bluuebunny/crossref_metadata_2025 huggingface dataset:
Minimally differentiated acute myeloid leukaemia (AML‐MO): cytochemical, immunophenotypic and cytogenetic analysis of 19 cases
Authors: Venditti Adriano , Poeta Giovanni Del , Stasi Roberto , Masi Mario , Bruno Antonio , Buccisano Francesco , Cox Cristina , Coppetelli Ugo , Aronica Germano , Simone Maria Domenica , Papa Giuseppe , Tribalto Maurizio , Amadori Sergio
Published: March 2008

Summary. We describe our experience in the identification of 19 cases of AML‐MO categorized among 200 consecutive AML cases. Leukaemic cells from our cases were morphologically marked by agranuler basophilic cytoplasm, finely dispersed chromatin and prominent nucleoli. In two cases heavily vacuolated and monocytoid‐shaped blasts were also observed. Cytochemistry (MPO, SBB, αANAE, αNBE, NASDCAE, AP, PAS) was negative in 14 cases, five cases expressing a very faint cytoplasmic positivity for αNBE (not exceeding 30% of the blasts) and αANAE (not exceeding 41%) which was sodium fluoride resistant. In these five cases other monocytic markers (e.g. CD14) were not in favour of myelomonocytic differentiation. All the cases were anti‐MPO positive at frequency > 10%. Phenotypic analysis also revealed myeloid features with all the patients having at least one myeloid antigen (CD13, CD33, CD15), Tdt was expressed in nine cases and CD7 in six cases. All cases but one were positive for CD34. Cytogenetic analysis, performed in 16 cases, showed no adequate growth in two cases and no consistent abnormality in four; among the remaining 10 cases no consistent abnormality was observed, the most common finding was trisomy 8 (two cases) and 4 (two cases) and aberrations of chromosomes 2, 3, 5, 7, 9, 12 and 21. No cases of (t9;22), Ph chromosome were observed. Interestingly three out of five patients with faint αNBE/αANAE positivity relapsed as typical M4 (one case) or M5a (two cases).
Molecular analysis of the erythropoietin receptor system in patients with polycythaemia vera
Authors: Hess Georg , Rose Petra , Gamm Heinold , Papadileris Stefan , Huber Christoph , Seliger Barbara
Published: March 2008

Summary Erythropoietin (EPO) is a potent regulator of the viability, proliferation and differentiation of erythroid progenitor cells. Its effect is mediated by binding to the erythropoietin receptor (EPO‐R), a member of a new cytokine receptor family. Alterations of the EPO/EPO‐R system have recently been shown to be involved in the pathogenesis of familial erythrocytosis and polycythaemia vera (PV). In order to define whether genetic changes in the EPO‐R gene and its ligand play a role in the development of PV, the structure and expression levels of the EPO‐R and EPO genes were examined in samples from bone marrow and/or peripheral blood mononuclear cells of 24 patients with PV. As expected, EPO serum levels were low and no detectable level of EPO mRNA was found by reverse polymerase chain reaction (RT‐PCR) in the peripheral blood mononuclear cells of our PV patients. To search for structural alterations of the EPO‐R, cDNA samples were subjected to PCR and SSCP analysis as well as sequencing. Heterogenous expression of EPO‐R mRNA was observed without any structural changes, as revealed by RT‐SSCP analysis using overlapping primers spanning the whole coding region of the EPO‐R gene. Structural integrity of the EPO‐R was further confirmed by sequencing of cloned PCR products. These data suggest that the mechanisms for the development of PV do not involve structural changes of the EPO‐R gene.
Cys209 Ser mutation in the platelet membrane glycoprotein ibα gene is associated with Bernard‐Soulier syndrome
Authors: Simsek S. , Noris P. , Lozano M. , Pico M. , von dem Borne A. E. G. Kr. , Ribera A. , Gallardo D.
Published: March 2008

Summary Molecular genetic analysis has been performed on a patient with Bernard‐Soulier syndrome (BSS). The patient had characteristically giant platelets and was deficient in the glycoprotein (GP) Ib/IX/V complex, the von Wllebrand factor (vWf) receptor on platelets. Previous studies with monoclonal antibodies directed against GP Ibα (CD 42b) and GP IX (CD 42a) demonstrated the absence of GP Ibα and presence of small amounts of GP IX on the surface of the patient's platelets. In ths study the presence of GP V (CD 42d) is also demonstrated. This indicates a defect in the α‐subunit of glycoprotein Ib. Therefore polymerase chain reaction (PCR)‐amplification Ib. Therefore polymerase chan reaction (PCR)‐amplification of the genomic DNA coding for GP Ibα was performed. Nucleotide sequence analysis of the entire coding region of GP Ibα revealed a homozygous single base pair mutation T A, leading to a single amino acid substitution cysteine serine at position 209 of the mature protein. We took advantage of the Mse I target site in the mutant allele, created by the T A mutation, to analyse all available family members. PCR‐ASRA (allele‐specific restriction enzyme analysis) using the restriction enzyme Mse I, revealed the heterozygosity of the mother and the two children of the patient, whereas homozygosity of the patient for the Cys209Ser mutation was confirmed. The sister of the patient was not found to the a carrier of the mutant allele.
Anti‐CD36 antibodies in patients with lupus anticoagulant and thrombotic complications
Authors: Rock G. , Chauhan K. , Jamieson G. A. , Tandon Narendra N.
Published: March 2008

