Biomedical subjects
Publications and source records attributed to A J Barrett.
Oral Cyclosporin A (CyA) was given for the prophylaxis of graft-versus-host disease (GVHD) to sixty-seven patients receiving a bone marrow transplant, and was found to be extremely effective in reducing the severity of acute GVHD. Side-effects of CyA included hypertrichosis, gum hypertrophy, and a rare but serious 'capillary leak' syndrome.
Between 1979 and 1982, 229 patients with acute nonlymphoblastic leukaemia in remission were given allogeneic bone marrow transplants from HLA-identical siblings. Data on 183 patients transplanted in first complete remission were compared with results of 46 patients transplanted in second or subsequent remission. The 2-year actuarial survival rate was 53% for patients transplanted in first remission and 36% for patients transplanted in second or subsequent remission (P = 0.02). The probability of survival was related to the age of the patient (P = 0.02) and of the donor (P = 0.0002). The 2-year actuarial leukaemia recurrence rate was 12% for patients transplanted in first remission and 44% for patients in a subsequent remission (P = 0.10). Other factors correlating with a higher probability of relapse after transplantation were the leukaemia subtypes M4 and M5 (P = 0.007), cyclosporin A used as prophylaxis of graft-versus-host disease (P = 0.008), and the absence of chronic (but not of acute) graft-versus-host disease (P = 0.003). These data indicate that prolonged survival can be achieved in approximately 50% of patients with acute nonlymphoblastic leukaemia who receive bone marrow transplants during their first complete remission. But modifications of pre- and post-transplant treatment should be considered to reduce the rate of relapse.
Between 1979 and 1982, 192 patients with acute lymphoblastic leukaemia were given allogenic bone marrow transplants from HLA-identical siblings. Data on 57 patients transplanted in first remission were compared with the results in 96 patients transplanted in second remission, and 39 in third or subsequent remission. The majority of these patients were of the non-B non-T-ALL and T-ALL subtypes. The 2-year actuarial survival for non-B non-T-ALL was 58% for patients transplanted in first remission, 34% for second remission patients and 33% for patients transplanted in third or subsequent remission (P = 0.60). For patients with T-ALL, the survivals at 2 years for first and second remission transplant patients were 61% and 10%, respectively (P = 0.04). Multifactorial analysis demonstrated a significantly higher probability of survival for patients grafted in first remission (compared with more advanced remission states) and with bone marrow from young (age less than or equal to 20 years) donors. The 2-year actuarial risk of relapse for patients with non-B non-T-ALL transplanted in first remission was 0%, 32% and 69% for second and third or subsequent remission patients, respectively (P = 0.04). For T-ALL the actuarial risk of relapse at 2 years is 10% for first remission grafts and 48% for second remission grafts (P = 0.07). Only patients (with both non-B non-T/and T-ALL) transplanted in first remission have a high and stable probability of remaining in remission.
This study shows that hitherto unexplained thrombocytopenia and neutropenia occurring after bone marrow allografting and autografting may have an immune basis. In the case of autografts, such antibodies are, by definition, auto-antibodies, and it is suggested that similar antibodies occurring after allografting are also auto-immune.
After bone marrow grafting, severe unexplained thrombocytopenia and granulocytopenia may complicate the post-graft recovery of the patient. The present study has shown the presence of antibodies to platelets and granulocytes of donor origin in recipients of both allogeneic and autologous bone marrow grafts. In the case of autografts, such antibodies are by definition autoantibodies, and similar antibodies after allografting may also have an autoimmune origin. It is likely that this is the result of transient immune system imbalance, common to both allo- and autografts, in the early post-graft period. The extent to which these antibodies affect the peripheral counts probably depends on the ability of the engrafted marrow to compensate for the rate of antibody-mediated cell destruction.
