bluuebunny/crossref_metadata_2025 huggingface dataset:
Prevalence of anti‐FVIII antibodies in severe haemophilia A patients with inversion of intron 22
Authors: Vianello Fabrizio , Radossi Paolo , Tison Tiziana , Dazzi Francesco , Tagariello Giuseppe , Giorgio Davoli Pier , Girolami Antonio
Published: November 2003

The aim of our study was to investigate whether haemophilia A patients with inversion of intron 22 are at high risk for non‐inhibitory anti‐FVIII antibodies development detected by ELISA. It is known that patients with severe forms of haemophilia A are more likely to develop anti‐FVIII antibodies. The incidence of inhibitory anti‐FVIII antibodies in patients with factor VIII gene inversion has been extensively evaluated, but if this defect has to be considered a predisposing factor is still debatable. Non‐inhibitory anti‐FVIII antibodies are attracting interest, due to the potential influence on FVIII half‐life. Our data show that FVIII gene inversion was a major predisposing factor for anti‐FVIII antibodies development.
Prognostic features of asymptomatic multiple myeloma
Authors: Weber Donna M. , Dimopoulos Meletios A. , Moulopoulos Lia A. , Delasalle Kay B. , Smith Terry , Alexanian Raymond
Published: November 2003

Approximately 20% of patients with multiple myeloma are recognized by chance without significant symptoms. In order to prevent morbidity with timely therapy, reliable criteria are needed that distinguish those likely to show early or late disease progression. Multiple clinical features were assessed in 101 consecutive, asymptomatic and previously untreated patients. Patients with one or more lytic bone lesions were excluded because this feature had been found previously to be associated with early progression. Multivariate analysis indicated that only serum myeloma globulin >30 g/l, IgA protein type, and Bence Jones protein excretion >50 mg/d remained as significant independent variables. The presence of two or more of these features signified high‐risk disease with early progression (median 17 months) whereas the absence of any adverse variable was associated with prolonged stability (median 95 months) (P <0.01. Magnetic resonance (MR) imaging of the spine was useful only in patients with one adverse feature and an intermediate time to progression (median 39 months). An abnormal pattern (40% of patients) helped to distinguish patients with an imminent complication from those with more stable disease. Because a serious complication (fracture, hypercalcaemia) occurred in 35% of patients with early disease progression, chemotherapy seems justified for selected patients with asymptomatic disease at diagnosis. The remaining patients were at such low risk for progression (median 6 years) that they may be followed safely at long intervals without treatment.
EPIDEMIOLOGY OF COAGULATION FACTORS, INHIBITORS AND ACTIVATION MARKERS: THE THIRD GLASGOW MONICA SURVEY II. RELATIONSHIPS TO CARDIOVASCULAR RISK FACTORS AND PREVALENT CARDIOVASCULAR DISEASE
Authors: Woodward M. , Lowe G. D. O. , Rumley A. , Tunstall‐Pedoe H. , Philippou H. , Lane D. A. , Morrison C. E.
Published: November 2003

Coagulation factor activity (fibrinogen, VII, VIII and IX), coagulation inhibitor activity (antithrombin, protein C, protein S), and coagulation activation markers (prothrombin fragment F1, 2; thrombin–antithrombin complexes) were measured in 746 men and 816 women aged 25–74 years, randomly sampled from the north Glasgow population in the Third MONICA Survey. After age‐adjustment, significant associations with cardiovascular risk factors were observed. Serum cholesterol and triglyceride were associated with increases in factors VII and IX, as well as antithrombin, protein C and protein S; and with increased fibrinogen and factor VIII in women. Apart from factor VIII (related to blood pressure in men, but not in women), similar associations were observed for blood pressure and body mass index. Smoking status and/or smoking markers were related to fibrinogen, factor IX, antithrombin and protein S. Alcohol intake was related to protein S, and inversely to fibrinogen and antithrombin in men. Low social class was associated with fibrinogen, factor VIII, factor IX, and with antithrombin, protein S, and low protein C in men. Serum vitamin C was associated inversely with coagulation factors and coagulation inhibitors. The only associations of activation markers were with low serum vitamin C, and with alcohol consumption and low social class in men. Prevalent cardiovascular disease was associated only with fibrinogen.

These associations of coagulation factors and inhibitors with cardiovascular risk factors are plausibly relevant to thrombotic risk in cardiovascular disease. In general, ‘worse’ values of risk factors are associated with increased plasma levels of both coagulation factors and inhibitors, without significant increase in coagulation activation markers. However, the association of lower serum vitamin C with increased coagulation activation markers is of potential therapeutic interest.
Is antenatal antibody screening worthwhile in Chinese?
Authors: Wong K. F. , Tse K. T. , Lee A. W. C. , Mak C. S. L. , So C. C.
Published: November 2003

A total of 1997 pregnant women were screened during their first antenatal visit for irregular antibodies for the prevention of haemolytic disease of the newborn. Patient sera were tested against a panel of group O screen cells including one with the expression of Miltenberger determinants GP.Mur. 17 women (0.85%) had irregular antibodies of which four were of potential clinical significance, including one with anti‐D, two with anti‐E and one with anti‐D, anti‐E and anti‐G. Although antenatal antibody screening is mandatory in Western populations, our results suggest that this may not be necessary in the Chinese population except for those who are Rh D‐negative or who have a history of haemolytic disease of the newborn.
Differential modulation of IL‐1‐induced endothelial adhesion molecules and transendothelial migration of granulocytes by G‐CSF
Authors: Eissner Günther , Lindner Heidrun , Reisbach Gilbert , Klauke Ines , Holler Ernst
Published: November 2003