Summary . Six patients with lupus anticoagulant with thrombotic complications, but not exhibiting systemic lupus erythematosus, demonstrated the presence in their plasma of antibodies directed against platelet antigens which were not detectable in two patients presenting with lupus anticoagulant but without thrombotic complications. Protein blotting of separated normal platelet proteins against patient plasma gave up to 18 bands of varying intensity indicative of multiple antiplatelet antibodies; one of these antibodies recognized a component with a mobility identical with CD36 (GPIV; m.w. 88,000) in 4/6 cases. Antibodies to CD36 and one or two other components were identified in 5/6 cases by immunoprecipitation from 125I‐labelled control platelets and 6/6 by dot blots against purified CD36. These results suggest that antiplatelet antibodies and, specifically, anti CD36 antibodies, occur frequently in the plasma of patients presenting with lupus anticoagulant and thrombotic complications.
Novel small deletions of the p53 gene in late‐stage B‐cell chronic lymphocytic leukaemia
Authors: Gandini D. , Aguiari G. L. , Cuneo A. , Piva R. , Castoldi G. L. , Sennoj Del
Published: March 2008

Summary . A group of 20 CLL patients selected for advanced clinical stage p53 mutations were analysed by single‐strand conformational polymorphism (SSCP) following PCR amplification of exons 5–9. In two patients abnormal SSCP of either exon 5 or exon 8 was found and PCR products were analysed by direct sequencing. A hemizygous or homozygous 12bp deletion at codon 135 and 3bp heterozygous deletion at codon 264 were detected; also, in the latter sample a heterozygous mutation at codon 282 (Arg to Gln) was found. To our knowledge, this is the first report of p53 deletions in B‐CLL. The two patients were elderly, and both had a rapidly progressive disease in the absence of unfavourable cytogenic abnormalities. These findings support a role for p53 alterations in the clinical course of some B‐CLL patients.
Erythropoietin response to anaemia is not altered by surgery or recombinant human erythropoietin therapy
Authors: Goodnough Lawrence T. , Price Thomas H. , Parvin Curtis A. , Friedman Kenneth D. , Vogler William R. , Khan Nurjehan , Sacher Ronald , Johnston Marilyn , Wissel Mary , Ciavarella David
Published: March 2008

Summary. Recombinant human erythropoietin (EPO) therapy has been shown to increase red blood cell (RBC) production and facilitate autologous blood donation before elective surgery. However, recent reports have suggested that surgery and/or EPO therapy may suppress endogenous erythropoietin secretion in response to anaemia. We therefore analysed the haemoglobin/erythropoietin relationship preoperatively and postoperatively in 71 autologous blood donors subjected to aggressive phlebotomy and six treatments with either EPO (150U/kg, n=16, 300U/kg, n=18, or 600 U/kg, n=19) or placebo (n=18). Using data from the three prepoerative study visit, the linear relationship between log erythropoietin and haemoglobin was determined for each of the 18 placebo patients. We found no significant differences in the slopes of the relationships in this group during aggressive phlebotomy. Furthermore, there was no evidence of a significant difference in the erythropoietin level recorded postoperatively for each patient to that predicted from the patient's postoperative haemoglobin level, based on the haemoglobin/log erythropoietin relationship preoperatively. Similarly, for each of the EPO‐treated groups, there was no evidence of a significant difference when comparing the recorded erythropoietin level to that predicted from each patient's postoperative haemoglobin level, based on the haemoglobin/log erythropoietin relationship preoperatively. We conclude that preoperative recombinant human erythropoietin therapy and/or surgery do not adversely affect the postoperative erythropoietin response to anaemia.
The chronic myeloid leukaemias: guidelines for distinguishing chronic granulocytic, atypical chronic myeloid, and chronic myelomonocytic leukaemia: Proposals by the French ‐ American ‐ British Cooperative Leukaemia Group
Authors: Bennett J. M. , Catovsky D. , Daniel M. T. , Flandrin G. , Galton D. A. G. , Gralnick H. , Sultan C. , Cox C.
Published: March 2008

Summary. We have reviewed our experience with four of the entities that are included under the generic term chronic myeloid leukaemia (CML), namely the classic Ph+ CGL, both BCR+ and BCR−, aCML and CMML. We have developed a statstical model that confirms that CGL, aCML and CMML can be distinguished from each other with reasonable success employing five quantitative parameters (WBC, percentage immature granulocytes, percentage monocytes, percentage basophils, percentage erythroid precursors in bone marrow) and one qualitative parameter (granulocytic dysplasia). It is hoped that these detailed recommendations will enable investigators to improve their diagnostic accuracy. This should permit more uniform comparisons of molecular biologic and clinical studies.
Peripheral blood stem cells (PBSCs) collected after recombinant granulocyte colony stimulating factor (rhG‐CSF): an analysis of factors correlating with the tempo of engraftment after transplantation
Authors: Bensinger W. I. , Longin K. , Appelbaum F. , Rowley S. , Weaver C. , Lilleby K. , Gooley T. , Lynch M. , Higano T. , Klarnet J. , Chauncey T. , Storb R. , Buckner C. D.
Published: March 2008

