bluuebunny/crossref_metadata_2025 huggingface dataset:
Erythrocyte membrane protein alterations underlying clinical heterogeneity in hereditary spherocytosis
Authors: del Giudice Emanuele Miraglla , Iolascon Achille , Pinto Luciano , Nobili Bruno , Perrotta Silverio
Published: March 2008

Summary. Hereditary spherocytosis (HS) is a very heterogenous condition both at clinical and biochemical level. To establish the relationship between these aspects we performed a clinical and biochemical study in 87 Italian HS subjcts. Patients were divided into three groups based on clinical severity (mild, typical and severe) and into five subgroups based on specific membrane abnormalities identified by polyacrylamide gel electrophoresis (isolated spectrin deficiency, spectrin deficiency combined with mild ankyrin reduction, spectin deficiency combined with severe ankyrin reduction, band 3 reduction and isolated protein 4.2 reduction). We were not able to assess any alteration in six HS patients. A good correlation between clinical HS forms and memberane protein defects is shown. We conclude that erythrocyte memberane analysis should be carried out after diagnosis of HS in order to predict the clinical course of the disease.
Human cytomegalovirus (HCMV) infection in paediatric patients given allogeneic bone allogeneic bone marrow transplantation: role of early antiviral treatment for HCMV antigenaemaia on Patients' outcome
Authors: Locatellil Franco , Percivalle Elena , Comoli Patrizila , Accario M. , Zecca Maco , Giorgiani Giovanna , Gerna Giuseppe
Published: March 2008

Summary. In a prospective study, we evaluated the role of early treatment with ganciclovir of human cytomegalvirus (HCMV) pp65‐antigenaemia, as well as the risk factors related to the infection in 48 paediatric patients given an allogeneic bone marrow transplantation (BMT). HCMV infection occurred in 24 children, the overall actuarial risk of infection at 120 d being 51%. Development of acute graft‐versus‐host disease (GVHD), steroid therapy and serological status of both recipient and donor were the most powerful predictors of HCMV infection, none of the six seronegative patient/donor pairs developing HCMV infection. Considering only the seropositive recipients and patients given a seropositive marrow (42 cases), the actuarial risk of developing HCMV antigenaemia in patients with acute GVHD was 76%v 27% in those with or without GVHD (P < 0.005) and 79%v 15% respectively in patients who did or did not receive steroid therapy (P < 0.001). HCMV disease developed in 5/24 children with pp65‐antigen‐aemia, which was detected before diagnosis in all cases but one. All patients with pp65‐positive cells were treated with ganciclovir at a dose of 5 mg/kg twice daily for 14d. In patients without acute GVHD no maintenance therapy was adminis‐tered, whereas children with active acute GVHD were given additional therapy with ganciclovir at a dose of 5 mg/kg/d for 14d. Ganciclovir produced complete clearing of viraemia and antigenaemia, with some patients presenting recurrences of antigenaemia, which were treated according to the above‐mentioned schedule. Likewise, HCMV disease completely resolved after treatment with ganciclovir and no patients died from HCMV‐related interstitial pneumonia. Our results suggest that an early short‐term therapy with ganciclovir after demonstration of antigenaemia can be effective in reducing or abolishing HCMV‐related mortality. This approach elim‐inates the use of ganciclovir in patients not presenting HCMV reactivation and therefore not benefiting from therapy. The administration of ganciclovir limited to the period needed to obtain antigenaemia clearance could also have the advantage of reducing myelotoxicity.
The relationship between bcl‐2 expression and response to chemotherapy in acute leukaemia
Authors: Maung Z. T. , MacLean F. R. , Reid M. M. , Pearson A. D. J. , Proctor S. J. , Hamilton P. J. , Hall A. G.
Published: March 2008

Summary. Immunocytochemistry was used to assess bcl‐2 expression in blasts obtained from the bone marrow of 28 patients with acute lymphoblastic leukaemia (ALL) (16 children and six adults at presentation and three children and three adults on relapse) and 20 with acute myeloid leukaemia (AML) (19 adults and one child, 13 with de novo AML, 11 at presentation and two on relapse, and seven secondary to myelodysplasia or chronic myeloid leukaemia). Slides were examined both for the percentage of positive cells and for the intensity of staining using a five‐point scale. There was a statistically significant increase in both the percentage of positive cells seen (P < 0.002) and the intensity of staining (P < 0.01) between samples obtained at relapse and those at presentation in ALL. There was a significantly greater intensity of staining in cells from patients with ALL (P < 0.05) and AML (P < 0.05) who failed to achieve remission after chemotherapy than in those who responded. The intensity of staining in cases of secondary AML was lower than that in de novo disease (P < 0.01). These results suggest that expression of bcl‐2 may be an important prognostic feature in both de novo AML and in ALL, but not in secondary AML.
Impact of response to treatment on survival in multiple myeloma: results in a series of 243 patients
Authors: Bladé Joan , López‐Guillermo Armando , Bosch Francesc , Cervantes Francisco , Reverter Joan‐Carles , Montserrat Emilio , Rozman Ciril
Published: March 2008

Summary.Two hundred and forty‐three patients diagnosed with multiple myeloma (MM) in a single institution over a 22‐year period and treated with standard chemotherapy were analysed in an attempt to determine the impact of response to therapy on survival. The overall response rate in 229 evaluable patients was 50.1% (34.9% objective response plus 15.2% partial response). Median survivals of patients with objective and partial response were 43.4 and 42.8 months, respectively, versus 19 months for nonresponders. Median survival of 14 patients who achieved a complete remission was 42 months, whereas in 21 rapid responders (≤ 2 months) median survival was 43.3 months. A significant correlation between response and survival was observed with the landmark (P = 0.0169), the Mantel & Byar (P = 0.0001) and the Cox regression model (P < 0.0001) methods. These results indicate that, in responding patients, neither the degree of response nor the response kinetics has a significant influence on survival. However, the response to therapy is associated with a significantly longer survival in MM patients.
Efficient gene transfer into human umbilical vein endothelial cells allows functional analysis of the human tissue factor gene promoter
Authors: Nathwani Amit C. , Gale Karen M. , Pemberton Kieran D. , Crossman David C. , Tuddenham Edward G. D. , McVey John H.
Published: March 2008