We have investigated the effect of tosyl-lysyl chloromethane (a serine proteinase inhibitor) on rat liver cytosolic glucocorticoid-receptor complex binding to isolated nuclei. Tosyl-lysyl chloromethane (1-2 mM) significantly blocked nuclear binding when added before (but not after) thermal activation. Fifty percent inhibition occurred at about 1 mM tosyl-lysyl chloromethane. Interestingly, several other serine proteinase inhibitors (tosyl-phenylalanyl chloromethane, phenylmethyl sulphonylfluoride, and diisopropyl fluorophosphate) also depressed glucocorticoid-receptor complex nuclear binding. Tosyl-lysyl chloromethane, like molybdate, inhibited dilution-induced nuclear binding at low temperature, altered the DEAE-cellulose binding characteristics of heat-treated glucocorticoid-receptor complexes, and caused glucocorticoid-receptor complexes to sediment at about 9-10S (control complexes sedimented at 7-8S) in low salt-sucrose density gradients. Overall, these results suggest that tosyl-lysyl chloromethane modulates several properties of the glucocorticoid-receptor complex, that tosyl-lysyl chloromethane effects resemble those of molybdate, and that a serine proteinase(s) could be involved in the mechanism of glucocorticoid-receptor complex activation into a nuclear binding form.
A representative selection of X-ray contrast media was tested for their effects on the action of chymopapain on cartilage proteoglycan in an in-vitro test system. Inhibition of the action of chymopapain was detected only at high concentrations of contrast media, and was more marked with ionic contrast media than with non-ionic media. A dose-response curve for the release of glycosaminoglycan by chymopapain was linear when the amount of enzyme was plotted on a logarithmic scale against glycosaminoglycan release. In view of our results, we suggest that there is no need for a delay between injection of contrast medium and enzyme, contrary to the instructions of the distributors of chymopapain, if small amounts of a non-ionic contrast medium are used.
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Patients with severe viral infections such as Lassa or Ebola may be denied adequate laboratory investigations because of justifiable fears among laboratory staff. This study in monkeys was designed to provide comprehensive haematological and biochemical monitoring in a contained environment during all stages of Ebola infection. Marked neutrophilia, depletion of lymphocytes, and early failure of platelet aggregation preceded a consumption coagulopathy with a microangiopathic haemolytic anaemia, thrombocytopenia, and failure of prostacyclin production by vascular endothelium. Liver dysfunction was moderate but in conjunction with the dehydration and hypoalbuminaemia could be expected to precipitate renal failure and shock. It seems reasonable to anticipate successful patient support with a patient management isolator and treatment with platelet transfusions, fresh frozen plasma, and possibly prostacyclin when haemostasis is defective during this otherwise self-limiting illness.
The lysosomal cysteine proteinase cathepsin B (from human liver) was tested for its peptide-bond specificity against the oxidized B-chain of insulin. Sixteen peptide degradation products were separated by high-pressure liquid chromatography and thin-layer chromatography and were analysed for their amino acid content and N-terminal amino acid residue. Five major and six minor cleavage sites were identified; the major cleavage sites were Gln(4)-His(5), Ser(9)-His(10), Glu(13)-Ala(14), Tyr(16)-Leu(17) and Gly(23)-Phe(24). The findings indicate that human cathepsin B has a broad specificity, with no clearly defined requirement for any particular amino acid residues in the vicinity of the cleavage sites. The enzyme did not display peptidyldipeptidase activity with this substrate, and showed a specificity different from those reported for two other cysteine proteinases, papain and rat cathepsin L.
The protein from chicken egg white that inhibits cysteine proteinases, and has been named 'cystatin', was purified by ovomucin precipitation, affinity chromatography on carboxymethylpapain-Sepharose and chromatofocusing. The final purification step separated two major forms of the protein (pI 6.5 and 5.6), with a total recovery of about 20% from egg white. By use of affinity chromatography and immunodiffusion it was shown that the inhibitor is also present at low concentrations in the serum of male and female chickens. Tryptic peptide maps of the separated forms 1 and 2 of egg-white cystatin were closely similar, and each form had the N-terminal sequence Ser-Glx-Asx. The two forms showed complete immunological identity, and neither contained carbohydrate. Ki values for the inhibition of cysteine proteinases were as follows: papain (less than 1 X 10(-11)M), cathepsin B (8 X 10(-10)M), cathepsin H (about 2 X 10(-8)M) and cathepsin L (about 3 X 10(-12)M). Some other cysteine proteinases, and several non-cysteine proteinases, were found not to be significantly inhibited by cystatin. The inhibition of the exopeptidase dipeptidyl peptidase I by cystatin was confirmed and the Ki found to be 2 X 10(-10)M. Inhibitor complexes with active cysteine proteinases and the inactive derivatives formed by treatment with iodoacetate, E-64 [L-trans-epoxysuccinylleucylamido(4-guanidino)butane] and benzyloxycarbonylphenylalanylalanyldiazomethane were demonstrated by isoelectric focusing and cation-exchange chromatography. The complexes dissociated in sodium dodecyl sulphate/polyacrylamide-gel electrophoresis (with or without reduction) with no sign of fragmentation of the inhibitor. Cystatin was found not to contain a free thiol group, and there was no indication that disulphide exchange plays any part in the mechanism of inhibition.
