Showing papers in "British Journal of Haematology in 1980"

Globin chain electrophoresis: a new approach to the determination of the G gamma/A gamma ratio in fetal haemoglobin and to studies of globin synthesis.

Abstract: Separation of globin chains by electrophoresis provides a simple and rapid method for the determination of the G gamma/A gamma ratio in human fetal haemoglobin, and of biosynthetic rates of the globin chains. Whole haemolysates were analysed by electrophoresis on polyacrylamide gels in urea, acetic acid and Triton X-100. Electrophoresis of haemolysates from newborn infants led to four bands: A gamma, G gamma, beta and alpha. The identity of these bands was indicated by examination of haemoglobins of known globin chain composition. In 15 samples, the % G gamma was similar by Triton gels and by amino acid analysis of the gamma CB-3 peptide. Some mutant globin chains were also separable with the electrophoretic technique. Triton gel electrophoresis provides rapid analysis of very small amounts of haemoglobin, and permits examination of globin chain composition as well as globin synthetic ratios.

The Anaemia of P. falciparum Malaria

Abstract: The haematological changes in a group of young Gambian children with P. falciparum malaria have been analysed. In children with acute infection anaemia was most marked during the period after treatment. Although many of these patients developed a positive direct Coombs test during this period of the illness it is not clear whether the anaemia which occurs after treatment has an immune basis. A second group of children showed quite different haematological findings. They appear to have a more chronic form of P. falciparum malaria infection, were profoundly anaemic at presentation, showed gross dyserythropoietic changes in their bone marrows, and had a full reticulocyte response and rise in haemoglobin after treatment. A third group of children were encountered whose haematological abnormalities were intermediate to those of the acute and chronic groups. These findings indicate that the patho-physiological mechanisms responsible for the anaemia of P. falciparum malaria are different at different stages of the illness.

Autoimmune thrombocytopenia: detection of platelet autoantibodies with the suspension immunofluorescence test.

Abstract: Summary An immunofluorescence test on paraformaldehyde-fixed platelets in suspension was used for the detection of antibodies on platelets and in the sera of patients with idiopathic or secondary thrombocytopenia. In idiopathic thrombocytopenia, a positive test was found on the patients' platelets in 55 out of 80 cases. The indirect immunofluorescence test with the patient's serum was positive in 28 out of 80 cases. In thrombocytopenia accompanied by other autoimmune diseases, the direct test was positive in 35 out of 39; the indirect test in 13 out of 39 patients. These results are similar to those obtained with other techniques recently described in the literature. By using FITC-labelled monospecific anti-immunoglobulin reagents, immunochemical characterization of the autoantibodies was possible. Of the 90 patients with a positive direct test, 80 were analysed in this way. Of these, platelet-bound IgG was detected in 76. This was accompanied by IgM in 17 and IgA in three. In four, only IgM was found. The IgG subclasses of platelet-bound IgG were studied in 48 patients. Mainly IgG1 was found (45), often together with IgG3 (22) or IgG4 (14), but rarely with IgG2 (1). Only a few patients had just IgG3 (2) or IgG4 (1) on their platelets. Platelet-bound autoantibodies could often be eluted (41 out of 86 cases) with ether and were demonstrable in the eluate with the indirect immunofluorescence test. This test was usually positive when the direct test was strongly positive (40 out of 44) and usually negative when it was weakly positive (1 out of 24) or negative (1 out of 18).

Superoxide dismutase, glutathione peroxidase, catalase and lipid peroxidation of normal and sickled erythrocytes.

Abstract: Summary. Sickled erythrocytes showed reduced glutathione peroxidase and catalase activities in comparison to normal erythrocytes. In addition, increased levels of superoxide dismutase and ‘peroxidation potential’as well as fluorescent lipid pigments and malonaldehyde suggestive of membrane lipid peroxidation were found in sickled erythrocytes. Finally, sickled erythrocytes showed membrane ‘bound’Heinz bodies as well as reduced membrane lipids and unsaturated fatty acids. From these observations, it is suggested that membrane lipid peroxidation occurs in sickled cells.

