bluuebunny/crossref_metadata_2025 huggingface dataset:
Neutrophil‐bound immunoglobulin in HIV infection is of autoantibody nature
Authors: Klaassen R. J. L. , Vlekke A. B. J. , Kr. vondem Borne A. E. G.
Published: March 2008

Here we report the serological and immunochemical characterization of neutrophil‐bound Ig (NBIg) and neutrophil‐binding Ig in the serum of 15 individuals infected by the human immunodeficiency virus (HIV). We found no correlation between the presence or amount of NBIg or neutrophil‐binding Ig in serum and either the serum concentration of IgG or the level of immune complexes (IC), as determined by the C1q‐binding test. Twelve of the 15 eluates prepared from the neutrophils of the HIV‐infected individuals reacted with donor neutrophils. These results indicate that NBIg in these men was not adhered IC nor passively absorbed Ig. This was supported by analysis of the sera of 13 of the 15 men by sucrose density‐gradient centrifugation: neutroPhil‐binding Ig consisted predominantly of monomeric IgG. However, also low levels of IC capable of binding to neutrophils were detected in nine sera. Immunofluorescence studies revealed that neutrophil‐binding Ig in the sera reacted with neutrophil‐specific, non‐polymorphic antigens that are not attached to the cell membrane via phosphatidylinositol linkage and, more specifically, not located on neutrophil FcRIII. None of the sera and eluates showed reactivity in the immunoprecipitation technique. Moreover, none of the eluates reacted with blotted neutrophil glycoproteins, whereas three sera reacted reproducibly and five sera reacted occasionally with a number of glycoproteins of different molecular weights in this technique. The latter results probably represent reactivity against cryptic and/or cytoplasmic antigens. Thus, NBIg in HIV infection has an autoantibody nature, but the full identity of the target antigens could not be clarified. These characteristics are not essentially different from those of the autoantibodies in classical autoimmune granulocytopenia.
The treatment of anaemia in the myelodysplastic syndromes with recombinant human erythropoietin
Authors: Bowen David , Culligan Dominc , Jacobs Allan
Published: March 2008

Recombinant human erythropoietin was administered subcutaneously to 10 patients with myelodysplasia (MDS) who had haemoglobin concentrations less than 10 g/dl. in an attempt to relieve their anaemia. Doses of 60 units/kg/d rising to 90 units/kg/d were given over a maximum period of 16 weeks.

Two out of 10 patients showed a steady rise in haemoglobin concentration during treatment. One patient with refractory anaemia had a sustained rise from 9.9 g/dl 11.3 g/dl. and one patient with refractory anaemia with excess blasts (RAEB) had a rise from 9.5 g/dl to 11.4 g/dl but then relapsed with the development of an iron deficient state. Serum concentrations of immunoreactive EPO varied considerably between patients, but both responders had relatively low baseline levels.

Both responders were also new diagnoses and had received no red cell transfusions. The criteria for response to recombinant human erythropoietin therapy, as well as the indications for therapy remain to be clarified.
Interferon‐γin vivo reverses the increased platelet levels of platelet‐derived growth factor and transforming growth factor‐β in patients with myelofibrosis with myeloid metaplasia
Authors: Martyré Marie‐Claire , Magdelenat Henri , Calvo Fabien
Published: March 2008

We previously reported high platelet‐derived growth factor (PDGF) and transforming growth factorβ (TGF‐β) levels in platelets from patients with myelofibrosis with myeloid metaplasia. We report here the effects of in vivo administration of recombinant human interferon‐γ on these growth factor levels. Of six patients who entered this study in our institution, four completed the 6‐month therapy trial with interferon‐γ PDGF and TGF‐β levels in platelets were evaluated before and after treatment. PDGF levels were determined using a radioimmunoassay and TGF‐β activity was measured using an inhibition growth assay on CC164 cells. In these four patients, IFN‐γ therapy led to a reduction of the increased PDGF and TGF‐β levels in platelets towards the normal range. PDGF and TGF‐β values decreased by 25–45% and 32–46%, respectively. An acute deterioration was observed in one patient 5 months after the end of IFN‐γ therapy. In this patient, the deterioration of his clinical state correlated with an increase in intraplatelet PDGF and TGF‐β levels.
Plasma factor VII and thrombin–antithrombin III levels indicate increased tissue factor activity in sickle cell patients
Authors: Kurantsin‐Mills Joseph , Ofosu Frederick A. , Safa Toufic K. , Siegel Robert S. , Lessin Lawrence S.
Published: March 2008

Although the mechanisms involved in the persistent clinical complications of sickle cell disease have not yet been fully delineated, previous studies suggest that sickle cell (HbSS) patients have a disposition to generate more thrombin and plasmin in vivo than normal subjects. The reasons for the impaired regulation of haemostasis in HbSS patients is poorly understood. We report studies evaluating the extent to which in vivo coagulation and fibrinolysis are altered in HbSS patients in steady state. The concentrations of total factor VII (F(VII)t), factor VII zymogen (F(VII)z), thrombin–antithrombin III (TAT), fibrinopeptide A(FPA), and fibrin D‐dimer in plasmas of 50 normal controls (HbAA) and 45 HbSS steady state patients, were measured using sensitive and specific enzyme‐linked immunoassays. The average plasma concentration of F(VII)t, in sickle cell plasma was significantly lower than that of the control subjects (0·70 ± 0·19 U/ml versus 1·16 ± 0·41 U/ml), whereas F(VII)z in the patients and controls were 0·47 ± 0·15 U/ml and 1·15 ± 0·33 U/ml respectively, P<0·001. Both measures of factor VII suggest a higher factor VII turnover in sickle cell disease. The mean concentration of TAT in the plasma of HbSS patients were significantly higher than those of HbAA controls (371 ± 44 pM versus 42 ± 2 pM) (P<0·001), a difference that is strongly indicative of higher rates of in vivo thrombin generation by HbSS patients. Plasmas of HbSS patients had significantly higher concentrations of FPA compared to those of the control subjects (12·85 ± 1·96 ng/ml versus 4·22 ± 0·37 ng/ml) (P<0·001). The D‐dimer levels were also higher in the HbSS than control plasmas (1029·6 ± 58·6 ng/ml versus 224·3 ± 27·6 g/ml) (P<0·001), with the patients’values being indicative of enhanced fibrinolysis. These results strongly suggest accelerated in vivo coagulation and fibrinolysis in HbSS patients even during steady state. They are consistent with the hypothesis that haemostasis is less tightly regulated in the HbSS patients than in HbAA controls. The altered regulation of haemostasis may contribute to the initiation of vaso‐occlusive processes associated with sickle cell painful episodes.
A variant of Glanzmann's thrombasthenia which fails to express a GPIIb–IIIa related epitope that is recognized by a specific monoclonal antibody (C17)
Authors: Escolar G. , Diaz‐Ricart M. , Garrido M. , Reverter J. C. , Villamor N. , Sanz C. , Maragall S. , Castillo R. , Ordinas A. , Nurden A. T.
Published: March 2008