Factors affecting mobilization and engraftment were analysed in 54 patients undergoing transplant using autologous PBSCs mobilized with high‐dose recombinant granulocyte stimulating factor (rhG‐CSF). Patients received 5‐7 d of rhG‐CSF. 16 μg/kg/d, administered subcutaneously. PBSCs were harvested by leukapheresis using automated continuous‐flow blood cell separators beginning on day 4 of rhG‐CSF, processing 10 litres of whole blood, for 2‐6 consecutive days. Transplants were performed for the following diseases: breast cancer (n = 22), non‐Hodgkin's lymphoma (n = 18), multiple myeloma (n = 7) and other (n = 7). Engraftment was rapid with patients reaching a neutrophil count of 1 × 109/1a median of 12 d (range 9‐22) after transplant. Platelets > 20 × 109/1 independent of transfusion support were achieved a median of day 10 (range 7‐60) after infusion. Multiple factors potentially influencing engraftment were examined using a Cox regression model. The number of CD34+ cells per kg was highly correlated with the time to achievement of granulocyte and platelet recovery (P < 0.012, 0.0001). The use of a post‐infusion growth factor and a radiation preparative regimen was important for neutrophil recovery, and a diagnosis of breast cancer was important for platelet recovery. In an analysis by linear regression of the logarithm of CD34+ cells collected, lower age, marrow without disease, no prior radiation, and lower number of prior chemotherapy regimens, were important factors influencing larger numbers of CD34+ cells in collections.
Dose of desferrioxamine and evolution of HIV‐1 infection in thalassaemic patients
Authors: Costagliola Dominique G. , Montalembert Mariane de , Lefrère Jean‐Jacques , Briand Catherine , Rebulla Paolo , Baruchel Sylvain , Dessi Carlo , Fondu Pierre , Karagiorga Markissia , Perrimond Henri , Girot Robert
Published: March 2008

To study the relationship between the dose of desferrioxamine (DFX) and the progression of the HIV‐1 disease in thalassaemia major patients (TMP), 64 seropositive TMP patients were studied. Cumulative incidence of CDC stage IV was calculated using a non‐parametric life‐table method. The association with the mean daily dose of DFX was tested with a Cox proportional hazards model which was also used to adjust for confounding variables. The median of the mean daily dose of DFX over the seropositive period was 40mg/kg (range 0‐65mg/kg). Age at seroconversion (P < 0.02) and splenectomy (P < 0.03) were found to be associated with the mean daily dose of DFX. 6.5 years after seroconversion, 11% of those who had been prescribed more than 40mg/kg of DFX daily had entered stage IV versus 35% of those who had been prescribed a lower dose (P < 0.01). When the dose was taken as a continuous variable it was found that the rate of progression was significantly smaller in TMP receiving a higher dose (P < 0.002). even after adjusting for age and splenectomy (P < 0.02). Although it should be noted that these results were obtained in an observational study, possibly biased by a non‐random allocation of the DFX dose, we believe that they are striking enough to support the claim that the role of DFX in the progression of HIV disease should be further evaluated.
Restoration of normal polyclonal haemopoiesis in patients with chronic myeloid leukaemia autografted with Ph‐negative peripheral stem cells
Authors: Bergamashi Gaetano , Podesta Marina , Frassoni Francesco , Rosti Vittorio , Carella Angelo Michele , Saglio Giuseppe , Cazzola Mario
Published: March 2008

Only a minority of patients with chronic myeloid leukaemia (CML) benefit from allogeneic bone marrow transplantation (BMT), a potentially curative therapy, or from treatment with interferon alpha, which prolongs survival in cytogenetic responders. In Genoa a programme has been initiated in which CML patients are autografted with Ph‐negative peripheral stem cells. To assess the pattern of marrow reconstitution, we studied the clonality of haemopoiesis in five females who engrafted and were Philadelphia chromosome negative. This was performed by evaluating the methylation patterns of the X‐linked hypervariable DXS255 locus with the probe M27β. All four analysable women showed polyclonal methylation patterns in both granulocytes and T lymphocytes, suggesting that marrow reconstitution occurred from normal residual stem cells.
Generation of anti‐tumour activity by OKT3‐stimulation in multiple myeloma: in vitro inhibition of autologous haemopoiesis
Authors: Attisano Carmela , Bianchi Alberto , Montacchini Laura , Carlesso Nadia , Peola Silvia , Bruno Benedetto , Roux Viviana , Ferrero Dario , Gallo Eugenio , Boccadoro Mario , Pileri Alessandro , Massaia Massimo
Published: March 2008

Summary. T cells in multiple myeloma (MM) patients are highly susceptible to activation with the anti‐CD3 monoclonal antibody (mAb) OKT3. When short‐term OKT3 stimulation is carried out on bone marrow mononuclear cells (BMMC), large numbers of CD3+CD25+HLA‐DR+ cells are rapidly generated and autologous malignant plasma cells are killed. OKT3 may thus be exploited in autologous bone marrow transplantation (ABMT) to purge residual plasma cells and simultaneously activate T cells to induce graftversus‐leukaemia‐like (GVL‐like) activity upon reinfusion. However, the possible impact of ex‐vivo short‐term OKT3 stimulation on haematological recovery is unknown. The aim of this work was to investigate the effect of OKT3 stimulation in vitro on autologous haemopoietic progenitor cells (HPC) of MM patients. Colony formation by granulocyte‐macrophage progenitor cells (granulocyte‐macrophage colony‐forming units, CFU‐GM) was highly suppressed, although supernatants of OKT3‐activated T cells contained up to 2500 pg/ml of granulocyte‐macrophage colonystimulating factor (GM‐CSF). T cell depletion completely prevented this suppression. Neutralizing antibodies against TNF‐α, TNF‐β and IFN‐γ (which are also produced by OKT3‐activated MM T cells) did not prevent it, and Transwell cultures showed that cell‐to‐cell contact was the main mechanism involved. OKT3‐activated T cells also suppressed erythroid burst‐forming units (BFU‐E) and CFU‐GM generation from HPC responsible for long‐term maintenance of in vitro myelopoiesis.

When tested on normal allogeneic BM, MM supernatants of OKT3‐stimulated BMMC partially suppressed the generation of day 7 CFU‐GM, but had no effect on day 14 CFU‐GM.

