bluuebunny/crossref_metadata_2025 huggingface dataset:
Platelet Adherence to Collagen: Role of Plasma, ADP, and Divalent Cations
Authors: Cowan Dale H. , Robertson Abel L. , Shook Patricia , Giroski Paul
Published: March 2008

Summary. The effect of varying methods of platelet preparation and the role of ADP and divalent cations in supporting platelet adherence to collagen and the release of 14C‐serotonin were assessed by affinity chromatography on collagen/Sepharose to define physiological conditions for this interaction. Platelets were separated by centrifugation or gel filtration from blood anticoagulated with EDTA or citrate and suspended in native plasma or buffer. After labelling with 51Cr or 14C‐serotonin, they were passed through columns of collagen covalently linked to cyanogen bromide‐activated Sepharose. Adherence to collagen was less in plasma as compared to buffer, was increased by centrifuging the platelets before testing, and was unaltered by addition of ADP. Removal of ADP with CP/CPK decreased the adherence of gel‐filtered citrated and EDTA platelets and washed EDTA platelets (P < 0.001) but not EDTA platelets in plasma. Adherence and release of citrated platelets in plasma or buffer containing CP/CPK were greater than that of EDTA platelets (P < 0.01); no difference existed with gel‐filtered platelets. The addition of 1 mM Mg++ to citrate or EDTA‐anticoagulated washed platelets increased adherence and release (P < 0.01). The results indicate that the choice of experimental conditions affects the assessment of factors which influence or promote platelet interaction with collagen. Platelet—collagen adherence is enhanced by laboratory manipulations and partially inhibited by normal plasma. Maximal adherence and release occur when divalent cations, particularly Mg++, and ADP are available. Their absence reduces but does not inhibit these reactions.
Methaemoglobinaemia Resulting from Heterozygosity for Two NADH‐Methaemoglobin Reductase Variants: Characterization as NADH‐Ferricyanide Reductase
Authors: Board P. G. , Pidcock M. E.
Published: March 2008

Summary. A case of congenital methaemoglobinaemia was found to result from heterozygosity for two different forms of NADH‐methaemoglobin reductase (NADH‐MR) deficiency. One of these variant forms was found to be electrophoretically fast, and to have sufficient residual activity to allow detailed characterization. This variant was characterized by its NADH‐ferricyanide reductase activity, and was found to have a normal pH optimum and normal affinity for K3Fe(CN)6, NADH and deamino NADH. The diminished activity associated with this variant appeared to result from its decreased stability.

It is suggested that the NADH‐ferricyanide reductase activity of NADH‐MR provides a convenient and standardized procedure for the characterization of NADH‐MR.
The Interaction between (Ca2++ Mg2+)‐ATPase and the Soluble Activator (Calmodulin) in Erythrocytes Containing Haemoglobin S
Authors: Dixon Earl , Winslow Robert M.
Published: March 2008

Summary. In normal erythrocytes, a membrane‐bound (Ca2++ Mg2+)‐ATPase is stimulated by a soluble activator, calmodulin. Since cells containing Hb S accumulate excessive Ca2+, the defect could lie in either the (Ca2++ Mg2+)‐ATPase or calmodulin. To decide between these two possibilities, we prepared (Ca2++ Mg2+)‐ATPase from erythrocytes of normal (AA), sickle cell trait (AS) and sickle cell disease (SS) individuals. Calmodulin was prepared from haemolysates from AA and SS erythrocytes. The enzyme prepared from SS ghosts had lower specific activity than that from AA membranes. Furthermore, calmodulin from either source did not stimulate the ATPase of SS erythrocytes. Enzyme from AS cells had specific activity similar to that of enzyme prepared from SS membranes. The enzymatic activity of a mixed cell population obtained from an SS patient 8 d following exchange‐transfusion was proportional to the per cent Hb A. These results indicate that calmodulin is unable to interact with the enzyme site on the SS membrane. This inability is believed to be due to a specific property of the membrane and not an abnormality of calmodulin itself.
Myoglobin Depletion in Childhood Iron Deficiency
Authors: Hargreaves R. M. , Street M. , Hoy T. , Jacobs A. , McLaren C.
Published: March 2008

Summary. Serum concentrations of myoglobin are significantly lower in iron‐deficient than in non‐iron‐deficient children. In the iron‐deficient subjects there is a correlation between haemoglobin concentration and serum myoglobin suggesting that the synthesis of both haem compounds is affected in parallel. No abnormality of serum myoglobin concentration was found in adults with iron deficiency anaemia.
Severe Aplastic Anaemia: Correlation of in Vitro Tests with Clinical Response to Immunosuppression in 20 Patients
Authors: Bacigalupo A. , Podesta M. , Van Lint M. T. , Vimercati R. , Cerri R. , Rossi E. , Risso M. , Carella A. , Santini G. , Damasio E. , Giordano D. , Marmont A. M.
Published: March 2008

Summary. Colony formation in agar (CFU‐c) was studied in 20 patients with severe aplastic anaemia by three different assays: (1) cultures of light density untreated marrow cells; (2) cultures of marrow cells manipulated in order to enhance colony formation (pretreatment with antilymphocytic globulin, ALG, or 6‐methylprednisolone, 6‐MPr, T cell depletion, adherent cell (AC) depletion, depletion of both AC and T cells), and (3) co‐culture of putative suppressor T cells with autologous T‐depleted marrow cells.