Binding of different antibodies to the GPIIb–IIIa complex in resting (AP2, EDU3, C17) or activated platelets (PAC1) was studied by flow cytometry in a patient with a platelet defect involving GPIIb–IIIa related functions. The patient has a mild history of bleeding. Aggregation induced by ADP and collagen were absent but normal response was obtained with ristocetin. Platelets from the patient do not bind fibrinogen. Perfusion studies with flowing blood showed that patient's platelets have a marked impairment in the process of spreading and aggregate formation on vascular subendothelium. Electrophoretic studies in SDS–polyacrylamide gels demonstrated the presence of normal amounts and normal mobility of GPIIb–IIIa. Fibrinogen was present in the patient's platelets (68–74% of controls). The binding of AP2 and EDU3 to patient's resting platelets was normal as assessed by flow cytometry. In contrast, a decreased presence of the C17 antigen (10 fold lower than control platelets) was detected in resting platelets and a markedly reduced binding of PAC1 was found in thrombin activated platelets. These studies suggest that C17 recognizes an epitope of the GPIIb–IIIa in resting platelets that is implicated in the regulation of adhesive and rdhesive properties of GPIIb–IIIa. Studies on this patient might be helpful for the understanding of GPIIb–IIIa functions.
The β‐thalassaemia mutations in the population of Cyprus
Authors: Baysal E. , Indrak K. , Bozkurt G. , Berkalp A. , Aritkan E. , Old J. M. , Ioannou P. , Angastiniotis M. , Droushiotou A. , Yüregir G. T. , Kilinç Y. , Huisman T. H. J.
Published: March 2008

We have identified the β‐thalassaemia alleles in nearly all known Turkish Cypriot β‐thalassaemia homozygotes and in over 700 Greek Cypriot β‐thalassaemia heterozygotes living on the island of Cyprus. The data confirmed earlier observations that the IVS‐I‐100 (G→A) mutation is present for about 74–80%, while three other alleles [IVS‐II‐745 (C→G), IVS‐I‐6 (T→C), IVS‐I‐1 (G→A)] occur at frequencies of 5–8%. Nearly identical percentages were observed for the two Cypriot groups, quite different from those for β‐thalassaemia patients from Greece and Turkey. This suggests close contacts between the two Cypriot communities during many centuries without a major recent influence from Greek or Turkish β‐thalassaemia carriers.
Subcutaneous injection of factor IX for the treatment of haemophilia B
Authors: Gerrard Ann J. , Austen Dennis E. G. , Brownlee George G.
Published: March 2008

Haemophilia B patients are normally treated, either prophylactically or in response to bleeding episodes, by frequent intravenous injections of factor IX purified from blood donors. Here we show in model animal experiments that purified human factor IX, when injected subcutaneously, is rapidly (in 3–11 h) and reasonably efficiently (30–40% of an equivalent intravenous dose) transported at least partly by the lymphatic drainage of the skin into the bloodstream, mostly in a biologically active form. This suggests that patients could be treated prophylactically by subcutaneous rather than intravenous injection, where the short delay in raising plasma factor IX to haemostatic levels would be clinically acceptable. More generally, our studies emphasize that the subcutaneous route of injection should be useful for other therapeutic proteins, including other clotting factors, which have to be delivered to the bloodstream, as long as their half‐life is at least a few hours allowing time for transport into the general circulation.
In vitro toxicity of 3′‐azido‐3′‐deoxythymidine, carbovir and 2′, 3′‐didehydro‐2′, 3′‐dideoxythymidine to human and murine haematopoietic progenitor cells
Authors: Du De‐Lin , Volpe Donna A. , Grieshaber Charles K. , Murphy Martin J.
Published: March 2008

Summary The myelotoxicities of three antiretroviral agents, 3′‐azido‐3′‐deoxythymidine (AZT), carbovir (CBV) and 2′,3′‐didehydro‐2′,3′‐dideoxythymidine (d4T), were evaluated in vitro with normal human and murine haematopoietic progenitor cells. These studies demonstrated that continuous AZT exposure was more inhibitory to human and murine colony formation than 1 h exposure, with murine and human progenitors similarly inhibited by continuous AZT exposure. These in vitro results on AZT's myelotoxicity correlate with both human and murine in vivo studies. CBV was only moderately toxic to human and murine cells following either 1 h or continuous exposure, with human and murine progenitors similarly suppressed by continuous CBV exposure. 1 h d4T exposure was less toxic to both human and murine marrow cells than continuous exposure and both species were equivalently inhibited when continuously exposed to d4T. In general, CBV was the least toxic agent to human and murine haematopoietic cells and AZT the most toxic. The study establishes CBV and d4T as less myelotoxic agents to human and murine haematopoietic progenitor cells in vitro than AZT which therefore could be considered as alternatives to AZT for the treatment of HIV infection.
Epidermal growth factor stimulates macrophage colony‐stimulating factor (M‐CSF) mRNA expression and M‐CSF release in cultured murine stromal cells
Authors: Abboud Sherry L.
Published: March 2008

