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Biomedical subjects

I R MacGregor

Publications and source records attributed to I R MacGregor.

At least 19 recordsLinked to original sources
PubMed2006

Extended storage of platelets in SSP platelet additive solution.

BACKGROUND AND OBJECTIVES: The aim of this study was to investigate the effects of extended storage of pooled random platelets in SSP+ additive solution (MacoPharma). MATERIALS AND METHODS: Eight buffy coat-derived, pooled, leucoreduced platelet concentrates were prepared in 75% SSP+, 25% plasma using Fresenius/NPBI Composelect thrombocyte polishing filter (TPF) systems. Platelet concentrates were stored for 19 days in polyolefin storage bags and samples for in vitro analysis were taken at various time-points during storage. RESULTS: Platelet yields were lower than seen routinely when platelets are prepared in 100% plasma. The in vitro quality of the platelets stored in SSP+ was maintained until day 9. Glucose was depleted by day 12 and this was accompanied by a rapid fall in pCO2, a rise in pO2 and a cessation of lactate production. ATP and bicarbonate concentrations fell, the platelets began to swell and the ability to swirl decreased. Soluble P-selectin, glycocalicin, and regulated on activation, normal, T-cell expressed, and secreted (RANTES) concentrations increased, as did P-selectin expression. Loss of platelets and an increase in lacate hydrogenase concentration indicated that lysis had occurred. However, the pH remained between 6.4 and 7.4. CONCLUSIONS: The results suggest that SSP+ could be used for platelet storage for up to 9 days. However, the preparation of platelets in the additive requires some optimization. In vivo studies are required to confirm these in vitro results.

Blood Platelets↗
PubMed2004

Distribution of a bovine spongiform encephalopathy-derived agent over ion-exchange chromatography used in the preparation of concentrates of fibrinogen and factor VIII.

BACKGROUND AND OBJECTIVES: The risk of haemophiliacs contracting variant Creutzfeldt-Jakob disease (vCJD) via treatment with factor VIII concentrates is not known. Therefore, in order to determine the extent to which the vCJD agent might be removed during the preparation of factor VIII concentrate, the partitioning of a bovine spongiform encephalopathy (BSE)-derived agent was measured over the main purification step used to prepare the Scottish National Blood Transfusion Service high-purity factor VIII concentrate (Liberate). MATERIALS AND METHODS: Murine-passaged BSE (strain 301V), in the form of a microsomal fraction prepared from infected brain, was used to 'spike' a solution of factor VIII of intermediate purity. The 'spiked' starting material was subjected to solvent-detergent treatment and then to anion-exchange chromatography with Toyopearl DEAE-650M. All fractions were tested for 301V infectivity using a murine bioassay, including the procedures used to clean the ion-exchange media after use. RESULTS: BSE 301V infectivity was reduced by 2.9 log(10) in the fibrinogen fraction and by 2.7 log(10) in the factor VIII fraction. Over 99% of the added 301V infectivity remained bound to the ion-exchange column after elution of factor VIII. A large quantity of infectivity was subsequently removed by washing the ion-exchange media with 2 m NaCl. No further BSE 301V infectivity was detected in column eluates after treatment with 0.1 m NaOH or a second wash with 2 m NaCl. CONCLUSIONS: Results using a BSE-derived agent suggest that vCJD infectivity would be substantially removed by the ion-exchange process used in the preparation of fibrinogen and factor VIII concentrate. Although 301V infectivity remained bound to the ion-exchange matrix following elution of factor VIII, this appeared to be eliminated by the procedure used for cleaning the ion-exchange media after each use.

Adsorption↗
PubMed2004

Impacts and concerns for vCJD in blood transfusion: current status.

