bluuebunny/crossref_metadata_2025 huggingface dataset:
Donor lymphocyte infusion enhances remission status in patients with persistent disease after allografting for multiple myeloma
Authors: Kröger Nicolaus , Krüger William , Renges Helmut , Zabelina Tatjana , Stute Norbert , Jung Roman , Wittkowsky Georg , Kuse Rolf , Zander Axel Rolf
Published: March 2003

Two patients with persistent disease after allografting for multiple myeloma received donor T‐cell lymphocyte infusion (DLI) (1·5 × 108 and 7 × 107) to induce a graft‐vs.‐myeloma effect for further tumour regression after withdrawal of immunosuppression. The interval between stem cell transplantation and DLI was 8 and 14 months respectively. Both patients converted from partial to complete remission, lasting 12+ and 28+ months. Immunofixation became negative after 3 and 4 months. The main toxicity was grade II and III acute graft‐vs.‐host disease (GvHD) and limited or extensive chronic GvHD in each patient. We conclude that DLI induced further tumour reduction in patients with persistent disease after allografting for multiple myeloma.
Platelet glycoprotein IIb polymorphism, traditional risk factors and non‐fatal myocardial infarction in young women
Authors: Reiner Alexander P. , Schwartz Stephen M. , Kumar Prasanna N. , Rosendaal Frits R. , Pearce Rachel M. , Aramaki Kaelen M. , Psaty Bruce M. , Siscovick David S.
Published: March 2003

Several platelet glycoprotein polymorphisms have been associated with an increased risk of myocardial infarction (MI) in studies that included predominantly men. In a population‐based sample of 68 Caucasian women < 45 years old with non‐fatal MI and 346 demographically similar control subjects, we found an increased risk of MI among women who possessed at least one copy of the glycoprotein IIb Ser843 allele compared with those lacking the Ser843 allele (odds ratio 1·85; 95% confidence interval = 1·03–3·33). The increased risk was present only in subgroups of women who smoked cigarettes, had hypercholesterolaemia or who had a family history of early onset MI. The Ser843 variant of glycoprotein IIb may be associated with an increased risk of MI in young women with other cardiovascular risk factors. Additional studies involving larger numbers of subjects are needed to confirm this preliminary finding.
Platelet activation via the collagen receptor GPVI is not altered in platelets from chronic myeloid leukaemia patients despite the presence of the constitutively phosphorylated adapter protein CrkL
Authors: Best Denise , Pasquet Sophie , Littlewood Tim J. , Brunskill Susan J. , Pallister Chris J. , Watson Steve P.
Published: March 2003

In this study, we show that the adapter proteins CrkL and Cbl undergo increases in tyrosine phosphorylation and form an intracellular complex in platelets stimulated with the snake venom toxin convulxin, a selective agonist at the collagen receptor glycoprotein VI (GPVI). Constitutive tyrosine phosphorylation of CrkL has previously been reported in platelets from chronic myeloid leukaemia (CML) patients. This was confirmed in the present study, and shown to result in a weak constitutive association of CrkL with Cbl and a number of other unidentified tyrosine‐phosphorylated proteins. There was no further increase in phosphorylation of CrkL in CML platelets in response to GPVI activation, whereas phosphorylation of Cbl and its association with CrkL were potentiated. In addition, this was accompanied by a small increase in p42/ 44 mapkinase (MAPK) activity in CML platelets. The functional consequence of the presence of constitutively phosphorylated proteins in CML platelets was investigated by measurement of aminophospholipid exposure and α‐granule secretion. This revealed little alteration in the concentration–response curves for either in CML platelets stimulated via GPVI, although maximal levels of P‐selectin were depressed. Despite the minimal effect on platelet activation in CML patients, we cannot exclude a role for CrkL or Cbl in signal transduction pathways stimulated via GPVI.
Automated CD61 immunoplatelet analysis of thrombocytopenic samples
Authors: Kunz D. , Kunz W. S. , Scott C. S. , Gressner A. M.
Published: March 2003

Revision of the current decision point for prophylactic platelet transfusion in thrombocytopenic patients requires the availability of a method that is able to provide accurate platelet counts to as low as 1 × 109/l. This study is the first to evaluate the immunoplatelet method (CD61‐Imm) of the haematological analyser Cell‐Dyn 4000 in direct comparison with the flow cytometric procedure. Additionally CD61‐Imm results were compared with CD4000 optical (Plto) counts in the ranges 20–547 × 109/l (n = 127) and 1–35 × 109/l (n = 107). The immunoplatelet and Plto results were in good agreement between 20 × 109/l and 547 × 109/l, but for samples of < 25 × 109/l the Plto tended to overestimate the counts. We determined the limits of detection (LD) and quantification (LLQ) for all three methods using standard statistical procedures. The LD for the flow cytometric CD41a method was 0·02 × 109/l compared with 0·009 × 109/l and 1·73 × 109/l for the CD61‐Imm and Plto methods respectively. The LLQCV = 15% for the CD41a method was 1·8 × 109/l compared with 1·6 × 109/l and 18·0 × 109/l for the CD61‐Imm and Plto procedures. In conclusion, (i) the CD61‐Imm method performance is at least equivalent to the reference flow cytometric method, and (ii) in severe thrombocytopenia the CD61‐Imm count is superior to the Plto count.
Transient adhesion refractoriness of circulating platelets under shear stress: the role of partial activation and microaggregate formation by suboptimal ADP concentration
Authors: Savion Naphtali , Shenkman Boris , Tamarin Ilya , Dardik Rima , Frojmovic Mony , Varon David
Published: March 2003