By the first assay, all patients showed poor colony formation (1 1.5 colonies/105 cells; normal controls 46.18 colonies/105 cells). By the second assay, ALG and 6‐MPr had no significant effect on colony formation. Removal of adherent cells proved equally without effect on colony growth. On the contrary, removal of T cells enhanced significantly (P < 0.001) colony formation in 10 out of 20 patients. By the third assay, colony formation of marrow cells (deprived of T lymphocytes) was inhibited by the addition of autologous T cells in six patients studied.

All patients were given high dose bolus 6‐MPr as first treatment on admission: only patients who had detectable suppressor T cells in their marrow achieved a complete autologous haematologic reconstitution after 6‐MPr or after 6‐MPr and ALG.

The results of this study indicate that detection of CFU‐c/suppressor T cells correlates with responses to immunosuppressive regimens, and may thus help to identify patients with immune mediated aplastic anaemia.
In Vitro Assay of Erythropoietin in Fetal Mouse: Liver Cultures
Authors: Brandan N. C. , Cotes P. M. , Espada J.
Published: March 2008

Summary. In a 13 d fetal mouse liver tissue culture system for assay of erythropoietin, stimulation of incorporation of H‐thymidine and of 125I‐iodo‐2′‐deoxy uridine into DNA and of 59Fe iron into haem were compared as metameters of erythropoietin effect. With both DNA and haem tracers, log‐dose response curves given by two preparations of human urinary erythopoietin purified to different extents (respectively 2 and 1300IU, by bioassay in vivo, per mg of protein) were essentially identical at low dose levels although at high dose levels the less pure preparation decreased stimulation of both DNA and haem synthesis. Pre‐incubation of erythropoietin with an antiserum to human erythropoietin abolished the effects of the hormone on both DNA and haem synthesis.
Acute Myelodysplasia with Myelofibrosis: a Report of Eight Cases
Authors: Sultan C. , Sigaux F. , Imbert M. , Reyes F.
Published: March 2008

Summary. Eight patients with acute myelodysplasia and myelofibrosis are described. Four cases were secondary to long‐term therapy with cytotoxic agents and four were idiopathic. All cases presented with an abrupt onset of the illness, absence of organomegaly and severe pancytopenia. Bone marrow aspirate yielded adequate material in four cases and showed myelodysplasic features. Study of histological sections indicated that the bone marrow was cellular in every case, including numerous dystrophic megakaryocytes, erythroblasts, immature cells of the granulocytic series and blast cells which were difficult to identify. The reticulin network was always increased. In each case the disease was rapidly fatal. No improvement was noted with chemotherapy. In three cases an overt leukaemia developed with marked pleomorphism of blast cells. The nosology of this syndrome is discussed.
Hypoxanthine‐Guanine Phosphoribosyltransferase Activity in Normal and Leukaemic Lymphocytes
Authors: Rambotti Pietro , Davis Stephen
Published: March 2008

Summary. The levels of hypoxanthine‐guanine phosphoribosyltransferase (HGPRT) were determined in lymphocytes from normal people and patients with chronic lymphocytic leukaemia (CLL). The HGPRT level in the total lymphocyte population from patients with CLL was lower than that from normal subjects. The HGPRT activity was higher in normal non‐T cells than normal T cells. The enzyme activity in CLL B cells was lower than in CLL T cells. The HGPRT level was higher in CLL T cells than in normal T cells; these data suggest CLL T cells differ biochemically from their normal counterparts.
Functional and Morphological Studies of Platelet Reactivity with Vessel Wall Subendothelium in Chronic Myeloproliferative Disease
Authors: Sacher Ronald A. , Jacobson Robert J. , McGill Manley
Published: March 2008

Summary. Mechanisms of thrombus formation in myeloproliferative disease were studied using a technique which visualized platelet‐vessel wall interactions under physiological conditions of blood flow. Whole blood from four patients with chronic myelogenous leukaemia and three with thrombocythaemia were pumped through perfusion chambers containing de‐endothelialized artery segments. Platelet reactivity with vessel walls (thrombus formation) was measured in sections of vessels by light microscopy and quantitative morphometric analysis. Five patients produced platelet reactivity values of 130‐258% of controls while two gave decreased values. The two highest platelet reactivity values occurred in samples with elevated platelet counts and normal haematocrits. In contrast, when anti‐platelet drugs were administered to three patients with high platelet counts, reactivity values decreased to 11‐34% of controls. Clinical correlations revealed that patients with highest platelet reactivity values (165‐258%) were those subjects who also exhibited thrombotic or haemorrhagic complications. Thus absolute platelet count and haematocrit may be major determinants in predicting these complications. Qualitative evaluations of thrombus formations by light and electron microscopy provide further evidence that platelets in myeloproliferative disease also possess qualitative abnormalities.
Bone Marrow Transplantation in Severe Aplastic Anaemia: a Survey of the European Group for Bone Marrow Transplantation (E.G.B.M.T.) *
Authors: Gluckman E. , Barrett A. J. , Arcese W. , Devergie A. , Degoulet P.
Published: March 2008

Summary. A survey of the results of bone marrow transplantation for severe aplastic anaemia in 13 European teams is reported. 159 questionnaires were analysed by univariate and multivariate analysis. The overall 1 year survival was 41.2%. The chance of survival was decreased by the presence of infection before grafting and by the use of female donors. The conditioning regimen did not influence survival. Graft rejection was influenced by the conditioning regimen and the sex of the donor. Graft‐versus‐host disease was influenced by the conditioning regimen, and the age and the sex of the recipient.