Granulocyte colony stimulating factor (G‐CSF) is widely used for mobilization of haemopoietic stem cells into the peripheral blood. However, little is known about the mechanisms involved in mobilization and the immune modulatory effects of this growth factor. In this report we show that G‐CSF down‐regulated intercellular adhesion molecule 1 (ICAM‐1) induced by Interleukin‐1 (IL‐1) on human endothelial cells. Interestingly, the G‐CSF‐mediated down‐modulation of IL‐1‐induced ICAM‐1 appeared to be biphasic. In pharmacological concentrations (>300 ng/ml), and in dose ranges of plasma G‐CSF levels above that of non‐febrile healthy individuals (30 pg/ml), a significant decrease in surface ICAM‐1 could be observed. This could be explained, at least in part, by an increased autocrine G‐CSF production by endothelial cells in response to IL‐1 and exogenous G‐CSF. In contrast to ICAM‐1, IL‐1‐triggered VCAM‐1 expression was superinduced by G‐CSF with the optimal concentration of 30 pg/ml. To evaluate the functional significance of these findings, 51Cr adhesion assays with peripheral blood mononuclear cells (PBMC) or granulocytes known to lack the VCAM‐1 counter‐receptor very late antigen 4 (VLA‐4) and IL‐1‐stimulated endothelial cells, in the presence or absence of G‐CSF, were performed. G‐CSF could not inhibit the IL‐1‐induced adhesion of PBMC to endothelial cells, which may be due to the differential adhesion molecule modulation. In contrast, granulocyte adhesion induced by IL‐1 could effectively be blocked by co‐incubation with G‐CSF. Finally, G‐CSF also inhibited transendothelial migration of granulocytes through IL‐1‐activated endothelial cells in a concentration‐dependent manner.
Lack of clonal BCRA2 gene deletion on chromosome 13in chronic lymphocytic leukaemia
Authors: PANAYIOTIDIS PANOS , GANESHAGURU K. , ROWNTREE CLARE , JABBAR SHIREEN A. B. , HOFFBRAND VICTOR A. , FORONI LETIZIA
Published: November 2003

Chromosome 13q deletion is among the most common cytogenetic abnormalities in chronic lymphocytic leukaemia (CLL). We investigated the 13q14.3 deletion in 44 CLL patients by Southern blotting following purification of clonal B CLL cells to >90%. Two sets of probes were used to investigate the site of clonal deletion, the D13S25 and D13S319 markers (at 13q14.3) and probes for exons 11 and 26–27 of the BRCA2 gene (at 13q12). Homozygous and heterozygous deletion at the 13q14.3 region was found in five and 17 patients, respectively. Despite the recent report of the BRCA2 gene involvement in >80% of CLL patients, we failed to detect a single case of homozygous or heterozygous deletion involving the 13q12 region. Our data support previous findings that the 13q14.3, and not the 13q12 region, is the major site of candidate tumour suppressor gene(s) in CLL.
Antiphospholipid antibody: functional specificity for inhibition of prothrombin activation by the prothrombinase complex
Authors: Pierangeli Silvia S. , Goldsmith George H. , Branch D. Ware , Harris E. Nigel
Published: November 2003

The antiphospholipid syndrome (APS) is associated with production of autoantibodies with lupus anticoagulant (LA) activity. These antibodies cause prolongation of in vitro clotting tests by inhibition of the conversion of prothrombin to thrombin in the presence of anionic phospholipid (PL). The extent to which this inhibition reflects antibody binding to, or functional inhibition of, phospholipids alone, prothrombin alone, or a prothrombin–phospholipid complex is pertinent to our understanding of the pathophysiology of this syndrome. Immunoglobulin fractions (IgG) from 18 patients with LA activity were tested for inhibitory activity against prothrombin activation by either factor Xa, in a purified prothrombinase system, or by purified fractions of snake venoms (E. carinatus, E. multisquamatus) which cleave prothrombin at the same initial site as the prothrombinase complex but do not require anionic phospholipid as a cofactor. Parallel testing of the same IgG samples for prothrombin binding by immunoassay was performed. Although all IgG samples inhibited the prothrombinase reaction, only three exhibited any inhibition of venom protease prothrombin activation in either the presence or absence of PL. Only one sample exhibited prothrombin binding by Western blot. These results suggest that lupus anticoagulant antibodies are heterogenous and that many, if not most, of the autoantibody populations responsible for LA activity impair prothrombin activation by interaction either with phospholipid alone or with a restricted range of prothrombin–phospholipid epitopes expressed by prothrombin only as part of the intact prothrombin–prothrombinase complex.
Polymorphism of the tumour necrosis factor‐alpha and lymphotoxin‐alpha genes in hairy cell leukaemia
Authors: Demeter Judith , Porzsolt Franz , RÄmisch Stephanie , Schmid Mathias , Messer Gerald
Published: November 2003

Hairy cell leukaemia (HCL) is a rare chronic B lymphoproliferative disorder which can lead to severe pancytopenia and several immunologic abnormalities. The pathogenetic role of tumour necrosis factor (TNF)‐α in HCL prompted us to study the potential contribution of functionally important genetic polymorphisms of the TNF gene cluster in a large group of patients with HCL. The TNF‐α (−308  bp) promoter/enhancer point mutation and two polymorphisms located within the first intron of the lymphotoxin (LT)‐α gene showed neither significant allelic deviation for the patient group nor, after analysis of clinical characteristics such as blood counts, stable or progressive disease or response to therapy.
Detection of drug‐dependent, platelet‐reactive antibodies by solid‐phase red cell adherence assays
Authors: Leach M. F. , Cooper L. K. , AuBuchon J. P.
Published: November 2003

