bluuebunny/crossref_metadata_2025 huggingface dataset:
Fibrin degradation product (FnDP) assays: analysis of standardization issues and target antigens in plasma
Authors: GAFFNEY P. J. , EDGELL T. , CREIGHTON‐KEMPSFORD L. J. , WHEELER S. , TARELLI E.
Published: March 2008

The in vivo formation of fibrin and its subsequent secondary fibrinolytic digestion yields a variety of crosslinked fibrin degradation products (XL/FnDP). One of these is known as D‐dimer and a variety of ELISA‐type commercial kits using monoclonal antibodies to D‐dimer have been developed. We have investigated the possibility of establishing a standard such that these various kits might indicate the same levels of D‐dimer in the same samples. We have concluded that because it seems that each individual monoclonal antibody to D‐dimer targets a unique and distinct epitope in the FnDP fraction the notion of introducing a standard D‐dimer which will respond uniformly to each D‐dimer monoclonal antibody is not feasible. Using various monoclonal and polyclonal anti‐ bodies in conjunction with gel exclusion chromatography we have examined human plasma derived from patients with disseminated intravascular coagulation (DIC) which contained high levels of fibrin degradation products (FnDP). The data suggested that the FnDP fraction in plasma contained mostly high molecular weight (>2xl06 daltons) crosslinked fragments which contain high levels of fibrino‐peptide A. Many of these crosslinked fragments crossreact with antibodies to D‐dimer because they each contain D‐dimer as a structural component. On the basis of this data, a novel sequence of events is proposed which may occur during the aggregation of fibrin in vivo.
Successful bone marrow transplantation for idiopathic hypereosinophilic syndrome
Authors: FOKUSHIMA TAKUYA , KURIYAMA KAZUTAKA , ITO HAJIME , MIYAZAKI YASUSHI , ARIMURA KOSEI , HATA TOMOKO , SAITOH MAMIKO , TOMONAGA MASAO
Published: March 2008

A 21‐year‐old man who had an increased number of eosinophils with morphological abnormalities, bone marrow fibrosis and multiple organ dysfunction failed to respond to methylprednisolone and hydroxyurea. He was diagnosed with hypereosinophilic syndrome (HES) probably due to myeloproliferative disorder, and underwent allogeneic bone marrow transplantation (allo‐BMT) from an HLA‐identical sibling. The engraftment was confirmed on day 21 after BMT, but the patient developed acute graft‐versus‐host disease (GVHD) with grade I veno‐occlusive disease, and transient increase of eosinophils of the donor type followed by chronic GVHD of the extensive type. These complications were eventually controlled with cyclosporin A. The patient survived free of disease for more than a year after BMT. Allo‐BMT seems to be a possible treatment of HES/MPD.
Expansion of CD8+CD57+ T cells after allogeneic BMT is related with a low incidence of relapse and with cytomegalovirus infection
Authors: DOLSTRA H. , PREIJERS F. , KEMENADE E. VAN DE WIEL‐VAN , SCHATTENBERG A. , GALAMA J. , WITTE T. DE
Published: March 2008

Summary. Peripheral blood lymphocytes of 46 recipients of lymphocyte‐depleted bone marrow allografts were pheno‐typically analysed over a period of 1 year. We investigated the repopulation of lymphocyte subpopulations and their relation with clinical parameters such as graft‐versus‐host disease (GVHD), graft‐versus‐leukaemia and cytomegalovirus (CMV) infection. The number of repopulated T cells varied strongly between the blood samples of the recipients. In 45% of the recipients the number of T cells recovered to or above normal levels within 3 months after bone marrow transplantation (BMT), whereas the other recipients remained below normal up to 1 year after BMT. In recipients with a high repopulation, the CD8+ T‐cell subset contributed more to this high repopulation than the CD4+ T‐cell subset. We showed that the majority of T cells of these recipients expressed the a/3 T‐cell receptor, CD8, CD57 and CDllb. HLA‐DR was also highly expressed reflecting the activation stage of T cells in these recipients. BMT recipients with a high repopulation of CD8+ T cells showed a lower incidence of leukaemic relapse than recipients with a low repopulation. The 3‐year probability of relapse was 19% versus 64% (P=O03), respectively. The relative high number of CD8+ T cells at 3 months after BMT was not associated with the incidence of GVHD. In contrast, occurrence of CMV infection after BMT was significantly higher in these recipients. Our results indicate that CD8+ T cells, predominantly CD57+, of BMT recipients with an expansion of these cells represent an in vivo activated cell population. This CD8+ T‐cell population may consist partially of cytotoxic cells with anti‐leukaemic activity as suggested by a low relapse rate. The signal for the strong expansion of these CD8+CD57+ T cells after BMT is still unclear, but association with CMV infection suggests that viral antigens are involved.
The determination of spontaneous megakaryocyte colony formation is an unequivocal test for discrimination between essential thrombocythaemia and reactive thrombocytosis
Authors: ROLOVIC ZORAN , BASARA NADEZDA , GOTIC MIRJANA , SEFER DIJANA , BOGDANOVIC ANDRIJA
Published: March 2008