Summary Macrophage colony‐stimulating factor (M‐CSF) released by stromal cells of the bone marrow microenvironment plays a crucial role in the growth and proliferation of mononuclear cells. Several peptide mitogens including interleukin‐1, tumour necrosis factor, platelet‐derived growth factor and fibroblast growth factor stimulate the release of M‐CSF and may be important in mediating the haematopoietic response to inflammation. Epidermal growth factor (EGF), released from platelets during aggregation, is mitogenic for a variety of cell types and may cause the release of certain cytokines. In this study we used the TC‐1 murine stromal cells which constitutively secrete M‐CSF as a model to study the regulation of M‐CSF in response to EGF. EGF markedly stimulated the steady state expression of M‐CSF mRNA with a peak effect observed at 3 h. This was associated with the release of M‐CSF protein as determined by radioimmunoassay. EGF also stimulated DNA synthesis in a concentration dependent manner. Although TC‐1 cells express GM‐CSF mRNA, this was not induced by EGF. These findings suggest that EGF is a key regulatory molecule for M‐CSF and may indirectly effect haematopoiesis via the release of M‐CSF from stromal cells.
Quality assessment of immunological marker analysis and the immunological diagnosis in leukaemia and lymphoma: a multi‐centre study
Authors: Van't Veer M. B. , Kluin‐Nelemans J. C. , Van Der Schoot C. E. , Van putten W. L. J. , Adriaansen H. J. , Van Wering E. R.
Published: March 2008

Summary In order to standardize and assess the quality of immunophenotyping of leukaemias and lymphomas for diagnostic purposes, a cooperative study group in the Netherlands, SIHON, has formulated guidelines for the composition of antibody panels to be applied and guidelines for the interpretation of the marker analysis. To assess the value of these guidelines frozen cell samples of three patients with different haematological malignancies were sent to the 26 participating laboratories twice a year. Here we present the results with respect to the marker analysis and to the immunological diagnosis on 387 samples from 18 patients. A large inter‐laboratory variation was seen in the percentage of positive cells for each marker, which influenced the valuation of a marker to be discordant positive in up to 23% and discordant negative in up to 40%. No single major factor could be traced to explain the large variation in the results. However, probably due to the balanced composition of the antibody panel and to the application of the guidelines for interpretation, this variation did not much influence the agreement in immunological diagnosis. In only 13/387 samples (3.3%) differences in the percentage of positive cells caused disagreement in the final diagnosis. In 23 samples (5.9%) the disagreement was due to an incorrect application of the guidelines. Quantitative data of single observations obtained from different laboratories, in which the materials and methods are not standardized, cannot be compared; but standardization of guidelines for marker sets and for interpretation contributes to a high grade of agreement in immunological diagnosis.
Acquired type II von Willebrand's disease associated with adrenal cortical carcinoma
Authors: Facon T. , Caron C. , Courtin P. , Wurtz A. , Deghaye M. , Bauters F. , Mazurier C. , Goudemand J.
Published: March 2008

Summary A case of acquired von Willebrand's disease (AvWD) associated with an adrenal cortical carcinoma is reported. The circulating highest molecular weight multimers (HMWM) of von Willebrand factor (vWF) were decreased when assessed by SDS‐agarose plasma electrophoresis, leading to the diagnosis of type II AvWD. No forms of inhibitor could be detected in the plasma of the patient. In contrast, indirect immunoperoxidase studies with a monoclonal antibody to vWF demonstrated an absorption of vWF into malignant cells. Infusion of a vWF‐FVIII concentrate, containing significant amounts of HMWM of vWF, allowed surgical resection of the tumour. After the first infusion of the concentrate, the vWF‐RCo recovery was found to be low (38%) compared to the vWF:Ag (75%) and FVIII:C (163%) recoveries. The resolution of all biological signs of vWD, including the abnormal multimeric pattern, in the postoperative period was prompt and permanent. Therefore, the absorption of the HMWM of vWF by carcinomatous cells appears to represent a likely pathophysiological mechanism responsible for the AvWD syndrome in this patient.
Antithrombotic activity, bleeding effect and pharmacodynamics of a succinyl derivative of dermatan sulphate in rabbits
Authors: Saivin S. , Caranobe C. , Petitou M. , Sie P. , Lormeau J. C. , Level M. , Crepon B. , Houin G. , Boneu B.
Published: March 2008

Summary This paper compares the pharmacological properties of a new succinyl dermatan sulphate derivative (Suc‐DS) to those of the natural dermatan sulphate (DS). Suc‐DS was on average 2–3 times more potent than DS in catalysing the inhibition of thrombin by heparin cofactor II and in prolonging the activated partial thromboplastin time and the thrombin clotting time. After bolus injection. Suc‐DS was also 2–3 times more potent than DS to prevent experimental venous thrombosis in a Wessler model. Thromboplastin or human serum were used as the thrombogenic stimulus. In contrast, the bleeding effect assessed by rat tail transection technique was comparable. After bolus intravenous injection, the pharmacodynamics of Suc‐DS indicated a lower volume of distribution, which was close to the plasma volume, and a slightly lower clearance of elimination. Therefore this chemical alteration of natural DS yields a new compound with an improved antithrombotic benefit/haemorrhagic risk ratio.
Use of several second generation serological assays to determine the true prevalence of hepatitis C virus infection in haemophiliacs treated with non‐virus inactivated factor VIII and IX concentrates
Authors: Watson Henry G. , Ludlam Christopher A. , Rebus Selma , Zhang Lin Qi , Peutherer John F. , Simmonds Peter
Published: March 2008