Exposure of whole blood (WB) to subendothelial extracellular matrix (ECM) under shear stress in the cone and plate(let) analyser (CPA) results in platelet adhesion, followed by release reaction and aggregation of circulating platelets on the adherent platelets. The properties of circulating non‐adhered platelets in the CPA was studied by exposure of WB to ECM at a high shear rate (1300/s) for 2 min (1st run), followed by transfer of the suspension to a new ECM‐coated well for a second run (2nd run) under similar conditions. The results of the 2nd run demonstrated transient adhesion refractoriness associated with platelet microaggregate formation in the suspension. The adhesion refractoriness was dependent on platelet activation during the 1st run and was prevented by addition of apyrase (ADP scavenger) or ADP receptor inhibitor, suggesting a role for ADP in mediating this response. Furthermore, exposure of WB samples to suboptimal concentrations of ADP (0·4–1 μmol/l) or a thrombin receptor activating peptide (TRAP) (5 μmol/l) for 2 min resulted in a similar transient platelet adhesion refractoriness to ECM under flow conditions. The transient platelet refractoriness and microaggregate formation induced by ADP was associated with a transient reduction in glycoprotein (GP)Ib, increased P‐selectin expression and increased fibrinogen binding by circulating platelets. These data suggest a role for platelet agonists at suboptimal concentrations in modulating platelet function and limiting the expansion of the thrombus.
Induction of Toll‐like receptor 4 in granulocytic and monocytic cells differentiated from HL‐60 cells
Authors: Mita Yoshinori , Dobashi Kunio , Nakazawa Tsugio , Mori Masatomo
Published: March 2003

Toll‐like receptor 4 (TLR4) is the main protein expressed on the cell surface and is an essential receptor for lipopolysaccharide (LPS) signalling in human peripheral blood leucocytes. We examined TLR4 expression and the functional response to LPS in retinoic acid‐treated HL‐60 cells (HL‐60‐derived granulocytic cells) and interferon‐γ‐treated HL‐60 cells (HL‐60‐derived monocytic cells). Slight TLR4 expression was induced in HL‐60‐derived granulocytic cells, while strong induction was seen in HL‐60‐derived monocytic cells. LPS induced interleukin 1β (IL‐1β) production and TLR4 expression in HL‐60‐derived monocytic cells, but not HL‐60‐derived granulocytic cells. These data indicate different responses to LPS in the cells. TLR4 surface expression paralleled LPS‐induced phagocytosis and TLR4‐neutralizing antibody partially inhibited LPS‐induced IL‐8 production in HL‐60‐derived monocytic cells, but not in HL‐60‐derived granulocytic cells. These results suggest that HL‐60‐derived monocytic cells are partially activated via TLR4, but that HL‐60‐derived granulocytic cells are not activated via TLR4.
Acute myeloid leukaemia in human immunodeficiency virus‐infected adults: epidemiology, treatment feasibility and outcome
Authors: Sutton Laurent , Guénel Pascal , Tanguy Marie‐Laure , Rio Bernard , Dhedin Nathalie , Casassus Philippe , Lortholary Olivier ,   for the French Study Group on Acute Myeloid Leukaemia in HIV‐infected Patients
Published: March 2003

The epidemiology and clinical outcome of acute myeloid leukaemia in human immunodeficiency virus (HIV)‐infected adults is poorly documented. We retrospectively surveyed all French haematology centres for adult acute myeloid leukaemia (AML) cases diagnosed between January 1990 and July 1996 who were found to be HIV‐seropositive before or at the time of AML diagnosis. Medical charts were reviewed to determine the stage of HIV infection, the characteristics of AML and the response of AML to chemotherapy. Sixteen cases of AML (13 men, three women) were reported by 12 haematology units. Based on assumptions on the size, age and sex distribution of the HIV‐infected population in France, the estimated risk of AML in 1990 to 1996 among HIV‐infected adults was twice that of the general population (standardized incidence ratio = 2·05; 95% confidence interval, 1·17–3·34). Two other cases occurring before 1990 were spontaneously notified to the authors and were included in the clinical analysis. At AML diagnosis, the median CD4+ cell count was 275 × 106/l and nine patients had acquired immune deficiency syndrome (AIDS). Fifteen patients were scheduled for remission‐induction therapy of AML. No deaths were related to AML treatment. Complete remission was obtained in 11 out of 15 patients. Three patients were long‐term survivors: two remain alive in complete remission at 8 years and 9 years, respectively, and the third died of AIDS at 8 years. A CD4+ cell count above 200 × 106/l at AML diagnosis was predictive of longer survival (log‐rank test: P = 0·004). Like many other malignancies, the incidence of AML appears to be increased in HIV‐infected patients. Our results show a twofold higher incidence, although this needs to be confirmed in a specifically designed prospective epidemiological study. Such patients, especially those with CD4+ cell counts above 200 × 106/l at AML diagnosis, should receive remission‐induction therapy, which can confer long‐term survival.
Analysis of the expression of critical activation/interaction markers on peripheral blood T cells in B‐cell chronic lymphocytic leukaemia: evidence of immune dysregulation
Authors: Scrivener Sophie , Kaminski Edward R. , Demaine Andrew , Prentice Archie G.
Published: March 2003

B‐cell chronic lymphocytic leukaemia (B‐CLL) is characterized by an accumulation of clonal malignant B cells. The intrinsic characteristics that permit this accumulation have been extensively studied and described. However, it is possible that proliferation and survival of this malignant clone is facilitated by a disruption in the interaction between B and T cells that normally regulate the immune system. In this study, using flow cytometry and cell culture techniques, marked abnormalities of the expression of certain key activation and interaction molecules on the peripheral blood T cells of patients with B‐CLL were demonstrated. In particular, on comparison with normal controls, there was a marked reduction in the number of circulating T cells expressing CD25 (interleukin 2 receptor) (P = 0·007), CD28 (P = 0·01) and CD152 (CTLA‐4) (P = 0·001). There was also a reduction in the number of circulating T cells expressing CD4 (P = 0·03), CD5 (P = 0·05) and CD11a (P = 0·01). There was no difference in the number expressing T‐cell receptor αβ (P = 0·1), CD8 (P = 0·4), CD54 (P = 0·4) and CD154 (P = 0·5), and the only marker expressed on a greater number of circulating T cells in B‐CLL patients was HLA‐DR (P = 0·05). These results suggest that there is a profound T‐cell dysregulation that may contribute to the survival of the malignant B cells in patients with B‐CLL and to the related autoimmune phenomena of the disease.
Haemophilia B mutations in Sweden: a population‐based study of mutational heterogeneity
Authors: Ljung Rolf , Petrini Pia , Tengborn Lilian , Sjörin Elsy
Published: March 2003