We developed a simple modification of the solid‐phase red cell adherence (SPRCA) assay system that can be used to identify drug‐dependent platelet antibodies (DDPAs) reactive by either the hapten or immune complex reaction mechanisms. Between January 1994 and August 1996 we tested sera from 173 patients [123 (71%) with unexplained thrombocytopenia and 50 (29%) because of poor responses to platelet transfusions not explicable by alloimmunization or the clinical situation] for DDPAs possibly associated with the receipt of 61 different drugs. We correlated positive results with patients’ clinical courses. DDPAs were identified in samples from 138 (80%) of the patients tested. Antibodies reactive only by the hapten mechanism were identified in 51 (37%) of those sera exhibiting positive reactions. The clinical courses of 108 (78%) patients were evaluable. Discontinuation of the implicated drug(s) resulted in prompt (<5 d) resolution of the thrombocytopenia or improvement in response to transfusion in all of these patients. In four cases thrombocytopenia returned upon re‐exposure to the implicated drug. This adaptation of SPRCA provides a simple means of investigating the possibility of DDPAs and documents a higher frequency of these antibodies than has previously been suspected.
Acute basophilic leukaemia and translocation t(X;6)(p11;q23)
Authors: Dastugue Nicole , Duchayne Eliane , Kuhlein Emilienne , Rubie Hervé , Demur Cécile , Aurich Joan , Robert Alain , Sie Pierre
Published: November 2003

We report two infants with acute basophilic leukaemia associated with a t(X;6)(p11;q23) as the sole abnormality. Morphologic evidence of basophilic lineage was provided by light and electron microscopy. Both patients also had a similar presentation on diagnosis, characterized by clinical signs consistent with a hyperhistaminaemia syndrome, i.e. urticarian rashes and gastro‐intestinal disorders evocative of peptic ulcer. Immunophenotypes differed in the two patients, one expressing CD24, CD13 and CD33, whereas only CD117 was found in the other.

Basophilic acute leukaemia, a rare group among acute leukaemias, might be nonrandomly associated with a specific chromosomal abnormality, t(X;6)(p11;q23). This new entity might also be identifiable by an uncommon clinical presentation and occurrence in infancy.
Activation during preparation of therapeutic platelets affects deterioration during storage: a comparative flow cytometric study of different production methods
Authors: METCALFE PAUL , WILLIAMSON LORNA M. , REUTELINGSPERGER CHRIS P. M. , SWANN IAN , OUWEHAND WILLEM H. , GOODALL ALISON H.
Published: November 2003

Three different separation methods, all using centrifugation, are routinely used to prepare therapeutic platelet concentrates from human donor blood. Platelet concentrates derived from platelet‐rich plasma (PRP‐PC), buffy coat (BC‐PC) and apheresis (AP‐PC) were investigated at the end of production, and over an 8 d storage period. Change in platelet surface markers were measured by flow cytometry, using fluorescein‐conjugated antibodies to fibrinogen, P‐selectin (CD62P), GPIIb–IIIa (CD41), GPIbα (CD42b) and GPV (CD42d), and fluorescein‐conjugated Annexin V was used to measure expression of anionic phospholipid.

All concentrates showed some changes during preparation but PRP‐PC underwent the greatest changes with significantly higher levels of P‐selectin (P < 0.001) and bound Annexin V (P = 0.001) than AP‐PC or BC‐PC, and lower levels of GPIbα (P = 0.002) and GPV (P < 0.001). These changes were attributable to component separation rather than venesection. These markers all continued to change on storage with a strong positive correlation between the changes seen during production and those after 5 d storage. PRP‐PC continued to show the greatest changes whereas BC‐PC showed the least. Fibrinogen was bound to 40–50% of platelets in all preparations and this did not alter significantly on storage whereas total expression of GPIIb–IIIa remained unchanged throughout.

There was no evidence that the platelet surface changes were thrombin‐mediated and leucocyte depletion of BP‐PC by filtration had no effect on the changes. It is proposed that the deterioration of platelet concentrates during storage may be related to activation occurring during preparation. ‘Whole blood’ flow cytometry using a panel of fluorescein‐labelled reagents provides an informative method for evaluating platelet concentrates.
Liposomal 1,25 (OH)2 vitamin D3 compounds block proliferation and induce differentiation in myelomonocytic leukaemia cells
Authors: FRANKENBERGER MARION , HOFMANN BARBARA , EMMERICH BERTOLD , NERL CHRISTOPH , SCHWENDENER RETO A. , LÖMS ZIEGLER‐HEITBROCK H.W.
Published: November 2003

The vitamin D3 derived hormone 1,25 (OH)2 vitamin D3 (1,25 D3) is able to induce growth arrest and differentiation in myelomonocytic leukaemia cells. In order to allow for specific delivery to leukaemic cells the lipophilic compound was incorporated into the lipid membranes of liposomes. Liposomal 1,25 D3 reduced proliferation as measured by 3H‐thymidine incorporation in HL60 leukaemia cells by up to 60%. When liposomes were prepared at different concentrations of 1,25 D3 65% inhibition was achieved at 48 nM. The MC 1288 stereoisomer of 1,25 D3 was more potent and had the same activity at 48 nM.

The effect of the liposomal compounds was specific to myeloid cells as they reduced proliferation in myelomonocytic HL60, monoblastic U937 and monocytic Mono Mac 6 cells but not in the T‐cell lines Jurkat and Molt 4.

The antiproliferative effect of liposomal 1,25 D3 was associated with an induction of differentiation since treated HL60 cells showed a monocytic morphology, increased expression of CD14 and decreased expression of CD33.

