bluuebunny/crossref_metadata_2025 huggingface dataset:
Retinoic acid stimulates plasminogen activator inhibitor 2 production by blood mononuclear cells and inhibits urokinase‐induced extracellular proteolysis
Authors: Montemurro P. , Barbuti G. , Conese M. , Gabriele S. , Petio M. , Colucci M. , Semeraro N.
Published: March 2003

Retinoids have been shown to modulate several functions of mononuclear phagocytes. We investigated the in vitro effect of all‐trans‐retinoic acid (ATRA) on the production of two major fibrinolytic components, urokinase‐type plasminogen activator (u‐PA) and PA inhibitor 2 (PAI‐2), by human blood mononuclear cells (MNC). ATRA caused a dose‐dependent (range 0.01–10 μm) accumulation of PAI‐2 antigen and activity into the cell culture medium, with a maximal increase (about 5‐fold over control) at a concentration of 1–10 μm. Similarly, a dose‐dependent increase in PAI‐2 antigen was observed in cell extracts upon ATRA stimulation. Northern blot analysis showed a parallel increase in the amount of PAI‐2 mRNA in ATRA‐treated cells. Time‐course experiments with 1 μm ATRA showed enhanced PAI‐2 mRNA expression as early as 2 h, reaching a maximum at 4–6 h and then declining at 18–24 h, and a time‐dependent increase in PAI‐2 antigen in the cell culture medium. At variance with PAI‐2, u‐PA was not influenced by the drug. To establish whether ATRA‐induced changes influenced the fibrinolytic process, we evaluated the effect of MNC stimulated with ATRA on u‐PA‐induced degradation of diluted plasma clots. ATRA‐treated cells markedly inhibited clot lysis induced by low concentrations of u‐PA. The effect was due to enhanced extracellular PAI‐2 accumulation since it was observed with conditioned medium from ATRA‐treated cells; it was abolished by the addition of neutralizing anti‐PAI‐2 antibodies and was negligible when single‐chain t‐PA was used instead of u‐PA. Since monocyte/macrophage‐mediated, plasminogen‐dependent extracellular proteolysis has been proposed as an important mechanism of tissue damage in several inflammatory states, our findings might contribute to better explain the anti‐inflammatory properties of retinoids.
Enhanced retroviral gene transfer into CML and normal bone marrow, and CML and mobilized peripheral blood CD34+ cells using the recombinant fibronectin fragment CH‐296
Authors: Garrett Elaine , Garin Marina I. , Miller Angela R‐M. , Goldman John M. , Melo Junia V. , Apperley Jane F.
Published: March 2003

Autologous stem cell transplantation is a therapeutic alternative for many chronic myeloid leukaemia (CML) patients ineligible for the only curative treatment of allogeneic bone marrow transplantation. In this study the retroviral transduction of CD34+ progenitor cells isolated from the bone marrow (BM) and peripheral blood (PB) of patients with CML was compared to that of CD34+ cells isolated from the BM and PB of normal individuals and patients with non‐haematological malignancies. A highly significant increase in transduction of all cell types was achieved in the presence of the recombinant fibronectin fragment, CH‐296 (P < 0.05). In the absence of fibronectin, centrifugation produced a marginal improvement in the transduction of all cell types, which was significant only for CMLBM progenitor cells (P < 0.05). There was no significant additive effect when centrifugation was included in the fibronectin infection protocol. In the presence of CH‐296, combinations of three or more cytokines improved transduction for all cell types. The same degree of transduction was observed for both normal and CML cells, irrespective of the variations employed in the infection protocol, suggesting that both leukaemic and non‐leukaemic progenitors are equally susceptible to retroviral infection. These results demonstrate that CH‐296 has a universal beneficial effect on the transduction of haemopoietic progenitor cells, with clear potential for future clinical trials.
Acute tumour lysis syndrome: a case in AL amyloidosis
Authors: Akasheh Marwan S. , Chang Chi P. , Vesole David H.
Published: March 2003

Tumour lysis syndrome (TLS) in plasma cell dyscrasias is extremely rare. TLS has been described in eight cases of multiple myeloma undergoing high‐dose therapy with autologous stem cell transplant (ASCT). Recently, clinical trials of intensive chemotherapy followed by autologous or allogeneic stem cell support has been shown to offer potential benefit in AL (amyloid light‐chain) amyloidosis. TLS in primary AL amyloidosis in this setting has not been previously reported. We report a case of TLS in a patient with AL amyloidosis which developed after high‐dose melphalan chemotherapy supported by ASCT.
Spatial and temporal mapping of c‐kit and its ligand, stem cell factor expression during human embryonic haemopoiesis
Authors: Teyssier‐Le Discorde Magali , Prost Stephane , Nandrot Emeline , Kirszenbaum Marek
Published: March 2003

Receptor tyrosine kinases (RTKs) mediate cellular responses to the extracellular signals involved in the regulation of cell differentiation and proliferation. Ligand binding initiates a cascade of events, such as receptor dimerization and tyrosine phosphorylation. Thec‐kit gene encodes an RTK for stem cell factor (SCF), (c‐kit ligand, KL), both of which play a critical role in the differentiation and growth of haemopoietic stem cells (HSCs). We investigated the expression of the c‐kit and SCF genes and the presence of the corresponding proteins in haemopoietic tissues during human embryogenesis. We have examined c‐kit and SCF transcripts levels in human embryonic yolk sac, the AGM region, and liver at different stages of gestation (days 25 to 63), using RT‐PCR amplification combined with PhosphorImager quantitative analysis and RNase Protection Assay (RPA). Weak levels of SCF gene expression were observed in the AGM region (days 25 to 34) and high levels were found in the early‐stage liver (day 34). The expression of c‐kit transcript was observed in all studied tissues, but at various levels. The restricted presence of SCF protein following mRNA expression was demonstrated in embryonic liver CD38+ haemopoietic cells by immunocytochemistry. These observations suggest that the biological function of the c‐kit receptor plays an important role in the early stages of human haemopoiesis, and that c‐kit/SCF signalling is particularly involved in early human definitive haemopoiesis.
Determinants of iron status and bilirubin levels in congenital dyserythropoietic anaemia type I
Authors: Wickramasinghe  , Thein  , Srichairatanakool  , Porter
Published: March 2003