The present series comprises all families (n = 77) with haemophilia B in Sweden and may be considered to be representative for the purposes of a population‐based study of mutational heterogeneity. The 77 families (38 severe, 10 moderate, 29 mild) had 51 different mutations in total. Thirteen families had total, partial or small deletions, two had mutations in the promoter, eight families had splice site mutations, 14 had nonsense and the remaining 41 had missense mutations. Ten of the mutations, all C→T or G→A, recurred in 1–6 other families. Using haplotype analysis of seven polymorphisms in the factor IX (FIX) gene, we found that the 77 families carried 65 unique, independent mutations. Of the 48 families with severe or moderate haemophilia, 23 (48%) had a sporadic case of haemophilia compared with 31 families out of 78 (40%) in the whole series. Five of those 23 sporadic cases carried de novo mutations, 11 out of 23 of the mothers were proven carriers and, in the remaining seven families, it was not possible to determine carriership. Eleven of the 48 patients (23%) with severe haemophilia B developed inhibitors and all of them had deletions or nonsense mutations. Thus, 11 out of 37 (30%) patients with severe haemophilia B as a result of deletion/nonsense mutations developed inhibitors compared with 0 out of 11 patients with missense mutations. The ratio of male to female mutation rates was 5·3 and the overall mutation rate was 5·4 × 10−6 per gamete per generation.
Intercellular adhesion molecule‐1 in extravasation of normal mononuclear and leukaemia cells
Authors: Mustjoki Satu , Alitalo Riitta , Elonen Erkki , Carpén Olli , Gahmberg Carl G. , Vaheri Antti
Published: March 2003

Interaction of intercellular adhesion molecules (ICAMs) with their receptors has a key role in normal leucocyte adhesion and migration, whereas in leukaemia this has not been well established. In this study, we have evaluated the roles of different adhesion molecules in normal and leukaemia cell extravasation in a novel organotypic model for vessel wall and measured plasma ICAM‐1 and ‐2 levels in acute leukaemia patients at diagnosis and during chemotherapy. We found that both normal mononuclear cells and blast cells from acute leukaemia patients, as well as retinoic acid‐treated promyelocytic leukaemia cells, rapidly extravasated through endothelial cell layers into the underlying collagen matrix. ICAM‐1 antibody prevented the extravasation, while antibodies to other adhesion molecules showed little (CD18, ICAM‐2) or no inhibition (CD11a and ICAM‐3). Soluble ICAM‐1 (sICAM‐1) protein had no effect. We also observed increased plasma sICAM‐1 and ‐2 levels in leukaemia patients and found that they correlated only weakly with the white blood cell count. No correlation was found between sICAM‐1 or ‐2 levels and the response to therapy. Although elevated sICAM‐2 levels decreased rapidly during chemotherapy, sICAM‐1 levels did not. Because sICAM‐1 protein had no effect on leukaemia cell extravasation in vitro, it is probable that the increased plasma sICAM‐1 levels in leukaemia patients may not play a role in leukaemia cell infiltration. However, as we showed that ICAM‐1 mediated leukaemia cell extravasation on the cell surface, it is possible that cellular ICAM‐1 has an important role in disease progression.
Two new alleles of the RHCE gene in Black individuals: the RHce allele ceMO and the RHcE allele cEMI
Authors: Noizat‐Pirenne France , Mouro Isabelle , Le Pennec Pierre‐Yves , Ansart‐Pirenne Hélène , Juszczak Geneviève , Patereau Claude , Verdier Martine , Babinet Jérôme , Roussel Michele , Rouger Philippe , Cartron Jean‐Pierre
Published: March 2003

Six unrelated individuals of Afro‐Caribbean origin, whose red cells have a marked reduction of the Rhe antigen expression, have been identified. All exhibited the same serological profile with anti‐e monoclonal antibodies and lacked expression of the high frequency e‐related antigen hrS. Transcripts and genomic analysis showed that these phenotypes resulted from the presence of two new RHCE alleles, ceMO and cEMI. The ceMO allele corresponded to a RHce gene carrying a G667T mutation (exon 5) and was detected at the homozygous state in sample 1 and at the heterozygous state in samples 2–6. The G667T mutation resulted in a Val223Phe substitution on the Rhce polypeptide, in close proximity to Ala226 (e‐antigen polymorphism), which might account for the altered expression of e. The ceMO allele is also associated with the lack of expression of the hrS antigen. The absence of the hrS antigen expression may have implications in transfusion as hrS‐negative individuals may develop clinically significant antibodies. The cEMI allele corresponded to a silent RHE allele carrying a nine nucleotide deletion within exon 3 and was detected at the heterozygous state in sample 2. This deletion resulted in a shortened polypeptide of 414 residues (instead of 417) that was absent (or severely reduced) at the red cell surface, as the E antigen was undetectable using serology and Western blot analysis with anti‐E reagents. In DNA‐based polymerase chain reaction genotyping for RHE determination, the cEMI allele provided a false positive result as the cells carrying this allele are serologically phenotyped as E‐negative. The incidence of this allele in the Black population is unknown but, as shown already for D genotyping, one must exercise caution when genotyping is performed to detect the e/E polymorphism.
Analysis of Vγ/Jβtrans‐rearrangements in paediatric patients undergoing chemotherapy
Authors: Lopes Luiz Fernando , Neto Emmanuel Dias , Lorand‐Metze Irene , Latorre Maria do Rosario D. O. , Simpson Andrew J. G.
Published: March 2003