The impact of vCJD upon blood transfusion practice hinges on its lymphoreticular involvement. B lymphocytes play a key supporting role for the capture and replication of infectivity by follicular dendritic cells of the lymphoid tissue in animal models of transmissible spongiform encephalopathies (TSE) and tonsils, spleen and appendix in man can harbour vCJD infectivity, a situation not seen with the other human TSEs. Leucodepletion of blood donations in the UK was implemented to reduce possible vCJD transmission and preliminary data suggests that white cell associated infectivity will be effectively removed although plasma infectivity will not. Blood screening assays are under development but none yet are ready for application. The conformation dependant immunoassay, based on differences in secondary and tertiary structure between normal and TSE-associated abnormal prion protein, has a sensitivity now approaching the best bioassay. Even so further development is needed to detect the fg/ml levels likely in the event that vCJD blood does contain abnormal prion, which is as yet unproven. Surrogate assays, such as for erythroid associated factor, may provide additional means of identifying donors harbouring vCJD. Validation of clearance of TSEs from pooled plasma products consistently demonstrates effective removal of the agents in downscaled systems and studies comparing vCJD, BSE and scrapie agents yield similar results. Many approaches to therapy are under investigation, in cell culture and animal models, targeted to normal or abnormal prion metabolism, including chemical and immunological interventions. Efficacy of quinacrine/chlorpromazine and pentosan polysulphate in a clinical setting, and agents yet to be used, will be more accurately known following recent agreement of clinical drug evaluation protocols.

Animals↗
PubMed2002

Studies on the removal of a bovine spongiform encephalopathy-derived agent by processes used in the manufacture of human immunoglobulin.

BACKGROUND AND OBJECTIVES: There is still uncertainty over how the agent of variant Creutzfeld-Jakob disease (vCJD) would partition during the manufacture of plasma derivatives. In this study, a BSE-derived agent was used as a vCJD model to determine the extent to which infectivity could be removed by selected steps used in the manufacture of intravenous immunoglobulin (IVIG). MATERIALS AND METHODS: Murine-passaged BSE (strain 301V), in the form of a microsomal fraction prepared from infected brain, was used to "spike" the starting material in three experiments. The partitioning of BSE infectivity was measured over Fraction I+III precipitation, borosilicate microfibre depth filtration and Seitz depth filtration, with these steps being examined individually and in series. RESULTS: Most 301V infectivity partitioned into Fraction I+III (log reduction 2.1). Infectivity remaining in Supernatant I+III was reduced by AP20 glass-fibre depth filtration (log reduction 0.6) and subsequently removed to below the limit of detection by Seitz KS80 depth filtration, giving an overall log reduction of > or = 2.9 for the three steps in series. By contrast, glass-fibre depth filtration gave a log reduction of 2.4 when challenged directly with "spiked" feedstock. Seitz KS80 depth filtration gave a log reduction of > or = 3.1 when challenged directly with 'spiked' feedstock and also removed residual infectivity to below the limit of detection when applied as the final step in series. CONCLUSIONS: Results using a BSE-derived agent suggest that vCJD infectivity should be substantially removed from immunoglobulin G (IgG) solutions by Fraction I+III precipitation and Seitz KS80 depth filtration. The three different process steps examined acted in a complementary manner to one another when operated in series. However, the data demonstrated that it would be inappropriate to add together the reduction factors that had been derived for each step in isolation.

Animals↗
PubMed2001

Species differences in the blood content of the normal cellular isoform of prion protein, PrP(c), measured by time-resolved fluoroimmunoassay.

BACKGROUND AND OBJECTIVES: The concern that variant Creutzfeldt-Jakob disease could be transmitted via blood transfusion has prompted studies of blood infectivity in animal models. As normal prion protein acts as a substrate for conversion to the abnormal form associated with infectivity, we have quantified its distribution in mice and hamsters, the most commonly used animal models. MATERIALS AND METHODS: A time-resolved fluoroimmunoassay was used to measure normal prion protein in hamster and mouse tissues, including blood. RESULTS: Levels of prion protein in hamster blood were remarkably low compared with human blood. In contrast, levels in mouse blood were quite similar to human blood; however, there were differences in the distribution of normal prion between cellular and cell-free fractions. CONCLUSION: Differences between levels of normal prion in blood of animal models and humans should be considered as a possible contributor to infectivity study outcomes in these models.

Animals↗
PubMed2000

Pulmonary endothelial permeability and circulating neutrophil-endothelial markers in patients undergoing esophagogastrectomy.