Summary To investigate the prevalence of hepatitis C virus infection in two risk groups, stored serum samples from treated haemophiliacs and intravenous drug users were tested for anti‐HCV by both anti‐C‐100 based and second generation ELISAs (Abbott and Ortho) followed by testing in two confirmatory immunoblot assays that incorporate core as well as other non‐structural antigens (Innogenetics LIA and Chiron RIBA‐HCV test). Clear evidence of HCV infection was found in all but one of 78 haemophiliacs treated with non‐virus inactivated clotting factor concentrates, but in none exposed only to super dry heat‐treated concentrates. Only four samples gave rise to conflicting serological results between the four tests, two of these occurred in patients with advanced HIV related disease and almost certainly reflected loss of humoral immunity associated with disease progression, and the others occurred in the only two patients tested who were chronic carriers of hepatitis B infection and may reflect an interaction between the two viruses. Comparison of anti‐C‐100 versus second generation tests in immunocompetent drug users revealed a false negative rate of 20% using C‐100 alone, indicating the advantage of using second generation assays for detection of past or current HCV infection. Of all of the antigens used in the confirmatory assay, positive sera showed strongest and most frequent reactivity with the C22 and C33c proteins (Ortho RIBA).
Post‐transfusion recovery of function of 5‐day stored platelet concentrates
Authors: Owens M. , Holme S. , Heaton A. , Sawyer S. , Cardinali S.
Published: March 2008

Summary Platelets show a rapid reduction in their responsiveness to aggregating agents during storage for transfusion, but little is known about reversal of this defect in vivo after transfusion. In this study, fresh and stored platelets from the same donor (n=12) were labelled with 111In or 51Cr, respectively, mixed, and simultaneously infused. Blood samples were taken for up to 5 d post‐infusion, and the functional behaviour of the labelled platelets ex vivo was measured by retention on glass bead columns, and by whole blood aggregability to ADP, epinephrine and ristocetin. Aggregation was determined by filtering aggregated samples through a column of cotton wool to remove the aggregates, and quantitated as per cent decrease in radioactive counts. The study showed that, although infused radiolabelled 5 d stored platelets had a significantly lower aggregability towards ADP and epinephrine immediately (1 h) after infusion (32% and 29%, respectively, of fresh platelet values), a complete restoration to fresh platelet levels was found 24–72 h post‐infusion, with no further change observed over the ensuing 5 d with either fresh or stored labelled platelets. A slightly (6–9%) lower adhesion to both uncoated and collagen‐coated beads was found for the stored platelets throughout the 5 d period of study post‐infusion.

In conclusion, these studies show that, with ex vivo testing, platelets stored for 5 d quickly recovered adhesion and aggregability capabilities similar to that of fresh platelets, suggesting that the functional lesion developed during storage is quickly and completely reversed after infusion.
Recombinant human erythropoietin is effective in correcting erythropoietin‐deficient anaemia after allogeneic bone marrow transplantation
Authors: Locatelli Franco , Pedrazzoli Paolo , Barosi Giovanni , Zecca Marco , Porta Fulvio , Liberato Lucio , Gambarana Daniela , Nespoli Luigi , Cazzola Mario
Published: March 2008

SummaryTwo children affected by severe aplastic anaemia (SAA) underwent allogeneic bone marrow transplantation (BMT) using partially matched family donors. In both cases there was a successful engraftment of donor haemopoietic stem cells. However, after an initial erythropoietic recovery, 5 months following BMT both children became severely anaemic. Although multiple factors were responsible for anaemia, in both cases there was a markedly impaired erythropoietin response to anaemia, as indicated by the inappropriately low levels of serum erythropoietin (EPO). Treatment with recombinant human erythropoietin (rHuEPO) induced a sustained erythropoietic response with complete correction of anaemia. This pilot study suggests that rHuEPO can be effective in correcting long‐lasting anaemia after marrow transplantation, characterized by inadequate erythropoietin production.
Effect of orally active hydroxypyridinone iron chelators on human lymphocyte function
Authors: Pattanapanyasat K. , Webster H. K. , Tongtawe P. , Kongcharoen P. , Hider R. C.
Published: March 2008

Summary Several iron chelators, 3‐hydroxypyridin‐4‐ones (CP) and desferrioxamine (DF) were compared for their effect on DNA synthesis, cell viability and expression of cell proliferation markers. Short‐term (4 h) exposure of human peripheral blood mononuclear cells to CP or DF inhibited the proliferative response of cells to concanavalin A (Con A). Inhibition by CP and DF showed a dose‐dependent effect with CP compounds more active than DF. Increased inhibitory activity of CP over DF was partly due to the lipophilic properties of CP. Pre‐saturation of CP and DF with exogenous ferric ion either diminished or prevented the inhibitory effect. At high chelator concentrations or prolonged (72 h) exposure of the cells to chelators, inhibition occurred but poor cell viability was observed.