The frequency of the hybrid Vγ/Jβtrans‐rearrangement in peripheral blood lymphocytes (PBLs) was analysed in a transversal study of paediatric patients (n = 210) with acute lymphoblastic leukaemia (ALL) and solid tumours (ST). Different amounts of DNA were used as the template for a nested polymerase chain reaction to evaluate the frequency of hybrid Vγ/Jβ genes, using silver‐stained gels. The frequency of the rearrangement was evaluated in groups before, during and after therapy. A greatly increased frequency of Vγ/Jβtrans‐rearrangement was found in PBLs of both groups of patients during exposure to chemotherapeutic agents compared with patients before chemotherapy. In patients who had finished treatment, the frequency of the rearrangement fell promptly to the baseline levels in ST but showed a slow decrease in ALL in those in whom increased levels could be found until 4 years after the end of treatment. We hypothesize that the chemotherapeutic agents are able to induce the Vγ/Jβtrans‐rearrangement, but this is transient in most cases. The exact relationship between the persistence of the rearrangement and the occurrence of secondary leukaemia remains to be determined.
High concentrations of coagulation factor VIII and thrombosis: is the factor VIII‐binding domain of von Willebrand factor implicated?
Authors: Bowen Derrick J. , Maclean Rhona M. , Pellard Sarah , Collins Peter W.
Published: March 2003

The possibility that high factor VIII (FVIII) levels in thrombosis patients is principally explained by a gain of function in the FVIII‐binding domain of von Willebrand factor (VWF), arising from amino acid substitution(s) or polymorphism(s), was investigated. Exons 18–24 of the VWF gene were sequenced in 13 thrombosis patients with high FVIII (> 1·50 IU/ml). No novel mutations were found. Four known polymorphisms were detected: G2615A and C2635T (Ex18), G2805A (Ex20) and G3130A (Ex22). Their frequencies showed no significant differences in a thrombosis vs. control cohort. The data suggest that amino acid substitutions/polymorphisms in the VWF‐FVIII‐binding domain are not the principal explanation for high FVIII in thrombosis patients.
Congenital sideroblastic anaemia successfully treated using allogeneic stem cell transplantation
Authors: Ayas Mouhab , Al‐Jefri Abdullah , Mustafa Mahmoud M. , Al‐Mahr Mohamad , Shalaby Loubna , Solh Hassan
Published: March 2003

Therapy for patients with congenital sideroblastic anaemia has been limited to blood transfusions and chelation. Three children with congenital sideroblastic anaemia (SA) who were blood transfusion dependent underwent stem cell transplantation (SCT) from matched sibling donors. Conditioning consisted of cyclophosphamide 50 mg/kg/d for 4 d, busulphan 4 mg/kg/d for 4 d and anti‐thymocyte globulin (ATG) 30 mg/kg for four doses pretransplant. Graft‐versus‐host disease (GVHD) prophylaxis was with cyclosporin A and methotrexate. All patients engrafted, and are alive and transfusion independent. SCT can be curative for patients with SA.
Evidence that natural killer cells express mini P‐glycoproteins but not classic 170 kDa P‐glycoprotein
Authors: Trambas Christina , Wang Zemin , Cianfriglia Maurizio , Woods Greg
Published: March 2003

Several lines of evidence including reverse transcription polymerase chain reaction, immunoreactivity and their ability to efflux rhodamine 123 have implied the existence of P‐glycoprotein in natural killer (NK) cells. It has been a natural tendency to assume that NK‐cell P‐glycoprotein is identical to the P‐glycoprotein of multidrug resistant (MDR) cell lines, however, the present study uncovered major differences. Functionally, NK cells demonstrated a restricted substrate profile, being unable to transport daunorubicin and calcein acetoxymethylester while efficiently transporting other P‐glycoprotein substrates. Furthermore, physical differences in NK‐cell P‐glycoprotein were established by differential reactivity with P‐glycoprotein antibodies. NK cells demonstrated strong reactivity with C494 and JSB‐1, but did not react appreciably with C219. In addition, NK cells were unable to bind to the antibody MM4·17 unless they had been fixed and permeabilized, yet this antibody normally recognizes an extracellular epitope of P‐glycoprotein. These differences culminated in the demonstration using Western analysis that NK cells did not express detectable levels of 170 kDa P‐glycoprotein. Instead, NK cells expressed small‐molecular‐weight ‘mini P‐glycoprotein’ products, of approximately 70 and 80 kDa. Collectively, these data indicate that the predominant P‐glycoprotein species of NK cells are novel mini P‐glycoproteins and not the classic P‐glycoprotein of MDR models.
Antisense oligodeoxynucleotides to latent membrane protein 1 induce growth inhibition, apoptosis and Bcl‐2 suppression in Epstein‐Barr virus (EBV)‐transformed B‐lymphoblastoid cells, but not in EBV‐positive natural killer cell lymphoma cells
Authors: Noguchi Toshio , Ikeda Kazuma , Yamamoto Kazuhiko , Yoshida Isao , Ashiba Atsuko , Tsuchiyama Junjiro , Shinagawa Katsuji , Yoshino Tadashi , Takata Minoru , Harada Mine
Published: March 2003