Seven untransfused patients with congenital dyserythropoietic anaemia type I were investigated to assess the determinants of both iron overload and serum bilirubin levels. The serum ferritin concentration was increased in all patients and non‐transferrin‐bound iron (NTBI) was increased in all but one patient. None of the patients showed the C282Y mutation in the hereditary haemochromatosis gene, HFE. One patient was homozygous for the H63D mutation in this gene. The data indicated that differences in the extent of iron overload were not mediated by co‐inheritance of the C282Y mutation in the HFE gene but could largely be explained by differences in the severity of anaemia and ineffective erythropoiesis, and in the age of the patient. In one patient an unusually high plasma bilirubin level was associated with the variant A[TA]7TAA configuration in the TATA box of the uridine diphosphate glucuronosyltransferase (UGT‐1A) gene promoter, the mutation found in most patients with mild Gilbert's syndrome.
A complete deficiency of coagulation factor XIII A‐subunit due to a novel compound heterozygote of Ser 413 Leu missense and an nt 389 (ins G) frameshift mutation
Authors: Niiya Toshiyuki , Osawa Haruhiko , Bando Shiro , Oto Yoshiko , Tokuda Kiriko , Takeda Namiko , Sumioka Maki , Murase Mitsuharu , Kida Kaichi , Makino Hideichi
Published: March 2003

Coagulation factor XIII consists of two A‐ and two B‐subunits, and either gene mutation can cause a complete deficiency. In a newborn patient with persistent bleeding from the umbilical cord stump, the plasma A‐subunit protein was not detectable. Direct PCR sequencing revealed an nt 389 (ins G) frameshift mutation in exon 4 resulting in a new stop codon and a Ser 413 Leu missense mutation in exon 10 in either allele. His mother and father were heterozygous for the nt 389 (ins G) and the Ser 413 Leu, respectively, with about 50% reduction of the plasma A‐subunit proteins. In all family members examined only those with either mutation showed the reduced subunit A protein levels. Thus, this complete deficiency of factor XIII was due to a novel compound heterozygous mutation in the A‐subunit gene.
Excessive apoptosis of bone marrow erythroblasts in a patient with autoimmune haemolytic anaemia with reticulocytopenia
Authors: Van De Loosdrecht  , Hendriks  , Blom  , Smit  , De Wolf  , Vellenga
Published: March 2003

We report a patient with autoimmune haemolytic anaemia (AIHA) with reticulocytopenia, who showed excessive apoptosis of erythroblasts. Ultrastructural analysis of bone marrow cells showed that 50% of erythroblasts had characteristic features of apoptosis, which was confirmed by staining with Annexin‐V. In addition, in contrast to normal erythroblasts, Ig staining of the entire erythroblast population could be shown. These data show that apoptosis may contribute to the mechanism of reticulocytopenia in AIHA.
Haematopoietic cell transplantation in patients with Fanconi anaemia using alternate donors: results of a total body irradiation dose escalation trial
Authors: MacMillan M. L. , Auerbach A. D. , Davies S. M. , DeFor T. E. , Gillio A. , Giller R. , Harris R. , Cairo M. , Dusenbery K. , Hirsch B. , Ramsay N. K. C. , Weisdorf D. J. , Wagner J. E.
Published: March 2003

Allogeneic haematopoietic cell transplantation (HCT) is the only therapeutic modality capable of correcting the haematologic manifestations of Fanconi anaemia (FA). However, HCT from alternative donors has been associated with poor survival. Between June 1993 and July 1998, 29 FA patients (median age 12·1 years; range 3·7–48·5 years) were enrolled in a prospective phase I–II dose escalation study. All patients were treated with cyclophosphamide 40 mg/kg, total body irradiation (TBI) 450 cGy or 600 cGy and antithymocyte globulin (ATG), followed by HCT from an alternative donor. Graft‐versus‐host disease (GVHD) prophylaxis consisted of cyclosporin A for 6 months, short course methylprednisolone (2 mg/kg/day) between days +5 and +19 and marrow T‐cell depletion by counterflow elutriation. The probability of developing grade III–IV toxicity was 17% (95% CI 3–31%). For the 25 marrow recipients, the probability of neutrophil engraftment (ANC̊  0·5 × 109/l by day 45) was 63% (95% CI 42–82%). Probabilities of grade II–IV acute GVHD and chronic GVHD were 32% (95%CI 10–54%) and 0% respectively. With a median follow‐up of 18 months, the probability of survival for the entire cohort at 1 year was 34% (95% CI 17–51%). The presence of lymphocyte somatic mosaicism [i.e. the presence of diepoxybutane (DEB)‐insensitive cells] was associated with a significantly increased risk of graft failure. Disappointingly, the use of higher dose TBI and post‐transplant ATG did not improve engraftment. More effective peritransplant immunosuppression, especially in FA patients with somatic mosaicism, was required to overcome the barrier of graft rejection. New conditioning regimens adapted to each individual's alkylator sensitivity are needed to improve the outcome of alternative donor HCT for FA.
Frequent good partial remissions from thalidomide including best response ever in patients with advanced refractory and relapsed myeloma
Authors: Juliusson Gunnar , Celsing Fredrik , Turesson Ingemar , Lenhoff Stig , Adriansson Magnus , Malm Claes
Published: March 2003