This study defines prognostic indices which may be helpful in determining the likelihood of success after marrow transplantation for severe aplastic anaemia.
Reactivity of Monoclonal Antibodies with Human Myeloid Precursor Cells
Authors: Crawford D. H. , Francis G. E. , Wing M. A. , Edwards A. J. , Janossy G. , Hoffbrand A. V. , Prentice H. G. , Secher D. , McConnell I. , Kung P. C. , Goldstein G.
Published: March 2008

Summary. Five monoclonal antibodies have been tested for their ability to bind to myeloid precursor cells in normal human bone marrow. Indirect immunofluores‐cence and the fluorescence activated cell sorter was used to separate cells according to their reactivity for trial culture in vitro in order to grow granulocyte‐macrophage colony forming cells (CFUc). Two antibodies (OKT3 and OKT11) which react strongly with bone marrow T lymphocytes were found to be unreactive with CFUc. YD 1/23 reacts very strongly with both T and B lymphocytes but is only weakly reactive with CFUc. In contrast, OKT10 and YE2/36 did react with CFUc. The consequences of these findings and the potential clinical use of these antibodies in bone marrow transplantation are discussed.
Dependence of Purine Loss from Human Erythrocytes on External pH
Authors: Schraufstätter Ingrid , Born G. V. R.
Published: March 2008

Summary. Human red cells were incubated at 37°C in physiological saline at pH 7.4 with 8‐3H‐adenine to label the adenine nucleotides. After washing, the cells were incubated in similar salines which were buffered to different values of pH. The loss of radioactivity from cells into medium was very slow below pH 7.4 and accelerated greatly with increasing pH up to about 7.8. At that pH the loss increased for at least 3 h when it represented more than 30% of the initial radioactivity of the cells. The loss could not be accounted for by haemolysis which remained minimal. The extracellular radioactivity was present mostly in hypoxanthine and some inosine. The loss of purine did not depend on oxygenation of the cells. These results, together with the high‐affinity uptake system for adenosine, could provide a partial explanation of the turnover of adenine nucleotide purines of red cells which is many times faster than that of the cells themselves.
Methotrexate Accumulation and Folate Depletion in Cells as a Possible Mechanism of Chronic Toxicity to the Drug
Authors: Kamen B. A. , Nylen P. A. , Camitta B. M. , Bertino J. R.
Published: March 2008

Summary. Methotrexate has been associated with chronic toxicities such as cirrhosis and neurological impairments ranging from mild learning disorders to a fatal leucoencephalopathy. The mechanism(s) for this toxicity is not completely understood. Certain tissues can convert methotrexate to polyglutamates. This results in a more persistent intracellular form of the drug. In this study the intracellular levels of folate and methotrexate were measured in the erythrocytes and liver of patients treated chronically with methotrexate. These tissues showed an accumulation of methotrexate as polyglutamates and a concomitant loss of folate. Folate concentrations were below normal in nine of 12 red cell and three of five liver samples. It is proposed that persistent methotrexate concentrations and/or the associated folate deficiency may be related to the toxicity of methotrexate, especially in time of cellular stress.
A Negative Direct Antiglobulin Test with Strong IgG Red Cell Autoantibodies Present in the Serum of a Patient with Autoimmune Haemolytic Anaemia
Authors: Van't Veer M. B. , Van Wieringen P. M. V. , Leeuwen I. van , Overbeeke M. A. M. , Von Dem Borne A. E. G. KR. , Engelfriet C. P.
Published: March 2008

Summary. The case is presented of a boy with an autoimmune haemolytic anaemia of 10 years duration. He had a positive direct antiglobulin test with IgG and complement detected on the red cells and with IgG autoantibodies in the serum. During a recent episode of severe haemolysis, the Hb level fell to 3.8 g/dl and the direct antiglobulin test became negative although his autoantibodies still reacted with all the red cells in a panel. The serum reacted more strongly with C‐ and e‐positive cells. The Rh phenotype of the patient was CcDee as it had always been. Possible explanations of the unexpected findings are discussed.
Unusual T‐Cell Phenotype in Advanced B‐Chronic Lymphocytic Leukaemia
Authors: Matutes Estela , Wechsler A. , Gomez R. , Cherchi Maria , Catovsky D.
Published: March 2008

Summary. T‐lymphocytes isolated by E‐rosetting from 22 patients with B‐cell chronic lymphocytic leukaemia (B‐CLL) showed membrane phenotype features distinct from those of normal T‐lymphocytes. These changes were particularly marked in advanced disease (Rai stages II, III, IV and WBC over 100 × 109/1). The most significant finding was the demonstration, in 10 cases, of a major population of E‐rosette positive cells (40—80%) unreactive with the OKT monoclonal antibodies against mature or immature T‐cells; 15—30% of the cells were unreactive in seven other cases. A significant reduction in OKT4 positive (helper) lymphocytes was seen in 18 cases. The proportion of OKT8 positive cells was increased in two and normal or low in the rest. Only four patients with early disease (stages 0 and 1) had a normal T‐cell phenotype. These findings could explain abnormalities previously described in the T‐lymphocytes of B‐CLL and provide new insights into the pathogenesis of the disease.
Activation of natural cytotoxicity of human peripheral blood mononuclear cells by interferon: a kinetic study and comparison of different interferon types
Authors: Claeys Hendrik , Van Damme Jozef , De Ley Marc , Vermylen Carl , Billiau Alfons
Published: March 2008