Platelet-activating factor (PAF-acether) secretion from platelets: effect of aggregating agents.

Abstract: Summary. Platelet-activating factor is a 1.0-alkyl-2 acetyl analogue of phosphatidylcholine (PAF-acether) which triggers platelet aggregation independently from ADP release and thromboxane A2 (T×A2) formation. PAF-acether was described initially as being secreted by rabbit basophils during an immunological challenge. We have now found that it is also formed by washed rabbit platelets stimulated by the calcium ionophore A23187, thrombin and collagen. These three agents are known to trigger platelet aggregation independently from the release of ADP and from the formation of T×A2. By contrast, ADP and arachidonic acid, the precursor of T×A2, which do not share these properties, and PAF-acether itself, were unable to induce PAF-acether formation. Our results suggest that PAF-acether may be the mediator responsible for AIIP and T×A2 independent-aggregation.

Shear Rate Dependent Inhibition of Platelet Adhesion and Aggregation on Collagenous Surfaces by Antibodies to Human Factor VIII/von Willebrand Factor

Abstract: Summary. The effect of heterologous and homologous antibodies to factor VIII/von Willebrand factor (F.VIII/VWF) and purified VWF on the interaction of human platelets with subendothelium and with a surface consisting of collagen fibrils was investigated using annular flow chambers. Wall shear rates of 200–5200 s−1 were produced by varying the flow rate of the citrated blood which was circulated through these chambers by a pump. Surface coverage with platelets and with platelet aggregates was measured morphometrically. The antibodies had no effect on platelet adhesion at low shear rates. However, an increasing inhibition of adhesion was observed with increasing shear rates. The antibodies to F.VIII/VWF and to purified VWF virtually abolished adhesion at shear rates which are present in the microvasculature (1300–5200 s−1). By contrast, antibodies to factor VIII:C which were isolated from two patients with haemophilia A had no effect on adhesion. In addition antibodies to F.VIII/VWF and to purified VWF inhibited adhesion-induced aggregation. We conclude that F.VIII/VWF plays an essential role as cofactor for platelet adhesion to subendothelium and collagen fibrils at high blood shear rates, i.e. when the time available for the establishment of a stable bond between platelets and collagenous surfaces is short.

Terminal Transferase Enzyme Assay and Immunological Membrane Markers in the Diagnosis of Leukaemia: a Multiparameter Analysis of 300 Cases

Abstract: Summary. Multiparameter analyses have been carried out with recently developed enzyme and membrane markers in 300 patients with various leukaemias including ALL, AML, but excluding Ph1 positive leukaemias. TdT enzyme levels were particularly valuable in the differential diagnosis of adult acute lymphoid and myeloid leukaemias. The levels were raised in 108 (94%) of the 115 patients who were considered to be non-T, non-B ALL on membrane marker and morphological analysis; all seven cases giving negative TdT results in this group were young children. Unexpectedly high levels were seen only in three (4.1%) of 73 cases of acute myeloid leukaemia verified by histochemistry and membrane markers. Anti-ALL serum was a most useful reagent in childhood leukaemias but blasts from 19 patients (10% of childhood ALL cases and 29% of adult ALL cases) failed to react with the serum in spite of TdT positivity. Strongly ALL+ blasts were seen only in non-T, non-B ALL and some undifferentiated leukaemias. Weakly ALL+ blasts were seen in seven of 32 cases of thymic ALL (Thy-ALL) but in other respects these blasts expressed Thy-ALL features, such as strong reactivity with anti-T cell (HuTLA) serum, negativity with anti-Ia-like serum and raised TdT. The combination of tests was particularly useful in 32 cases of undifferentiated leukaemia: in 10 of these cases TdT positivity indicated the probable ‘lymphoblast’, nature of blast cells: the remaining 22 cases remained unclassifiable with the markers used. The analysis revealed other interesting variant forms of leukaemias.