When peripheral blood leukaemic cells from M4 and M5 acute myeloid leukaemia (AML) patients were admixed with liposomal compounds an antiproliferative effect was seen in all five cases, including the two cases where free compounds led to enhanced growth. Liposomal delivery of 1,25 (OH)2 vitamin D3 may offer a novel approach to treatment of myelomonocytic leukaemia.
Cytogenetic abnormalities and their prognostic significance in idiopathic myelofibrosis: a study of 106 cases
Authors: Reilly J. T. , Snowden J. A. , Spearing R. L. , Fitzgerald P. M. , Jones N. , Watmore A. , Potter A.
Published: November 2003

The prognostic significance of cytogenetic abnormalities was determined in 106 patients with well‐characterized idiopathic myelofibrosis who were successfully karyotyped at diagnosis. 35% of the cases exhibited a clonal abnormality (37/106), whereas 65% (69/106) had a normal karyotype. Three characteristic defects, namely del(13q) (nine cases), del(20q) (eight cases) and partial trisomy 1q (seven cases), were present in 64.8% (24/37) of patients with clonal abnormalities. Kaplan‐Meier plots and log rank analysis demonstrated an abnormal karyotype to be an adverse prognostic variable (P < 0.001). Of the eight additional clinical and haematological parameters recorded at diagnosis, age (P < 0.01), anaemia (haemoglobin ≤10 g/dl; P < 0.001), platelet (≤100 × 109/l, P < 0.0001) and leucocyte count (>10.3 × 109/l; P = 0.06) were also associated with a shorter survival. In contrast, sex, spleen and liver size, and percentage blast cells were not found to be significant. Multivariate analysis, using Cox's regression, revealed karyotype, haemoglobin concentration, platelet and leucocyte counts to retain their unfavourable prognostic significance. A simple and useful schema for predicting survival in idiopathic myelofibrosis has been produced by combining age, haemoglobin concentration and karyotype with median survival times varying from 180 months (good‐risk group) to 16 months (poor‐risk group).
Recurrent PIG‐A mutation (IVS5+1G→A) in a paediatric case of paroxysmal nocturnal haemoglobinuria: detection by the protein truncation test
Authors: Maugard Claude , Margueritte Geneviève , Tuffery Sylvie , Rabesandratana Heri , Demaille Jacques , Claustres Mireille
Published: November 2003

Paroxysmal nocturnal haemoglobinuria (PNH) is an acquired haemopoietic stem cell disorder caused by the absence of glycosyl phosphatidylinositol (GPI)‐anchored surface proteins due to a deficient biosynthesis of GPI‐anchor. The disease occurs predominantly in adults, and very few cases have been described in children and adolescents. Recent analyses have shown that null mutations in the X‐linked PIG‐A (phosphatidylinositol glycan‐class A) gene are responsible for GPI‐anchor deficiency in most PNH adult patients analysed. We report a young male from southern France who was diagnosed with PNH at 12 years of age during follow‐up of aplastic anaemia. To further elucidate the molecular basis of PNH occurring in childhood, we used the powerful and rapid protein truncation test to scan for truncative mutations in the entire PIG‐A mRNA reverse transcribed and amplified from blood mononuclear cells. The somatic defect responsible for PNH in the patient was found to be a splicing mutation, IVS5+1G→A, which has previously been described in two Asiatic adults with PNH.
Should cytomegalovirus be tested for in both blood and bronchoalveolar lavage fluid of patients at a high risk of CMV pneumonia after bone marrow transplantation?
Authors: Ibrahim A. , Gautier E. , Roittmann S. , Bourhis J.‐H. , Fajac A. , Charnoz I. , Terrier P. , Salord J‐M. , Tancrède C. , Hayat M. , Bernaudin J.‐F. , Pico J.‐L.
Published: November 2003

To identify and treat patients at high risk of cytomegalovirus (CMV) pneumonia after bone marrow transplantation (BMT), we tested for CMV viraemia weekly, and performed broncho‐alveolar lavage (BAL) on day 35 post‐transplant in 63 recipients. 36 allogeneic BMT recipients were at a high risk of CMV pneumonia (25 CMV‐seropositive recipients and 11 patients receiving marrow from a CMV‐seropositive donor). Patients with a positive BAL or viraemia received a 14 d course of ganciclovir or foscarnet. CMV was detected in 29 (46%) of the 63 BMT recipients and excretion of CMV in blood and BAL was significantly linked. However, among the 29 patients who excreted the virus, only 10 (35%) shed CMV in blood and BAL at the same time; 19 patients (65%) had detectable CMV in blood (11 patients) or BAL (eight patients) only. Therefore, on the basis of viraemia or BAL alone, 21/29 patients (70%) and 18/29 patients (60%), respectively, would have received antiviral treatment. BAL increased the CMV detection rate by 13% (8/63 patients) relative to viraemia. With this strategy, the incidence of CMV pneumonia was reduced to 3% in allografted patients. Only two of the 19 autografted patients developed fatal CMV pneumonia. We avoided anti‐CMV treatment in 54% of all the BMT recipients. In conclusion, CMV should be tested for in both blood and BAL fluid of BMT recipients at high risk of CMV pneumonia.
Interferon‐γ+ and interleukin‐2+ T cells in peripheral blood from multiple myeloma patients in relation to disease status and maintenance therapy with interferon‐α2b (intron A)
Authors: Millar Barbara C. , Millar J. L. , Powles R. L. , Catovsky D.
Published: November 2003

Peripheral blood T cells from 83 patients with multiple myeloma (MM) were examined for the production of interferon‐γ (INFγ) and interleukin‐2 (IL‐2) using three‐colour flow cytometry. Comparisons were made between the percentage of cytokine‐positive lymphocytes in normal donors and in patients during remission or relapse. Patients were divided into those who were on maintenance therapy with interferon‐α2b (intron A) and those who had no further treatment after high‐dose melphalan (HDM) with or without autologous bone marrow (ABMR) or peripheral blood stem cell rescue (PBSCR).

The percentage of INFγ+/CD3+, INFγ+/CD45R0+/CD3+ and IL‐2+/CD8+ was higher in patients on INFα2b during remission and relapse compared with normal donors (P < 0.005). During remission INFγ+/CD45R0+/CD3+ and IL‐2+/CD8+ lymphocytes were higher in patients not on INFα2b (P < 0.05 and P < 0.005, respectively). In relapsed patients INFγ+/CD3+ and INFγ+/CD45R0+/CD3+ were increased in patients not taking INFα2b (P < 0.005). There was no significant difference between the percentages of cytokine‐positive lymphocytes in patients taking or not taking INFα2b either during remission or relapse.