Twenty‐three patients with advanced and heavily pretreated myeloma were treated with thalidomide. Starting dose was 200 mg/d, and 20 patients had dose escalations up to 400 (n = 5), 600 (n = 12) or 800 mg/d (n = 3), usually in divided doses. Nineteen patients were refractory to recent chemotherapy, and four had untreated relapse after prior intensive therapy. Ten out of 23 patients (43%) achieved partial response (PR; nine with refractory and one with relapsed disease), six patients had minor response or stabilization of the disease and four had disease progression. Another three patients died early from advanced myeloma at less than 3 weeks of thalidomide therapy. Of the 10 patients with PR, seven had a better response than after any prior therapy, despite vincristine–doxorubicin–dexamethasone (VAD)‐based treatment in all but one and high‐dose melphalan with autologous stem cell support in four. Time to achieve PR was rapid in patients receiving thalidomide in divided doses (median 31 d). Responses also included reduced bone marrow plasma cell infiltration and improved general status. Normalized polyclonal gammaglobulin levels were seen in four cases. Six out of 10 patients with PR remained in remission with a median time on treatment of 23 weeks (range 15–50 weeks). Sedation was common but usually tolerable, and some patients continued full‐ or part‐time work. Four patients had skin problems, three patients had pneumonia, one hypothyrosis, one sinus bradycardia and one minor sensory neuropathy. Thalidomide may induce good partial remissions in advanced refractory myeloma with tolerable toxicity, and should be evaluated in other settings for myeloma patients. Divided thalidomide doses seem to reduce time to achieve remission and may improve response rate.
Ex vivo expanded mobilized peripheral blood CD34+ cells accelerate haematological recovery in a baboon model of autologous transplantation
Authors: Norol Françoise , Drouet Michel , Mathieu Jacques , Debili Najet , Jouault Héléne , Grenier Nancy , Laplanche Agnes , Vainchenker William , Hérodin Francis
Published: March 2003

To address the value of ex vivo expanded haematopoietic cells for shortening cytopenia in autologous haematopoietic transplantation, we designed an ex vivo expansion protocol based on a cocktail of early acting cytokines and short‐term culture and tested it in a baboon model. Expansion involved enriched CD34+ peripheral blood haematopoietic cells cultured for 6 d with a combination of FLT3‐L, stem cell factor (SCF), thrombopoietin (TPO) and interleukin (IL)‐3 (50 ng/ml each); CD34+ cells, granulocyte–macrophage colony‐forming units (GM‐CFU) and megakaryocytic colony‐forming units (MK‐CFU) were amplified, respectively, 10·5‐, 20·5‐ and 17·9‐fold. Baboons were submitted to a myeloablative regimen consisting of cyclophosphamide plus total body irradiation (TBI; 6 Gy) and were then grafted with either 2 × 106/kg unmanipulated CD34+ cells (control group, n = 4) or cells cultured from 2 × 106/kg CD34+ cells (expansion group, n = 4). No cytokines were administered after transplantation. All the animals engrafted. The mean times to white blood cell (WBC), granulocyte and platelet recovery were significantly shorter in the expansion group than in the control group: WBC (> 1 × 109/l) and neutrophil (> 0·5 × 109/l) recovery occurred on days 8 (range 6–9) and 9 (range 6–11), respectively, compared with days 12 (range 10–15) and 14 (range 11–16); platelets recovered (> 20 × 109/l) on day 9 (range 7–12) compared with day 13 (range 11–15) in the control group (P < 0·05). No toxicity was observed after reinfusion. No secondary hypoplasia was observed during more than 12 months of follow‐up. Functions of both neutrophils and platelets produced from expanded cells were normal in terms of oxidative metabolism, chemotaxis and the bleeding time. This study shows that in comparison with unmanipulated cells peripheral blood haematopoietic cells expanded from similar doses of CD34+ cells, under the conditions defined here, accelerated both neutrophil and platelet recovery without impairing long‐term haematopoiesis.
Establishment and characterization of an arsenic‐sensitive monoblastic leukaemia cell line (SigM5)
Authors: Walter Roland , Schoedon Gabriele , Bächli Esther , Betts David R. , Hossle Johann P. , Calandra Thierry , Joller‐Jemelka Helen I. , Fehr Jörg , Schaffner Andreas
Published: March 2003

Few human monoblastic cell lines have been characterized to date. We have established the SigM5 cell line from a patient with acute monoblastic leukaemia (FAB M5a). Original leukaemic cells had a karyotype of 47,XY,+8, whereas the cell line showed a stemline clone of 81,XX,Y,Y,1,4,6,7,+8,+8,9,10,10,11,13,16,19[cp], with a minor sideline also present. Cytochemical staining was strongly positive with α‐naphthylbutyrate acetate esterase, particulate positive with Sudan black and weakly positive for myeloperoxidase. Cells were positive for CD13, CD15, CD18, CD23, CD33, CD38, CD45, CD68 and myeloperoxidase. CD14 expression was 3–15%. SigM5 constitutively secreted interleukin (IL)‐2, IL‐8, IL‐10, tumour necrosis factor (TNF)‐α, ferritin, lysozyme, N‐elastase and neopterin upon stimulation with interferon (IFN)‐γ. Cells expressed the proinflammatory mediator macrophage migration inhibitory factor (MIF). All NADPH oxidase subunits were constitutively present, but nitroblue tetrazolium reduction was only detectable upon activation with IFN‐γ. SigM5 monoblasts were sensitive to arsenic trioxide (As2O3) previously not described to induce apoptosis in monoblastic cells. Differing considerably in morphology, immunophenotype and sensitivity to arsenics from the widely used cell lines U937, HL‐60 and THP‐1, SigM5 is a new monoblastic cell line useful for studying leukaemogenesis, monocyte differentiation and tumour cell susceptibility to arsenic compounds.
A novel BCR–ABL transcript (e8a2) with the insertion of an inverted sequence of ABL intron 1b in a patient with Philadelphia‐positive chronic myeloid leukaemia
Authors: Branford S. , Rudzki Z. , Hughes T. P.
Published: March 2003

