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Biomedical subjects

H Ireland

Publications and source records attributed to H Ireland.

70 records · Page 4Linked to original sources

Haemodialysis with low MW heparin: dosage requirements for the elimination of extracorporeal fibrin formation.

We have performed a dose ranging study of a low MW heparin, Kabi 2165, during haemodialysis in humans (n = 16) and compared it to a dose of unfractionated commercial heparin that has already been shown to inhibit fibrin formation. Low MW heparin administered as 5000 or 10,000 anti-factor Xa units, s.c., half an hour prior to the initiation of dialysis was unable to prevent fibrin formation in the dialyser circuit. A single bolus injection of 5000 anti-factor Xa units of low MW heparin given i.v. inhibited fibrin formation, as determined by FPA generation, for up to 4 h and permitted dialysis for 6 h. Such a bolus injection may be useful for short frequent dialyses. Infusion of low MW heparin in the same dosage regimen as unfractionated heparin, 5000 anti-factor Xa units bolus plus 1500 anti-factor Xa units/h, resulted in a progressive rise in heparin, caused by its longer half-life of elimination from the circulation, and almost completely suppressed both FPA generation and fibrin clot formation for 6 h dialysis. From these studies we calculate that infusion of this low MW heparin at a dose of approximately 4000 anti-factor Xa units bolus plus 750 anti-factor Xa units/h should be a useful regimen that will be effective in suppressing fibrin formation during prolonged dialysis, and the plasma anti-factor Xa level of low MW heparin may reflect its ability to inhibit fibrin formation, although exactly comparable anti-factor Xa levels of unfractionated commercial heparin and low MW heparins may not have identical inhibitory effects.

Clinical Trials as Topic↗

Fibrinogen Manchester. Detection of a heterozygous phenotype in the intraplatelet pool.

Family members heterozygous for the congenitally abnormal fibrinogen designated fibrinogen Manchester, A alpha 16Arg----His, have previously been shown by h.p.l.c. and amino acid analysis to release a variant fibrinopeptide, [His16]fibrinopeptide A, from plasma fibrinogen after the addition of thrombin. The present study was designed to determine if the same abnormal phenotype was also present in the intraplatelet fibrinogen pool. Fresh platelets were washed in buffers containing EDTA until it could be shown that all washable plasma fibrinogen was removed. Normal platelets were then lysed by freezing and thawing to release their intracellular proteins, which were then treated with thrombin. The fibrinopeptides, cleaved from the intraplatelet fibrinogen, could be detected by an optimized h.p.l.c. technique. Quantification of the intraplatelet fibrinogen gave a result (means +/- S.D., n = 5) of 110 +/- 30 and 90 +/- 30 micrograms/10(9) platelets, when determined by h.p.l.c. quantification of fibrinopeptide B content and fibrinogen fragment E radioimmunoassay respectively. Examination of fibrinopeptides released from the platelet fibrinogen from the family with fibrinogen Manchester with the same techniques showed elution peaks in the same positions as both [His16]fibrinopeptide A and normal fibrinopeptide A. The identity of these peaks was further substantiated by analysis of the h.p.l.c. peaks by using specific radioimmunoassay to fibrinopeptide A. Our results therefore demonstrate that platelet fibrinogen expresses the heterozygous A alpha 16His phenotype. This supports the view that the A alpha chains of platelet and plasma fibrinogen are produced from a single genetic locus.

Blood Platelets↗

Fibrinogen derivatives and platelet activation products in acute and chronic liver disease.

1. The concentration in plasma of fibrinogen derivatives fibrinopeptide A (FPA) and B beta 1-42 and the platelet release products beta-thromboglobulin (beta TG) and platelet factor 4 (PF4) have been determined in patients with acute and chronic liver disease. 2. In 21 patients with fulmiant hepatic failure on admission in grade III or IV coma the plasma FPA, B beta 1-42, beta TG and PF4 levels were significantly increased compared with those in normal control subjects. On heparinization before haemoperfusion the FPA levels returned to the normal range and during resin and charcoal haemoperfusion there were no significant changes in the coagulation or platelet factors, except for a small increase in FPA with charcoal haemoperfusion. 3. In ten patients with compensated chronic liver disease there was a significant increase in B beta 1-42 and beta TG levels but not FPA and PF4 as compared with normal controls. 4. Interpretation of the results is complicated by the possible reduced clearance of these proteins as a result of renal failure in some of the patients with fulminant hepatic failure and also by the damaged liver itself. However, these results have confirmed that disseminated intravascular coagulation can occur in both acute and chronic liver disease.