NK cells from young normal mice are cytolytic in vitro for a virus-induced tumour cell line, YAC-1. Cytotoxicity is inhibited by the addition of unlabelled homologous YAC-1 cells and by regenerating bone marrow cells from the spleens of lethally irradiated, bone-marrow-grafted mice. Quiescent marrow from syngeneic and allogeneic mice produces little or no competition. This suggests that NK cells recognize, and may regulate, marrow progenitor cells.
Twenty-four patients have received bone marrow transplantation for severe aplastic anaemia at the Westminster Hospitals since 1974. Twelve patients are long term survivors. Infectious complications in association with graft rejection, graft versus host disease or prolonged neutropenia were the major cause of death. In the last 18 months the introduction of more effective conditioning regimes and Cyclosporin A as graft versus host disease prophylaxis has improved the survival rate to 85%. One patient has required regrafting for late graft failure without evidence of graft rejection.
The cytotoxicity of radiolabelled YAC-1 target cells by natural killer (NK) cells from the spleens of immunocompetent CBA mice is inhibited by unlabelled YAC-1 competitor cells, but not by resting bone marrow from syngeneic or allogeneic adult mice. Rapidly proliferating haemopoietic cells recovered from the spleens of lethally irradiated, bone marrow-reconstituted CBA mice, however, compete strongly in the NK assay. The competitive ability of early regenerating marrow correlates with the presence of an increased percentage of morphologically immature cells of mixed lineages. Competition declines in reconstituted spleens recovered more than 10 days after engraftment, as the proportion of immature elements falls towards that of resting marrow. Although the numbers of unlabelled YAC-1 cells required to produce equivalent competition of unstimulated and interferon-activated NK killing are similar, 10 times fewer regenerating marrow competitors compete cytotoxicity by unstimulated NK effectors to the same degree as interferon activated cells. The numbers of granulocyte-macrophage colonies formed in soft agar by regenerating marrow is also influenced by prior incubation of the marrow cells with NK effector populations. Spleen cells from homozygous athymic mice produce the same effect as cells from their heterozygous littermates. These data suggest that NK cells recognize and regulate the differentiation of progenitor elements within the marrow.
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Severe thrombocytopenia developed after an allogeneic bone-marrow graft in a 40-year-old woman in first remission from acute myeloblastic leukaemia. The thrombocytopenia had an autoimmune basis and was acquired from the bone-marrow donor who had an identical platelet autoantibody. The patient responded to high-dose intravenous IgG, and she has a normal platelet count 10 months after the graft. This study suggests that the immune defect in autoimmune thrombocytopenia can be transferred in cells capable of proliferating in a bone-marrow graft.
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Oral cyclosporin A was used as prophylaxis against graft-versus-host disease in (a) 31 patients with acute leukaemia or aplastic anaemia given transplants of HLA-matched bone marrow and (b) five patients with inborn errors of metabolism given transplants of haplotype-identical (parental) bone marrow. Twenty-six patients survived longer than two months after the operation. Despite the cyclosporin A, 31 patients (86%) suffered an acute form of graft-versus-host disease and 22 (61%) a chronic form. Nevertheless, the disease was usually treatable with immunosuppressive agents and caused the death of only one patient. Cyclosporin A caused renal toxicity in all cases; occasionally this was associated with a "capillary leak" syndrome, fatal in two patients. In children hypertension, fits, and fluid retention were common side effects. Blood concentrations of cyclosporin A correlated with blood urea values and blood pressure but did not predict the occurrence of graft-versus-host disease. Four different dose schedules were used to find the optimum way to administer this drug. Oral cyclosporin A is extremely effective at reducing the severity of graft-versus-host disease, but prevention of the disease is limited by toxicity of the drug and variable absorption. Better results might be achieved with parenteral administration or by using the drug in combination with other methods.