These data indicate that short‐term stimulation of BMMC with OKT3 can be used to generate anti‐tumour effector T cells for autologous adoptive immunotherapy. It is not a feasable approach for ex‐vivo purging and activation procedures in ABMT because of its potent inhibition of autologous haemopoiesis.
P‐glycoprotein expression on acute myeloid leukaemia blast cells at diagnosis predicts response to chemotherapy and survival
Authors: Wood P. , Burgess R. , MacGregor A. , Yin J. A. Liu
Published: March 2008

P‐glycoprotein (Pgp) expression, which is associated with the multi‐drug resistance (MDR) phenotype, has been reported to be a useful predictor of treatment outcome in acute leukaemia. We have examined the expression of Pgp on acute myeloid leukaemia (AML) cells in 54 newly diagnosed patients, using a novel streptavidin‐biotin complex (ABC) technique, 56% of patients at diagnosis were positive for Pgp with JSB‐1, a monoclonal antibody that binds to an internal epitope of Pgp. All patients received intensive induction chemotherapy. Post‐remission treatment consisted of futher chemotherapy ± bone marrow transplantation. Complete remission (CR) rates were significantly lower in the Pgp positive group than in the Pgp negative group (60%v 92%; P= 0.02). The overall survival for Pgp‐positive patients was significantly shorter (329 v 534d, P= 0.004), disease‐free survival was also reduced but the difference was not statistically significant (median 277 v 522d, P= 0.16). In this study CD34 expression was not predictive of response to chemotherapy nor was it associated with Pgp expression. Our results confirm the prognostic value of Pgp expression in AML at diagnosis and we suggest that Pgp could be a useful therapeutic target for reversing multi‐drug resistance. Furthermore, our simple and sensitive method of detecting Pgp should enable widespread testing to be performed.
Peripheral T cell receptors αβ and γδ in patients with Hodgkin's lymphoma
Authors: Akan Hamdi , Beksaçl Meral , Aydoǧgdu Ismet , Koçl Haluk , Ilhan Osman , ÖZcan Muhit
Published: March 2008

Antigen recognition by T cells is determined by an antigen specific T cell receptor (TCR). Two heterodimeric TCR structures associated with CD3 have been defined: TCR αβ and TCR γδ. TCR αβ and its function are well described but the role of TCR γδ in normal and lymphoproliferative disorders is not well established. In newly diagnosed or relapsed/refractory Hodgkin's disease (HD), a disease associated with defective T cell functions and increased sIL‐2R, We determined levels of seven TCR αβ variable regions [βV5(a), βV5(b), βV6(a), βV12(a), αβV(a), αV2(a)] and TCR γδ by using monoclonal antibodies (MCA). TCR γδ levels did not show any difference, but several variable regions of the TCR αβ differed when groups are compared with each other and the control group.
A case of severe neonatal thrombocytopenia with schizencephaly associated with anti‐HPA‐1 b and anti‐HPA‐2a
Authors: Kuijpers Robert W. A. M. , Anker John N. van den , Baerts Wim , Borne Albert E. G. Kr. von dem
Published: March 2008

We report a family with a neonate who was severely damaged by intracranial haemorrhages. These probably occurred before the 20th week of gestation. The neonate had a moderate thrombocytopenia. In the maternal serum anti‐HPA‐1b and anti‐HPA‐2a alloantibodies were detected. Third‐generation assays were applied to identify the alloantibodies. No other cause for the bleeding was found. Probably the combination of anti‐HPA‐1b and anti‐HPA‐2a alloantibodies, directed against the platelet fibrinogen receptor and the von Willebrand receptor, respectively, induced a thrombocytopenia and a thrombocytopathy.
The painful crisis of homozygous sickle cell disease: clinical features
Authors: Serjeant Graham R. , Ceulaer Charles D. E. , Lethbridge Rosemary , Morris Joanne , And Atul Singhal , Thomas Peter W.
Published: March 2008

The details of onset, perceived precipitating factors, associated symptoms, and pain distribution in the painful crisis of homozygous sickle cell (SS) disease have been prospectively recorded in 183 painful crises in 118 patients admitted to a day‐care centre in Kingston, Jamaica. Painful crises developed most frequently between 3 p.m. and midnight, most commonly affected patients aged 15‐29 years, affected the sexes equally, and were not obviously influenced by menstrual cycle. Of the perceived precipitating factors, skin cooling occurred in 34%, emotional stress in 10%, physical exertion in 7%, and pregnancy in 5% of women of child‐bearing age. Cold as a precipitant was not less common in patients with more subcutaneous fat. Pain affected the lumbar spine in 49%, abdomen in 32%, femoral shaft in 30%, and knees in 21%. There was a highly significant excess of bilateral involvement in limb and rib pain. Recurrent painful crises occurred in 40 patients but showed no evidence of involving similar sites on successive occasions. Abdominal painful crises were associated with abdominal distention in 18 (31%) and with referred rib pain in a further 15 (26%) of crises. Fever was common even in apparently uncomplicated painful crises, suggesting that fever is characteristic of the painful crisis itself and not necessarily indicative of infection. Following investigation and treatment in a day‐care centre, over 90% of patients returned home.
rh‐Erythropoietin stimulates immature reticulocyte release in man
Authors: Major Attila , Bauer Christian , Breymann Christian , Huch Albert , Huch Renate
Published: March 2008

The pharmacodynamics of single intravenous dosing with recombinant human erthropoietin (rhEPO) was investigated in eight healthy volunteers (150U/kg, n = 2; 300 U/kg, n = 6) with respect to reticulocyte subdivisions (by fluorescence flow cytometry) and serum ferritin over 6.5 d. The present study shows that bolus rhEPO injection produces an immediate release of high and middle fluorescence (immature) reticulocytes with a high RNA content from the marrow into the circulation, whereas the low fluorescence (more mature) reticulocytes were at first not affected. Serum ferritin decreased markedly within 24 h, reaching a nadir 50% of baseline after 120h (5 d), with no increase in haemoglobin. Our data suggests that rhEPO triggers premature expulsion of immature reticulocytes from the bone marrow into the circulation independent of its effect in stimulating erythropoiesis and that rhEPO has an effect on serum ferritin concentration which in this dynamic situation is dependent not only on the iron stores.
Detection and significance of bone marrow infiltration at the time of autologous bone marrow transplantation in Hodgkin's disease
Authors: Chopra R. , Wotherspoon A. C. , Blair S. , McMillan A. K. , Mills W. , Fielding A. K. , Linch D. C. , Goldstone A. H.
Published: March 2008