Bone Marrow Transplantation in Fanconi Anaemia

Abstract: Summary Five patients with Fanconi anaemia have been treated by bone marrow transplantation from HLA identical donors. Only one patient survived for more than 3 years. She is now perfectly healthy with complete haematological reconstitution with chimaerism and disparition of chromosomal abnormalities. In contrast, four patients died of acute severe GVHD soon after grafting. In addition, all had signs of severe cyclophosphamide toxicity. This evolution could be explained by a special sensitivity of FA cells to alkylating agents and may indicate the need to modify the conditioning regimen in FA patients.

Platelet function in diabetes mellitus.

Abstract: In spite of great advances in treatment of diabetes mellitus, accelerated disease of the microcirculation and the large vessels accounts for the majority of cases of blindness, renal failure, and sudden cardiac death. In insulin-dependent diabetes, life expectancy is 17 years less than the general population (Marks & Krall, 1973). Renal failure and sudden cardiac deaths contribute maximally to the mortality figures, while blindness from obliterative retinal vascular disease occurs in many diabetics of long duration. In non-insulin dependent diabetics, life expectancy is also shortened, either by atherosclerosis of the larger vessels of heart, legs or brain or by associated microvascular disease. Recent studies have improved our understanding of the pathogenesis ofatherosclerosis (Ross 8i Glomset, 1975) and of microvascular disease (McMillan, 1975; Colwell et al, 1979a, b) and an important role for the platelet has been postulated (Colwell et al , 1976; Sage1 et al , 1975). Many studies have now shown that altered platelet function occurs in diabetes mellitus, and the biochemical mechanisms which may underly these platelet functional abnormalities are becoming clear. Therapy based on these studies has been proposed, and is under investigation in large-scale cooperative studies of patients with diabetes mellitus.

Kinetics, Distribution and Sites of Destruction of 111Indium‐labelled Human Platelets

Abstract: Summary. The survival, tissue distribution and fate of 111In-oxine labelled autologous platelets in six normal humans were studied with serial blood sampling, scintillation camera and computer-assisted imaging, whole body profile scanning, and rectilinear scanning. 111In-platelets recovery in the circulation was 72±16% and survival was 216±17 h. Platelet survival curves fitted a linear function best. Initially platelets pooled rapidly in the spleen as a single exponential function, and at 90 min 26% of the injected 111In was located in this organ. Early hepatic uptake was also significant and at 90 min constituted 16% of total body 111In-activity. As labelled platelets disappeared from the circulation there was a threefold increase of radioactivity in the liver to reach 39% of whole body activity at 216 h. Radioactivity also increased significantly in the spleen (33±3% at 216 h). There was significant residual radioactivity in the thoracic and lower abdominal regions at 216 h, suggesting that platelets are also sequestered in the bone marrow. Radioactivity in the lower limbs almost disappeared with time (0±7% at 216 h), indicating that utilization of platelets in the peripheral vasculature is not marked in normal subjects.

Studies on the Concentration and Intracellular Localization of Iron Proteins in Liver Biopsy Specimens from Patients with Iron Overload with Special Reference to their Role in Lysosomal Disruption

Abstract: Summary. Liver biopsies were collected from control subjects and patients with iron overload due to either primary or secondary haemochromatosis. They were analysed for iron proteins by cation exchange chromatography and flameless atomic absorption spectrophotometry. In control tissue the transferrin fraction contains 25%, ferritin 50% and haemprotein and haemosiderin 10–15% each, of the total iron. In iron overloaded tissue the ferritin and haemosiderin iron increases approximately 10- and 100-fold, respectively, compared with control tissue. There was a close positive correlation between enhanced lysosomal fragility as determined by measurements of latent N-acetyl-β-glucosaminidase and haemosiderin content of the tissue; it is suggested that the haemosiderin is responsible for the lysosomal disruption and hence the tissue damage in iron overload. Studies were performed on the intracellular localization of ferritin and of total iron in biopsy extracts from control subjects and from patients with iron overload. In control tissue, ferritin contains most of the iron and is apparently free in the cytosol. In iron overload, ferritin is the major iron protein in the post-nuclear supernatant sedimenting into the gradient as the free protein. There are, however, significant amounts of immunoreactive ferritin deeper in the gradients but this cannot be assigned to any particular subcellular organelle. The extreme fragility of lysosomes in iron overloaded human tissue makes isolation of these organelles for detailed biochemical analysis extremely difficult.