Epstein‐Barr virus (EBV)‐encoded latent membrane protein 1 (LMP‐1) is essential for immortalization of B cells by EBV, protects the infected cells from apoptotic cell death and induces Bcl‐2 expression. Suppression of LMP‐1 expression by antisense oligodeoxynucleotides (AS‐oligo) to LMP‐1 inhibits proliferation, promotes apoptosis and suppresses Bcl‐2 expression in EBV‐transformed B cells. However, the function of LMP‐1 expression in EBV‐positive natural killer (NK) cell lymphoma cells has not been reported previously. We examined the function of LMP‐1 in two EBV‐positive NK cell lymphoma cell lines (NK‐YS and YT) through suppressing LMP‐1 expression by AS‐oligo to LMP‐1. The AS‐oligo to LMP‐1 suppressed LMP‐1 mRNA and protein expression in two EBV‐positive NK cell lymphoma cell lines, as well as in an EBV‐transformed B‐cell line (CMG‐1). Proliferation was inhibited, apoptosis was induced and Bcl‐2 expression was suppressed in CMG‐1 cells, but none of these events were observed in NK‐YS or YT cells. These results suggest that proliferation, inhibition of apoptosis and Bcl‐2 expression in EBV‐positive NK cell lymphoma cells are not directly regulated by LMP‐1 as in EBV‐transformed B‐cell lines, but are probably mediated through other signal transducing systems.
Haemolytic onset of Wilson disease in a patient with homozygous truncation of ATP7B at Arg1319
Authors: Prella Maura , Baccalà Roberto , Horisberger Jean‐Daniel , Belin Dominique , Di Raimondo Francesco , Invernizzi Rosangela , Garozzo Rosaria , Schapira Marc
Published: March 2003

We describe a 19‐year‐old woman with haemolytic anaemia and thrombocytopenia as the initial manifestation of Wilson disease (WD). There are two reasons for reporting such an improbable case. First, it emphasizes the importance of recognizing atypical clinical presentations of potentially lethal recessive traits for which therapy is available. Second, it shows that, even in a monogenic disorder like WD, the phenotype cannot be extrapolated from the mutated genotype in a simple fashion; this patient had a relatively late‐onset form of WD despite homozygosity for a genetic lesion leading to an apparent complete loss of function of the WD copper transporter.
Clearance of maternal leukaemic cells in a neonate
Authors: Van Der Velden V. H. J. , Willemse M. J. , Mulder M. F. , Szczepański T. , Langerak A. W. , Wijkhuijs J. M. , Van Dongen J. J. M.
Published: March 2003

A 36‐week pregnant woman was diagnosed with acute lymphoblastic leukaemia. Delivery was initiated prematurely, and a healthy child was born. Cord blood and peripheral blood samples from the neonate (obtained at 6 weeks, 3 months and 6 months) were analysed for the presence of minimal residual disease by polymerase chain reaction analysis of a leukaemia‐specific IGH gene rearrangement and the E2A–PBX1 fusion gene transcript. In the cord blood sample, a tumour load of ≈ 4 × 10−4 was found, whereas all later blood samples were negative. Our data indicate that the maternal leukaemic cells did not engraft in the neonate.
High factor VIII levels contribute to the thrombotic risk in families with factor V Leiden
Authors: Lensen Ruud , Bertina Rogier M. , Vandenbroucke Jan P. , Rosendaal Frits R.
Published: March 2003

Factor V Leiden (FVL)‐carrying relatives of selected patients with venous thromboembolism (VTE) have much higher venous thrombotic risks than FVL‐carrying relatives of unselected consecutive patients with VTE. To find an explanation for this, we explored other risk factors of VTE, in particular the presence of high factor VIII levels, in a retrospective follow‐up study. We assessed levels of factor VIII, factor IX, fibrinogen, protein C, protein S, antithrombin, the presence of prothrombin 20210A, and the occurrence of VTE in 61 first‐degree relatives of 12 selected thrombophilic families harbouring FVL, and 183 first‐degree relatives of 47 unselected families of FVL carriers with a first VTE. In all families, FVL appeared to be an independent risk factor for VTE. Higher thrombosis incidence rates were found in carriers of both FVL and high factor VIII levels (≥ 150 IU/dl), while high levels of factor VIII appeared to be an independent thrombotic risk factor only in selected thrombophilic families. The fraction of individuals with more than one prothrombotic coagulation disorder was 10% higher in selected families. These results and the higher thrombotic risks we found in the thrombophilic families favour the hypothesis that other unknown co‐existing genetic defects contribute to thrombophilia.
Anti‐prothrombin antibodies combined with lupus anti‐coagulant activity is an essential risk factor for venous thromboembolism in patients with systemic lupus erythematosus
Authors: Nojima Junzo , Kuratsune Hirohiko , Suehisa Etsuji , Futsukaichi Yoshiaki , Yamanishi Hachiro , Machii Takashi , Kitani Teruo , Iwatani Yoshinori , Kanakura Yuzuru
Published: March 2003

Anti‐prothrombin antibodies (anti‐prothrombin) and anti‐β2‐glycoprotein I antibodies (anti‐β2‐GP I) are the most common and characterized anti‐phospholipid antibodies (aPL) detected using specific enzyme‐linked immunosorbent assay (ELISA) systems. Recently, lupus anti‐coagulant (LA) activity detected by a phospholipid‐dependent coagulation assay was reported to be associated with anti‐prothrombin and/or anti–β2‐GP I. Here we show that the co‐existence of IgG anti‐prothrombin and LA activity might be an essential risk factor for venous thromboembolism (VTE) in patients with systemic lupus erythematosus (SLE). We examined not only the levels of antibodies to prothrombin and anti‐β2‐GP I (both IgG and IgM isotypes) using an ELISA system, but also LA activity detected using both diluted Russell's viper venom time (dRVVT) and STACLOT LA test in 124 patients with SLE. The SLE patients were divided into four groups according to the results of ELISA and LA assay results for each aPL: group A, ELISA+ and LA+ group B, ELISA+ and LA−; group C, ELISA− and LA+ group D, ELISA− and LA−. Regarding IgG anti‐prothrombin, the prevalence of VTE was significantly higher in group A (16/35 cases, 45·7%, P < 0·001, Fisher's exact probability test) than in the other groups (B, 2/30, 6·7%; C, 1/22, 4·5%; D, 1/37, 2·7%). With respect to IgM anti‐prothrombin and IgG or IgM anti‐β2‐GP I, the prevalence of VTE was higher in both groups A and C than in group D, but no statistical difference in prevalence was found between groups A and C. Multivariate logistic regression analysis of risk factors for VTE confirmed that the co‐existence of IgG anti‐prothrombin and LA activity was the only significant risk factor for VTE (odds ratio, 19·13; 95% confidence intervals, 4·74–77·18).
Calcium ionophore: a single reagent for the differentiation of primary human acute myelogenous leukaemia cells towards dendritic cells
Authors: Waclavicek M. , Berer A. , Oehler L. , Stöckl J. , Schloegl E. , Majdic O. , Knapp W.
Published: March 2003