An atypical BCR–ABL transcript in a patient with Philadelphia (Ph)‐positive chronic myeloid leukaemia (CML) was detected using a long polymerase chain reaction technique. Sequence analysis revealed a joining of BCR exon e8 and ABL exon a2, with a 55‐base pair (bp) insert of an inverted sequence of ABL intron 1b between them, giving rise to an in frame e8a2 BCR–ABL transcript. This is the first report of a BCR–ABL transcript with the insertion of an inverted sequence. The long PCR technique is a useful screen for atypical BCR–ABL transcripts not detected by standard techniques.
Anaplastic large‐cell lymphomas of B‐cell phenotype are anaplastic lymphoma kinase (ALK) negative and belong to the spectrum of diffuse large B‐cell lymphomas
Authors: Haralambieva Eugenia , Pulford Karen A. F. , Lamant Laurence , Pileri Stefano , Roncador Giovanna , Gatter Kevin C. , Delsol Georges , Mason David Y.
Published: March 2003

There is controversy in the literature as to whether anaplastic large‐cell lymphoma of B‐cell phenotype is related to the t(2;5)‐positive T‐ or ‘null’ cell lymphoma of the same morphology. We report a study of 24 lymphomas with morphological features of anaplastic large‐cell lymphoma which expressed one or more B‐cell markers and lacked T‐lineage markers. Clinical features were more in keeping with large B‐cell lymphoma than with classical t(2;5)‐positive anaplastic large‐cell lymphoma, and immunostaining for anaplastic lymphoma kinase (ALK) protein provided no evidence for the (2;5) translocation (or one of its variants). The staining patterns for CD20 and CD79 were typical of diffuse large B‐cell lymphoma, CD30 expression was variable, and most cases (15/22) lacked epithelial membrane antigen (EMA). These findings support the view that ‘B‐cell anaplastic large‐cell lymphoma’ is unrelated to t(2;5)‐positive (ALK‐positive) lymphoma, and that it represents a morphological pattern occasionally encountered among diffuse large B‐cell lymphomas. By the same reasoning, most tumours diagnosed as ‘ALK‐negative anaplastic large‐cell lymphoma of T‐cell or null phenotype’ probably belong to the spectrum of peripheral T‐cell lymphomas.
Ex vivo expansion of mature human neutrophils with normal functions from purified peripheral blood CD34+ haematopoietic progenitor cells
Authors: Hino Masayuki , Suzuki Kenichi , Yamane Takahisa , Sakai Nobuaki , Kubota Hiroshi , Koh Ki Ryang , Ohta Kensuke , Hato Fumihiko , Kitagawa Seiichi , Tatsumi Noriyuki
Published: March 2003

Purified CD34+ haematopoietic progenitor cells were cultivated with stem cell factor, interleukin 3 (IL‐3), granulocyte–macrophage colony‐stimulating factor (GM‐CSF) and granulocyte CSF (G‐CSF) for 7 d, and thereafter non‐adherent cells were divided into two groups. Cells in one group (group A) were further cultivated for 7 d with four cytokines, and cells in the other group (group B) were further cultivated for 7 d with G‐CSF alone. On day 14, 220‐fold and 130‐fold increases in the numbers of non‐adherent cells were achieved for groups A and B respectively. These cell preparations contained 65% granulocytes for group A and 95% granulocytes for group B. These cells gained the ability to respond effectively with chemotaxis, phagocytosis and superoxide (O2−) release. Cells in group B were appropriately primed by G‐CSF, GM‐CSF, tumour necrosis factor α and IL‐1β for enhanced release of

. The responsiveness of these cells was identical to that of peripheral blood neutrophils, indicating that cells in group B may be in the resting state. In contrast, cells in group A were not primed by these cytokines for enhanced release of O2− and released a large amount of O2− spontaneously, indicating that cells in group A may be in the activated state. These findings indicate that mature neutrophils with normal functions were expanded ex vivo in group B and suggest that these cells could be used for possible autologous neutrophil transfusion to prevent bacterial infections during severe neutropenia after cytotoxic chemotherapy.
Plasminogen activator inhibitor 2 and urokinase‐type plasminogen activator in plasma and leucocytes in patients with severe sepsis
Authors: Robbie L. A. , Dummer S. , Booth N. A. , Adey G. D. , Bennett B.
Published: March 2003

Proteins influencing plasminogen activation to plasmin, namely plasminogen activators tissue‐type plasminogen activator (t‐PA) and urokinase‐type plasminogen activator (u‐PA) and their principal inhibitors, plasminogen activator inhibitor 1 (PAI‐1) and PAI‐2, were measured in the plasma, the polymorph and mononuclear cell fractions taken from patients with major sepsis who were entering a general intensive care unit. The purpose of this study was to elucidate the factors favouring the persistence of fibrin in the microvasculature and thus contributing to multiple organ failure. Levels of u‐PA antigen in plasma rose in sepsis and u‐PA activity, not detectable in normal plasma, appeared. Levels of u‐PA antigen in the cell fractions fell concomitantly. t‐PA antigen in plasma and in the mononuclear cell fraction rose in sepsis, but t‐PA activity was not detectable. Plasma PAI‐1 antigen levels were strikingly raised in sepsis, presumably accounting for the complete neutralization of t‐PA activity. PAI‐2 antigen, not normally detected in plasma, appeared in the plasma of some patients, whereas it disappeared from the cellular fractions. Appearance of PAI‐2 in plasma was associated with non‐survival of the patient. The observations indicate that all the agents involved in plasminogen activation are released into the plasma in major sepsis. The levels of PAI‐1 reached were quantitatively sufficient to suppress all activity of the released t‐PA, but the inhibitors did not prevent expression of u‐PA activity in the circulation. Circulating active u‐PA and PAI‐2 in the plasma of patients with severe sepsis may represent material originating from leucocytes. Leucocyte release of these agents within fibrin deposits may influence the persistence of fibrin and thus the development of multiple organ failure.
Recombinant fragment of von Willebrand factor AR545C inhibits platelet binding to thrombin and platelet adhesion to thrombin‐treated endothelial cells
Authors: Dardik Rima , Varon David , Eskaraev Regina , Tamarin Ilya , Inbal Aida
Published: March 2003