Summary. Overnight incubation of human peripheral blood mononuclear (PBMN) cells with leucocyte interferon (leucocyte IFN) resulted in a 2–5‐fold increase in natural cytotoxicity (NC) against the erythroleukaemic cell line K562. Fibroblast IFN, purified to homogeneity by zinc‐chelate chromatography, stimulated NC to the same extent, while partially purified immune IFN was about twice as active. Upon gel filtration of immune IFN, NC stimulating and antiviral activity co‐eluted. Treatment of PBMN cells with ammonium chloride buffer (AmCl) abrogated NC nearly completely. Incubation of AmCl‐treated cells with leucocyte IFN resulted in a partial regeneration of NC. Kinetic studies revealed that this regeneration required only a short exposure to IFN followed by a longer incubation period. The data are interpreted as indicating that in the process of NC activation IFN mainly acts as a trigger for precursor cells to mature into cells with NC.
Precipitating anti‐VIII:C antibodies in two patients with haemophilia A
Authors: Lavergne Jean‐Maurice , Meyer Dominique , Girma Jean‐Pierre , Larrieu Marie‐José
Published: March 2008

Summary. IgG from the plasmas of two haemophilia A patients with anti‐VIII:CAg antibodies (1000 and 200 u/ml) was isolated and labelled with 125I. The specific labelled anti‐VIII:CAg IgG was further purified by binding to and elution from immobilized factor VIII/von Willebrand factor (F.VIII/vWF). When studied by immunodiffusion and autoradiography, both antibodies gave a precipitin line with normal plasma, serum, cryoprecipitate, purified F.VIII/vWF and the plasmas of two patients with haemophilia A+. No precipitin line was observed with the plasmas of 11 patients with haemophilia A− or four patients with severe von Willebrand's disease. Levels of VIII:CAg obtained by radioelectroimmunoassay were in agreement with those obtained by immunoradiometric assay. This study demonstrates that, contrary to previous evidence, human anti‐VIII:CAg antibodies are precipitating as well as neutralizing when studied by highly sensitive techniques.
Acute promyelocytic leukaemia: a study of 39 cases with identification of a hyperbasophilic microgranular variant
Authors: McKenna Robert W. , Parkin Janet , Bloomfield Clara D. , Dorothy Sundberg R. , Brunning Richard D.
Published: March 2008

Thirty‐nine cases of acute promyelocytic leukaemia (APL) were divided into two morphological subgroups, typical hypergranular APL (31 cases) and microgranular APL (eight cases, 21%). The leukaemic cells in the microgranular APL cases were characterized by striking nuclear folding or lobulation; granulation was present in most of these cells but was less abundant and finer than in typical APL. In three microgranular APL cases a distinctive small leukaemic promyelocyte with unusual nuclear lobulation and deeply basophilic cytoplasm containing few or no visable granules was the predominant leukaemic cell. This small hyperbasophilic promyelocyte was also present as a minor population of cells in the other five microgranular APL cases and in 28 of the 31 typical APL cases. Ultrastructurally the most abundant promyelocytes in microgranular APL had smaller and usually fewer granules than in typical APL; other characteristic ultrastructural features of APL were found with equal frequency. The median blood leucocyte count was significantly higher in microgranular APL, 83·0 × 109/1, than in typical APL, 1 · 8 × 109/1 (P < 0 · 01). The median duration of complete remission (CR) for microgranular APL, 6 · 5 months, was shorter than the 21 + month median CR for typical APL. The morphological characteristics of microgranular APL may mimic those of myelomonocytic leukaemia; however, the presence of cells with multiple Auer rods, large inclusions of Auer‐like material and the small hyperbasophilic promyelocytes are important distinguishing features. In equivocal cases cytochemistry, electron microscopy and cytogenetic studies may verify the diagnosis.
Production of immunoreactive calcitonin by myeloid leukaemia cells
Authors: Foa R. , Oscier D. G. , Hillyard C. J. , Incarbone E. , McIntyre I. , Goldman J. M.
Published: March 2008

Raised plasma levels of immunoreactive human calcitonin (i‐HCT) have been found in patients with chronic granulocytic leukaemia (CGL) in chronic phase and myeloblastic transformation and in patients with acute myeloid leukaemia at presentation and in relapse. In CGL levels were significantly higher in myeloblastic transformation than in the chronic phase. Leukaemia cells were cultured in a short‐term liquid culture system in which little cell proliferation occurs and in a two layer blast‐cell colony system which permits blast‐cell proliferation. i‐HCT was identified in supernatant media from cells cultured in both systems but levels were substantially higher in media collected from cells cultured in the latter system. These results suggest that i‐HCT is synthesized by proliferating blast cells.
Lymphocyte sub‐populations following splenic irradiation in patients with chronic lymphocytic leukaemia
Authors: McCann S. R. , Whelan C. A. , Breslin B. , Temperley I. J.
Published: March 2008

Four patients with chronic lymphocytic leukaemia were treated with splenic irradiation, 100 rads per week for 10 weeks. No patient had received prior therapy of any kind. All patients showed an excellent clinical response in terms of decrease in lymphocyte count in blood and bone marrow and decrease in lymph node and spleen size. B and T cell populations returned to normal in all patients as did the ratio of TG to TM cells. Two patients showed an increase in IgM levels, but the overall pattern of immunoglobulins showed no consistent change following treatment. No toxicity was incurred during treatment. We conclude that splenic irradiation is a safe and effective treatment for previously untreated patients with CLL and is associated with a return to normal of lymphocyte sub‐populations. The role of the abnormal T cell population in the pathogenesis of the hypoimmuno‐globulinaemia remains unclear.
Acetylcholinesterase activity of lymphocytes: an enzyme characteristic of T‐cells
Authors: Szelényi Judith G. , Bartha Emilia , Hollán Susan R.
Published: March 2008