Summary. Spontaneous colony formation from bone marrow megakaryocyte progenitors (BMsCFU‐Mk) was studied in 24 patients with essential thrombocythaemia (ET), 20 patients with reactive thrombocytosis (RT), 20 patients with polycthaemia rubra vera with thrombocytosis (PRVtr), 16 patients with chronic myeloid leukaemia with thrombocytosis (CMLtr) and 18 normal control subjects (C).

The culture medium which was used in the methylcellu‐lose assay in vitro contained 30% of plasma from a single patient with hereditary haemochromatosis. Remarkable BMsCFU‐Mk growth was recorded in all patients with ET but in none with RT or in C. BMs‐CFU‐Mk were present in 11/20 patients with PRVtr and 7/16 patients with CMLtr.

Spontaneous bone marrow erythroid progenitors (BMsBFU‐E) were also determined in these patients. BMsBFU‐E were found in 21/24 patients with ET and none in the patients with RT and C. All patients with PRVtr and one patient with CMLtr showed BMsBFU‐E.

We conclude that our implementation of the in vitro methylcellulose assay allows the BMsCFU‐Mk to be used as an unequivocal test for discrimination between ET and RT which has not been shown in previously published studies.

In addition, we present evidence that in 10 patients BMsCFU‐Mk and/or BMsBFU‐E growth in the test persisted after long‐lasting haematological remission.
Sustained increase in haemoglobin and RBC following long‐term administration of recombinant human erythropoietin to patients with homozygous beta‐thalassaemia
Authors: RACHMILEWITZ E. A. , AKER M. , PERRY D. , DOVER G.
Published: March 2008

Summary.Recombinant human erythropoietin (rHuEPO) was given subcutaneously three times per week in an escalating dose from 500u/kg to 950u/kg together with ferrous fumarate 305 mg and folic acid 5mg/d, to 10 patients from four unrelated Arab families with homozygous /3‐thaIassaemia. Six splenectomized patients showed a mean (istandard error) increase in haemoglobin from 7‐1 – 0‐1 to 9‐3 – 0‐1 g/dl (P = 0‐0001), in RBC from 4‐0 to 5‐0 × 1012/1 (P = 0‐0001) and in nucleated RBC from 32 – 7 × 1010/1 to 82–6 × 1010/1 while receiving 750u/kg three times per week which persisted for 4‐11 months. In two patients there was no need for further blood transfusions. In three out of four unsplenectomized patients there were no changes in Hb and RBC despite dose escalation. There were no significant changes in MVC, MCH and reticulocyte count, serum bilirubin, LDH, malonyldialdehyde (MDA) and vitamin E levels. After 13 weeks of rHuEPO there was a mean increase in the percentage of F cells from 31 –10% to 86 –6% (P< 0‐003) in three splenectomized patients and in one unsplenectomized patient from 56‐4% to 80% without changes in the levels of Hb F. Globin chain synthesis ratios did not change in four responding patients. Mean serum iron and transferrin saturation index did not change, whereas mean serum ferritin increased from 299–45/ng/I to 480–20/ig/l (P< 0‐001). In seven responding patients an accelerated linear growth was indicated by positive changes in height standard deviation score for chronological age. Side‐effects were minimal throughout the treatment period.
Rearrangement of the immunoglobulin heavy chain gene in juvenile chronic myeloid leukaemia
Authors: MARK ZIVA , TOREN AMOS , AMARIGLIO NINETTE , VONSOVER AMI , RECHAVI GIDEON , BROK‐SIMONI FRIDA
Published: March 2008