Activated, but not resting, platelets are capable of adhering to intact endothelial cells (ECs). We evaluated the effect of a recombinant von Willebrand factor (VWF) fragment AR545C, which inhibits glycoprotein Ib (GPIb)/VWF binding, on platelet adhesion to human ECs under static or flow conditions. Incubation of resting platelets with intact endothelium under flow conditions (350/s) resulted in minimal platelet adhesion. The adhesion was enhanced two‐ to threefold after either platelet activation by thrombin receptor agonist peptide (TRAP) or EC pretreatment with thrombin. The enhancing effect of thrombin was abolished by addition of either hirudin (10 u/ml) or PGE1 (1 µg/ml). Preincubation
of resting platelets with increasing concentrations of AR545C under static or flow conditions resulted in a dose‐dependent inhibition of thrombin‐induced enhanced adhesion to ECs. AR545C (0·3 µm) completely abolished the effect of thrombin, reducing platelet adhesion to the control level observed with non‐treated ECs. In contrast, the same concentration of AR545C had no effect on the adhesion of TRAP‐activated platelets to ECs. AR545C also inhibited thrombin‐induced platelet aggregation and binding in a dose‐dependent manner. In addition, 0·3 µm of AR545C reduced thrombin‐induced serotonin release by 57%, whereas monoclonal antibody AN51, which inhibits ristocetin‐induced platelet aggregation, had no effect on either thrombin‐induced platelet aggregation or binding or on serotonin release. Similarly, AR545C had no effect on TRAP‐induced serotonin release. These findings suggest that (i) AR545C inhibits platelet activation mediated by thrombin and this inhibition occurs through blocking the high‐affinity thrombin binding sites on the GPIb/IX complex and (ii) AR545C has no effect on the moderate affinity thrombin receptor (seven transmembrane domain thrombin receptor; STDR).
HCV‐associated thrombocytopenia: clinical characteristics and platelet response after recombinant α2b‐interferon therapy
Authors: García‐Suárez Julio , Burgaleta Carmen , Hernanz Nuria , Albarran Fernando , Tobaruela Pedro , Alvarez‐Mon Melchor
Published: March 2003

Hepatitis C virus (HCV) has been proposed as a possible causative agent of chronic thrombocytopenia. We investigated HCV infection in a series of 51 unselected Spanish patients with chronic acquired thrombocytopenia. Anti‐HCV and HCV viraemia were detected in 13/51 (22·5%) of cases; this prevalence was particularly significant when compared with HCV seropositivity in age‐matched controls (0·4%). Anti‐HCV‐positive patients, four men and nine women with a median age of 59·3 years (range 36–72), had a mean platelet count of 55·8 × 109/l (range 12–96). Only one of our HCV‐positive thrombocytopenic patients had hypersplenism. Platelet‐associated IgG (PAIgG) was negative, as measured by immunofluorescent flow cytometric analysis in 11/13 HCV‐positive thrombocytopenic patients. Thus, thrombocytopenia in our HCV‐positive patients appeared to be non‐autoimmune mediated. In six patients, a trial of recombinant α2b‐interferon (IFN‐α) given at a dose of 3 MU three times per week for 6–24 months gave a durable (> 1 year) and significant increase in platelet count in all six patients. The maximum increase occurred after 6 months of IFN‐α therapy. In conclusion, the ability of IFN‐α to increase platelet counts in HCV‐positive thrombocytopenic patients supports mechanisms involving a direct role for HCV inhibiting platelet production.
Post‐transplantation lymphoproliferative disease of natural killer cell lineage: a clinicopathological and molecular analysis
Authors: Kwong Y. L. , Lam C. C. K. , Chan T. M.
Published: March 2003

Post‐transplantation lymphoproliferative disorders (PTLD) occur after solid organ and bone marrow transplantation. They are predominantly of B‐cell and occasionally of T‐cell lineage. We report a case of PTLD of natural killer (NK) cell lineage. A renal allograft recipient developed progressive pancytopenia 1 year after transplantation. Serial bone marrow biopsies showed an increasing infiltration by large granular lymphoid cells. A subsequent leukaemic phase also developed with systemic infiltration of other organs. Immunophenotyping showed that these cells were CD2+, CD3−, CD3ε+, CD56+, CD94+, CD158a− and CD158b−. In situ hybridization showed Epstein‐Barr virus (EBV) infection of the neoplastic cells. Genotypical analysis showed the T‐cell receptor gene in germline configuration and clonal EBV episomal integration. The overall features were consistent with NK cell lymphoma/leukaemia. The patient did not respond to cessation of immunosuppression or anti‐EBV treatment. Combination chemotherapy was given, but the patient died ultimately of disseminated fungal infection. In conclusion, we have demonstrated that NK cell lymphoma is another rare type of PTLD that appears to be highly aggressive and therefore may require early chemotherapy to improve treatment outcome.
Monoclonal antibody‐mediated inhibition of the human HLA alloimmune response to platelet transfusion is antigen specific and independent of Fcγ receptor‐mediated immune suppression
Authors: Crow Andrew R. , Freedman John , Hannach Barbara , Lazarus Alan H.
Published: March 2003