Acute Disease↗

Haemodialysis with prostacyclin (Epoprostenol) alone.

Dialysis with prostacyclin (Epoprostenol, PGI2) alone prevents platelet activation and endothelial cell stimulation but not the elevation of fibrinopeptide A (FPA), a sensitive marker of fibrin generation. The generation of FPA may explain why some patients develop clot in the dialysis circuit during PGI2-only dialysis. In combination with heparin, PGI2 augments the anticoagulant effect of the heparin as well as providing platelet protection.

Blood Platelets↗

Detection of enhanced in vivo platelet alpha-granule release in different patient groups--comparison of beta-thromboglobulin, platelet factor 4 and thrombospondin assays.

During the platelet release reaction beta-thromboglobulin (beta TG), platelet factor 4 (PF4) and thrombospondin (TSP) are released from the platelet into plasma and assays of these proteins can be used to monitor in vivo platelet activation. We have assessed their relative merits as markers of the in vivo platelet alpha-granule release reaction in a number of patient groups which have previously been shown to have elevated plasma beta TG and/or PF4 levels. It is concluded that in diseases or conditions not complicated by its reduced clearance, beta TG is the most sensitive marker of in vivo platelet alpha-granule release. However, the TSP assays may be the least ambiguous when monitoring the platelet alpha-granule release reaction in patients with renal failure who are undergoing haemodialysis with heparin anticoagulation. Under these circumstances plasma beta TG, but not PF4 or TSP, levels are elevated because of impaired renal catabolism, and the presence of a heparin-releasable reservoir of PF4 on the endothelium complicates the use of the PF4 assay. In liver failure none of these assays may accurately reflect platelet alpha-granule release because of impaired hepatic or renal elimination of the proteins.

Adolescent↗

Experimental evidence for renal catabolism of fibrin fragment beta 15-42.

The elimination of labelled human fibrin fragment beta 15-42 from the circulation has been studied in an experimental rat model. When 125I beta 15-42 was injected as a bolus into a control group of rats, its elimination from the circulation could be fitted to a two compartment exponential model. A fast initial elimination had a t1/2 of less than 2 min and this was followed by a slower component with a calculated t1/2 of 135 min. The fast component was caused by equilibration of peptide throughout the intra and extra vascular spaces and the slow component reflected the action of catabolic processes once equilibration had been attained. Rats that had undergone bilateral nephrectomy eliminated significantly less of the labelled peptide than the control animals at a given time and the t1/2 of the slow component was prolonged, but not significantly, to 159 min. Rats with ligated ureters had statistically indistinguishable elimination curves from control rats. Examination of the heterogeneity of labelled peptide in plasma samples taken during the experiments by immunoprecipitation and by gel filtration revealed progressive extensive degradation in the control and ureteral ligated rats, but less degradation in nephrectomized rats. These results suggest that beta 15-42 is eliminated, in part, from the circulation by uptake and catabolism by the kidney. It is concluded that impaired renal function may results in elevated plasma levels of beta 15-42 antigen in human renal failure without the need for an increased rate of production of the peptide.

Animals↗

The significance of fibrinogen derivatives in plasma in human renal failure.

The concentrations in plasma of fibrinogen derivatives fibrinopeptide A (FPA), beta 15-42 antigen and fragment E (FgE) antigen have been determined in patients with renal failure and compared to the concentrations of the platelet release products, beta-thromboglobulin (beta TG) and platelet factor 4 (PF4). In 'partial renal failure' (51Cr-EDTA clearance rate 4-60 ml/min) FPA, beta 15-42 antigen, FgE antigen and beta TG levels were significantly raised above a normal laboratory control group. These levels were further raised in a group of patients whose disease required regular maintenance haemodialysis (51Cr-EDTA clearance rate less than 4 ml/min). PF4 levels were not significantly raised in either group. A statistical analysis of all patient results revealed that FPA, beta 15-42 antigen and FgE antigen levels all correlated with beta TG levels but not with PF4 levels. It is known that beta TG is catabolized by the kidney but PF4 is not and that elevated beta TG levels in renal failure are caused by impaired elimination rather than increased production. These results suggest that the plasma levels of these three fibrinogen derivatives are elevated in renal disease at least in part by decreased elimination rather than by increased thrombin and plasmin activities alone.

Adolescent↗

Objective assessment of heparin requirements for hemodialysis in humans.