Summary. Five patients with clinical and laboratory features typical for juvenile chronic myeloid leukaemia (JCML) are presented. Rearrangement of the j joining region of the immunoglobulin heavy chain (Jh) was demonstrated in three children out of five analysed. As no Vh to Dhjh nor kappa light chain rearrangements were demonstrated, it is reasonable to speculate that the transforming event of the stem cell happened at the stage when Dh to Jh rearrangement took place. As the monocytic lineage is prominent in JCML, it is suggested that the transforming event happens in a unique stem cell with intermediate differentiation towards the myelomonocytic as well as the B‐lymphatic lineage. This stem cell, which is present at a certain stage of embryogen‐esis, disappears later. Such an early‘hybrid’cell is sometimes involved in leukaemias of early infancy, and may be the transformed cell in some cases of infantile leukaemia.
Abnormal TCR V/3 repertoire in patients with chronic myeloid leukaemia
Authors: COLEMAN S. , PERERA P. , FISHER J. , HOY T. , BURNETT A. K. , LIM S. H.
Published: March 2008

Summary. We have used 25 sets of oligonucleotide primers specific for the 24 known major human T‐cell receptor (TCR) V/? families in polymerase chain reactions to analyse the T‐cell repertoire of the peripheral blood in seven patients with chronic myeloid leukaemia (CML). In contrast to normal healthy individuals, all seven patients exhibited variable degrees of TCR V/3‐specific T‐cell deletion, ranging from two to eight of the 24 major families. T cells bearing V/3 17 and 8 were most commonly deleted. These results suggest a superantigen effect associated with CML. The patterns of deletion did not appear to correlate with either of the two bcr‐abl transcripts. The reason and aetiological agent responsible for the T‐cell deletion remain speculative. Further work is ongoing to characterize this phenomenon in animal models and patients with CML.
GST‐7T and P‐170 co‐expression in multiple myeloma
Authors: PETRINI MARIO , SIMONE DANIELA DI , FAVATI ADRIANA , MATTII LETIZIA , VALENTINI PAOLA , GRASSI BRUNO
Published: March 2008

Summary. Bone marrow samples from 40 patients affected by multiple myeloma either treated or untreated were examined for expression of glutathione‐S‐transferase n (GST‐TT), P‐glycoprotein and the protein product of ras oncogenes family, p‐21, on plasma cells, by immuno‐cytochemical detection. 72% of evaluated samples were positive for P‐170 and 82% for GST‐7T without any correlation with clinical or prognostic parameters. A significant relationship between GST‐7T expression and P‐l 70 positivity was found and co‐expression was observed in 91% of evaluated samples.

Expression of P‐170 and GST‐TT was found both in treated and untreated patients. However, patients evaluated before and after therapy showed an increase in the percentage of plasma cells positive for GST‐7T or P‐170 or both. Expression of p‐21 was not associated with these mechanisms of drug resistance.

These data suggest that different resistance mechanisms are present in multiple myeloma.
Characterization of protein disulphide isomerase released from activated platelets
Authors: CHEN Km , DETWILER THOMAS C. , ESSEX DAVID W.
Published: March 2008

Summary. Protein disulphide isomerase (PDI) activity is released by activated platelets. In this study, PDI was purified from platelets and found to have an apparent mass, pi and N‐terminal sequence similar to those for other human PDIs. Rabbit antibodies were generated and used to establish that, on activation, platelets release a protein immunologically identical to PDI in platelets. Approximately 10% of total platelet PDI was released by thrombin and 20% by calcium ionophore. The antibody was used to demonstrate PDI on the external surface of platelets by electron microscopy. Flow cytometry was used to demonstrate that upon activation of platelets with ionophore PDI was released by vesiculation. Since platelets are present and become activated at sites of vascular injury, platelet PDI may play a role in the various haemostatic and tissue remodelling processes in which platelets are involved.
The effect of endogenous oestradiol levels on protein S concentration during a menstrual cycle and after GnRH analogues and gonadotropin therapy
Authors: FALKON L. , GARI M. , MORA J. L. GARCIA , CALAF J. , ESPINOSA J. RODRIGUEZ , OLIVER A. , FONTCUBERTA J.
Published: March 2008

Summary. We studied two groups of females to investigate the effect of endogenous oestradiol levels on total and free protein S (tPS, fPS) plasma concentrations. One group (group I) consisted of 12 healthy volunteers who were studied throughout one menstrual cycle; the other group (group II) consisted of 16 young women who were treated with GnRH analogues and gonadotropins before undergoing in vitro fertilization. Neither tPS nor fPS varied significantly with respect to the physiological changes of oestradiol or to the very low and high levels of oestradiol, achieved after GnRH analogues suppression and gonadotropin stimulation. These results indicate that endogenous oestradiol does not affect PS concentration.
Macrophage colony‐stimulating factor‐producing malignant histiocytosis
Authors: OZAKI SHUJI , MATSUSHITA TAKAYA , IDE MAKOTO , OZAKI KEIJI , SANO TOSHIAKI , KOSAKA MASAAKI , SAITO SHIRO
Published: March 2008

