < 0.0001 Handel Maenzel test). Patients with antibody detected preoperatively were tested again, either at the time of graft failure or at 5 years posttransplantation. The sera were tested against stored donor cells and the intensity of surface IgG compared with the preoperative levels. In those recipients who lost their grafts, the levels increased in 60% of cases but those that retained their grafts also had an increase in levels of donor-directed antibody in 50% of cases. The changing levels of antibody therefore appeared to have little relevance to outcome. However, when IgG isotypes were considered, for those who experienced graft failure and also had a gamma 3 isotype, a rise in IgG was demonstrated in all cases. Conversely, successful grafts with gamma 3 had a decline in levels between preoperative and 5-year samples in three of the four cases (p not significant).
The impetus for the development of living related liver programmes lies with donor shortage, which relates inversely to the success of generating cadaveric donors. A shrinking or non-existent cadaveric donor pool leads to an increased death rate among potential recipients awaiting transplantation. The living related liver programmes have by and large been successful, though it is accepted that there is potentially a significant risk to the donors. The technique of live donor liver transplantation is clearly here to stay, but the selection of suitable donors is between the family and the unit. Consequently, because of the lack of international guidelines, the programmes are open to abuse. Steps should be taken to establish either mechanisms of control or a worldwide register to combat this potential.
Heparan sulphate is a common glycosaminoglycan component of proteoglycans present on the luminal surface of vascular endothelium. It has been proposed that an important function of these molecules is the sequestration of a range of proinflammatory and proadhesive cytokines. Such cytokines play a vital role during lymphocyte recruitment from the blood at sites of inflammation. In this study it is shown that the effects of interferon-gamma (IFN-gamma), but not of tumour necrosis factor-alpha (TNF-alpha), are inhibited by treatment with soluble heparin. Specifically, heparin was shown to inhibit the induction of class II MHC antigens and the up-regulation of intercellular adhesion molecule-1 (ICAM-1) produced by treatment of cultured human endothelial cells with IFN-gamma. Furthermore, it was shown that heparin blocked the enhanced adhesion of T lymphocytes to IFN-gamma-treated endothelial cells. Investigation of the inhibitory effects of other GAG molecules demonstrated a requirement for heparin-like structural domains as chondroitin sulphate was unable to inhibit the function of IFN-gamma. These results may explain reported immunosuppressive properties of heparin, and are consistent with the model that heparin may compete with cell surface GAGs to bind IFN-gamma, thereby reducing effective biological activity.
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PURPOSE: To investigate the feasibility of therapeutic monitoring of etoposide at different plasma concentrations of the drug, and the resulting pharmacodynamic effects of such an approach. PATIENTS AND METHODS: Forty-nine previously untreated small-cell lung cancer (SCLC) patients received single-agent etoposide every 3 weeks by continuous infusion over 5 days. Plasma etoposide concentrations were monitored 18 and 66 hours into the infusion to permit dose modification. The first cohort of 15 patients began treatment with etoposide 2 micrograms/mL, with dose escalation to 3 micrograms/mL for cycles 3 and 4 and 4 micrograms/mL for cycles 5 and 6, toxicity permitting. The second cohort of 34 patients commenced at 3 micrograms/mL, with dose escalation to 4 and 5 micrograms/mL on cycles 3 and 5, respectively. RESULTS: Mean plasma etoposide concentration during the first treatment cycle was 93.4% +/- 26.6% of the target level at 18 hours (57% of patients within +/- 20% of the target) and 98.9% +/- 14.5% of the target level at 66 hours (82% of patients within +/- 20%). Hematologic toxicity was more pronounced in those treated with 3 micrograms/mL versus 2 micrograms/mL (median nadir neutrophil count, 1.3 v 2.6 x 10(9)/L, P = .032). Tumor responses, typically documented by the third cycle, were similar in each cohort (71% in patients commenced at 2 micrograms/mL and 70% at 3 micrograms/mL). Treatment cohort was not independently predictive of survival. CONCLUSION: Therapeutic monitoring of infusional etoposide is feasible and dramatically reduces interpatient pharmacokinetic variability. Although this was a small nonrandomized trial, the observation of different hematologic toxicity at the two starting concentrations but similar antitumor activity further suggests that these effects may be associated with different plasma etoposide concentrations.
Sixty-six patients with advanced non-small cell lung cancer have been entered into a phase I/II study of a combination of gemcitabine and cisplatin. An initial phase I portion of the study has been completed, and 16 patients have been entered using a fixed dose of gemcitabine 1,000 mg/m2 as a 30-minute intravenous infusion weekly for 3 weeks. Cisplatin was administered on day 15 following gemcitabine with appropriate prehydration and posthydration. The study was designed to allow for sequential groups of three patients to receive three dose levels of cisplatin (60 mg/m2, 75 mg/ m2, and 100 mg/m2). Dose modification and patient numbers were to be increased at any dose level if significant toxicity was observed. The regimen was well tolerated at all doses, and the final level of cisplatin 100 mg/m2 was expanded to 10 patients before the phase II portion was opened. Neutropenia (World Health Organization grade 4 in three patients) and thrombocytopenia (grade 3 or 4 in five patients) were the main hematologic toxicities recorded. These episodes were brief and uncomplicated. Grade 3 nausea and vomiting occurred in 12 patients, but was no worse than would be expected from cisplatin alone. Alopecia, when it occurred, was minimal (no hair loss in 10 patients and grade 1 or 2 in six patients). No significant renal toxicity or neurotoxicity was seen. A phase II study with cisplatin 100 mg/m2 and gemcitabine 1,000 mg/m2 has been opened, and to date 43 patients are evaluable for response. Eighteen (42%) patients have achieved partial remissions. The study will close when 50 evaluable patients have been entered.
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