Summary. Tissue factor (TF) is a cellular receptor and cofactor for factor VII/VIIa which initiates the blood coagulation cascade. We have investigated the role of 5′‐flanking DNA sequences in regulating the expression of the human TF gene in human umbilical vein endothelial cells (HUVEC). Using a chloramphenicol acetyltransferase (CAT) reporter gene, we attempted to transfect primary cultured HUVEC (passage 3–4) with calcium phosphate coprecipitation, DEAE Dextran, lipopolyamine‐coated DNA or electroporation. Electroporation in HEPES‐buffered saline of 1 × 107 cells at 200V and 250 μF was found to be optimal. Using these conditions, varying lengths of TF 5′‐flanking sequences coupled to the CAT reporter gene were tested in transient expression studies. CAT expression corrected for variation in transfection efficiency and cell viability revealed that the sequences between −111 and +14 base pairs are essential for minimal transcriptional activity. This region contains consensus sequences for a TATA box and three Sp 1 binding sites. A domain from −382 to −111 bp, which contains two AP‐1 consensus elements, promoted high levels of gene expression. This transcriptional activity was repressed by 50% with constructs containing sequences between −550 and −382 bp. A further 2‐fold drop in transcription activity was attributed to the region between −948 and −550 bp. These results suggest that the basal transcription of the human TF gene in HUVEC is mediated through at least two negative regulatory elements upstream of the proximal promoter domain. The proximal promoter region which contains two AP‐1 sites is essential for efficient transcription.
Epitope mapping of human factor IX inhibitor antibodies
Authors: Takahashi Isao , Mizumo Shinichi , Kamiya Tadashi , Takamatsu Junki , Saito Hidehiko
Published: March 2008

Summary. We have determined the location of epitopes on the factor IX for three haemophilia B inhibitor antibodies (HB‐1, HB‐3, HB‐7) and a monoclonal anti‐factor IX inhibitory antibody (designated 65–10). The main binding region of HB‐1, HB‐3 and HB‐7 was 155YVNSTEAETI164 (residues 155–164), 167NITQSTQSFN176 and 156VNSTEAETI164, respectively. The binding region of 65–10 was 168ITQSTQSFNDFTRVV182, which included the cleavage site (180R‐V181) for activation by factor XIa. By neutralization experiments using two peptides, 156VNSTEAETI164 and 167NITQSTQSFN176, the degree of neutralization of anti‐factor IX IgG purified by protein A was determined. Neutralization of three antibodies, HB‐1, HB‐3 and HB‐7, in the presence of 10mm of the peptides 156VNSTEAETI164 was 30.1%, 0% and 10.8%, respectively, and in the presence of 4 mm of 167NITQSTQSFN176 it was 0%, 13.5% and 17.3%, respectively. On the other hand, when plasmas of patients instead of purified IgG were used for neutralization, 10 mm of 156VNSTEAETI164 and 4 mm of 167NITQSTQSFN176 failed to neutralize the inhibitor in the plasmas.
High prevalence of a mutation in the factor V gene within the U.K. population: relationship to activated protein C resistance and familial thrombosis
Authors: Beauchamp Nicholas J. , Daly Martina E. , Hampton K. K. , Cooper Peter C. , Preston F. Eric , Peake Ian R.
Published: March 2008

Summary. Recent findings have indicated the importance of factor V (FV) in causing resistance to activated protein C (APC) in a high proportion of patients with venous thrombosis. This prompted us to investigate whether resistance could be due to defective inactivation of FVa by APC. Consequently, we amplified a 3.2 kb fragment of the FV gene sequence encoding the heavy chain APC cleavage site. DNA analysis showed a guanine to adenine transition at nucleotide 1691 in all affected members of two families with inherited APC resistance associated with thrombosis and confirmed suspected homozygosity in two individuals. The mutation, in heterozygous form, was also found in ˜3.5% of our normal population (n = 144) and correlated with low APC resistance. The high prevalence of this mutation suggests that it may be a major contributory factor in early thrombosis.
Retrovirus‐mediated transfer of the multidrug resistance gene into human haemopoietic progenitor cells
Authors: Bertolini Francesco , Monte Lucia De , Corsini Chiara , Lazzari Lorenza , Lauri Eleonora , Soligo Davide , Ward Maureen , Bank Arthur , Malavasi Fabio
Published: March 2008