Bilateral iliac crest biopsies were performed at the time of bone marrow (BM) harvest in patients with poor‐risk Hodgkin's disease (HD) undergoing autologous bone marrow transplantation (ABMT). 67 consecutive patients with normal trephine biopsies (58 unilateral; nine bilateral) taken 2 weeks prior to harvest were studied. Nine (13%) of patients had BM infiltration with HD on the harvest biopsies. These patients had all previously had only unilateral biopsies. Three patients did not proceed to ABMT because disease progression became clinically apparent, but the remaining six patients were infused with involved marrow. One patient died 6 weeks post ABMT, with no evidence of disease at post‐mortem. One patient showed no response to ABMT and four patients either had a partial or complete response to ABMT. Of the responders, three patients are now in complete remission at 53, 39 and 33 months past ABMT.
Serum thymidine kinase in congenital dyserythropoietic anaemia type III
Authors: Sandströum Herbert , Wahlin Anders , Eriksson Mikael , Bergström Ingmar
Published: March 2008

Serum thymidine kinase (TK) was determined in a family with congenital dyserythropoietic anaemia type III (CDA, type III). 20 patients and 10 of their healthy siblings were investigated. Elevated TK was found in all 20 patients (median 56.2 U) but their healthy siblings had normal values (median 2.65 U). We suggest that determination of TK should be used for discrimination between healthy siblings and individuals affected by CDA type III when bone marrow examination is not suitable.
A function of CD10 on bone marrow stroma
Authors: Delikat Sylvie E. , Galvani David W. , Zuzel Mirko
Published: March 2008

Summary. Bone marrow (BM) stromal cells express CD10 (cALLA), a surface antigen now known to be a neutral endopeptidase (NEP‐24.11). The function of CD10 in BM stroma is unknown, although purified NEP‐24.11 is known to degrade different substrates including interleukin 1β (IL‐1β).

We have therefore employed a CD10‐positive BM stromal cell line (L2AK) which proliferates in response to IL‐1β to test the hypothesis that degradation of this cytokine is one of the functions of stromal CD10.

We first showed that [3H]thymidine incorporation by L2AK cells is enhanced by IL‐1β in a clear dose‐dependent manner. Addition of the CD10 inhibitor, phosphoramidon, together with IL‐1β resulted in a left shift in the dose‐response curve which corresponded to a 10‐fold potentiation of the IL‐1β effect.

These results indicate that CD10 on bone marrow stromal cells can degrade IL‐1β and therefore provide a local control of the effects of this, and possibly other, growth factor(s).
A patient with multiple myeloma and respiratory insufficiency due to accumulation of paraprotein in the alveolar space
Authors: Meijer W. G. , Kooy M. Van Marwijk , Ladde B. E.
Published: March 2008

We describe a patient with multiple myeloma and respiratory insufficiency. Autopsy revealsed acumulation of igG‐kappa paraprotein in the alveolar space. The clinical and pathological presentation are typical for secondary pulmonary alveolar proteinosis. Attention is focussed on the possibility of secondary pulmonary alveolar proteinosis as cause for dyspnoea in patients with haematological malignancies.
A new Turkish type of β‐thalassaemia major with homozygosity for two non‐consecutive 7.6 kb deletions of the Ψβ and β genes and an intact δ gene
Authors: ÖNer C. , ÖNer R. , Guurgey A. , Altay Ç.
Published: March 2008

Summary. In a 2.5‐month‐old infant with β‐thalassaemia major, DNA analysis of the gamma‐beta region revealed homozygosity for two large deletions removing the entire β and β regions including their 5’ promoter regions but leaving the delta gene intact. The downstream deletion was predicted to be 7.6 kb in length extending from a point 1.5 kb on the 3’ side of the δ‐globin gene to about 1.8 kb on the 3’ side of the β‐globin gene. The upstream deletion, which was also about 7.6 kb, extended from a point 1.5 kb on the 5’ side of the β‐globin gene to about 4.5 kb on the 3’ of the β gene. The δ‐giobin gene was intact. From the phenotypic expression of the disease it is concluded that removal of the β gene probably prevents derepression of the γ gene that has previously been observed in the absence of the promoter region of the β gene and the switch mechanism from gamma to beta gene expression may take place earlier than expected.
Immunophenotype of c‐kit cells in normal human bone marrow: implications for the detection of minimal residual disease in AML
Authors: Macedo A. , Orfao A. , Martianez A. , Vidriales M. B. , Valverde B. , Loapez‐Berges M. C. , Miguel J. F. San
Published: March 2008

Summary. In the present study, seven normal human bone marrow samples from healthy volunteers have been analysed in order to investigate the immunophenotypic characteristics of the normal CD117+ cells and their utility for the detection of minimal residual disease in 71 acute myeloid leukaemia patients.

Our results show that most of normal BM CD117+ cells coexpress the HLADR and the myeloid associated CD33 antigen. In addition, almost half of CD117+ cells are CD34+, these cells displaying a different FSC/SSC distribution when compared to the CD117+/CD34− cells. No CD117+/CD15+ and CD117+/CD10+ cells were detected and very few CD117+ cells (<1 × 10−3) expressing the HLADR−/CD34−, CD33+/HLADR− and CD34+/HLADR− phenotypes were found to be present in normal BM. In contrast, from the 71 AML patients analysed, 34 had CD117+/CD15+ blast cells and eight had the CD117+ phenotypes detected at low frequencies (<1 × 10−3) in normal BM.