In contrast to their inhibition of DNA synthesis, these iron chelators showed little effect on protein synthesis and the expression of transferrin receptors and interleukin‐2 (IL‐2) receptors. These findings suggest that both DF and CP compounds exert their effect by chelation of ferric ion with subsequent inhibition of DNA synthesis.
Establishment of a novel granulocytic sarcoma cell line which can adhere to dermal fibroblasts from a patient with granulocytic sarcoma in dermal tissues and myelofibrosis
Authors: Kobayashi Masanobu , Imamura Masahiro , Soga Ryosuke , Tsuda Yuuji , Maeda Shiro , Iwasaki Hiroshi , Sugiura Makoto , Ohizumi Hiroko , Musashi Manabu , Morioka Masanobu , Okabe Mihiro , Sakurada Keisuke , Kasai Masaharu , Miyazaki Tamotsu
Published: March 2008

Summary A novel human myeloid cell line, designated HSM‐1, has been established from the pleural effusion of a patient with granulocytic sarcoma (GS) who had been followed as having primary myeiofibrosis for 10 years. When he was diagnosed as having granulocytic sarcoma in dermal tissues, no evidence of malignant transformation into leukaemia was found in both the peripheral blood and bone marrow. The established cell line was positive for myeloperoxidase, Sudan black B. Naphthol AS‐D chloroacetate esterase. Surface marker analysis revealed that HSM‐1 expressed CD4. CD13, CD11a, CDllb, Leu8. CD49b. CD49d, CD49e, CD29 and HLA‐DR.

To clarify why the unusual myeloid tumours developed in non‐haematopoietic tissues, we examined the capability of HSM‐1 to bind to skin fibroblast layers. The HSM‐1 cells were found to bind to both bone marrow stromal layers and skin fibrobiast layers. Among the other myeloid ceil lines tested, none was found to bind to skin flbroblast layers. These findings suggest that the GS cell line may be derived from a haematopoietic precursor cell which can bind to skin fibroblasts and is localized in non‐haematopoietic tissues resulting in the formation of extramedullary myeloid metaplasia. HSM‐1 is a useful tool for analysing the characteristics of granuiocytic sarcoma and homing receptors for haematopoietic stem cells.
Comparison of plasma cell infiltration in bone marrow biopsies and aspirates in patients with multiple myeloma
Authors: Terpstra W. E. , Lokhorst H. M. , Blomjous F. , Th. Meuwissen O. J. A. , Dekker A. W.
Published: March 2008

Summary In 54 patients with multiple myeloma plasma cell infiltration was compared in bone marrow biopsies and aspirates. In 48% of cases plasma cell infiltration was comparable, in 48% infiltration in the aspirate was lower than in the biopsy. In only two cases more plasma cells were found in the aspirate. Eleven patients (20%) had less than 20% plasma cells in the aspirate and more than 50% in the biopsy. Underestimation of plasma cell load especially seems to occur in patients with a focal growth pattern of multiple myeloma or when strong fibrosis is present. 69% of patients with stage III. according to Durie & Salmon (1975), and 76% of patients with a high β2‐microglobulin had more than 50% plasma cells in the biopsy, indicating that these parameters, which are based on tumour load, are influenced by other factors as well.

The bone marrow biopsy is of superior value for direct estimation of the tumour load in multiple myeloma compared to bone marrow aspirates. A prospective study is needed to determine its prognostic significance.
Enhancement of human T cell responses to allogeneic stimuli by factor VIII concentrates
Authors: Batchelor Alison , Michael Steel C. , Ludlam Christopher A.
Published: March 2008

Summary The effect of factor VIII concentrate, from commercial and National Health Services manufacturers, on in vitro lymphocyte proliferative response to allogeneic stimulator cells was investigated.

Factor VIII preparations‘purified’by ion exchange or by monoclonal antibody affinity had no effect in this assay but lymphocyte proliferation in response to allogeneic cells was markedly and consistently enhanced by some intermediate purity factor VIII concentrates and. to a lesser extent, by a factor IX preparation. These preparations did not stimulate lymphocyte proliferation in the absence of other mitogens. The co‐mitogenic factor(s) present in intermediate purity factor VIII was not identified. However, enhanced proliferation was not due to factor VIII itself, nor to albumin or fibronectin. The clinical relevance of the immunomodulatory activity of intermediate purity factor VIII concentrates in vitro is discussed.
The production of human monoclonal antiplatelet auto‐antibodies derived from human lymphocytes of normal origin: reactivity to DNA, anionic phospholipids and platelet proteins
Authors: Denomme Gregory A. , Kelton John G. , Bell David A.
Published: March 2008

Summary Human hybridoma monoclonal antiplatelet antibodies were produced using tonsillar lymphocytes from a nonthrombocytopenic male fused to the lymphoblastoid cell line GM 4672. Twenty of 472 (4%) IgM producing hybridomas had antiplatelet reactivity as detected by ELISA. Thirteen of these antiplatelet antibody producing hybridomas with clonality ensured by limiting dilution were tested for anti‐genie specificity. Two different and mutually exclusive groups of antiplatelet antibodies were identified. The first group of antiplatelet antibodies (four clones) showed reactivity that was limited to DNA and anionic phospholipids. Antibodies from the second group (seven clones) showed reactivity by immunoblotting to a variety of platelet proteins including platelet glycoprotein IIb. These antibodies did not bind DNA nor anionic phospholipids.

These studies indicate that lymphocytes of normal human origin have the genetic potential to produce antiplatelet autoantibodies. These antiplatelet antibodies segregate on the basis of their target antigens into two major groups, which mimic the target antigens held responsible for antiplatelet autoantibodies in disease. These include glycoproteins (typical of chronic idiopathic thrombocytopenic purpura) and DNA and/or anionic phospholipids (typical of the lupus anticoagulant syndrome).
Cytotoxicity of activated platelets to autologous red blood cells
Authors: Okada Mariko , Kodama Takeshi , Tominaga Akio , Kon Kazunori , And Terutaka Sagawa , Utsumi Sayaka
Published: March 2008