The increased susceptibility of young red cells to invasion by the malarial parasite Plasmodium falciparum.

Abstract: Summary The relationship between red cell age and susceptibility to invasion by the malarial parasite Plasmodium falciparum has been examined by several different methods including short-term cultures of parasitized human blood. The results indicate that reticulocytes and young red cells are more susceptible to invasion by this parasite as compared with metabolically older cell populations. This is contrary to the current belief that red cells of all ages are invaded indiscriminately by P. falciparum. This observation has important theoretical, clinical and practical implications; its mechanism remains as yet unclear.

Origin of Human Bone Marrow Fibroblasts

Abstract: Summary. We investigated the origin of bone marrow fibroblasts in three bone marrow transplant recipients with aplastic anaemia and leukaemia who received grafts from HLA-identical siblings of opposite sex. The patients were conditioned for transplantation with high doses of cytotoxic drugs and 300–1000 rads total body irradiation. After transplantation, bone marrow cells wrere cultured in T flasks for 3 weeks and the adherent cells were then trypsinized and passaged weekly. After several passages the cells had the typical morphology and growth pattern of fibroblasts. Metaphases from these cells were all of recipient sex type. In contrast, haematopoietic cells and lymphocytes obtained at the same time were of donor sex type. Our findings indicate that the human bone marrow fibroblast is not derived from a precursor common to haematopoietic cells or lymphocytes. The bone marrow fibroblast appears to be a mesenchymal cell, unrelated to haematopoietic stem cells, which is capable of in vitro proliferation after as much as 1000 rads of total body irradiation.

Fetal Hb production during acute erythroid expansion. I. Observations in patients with transient erythroblastopenia and post-phlebotomy.

Abstract: In order to study fetal haemoglobin production during acute erythroid expansion we did sequential measurements of Hb F-containing erythrocytes (F-cells) and of relevant haematological parameters in 10 subjects recovering from eyrthroid aplasia, iron deficiency anaemia or following phlebotomy. An increased production of F-cells was consistently observed during the acute marrow expansion, but there were significant differences in the maximum F-cell response among individuals. These differences could not be explained by differences in the degree of anaemia alone, nor could they be correlated with the level of peak reticulocytosis. Two patients who reached the highest F-cell numbers were probably carriers of heterocellular hereditary persistence of Hb F, suggesting that this gene may play a role in determining the magnitude of F-cell production in anaemic patients. It is speculated that the main reason for the consistently observed increase in F cell production during acute marrow expansion is the premature terminal differentiation of earlier erythroid precursos (burst forming units; BFUe). This proposition is in accord with observations in vitro which suggest that Hb F is expressed en eyrthroid clones derived from BFUne's. It is further proposed that differences in the degree of BFUe recruitment may underly the disparities in Hb F response among individuals subjected to similar anaemic stimuli.

Malignancy and haemostasis.

Abstract: I t has been known for a long time that malignant tumours are often associated with the development of deep vein thrombosis and several years ago procoagulant and/or fibrinolytic Activity was described in extracts from human and experimental tumours (Cliffton & Grossi, 1956; O’Meara, 1958). During the last decade new evidence has accumulated indicating possible interactions of cancer cells with the haemostatic system and some concepts of the pathogenesis and clinical relevance of such interactions have developed. These stem from investigations on in vi tro systems and experimental models, as well as from some clinical studies. All the steps leading to cancer cell growth and dissemination, such as local growth, detachmcnt from the primary tumour, local invasion and penetration, transport in blood or lymph, re-entry in extravascular spaces and lodgement in target tissues can, i t seems, be influenced by components of the haemostatic system, such as endothelial cells, platelets, coagulation and/or fibrinolysis.