Summary. We report the utilization of cord blood (CB) or bone marrow (BM) derived low density or purified CD34+ cells as a target for human multidrug resistance (MDR1) gene transfer, Cells were cocultivated for 48 h with an irradiated MDR1 retroviral producer line. Since some degree of MDR1 gene expression has been reported to occur in haemopoietic progenitor cells and in peripheral blood cells, effciency of MDR1 gene transfer was assessed by: (1) Drug selection and culture in presence of 50 ng/ml doxorubicin, 10 ng/ml colchicine and 0.85 μg/ml taxol. In uninfected control, 1–2% of CFU‐GM and CFU‐GEMM were found to be drug‐resistant, while 14–31% of original clonogenic activity was found after 2 weeks of culture of transduced cells. Efficiency of MDR1 transfer was significantly enhanced by prestimulation with cytokines, and found to be significantly superior in CB‐derived compared to BM‐derived progenitors. (2) Analysis of MDR1 gene expression by evaluating MDR1 mRNA through polymerase chain reaction. MDR1 expression was very low in cultures of uninfected controls, whereas, after drug selection, MDR1 mRNA levels in transduced cells was as high as in the MDR1 retroviral producer line (positive controls). (3) Flow cytometiric analysis of the expression of CD34 and P‐glycoprotein, the product of the MDR1 gene. After MDR1 transduction and 2 weeks of culture, membrane expression of P‐glycoprotien, was found on 17–25% of viable CD34+ cells. (4) Cytochemical localization by APAAP staining of P‐glycoprotein. No specific localization was found in untransduced controls, whereas transduced and cultured CB‐cells expressed P‐glycoprotein on plasma and nuclei membrane. In conclusion, MDR1 gene transfer into CB‐ and BM‐derived progenitor cells seems a feasible and attractive approach to generate a drug‐resistant haemopoiesis.
Ultrastructural localization of the small GTP‐binding protein Rap 1 in human platelets and megakaryocytes
Authors: Berger Gaëtan , Quarck Rozenn , Tenza Danié;le , Levy‐Toledano Sylviane , Gunzburg Jean de , Cramer Elisabeth Martin
Published: March 2008

Summary. Several functions have been proposed for Rap 1B in human platelets, including the regulation of phospholipase (PL) Cγ and Ca2+ ATPase. However, its localization is largely unknown. In the present study we have investigated the subcellular distribution of Rap1 by immunocytochemical techniques using affinity purified polyclonal antibodies raised against residues 121–137 common to the 95% homologous Rap1A and Rap1B proteins. By immuno‐fluorescence, a positive labelling was obtained on intact resting platelets and was abolished after adsorption of the antibodies with the control peptide. Immunoelectron microscopy was then used to further define the subcellular localization of Rap1B in platelets and megakaryocytes (MK). In resting cells, immunolabelling for Rap1B was associated with the plasma membrane, mostly at its inner face, and lined the membrane of the open canalicular system (OCS). Some labelling was also found outlining the α‐granules, identified as such by a double labelling with an anti‐GPIIb‐IIIa. On thrombasthenic platelets the same localization was observed. When platelets were stimulated by thrombin, immunolabelling for Rap1B was redistributed to the zones of fusion of the granules with the OCS, and to the plasma membrane with a higher concentration on pseudopods. Human MK expressed Rap1 and the staining revealed the association of the protein with the demarcation membranes and α‐granules.

This study presents a first approach to the localization of a small GTP binding‐protein Rap1B in whole platelets and MK, and shows its association with both the plasma and OCS membranes, as well as with the α‐granule membranes.
Involvement of eosinophils in acute myeloid leukaemia with monosomy 7 demonstrated by fluorescence in situ hybridization
Authors: Kwong Y. L. , Chan L. C.
Published: March 2008

Summary. Monosomy 7 is the commonest monosomy in acute myeloid leukaemia (AML). A patient with AML and monosomy 7 was investigated by fluorescence in situ hybridization. Monosomy 7 could be demonstrated in the blasts and also in mature eosinophils, but apparently not in lymphocytes. Monosomy 7 may occur in a multipotential myeloid progenitor cell as part of the multi‐step leukaemogenic process.
Location and PCR‐based detection of three polymorphisms of the human erythrocyte β‐spectrin gene (SPTB)
Authors: Gallagher Patrick G. , Lecomte Marie‐Christine , Galand Colette , Wang Yong‐Ping , Tse William T. , Forget Bernard G.
Published: March 2008

Summary. Defects of β spectrin, a structural protein of the erythrocyte membrane skeleton, have been identified in many cases of inherited disorders of red blood cell shape such as hereditary elliptocytosis and spherocytosis. To aid in genetic analyses of families with these disorders, the locations of three β‐spectrin gene (SPTB) polymorphisms were mapped and PCR‐based assays designed for their identification. Using these PCR‐based assays, the frequencies of these polymorphisms were determined in two populations.
Cytosine arabinoside (Ara‐C) plus alpha‐interferon (αIFN) determine prolonged complete remissions in patients with aggressive non‐Hodgkin's lymphoma partially responsive to first‐line doxorubicin‐containing regimens
Authors: Palmieri Giovannella , Morabito Alessandro , Rea Antonio , Caraglia Michele , Tafliaferri Pierosandro , Bianco Angelo Raffaele
Published: March 2008

Summary. Ten patients with aggressive NHL who failed to achieve a complete remission with first‐line chemotherapy were treated with Ara‐C and αIFN. Ara‐C was administered subcutaneously at 100mg on day 1, 150mg on day 2 and 200 mg on days 3,4 and 5 of a 28 d cycle; αIFN was given at 3 million International Units three times a week and continued for 2 years in CR patients. Six CR were attained with a median duration of 36+ months. Toxicity was mild. A new approach to second‐line therapy for aggressive NHL is proposed.
Chronic granulocytic leukaemia presenting with an extramedullary T lymphoblastic crisis
Authors: Jacob A. , Rowlands D. C. , Patton N. , Holmes J. A.
Published: March 2008

Summary. A 33‐year‐old man presented with hepatosplenomegaly and lymphadenopathy. bone marow findings were consistent with Philadelphia‐positive acceleratedphase chronic granulocytic leukaemia (CGL). Lymph node histology and immunhistochemistry were diagnostic of T lymphoblastic lymphoma. Molecular studies on lymph node DNA identified rearrangement within the major breakpoint cluster region (M‐ber), indicating a common clonal origin of CGL and the T lymphoblastic lymphoma.