The abilities of four heparin regimens to inhibit activation of blood coagulation, fibrinolysis, and the platelet release reaction in humans during renal dialysis have been assessed by visible examination of the extracorporeal circulation and by use of radioimmunoassays to FPA, beta 15-42 antigen, beta gamma G, and PF4. A subcutaneous injection of 5000 IU of heparin administered 1 hour before dialysis in two patients was either unable to sustain dialysis because of excess fibrin formation or allowed elevated plasma FPA and beta gamma G concentrations during dialysis. Injection of 10,000 IU heparin s.c. in five patients could sustain dialysis in only three patients for 5 hours, and allowed progressively increasing concentrations of FPA, beta gamma G, and PF4, as well as fibrin formation, in the extracorporeal circulation. A lower dose of heparin administered intravenously (2500 IU bolus plus 1000 IU/hr) to six patients was also unable to prevent elevations in FPA and beta gamma G 4 hours and 5 hours after the initiation of dialysis. Intravenous administration of heparin at a dose of 5000 IU bolus plus 1500 IU/hr completely suppressed generation of FPA during 5-hour dialysis and was free of visible fibrin formation in the extracorporeal circulation in 13 patients. The concentration of the fibrinolytic system marker beta 15-42 antigen did not change significantly in any of the regimens, and the concentration of PF4 altered in response to infused heparin as well as to inadequate heparinization. We conclude that complete inhibition of activation of coagulation and suppression of the platelet release reaction occurs when high plasma heparin levels (greater than or equal to 0.5 IU/ml) are maintained during dialysis: lower heparin levels may be compatible with dialysis in some patients, but they allow generation of fibrin and the platelet release reaction, which pose a potential risk to the dialysis procedure and to the patient.

Aged↗

Delayed release of an abnormal fibrinopeptide A from fibrinogen Manchester: effect of the A alpha 16 Arg leads to His substitution upon fibrin monomer polymerization and the immunological crossreactivity of the peptide.

Fibrinogen Manchester is an abnormal fibrinogen with an impaired release of fibrinopeptide A (FPA) and a polymerization abnormality. In the accompanying article we have identified the amino acid substitution in fibrinogen Manchester as A alpha 16 Arg leads to His. When fibrinogen Manchester was digested with low thrombin concentrations approximately 40-50% of the total FPA content was release at a rate similar to FPA release from normal fibrinogen. The fibrin so formed exhibited an impaired polymerization of monomers. Digestion of fibrinogen Manchester with high concentrations of thrombin for prolonged times released the remaining FPA which had an abnormal retention time when studied by high performance liquid chromatography (HPLC). This fibrinopeptide has been shown previously to contain the A alpha 16 Arg leads to His substitution. fibrin resulting from this exhaustive digestion had normal polymerization of monomers. The normal and substituted FPAs were isolated by HPLC and compared in a double antibody competitive-binding assay for normal FPA. The immunological cross-reactivity of the abnormal peptide was reduced, so that approximately 5 times more abnormal peptide was required on a molar basis to displace labelled normal FPA. Normal intact fibrinogen was 10-fold less reactive (on a half molar basis) than free normal FPA and the crossreactivity of fibrinogen Manchester was measurably less than that of normal fibrinogen. It is concluded that immunological measurement alone of FPA released from abnormal fibrinogens may not give a complete description of the kinetics of peptide release if the amino acid substitution lies within the FPA sequence. The combination of radioimmunoassay and HPLC, however, provides a powerful analytical approach that should be useful in classifying and characterizing abnormal fibrinogens.

Blood Coagulation Disorders↗

Activation of coagulation and fibrinolytic systems following stroke.

When the human blood coagulation and fibrinolytic systems are activated thrombin cleaves fibrinopeptide A (FPA) and plasmin cleaves b beta1 leads to 42 from fibrin(ogen). Elevated plasma concentrations of FPA and B beta 1 leads to 42 are evidence for enhanced thrombin and plasma activities in plasma. We have determined the plasma concentrations of FPA and B beta 1 leads to 42 in patients who have had thrombotic stroke. Patients who were studied immediately following stroke were found to have greatly elevated plasma FPA and B beta 1 leads to 42 levels, but these decreased to the concentrations found in an apparently healthy age-matched control group 1 month after the infarct. In contrast, the plasma concentrations of the platelet release product beta-thromboglobulin (beta TG) were slightly, but significantly, elevated immediately following the stroke and these did not alter with time after the infarct. It is concluded that following thrombotic stroke increased thrombin and plasmin activities are to be found in plasma. These increased protease activities are probably not directly associated with an increased in vivo platelet release reaction and may be useful in deciding which patients are at risk of reinfarction or stroke progression.