Blood monocytes and CD34+ haemopoietic progenitor cells, as well as certain leukaemic cell lines, acquire characteristics of mature dendritic cells (DC) after stimulation with calcium ionophore (CI). We studied whether the in vitro treatment of primary human acute myelogenous leukaemia (AML) cells with CI leads to differentiation towards DC. Blast cells derived from nine AML patients were cultured in the presence of either CI or an established differentiation cocktail consisting of granulocyte‐macrophage colony‐stimulating factor plus interleukin 4 and tumour necrosis factor‐α for 5–7 d. Microscopic examination revealed that under both conditions, AML cells were shifted along the DC pathway. In seven out of nine cases, CI‐cultivation led to a higher proportion of cells with dendritic morphology. The percentage of CD40 and CD86 expressing cells was significantly increased upon CI treatment compared with cytokine‐cultured cells. DC molecules as CD80 and CD83 were up‐regulated upon calcium mobilization of AML cells in four out of nine samples. In four cases, CI‐treated stimulator cells induced an enhanced proliferative allogeneic T‐cell response compared with cytokine‐treated stimulator cells. In conclusion, these data demonstrate that CI treatment is an alternative in vitro strategy to differentiate human AML cells into DC.
Transplant dose of CD34+ and CD3+ cells predicts outcome in patients with haematological malignancies undergoing T cell‐depleted peripheral blood stem cell transplants with delayed donor lymphocyte add‐back
Authors: Nakamura Ryotaro , Bahceci Erkut , Read Elizabeth J. , Leitman Susan F. , Carter Charles S. , Childs Richard , Dunbar Cynthia E. , Gress Ronald , Altemus Rosemary , Young Neal S. , Barrett A. John
Published: March 2003

We sought to optimize and standardize stem cell and lymphocyte doses of T cell‐depleted peripheral blood stem cell transplants (T‐PBSCT), using delayed add‐back of donor T cells (DLI) to prevent relapse and enhance donor immune recovery. Fifty‐one patients with haematological malignancies received a T‐PBSCT from an HLA‐identical sibling, followed by DLI of 1 × 107 and 5 × 107 CD3+ cells/kg on d +45 and +100 respectively. Twenty‐four patients were designated as standard risk and twenty‐seven patients with more advanced leukaemia were designated as high risk. Median recipient age was 38 years (range 10–56). Median (range) of CD34+ and CD3+ cell transplant doses were 4·6 (2·3–10·9) × 106/kg and 0·83 (0·38–2) × 105/kg respectively. The cumulative probability of acute GVHD was 39%. No patient died from GVHD or its consequences. The probability of developing chronic GVHD was 54% (18% extensive). The probability of relapse was 12% for the standard‐risk patients and 66% for high‐risk patients. In multivariate analysis, the risk factors for lower disease‐free survival and overall survival were high‐risk disease, CD34+ dose < 4·6 × 106/kg and CD3+ dose < 0·83 × 105/kg. Predictive factors for chronic GVHD were a T‐cell dose at transplant > 0·83 × 105 CD3+ cells/kg. These results further define the impact of CD34 and CD3 cell dose on transplant outcome and show that careful dosing of stem cells and lymphocytes may permit the control and optimization of transplant outcome.
The leukaemia‐associated transcription factors EVI‐1 and MDS1/EVI1 repress transcription and interact with histone deacetylase
Authors: Vinatzer Ursula , Taplick Jan , Seiser Christian , Fonatsch Christa , Wieser Rotraud
Published: March 2003

EVI‐1 and its variant form, MDS1/EVI1, have been reported to act in an antagonistic manner and be differentially regulated in samples from patients with acute myeloid leukaemia and rearrangements of the long arm of chromosome 3. Here, we show that both EVI‐1 and MDS1/EVI1 can repress transcription from a reporter construct containing EVI‐1 binding sites and interact with histone deacetylase in mammalian cells. This interaction can be recapitulated in vitro and is mediated by a previously characterized transcription repression domain, whose activity is alleviated by the histone deacetylase inhibitor trichostatin A.
Role of methylene blue‐treated or fresh‐frozen plasma in the response to plasma exchange in patients with thrombotic thrombocytopenic purpura
Authors: De La Rubia Javier , Arriaga Francisco , Linares Dolores , Larrea Luis , Carpio Nelly , Marty María L. , Sanz Miguel A.
Published: March 2003

Twenty patients with thrombotic thrombocytopenic purpura (TTP) underwent plasma exchange using either standard fresh‐frozen plasma (Group A, n = 13) or methylene blue‐treated plasma (Group B, n = 7). Both groups presented similar characteristics except that bilirubin values were higher in Group A (P < 0·05). The complete remission rate was higher in Group A than B (69% versus 57%). The mean number of procedures was higher in Group B (21 ± 7 versus 11 ± 3, P < 0·01) and the mean duration of hospitalization was also longer (37 ± 12 d versus 22 ± 11 d; P < 0·01). Our study shows that the use of methylene blue‐treated fresh‐frozen plasma to treat TTP is associated with a higher number of plasma exchanges and greater transfusion requirements without improving clinical results.
A novel multilocus genotyping assay to identify genetic predictors of stroke in sickle cell anaemia
Authors: Hoppe Carolyn , Cheng Suzanne , Grow Michael , Silbergleit Arkadiy , Klitz William , Trachtenberg Elizabeth , Erlich Henry , Vichinsky Elliott , Styles Lori
Published: March 2003