We report the second case where extramedullary T lymphoblastic crisis was the presenting feature of CGL.
A deletional frameshift mutation in spectrin β‐gene associated with hereditary elliptocytosis in spectrin Napoli
Authors: Wilmotte R. , Miraglia del Giudice E. , Marechal J. , Perrotta S. , Mattia D. de , Delaunay J. , Iolascon A.
Published: March 2008

Summary. We studied a clinically manifest. dominantly transmitted elliptocytosis in an Italian family. We found a new spectrin variant, designated spectrin Napoli. Its β‐chain was truncated in its C‐terminal region (apparent MW 216kD. It its C‐terminal region (apparent MW 216kD. It displayed a low expression level (15%). There was a 8 nt deletion; CTTTTGAGAAGT → CTGT (nt 62556262), starting after condon 2053. This deletion was followed by a 54nt (18 amino acids missense sequence and terminated by the TGA triplet which normally overlaps codons 2074 and 2075 (CTTGAG0. The overall length of the mutrated β‐chain was comparable to that found in spectrin Nice, spectrin Tokyo and spectrin Tandil, which are other variants with truncated β‐chains; however, a distinct nonsense codon was used in spectrin Napoli.
Haemopoietic colony‐forming cells from peripheral blood stem cells harvests: cytokine requirements and lineage potential
Authors: Baines P. , Truran L. , Bailey‐Wood R. , Hoy T. , Lake H. , Ponton C. H. , Burnett A.
Published: March 2008

Summary. We have measured the in vitro growth requirements of progenitor cells released into the blood of cancer patients following administration of chemotherapy and cytokines. In order to distinguish the direct effects of cytokines on progenitors from those activating acessory cells, we have comparied clonogenic grwoth before and after CD34‐positive selection of progenitors, in serum‐free conditions. CD34 selection had little effect on the cytokine requirements of erythroid colony‐forming cells and single cytokines, particularly interleukin‐3, could support considerable colony growth in both mononulear and CD34+ cell suspensions. Optimal erythroid colony grwoth, however, usually required the addition of a combination of stem cells factor and interleukin‐3, in addition to erythoropoietin, which was was always required. Maximal numbers of granulocyte monocyte progenitors in mononuclear cell cultures, could be achieved with a mixture of stem cell factros, ionterleukin‐3 and granulocyte‐monocyte colony stimulating factor. How ever, after CD34 selection, full myeloid colony growth was only achieved when granulocyte colony stimulating factor was added to the above mixture. This presumably reflects loss of accessory cells, during CD34 selection, which produced this cytokine. When transplanted after 8 d of culutre. 16/22 myeloid colonies from erythorpoietin‐free cultures of peripheral blood stem cell harvests, could generate secondary multipotential. However, surface marker analysis of individual erythroid colonies revealed only the occasional presence of granulocytes and monocytes. These date demonstrate that cytokine mixtures are required for optimal colony growth, particuarly after CD34 selection, a nd that most mobilizied, blood clonogenic cells are multipotential.
Polycythaemia vera. IV. Specific binding of stem cell factor to normal and polycythaemia vera highly purified erythroid progenitor cells
Authors: Dai Chun‐Hua , Krantz Sanford B. , Koury Stephen T. , Kollar Kathleen
Published: March 2008

Summary. Polycythaemia vera (PV) patients' blood burstforming units‐erythroid (BFU‐E) have an enhanced sensitivity to stem cell factor (SCF) compared to normal BFU‐E. To characterize SCF receptors on erythroid progenitors from normal individuals and PV patients, we performed binding experiments using radioiodinated recombinant SCF (rSCF), day 1 BFU‐E and day 8 erythroid colony‐forming cells (ECFC), which are mostly colony‐forming units‐erythroid (CFU‐E). 125I‐rSCF binds to a single class of cell surface receptors (23 000/ECFC) at 0d́;C with a high‐binding affinity (Kd= 17pm). Saturation occurred at 0.5 nm (10ng/ml) which produces a nearly maximum biological effect. One half of the radiolabelled rSCF was internalized by the cells after 30 min at 37d́;C. No significant differences in the receptor number, dissociation constant, or internalization rate were found between normal and PV ECFC. Autoradiographic analysis of 125I‐rSCF binding to normal BFU‐E and ECFC showed that no differences were present in either the percentage of positive cells or the number of radioactive grains/cell between the normal and PV erythroid progenitors. The enhanced sensitivity of PV BFU‐E and CFU‐E to SCF does not appear to be related to changes in SCF receptor number, binding affinity or internalization and the hypersensitivity of PV erythroid progenitors to SCF must reside in a further internal cellular abnormality.
Interleukin‐3 administration enhances human monocyte function in vivo
Authors: Khwaja Asim , Addison Ian E. , Johnson Beryl , Yong Kwee , Linch David C.
Published: March 2008

Summary. In addition to its haemopoietic effects, interleukin‐3 (IL‐3) enhances leucocyte function in vitro. In this study we examined the effects on haematological variables and monocyte function of a single IL‐3 infusion in five haematologically normal individuals. There was a rapid fall in circulating monocyte (to 24 ± 6% of pre‐infusion value) and eosinophil numbers (to 3 ± 2%) with a nadir at 30 min and gradual return to baseline over 6h. No significant changes in monocyte expression of the adhesion molecules CD11b or L‐selectin or of monocyte respiratory burst activity were detected. There was a significant increase in monocyte phagocytosis and killing of Canadida after IL‐3 infusion: the percentage of monocytes which had ingested Candida increased from 39 ± 10% to 62 ± 12% and the total number of Candida killed per 100 monocytes increased from 63 ± 34 to 210 ± 59 (P < 0.05 and P < 0.01 respectively). There was no inhibition of neutrophil migration into a ‘skin window’ site and monocyte migration was moderately enhanced (peak increase of 260 ± 47%). These results show that IL‐3 has significant effects on monocyte function in vivo and could be of use in augmenting host defence mechanisms in immunocompromised patients.
Impaired TCR signal transduction, but normal antigen presentation, in a patient with common variable immunodeficiency
Authors: Fischer Michael B. , Hauber Ilona , Wolf Hermann M. , Vogel Erich , Mannhalter Josef W. , Eibl Martha M.
Published: March 2008