Acetylcholinesterase (AchE) activity of normal human peripheral blood lymphocytes was determined by an adaptation of the colorimetric and radiometric techniques described for other cells. The enzyme activity seemed to be correlated to the number of T‐lymphocytes in the blood. To decide whether only the T‐cells possess AchE activity the lymphocytes were separated on Leucopac filter and on Percoll density gradient. B‐lymphocytes had no detectable enzyme activity, while the T‐lymphocyte fraction represented the total activity measured in the unseparated sample. The majority of AchE activity could be demonstrated in T‐lymphocytes of lower density (TLD). The role of AchE in the plasma membrane of various blood cells is not known. Nevertheless, the enzyme is a good marker of the integrity and functional state of the membrane. The difference observed in AchE activity of the lymphocyte populations seems to be suitable for using it to characterize T‐lymphocytes.
Further studies on the mechanism of marrow granulocytic hyperplasia in mice chronically injected with endotoxin
Authors: Levitt Lee J. , Quesenberry Peter J.
Published: March 2008

Marrow granulocytic hyperplasia occurs regularly in mice injected with endotoxin for 7‐30 d, despite minimal elevations of serum colony‐stimulating activity (CSA). Alterations in marrow granulocyte‐monocyte progenitor (CFU‐C) number or changes in marrow cell cycle status do not explain this hyperplasia. We have studied other mechanisms which may explain this increased granulopoiesis. CF1, BDF1 or C57bl/6J mice were injected with 10 μg of S. typhosa, endotoxin i.p. daily for 7‐20 d. Control and endotoxin injected (tolerant) sera, each with identical levels of CSA, were assayed against control marrow cells stimulated with supramaximal amounts of CSA to assess the role of serum potentiators in augmenting granulopoiesis. In six separate experiments, tolerant sera, over a 30‐fold concentration range, produced a 1.7–4.0‐fold potentiation of colony growth compared to control sera (P<0.001). No increased tolerant sera potentiation was seen over a similar concentration range when assayed against tolerant marrow. Tolerant and control splenic conditioned media, both dialysed and non‐dialysed, failed to potentiate control marrow colony growth. Tolerant marrow stem cells did not show changes in CSA sensitivity, colony size distribution or differentiation, and tolerant bone or bone marrow cells did not produce increased amounts of CSA. We conclude that serum factors separate from CSA may in part explain the increased granulopoiesis seen in endotoxin injected mice. The failure of tolerant sera to potentiate tolerant marrow growth in vitro, may reflect prior in vivo, exposure of marrow to these potentiating factor(s).
Canine cyclic haematopoiesis: the effect of endotoxin on erythropoiesis
Authors: Hammond William P. , Adamson John W. , Dale David C. , Rodger Elin , Wise Cathy
Published: March 2008

We have examined the effects of chronic endotoxin treatment on erythropoiesis in six grey collies with cyclic haematopoiesis. Blood reticulocytes, bone marrow erythroid colony (EC) forming cells, serum iron and erythropoietin (ESF) values showed regular periodic fluctuations in untreated grey collies. Daily endotoxin injections eliminated the cyclic fluctuations of reticulocytes and EC. The mean serum iron values were increased and recurrent hypoferraemia eliminated, while the mean serum ESF values were reduced. The cyclic fluctuations of serum ESF values were no longer apparent in the endotoxin treated grey collies. Tritiated thymidine suicide of the marrow EC forming cells failed to show cyclic changes either in untreated or endotoxin treated dogs. The ESF sensitivity of EC in the grey collie was unchanged during endotoxin treatment and was not different from normal dogs. Endotoxin appears to alter periodic erythropoiesis by stabilizing the flux of cells into the committed erythroid precursor cell pool from a more primitive stem cell compartment.
Erythroid progenitors in paroxysmal nocturnal haemoglobinuria
Authors: Urabe Akio , Fujioka Seitoku
Published: March 2008

In vitro colony formation of bone‐marrow erythroid progenitor cells in patients with paroxysmal nocturnal haemoglobinuria (PNH) was examined. The numbers of early and late erythroid progenitors (BFU‐E and CFU‐E) showed wide variations; two cases out of eight cases of PNH showed decreased erythroid colony formation, but other cases showed normal or rather increased colony formation of BFU‐E and CFU‐E. The number of erythroid progenitors in patients with PNH may be related to the marrow cellularity.
DNA chain elongation rates in marrow cells from vitamin B12‐deficient patients and methotrexate‐treated mice
Authors: Bond A. N. , Harris G. , Wickramasinghe S. N.
Published: March 2008

SUMMARY. The DNA synthesized by marrow cells from patients with vitamin B12 deficiency and mice previously given methotrexate (MTX), has been investigated. Suspensions of bone marrow cells were pulse‐labelled with [methyl‐3H]thymidine or deoxy [5‐3H]cytidine for 30 s and the radioactivity in the DNA was chased thereafter in the presence of 10 μM non‐radioactive nucleoside for periods up to 60 min. The rates of elongation of new daughter strands were then assessed by hydroxyapatite chromatography of alkali‐denatured DNA samples. No significant differences were found between the average rates of elongation of daughter strands from control marrow cells on the one hand and the vitamin B12‐deficient or the methotrexate‐affected cells on the other. This is to be contrasted with the results of previous studies which have shown a retardation in the rates of movement of replication forks in stimulated, cultured lymphocytes obtained from vitamin B12‐ or folate‐deficient patients.
Serological and biochemical analysis of the PlA1 alloantigen of human platelets
Authors: Lane Judy , Brown Melissa , Bernstein Irwin , Wilcox Paula K. , Slichter Sherrill J. , Nowinski Robert C.
Published: March 2008

A solid phase platelet antibody assay has been developed which rapidly and sensitively detects PlA1 antibodies. The three‐step assay is performed by: (1) adhering platelets to the wells of a microtitre plate, (2) incubating the platelets with test serum, and (3) adding radiolabelled Staphylococcal protein A which binds to the Fc domain of IgG antibodies. Immune reactions are detected by overnight autoradiography.