In summary, the present study shows that the use of the CD117 antigen in different monoclonal antibodies combinations may be of great help for the detection of minimal residual disease in a high proportion of AML cases, especially in those patients displaying the CD117+/CD15+ phenotype, because cells coexpressing both antigens in normal BM, if present, are at very low frequencies.
Prognostic value of S‐phase cells in AML patients
Authors: Vidriales M. B. , Orfao A. , López‐Berces M. C. , Gonzaalez M. , Loapez‐Macedo A. , Ciudad J. , Loapez A. , Garciaa M. A. , Hernández J. , Borrego D. , Miguel J. F. San
Published: March 2008

Summary. Until now, reports on the cell cycle distribution of AML patients have shown highly variable results which are probably related to the technical heterogeneity of such studies. The aim of the present paper was to analyse in 204 AML patients at diagnosis the proliferative rate (assessed by the number of S‐phase cells) of peripheral blood (PB) (126 cases) and bone marrow (BM) (78 cases) cells in order to explore its relationship with disease characteristics. A strong parallelism was observed regarding the relationship between the proliferative rate of the blast cells both in PB and in BM and the disease characteristics. Cases with a high proliferative rate were associated with advanced age, high WBC counts and LDH serum levels, monocytic leukaemias (assessed both by morphological and immunophenotypic criteria) and NK‐associated antigens (CD56, CD16). The proliferative activity did not influence the CR rate; in contrast, cases with a high number of S‐phase cells had shorter survival. In addition, multivariate analysis showed that age, the expression of CD11b, and the S‐phase counts are the only three prognostic factors displaying an independent impact on survival.
Gender and treatment outcome in childhood lymphoblastic leukaemia: report from the MRC UKALL trials*
Authors: Chessells J. M. , Richards S. M. , Bailey C. C. , Lilleyman J. S. , Eden O. B.
Published: March 2008

Summary. We have examined the factors influencing prognosis in over 4000 children with acute lymphoblastic leukaemia (ALL) aged 1–14 who have been treated on consecutive MRC UKALL trials from 1972 to 1990. During this time the results of treatment have improved steadily but are consistently superior in girls when compared with boys; the 5‐year event‐free survival in girls improving from 51% to 71% and in boys from 31% to 57%. These results were independent of age and presenting leucocyte count. Boys not only had a testicular relapse rate of 10% but an excess of bone marrow relapse, particularly evident after 2 years from diagnosis. Other prognostic factors included organomegaly and the morphology of leukaemic blast cells; immuno‐phenotype of the leukaemia, however, had no independent significance after allowance for age, sex and leucocyte count.

The influence of sex on prognosis was reaffirmed when we examined various methods of identifying children at highest risk of treatment failure for whom alternative therapy such as bone marrow transplantation might be justified. In MRC UKALL X children had been deemed ‘high risk’ on the basis of leucocyte count alone, but with further follow‐up it has become apparent that girls with an initial leucocyte count of > 100 × 109/1 have a similar prognosis to boys with a lower count. We therefore derived a risk score based on sex, age and count which has given better discrimination between standard risk (66% 5‐year survival) and poor risk (39%) survival than other methods. This group of worse‐risk children includes 16% of boys but only 3% of all girls.

Gender remains an important prognostic factor in UKALL trials and there are very few girls who are at highest risk of treatment failure. The reasons for this remain unclear, but the pattern of relapses suggests that boys more often get inadequate systemic therapy. We postulate that the reasons for treatment failure may relate to sensitivity to continuing (maintenance) chemotherapy.
A three‐base deletion removing a leucine residue in a leucine‐rich repeat of platelet glycoprotein Ibα associated with a variant of Bernard‐Soulier syndrome (Nancy I)
Authors: de la Salle Corinne , Baas Marie‐Jeanne , Lanza Franccois , Schwartz Agnegs , Hanau Daniel , Chevalier Jerotme , Gachet Christian , Briquel Marie‐Elisabeth , Cazenave Jean‐Pierre
Published: March 2008

Summary. Leucine‐rich repeats are conserved structural motifs present in the four components of the human platelet glycoprotein Ib/IX/V complex receptor for the adhesive protein von Willebrand factor. The absence or abnormality of this complex is responsible for Bernard‐Soulier disease, an autosomal recessive bleeding disorder. We report a deletion of leucine 179, located in a highly conserved position of the seventh leucine‐rich repeat of GPIbα, found in a variant form of Bernard‐Soulier disease (Bernard‐Soulier Nancy I). Three affected siblings of a family were characterized by absence of ristocetin‐induced platelet agglutination, although ADP aggregation was normal. Flow cytometry studies showed detectable amounts of all four members of the GPIb/IX/V complex on the surface of the patients’ platelets. Western blotting revealed normal levels of GPIX, decreased levels of GPIbβ and GPV, and <1% of GPIbα. RT‐PCR studies showed the presence of mRNA coding for GPIbα. GPIbβ, GPIX and GPV. Sequencing showed a three‐base deletion which results in the absence of a leucine residue, highly conserved across the seven leucine‐rich repeats of GPIbα and also within the other members of the leucine‐rich glycoprotein family. The absence of the leucine 179 in a patient's GPIbα is believed to cause a conformational change in the protein which would account for the lack of binding of most of the MoAbs tested and would be responsible for the absence of von Willebrand factor binding. These results point to the leucine‐rich region of GPIbα as being required for the correct exposure of the von Willebrand binding site as well as for the correct assembly and stability of the GPIb IX V complex on the platelet surface.
Heteroduplex screening for molecular defects in factor IX genes from haemophilia B families
Authors: Chen Shi‐Han , Schoof Jon M. , Weinmann Ann F. , Thompson Arthur R.
Published: March 2008