Summary. We describe a patient with malignant histiocytosis who had a prominent neutrophilia uncommonly observed in this disease. Serum concentration of macrophage colony‐stimulating factor (M‐CSF) was markedly elevated, and correlated with clinical symptoms and neutrophil counts during clinical course. Moreover, M‐CSF was detected in the cytoplasm of malignant histiocytes of the lymph node and bone marrow by immunohistochemical staining. These results indicate that the neutrophilia observed in this patient was caused by M‐CSF produced by the malignant histiocytes.
CMV viraemia demonstrated in the serum of a patient with cytomegalovirus pneumonia
Authors: WATANABE NAOKI , TAKEDA MITSUAKI , ISHIGAKI SEISHI , TSUJI NAOKI , SAKAMAKI SUMIO , KATO JUNJI , KOHDA KYUHEI , MORI YOSHIHIRO , NIITSII YOSHIRO
Published: March 2008

Summary. We attempted to demonstrate the expression of cytomegalovirus (CMV) particles in the serum of an acute lymphocytic leukaemia patient with CMV pneumonia. The serum sample was applied to an affinity column coupled with human monoclonal antibody C2 3 which recognizes the envelope glycoproteins of CMV virus and neutralizes the viral activity. The DNA obtained from each fraction was amplified by double polymerase chain reaction (PCR) and analysed by gel electrophoresis. Bands were clearly observed in the eluted fraction. These results strongly suggest that CMV particles exist in the sera of patients with CMV pneumonia.
Clinicopathological correlates of CD56 expression in multiple myeloma: a unique entity?
Authors: MATHEW PAUL , AHMANN GREGORY J. , WITZIG THOMAS E. , ROCHE PATRICK C. , KYLE ROBERT A. , GREIPP PHILIP R.
Published: March 2008

Summary. Prior studies have suggested that loss of plasma cell CD 5 6 expression in multiple myeloma defines a unique patient subset and that CD56 expression reliably discriminates between monoclonal gammopathy of undetermined significance (MGUS) and multiple myeloma (MM). We conducted a study of 68 untreated patients with MM from a single institution to define the clinicopathological correlates of CD56 expression. We find CD56 expression in 55% of MM. Lack of CD56 expression does not define a unique clinicopathological or prognostic entity in MM. Strong CD 5 6 expression can also be found in MGUS and does not help to distinguish from MM.
Immunohistochemical characterization of intact stromal layers in long‐term cultures of human bone marrow
Authors: WILKINs B. S. , JONES D. B.
Published: March 2008

Summary. We have performed an immunohistochemical study of intact adherent layers of human long‐term bone marrow cultures (hLTBMC) in order to characterize the cell types present. Our panel of antibodies was selected to demonstrate various mesenchymal and haemopoietically derived cell types and to assess the presence of molecules of potential importance as adhesive ligands between haemopoietic cells and stroma.

Subpopulations of fibroblasts and macrophages were identified which differed in immunophenotype. We were able to demonstrate modulation of fibroblast and extracellular matrix immunophenotypes between 2 and 12 weeks in culture. Stromal cells and matrix expressed a wide variety of antigens of potential importance in haemopoietic cell adhesion, but no ICAM‐1, 2 or 3 could be demonstrated to correspond to the strong LFA‐1 expression seen in haemopoietic precursor cells.

No localization of antigen expression by stromal elements was found to account for the formation of haemopoietic foci at particular sites. However, granulocyte‐predominant foci preferentially occupied the interstices and margins of structures which appeared to be vascular arrays.
Transplanted sickle‐cell disease patients with autologous bone marrow recovery after graft failure develop increased levels of fetal haemoglobin which corrects disease severity
Authors: FERSTER A. , CORAZZA F. , VERTONGEN F. , BUJAN W. , DEVALCK C. , FONDU P. , COCHAUX P. , LAMBERMONT M. , KHALADJI Z. , SARIBAN E.
Published: March 2008

Summary. Bone marrow transplantation (BMT) is the only curative therapy for sickle‐cell disease (SCD), but is not devoid of failure risk. Nine patients with severe SCD were grafted in our institution between 1988 and 1993. Six patients successfully engrafted, but three failed to engraft and had delayed autologous recovery.