Characterization of the PlA1 antigen was performed by using PlA1 antisera in immune precipitation assays. A 90 000 dalton molecular weight species was precipitated from PlA1 positive human and dog platelets.
Congenital dyserythropoietic anaemia type II (CDA‐II): chromosomal banding studies and adherent cell effects on erythroid colony (CFU‐E) and burst (BFU‐E) formation
Authors: Roodman G. D. , Clare C. N. , Mills G.
Published: March 2008

Bone marrow CFU‐E and BFU‐E from a patient with CDA‐II formed erythroid colonies and bursts which contained multinucleated erythroblasts in vitro. Adherent cell depletion of the patient's marrow increased CFU‐E derived colonies six‐fold (98 ± 17 v. 640 ± 15 per 105 marrow cells plated) and co‐culture of CDA‐II marrow adherent cells with CDA‐II adherent cell depleted marrow significantly suppressed erythroid colony formation. Similar adherent cell suppression of the patient's BFU‐E also occurred. Adherent cell depletion of normal marrow did not increase CFU‐E derived colony formation (488 ± 63 v. 495 ± 108) and decreased BFU‐E derived burst formation. Addition of normal adherent cells to normal marrow increased erythroid colony and burst formation. Karyotype and chromosomal banding studies of the patient's multinucleated cells did not show chromosomal inversions, deletions or translocations.
Erythropoiesis in the anaemia of rheumatoid arthritis
Authors: Cavill I. , Bentley D. P.
Published: March 2008

Erythropoietic activity and mean red‐cell lifespan were measured using 59Fe‐transferrin in 32 anaemic patients with active rheumatoid arthritis (RA) and in 20 haematologically normal subjects. Marrow iron turnover (MIT) in the patients was normal and in only 11 was red‐cell lifespan less than 70 d. Ineffective iron turnover (IIT) was significantly increased in those patients in whom there was evidence of iron deficiency (serum ferritin less than 12 μg/l) but in the remaining patients IIT was significantly less than in the normal subjects. In a further 19 patients with simple iron deficiency anaemia IIT was significantly increased. The marrow response to chronic inflammatory disease is clearly distinct from that seen in simple iron deficiency and this suggests different pathogenic mechanisms for these two anaemias.
A technique for rapid isolation of bone marrow mononuclear cells using Ficoll‐Metrizoate and the IBM 2991 blood cell processor
Authors: Gilmore M. J. M. L. , Prentice H. G. , Blacklock H. A. , Ma D. D. F. , Janossy G. , Hoffbrand A. V.
Published: March 2008

Marrow from seven normal donors and patients has been layered onto Ficoll‐Metrizoate (FM) under pressure in the IBM 2991 blood cell processor to isolate the mononuclear cell (MNC) population prior to allogeneic transplantation or cryopreservation. This separation method, which takes less than 90 min, is a further development since our previous report detailing the use of the IBM 2991 to produce a concentrated marrow‘buffy coat’for infusion (Gilmore & Prentice, 1981). By adding FM to the system, marrow stem cells are further concentrated in a small volume with removal of unwanted granulocytes and red blood cells. This facilitates the in vitro treatment of marrow with monoclonal antibodies (Granger et al, 1982) or drugs, for either the selective elimination of malignant cells prior to autologous bone marrow transplantation (BMT), or T lymphocytes in an attempt to prevent graft versus host disease (GvHD) in allogeneic BMT.

Five of the seven marrows processed by this procedure have thus far been infused into lethally irradiated recipients with engraftment (allogeneic); the other two marrows have been cryopreserved.
Factor IX antigen by a rapid staphylococcal protein A‐membrane binding radioimmunoassay: results in haemophilia B patients and carriers and in fetal samples
Authors: Suzuki Lucy A. , Thompson Arthur R.
Published: March 2008

Staphylococcal protein A‐membranes have been used with isolated, radiolabelled factor IX and specific rabbit antisera for modification of a radioimmunoassay. The current method is a rapid 4 h procedure and dilution curves of plasma parallel those of isolated, unlabelled protein. Non‐specific binding is 5%; the assay readily detects concentrations as low as 0.6 u/dl. Carrier detection of haemophilia B was improved and/or confirmed by the demonstration of factor IX antigen in excess of clotting activity in nine of 15 women tested from pedigrees in which the affected members had excess circulating antigen. Of 15 new haemophilia B pedigrees examined, 13 had antigen levels which were in two‐fold or greater excess over their clotting activities; all but three were considerably below normal, however. To diagnose haemophilia B in newborns at risk, levels in three cord blood samples were tested; two were positive and the third was normal. Six fetal blood samples were assayed and contained from 4 to 20 u/dl factor IX antigen; levels correlated with fetal age.
Plasma fibronectin in acute leukaemia
Authors: Boughton B. J. , Simpson A.
Published: March 2008

Summary.

In acute leukaemia, low plasma fibronectin was seen at diagnosis and when the disease relapsed. This was not directly related to tumour load, chemotherapy or neutropenia, but correlated well with episodes of intercurrent infection. In severe infections, fibronectin fell rapidly to very low levels, and was sometimes not restored to normal up to 2 weeks later. Preliminary attempts at replacement therapy were successful and the possible importance of this is discussed.
Joint involvement in patients with severe haemophilia A in 1957–59 and 1978–79
Authors: Helske T. , Ikkala E. , Myllylä G. , Nevanlinna H. R. , Rasi V.
Published: March 2008

Summary.