Summary. We describe a 27‐year‐old white man with common variable immunodeficiency (CVID) who has two healthy histoidentical brothers and one IgA‐deficient sister who shares one HLA haplotype with the patient. T cells from the patient with CVID showed an impaired response to recall antigens (tetanus toxoid, E. coli), whereas his IgA‐deficient sister and his two healthy histoidentical brothers responded normally. Cross‐mixing experiments using isolated monocytes and T cells from the CVID patient and one histoidentical brother revealed that the patient's monocytes were fully functional in processing and presenting antigen to resting T cells of his brother, and provided normal accessory cell function for superantigen‐induced activation of his brother's resting T cells. In contrast, the patient's T cells were unable to respond to antigen presented by the brother's monocytes and failed to respond with an increase in intracellular free Ca++ to stimulation with superantigen, which is known to bind to the TCR Vβ‐chain outside the antigen‐binding groove. However, stimulation with a combination of PMA and IM, directly activating protein kinase C and increasing intracellular free Ca++ by bypassing membrane receptors, induced normal Ca++ flux. These data indicate that the patient with CVID has a defect in TCR‐mediated signalling at the level of the T cells which is not present in his histoidentical healthy brothers or in his haploidentical IgA‐deficient sister.
A variant of spectrin low‐expression allele αLELY carrying a hereditary elliptocytosis mutation in codon 28
Authors: Randon J. , Boulanger L. , Marechal J. , Garbarz M. , Vallier A. , Ribeiro L. , Tamagnini G. , Dhermy D. , Delaunay J.
Published: March 2008

Summary. Allele αLELY is a low‐expression allele of the erythroid spectrin α‐gene. It carries mutations in exon 40 (αV/41 polymorphism) and intron 45, respectively, and is associated with partial skipping of exon 46. The latter phenomenon is thought to impair the recruitment of α‐chains by β‐chains, and would eventually account for the low‐expression character. When it occurs in trans to an α‐allele responsible for hereditary elliptocytosis (αHE allele; αHE/αLELY diplotype), allele αLELY enhances the severity of elliptocytosis. Because allele αLELY is widespread, we anticipated that it would occasionally carry HE determinants. These variants of allele αLELY will be designated αHE‐LELY alleles. We report two families with the same αHE‐LELY allele. The HE component was the known α28 Arg → His mutation. This αHE‐LELY allele was investigated within the αHE‐LELY/αLELY diplotype, a diplotye not described before. Except for the neonatal period, the presentation was mild. In a consistent manner, the αLELY component in cis of the αHE mutation counteracted the like component in trans.
Development of extended multidrug resistance in HL60 promyelocytic leukaemia cells
Authors: Su Gloria M. I. , Davey Mary W. , Davey Ross A. , Kidman Antony D.
Published: March 2008

Summary. In an attempt to mimic clinical conditions for the treatment of leukaemia, the HL60 promyelocytic cell line was treated for 18 h with low, clinically relevant, levels of the anthracycline epirubicin and the Vinca alkaloid vinblastine. The resulting drug‐resistant sublines not only expressed P‐glycoprotein and the MDR phenotype but were also cross‐resistant to chlorambucil, methotrexate but were also cross‐resistant to chlorambucil, methotrexate and cisplatinum, and had increased resistance to radiation. Development of resistance was associatted with an aberrant differentiation phenotype with decreased expression of myeloid antigens and expression of glycophorin A. an antigen normally associated with erythroid differentiation. The ability of HL60 cells to terminally differentiate in response to all‐trans‐retinoic acid (vitamin A acid) was lost in the sublines. These results suggest that either a single novel mechanism is responsible for multiple drug resistance or the initial response to drug treatment is the co‐induction of multiple mechanisms. These cells and the method by which they were generated therefore provide a clinically relevant model for the study of the initial events in the development of not only multidrug resistance but also the extended multiple drug resistance usually encountered in the treatment of leukaemia.
Possible misdiagnosis using the Xba I polymorphism for prenatal diagnosis of haemophilia A
Authors: Van De Water Neil S. , May Stephen J. , Browett Peter J.
Published: March 2008

Summary. A mother of two haemophilia A sons presented at 7 weeks pregnancy for a prenatal diagnosis. Sourthern blot analysis of haemophilia DNA at the factor VII intron 22 Xba I restriction fragment lenght polymorphism (RFLP) site revealed an Xba I haplotype of A‐B+C‐. This haplotype has been alluded to but not reported before, and when a 1.6kb Bst XI fragment of p482.6 is used as a probe the resulting band pattern is similar to that of females heterozygous at site A.
Reactivation of antibodies of donor and recipient origin to platelet antigens early after allogeneic bone marrow transplantation: a case report
Authors: Wernet Dorothee , Northoff Hinnak , Schnaidt Martina
Published: March 2008

Summary. Reactivation of platelet‐reactive antibodies of donor and recipient origin is described in a patient following allogeneic BMT (donor: anti‐HPA‐5b; recipient: anti‐HLA, anti‐HPA‐1b). The antibodies were detected around day 15 after BMT, peaked around day 25, and then decreased. These antibodies are interpreted as an antigen‐independent reactivation of secondary B‐cell responses, activated in the context of recognition of host antigens by the graft.
A spontaneous remission of lymphoid blast crisis in chronic myelogenous leukaemia following blood transfusion and infection
Authors: Lefrere Franclois , Hermine Olivier , Radford‐Weiss Isabelle , Veil Agnes , Picard Francoise , Dreyfus Francois , Flandrin Georges , Varet Bruno
Published: March 2008