Summary Gel‐filtered human platelets exerted lytic activity on autologous red blood cells (RBC) when they were co‐incubated at 37°C with platelet‐activating agents, such as thrombin, collagen, ADP. LPS or PMA in the absence of plasma. Lysis of activated platelets themselves did not occur during the incubation period examined. Morphological observations showed that RBC exposed to thrombin‐activated platelets were fragmented and/or transformed into spherocytes. This haemolytic reaction by thrombin‐activated platelets did not occur at 4°C, or in the presence of agents which inhibited glycolysis or elevated intracellular levels of cAMP, indicating that energy‐dependent and cAMP‐regulated platelet metabolism was required for this reaction. When platelets and RBC were incubated in the same vessel, but were prevented from coming into direct cell to cell contact by means of a membrane barrier, their cytotoxicity was reduced but not eliminated completely. No cytotoxic activity against RBC was detected in platelet‐free supernatants obtained by centrifugation after activation of platelets with thrombin. On the contrary, activated and washed platelets retained the activity. These observations suggested that the cytotoxic activity was carried by some diffusible and easily inactivated factors, which were continuously produced and liberated from activated platelets. Cyclo‐oxygenase inhibitors inhibited the haemolytic activity of thrombin‐activated platelets, suggesting a role for some products of platelet‐cyclooxygenase pathway in platelet‐mediated haemolysis. These results provide the first evidence for a direct role of activated platelets in mediation of RBC‐damage in the absence of any plasma factors.
Mitogenic role for extracellular calmodulin‐like activity in normal human umbilical vein endothelial cells
Authors: Dawson R. A. , Neil S. Mac
Published: March 2008

Summary Normal human umbilical vein endothelial cells cultured on gelatin‐coated plastic dishes were found to produce a protein in their media which had calmodulin‐like immunoreactivity and biological activity. Further identification of the protein was achieved by examining the incorporation of 14C leucine into protein found in the conditioned medium. Cells produced 14C labelled protein in their medium which specifically bound to an affinity column for calmodulin. This latter material stimulated calmodulin dependent phosphodiesterase activity in vitro and this stimulation was inhibited by the addition of the calmodulin antagonist W7. The presence of calmodulin‐like activity and immunoreactivity in the media varied as the cells grew from low to high density, a peak of extracellular calmodulin‐like activity preceding an increase in cell number. Extracellular calmodulin‐like activity did not correlate with the presence of lactate dehydrogenase in the medium. The addition of pure pig brain calmodulin affected the rate of cell proliferation; significant proliferation to pure calmodulin was only seen in cells at low density, at higher density calmodulin either had no effect or inhibited proliferation. Inhibition of extracellular calmodulin activity by a calmodulin antagonist immobilized on agarose beads, or by an antibody to calmodulin significantly decreased proliferation in all dividing cultures. Taken together this data suggests that, in vitro, calmodulin, or a very closely related protein, influences endothelial cell proliferation through an autocrine mechanism.
A pilot study of low‐dose cyclosporin for graft‐versus‐host prophylaxis in marrow transplantation
Authors: Stockschlaeder Marcus , Storb Rainer , Pepe Margaret , Longton Gary , McDonald George , Anasetti Claudio , Appelbaum Frederick , Doney Kris , Martin Paul , Sullivan Keith , Witherspoon Robert
Published: March 2008

Summary. Nineteen patients with leukaemia, preleukaemia, and aplastic anaemia were treated by marrow transplantation from HLA‐identical siblings. All were given postgrafting immunosuppression with a combination of methotrexate (days 1,3,6 and 11) and cyclosporin (days ‐1 to 180). In an attempt at reducing cyclosporin‐associated toxicity, we explored whether the cyclosporin dose during the first 2 weeks could be decreased by 50% (from 3.0 to 1.5 mg/kg/d intravenously) without adversely affecting the incidence, onset, and severity of acute graft‐versus‐host disease (GVHD) and overall survival. Results from this pilot study were compared to those of a matched cohort of 38 patients given a standard dose of 3.0 mg cyclosporin/kg/d starting on day ‐1. The cumulative incidence of grade II and III acute GVHD in the ‘low dose’ cyclosporin group was 42% compared to 51% in the ‘standard dose’ group (P= 0.60). Three‐year survival was 63% and 54% respectively (P=0.59). Patients receiving the reduced cyclosporin dose during the first 14 d appeared to have less hepatotoxicity, and the methotrexate and cyclosporin doses administered were closer to the doses intended per protocol. We suggest that‘low dose’cyclosporin from day ‐1 to day 15 postgrafting might be as effective as ‘standard dose’ cyclosporin during that time period for the prevention of acute GVHD.
The role of an anti‐myeloperoxidase antibody in the diagnosis and classification of acute leukaemia: a comparison with light and electron microscopy cytochemistry
Authors: Buccheri V. , Shetty V. , Yoshida N. , Morilla R. , Matutes E. , Catovsky D.
Published: March 2008

Summary. The enzyme myeloperoxidase (MPO) is the hallmark of the myeloid lineage. We have analysed the presence of MPO in blasts from 180 cases of acute leukaemia (103 acute myeloid leukaemia (AML) and 77 acute lymphoid leukaemia (ALL) by means of monoclonal antibodies anti‐MPO and immunocytochemistry (alkaline phosphatase anti‐alkaline phosphatase method). The aim of the study was to investigate the specificity and sensitivity of this marker compared with MPO cytochemistry by light (LM) and electron microscopy (EM), and with the expression of myeloid antigens. Anti‐MPO was positive (> 3% blasts) in all but one of the 90 AML positive by LM cytochemistry. Of 13 AML cases negative by MPO cytochemistry, six showed 3.10% blasts reactive with anti‐MPO and were also positive with antibodies to CD13 arid/or CD33. The presence of MPO was confirmed in four of these by EM. The overall positivity of anti‐MPO in AML was 92%. Anti‐MPO was negative in all but two ALL (6% and 8% positive blasts). The blasts in these two cases were also GD13, CD33 and MPO positive by EM; both were thus reclassified as biphenotypic. Another two ALL reinterpreted as biphenotypic were negative by MPO cytochemistry and anti‐MPO but were MPO positive by EM and with CD13 and/or CD33. We conclude that anti‐MPO is a sensitive and specific early marker of myeloid blasts and should be incorporated in the routine immunophenotyping of acute leukaemia.
A trial of recombinant human interleukin‐1 in patients with severe refractory aplastic anaemia
Authors: Walsh Christopher E. , Liu Johnson M. , Anderson Stacie M. , Rossio Jeffrey L. , Nienhuis Arthur W. , Young Neal S.
Published: March 2008