Plasma IL‐6 levels were similar in both groups of patients during remission. The data suggest that if maintenance therapy with INFα2b induces the synthesis of INFγ and IL‐2 in vivo, the magnitude of the effect is small and may be unimportant in providing an anti‐tumour effect in the majority of patients.
FIRST DESCRIPTION OF A FRAMESHIFT MUTATION IN THE α1‐GLOBIN GENE ASSOCIATED WITH α‐THALASSAEMIA
Authors: Ayala Sonia , Colomer Dolors , Aymerich Marta , Abella Eugenia , Vives Corrons Joan Lluís
Published: November 2003

A frameshift mutation in the α1‐globin gene, responsible for a clinically mild α‐thalassaemia phenotype, has been characterized in a Spanish woman. After excluding the most common forms of α‐thalassaemia found in the Mediterranean area, both α‐globin genes (α1 and α2) were amplified and analysed selectively by non‐radioactive single‐strand conformation polymorphism (SSCP). An abnormal SSCP mobility was present in the second exon of the α1‐globin gene and direct sequence analysis revealed a 13 bp deletion (between codons 51 and 55) affecting a single allele. The consequence of this mutation is a reading frameshift leading to a novel amino acid coding sequence from codons 51–61 and a premature stop signal at new position 62, which results in a net reduction of the affected α‐globin chain output. The presence of this new mutation was confirmed by restriction enzyme analysis of the specific PCR product.
Expression of CD8β and alteration of cell surface phenotype in adult T‐cell leukaemia cells
Authors: Joh Tatsuroh , Yamada Yasuaki , Seto Masao , Kamihira Shimeru , Tomonaga Masao
Published: November 2003

Typical adult T‐cell leukaemia (ATL) cells have a CD4+CD8− cell surface phenotype, but atypical phenotypes such as CD4+CD8+ and CD4−CD8+ have also been reported. The CD8 molecule is composed of α and β chains and commonly used monoclonal antibodies against CD8 molecule detect only CD8α. Since it has been reported that CD8α can be induced in mature CD4+ T cells by cell activation, but not CD8β, we studied whether ATL cells which express CD8α may also express CD8β. We found some cases of CD8α+ ATL were also positive for CD8β. Furthermore, we experienced a case whose ATL cell surface phenotype changed from CD4+CD8α+CD8β+ to CD4−CD8α+CD8β+ and finally to CD4+CD8α−CD8β−. Southern blot analysis revealed that the monoclonal integration of human T lymphotropic virus type I (HTLV‐I) was identical throughout the course of the study, indicating that a single clone had demonstrated the alterations. These data suggest that peripheral CD4+CD8+ ATL cells can express not only CD8α, but also CD8β and that a single ATL cell clone has the potential to change its surface phenotype in vivo as well as in vitro.
Trisomy 14 is a non‐random karyotypic abnormality associated with myeloid malignancies
Authors: Toze C. L. , Barnett M. J. , Naiman S. C. , Horsman D. E.
Published: November 2003

Isolated gain of chromosome 14 (trisomy 14 or +14) has been reported in myeloid malignancy. Seven cases were identified by review of all diagnostic bone marrow specimens with cytogenetics performed at our institution from 1983 to 1995. Median age was older (72 years) and diagnosis was myelodysplasia in the majority of cases. Although trilineage dysplasia occurred, platelet counts were relatively well preserved (median 131×109/l). Mosaic karyotype (normal plus abnormal metaphases) was seen in the majority of cases, and survival from diagnosis was short (<2 years). These features are consistent with data from 30 previously published cases, and support the hypothesis that trisomy 14 occurs as a non‐random cytogenetic abnormality in association with myeloid malignancy.
Immunoglobulins targeting both GPIIb/IIIa and GPIb/IX in chronic idiopathic thrombocytopenic purpura (ITP): evidence for at least two different IgG antibodies
Authors: HOU MING , STOCKELBERG DICK , KUTTI JACK , WADENVIK HANS
Published: November 2003

Antiplatelet antibodies in chronic idiopathic thrombocytopenic purpura (ITP) mainly target glycoprotein (GP) IIb/IIIa and GPIb/IX. Previous studies, employing modern antigen‐specific assays, indicate that serum reactive with both GPIIb/IIIa and GPIb/IX is not an uncommon finding in chronic ITP. However, the mechanism behind this dual reactivity remains unclear. We studied sera from 72 patients with chronic ITP using modified GPIIb/IIIa‐ and GPIb/IX‐specific MAIPA assays. Among the 34 positive sera, seven showed strong reactivity against both GPIIb/IIIa and GPIb/IX. These seven dual reactive ITP sera were further analysed by absorption studies. It was found that sera absorbed with immobilized GPIb/IX lost nearly all serum IgG specific for GPIb/IX but fully retained the IgG specific for GPIIb/IIIa. Conversely, sera absorbed with immobilized GPIIb/IIIa retained their reactivity only with GPIb/IX. These findings demonstrate that ITP sera, reactive with both GPIIb/IIIa and GPIb/IX, contain at least two different IgG antibody populations, each reactive with only one of the GP complexes.
The effects on growth and survival of IL‐6 can be dissociated in the U‐266‐1970/U‐266‐1984 and HL407E/HL407L human multiple myeloma cell lines
Authors: Spets Helena , Jernberg‐Wiklund Helena , Sambade Clara , Söderberg Ola , Nilsson Kenneth
Published: November 2003