Summary. We report a case of spontaneous remission of lymphoid blast crisis in chronic myelogenous leukaemia (CML) which returned to chronic phase, without the use of cytostatic chemotherapy. following an episode of viral infection and blood transfusion. Although complete remissions of acute leukaemia have been described, this evolution is extremely rare and has never been reported in CML blast crisis. The role of hypothetical factors leading to such a rare event are briefly discussed.
Transfusion‐associated graft‐versus‐host disease in fludarabine‐treated B‐chronic lymphocytic leukaemia
Authors: Maung Z. T. , Wood A. C. , Jackson G. H. , Turner G. E. , Appleton A. L. , Hamilton P. J.
Published: March 2008

Summary. Transfusion‐associated graft‐versus‐host disease (TA‐GVHD) is a rare but serious complication of blood component therapy in patients with haematological malignancies. B‐chronic lymphocytic leukaemia (B‐CLL), however, has rarely been associated with TA‐GVHD. We report three patients with advanced B‐CLL who developed TA‐GVHD. All these had been treated with fludarabine. Suppression of T cells by fludarabine may have contributed to an increased susceptibility to TA‐GVHD. The use of irradiated blood products to prevent this complication should be considered for patients with advanced B‐CLL treated with fludarabine or other purine analogues.
Protective effect of granulocyte‐colony stimulating factor against amphotericin B‐induced myelosuppression in vitro
Authors: Charak Bishan S. , Brown Ernest G. , Mazumder Amitabha
Published: March 2008

Summary Amphotericin B causes suppression of bone marrow (BM) progenitor cells in vitro. Granulocyte colony stimulating factor (G‐CSF) enhances the proliferation of myeloid cells. The present study defines the role of G‐CSF in preventing amphotericin B‐induced myelosuppression. G‐CSF increased the proliferative potential of BM and protected against amphotericin B‐induced myelosuppression if it was added to the medium during the early phase of exposure of BM to amphotericin B. Monoclonal antibodies to tumour necrosis factor‐α (TNF) or interferon‐γ (IFN) inhibited the myelosuppression partially: simultaneous presence of both these antibodies completely abrogated this suppression, suggesting that both TNFα and IFNγ were involved in amphotericin‐induced myelosuppression. TNF‐ or IFN‐ induced suppression of BM was also inhibited by G‐CSF. These data suggest that G‐CSF prevents the amphotericin B‐induced myelosuppression by antagonizing the suppressive effects of TNF and IFN and by enhancing the proliferative activity of BM.
Human resident peritoneal macrophages: phenotype and biology
Authors: Eischen A. , Duclos B. , Schmitt‐Goguel M. , Rouyer N. , Bergerat J. P. , Hummel M. , Oskam R. , Oberling Fr.
Published: March 2008

Summary . Macrophages represent the primary line of host defences in the peritoneal cavity. In order to study the metabolic activity and maturation stage of human resident peritoneal macrophages (PMø), peritoneal fluid (PF) was taken by Douglas puncture from healthy hyperstimulated infertile women undergoing oocyte retrieval for in vitro fertilization. Peritoneal fluid and macrophage culture fluids were studied for different inflammatory mediators such as interleukin‐1 (IL‐1), tumour necrosis factor (TNF) and interleukin‐6 (IL‐6). The level of macrophage colony‐stimulating factor (M‐CSF), which represents a macrophage proliferation and differentiation factor, was determined in the PF and in the serum. Furthermore, the macrophage phenotypic profile was analysed, in particular the expression of sex steriod hormone receptors. IL‐1, IL‐6 and TNF were detectable in the PF and in the culture supernatants of PMø whether stimulated or not by IFN‐γ and LPS. The mean level of M‐CSF in the PF was 6.37 ± 2.02 ng/ml as measured by RIA; this level did not correlate with the concentration of PMø. The mean PF‐M‐CSF level was 1.4‐fold higher than in the sera as measured by a EIA. Oestrogen and progesterone receptors could not be demonstrated on the PMø analysed, so that a direct relationship between the ovarian steroid concentration in these women and the function of PMø was unlikely. As compared to peripheral blood monocytes (Mo), PMø showed a phenotypic profile, with some more mature features, e.g. increased expression of CD14, CD68, FcRII, FcRIII, CR3, CR4 and MHC class II determinants. These results indicate that resident PMø have acquired in vivo a certain differentiation and/or activation state under micro‐environmental factors where cytokines secreted by the Mø themselves or by other cells such as the mesothelium may play important roles.
Prognostic value of serum IL‐10 and soluble IL‐2 receptor levels in aggressive non‐Hodgkin's lymphoma
Authors: Stasi Roberto , Zinzani Pier Luigi , Galieni Piero , Lauto Vito Michele , Damasio Eugenio , Dispensa Egidio , Dammacco Franco , Papa Giuseppe , Tura Sante
Published: March 2008

SummaryWe investigated the prognostic significance of interleukin‐10 (IL‐10) and soluble interleuckin‐2 receptor (sIl‐2r) levles in the pretreatment serum of 105 individuals with newly‐diagnosed aggressive non‐Hodgkin's lymphoma (NHL). Commercially available enzyem‐linked immunoassay kits were used for cytokine and receptor measurements. Detectable levels of IL‐10 were found in 42 (40%) patients at diagnosis, with no correlation with clinico‐haematological parameters, but in no control samples (P < 0.001).