Summary. We report here the effects of in vivo administration of recombinant interleukin‐1 alpha (rIL‐1α) to patients with severe, idiopathic aplastic anaemia. Four patients who were refractory to immunosuppressive therapy and were not bone marrow transplantation candidates received daily doses of 0.03 μg/kg and 0.10 μg/kg intravenously as 5 d courses. No significant changes in either peripheral blood counts or bone marrow cellularity were observed at either dose during or following therapy. Two patients showed increased numbers of bone marrow progenitor colonies. Lymphocyte pheno typing demonstrated an elevated percentage of CD8 +/DR+ activated suppressor T lymphocytes prior to therapy. After rIL‐1α administration, the percentage of CD8 +/DR+ cells was reduced or returned to normal in all patients. Significant side‐effects included fever, rigours, fatigue, headache and nausea. Transient hypotension was observed at both doses in all patients. These results suggest that while rIL‐1α can be safely administered, no significant haematologic improvement was observed in patients with severe aplastic anaemia.
Interleukin‐2 induces activation of coagulation and fibrinolysis: resemblance to the changes seen during experimental endotoxaemia
Authors: Baars Johanna W. , de Boer Jan Paul , Wagstaff John , Roem Dorina , Eerenberg‐Belmer Anke J. M. , Nauta Jos , Pinedo Herbert M. , Hack C. Erik
Published: March 2008

Summary. The administration of Interleukin‐2 (IL‐2) causes the release or generation of other cytokines such as tumour necrosis factor (TNF) which, by disturbing the anticoagulant properties of the endothelium, may induce a procoagulant state in patients receiving this drug.

We therefore evaluated the effects of IL‐2 on coagulation and fibrinolysis in 14 patients receiving 12 or 18 × 106 IU/m2/d of IL‐2 given as a 15 min infusion for 5 d. Blood samples were drawn at short intervals after the first IL‐2 infusion. The parameters were analysed by way of analysis for repeated measures (F tests rather than t tests).

During the first day, thrombin‐antithrombin (TAT) complexes started to increase 2 h after the IL‐2 infusion, reaching peak levels at 4 h (n= 14; 11.2±6.4 μg/l v 49.8±49.2 μg/l, P < 0.01). Plasmin α2 antiplasmin (PAP) complexes showed a similar pattern rising from a mean baseline value of 17.5±7.6 nmol/l to 66.48±47.7 nmol at 4 h (P < 0.01). In four patients the peak of PAP preceeded that of TAT. Tissue plasminogen activator (tPA) rose from a mean baseline value of 4.9±3.7 μg/l to 26.3±13.5 μg/l at 4 h (P < 0.01). Plasminogen‐activator‐inhibitor‐1 (PAI‐1) levels increased from 59.35 μg/l to 113.39 μg/l at 6 h (P < 0.01). tPA PAI‐1 complexes increased from 0.15 ±0.07 to 0.69±0.21 nmol/l at 6 h (P < 0.01).

Our study indicates that IL‐2 activates the coagulation and fibrinolytic systems in vivo. The changes resemble the perturbations observed after endotoxin/TNF administration. These abnormalities may play a role in the side‐effects induced by IL‐2 therapy.
Immunophenotyping of fetal haemopoietic cells permissive for human parvovirus B19 replication in vitro
Authors: Morey Adrienne L. , Fleming Kenneth A.
Published: March 2008

Summary. Human parvovirus B19 is known to inhibit erythroid colony formation in vitro, but the precise stage of differentiation at which erythroid precursors become capable of supporting viral replication has not been accurately determined. In order to address this issue, haemopoietic cells derived from first trimester fetal liver were cultured in medium containing B19 antigen‐positive serum. Infected cells were phenotyped by combining immunohistology for cell‐type specific antigens with non‐isotopic in situ hybridization for B19 nucleic acid. Strong nuclear hybridization signal was detected as early as 8 h after infection in erythroid precursors labelling with antibodies to glycophorin A. glycophorin C, CD43, CD36 and HLA‐ABC (pronormoblast or normoblast phenotype). Giant erythroid precursors labelling with the same five antibodies were a pathognomonic feature of infected cultures, but contained relatively little B19 nucleic acid. Hybridization signal was not detected in progenitor cells of more primitive erythroid phenotype or in nuclei of cells of other lineages, though B19 DNA was occasionally localized within the cytoplasm of macrophages. Double‐labelling with antibody Ki‐67 confirmed that proliferating cells were targets for B19 infection. Co‐detection of cell‐type specific antigens and viral nucleic acid is a powerful tool for investigating host cell specificity, and suggests that proliferating late erythroid precursors are the only haemopoietic cells fully permissive for B19 infection.
The selective use of AMSA following high‐dose cytarabine in patients with acute myeloid leukaemia in relapse: a Leukemia Intergroup Study
Authors: Larson Richard A. , Day Roger S. , Azarnia Nozar , Bennett John M. , Browman George , Goldberg Jack , Gottlieb Arlan , Grunwald Hans , Miller Kenneth , Raza Azra , Vogler Ralph , Winton Elliot , Preisler Harvey D.
Published: March 2008