OBJECTIVE: Esophagogastrectomy is an established surgical treatment for esophageal malignancy. The postoperative period may be complicated by the development of acute lung injury syndromes and thus, may provide a useful model in which to study the early pathogenic mechanisms of inflammatory lung injury. DESIGN: Open, prospective study. SETTING: High dependency and intensive therapy units. PATIENTS: Eight healthy male volunteers and 20 patients in the early postoperative period INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: The lung protein accumulation index (PAI) of radiolabeled transferrin was determined by using a portable, double-isotope system. The following circulating inflammatory markers-thought to reflect neutrophil-endothelial activation and injury including circulating neutrophil elastase-soluble L-, E-, and P-selectins and thrombomodulin and von Willebrand factor antigen were assayed from venous blood samples The PAI for healthy volunteers was median -0.5 (range, -1.73 to 0.27) x 10(-3)/min and for patients undergoing esophagogastrectomy -0.005 (range, -1.53 to 2.28) x 10(-3)/min. There was no statistical difference between the two groups. In the postesophagogastrectomy group, a significant elevation in circulating levels of neutrophil elastase, soluble P- and E-selectin, thrombomodulin, and von Willebrand factor antigen were observed relative to the control group but only circulating plasma elastase demonstrated a significant correlation with the PAI (r2 = .23, p =.03). CONCLUSIONS: The data suggest patients undergoing esophagogastrectomy develop a inflammatory response but this is not a surrogate of permeability and other factors are likely to determine persistent injury to the alveolar-capillary barrier function in this patient group.

Aged↗
PubMed1999

Preliminary assessment of whole-blood, red-cell and platelet- leucodepleting filters for possible induction of prion release by leucocyte fragmentation during room temperature processing.

Universal leucodepletion is being introduced in the U.K. to reduce a theoretical risk of Creutzfeldt-Jakob disease (CJD) transmission. If CJD infectivity is associated with leucocytes, any cell fragmentation associated with filtration could reduce the potential benefit. Four types each of whole blood, red cell and platelet leucodepletion filters were assessed after holding of blood units for at least 4 h at 22 degrees C. In all cases the mean residual leucocyte content was <1 000 000 per unit, with only two individual filtered whole blood units having a leucocyte content exceeding this. Evidence of leucocyte fragmentation during filtration was sought but not found by assay of soluble elastase, beta-thromboglobulin and normal prion protein, as well as by isotopic labelling of leucocyte external membrane. These preliminary studies indicate that it was possible to prepare leucodepleted blood components by filtration at room temperature, and that this appeared not to be associated with overt cell fragmentation. Definitive demonstration that fragmentation does not occur requires the development of improved general (non-specific) assays for cell membrane fragments.

Blood Component Removal↗
PubMed1998

Neoantigens and antibodies to factor VIII.

It is well known that 10-20% of severe haemophiliacs are likely to develop inhibitors to factor VIII, usually soon after the commencement of therapy. Two recent inhibitor outbreaks have occurred in patients treated for a number of years on switching to a product subjected to additional virus inactivation. Hence the incidence of inhibitor formation may be affected by the type of product used for treatment and the potential for processing to result in 'neoantigens'. Examination of the parts of factor VIII interacting with inhibiting antibodies, and the effect of various therapies on these, can teach us something about the mechanisms involved in antibody formation. However, the development of pre-clinical assays to assess products and processes for neoantigen formation should allow the prevention of inhibitor outbreaks. This review summarizes current in vitro and in vivo approaches to this problem, concluding that most available assays are inadequate for this purpose, with competitive immunoassay and phospholipid binding providing the most hopeful route forward.

Amino Acid Sequence↗
PubMed1997

Modulation of human endothelial thrombomodulin by neutrophils and their release products.