Pretreatment concentrations of sIL‐2r were markedly increased in individuals with NHL when compared to controls (2614 ± 893 U/ml v 219 ± 65U/ml, P < 0.001), patients with stage III/IV presenting higher values than those with stage II disase (3885 ± 1196U/ml v 1732 ± 646U/ml, P < 0.001). No single parameter was associated with the achiveement of complete remission, but the combination of elevated IL‐10 and of sIL‐2r greater than 3000U/ml selected a subset of patients with a high probability of failing induction therapy (P < 0.001). Lifetable analysis also indicated thatj patients with these characteristics have a significantly shorter event‐free survival. In a multivariate analysis the combination of IL‐10 with sIL‐2r was found to have greater predictive strength than the combination of IL‐10 with β2‐micro‐globulin. We conclude that IL‐10 and sIL‐2r measurements can be expected to improve existing methods of risk assignment in aggressive NHL.
Anti‐CD36 antibodies in thrombotic thrombocytopenic purpura
Authors: Tandon Narendra N. , Rock G. , Jamieson G. A.
Published: March 2008

Summary. The membrane glycoprotein CD36 (GPIV, Mr 88000) is found on platelets, monocytes and endothelial cells of the microvasculature. In the present study, anti CD36 antibodies have been identified as occurring with high frequency in patients with thrombotic thrombocytopenic purpura. The presence of anti CD36 antibodies in 15 TTP plasma samples thought to contain them on the basis of an initial screening by protein blots was confirmed by re‐screening against a standard of purified CD36, by immunoprecipitation from 125I‐labelled control platelets and by dot blots against purified CD36. In a further 28 random samples examined, 23/27 (85%) were CD36‐positive by immunoprecipitation, 21/28 (75%) by protein blotting, and 17/28 (60%) by dot blots against purified CD36. On protein blots following SDS‐PAGE, immunoprecipitates produced from normal platelets by TTP plasma gave positive reactions with the anti CD36 monoclonal antibody 125I‐Mo91. One half of the total TTP samples examined (21/42) caused 70% release in control platelets loaded with 14C‐serotonin. Of samples causing release 70%, one‐half (8/15) failed to cause release from Naka‐negative platelets which constitutively lack CD36 showing that CD36 was the sole target for platelet activation in these TTP samples. These studies demonstrate that antibodies directed against CD36 occur frequently in TTP patients and could cause thrombotic complications and vascular damage by reacting with the parent antigen present in platelets and endothelial cells.
Blast cells in child and adult AML: comparative study of morphocytochemical, immunological and cytogenetic characteristics
Authors: Frenkel M. A. , Tupitsyn N. N. , Protasova A. K. , Kadagidze Z. G. , Konstantinova L. N. , Fleischman E. W.
Published: March 2008

Bone marrow blast cells of 174 child and 188 adult patients with AML were examined and characterized in terms of their FAB type, immunological phenotype (102 children, 123 adults) and karyotype (69 children, 95 adults). The incidence of FAB variants of AML proved similar in children and adults. In patients under 15 and over 60, peroxidase activity in myeloblasts was lower than in middle‐aged patients. Similar rates of HLA‐Dr, Thy‐1, CD11a, T‐cell antigens, CD19, Gly‐A and Eb antigens were found in cells of child and adult patients. The frequency of CD11b, CD38 and CD10 antigen expression on blast cells was higher in children than in adults. Abnormal blast karyotype was noted in 81.8% of children and 73.7% of adults. Translocation (8;21) was usually found in cases of M2 type (82%), significantly more frequently in children, predominantly in the group aged 6‐10. t(15;17) was detected in all age groups only in M3 type of cells (86%), t(9;22) occurred more frequently in adults than in children; t(11q23) incidence rates were somewhat higher in children than in adults. Three cases of AML in children are described with deletion of chromosome 5 in their leukaemic cells. The data obtained indicate different biological characteristics of blast cells in children and adults. It is likely that haemopoietic cell involvement in children under 2 years and adult patients over 60 occurs at earlier stages than in middle‐aged patients.
Evidence for a selective inhibitory effect of thrombin on megakaryocyte progenitor growth mediated by the thrombin receptor
Authors: Plantier Jean‐Luc , Berthier Rolande , Rival Yves , Schweitzer Annie , Rabiet Marie‐Josèphe
Published: March 2008

Summary. An efficient method for the culture of human megakaryocyte precursors in serum‐free medium has been developed facilitating study of the effect of regulators of megakaryocyte growth and maturation without interference by serum‐derived factors. We have investigated how megakaryocytes and their precursors respond to the procoagulant enzyme, thrombin. In addition to its already documented agonist effect on mature megakaryocytes, thrombin was found to have a marked inhibitory effect on the growth of megakaryocyte colonies from CD34+ bone marrow cells stimulated by IL3. This inhibitory effect, not previously reported, was selective for megakaryocytic cells. The growth of granulomonocytic and erythroid colonies was not affected. A monoclonal antibody which neutralized the effect of exogenous transforming growth factor β (TGFβ) was unable to fully neutralize the inhibitory effect of thrombin. With the use of a synthetic peptide, corresponding to the tethered thrombin receptor ligand, and of a recombinant inactive form of thrombin, we provide direct evidence that both the inhibitory effect of thrombin on megakaryocyte proliferation and its agonist effect on mature magakaryocytes are mediated by a receptor analogous to the recently cloned platelet thrombin receptor.
Evidence implicating calpain (Ca2+ ‐dependent neutral protease) in the destructive thrombocytopenia of the Wiskott‐Aldrich syndrome
Authors: Kenney D. M. , Reid R. , Parent D. W. , Rosen F. S. , Remold‐O'Donnell E.
Published: March 2008