The association of eosinophilia with lymphoblastic leukaemia or lymphoma: a study of seven patients.

Abstract: Seven patients with hypereosinophilia in association with a lymphoblastic malignancy are described. The eosinophilia preceded or was present at diagnosis in all patients. Eosinophil counts fell during complete remission but rose significantly before or during relapses in five patients. Hypogranular and sometimes Pelger-eosinophils were seen in five cases. Surface and enzyme markers defined the malignancy in six cases as common-ALL (three), T-ALL (two) and T-lymphoblastic lymphoma (one). Although a diagnosis of eosinophilic leukaemia or acute myeloid leukaemia with eosinophil differentiation was considered in three patients, cytochemical and ultrastructural studies failed to show any evidence of myeloid differentiation in the blast cells. The bone marrow karyotype was normal in the four patients studied. All seven patients had one or more relapses and six died 6-62 months from diagnosis. Severe complications of the hypereosinophilic syndrome developed in one patient. As T-lymphocytes have been shown to be involved in the induction of eosinophilia in rodents, it is suggested that the hypereosinophilia in these patients was induced by eosinopoietic stimuli produced by lymphoblasts.

In Vitro Effect of Cyclophosphamide Metabolites on Chromosomes of Fanconi Anaemia Patients

Abstract: Summary The effect of cyclophosphamide metabolites was studied on chromosomes of Fanconi anaemia patients, parents and controls. A high susceptibility of FA patients chromosomes was observed when low concentrations of sera of a cyclophosphamide-treated patient was added to PHA-stimulated lymphocyte cultures. No effect was observed with comparable concentrations on cells from FA patients or controls. The high susceptibility of FA cells is discussed in relation to the high sensitivity of FA patients to cyclophosphamide when used as a conditioning drug for bone marrow graft.

Acidified glycerol lysis test: a screening test for spherocytosis.

Abstract: Acidified glycerol lysis test (AGLT) is a screening procedure which has been developed for spherocytosis. AGLT was found positive in 100% of 48 patients suffering from hereditary spherocytosis, 100% of nine couples of affected parents, and 86% of 14 couples of clinically healthy parents. The test was positive in acquired spherocytosis and negative in normal controls. AGLT appears to have a predictive value higher than the entire battery of conventional tests. It is simple, rapid, inexpensive, gives clear-cut results and requires minute amounts of blood. It can be performed on blood stored for up to 24 h.

Sequential changes in factor VIII and platelets preceding deep vein thrombosis in patients with spinal cord injury.

Abstract: Summary. The relationships between factor VIII associated activities, platelet function, and venous thrombosis were studied in 18 patients with lower limb paralysis following acute spinal cord injury (SCI). Deep vein thrombosis (DVT) was detected in 13 patients (72%). Eight of the 13 thromboses were documented between 6 and 8 d following injury while the other five episodes were noted on days 11 (two), 13,18 and 22. The detection of thrombosis was preceded by marked increases in VIIIR:Ag and VIII:RCoF whereas VIII:C was only marginally increased. The platelet aggregation response to collagen was hyperactive by the sixth day while the platelet aggregate ratio (PAR) did not become abnormal until after DVT was detected. These studies suggest a chronology in the series of events leading to DVT in patients with lower limb paralysis following SCI. Initial elevations in VIIIR:Ag and VIII:R-CoF are followed in sequence by increased platelet responsiveness to collagen, the occurrence of DVT, and the appearance of circulating platelet aggregates. Conceivably, VIIIR:Ag elaborated by endothelial cells alters platelet reactivity and provides an important determinant for venous thrombosis.