Summary. Heteroduplex screening of amplified fragments containing sequences of all known small haemophilic mutations in the factor IX gene localized mutations in 18 new families: 12 were at common recurrent sites; three were novel. Carriers and/or patients from each of 41 families with mutation in 7 exons and 5’ and 3’ non‐coding regions were positive.
IL‐3 increases marrow and peripheral erythroid precursors in chronic pure red cell aplasia presenting in childhood
Authors: Bastion Yves , Campos Lydia , Roubi Nora , Bienvenu Jacques , Felman Pascale , Dumontet Charles , Coiffier Bertrand
Published: March 2008

Summary. In most patients with Diamond‐Blackfan anaemia (DBA) the addition of IL‐3 to in vitro cultures increases the number of erythroid progenitors. However, a small proportion of patients show an erythroid response to in vivo administration of IL‐3. We treated two adult patients with corticosteroid‐refractory DBA, who had shown a clear in vitro erythroid response to IL‐3, with increasing doses of IL‐3 (from 2.5 to 10μg/kg/d) by subcutaneous injections for 7 and 9 weeks. Monitoring of marrow and circulating progenitors showed a sustained elevation of BFU‐E and CFU‐E as well as an increase of other progenitors and CD34+ cells during therapy. However, this effect did not translate into a clear clinical response, although transfusion requirements decreased transiently in one patient. This report shows that a sustained elevation of erythroid progenitors induced by in vivo IL‐3 administration may not translate into an increase of mature red cells production in DBA patients.
Lack of G blood group antigen in DIIIb erythrocytes is associated with segmental DNA exchange between RH genes
Authors: Rouillac Christelle , Kim Caroline Le Van , Blancher Antoine , Roubinet Francis , Cartron Jean Pierre , Colin Yves
Published: March 2008

Summary. The Rh blood group antigens D, Cc and Ee are encoded by two highly related genes, RHD and RHCE. Almost all red cells which carry D and all cells which carry C also express the G (Rh12) antigen. In this report we have determined the molecular basis of the DIIIb category phenotype which represents a very rare condition characterized by the presence of most of the D epitopes and the total absence of the antigen G. mRNA sequencing and Southern blot analysis of two unrelated samples indicated that the DIIIb category phenotype is associated with a segmental DNA exchange between exon 2 of the RHD and RHCE genes resulting in three D c amino acid substitutions (Ile60Leu, Ser68Asn Ser103Pro).
Modulation of cell kinetics and cell cycle status by treating CD34+ chronic myeloid leukaemia cells with p53 antisense phosphorothioate oligonucleotides
Authors: LANZA FRANCESCO , BI SUCAI , MORETTI SABRINA , CASTOLDI GIANLUIGI , GOLDMAN JOHN M.
Published: March 2008

Mutations of the p53 tumour suppressor gene occur in 20% of chronic myeloid leukaemia (CML) patients in blastic crisis, but it is still uncertain whether this inactivation plays a role in the pathogenesis of blastic transformation or in maintaining the leukaemic proliferation in CML, as it does in several solid tumours. We have previously shown that more than 50% of both normal and CML CD34+ cells express the p53 protein. However, haemopoietic cells at different phases of the cell cycle express p53 with different conformations, suggesting that the function of p53 may be closely regulated during the cell cycle. In order to elucidate the mechanism by which p53 suppresses cell proliferation, we evaluated the effects of inhibiting p53 expression on cell cycle and cell kinetics of chronic phase CML (n = 12) and normal (n = 7) bone marrow light‐density cells and purified CD34+ progenitors by using an 18‐mer modified antisense oligonucleotide which targets the region covering the six base pairs immediately before the first codon and the first four coding codons of p53, We found that the number of cells positive for the cell cycle‐specific nuclear antigen Ki67 and for the BrdU monoclonal antibody (McAb) was significantly increased after p53 antisense oligonucleotide treatment. At the same time, p53 protein expression was completely abrogated in both light‐density and CD34+ cells. In addition, DNA analysis by flow cytometry demonstrated that the number of cells in quiescent phases of the cell cycle (G0‐G1) was significantly decreased after exposure of light‐density cells to p53 antisense oligomers, whereas the number of cells in S or G2‐M phases was increased. Furthermore, the longer the incubation time the higher the increase in cell proliferation. Treatment of CML cells with p53 antisense oligomers also resulted in significantly increased numbers of CFU‐GM colonies. Our data suggest that p53 is a negative regulator of cell proliferation and its action is mediated through changes in cell cycle kinetics, mainly before the S phase. We can further speculate that the loss of p53 function, at the time of blastic crisis of CML, may play a role, in combination with other genetic changes (p210 BCR/ABL, Rb gene abnormality, others to be defined), in inducing disturbances in cell proliferation, differentiation, and apoptosis.
Scintigraphic evaluation of the haemopoietic bone marrow using a 99mTc‐anti‐granulocyte antibody: a validation study with 52Fe
Authors: JAMAR FRANCOIS , FIELD CATHERINE , LENERS NORBERT , FERRANT AUGUSTIN
Published: March 2008