We describe a novel multilocus genotyping assay permitting simultaneous identification of 60 candidate markers for stroke in sickle cell anaemia (SCA). Based on cerebral magnetic resonance imaging (MRI), 69 patients were divided into stroke and control groups. The variant allele, CBS 278thr, showed protection from stroke, whereas the apoE3 allele showed a trend towards association with increased stroke risk. Several other variant alleles [TNFα (−308)A, CETP (−628)A, apoCIII (−641)A] showed a trend towards significant associations with stroke risk. These preliminary results on a small group of patients suggest that a multilocus genotyping assay may be valuable in identifying genes that increase the risk of stroke in SCA.
Pharmacokinetics of native Escherichia coli asparaginase (Asparaginase medac) and hypersensitivity reactions in ALL‐BFM 95 reinduction treatment
Authors: Müller Hans‐Joachim , Beier Rita , Löning Lutz , Blütters‐Sawatzki Renate , Dörffel Wolfgang , Maass Eberhard , Müller‐Weihrich Stefan , Scheel‐Walter Hans‐Georg , Scherer Franziska , Stahnke Karsten , Schrappe Martin , Horn Alexander , Lümkemann Karen , Boos Joachim
Published: March 2003

Repeated asparaginase treatment has been associated with hypersensitivity reactions against the bacterial macromolecule in a considerable number of patients. Immunological reactions may range from anaphylaxis without impairment of serum asparaginase activity to a very fast decline in enzyme activity without any clinical symptoms. Previous investigations on a limited number of patients have shown high interindividual variability of asparaginase activity time courses and hypersensitivity reactions in about 30% of patients during reinduction treatment. Therefore, monitoring of reinduction treatment was performed prospectively in 76 children with newly diagnosed acute lymphoblastic leukaemia (ALL). According to the ALL‐Berlin–Frankfurt–Münster (BFM) 95 protocol, 10 000 U/m2 body surface area of native Escherichia coli asparaginase (Asparaginase medac) was given on d 8, 11, 15 and 18. In 45/76 children, trough and peak activities were determined with every dose, and also on d 4 and d 11 after the last administration. Data on asparaginase activity were not available from the remaining 31 patients, but information with regard to hypersensitivity reactions only was given. Eighteen out of 76 patients (24%) suffered a clinical hypersensitivity reaction; however, no silent inactivation was observed. Activity in the therapeutic range of greater than 100 U/l for at least 14 d was determined in 43 of the 45 patients who were analysed for enzyme activity.
The inhibition of platelet aggregation and blood coagulation by Micropechis ikaheka venom
Authors: Sundell I. B. , Theakston R. D. G. , Kamiguti A. S. , Harris R. J. , Treweeke A. T. , Laing G. D. , Fox J. W. , Warrell D. A. , Zuzel M.
Published: March 2003

Uncoagulable blood and life‐threatening bleeding can result from the action of some snake venom toxins on haemostatic components of blood and vessel walls. Although envenoming by Micropechis ikaheka primarily affects neurones and muscle cells causing post‐synaptic neuromuscular blockade and rhabdomyolysis, disturbances of haemostasis also occur. Therefore, the present study explored the effects of M. ikaheka venom on platelets and endothelium, which are important components of the haemostatic mechanism. The venom inhibited platelet aggregation in response to ADP and collagen, and also delayed clotting dependent on platelet activation or endothelial cell tissue factor expression. Some of these effects were reduced by the incubation of venom with a phospholipase A2 (PLA2) inhibitor and could be reproduced by a 17 kDa venom fraction containing a PLA2. In addition, an 11 kDa fraction containing a long‐chain neurotoxin reduced ADP‐induced aggregation. The venom was also found to reduce endothelial cell adherence to vitronectin‐, fibronectin‐ and collagen‐coated surfaces. These results suggest that, by inhibiting procoagulant activities of platelets and endothelial cells, a 17 kDa PLA2 plays an important role in the anticoagulant action of M. ikaheka venom.
Glycoconjugate abnormalities in patients with congenital dyserythropoietic anaemia type I, II and III
Authors: Zdebska Ewa , Gołaszewska Ewa , Fabijańska‐Mitek Jadwiga , Schachter Harry , Shalev Hannah , Tamary Hannah , Sandström Herbert , Wahlin Anders , Kościelak Jerzy
Published: March 2003

Congenital dyserythropoietic anaemia type II (CDA II) is well known for glycosylation abnormalities affecting erythrocyte membrane glycoconjugates that encompass hypoglycosylation of band 3 glycoprotein and accumulation of glycosphingolipids: lactotriaosylceramides, neolactotriaosylceramide and polyglycosylceramides. These abnormalities were not observed in erythrocytes from patients with CDA of either type I or III. Recently, however, we have described a CDA type I patient in Poland with identical, though less pronounced, glycoconjugate abnormalities to those observed in patients with CDA type II. The abnormalities included partial unglycosylation of O‐linked glycosylation sites in glycophorin A. These abnormalities are now reported in three Bedouin patients from Israel with CDA type I. In addition, the erythrocyte membranes of these patients exhibited highly increased globotetraosylceramide content. Glycoconjugate abnormalities were also present in erythrocyte membranes from three patients from Northern Sweden with CDA type III but they almost exclusively affected glycosphingolipids. In erythrocytes of all patients examined including one with CDA type II, polyglycosylceramides were significantly hypoglycosylated although, on a molar basis, their contents in erythrocyte membranes were increased. Thus, glycoconjugate abnormalities of varying intensity occur in erythrocyte membranes from all patients with CDA that were investigated.
TAFI antigen and D‐dimer levels during normal pregnancy and at delivery
Authors: Chabloz Patrick , Reber Guido , Boehlen Françoise , Hohlfeld Patrick , De Moerloose Philippe
Published: March 2003