Aged↗

In vivo platelet release in myeloproliferative disorders.

The in vivo platelet release reaction in 22 patients with myeloproliferative disorders has been studied by measuring plasma concentrations of the platelet release product beta-thromboglobulin (beta TG). Mean beta TG and mean beta TG: whole blood platelet count ratio were significantly raised in the patient group taken as a whole compared to an age matched control group. No significant increases were observed in the plasma concentrations of thrombin and plasmin sensitive fibrinogen fragments fibrinopeptide A (FpA) and B beta 1-42. The patients were divided into those who had normal, increased or decreased responses to in vitro ADP-induced platelet aggregation. Mean beta TG and the mean beta TG: whole blood platelet count ratio were higher in the increased and decreased responders to ADP than in the normal aggregation group, but the differences in means were not statistically significant. Aspirin given to six patients at a dose sufficient to eliminate the secondary phase of ADP-induced platelet aggregation reduced mean beta TG and the mean beta TG: whole blood platelet count ratio but did not alter mean FpA and B beta 1-42. It is concluded that the enhanced platelet release reaction seen in myeloproliferative disorders is independent of plasma protease activity that arises when coagulation and fibrinolytic systems are activated.

Aged↗

Proteolysis of fibrinogen in healthy volunteers following major and minor vivo plasminogen activation.

Changes in immunoreactive fibrinopeptide A, fragment B beta 1-42 and fragment E were followed after major and minor in vivo plasminogen activation, after infusion of streptokinase and acylated streptokinase-plasminogen complex respectively, in healthy male volunteers. Major activation resulted in a dramatic rise in all three peptides, fragment E persisting in the circulation longest. Fragment B beta 1-42 was also markedly increased after minor systemic plasminogen activation despite apparently adequate levels of immediate antiplasmin.

Anistreplase↗

Plasma concentrations of fibrinopeptide A, fibrinogen fragment B beta 1-42 and beta-thromboglobulin following total hip replacement.

Plasma concentrations of thrombin sensitive peptide fibrinopeptide A (FpA), plasmin sensitive fibrinogen fragment B beta 1-42 and the platelet release product beta-thromboglobulin (beta TG) have been measured in 36 patients before and after total hip replacement. Statistically significant elevations of all three activation products were observed in the days following operation. There were small differences in plasma concentrations of FpA, B beta 1-42 and beta TG in patients who did (n = 13) and did not (n = 23) develop post operative deep vein thrombosis, as assessed by ascending venography on post operative day 10, but these differences were not statistically significant. It is concluded that coagulation and fibrinolytic systems and also blood platelets are activated following total hip replacement operations. However, the formation of post operative deep vein thrombosis can not be effectively monitored by measurement of the activation products.

Adult↗

Quantitative histochemical investigations of semipermeable membrane techniques for the assay of acid phosphatase in skeletal muscle. II. Non-specific reactions in control sections.

The absorbances of skeletal muscle fibres at 530 nm in control sections of either fresh or boiled tissue, incubated according to Meijer's semipermeable membrane technique for acid phosphatase, increase linearly with incubation time. Moreover, the absorbances continue to increase even after the incubation has been terminated by immersion of the sections in formalin. The comparatively weakly absorbing yellow reaction product originally formed is transformed with 24 h of mounting the sections into a strongly absorbing purple material with an absorption maximum at 570 nm. The original yellow product seems to be hexazotised Pararosanaline adsorbed onto tissue proteins. The formation of the purple product can be prevented by treating control sections immediately after incubation with 70% ethanol for 30 min at room temperature in place of formalin. Ethanol stabilises the yellow non-specific reaction product but does not extract it from tissue sections. The ethanol treatment thus seems useful for improving the validity of Meijer's technique.

Acid Phosphatase↗

The thrombomodulin gene mutation G(127)-->A (Ala25Thr) and cerebrovascular disease.

BACKGROUND AND PURPOSE: Thrombomodulin is an integral part of the protein C anticoagulation pathway, and polymorphisms of its gene have been implicated in thrombosis. The point mutation G(127)-->A has recently been found to be associated with myocardial infarction. METHODS: We investigated this mutation in 465 patients with acute stroke and 353 control subjects. Genomic DNA containing the region of interest was amplified by PCR, and differing genotypes were identified by RFLP. RESULTS: The A allele frequency was not statistically significantly different in the two groups, being 0.5% in the stroke group and 0.7% in the control group. CONCLUSIONS: The point mutation G(127)-->A is an uncommon finding and, in this population, is unlikely to be a major risk factor for cerebrovascular disease.

Aged↗