Summary. The Wiskott‐Aldrich syndrome (WAS) is an inherited platelet/T‐lymphocyte disease characterized by small platelets, thrombocytopenia and immunodeficiency. Because degradative events have a significant role, we directly examined calpain (Ca2+‐dependent neutral protease), a prominent protease in the affected cells, by functional and antigenic quantitation. Calpain activity in platelets of seven WAS patients was decreased to 59 ± 3.7% (P < 0.01) relative to platelets of 11 normals. Platelets of two patients with immune thrombocytopenia had normal calpain activity. By immunoblotting, μ‐procalpain, the μ‐calpain species in resting (unstimulated) blood cells, was decreased in platelets of nine WAS patients to 58 ± 14.6% (P < 0.01) relative to paired normals. In contrast, μ‐procalpain levels in lymphocytes of seven WAS patients did not differ from normal lymphocytes. Normal platelets and lymphocytes have different mechanisms for Ca2+‐dependent μ‐procalpain activation. On addition of ionophore and Ca2+ to stirred platelets, 80kD μ‐procalpain was rapidly (0.5 min) and quantitatively converted to 76 kD active μ‐calpain; this process was the same in WAS platelets. In lymphocytes, μ‐procalpain activation was slow, only partially complete (40min), and the active species was 78kD. The marked depletion of calpain in WAS platelets demonstrated in this study may result from inappropriate stimulation of platelets and be related to the severe thrombocytopenia that characterizes this disease.
Nondeletional type of hereditary persistence of fetal haemoglobin: molecular characterization of three unrelated Thai HPFH
Authors: Winichagoon Pranee , Fucharoen Suthat , Wilairat Prapon , Chihara Kazuo , Fukumaki Yasuyuki
Published: March 2008

The β‐globin gene clusters of three unrelated Thai families with a nondeletional type of hereditary persistence of fetal haemoglobin (HPFH) were studied using polymerase chain reaction‐related techniques. All appeared to have normal nucleotide sequences from the Cap site to position ‐400 of both the Gγ‐ and Aγ‐globin genes. Two individuals suspected of having a β‐thalassaemia gene linked to the high HbF condition also had a normal β‐globin gene sequence, spanning from position ‐108 from the Cap site to the polyadenylation site. Deletion of four nucleotides, AGCA, at positions ‐225 to ‐222 of one Aγ‐globin allele was detected in one subject and was confirmed by dot‐blot hybridization. Restriction fragment length polymorphisms in the β‐globin gene cluster showed that the 5’ haplotype (‐ + ‐ ++) and the presence (+) of an Xmn 1 polymorphic site at ‐158 of the Gγ‐globin gene are associated with the high F phenotype in these families. Direct sequencing of the 5’ hypersensitive‐2 (5’ HS‐2) site of the locus control region (LCR) showed that this Xmn, 1 (+) site is also linked to a specific rearrangement of TA repeats (TA)9CACATATACG(TA)10, in HS‐2 segment.
Analysis of immunoglobulin class, IgG subclass and titre of HPA‐1a antibodies in alloimmunized mothers giving birth to babies with or without neonatal alloimmune thrombocytopenia
Authors: Proulx Chantal , Filion Mario , Goldman Mindy , Bradley Amanda , Devine Dana , Décary Francine , Chartrand Pierre
Published: March 2008

We analysed the titre and isotype composition of antibodies produced by mothers giving birth to babies with or without neonatal alloimmune thrombocytopenic purpura (NAITP) and patients with post‐transfusion purpura (PTP). All these individuals produced an antibody specific for the HPA‐1a allotype present on the platelet glycoprotein IIb‐IIIa (GPIIb‐IIIa). Sera from mothers who gave birth to thrombocytopenic babies (group 1, n = 36), non‐thrombo‐cytopenic babies (group 2, n = 4) or from PTP patients (group 3, n = 3) were tested by an indirect‐ELISA. Results indicated no evident differences in the isotype composition or titre of the antibodies from the three groups of sera. The antibody titre ranged from 1: 120 to 1: 3500. Antibodies with the IgGl subclass were present in all sera. Most sera contained IgGl alone (24/43 sera tested) or in combination with IgG3 (10/43). IgG2 was never present and only three sera showed intermediate reactivity with anti‐IgG4 MAb. Few sera (nine sera from groups 1 and 2) were weakly positive when tested with the anti‐IgM antibodies. These results suggest that neither the titre nor the isotype composition can be used to predict the severity or the occurrence of thrombocytopenia in newborns.
Multiple aberrant splicing of the p53 transcript without genomic mutations around exon‐intron junctions in a case of chronic myelogenous leukaemia in blast crisis: a possible novel mechanism of p53 inactivation
Authors: Nakai Hiroyuki , Kaneko Hiroto , Horiike Shigeo , Ariyama Yuko , Misawa Shinichi , Kashima Kei , Ishizaki Kanji
Published: March 2008

Summary. We found three truncated p53 transcipts in a patient with chronic myelogenous leukaemia in blast crisis carrying chromosome 17 abnormalities. Sequencing of these transcripts revealed complete absence of the entire exons 7, 8 and 9 in one, exons 8 and 9 in another, and exon 10 in the other. Sequencing analysis of genomic DNA, however, revealed no mutation in exons 6‐10 and their flanking introns. These results suggest that the aberrant p53 transcripts in this case might not result from splicing mutations but from an unknown affected splicing process.
Acquired sideroblastic anaemia following progesterone therapy
Authors: Brodsky Robert A. , Hasegawa Setsuo , Fibach Eitan , Dunber Cynthia E. , Young Neal S. , Rodgers Griffin P.
Published: March 2008

We report a case of acquired sideroblastic anaemia precipitated by progesterone. On two separate occasions, over 15 years apart, the patient developed sideroblastic anaemia with iron overload shortly after the administration of progesterone. No other cause for sideroblastic anaemia was found, and treatment with folic acid, pyridoxine or androgens corrected the anaemia. In both instances removal of the progestational agent led to prompt disappearance of the anaemia as well as the ringed sideroblasts. Using a two‐phase liquid culture procedure in which human peripheral blood‐derived progenitor cells undergo erythroid proliferation and differentiation. we demonstrated enhanced sensitivity of the patient's erythroid progenitors to progesterone. We conclude that progesterone should be added to the list of medications known to be associated with acquired sideroblastic anaemia.