Presensitization of donor platelets with allo‐specific immunoglobulin (Ig)G results in a diminished immune response against subsequent transfusions of platelets. To understand better the mechanism of how alloantibody presensitization results in a decreased alloimmune response, we have used murine monoclonal antibodies directed to polymorphic and non‐polymorphic regions of human leucocyte antigen (HLA) as well as platelet‐specific molecules. Here, we demonstrated that presensitization with anti‐human HLA class I antibodies, as well as β2‐microglobulin‐specific antibody, protected against alloantibody production to five subsequent untreated platelet challenges. Use of complement fixing, non‐fixing or F(ab′)2 fragments of HLA‐specific antibody also resulted in complete inhibition of alloantibody production. This protection was not seen when the platelets were presensitized with monoclonal antibodies to CD42a (GPIX), CD32 (low‐affinity IgG/Fcγ receptor) or murine IgG and was thus independent of B‐cell FcγRII‐mediated immune suppression. The inhibition observed was independent of HLA alloantigenic specificity as antibodies directed at the β2‐microglobulin portion of HLA class I were as effective as antibodies against any of the HLA‐α regions (either polymorphic or non‐polymorphic) of class I. This work demonstrates that monoclonal antibody‐mediated suppression of the human HLA alloimmune response to platelet transfusion is antigen specific and is independent of FcγRII‐mediated immune regulation, complement fixing or HLA alloantigenic specificity.
Stem cell transplantation from HLA‐matched related donor for Fanconi's anaemia: a retrospective review of the multicentric Italian experience on behalf of Associazione Italiana di Ematologia ed Oncologia Pediatrica (AIEOP)–Gruppo Italiano Trapianto di Midollo Osseo (GITMO)
Authors: Dufour Carlo , Rondelli Roberto , Locatelli Franco , Miano Maurizio , Di Girolamo Gabriele , Bacigalupo Andrea , Messina Chiara , Porta Fulvio , Balduzzi Adriana , Iorio Anna Paola , Buket Erer , Madon Enrico , Pession Andrea , Dini Giorgio , Di Bartolomeo Paolo
Published: March 2003

Twenty‐seven consecutive Italian patients with Fanconi's anaemia (FA) underwent stem cell transplantation (SCT) from an HLA‐matched related donor in 10 Italian centres of the Associazione Italiana Ematologia ed Oncologia Pediatrica (AIEOP), Gruppo Italiano di Trapianto di Midollo Osseo (GITMO). Twenty‐two patients (81·5%) were conditioned with low‐dose (median 20 mg/kg) cyclophosphamide (Cy) and thoraco‐abdominal or total body irradiation (median dose 500 cGy), five patients (18·5%) with high‐dose Cy (median 120 mg/kg). Graft‐vs.‐host disease (GVHD) prophylaxis was carried out with cyclosporin A in 26 cases; methotrexate (MTX) was added in eight cases. One patient received MTX alone. The median follow‐up was 36 months. Ninety‐two percent of patients (25 out of 27) engrafted, grade II and III acute GVHD occurred in 28% and 8% of patients, respectively, with chronic GVHD in 12·5%. Conditioning‐related toxicity was mild: 4% of patients had grade III mucositis, 7·4% had grade II haemorrhagic cystitis, 14·8% had grade III liver toxicity and 11·1% had grade III renal toxicity. Transplant‐related mortality at 12 months was 19·2%, survival at 36 months was 81·5%, with a median Karnofsky score of 100%. No late tumours occurred after a mean follow‐up of the survivors of 5 years. None of the studied variables significantly affected the survival, including conditioning regimen, acute GVHD and clinical non‐haematological phenotype. Among the studied variables, only conditioning regimens containing high‐dose Cy and the presence of genital abnormalities were significantly (P < 0·05) associated with an increased rate of acute GVHD. Our study demonstrates that the Italian FA patients undergoing SCT from an HLA‐matched related donor have a very good outcome. These patients, when compared with others of different ethnic origin who underwent allogeneic bone marrow transplantation, showed a less severe non‐haematological phenotype, raising the possibility that this milder phenotype may have, at least in part, contributed to the outcome. Our data may provide a useful tool for further studies aiming to correlate genotype with phenotype.
Somatostatin receptor scintigraphy useful in stage I–II Hodgkin's disease: more extended disease identified
Authors: Lugtenburg P. J. , Krenning E. P. , Valkema R. , Oei H. Y. , Lamberts S. W. J. , Eijkemans M. J. C. , Van Putten W. L. J. , Löwenberg B.
Published: March 2003

Somatostatin receptor (SS‐R) scintigraphy successfully  shows primary cancers and metastases in patients with a variety of SS‐R‐positive tumours. In vitro studies have shown that SS‐Rs are present in lymph nodes from patients with Hodgkin's disease (HD). We performed a prospective study in 126 newly diagnosed patients with HD and compared the results of SS‐R scintigraphy with conventional staging procedures, i.e. physical examination, computerized tomography (CT) scanning and other imaging techniques. We report positive scintigraphy in all patients. The lesion‐related sensitivity was 94% and varied from 98% for supradiaphragmatic lesions to 67% for infradiaphragmatic lesions. In comparison with CT scanning and ultrasonography, SS‐R scintigraphy provided superior results for the detection of Hodgkin's localizations above the diaphragm. In the intra‐abdominal region, the CT scan was more sensitive than the SS‐R scan. A false‐positive scan was rarely seen. In stages I and II supradiaphragmatic HD patients, SS‐R scintigraphy detected more advanced disease in 18% (15 out of 83) of patients, resulting in an upstaging to stage III or IV, thus directly influencing patient management. Our data would support the validity of SS‐R scanning as a powerful imaging technique for the staging of patients with HD.
Detection of haemoglobin variants and inference of their functional properties using complete oxygen dissociation curve measurements
Authors: Imai K. , Tientadakul P. , Opartkiattikul N. , Luenee P. , Winichagoon P. , Svasti J. , Fucharoen S.
Published: March 2003