Summary. This clinical trial was designed to evaluate the role of high‐dose cytarabine (ara‐C) in the treatment of adults with acute myeloid leukaemia (AML) in first relapse. We also tested the hypothesis that the selective use of AMSA (100 mg/m2/d on days 7, 8 and 9) would increase the complete remission (CR) rate when leukaemia cells remained in the bone marrow immediately following 6 d of Ara‐C (2–3 g/m2/12 h) alone. Of 155 patients evaluable for response, 115 (74%) experienced marked cytoreduction by day 6 and received no further induction chemotherapy; 53 (45%) of these patients achieved CR after one course and 45 (38%) had resistant disease. The 36 patients (23%) with inadequate cytoreduction after the 6 d of ara‐C alone were randomly assigned either to no further chemotherapy (21 patients) or to 3 d of AMSA (15 patients). The CR rates after one course were 14% and 53%, respectively (P= 0001). and the fractions with resistant disease were 76% and 40%, respectively. The fractional reduction of leukaemia cells in the day 6 bone marrow aspirate specimen (P<0.0001) and the reduction in the leukaemia cell mass measured in the day 6 marrow biopsy (P= 0.001) were the strongest predictors for achieving CR versus having residual disease in univariate analyses. The median duration of remission was 5 months, but seven patients (10%) remain in CR after 30–92+ months. Among the 140 patients who received only the 6 d of ara‐C, the pretreatment albumin (P= 0.002) and lactate dehydrogenase (P= 0.01) levels were the strongest predictors of response in univariate analyses, but only the albumin remained significant (P= 0.01) in a stepwise logistic regression analysis. Those patients with albumin >4.0 mg/dl and LDH <125% of normal had a 71% CR rate, and only 16% had resistant disease. Among patients lacking both of these favourable characteristics, only 38% had a CR and 39% had resistant disease. Thus, pretreatment characteristics and rapid cytoreduction in the day 6 bone marrow sample identified a favourable subset of patients with AML in first relapse, some of whom responded quite well to 6 d of ara‐C alone and have had long disease‐free remissions.
Age‐related incidence and other epidemiological aspects of myelodysplastic syndromes
Authors: Aul C. , Gattermann N. , Schneider W.
Published: March 2008

Summary. Although most haematologists perceive a rising prevalence and incidence of myelodysplastic syndromes (MDS), reliable epidemiological data on these disorders are largely lacking. The bone marrow register of the University of Düsseldorf allowed us to assess among other epidemiological features the incidence of MDS. which was compared to that of acute myeloid leukaemia (AML). Among a total of 18416 different patients registered between 1975 and 1990, 584 cases of MDS (3.2%) and 506 cases of AML (2.8%) were identified. Over the study period, the percentage of newly diagnosed MDS rose from 1.3% to 4.5%, while there was no upward trend for AML. Among all patients undergoing bone marrow biopsy, the proportion of those over 60 years of age increased from 41.9% in 1975 to 54.1%in 1990. Wefounda strong correlation between the proportion of elderly patients and the relative frequency of MDS diagnoses. Thirty‐one patients (5.3%) were classified as a secondary MDS because of previous treatment with cytotoxic chemotherapy and/or irradiation for a variety of malignancies. Twelve patients were identified in whom occupational exposure to organic solvents could not be ruled out.

For calculating age‐specific incidence rates, the analysis was confined to the town district of Düsseldorf (575000 inhabitants), because exact demographical data were available for this population. In the last quinquennium of the study period (1986–90), myelodysplastic syndromes were more frequent than AML in the age group 50–70 years (4.9 v 1.8/100000/year). In patients over 70, the incidence of MDS was more than 3 times that of AML (22.8 v 6.7/100000). In this group, men had a higher incidence of MDS (33.9/100000) than women (18/100000). Crude annual incidence (all age groups) was also higher for MDS (4.1/100000) than for AML (2.1/100000) in recent years.

We conclude that MDS are relatively common haematological neoplasias. The rising incidence in recent years is probably not due to changes in aetiological factors, but may reflect increased awareness on the part of physicians and extended use of diagnostic procedures in elderly patients.
Treatment of haemophilia flexion deformities using the Flowtron intermittent compression system
Authors: Yates P. , Cornwell J. , Scott G. L. , Atkins R. M.
Published: March 2008

Summary. We have studied prospectively the use of the Flowtron intermittent compression system for the home treatment of fixed flexion deformities of elbows and knees, resulting from chronic haemophilic arthropathy. A total of 16 elbows (six unilateral and five bilateral), and 10 knees (six unilateral and two bilateral) in 11 patients were studied. Treatment caused a decrease in flexion contracture of the elbows from 41 ± 3° to 28 ± 3°(P<0.01), an improvement of 32%. The knees showed a decrease in flexion contracture from 31±3° to 20 ± 2° (P<0.0001), an improvement of 35%. Prophylactic factor replacement was not used during the study and there were no bleeds attributable to the treatment. We conclude that the use of the Flowtron in home treatment of fixed flexion deformities in haemophiliac patients is safe, effective and cheap.
A reversed activity staining procedure for detection of an acquired antibody against factor XIII in a girl with factor XIII deficiency
Authors: Seiving Birgitta , Henriksson Per , Stenberg Pål , Nilsson Inga Marie
Published: March 2008

Summary. A new method for detection of inhibitors against factor XIII is described. Agarose gel electrophoresis with normal platelet poor plasma as a factor XIII source included in the gel was combined with reversed activity staining. The procedure was applied to a 10‐year‐old girl with factor XIII deficiency where substitution therapy had failed. Plasma samples from the patient inhibited the factor XIII catalysed incorporation of the fluorescent monodansylthiacadaverine into casein at a position corresponding to the migration of γ‐globulins. Treatment with Sepharose‐bound protein A effectively adsorbed the factor XIII antibody in the patient's plasma.