Clinical findings of joint involvement in patients with severe haemophilia A were compared between two surveys made in 1957‐59 and 1978–79. Permanent joint changes were seen in 50 (81%) of 62 patients in 1957‐59 and in 70 (86%) of 81 patients in 1978–79. The incidence of permanent changes was the same in the two surveys in all 10‐year age groups, but the degree of involvement was less severe in the two youngest age groups in 1978–79. It is concluded that effective replacement therapy can delay but not prevent completely the development of progressive arthropathy in patients with severe haemophilia A.
Antithrombin III transfusion in patients with hepatic cirrhosis
Authors: Schipper H. G. , Ten Cate J. W.
Published: March 2008

Summary. This study was designed to examine the effect of selective correction of antithrombin III activity on the increased turnover of fibrinogen, which occurs in patients with liver cirrhosis supposedly due to disseminated intravascular coagulation. Human antithrombin III concentrates were therefore transfused in seven patients with cirrhosis and antithrombin III deficiency (<80%). Fibrinogen half‐life and the fractional catabolic rate constant were calculated from the turnover of 125I‐fibrinogen which was represented by a two‐compartment model. Prior to antithrombin III transfusion, 125I‐fibrinogen half‐life was 76.7 ± 15.2 h and the fractional catabolic rate constant was 0.33 ± 0.11 of the plasma fibrinogen pool per day. In six healthy adult controls these values were significantly different: 109.4 ± 8.8 h and 0.19 ± 0.01 respectively. Correction of antithrombin III activity with human antithrombin III concentrate reduced the increased turnover of radiolabelled fibrinogen to normal. The 125I‐fibrinogen half‐life became 108.4 ± 17.6 h and the fractional catabolic rate constant decreased to 0.23±0.06. These observations indicate that decreased antithrombin III activity contributes in an important way to the increased 125I‐fibrinogen turnover in patients with cirrhosis and this might reflect intravascular coagulation.
Serological studies in patients on platelet‐ and granulocyte‐substitution therapy
Authors: Pegels J.G. , Bruynes E.C.E. , Engelriet C.P. , Kr. vondem Borne A.E.G.
Published: March 2008

Summary. A serological follow‐up study was undertaken in 47 patients with bone‐marrow failure, who were repeatedly transfused with random donor granulocytes and/or platelets. Sera, obtained at regular intervals, were investigated in the leucoagglutination test, the lymphocytotoxic test and the immunofluorescence test on paraformaldehyde‐fixed platelets, granulocytes and lymphocytes. The frequency of alloimmunization was high (73%). Not only HLA antibodies, but also blood‐cell‐specific alloantibodies were detected in the sera of the alloimmunized patients, e.g. lymphocyte‐specific, platelet‐specific and granulocyte‐specific antibodies.

The immunofluorescence test on platelets was also used as a crossmatch, and when this test was positive it was always found that after platelet transfusion the increment value was nil.
Relationship between platelet aggregation, thromboxane synthesis and albumin concentration in nephrotic syndrome
Authors: Jackson C. A. , Greaves M. , Patterson A. D. , Brown C. B. , Preston F. E.
Published: March 2008

Summary. Increased platelet aggregability on stimulation with sodium arachidonate (NaAA), collagen and ADP was found in a group of 14 patients with nephrotic syndrome when compared with age and sex matched controls. Five of the group also exhibited spontaneous platelet aggregation (SPA), associated with synthesis of thromboxane B2 (TxB2), and which appeared to correlate with a markedly increased serum triglyceride concentration.

Thromboxane B2 generation in response to NaAA was increased and reflected both the low serum albumin concentration and the increased platelet aggregation response to this agent. Addition of albumin in vitro decreased the amount of TxB2 generated for a given dose of NaAA and increased NaAA and collagen‐induced platelet aggregation thresholds. However, albumin had no significant effect on collagen‐induced TxB2 production.

The results suggest that the hypoalbuminaemia and associated reduced binding of arachidonic acid and increased synthesis of TxA2 account in part for the increased platelet aggregability seen in the nephrotic syndrome but that other mechanisms are also involved.
Immunohistological comparison of platelet factor 4 (PF4), fibronectin (Fn) and factor VIII related antigen (VIIIR:Ag) in human platelet granules
Authors: Giddings J. C. , Brookes L. R. , Piovella F. , Bloom A. L.
Published: March 2008

Summary. Immunofluorescence microscopy was used to study the localization of platelet factor 4 (PF4), fibronectin (Fn) and factor VIII related antigen in washed normal platelets and those from patients with severe von Willebrand's disease (vWd). Platelets were prepared by an improved multi‐unit modification of the Sayk chamber. This facilitated the preparation of samples with minimum disruption of platelets and readily permitted the demonstration of intact platelet granules by the immunological techniques used. The antigens were demonstrated by treating the same preparations sequentially with appropriate heterologous antisera and species specific fluorescein or rhodamine conjugated antisera. Comparison of the different antigens in identical platelets indicated that Fn and VIIIR:Ag were localized to the same granules as PF4 and the results were thus consistent with their presence in alpha granules. Fn and PF4, but not VIIIR:Ag, were present in platelet granules of patients with severe vWd. The antigens were always detected in the same granules, and major sub‐populations of differently stained granules were not observed. The methods were applied to investigate normal platelets aggregated with collagen. Fn and VIIIR:Ag were detected in platelet granules after aggregation although the granules themselves were possibly differently distributed in these samples compared with the non‐aggregated platelets. The localization of PF4 could not be reliably assessed in aggregated platelets by these methods. The techniques may be useful in localizing platelet antigens and studying release during aggregation.
4‐Hydroperoxyclophosphamide: a model for eliminating residual human tumour cells and T‐lymphocytes from the bone marrow graft
Authors: Körbling Martin , Hess Allan D. , Tutschka Peter J. , Kaizer Herbert , Colvin Michael O. , Santos George W.
Published: March 2008