Complete oxygen dissociation curves for red cell suspensions of three haemoglobinopathies, namely haemoglobin (Hb) H, Hb Köln and Hb Tak/β thalassaemia diseases, were measured using automatic recording methods. These curves were left‐shifted compared with the normal red cell curve and showed a biphasic shape as a result of co‐existence of the high and normal affinity haemoglobin components. Computer‐assisted simulation of these biphasic curves enabled us to infer the curves for the pure abnormal haemoglobins and their fraction in the total haemoglobin of the red cell. The inferred values of fraction agreed with those determined by haemoglobin type analysis or the literature values. The curve for Hb Köln red cells deviated from the normal red cell curve in the whole range of oxygen saturation, whereas the curve for Hb H was close to the normal curve at the middle and upper portions. This difference in deviation was ascribed to a possible interaction between Hb Köln and Hb A through subunit exchange, and its absence between Hb H and Hb A. The present results indicate that measurement of the complete oxygen dissociation curve is important for the detection of non‐interacting variants such as Hb H and is useful for inferring the functional properties of haemoglobin components that are not easily isolated.
Induction of the monocytic differentiation of myeloid leukaemia cells by cotylenin A, a plant growth regulator
Authors: Yamamoto‐Yamaguchi Yuri , Yamada Kazunari , Ishii Yuki , Asahi Ken‐ichi , Tomoyasu Shigeru , Honma Yoshio
Published: March 2003

Regulators that play an important role in the differentiation and development of plants or invertebrates may also affect the differentiation of human leukaemia cells through a common signal transduction system, and might be clinically useful for treating acute myeloid leukaemia. Cotylenin A has been isolated as a plant growth regulator. We examined the effects of cotylenin A on the differentiation of several myelogenous leukaemia cells, and found that cotylenin A is a potent and novel inducer of the monocytic differentiation of human myeloid leukaemia cells. Cotylenin A induced the functional and morphological differentiation of myeloblastic and promyelocytic leukaemia cells, but did not effectively induce the differentiation of monocytoid leukaemia cells. Cotylenin A‐induced differentiation was not affected by several inhibitors of signal transduction, suggesting that this inducer exhibits a unique mode of action.
Interleukin 4 content in chronic lymphocytic leukaemia (CLL) B cells and blood CD8+ T cells from B‐CLL patients: impact on clonal B‐cell apoptosis
Authors: Kay Neil E. , Han Lei , Bone Nancy , Williams Gary
Published: March 2003

B‐chronic lymphocytic leukaemia (CLL) clonal B cells are characterized by resistance to apoptosis. We evaluated clonal B cells and blood T cells for interleukin 4 (IL‐4) content as IL‐4 is able to increase CLL cell resistance to apoptosis. The content of IL‐4 in CD8+ T cells of CLL patients (n = 9) ranged from 37% to 63% of the total CD8+ T cells (mean level of 49% ± 3·4) compared with a range of 5–10% for control CD8+ T cells. Clonal B cells positive for cytoplasmic IL‐4 ranged from 1% to 97% (mean value 57·8 ± 6·9%). CD8+ T cells and clonal B cells secreted detectable levels of IL‐4, but only clonal CLL B cells (n = 4) secreted IL−4 in association with increasing cell numbers. Fludarabine (F‐ara‐AMP, 0·1–100 μmol/ml) was able to downregulate the IL‐4 content of CD8+ T cells, but not clonal B‐cell IL‐4. Culture supernatant from CLL CD8+ T cells decreased the spontaneous apoptotic rate of clonal B cells that was reversed with anti‐IL‐4 and soluble IL‐4 receptor. These findings show that IL‐4 is present in the microenvironment of B‐CLL. In addition, use of agents that can interfere with IL‐4 presentation to clonal B cells can be effective in increasing clonal B‐cell apoptosis.
Polymorphonuclear leucocytes mediate endogenous thrombus lysis via a u‐PA‐dependent mechanism
Authors: Moir E. , Booth N. A. , Bennett B. , Robbie L. A.
Published: March 2003

Many human thrombi lyse spontaneously without the administration of lytic drugs and cause no clinical symptoms. The mechanisms by which this occurs are incompletely understood. We found that model thrombi prepared from whole human blood in a Chandler loop also exhibited significant spontaneous lysis. Lysis was inhibited by chemical protease inhibitors, consistent with proteolysis resulting primarily from serine proteases, with a small contribution from matrix metalloproteinases. Whole blood was fractionated into platelet‐rich plasma and cell populations. Significant spontaneous lysis was observed in platelet‐rich thrombi enriched with polymorphonuclear leucocytes (PMNs), whereas mononuclear cells (MCs) and erythrocytes did not contribute to lysis. Incorporation of antibodies to urokinase (u‐PA) and its receptor u‐PAR neutralized a large proportion of the activity. Incubation of plasma with PMNs generated free u‐PA activity, which was also detectable in model thrombi and in vivo human thrombi. Purified neutrophils, free of eosinophils, generated activity identical to PMNs. Smaller contributions to lysis by tissue‐type plasminogen activator (t‐PA), elastase and cathepsin G were also identified. These findings suggest a major role for circulating PMNs in endogenous thrombus lysis.
Autotransplants for histologically transformed follicular non‐Hodgkin's lymphoma
Authors: Chen Christine I. , Crump Michael , Tsang Richard , Stewart A. Keith , Keating Armand
Published: March 2003