To specify the validity of bone marrow scanning using a monoclonal anti‐granulocyte antibody labelled with 99mTc (BW 250/183) for the functional assessment of haemopoiesis, we compared this method with 52Fe scan in 16 patients with haematological disorders. The examinations were performed using a rectilinear whole‐body scanner and the distribution of the two tracers was assessed visually and quantitatively in anatomical bone marrow segments, the spleen and liver. Qualitative comparison showed concordance in the bone marrow distribution of the two tracers in 83% of the segments. Discrepancies were found in six patients with hypoplastic or aplastic marrow. The spleen was visualized in all cases with the 99mTc‐Moab, including nine patients without splenic haemopoiesis (i.e. without spleen uptake of 52Fe). The uptake of the two tracers, quantified in bone marrow segments and the spleen, correlated well (PO‐0001), but not in the liver (NS). The correlation between the uptake values for each patient was excellent, except in cases of aplastic bone marrow. In conclusion, bone marrow scanning using a 99mTc labelled anti‐granulocyte monoclonal antibody enables functional evaluation of the distribution of haemopoiesis. Limitations include the evaluation of bone marrow aplasia and identification of splenic haemopoiesis, for which 52Fe remains the tracer of choice.
The effect of continuous G‐CSF application in human cyclic neutropenia: a model analysis
Authors: SCHMITZ STEPHAN , FRANKE HORST , WICHMANN H. ERICH , DIEHL VOLKER
Published: March 2008

Human cyclic neutropenia (CN) is a rare haematological disorder characterized by oscillations of blood neutrophils at subnormal levels with a stable period of approximately 21 d. During the phase of severe neutropenia (neutrophils <250 cells/yul), which last 4–10 d, the patients are endangered by serious infections. Several authors report that continuous G‐CSF application can elevate the blood neutrophils to such a level that the risk of infections is significantly reduced. Although the characteristic cycles are not eliminated by G‐CSF, the period of the oscillations is shortened to 12–14 d. Based on a previously proposed computer‐simulation model of human CN, the effects of continuous G‐CSF application on CN are studied. It is shown how the known different cell‐kinetic effects of G‐CSF on granulopoiesis explain the clinical data in CN. The reduced length of the cycles emerges as a result of the transit time reduction of the post‐mitotic granulopoietic cells by G‐CSF. The measured increase of the neutrophil maxima is reproduced by the additional mitoses of the immature granulopoietic bone marrow cells induced by G‐CSF. The slight elevation of the neutrophil nadirs can be attributed to a weak effect of G‐CSF on the assumed underlying defect in CN (an abnormally small variance of the granulopoietic bone‐marrow transit time).
Haemoglobin Tunis‐Bizerte: a new a globin 129 Leu–> Pro unstable variant with thalassaemic phenotype
Authors: DARBELLAY R. , MACH‐PASCUAL S. , ROSE K. , GRAF J. , BERIS PH.
Published: March 2008

A Leu– Pro substitution at position 129 of the ci globin gene was detected in three members of a Tunisian family by sequencing the whole a2 and i DNA. The mutation was verified by dot‐blot allele‐specific hybridization as well as by digestion of PCR and RT‐PCR products with Nci I, since the aj129T_>c mutation creates an additional recognition site for the above‐mentioned enzyme. The Q,ii29(Hi2)Leupro substitution disturbs helix H resulting in a‐thal trait most probably because the unstable a‐globin chain variant cannot form a/3 dimers. A search for the abnormal Hb and for the abnormal a globin chain by isoelectric focusing, carboxymethyl cellulose chromatography and electrospray ionization mass spectrometry was negative. In the heterozygous state, the Qii29(Hi2) Leu‐Pro varjant is manifested by microcytosis (MCV73fl), whereas in the homozygous state there is moderate anaemia with marked microcytosis (Hb 11–6 g/dl, MCV 65 fl).
Prognostic relevance of serum thymidine kinase in primary myelodysplastic syndromes: relationship to development of acute myeloid leukaemia
Authors: MUSTO PELLEGRINO , BODENIZZA CARLO , FALCONE ANTONIETTA , D'ARENA GIOVANNI , SCALZULLI POTITO , PERLA GIANNI , MODONI SERGIO , PARLATORE LUIGI , VALVANO MARIA ROSA , CAROTENUTO MARIO
Published: March 2008

The aim of this study was to evaluate the possible prognostic relevance of thymidine kinase serum levels (s‐TK), an indirect marker of proliferative activity, in myelodysplastic syndromes (MDS). S‐TK levels were monitored by means of a radioenzyme assay in 90 patients affected by MDS (22 refractory anaemia, RA; 17 RA with ring sideroblasts, RARS; 21 RA with blast excess, RAEB; 15 RAEB in transformation, RAEB‐T; 15 chronic myelomono‐cytic leukaemia, CMMoL). Mean s‐TK levels (U//tl) measured at diagnosis were 11–9 –12–6 for RA, 11–4–13′6 for RARS, 19–9 – 28–4 for RAEB, 39–6 – 34–3 for RAEB‐T and 77–7 – 69–7 for CMMoL (normal values <5U//LI1). With the only exception of a weak relationship with lactate dehydrogenase, no correlation was found between initial s‐TK values and other clinical or laboratory parameters, such as age, haemoglobin, white blood cell or platelet count, percentage of bone marrow blasts. MDS patients with s‐TK >38 V/fA, a cut‐off level selected by means of ROC statistical analysis, showed a significantly shorter survival than those with s‐TK <38U//xl (8–2 v 37–4 months, respectively; P < 0–0001). In particular, transformation in acute myeloid leukaemia (AML) occurred in 17/21 (81%) of patients with s‐TK >38U//d and 9/69 (13%) of those with lower levels at diagnosis (P < 00001), independently of FAB subtype. High s‐TK levels were also useful to predict evolution in AML during the course of the disease in patients with normal initial values. Multivariate analysis confirmed the independent prognostic value of s‐TK on both overall survival and risk of acute transformation. We conclude that s‐TK may be an important prognostic factor in MDS, strongly correlated with development of AML.