Several studies have documented IL‐6‐dependent growth promotion of murine and human neoplastic plasma cells. However, it is well known that human multiple myeloma (MM) cells in vitro show a considerable degree of heterogeneity concerning growth and survival requirements. This heterogeneity, which probably reflects overlapping effects of feeder cells, interleukin 6 (IL‐6) and components of fetal calf serum (FCS) as well as tumour heterogeneity in vivo, has hampered the elucidation of molecular mechanisms underlying the effects of IL‐6. In an attempt to dissociate growth and survival promotion of IL‐6, we have studied two pairs of human MM cell lines, HL407E/HL407L and U‐266‐1970/U‐266‐1984, selected to represent different stages of in vitro tumour progression and dependence of feeder cells and exogenous IL‐6. We demonstrated that exogenous IL‐6, in the presence of FCS, conveyed: (a) a strong growth stimulatory effect with weak or no survival promotion in HL407L and U‐266‐1970 cells; (b) promotion of survival with no effects on growth in HL407E cells; (c) no growth or survival promotion to U‐266‐1984. Moreover, our results suggested that IL‐6 may enhance apoptosis in U‐266‐1970/U‐266‐1984 cells, and that FCS may interfere with IL‐6 in its growth stimulatory effect. The relative dissociation of growth, survival and apoptotic effects of IL‐6 leads to the conclusion that the HL407E/HL407L and U‐266‐1970/U‐266‐1984 pairs of cell lines provide a useful human model system to study molecular mechanisms underlying these separate events.
Therapeutic leukapheresis in hyperleucocytic leukaemias: lack of correlation between degree of cytoreduction and early mortality rate
Authors: Porcu Pierluigi , Danielson Constance F. , Orazi Attilio , Heerema Nyla A. , Gabig Theodore G. , McCarthy Leo J.
Published: November 2003

The clinical and laboratory data of 48 leukapheresis‐treated patients with hyperleucocytic leukaemia (HL) was reviewed to assess the correlation between the degree of leucoreduction and early mortality. Leukapheresis resulted in > 50% leucoreductions and postapheresis WBC counts <100 × 109/l in most patients (64.5%). Patients presenting with neurological, respiratory or renal complications had higher early mortality rates than patients without such complications, despite similar initial WBC counts and comparable leucoreductions. Thus, in these patients, more efficient leucoreduction was not associated with improved early survival.
Detection of melanoma cells in peripheral blood stem cell harvests of patients with progressive metastatic malignant melanoma
Authors: Probst‐Kepper Michael , Schrader Andres , Buer Jan , Grosse Jens , Volkenandt Matthias , Illiger Hans‐Joachim , Metzner Bernd , Kadar Janosch , Duensing Stefan , Hertenstein Bernd , Ganser Arnold , Atzpodien Jens
Published: November 2003

The detection of melanocyte‐specific messenger RNA in patients with malignant melanoma suggests the potential contamination of peripheral blood stem cell (PBSC) harvests by neoplastic cells. In this study, the melanocyte‐specific transcripts of tyrosinase and Melan‐A/MART‐1 were used to detect neoplastic cells in PBSC harvests of nine metastatic malignant melanoma patients. Only one patient’s PBSC harvest tested positive for tyrosinase. All harvests were negative for Melan‐A/MART‐1. Our results suggest that contamination of PBSC harvests with neoplastic cells may not contribute to disease progression following high‐dose chemotherapy in advanced malignant melanoma.
Analysis of ETV6 and ETV6‐AML1 proteins in acute lymphoblastic leukaemia
Authors: AGAPE PHILIPPE , GERARD BENEDICTE , CAVE HELENE , DEVAUX ISABELLE , VILMER ETIENNE , LECOMTE MARIE‐CHRISTINE , GRANDCHAMP BERNARD
Published: November 2003

The t(12;21) is a recurring chromosomal abnormality in acute lymphoblastic leukaemias (ALLs) which results in the production of an ETV6‐AML1 fusion gene. The association between t(12;21) and the deletion of the untranslocated allele of ETV6 is among the most frequent abnormalities observed in B‐lineage ALLs in children. In order to study the proteins encoded by ETV6ETV6‐AML1, we raised polyclonal antibodies directed against a recombinant peptide corresponding to the junctional region of ETV6‐AML1. Cell lysates from various leukaemic cell lines, and from children with B‐ and T‐lineage ALLs, were studied by Western blot.

Two isoforms of ETV6 protein were detected in normal bone marrow cells and in leukaemic cells without 12p alteration: a major form (apparent m.w. 63 kD) and a minor one (apparent m.w. 53 kD).

In the REH cell line, which expresses the ETV6‐AML1 fusion transcript and no normal ETV6 mRNA, the ETV6 isoforms were absent and two new bands were detected corresponding to ETV6‐AML1 protein products (apparent m.w. 95 and 105 kD). A similar pattern was obtained with blast cells from patients with a t(12;21) and a deletion of ETV6.

In two patients with a t(12;21) but no deletion of ETV6, four bands were detected corresponding to both the normal ETV6 and ETV6‐AML1 proteins, suggesting that in these cases the second ETV6 allele was not inactivated.