Factor VIII and Fibrinolytic Response to Deamino‐8‐d‐Argenine Vasopressin in Normal Subjects and Dissociate Response in Some Patients with Haemophilia and von Willebrand's Disease

Abstract: Summary Deamino-8-d-argenine vasopressin (DDAVP) was given by intravenous infusion to normal subjects, haemophiliacs and patients with von Willebrand's disease (vWd) and the factor VIII and plasminogen activator response was studied. In normal subjects and most patients with mild haemophilia and mild (intermediate) von Willebrand's disease there was an increase in plasminogen activator and all factor VIII related activities. In patients with mild vWd the prolonged bleeding time was shortened by DDAVP despite only a modest rise in factor VIII related Ristocetin cofactor activity (VIIIR:RiCoF). Sub-groups of patients have been characterized in whom atypical responses were observed. In two brothers with clinically severe haemophilia, but with 5–6 u/dl procoagulant factor VIII (VIIIC), there was an increase in VIIIC but no rise of the corresponding antigen, suggesting increased release of an antigenically abnormal poorly functioning molecule. A patient with intermediate vWd was studied in whom neither DDAVP, adrenaline infusion, nor venous occlusion resulted in an increase in either plasminogen activator or factor VIII related antigen (VIIIRAg), although there was a significant increase in VIIIC. In a further patient with severe vWd, DDAVP failed to elicit any plasminogen activator or VIII response. The results obtained from these two patients suggested that in some individuals the presumed endothelial cell abnormality in vWd may be more extensive than a defect in VIIIRAg synthesis. Sub-groups of patients have been identified for whom treatment with factor VIII concentrates would be more appropriate than DDAVP prior to minor surgery.

Cytochemical study of thymocytes and T lymphocytes.

Abstract: Fetal and postnatal thymocytes and circulating T lymphocytes were evaluated for six cytochemical reactions Acid phosphatase activity was present in a high percentage of cells in all threee groups Beta-glucoronidase and alpha-naphthyl acetate acid esterase were negative in the most immature fetal thymocytes, but become increasingly positive with T-cell maturation Only the circulating lymphocytes presented a high percentage of N-acetyl beta glucosaminidase, alpha-naphthyl acetate esterase and alpha-naphthyl butyrate esterase positive cells This study discusses the presence of these enzymes as proportional to different stages in T-cell maturation, and also of certain cytochemical phenotypes characteristic of these stages

Isolation and characterization of normal human megakaryocytes.

Abstract: Human megakaryocytes have been isolated from marrow obtained from ribs removed at thoracotomy. All but one of the patients had normal pre-operative platelet and leucocyte counts. Megakaryocytes averaged 0.37% of all cells in marrow cell suspensions from nine consecutive subjects. A 283-fold purification (to 10.3%) was achieved by a density gradient centrifugation followed by two successive velocity sedimentations at unit gravity. The net yield, 12 800 megakaryocytes per specimen, was sufficient for many kinds of morphological study. Bright-field, phase contrast, and electron microscopy were used to characterize the younger and smaller megakaryocytes. Ploidy analyses were carried out on 100--235 megakaryocytes per specimen; 8N was the predominant ploidy class in isolated megakaryocyte populations from three individuals. The mean megakaryocyte diameter was 24 micrometers in three other specimens and the range was 10--48 micrometers. This data had a normal distribution and overlapped minimally with the size range of all other marrow cells. The presence of a distinct size threshold (at 11.5 micrometers) implied that size alone may be a sufficient objective criterion for identification of human megakaryocytes.

Intensive plasma exchange in the management of severe Rh disease.

Abstract: Fourteen high-risk cases of Rh alloimmunized women were treated by intensive plasma exchange on the cell separator throughout their pregnancies. The mean volume of plasma exchanged per week was 3.21 with a total volume of 10-123 1. The replacement fluid was mainly plasma protein fraction (PPF) supplemented with fresh frozen plasma (FFP). The mean duration of treatment was 131/2 weeks, ranging from 2 to 22 weeks, commencing at 12-30 weeks gestation. The complications encountered and the selection of the optimal time for delivery are discussed. The expected stillbirth rate in this series as determined by their past obstetric histories and anit-D levels was 62%. Intrauterine transfusion was given to only two of the infants and both were later stillborn. It was possible to reduce and maintain a lower level of anti-D in the serum of most of the patients and a successful outcome was achieved in nine of the 12 cases included for analysis (75%). Plasma exchange commenced early in pregnancy is recommended as a non-hazardous form of treatment in the management of severe Rh haemolytic disease. However, if the mean level of anit-D cannot be maintained at less than 35 iu/ml then the outcome is more likely to be fatal. Amniocentesis should be delayed where possible until 28 weeks gestation and intrauterine transfusion reserved for those cases where the anti-D level becomes uncontrollably high and the fetal death is imminent.