Summary. Autologous bone marrow transplantation in acute leukaemia carries the risk of relapse from reinfusion of tumour cells present in marrow collected in remission and cryopreserved. An effective method for clearing marrow of tumour cells in required for a successful outcome. In the animal model 4‐hydroperoxycyclophosphamide (4‐HC) has proved to be effective in eliminating tumour cells from an autologous marrow graft. In the present studies, the in vitro effect of short‐and long‐term marrow cell incubation with 4‐HC on haemopoietic stem cells was investigated to determine the maximum concentration of 4‐HC that can be used for in vitro incubation without destroying the capacity of the marrow to effect complete haematological recovery as judged by residual CFUc content. However, loss of CFUc may not necessarily parallel survival of pluripotential stem cells. 4‐HC was also shown to be effective against peripheral T‐lymphocytes. Its possible therapeutic use in preventing or ameliorating graft‐versus‐host disease in allogeneic marrow grafts by preincubation with 4‐HC prior to transplantation is discussed.
Initial characterization of human platelet mepacrine‐labelled granules isolated using a short metrizamide gradient
Authors: Rendu F. , Lebret M. , Nurden A. T. , Caen J. P.
Published: March 2008

Summary. A method for the purification of human platelet mepacrine‐labelled granules is described. Characterization of these isolated granules allowed them to be identified as the serotonin storage organelles or dense bodies. Each step of the purification procedure has been controlled in order to obtain a minimum of leakage of the granule content during initial isolation of the platelets from the blood, the platelet washing procedures, and platelet lysis and the subcellular separation. A key step in the procedure was the centrifugation of the labelled granules across a short, discontinuous metrizamide gradient. The pellet of isolated mepacrine‐fluorescent granules consisted almost entirely of granules with the typical appearance of dense bodies, as shown by electron microscopy, and was relatively free from membranes and other granule populations as evaluated by the presence of the different markers (tritiated lectin, beta‐glucuronidase, monoamine oxidase, platelet factor 4). The method is simple, reproducible and allows the highest enrichment in dense bodies obtained hitherto with human platelets: x 177 in calcium and x 115 in [14C]serotonin after fractionation of [14C]serotonin‐labelled whole platelets. Functional studies performed with the isolated granules showed that they rapidly accumulated [14C]serotonin.
Self‐renewal and differentiation capacity of bone marrow and fetal liver stem cells
Authors: Zucali J. R.
Published: March 2008

Summary. An operational definition of the pluripotent stem cell (CFC‐S) requires that it have both the capacity for self‐renewal and the potential for differentiation into more than one class of formed blood elements. Because the CFC‐S compartment is heterogeneous, younger stem cells would be expected to be less committed to differentiation and have a higher rate of self‐renewal; whereas, older stem cells would be more committed to differentiation and have a lower rate of self‐renewal. In this study, the self‐renewal capacity versus the differentiation potential of adult bone marrow and fetal liver stem cells were compared. The self‐renewal potential was estimated by determining the number of CFC‐S which develop during growth in the spleen or femur of primary recipients. The differentiation potential was estimated by determining the total number of nucleated cells or committed progenitor cells (GM‐CFU‐C and BFU‐E) which develop during growth in the spleen or femur of primary recipients. In order to circumvent possible differences in self‐renewal or differentiation pressures due to the presence of differing numbers of CFC‐S, an equivalent number of bone marrow and fetal liver CFC‐S were allowed to seed the spleen and femur. While both adult bone marrow and fetal liver stem cells showed an extensive capacity for self‐renewal, fetal liver CFC‐S displayed a greater potential for self‐renewal in both the spleen and femur and at all growth intervals measured when compared to adult bone marrow CFC‐S. In contrast, no differences were seen in the number of nucleated cells or committed stem cells found per CFC‐S when comparing adult bone marrow and fetal liver stem cells.
Erythropoietin‐independent erythroid colony formation in patients with erythroleukaemia (M6) and related disorders
Authors: Anderson W. F. , Beckman B. , Beltran German , Fisher J. W. , Stuckey W. J.
Published: March 2008

Summary. In vitro erythropoietin (Ep) responsiveness of human bone marrow mononuclear cells was determined in 12 normal human volunteers and four patients with erythroleukaemia (EL), two patients with refractory anaemia with excess blasts (RAEB), and one patient with de novo acute myelogenous leukaemia (AML). The bone marrow cells were cultured in a microtitre methylcellulose system containing 30% human AB serum and human urinary Ep in concentrations ranging from 0 to 2 units/ml. Erythroid colony growth from normal marrow cultures was Ep‐dependent. It was augmented by added Ep and inhibited by Ep antiserum. Marrow cells from one patient with EL and one patient with RAEB after transformation to AML had no erythroid colony formation with or without added Ep. All of the remaining patients formed ‘spontaneous’or endogenous erythroid colonies (EEC) without the addition of Ep. In three of these (two with EL and one with de novo AML), the erythroid colony formation was augmented by added Ep. In three other patients (one with EL and two with RAEB), erythroid colony growth was unaffected by added Ep or Ep antiserum, and thus appeared to be Ep‐independent.