Surprisingly, the expression pattern of ETV6 differed widely from patient to patient. In three out of 13 patients without t(12;21), the relative intensity of the bands corresponding to ETV6 isoforms in blast cells from patients was completely different from normal cells, with a marked predominance of the 53 kD isoform. The pattern of ETV6 expression was normal in bone marrow from the same patients during remission. These finding suggest that ETV6 abnormalities are not restricted to patients with translocations or deletions involving this gene.
Reduced interferon‐alpha production by Epstein‐Barr virus transformed B‐lymphoblastoid cell lines and lectin‐stimulated lymphocytes in congenital dyserythropoietic anaemia type I
Authors: WICKRAMASINGHE S. N. , HASAN R. , SMYTHE J.
Published: November 2003

The concentrations of interferon‐α (IFN‐α) in supernatants from cultures of Epstein‐Barr virus (EBV) transformed B‐lymphoblastoid cell lines derived from seven patients with congenital dyserythropoietic anaemia (CDA) type I were below the 95% confidence limits for those derived from six healthy subjects. In contrast, the concentrations of IFN‐α in supernatants from cultures of EBV‐transformed lymphoblastoid cell lines derived from four patients with other types of CDA and four patients with hereditary sideroblastic anaemia were normal. Supernatants from cultures of peripheral blood lymphocytes stimulated with phytohaemagglutinin or pokeweed mitogen contained less IFN‐α when the cells were derived from patients with CDA type I than when derived from healthy subjects. Since patients with CDA type I show a substantial haematological response to treatment with IFN‐α, the data suggest that impaired IFN‐α production may be an important pathogenetic mechanism in CDA type I.
Ex vivo cytokine expansion of peripheral blood 5‐fluorouracil‐treated CD34‐positive chronic myeloid leukaemia cells increases the selection of Ph‐negative cells
Authors: Mahon FranÇois‐Xavier , Pigeonnier‐Lagarde Valérie , Chahine Hikmat , Barbot Caroline , Jazwiec Bozena , Ripoche Jean , Reiffers Josy
Published: November 2003

It has been shown that normal early progenitor or stem cells persist in the blood and bone marrow of patients with Philadelphia chromosome (Ph)‐positive chronic myeloid leukaemia (CML); it is also known that normal haemopoietic progenitors can be expanded ex vivo in the presence of various cytokine combinations. The selection of normal (Ph‐negative) progenitor cells from CML patients would potentially be of considerable clinical value for ex vivo purging and autologous transplantation. To obtain these cells, CD34‐positive (progenitor) cells from the peripheral blood (PB) of CML patients were pretreated with 5‐fluorouracil (5FU) (5 μg/ml) to suppress Ph‐positive cells and then grown in suspension culture for 7 d with a combination of cytokines. We compared two combinations of cytokines: interleukin‐1α (IL1α) and interleukin‐3 (IL3) with either Flt3‐ligand (FLT3L) or stem cell factor (SCF). Using these two combinations, we obtained the same degree of day 14 CFU‐GM expansion (3.1 ± 0.5 and 3.4 ± 0.7 fold expansion). FISH analysis showed that 5FU pretreatment significantly favoured a higher frequency of Ph‐negative cells after expansion in liquid culture. Moreover, after 5FU pretreatment, the mean (±SEM) percentage of Ph negativity was significantly greater for IL1α‐IL3‐FLT3L compared to IL1α‐IL3‐SCF (19.1 ± 2.5% v 14.8 ± 2.3%, P = 0.009, n = 7). The output long‐term culture initiating cells (LTC‐IC) which could only be detected after 5FU pretreatment and the combination, IL1α‐IL3‐FLT3L were all Ph negative by FISH analysis. Thus, a subset of Ph‐negative cells was selected from CML PB by 5FU and expanded using the combination of cytokines IL1α‐IL3‐FLT3L and IL1α‐IL3‐SCF. Ex vivo expansion of putatively normal haemopoietic progenitor cells is feasible in CML.
Establishment of a novel human myeloid leukaemia cell line (HNT‐34) with t(3;3)(q21;q26), t(9;22)(q34;q11) and the expression of EVI1 gene, P210 and P190 BCR/ABL chimaeric transcripts from a patient with AML after MDS with 3q21q26 syndrome
Authors: Hamaguchi Hiroyuki , Suzukawa Kazumi , Nagata Kaoru , Yamamoto Katsuya , Yagasaki Fumiharu , Morishita Kazuhiro
Published: November 2003

A novel human myeloid leukaemia cell line (HNT‐34) was established from the peripheral blood of a 45‐year‐old female patient with acute myelogenous leukaemia (AML) transformed from chronic myelomonocytic leukaemia (CMMoL) with 3q21q26 syndrome. Morphologically, the HNT‐34 cells were undifferentiated blasts which were negative for myeloperoxidase. The HNT‐34 cells were positive for CD4, CD13, CD33 and CD34, but negative for CD41a and CD42b. The cells actively proliferated in suspension with a doubling time of 26–27 h in the absence of any growth factors. Neither proliferative advantage nor differentiation was observed with the addition of G‐CSF, GM‐CSF, IL‐3, TPO, DMSO or PMA. Cytogenetic analysis showed 46,XX, t(3;3)(q21;q26), t(9;22)(q34;q11),20q−. Molecular analysis showed expression of EVI1 gene, P210 and P190 BCR/ABL chimaeric transcripts. The chromosomal breakpoint at 3q26 of HNT‐34 cell line was located to approximately 200 kb 5′ of FIM3 locus and more upstream of the MDS1, which is the same region as that of somatic cell hybrid line H10C. The breakpoint at 3q21 was located within the 390 kb centromeric from the breakpoint cluster region. These results suggest that the HNT‐34 cell line may be a useful tool for the elucidation of the mechanisms of leukaemogenesis which involve the 3q21q26 syndrome and Ph1 chromosome.
The eosinophil granule proteins in serum, but not the oxidative metabolism of the blood eosinophils, are increased in cancer
Authors: Trulson A. , Nilsson S. , Venge P.
Published: November 2003

The eosinophil activity in patients with renal cell adenocarcinoma during treatment with interleukin‐2 (IL‐2) and interferon‐alpha (IFN‐α) was reduced when measured as zymosan‐induced lucigenin‐enhanced chemiluminescence (CL). Eosinophil cationic protein (ECP), eosinophil peroxidase (EPO) and eosinophil protein‐X (EPX) were significantly elevated before and during treatment (P < 0.001) when compared with the controls. ECP and EPO were unaffected by the treatment whereas it induced an increased EPX level compared with values measured before treatment (P < 0.05). The propensity of eosinophils to secrete their granule proteins may reflect the fact that eosinophils in cancer patients have an enhanced capacity to kill cancer cells.
Missense mutations at ALA‐10 in the factor IX propeptide: an insignificant variant in normal life but a decisive cause of bleeding during oral anticoagulant therapy
Authors: Oldenburg Johannes , Quenzel Ernst‐Markus , Harbrecht Uschi , Fregin Andreas , Kress Wolfram , Müller Clemens Rainer , Hertfelder Hans‐Jörg , Schwaab Rainer , Brackmann Hans‐Hermann , Hanfland Peter
Published: November 2003