Renal and Extrarenal Erythropoietin Production in Anaemic Rats

Abstract: It has been shown in anephric animals and man that small amounts of extrarenal erythropoietin is generated in response to severe anemia or hypoxia. Although this response is inadequate to do more than maintain a hematocrit of 10 to 20%, it seems possible that a stimulated production of extrarenal erythropoietin could replace renal production in anephric patients and provide a new therapeutic approach to patients with impaired renal endocrine function.

Binding of Serum Ferritin to Concanavalin A: Patients with Hornozygous β Thalassaemia and Transfusional Iron Overload

Abstract: Serum ferritin concentrations have been measured in 124 patients with homozygous beta thalassaemia who were between 2 and 21 years old, had received 11--504 units of blood but had not undergone splenectomy. There were highly significant correlations between serum ferritin concentration and both the amount of blood transfused and alanine amino-transferase (ALT) activity. However, multivariate analysis showed that units of blood and ALT activity together only accounted for about 30% of the variation in serum ferritin concentration. Little of the remaining variation could be explained by other variables related to iron metabolism or liver damage. The concentration of concanavalin A binding ferritin increased rapidly with the number of units of blood up to 100 units but thereafter showed no further increase with number of transfusions. The concentration of non-binding ferritin was more closely related to transfusion load. These results suggest that the secretion of glycosylated ferritin from reticuloendothelial cells reaches a maximum with increasing iron accumulation, perhaps reflecting a maximum rate of synthesis. Ferritinaemia in patients with transfusional iron overload therefore seems to be the result of the combined effects of increased ferritin synthesis and the release of intracellular ferritin from damaged cells. A simple relationship between serum ferritin and iron stores cannot be assumed when ferritin concentrations exceed 4000 microgram/l or in patients who have received more than 100 units of transfused blood.

Positive Interaction between Agonists in the Aggregation Response of Human Blood Platelets: Interaction between ADP, Adrenaline and Vasopressin

Abstract: Summary. ADP, adrenaline and vasopressin interact positively as agonists in aggregating human blood platelets in vitro. This interaction is maximal if the addition of two of the agonists is separated by 10–20 s but decreases rapidly at longer intervals especially at low agonist concentrations. The agonist concentrations at which positive interaction gives full aggregation are significantly less than those required for such a response to each agonist alone. The lowest concentrations at which adrenaline and vasopressin interact positively are at least two orders of magnitude greater than the normal blood concentrations of these hormones, and at least an order of magnitude greater than the concentrations achieved in pathological states. Specifically antagonizing the adrenaline and ADP receptors showed that the response was to the second agonist added to the system. An inhibitor of intracellular Ca2+ movement (tetracaine) is equally effective in blocking the responses generated by a single agonist or by interaction of two agonists. Inhibitors which increase cyclic-3′,5′-AMP concentration (adenosine, prostaglandin E1, dipyridamole) are more effective against the response to a single agonist than that to agonist interaction. These data suggest that positive agonist interaction results from effects on the concentrations of second messengers within the platelet rather than from a direct interaction on the membrane receptors or the transmembrane coupling mechanisms.