Pulmonary microvascular injury, one of the earliest events in adult respiratory distress syndrome (ARDS), is caused by the release of injurious products from stimulated neutrophils and other inflammatory cells in the lung microvessels. An increased level of the endothelial cell-surface anticoagulant protein thrombomodulin (TM) in plasma from patients with ARDS as shown in this study may be one consequence of this, and our objective in this investigation was to define the mechanisms by which TM is modulated by neutrophils, using an endothelial tissue culture system. Human neutrophils in contact with endothelium caused a fourfold reduction in cell-surface TM activity, but only after prior neutrophil priming and activation. Neutrophil release products elastase and cathepsin G caused rapid dose-related reductions in endothelial cell-surface TM activity, with complete abolition at 5 microg/ml in the absence of endothelial detachment. H2O2 also reduced TM activity. TM antigen accumulated in culture supernatant after treatment of endothelium with cathepsin G, indicating proteolytic release of cell-surface TM. We conclude that primed activated neutrophils are potent modulators of endothelial TM in vitro.

Adolescent↗
PubMed1996

Neutrophil activation in chronic liver disease.

OBJECTIVE: To assess the relationship between neutrophil activation and indices of disease severity in patients with chronic liver disease. METHODS: Plasma neutrophil elastase was measured by radioimmunoassay as a marker of neutrophil activation, and disease severity assessed by standard clinical, biochemical, haematological and histological techniques. PATIENTS: Eighty-eight patients with chronic liver disease were studied, Thirty-nine had alcohol-induced liver disease (ALD), 18 autoimmune chronic hepatitis, 13 cryptogenic cirrhosis, seven primary biliary cirrhosis, six primary sclerosing cholangitis, three haemochromatosis and two secondary biliary cirrhosis. Seventy-three patients were cirrhotic and 15 were non-cirrhotic, confirmed by biopsy. RESULTS: Levels of neutrophil elastase were raised in Childs C cirrhotic patients with ALD compared with Childs A or B patients with ALD (P < 0.01), Childs A or B patients with non-ALD (P < 0.01), and Childs C patients with non-ALD (P = 0.02). In patients with ALD, neutrophil elastase correlated with prothrombin time (r = 0.679, P = 0.001), bilirubin (r = 0.587, P < 0.001), Child-Pugh score (r = 0.546, P < 0.001) and inversely with serum albumin (r = -0.511, P < 0.001). In patients with non-ALD, there were no correlations with the measurements of with transaminase levels. CONCLUSION: Neutrophil activation, as measured by plasma neutrophil elastase, is a marker of disease severity in patients with alcohol-induced chronic liver damage, but not in those with other causes of liver disease.

Autoimmune Diseases↗
PubMed1996

High purity factor IX and prothrombin complex concentrate (PCC): pharmacokinetics and evidence that factor IXa is the thrombogenic trigger in PCC.

Recent studies using assays for surrogate markers of thrombogenicity in man have demonstrated that activation of the coagulation system occurs following infusion of clinical doses of prothrombin complex concentrates (PCC) but not after the same doses of high-purity factor IX concentrates (HP-FIX) in patients with haemophilia B. Here we have investigated the mechanism of such thrombogenesis by applying assays that detect early-through to late-events in coagulation system activation in a pharmacokinetic cross-over study of 50 IU/kg PCC and a new HP-FIX product in haemophilia B patients. Satisfactory recoveries and half-lives were observed for both concentrates. HP-FIX caused no increases in thrombin-antithrombin III complex (TAT), prothrombin activation peptide fragment F1+2 (F1+2), factor X activation peptide (FXAP) or factor VIIa (FVIIa). In contrast the same dose of factor IX in the form of PCC was followed by significant increases over pre-infusion levels of TAT, F1+2 and FXAP, but not FVIIa. Elevations of FIXAP occurred after both HP-FIX and PCC but did not reach normal levels and were attributed to normalisation of the FIX concentration in those patients whose levels of FIXAP were initially low. We conclude that the thrombogenic trigger associated with PCC infusion occurs at the level of factor X activation. In the absence of any increase in FVIIa, we would attribute this to the likely presence of FIXa in the PCC.

Adult↗
PubMed1995

Low proteins C and S and activation of fibrinolysis in treated essential thrombocythemia.