Bleeding complications are the most common and unwanted side‐effect of oral anticoagulant therapy. We report three patients in whom mutations in the factor IX (FIX) propeptide were found to cause severe bleeding during coumarin therapy. Strikingly, the bleeding occurred within the therapeutic ranges of the prothrombin time (PT) and international normalized ratio (INR). In all three patients coumarin therapy caused an unusually selective decrease of FIX activity (FIX:C) to levels below 1–3%. Upon withdrawal of coumarin, FIX:C increased to subnormal or normal values of 55%, 85% and 125%, respectively. Analysis of the FIX gene revealed two different missense mutations affecting the Ala‐10 residue in the propeptide coding region: Ala[GCC] to Val[GTC] in two patients and Ala[GCC] to Thr[ACC] in one patient. No further mutation was detected by screening 195 random blood donors for mutations at Ala‐10, thus excluding a frequent polymorphism at this position. The mutation in the FIX propeptide at a position which is essential for the carboxylase recognition site causes a reduced affinity of the carboxylase enzyme to the propeptide. This effect leads to an impaired carboxylase epoxidase reaction which is decisively triggered by the vitamin K concentration. Determination of FIX and APTT in addition to PT and INR is therefore recommended in coumarin‐treated patients with an uncommon bleeding pattern.
Health‐related quality of life assessed before and during chemotherapy predicts for survival in multiple myeloma
Authors: WislØff Finn , Hjorth Martin
Published: November 2003

Measurement of health‐related quality of life was integrated into a randomized trial (NMSG 4/90) comparing melphalan/prednisone to melphalan/prednisone + interferon α‐2b in newly diagnosed multiple myeloma. One of the aims of the study was to assess the prognostic significance of quality‐of‐life scores, using the EORTC QLQ‐C30 questionnaire. Univariate analysis showed a highly significant association with survival from the start of therapy for physical functioning as well as role and cognitive functioning, global quality of life, fatigue and pain. In multivariate analysis, physical functioning and W.H.O. performance status were independent prognostic factors (P values=0.001 for both) when analysed in a Cox regression model with the somatic variables β‐2 microglobulin, skeletal disease and age. The best prediction for survival from the start of therapy was obtained by combining the β‐2 microglobulin and physical functioning scores in a variable consisting of three risk factor levels with an estimated median survival of 17, 29 and 49 months, respectively. At a 12 months landmark analysis, the relative risk for patients with physical functioning score 0–20 v 80–100 was 5.63 (99% CI 2.76–11.49), whereas the relative risk for patients without an objective response to chemotherapy compared to those with at least a minor response was 2.32 (99% CI 1.44–3.74). Quality‐of‐life assessment may be an independent and valuable addition to the known prognostic factors in multiple myeloma.
Anti‐platelet antibodies in patients with systemic lupus erythematosus and the primary antiphospholipid antibody syndrome: their relationship with the observed thrombocytopenia
Authors: Macchi Laurent , Rispal Patrick , Clofent‐Sanchez Gisèle , Pellegrin Jean‐Luc , Nurden Paquita , Leng Bernard , Nurden Alan T.
Published: November 2003

The role of antiphospholipid antibodies in the pathogenesis of the thrombocytopenia observed during primary antiphospholipid antibody syndrome (APAS) and systemic lupus erythematosus (SLE) remains controversial. We have used the MAIPA test to examine the frequency and specificity of anti‐platelet antibodies directed against the major platelet membrane glycoproteins (GP IIb–IIIa, GP Ib–IX, GP Ia–IIa and GP IV) in patients where SLE and APAS were associated or not with thrombocytopenia. Results were compared with a series of 26 ITP patients, 46% of whom were shown to possess anti‐platelet antibodies directed against one or more of the platelet surface glycoproteins. When APAS was associated with thrombocytopenia, 7/10 patients possessed antibodies against GP IIb–IIIa and/or GP Ib–IX. For SLE patients with thrombocytopenia, 6/10 patients were shown to have antiplatelet antibodies against GP IIb–IIIa, GP Ib–IX or GP IV. In contrast, for APAS (n=11) and SLE patients (n=11) without thrombocytopenia, only one patient had an antibody directed against GP IIb–IIIa and one patient had an antibody to GP IV. Our results suggest that antibodies directed against major platelet membrane glycoproteins may play a role in the thrombocytopenia that is seen during SLE and APAS.
PML/RARα rearrangement in acute promyelocytic leukaemia with t(1;17) elucidated using fluorescence in situ hybridization
Authors: Eclache Virginie , Benzacken Brigitte , Le Roux Genevieve , Casassus Philippe , Chomienne Christine
Published: November 2003

Acute promyelocytic leukaemia (APL) is characterized by t(15;17)(q22;q21) which results in the formation of two chimaeric genes, PML/RARα and RARα/PML, thought to play a role in leukaemogenesis. We report a case of a patient with APL apparently lacking the t(15;17) but with t(1;17) translocation identified by fluorescence in situ hybridization (FISH). Chromosome 15 seemed intact but PML/RARα fusion transcript was detected by molecular analysis. The patient achieved complete remission with all‐trans retinoic acid treatment associated with chemotherapy. This case illustrates the usefulness of combined cytogenetics, FISH and molecular biology in cases with no evident t(15;17) to predict response to treatment.