Biliary Iron Excretion in Rats following Pyridoxal Isonicotinoyl Hydrazone

Abstract: Biliary excretion of iron after administration of pyridoxal isonicotinoyl hydrazone (PIH), a recently identified effective iron-chelating agent, was investigated in rats. PIH administered both intraperitoneally and orally was shown to increase significantly 59Fe excretion into bile of rats which had previously been injected with 59Fe-transferrin to label hepatic parenchymal cells. 59Fe-PIH appears in bile as early as 15 min after chelator administration and the peak of 59Fe-radioactivity in bile is seen 1--5 h following intraperitoneal PIH injection. PIH, administered intraperitoneally, 125--250 mg/kg, increased 24 h biliary radioiron excretion about 35 times and in addition increased urinary and faecal iron excretion. When PIH was given immediately before 59Fe-transferrin, 24 h cumulative biliary 59Fe excretion was even higher. PIH was also demonstrated to increase biliary excretion of radioiron released from 59Fe-haemoglobin catabolysed in reticuloendothelial cells. The effect of PIH was confirmed by estimation of biliary iron concentration using the method of atomic absorption spectrophotometry. Repeated PIH administration to rats decreased 59Fe radioactivity in liver and kidney and increased urinary and faecal iron excretion.

Immune complexes in thrombocytopenic patients: cause or effect?

Abstract: Summary. Immune complexes (ICs) in the serum of 43 patients with chronic idiopathic thrombocytopenic purpura (ITP) were measured by the C1q deviation assay during the active and inactive phases of the disease. An inverse relationship between platelet count and levels of ICs was demonstrated in all but one patient. To test whether this phenomenon was specific for chronic ITP, ICs were assayed in sera from two groups of control patients with thrombocytopenia. Group 1 had thrombocytopenia due to recognized immune mechanisms while group 2 had thrombocytopenia secondary to non-immune mechanisms. In both these groups the degree of thrombocytopenia proved to be inversely proportional to IC levels, which was similar to the pattern observed in chronic ITP. The specificity of the assay for detection of ICs was confirmed by demonstrating a positivity rate of 65% in sera of patients with systemic lupus erythematosus, a known IC disease. On analysis the ICs were shown to have molecular weights in excess of 500000 daltons and contain variable immunoglobulin classes. The findings implicate ICs in immune destruction of platelets both in chronic ITP (as has been suggested previously) and also in thrombocytopenia secondary to known immune mechanisms. In addition the association of ICs with non-immune thrombocytopenias is consistent with the hypothesis that platelets play an important role in clearance of ICs from the circulation, thereby protecting the vascular endothelium from damage.

Polycythaemia vera: in vitro studies of circulating erythroid progenitors.

Abstract: Summary. Peripheral blood mononuclear cells from healthy subjects, patients with secondary polycythaemia and patients with polycythaemia vera (PV) were cultured using the plasma clot technique, and large erythroid colonies or bursts were scored after 14 d. Erythroid bursts appeared without addition of erythropoietin in all 20 patients with PV but never in normal subjects (n=10) nor in patients with secondary polycythaemia (n=6). The erythropoietin dose-response curves in six PV patients were characterized by an initial plateau followed by a nearly normal response. This suggests the coexistence of two populations of erythroid progenitors in PV, one abnormally sensitive, the other normally responsive to erythropoietin. In nine patients the ratio of spontaneous bursts to bursts induced by high doses of erythropoietin from the blood on day 14 was identical with the ratio of spontaneous colonies to colonies induced by high doses of erythropoietin from the bone marrow of the same patients on day 7. This argues against the hypothesis that spontaneous erythroid bursts are derived from erythroid progenitors (BFUE) abnormally sensitive to erythropoietin. Moreover blood mononuclear cells from healthy subjects were found to give rise to erythroid bursts when erythropoietin addition was delayed until 6 d in culture. This is consistent with the concept that erythropoietin is not normally required in vivo for the differentiation of early (BFUE) to late (CFUE) erythroid progenitors. These findings suggest that in PV all BFUE are normally responsive to the physiological factor(s) different from erythropoietin and responsible for their differentiation into CFUE, and that the abnormal response to erythropoietin, distinctive of a population of erythroid progenitors, is expressed only at the most mature (CFUE) level.