We have investigated whether the levels of thrombin‐activatable fibrinolysis inhibitor (TAFI) were correlated with d‐dimer levels during pregnancy and at delivery. From the 10th week of pregnancy to delivery, 519 samples from 144 women (mean age 29·3 ± 5, range 19–43) were obtained. We confirm the gradual increase of d‐dimer levels, and provide reference intervals for d‐dimer measurements throughout normal pregnancy. TAFI levels increased moderately during pregnancy but no inverse correlation with d‐dimer levels was observed.
Expression of co‐stimulatory and adhesion molecules and chemokine or apoptosis receptors on acute myeloid leukaemia: high CD40 and CD11a expression correlates with poor prognosis
Authors: Brouwer Rolf E. , Hoefnagel Juliette , Van Der Burg Boudewijn Borger , Jedema Inge , Zwinderman Koos H. , Starrenburg Ingrid C. W. , Kluin‐Nelemans Hanneke C. , Barge Renee M. Y. , Willemze Roel , Falkenburg J. H. Frederik
Published: March 2003

The expression of adhesion and co‐stimulatory molecules, and chemokine and death receptors such as tumour necrosis factor (TNF) and FAS on acute myeloid leukaemia (AML) may influence the biology of the disease and response to chemotherapy and immunotherapy. In this study, we analysed the expression of these molecules in 99 AML patients using monoclonal antibodies and flow cytometry, and correlated the expression with French–American–British (FAB) classification and survival. The following molecules were studied: the co‐stimulatory molecules CD80, CD86 and CD40; the adhesion molecules CD11a‐c, CD31, CD43, CD50, CD54, CD102, CD58 and CD62L; the chemokine receptor CXCR4; and the death receptors TNFR1 and TNFR2 and FAS. The expression of all molecules was significantly higher in the M4/M5 FAB subgroups except for CD80, CD43, CD54 and CD62L. The AML M3 subgroup had a significant lower expression of CD11a (P = 0·02) and CD11c (P = 0·03). Five‐year survival was significantly shorter in cases of high CD40 expression [> 20% positive cells, relative risk (RR) 2·56, P = 0·02] or high CD11a expression (> 80% positive cells, RR 2·6, P = 0·03). This effect was most prominently present in the AML M4/M5 FAB subgroups. We conclude that the expression levels of adhesion and co‐stimulatory molecules, CXCR4 and apoptosis‐receptors are predominantly FAB subtype‐related with high CD40 and CD11a expression as poor prognostic factors.
Protamine inhibits tissue factor‐initiated extrinsic coagulation
Authors: Chu Arthur J. , Wang Zhen‐Guo , Raicu M. , Beydoun Salwa , Ramos Nilda
Published: March 2003

The enhanced extrinsic coagulation in response to inflammation could contribute to disseminated intravascular coagulation, often manifesting cardiovascular complications. The complex mechanism remains unclear and effective management is not well established. The ability of protamine to offset bacterial endotoxin (LPS)‐induced tissue factor (TF)‐initiated extrinsic coagulation was demonstrated in human peripheral blood monocytes and cultured human leukaemia THP‐1 monocytes, which was consistent with the inhibition of rabbit brain thromboplastin (rbTF) procoagulant activity in a cell‐free in vitro model. Protamine significantly prolonged prothrombin time, further confirming the downregulation of the extrinsic pathway. However, thrombin time remained unaltered. Chromogenic assays were performed to dissect the extrinsic pathway, identifying inhibitory site(s). Protamine significantly inhibited factor VII (FVII) activation but not the dissected FX activation. The amidolytic activities of FVIIa and FXa were unaffected. The inhibited FVII activation in the presence of protamine was confirmed by the diminished FVIIa formation on Western blot analyses. Protamine preferentially inhibited TF‐catalysed FVII activation, downregulating the extrinsic cascade. Protamine could be of anticoagulant significance in the management of the extrinsic hypercoagulation.
Subcellular localization of thrombopoietin in human blood platelets and its release upon thrombin stimulation
Authors: Klinger Matthias H. F. , Jelkmann Wolfgang
Published: March 2003

Thrombopoietin (TPO) is a major regulator of platelet production. The concentration of circulating TPO seems to be determined by its binding and internalization by megakaryocytes and platelets. To elucidate the platelet compartments involved in TPO metabolism, we investigated intraplatelet TPO by post‐embedding immunoelectron microscopy, incubated platelets with recombinant human (rh)TPO coupled to colloidal gold and visualized the TPO uptake using electron microscopy. TPO concentrations were measured in 12 platelet concentrates (PC) before and after stimulation with thrombin and after disruption of platelets by freezing–thawing. In resting platelets, immunogold labelling revealed a prevailing cytoplasmic localization of TPO antigen and minor labelling within the surface‐connected canalicular system (SCCS); storage granules were devoid of labelling. In tracer experiments, TPO–gold was observed on the plasma and SCCS membranes and within the cytoplasm. Upon thrombin stimulation, endogenous TPO was still detected within the cytoplasm by immunolabelling, and tracer experiments revealed TPO–gold within the cytoplasm and on fibrin fibres. After thrombin stimulation of PC, the plasma TPO levels increased to an average of 535%, and after platelet lysis to an average of 1625% compared with plasma values in unstimulated PC. We conclude that platelets contain releasable immunoreactive TPO within the SCCS and within their cytoplasm, but not within granular compartments. Stored immunoreactive TPO is released upon thrombin stimulation, but only to a minor degree.
A spontaneous novel XK gene mutation in a patient with McLeod syndrome
Authors: Supple S. G. , Iland H. J. , Barnett M. H. , Pollard J. D.
Published: March 2003

A 29‐year‐old man with a history of elevated creatine kinase and necrotizing myopathy was reviewed. Prominent red cell acanthocytosis in association with reduced Kell antigen expression was present, findings consistent with the McLeod syndrome. Investigation of the patient's XK gene revealed a novel TGG‐ to‐TAG transition at position 1023 in exon 3. This point mutation creates an in‐frame stop codon (W314X), and predicts a truncated XK protein of 313 amino acids, compared with 444 amino acids in the normal XK protein. The mutation was not identified in the patient's mother or sister indicating that this mutation was spontaneous.