The aim of this study was to investigate whether abnormalities in the fibrinolytic system and in the naturally occurring anticoagulant proteins could contribute to the thrombotic risk in essential thrombocythemia. Euglobulin lysis time, fibrin plate lysis area, tissue plasminogen activator antigen, and activity and plasminogen activator inhibitor antigen were measured before and after venous occlusion in a group of 16 patients with essential thrombocythemia and in 16 healthy age and sex matched controls. In addition, resting levels of antithrombin III, D-dimer, prothrombin fragment 1 + 2, and protein C and S were assessed. The results were related to the presence or absence of a thrombotic history. The results demonstrated that the patients had a significantly elevated fibrin plate lysis area and significantly decreased plasminogen activator antigen, both at baseline and after venous occlusion. They also had significantly decreased levels of plasma protein C and total protein S. There was a modest, non-significant elevation in the plasma concentration of D-Dimer and F 1 + 2. Those patients with a history of thrombosis had significantly lower protein C levels compared with individuals without a thrombotic history. We conclude that patients with essential thrombocythemia have evidence of activated fibrinolysis in the resting state and after stimulation. This, and the decreased levels of protein C and total protein S, may be secondary to chronic clinically occult thrombosis occurring in myeloproliferative disorders.

Adult↗
PubMed1995

No detectable alterations in immunogenicity following terminal severe dry-heat treatment of high-purity factor VIII (Liberate) and factor IX (HP9) concentrates.

We used monoclonal antibody ELISAs, antigen molecular size distribution, competition ELISA and neonatal mouse immune tolerance methods to detect potential neoantigen formation and increased immunogenicity following severe dry-heat treatment of high-purity factor VIII (Liberate) and factor IX concentrates. To provide positive controls, concentrates were heated in solution (70 degrees C for 2 h) to produce denaturation on purpose. The competition ELISA applied to factor IX proved particularly useful for quantifying differences between the positive control and the dry-heated/unheated concentrates. None of the test systems employed by us indicated any detectable neoantigen formation or any alteration in immunogenicity following terminal severe dry-heat treatment of the high-purity concentrates, and this finding is supported by clinical experience so far.

Antigens↗
PubMed1994

The long-term effects of metoprolol and epanolol on tissue-type plasminogen activator and plasminogen activator inhibitor 1 in patients with ischaemic heart disease.

This double-blind, randomized parallel group study investigated the effect of 6 months beta-adrenoceptor antagonist therapy with either metoprolol (beta 1-selective without intrinsic sympathomimetic activity [ISA]) or epanolol (beta 1-selective with ISA) on markers of endogenous fibrinolysis in 20 patients with chronic stable angina receiving concurrent treatment with nifedipine. Neither drug had an effect on tissue-type plasminogen activator or plasminogen activator inhibitor type 1 (PAI-1). A significant correlation between fasting insulin and PAI-1 has previously been described and was confirmed in this study. The group treated with metoprolol showed a significant rise in fasting insulin after 6 months with no change in PAI-1. This suggests that the previously described link between these two may not be causal.

Adrenergic beta-Antagonists↗
PubMed1994

Study of endothelial t-PA and vWf in normal subjects and in von Willebrand's disease.

Some well-described similarities exist between tissue plasminogen activator (t-PA) and von Willebrand factor (vWf) which may suggest a link in either the synthesis or release of both proteins from endothelial cells. To investigate this relationship further immunocytochemical localization of t-PA and vWf was performed in normal tissues and in skin obtained from patients with type I and type III von Willebrand's disease (vWd). Components of the fibrinolytic system were measured at baseline and after venous occlusion in healthy controls and patients with vWd. Patients with severe vWd received intravenous vWf concentrate, followed by desmopressin (DDAVP), to study the plasma response of vWf and t-PA. By immunocytochemical staining, t-PA was demonstrated in endothelial cells of normal skin, kidney and liver and also in the skin of patients with type I and type III vWd. vWf was localized in endothelial cells of all tissues except the specimens from an individual with severe vWd. Basal plasma levels of fibrinolytic components were normal in patients with vWd. Venous occlusion resulted in a rise of fibrinolytic activity in controls and patients with type I, but not type III, vWd. No rise in plasma t-PA was observed following DDAVP in severe vWd, even though near-normalization of plasma vWf levels had been obtained by prior infusion of vWf concentrate. It is concluded that the synthesis of t-PA and vWf is probably regulated by independent processes. Constitutive and regulated release of both proteins occur through different mechanisms and the basal secretion of t-PA is intact in severe vWd.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