Histological transformation from a follicular non‐Hodgkin's lymphoma (NHL) to a higher grade lymphoma carries a poor prognosis despite treatment with aggressive anthracycline‐based chemotherapy. We retrospectively analysed 35 patients with histologically transformed NHL who underwent high‐dose therapy and autotransplantation at our centre. Patients up to 65 years old were eligible for autotransplant at the time of transformation or with subsequent relapses, provided that chemosensitivity to a salvage regimen could be demonstrated. All patients received high‐dose therapy [etoposide 60 mg/kg, melphalan 160 mg/m2 and fractionated total body irradiation (TBI) 12 Gy] followed by unpurged autologous bone marrow or blood stem cell rescue. Most patients (69%) had advanced stage disease (stages 3–4) at transformation and bone marrow involvement was common (49%). Twenty‐six (74%) patients were in partial remission (PR) and nine (26%) in complete remission (CR) at the time of transplant. Median duration from transformation to transplant was 10·9 months (range, 5·2 months−4·6 years). At a median follow up of 52 months after autotransplant, 19 (54%) patients had died. Causes of death were progressive lymphoma in nine patients (26%), treatment‐related mortality (TRM) in seven (20%) and myelodysplasia in three (8%). Only five patients in our cohort were > 60 years old, but all died as a result of treatment‐related causes (mostly pulmonary infections). Five‐year overall survival and progression‐free survival from time of transplant were 37% and 36% respectively. Using multivariate analysis of factors including gender, age, stage, extranodal disease, disease bulk, B symptoms, number of prior therapies, relapse status and CR/PR status at transplant, only advanced age significantly predicted for survival from autotransplant (P = 0·002). Our survival data are comparable to previous reports of autotransplantation for transformed NHL and suggest a benefit over standard chemotherapy alone in selected patients. However, our high TRM cautions the use of aggressive therapy, including TBI, in patients over 60 years old.
Decreased expression of transcription factor GATA‐2 in haematopoietic stem cells in patients with aplastic anaemia
Authors: Fujimaki Shin‐ichi , Harigae Hideo , Sugawara Tomohiro , Takasawa Naruhiko , Sasaki Takeshi , Kaku Mitsuo
Published: March 2003

Aplastic anaemia is characterized by reduced haematopoiesis resulting in pancytopenia. It has been speculated that there is an injury in haematopoietic stem cells in the bone marrow; however, the precise nature of the injury has not been elucidated. In this study, the levels of expression of mRNAs for three transcription factors, GATA‐2, SCL and AML1, which function in the early stages of haematopoiesis, were examined by quantitative polymerase chain reaction in patients with aplastic anaemia, idiopathic thrombocytopenic purpura (ITP) and normal subjects. Among these factors, expression of GATA‐2 mRNA in purified CD34‐positive cells was markedly decreased in aplastic anaemia compared with that in ITP and in normal subjects. The expression levels of SCL and AML1 mRNA in CD34‐positive cells in aplastic anaemia were not different from those in normal subjects. When the expression of GATA‐2 protein in CD34‐positive cells was examined by immunocytochemical analysis, the percentage of GATA‐2‐positive cells in aplastic anaemia was lower than that in normal subjects. These findings strongly suggest that there is an aberrant expression of transcription factors in stem cells in aplastic anaemia, which may be responsible for the development of the disease.
Patterns of failure with increasing intensification of induction chemotherapy for acute myeloid leukaemia
Authors: Matthews J. P. , Bishop J. F. , Young G. A. R. , Juneja S. K. , Lowenthal R. M. , Garson O. M. , Cobcroft R. G. , Dodds A. J. , Enno A. , Gillett E. A. , Hermann R. P. , Joshua D. E. , Ma D. D. , Szer J. , Taylor K. M. , Wolf M. , Bradstock K. F. ,   for the Australian Leukaemia Study Group
Published: March 2003

Patterns of failure were studied in two consecutive randomized trials of intensified induction therapy carried out by the Australian Leukaemia Study Group (ALSG) between 1984 and 1991 to determine the impact of dose intensification. Patients received standard dose cytarabine and daunorubicin (7‐3), 7‐3 plus etoposide (7‐3‐7) or 7‐3 plus high‐dose cytarabine (HIDAC‐3‐7) chemotherapy. Patients with FAB M3 morphology were excluded. Time to failure (TTF) was defined as the time from randomization to induction death or removal from study for non‐responders, or to relapse or death in complete response (CR) for complete responders. An estimated 86% of 470 de novo patients with acute myeloid leukaemia failed within 10 years of randomization, as a result of death in induction in 17% of the randomized patients, failure to achieve CR in a further 17%, relapse in 44% and death in CR in 8% of patients. An estimated 66% of patients failed as a result of refractory disease or relapse within that period (disease‐related failures). Multifactor analysis identified age and peripheral blast count as the most significant pretreatment factors associated with overall TTF. These factors, together with cytogenetics, were significantly associated with disease‐related failures. High‐dose cytarabine in induction significantly decreased the disease‐related failure rate as did allogeneic transplantation in first CR. The impact of high‐dose cytarabine did not depend on the cytogenetic risk group.
Relationship of p15 and p16 gene alterations to elevated dihydrofolate reductase in childhood acute lymphoblastic leukaemia
Authors: Shah Sanjay J. , Taub Jeffrey W. , Witt Teah L. , Pollock Brad H. , Ding Bee Ching , Moore Daniel S. , Amylon Michael , Pullen Jeanette , Ravindranath Yaddanapudi , Matherly Larry H.
Published: March 2003

The downstream effects of p15 and p16 gene deletions and loss of transcripts on dihydrofolate reductase (DHFR) were examined in 63 B‐precursor (BP) acute lymphoblastic leukaemia (ALL) samples. p15 and/or p16 gene deletions were seen in 6% and 8%, respectively, of BP‐ALL samples; however, losses of p15 and/or p16 transcripts were seen in 26 out of 63 (41%) samples. Loss of p15 transcripts (36·5%) exceeded that for p16 (17·5%). For the 26 BP‐ALLs that lacked p15 and/or p16 transcripts, only six (23%) exhibited low levels of DHFR by flow cytometry assay with Pt430, a fluorescent anti‐folate. Conversely, 18 out of 37 (49%) BP‐ALL samples with intact p15 and/or p16 genes and transcripts showed low levels of DHFR (P = 0·04). In p15‐ and p16‐null K562 cells transfected with a tetracycline‐inducible p15 cDNA construct, induction of p15 transcripts and protein was accompanied by decreased growth rates, decreased S‐phase fraction, decreased retinoblastoma protein phosphorylation, and markedly reduced levels of DHFR transcripts and protein. Collectively, our results suggest that losses of p15 and/or p16 gene expression result in elevated levels of DHFR in BP‐ALL in children. However, additional downstream factors undoubtedly also contribute to elevated levels of this enzyme target.