All patients had, prior to BMT, low levels of fetal haemoglobin (HbF ≤3.5%). No change in HbF occurred in successfully grafted patients. In the three patients with graft failure HbF increased and remained persistently present at a high level (≥22%) 14 months, 16 months and 39 months post BMT, although two of the three patients were homozygous for either the Benin or the Central African Republic haplotype, a characteristic associated with low HbF level. Of interest, these three previously severely affected patients remain free of vaso‐occlusive events.

The mechanism responsible for the expression of high levels of HbF in our three patients with graft failure is not understood, but it protects them from the recurrence of severe vaso‐occlusive crises.
A specific chromosome abnormality of t(4;12)(qll‐12;pl3) in CD7+ acute leukaemia
Authors: HARADA HIRONORT , ASOU HIROYA , KYO TAIICHL , ASAOKU HIDEKI , IWATO KOJI , DOHY HIROO , ODA KENJI , HARADA YUKA , KITA KENKICHI , KAMADA NANAO
Published: March 2008

Summary. Three cases of acute leukaemia with t(4;12) (qll‐12;pl3) karyotypic abnormalities were analysed. They had the following common clinical and biological characteristics: (1) dysplasia of three haemopoietic lineages; (2) absent or low myeloperoxidase activity; and (3) retention of platelets in the peripheral blood and megakaryocytes in the bone marrow. There were increased numbers of basophils in the bone marrow and peripheral blood in two of the cases. In all, the blast cells displayed the unique immunophenotype CD7+CD13+CD34+HLA‐DR+. The blasts analysed in one case expressed c‐kit on the membrane surface. These findings suggest that the t(4:12) (qll‐12:pl3) abnormality is associated with a particular type of acute leukaemia, one in which the morphology and immunophenotype suggest that the translocation may have occurred at an early stage of haemopoiesis.
Transduction of MDR1 into human and mouse haemopoietic progenitor cells: use of rhodamine (Rhl23) to determine transduction frequency and in vivo selection
Authors: HEGEWISCH‐BECKER SDSANNA , HANANIA ELTE G. , FU SIQUING , KORBLING MARTIN , DEISSEROTII ALBERT B. , ANDREEFF MICHAEL
Published: March 2008

Summary. The MDR1 gene product P‐glycoprotein (P‐gp) extrudes several anticancer drugs including taxol and fluorescent dyes such as rhodamine (Rhl23). Modulation of the level of P‐gp expression has the potential of overcoming multidrug resistance. One possible approach is the retroviral transfer of the human MDR1 gene into murine and human bone marrow (BM) progenitor cells. The rationale for this approach is increased chemoprotection, which allows chemotherapy of a greater level of intensity to be delivered. In this study, flow cytometric measurement of Rhl23 extrusion was used to test P‐gp function in human and mouse haemopoietic progenitor cells, which had been transduced with a virus containing the human MDR1 transcription unit. Human CD34+ selected cells were analysed immediately following transduction. In two successive experiments MDR1 cDNA transduction resulted in a 7% and 11% increase of P‐gp expressing Rhl23 dull cells. To monitor transduction efficiency over time as well as the possibility of in vivo selection of drug‐resistant BM cells in mice treated with increasing numbers of taxol cycles, the assay was also successfully applied to peripheral blood lymphocytes of mice transplanted with MDR1 transduced BM cells, demonstrating increased Rhl23 efflux in transduced cells. Analysis of another fluorescence assay using fluorescein di‐beta galactopyranoside as a substrate for β‐galactosidase in cells transduced with a MDRl:d‐gal double transcription unit was hindered by background staining due to endogenous β‐gal activity. We conclude that the Rhl23 efflux assay is a sensitive method to monitor P‐gp function in MDR1 cDNA transduced cells, and may be used to enrich transduced cells via flow cytometric cell sorting for Rhl23 dull cells.
Severe haemophilia A in a female resulting from two de novo factor VIII mutations
Authors: WINDSOR SHARON , LYNG ANN , TAYLOR SHERRYL A. M. , EWENSTEIN BRUCE M. , NEUFELD ELLIS J. , LILLICRAP DAVID
Published: March 2008

Summary. A 2‐year‐old girl is described with severe haemophilia A (factor VIII: C <0‐01 units/ml). Both of her parents were phenotypically normal. Cytogenetic analysis on the proband demonstrated an interstitial X chromosome deletion encompassing Xq26‐q28. Molecular studies with several polymorphic markers close to and within the factor VIII gene showed that the proband had inherited only the paternal factor VIII gene, indicating that the X chromosome deletion had occurred de novo in the maternal germ line. Further study of the factor VIII gene inherited by the proband from her father showed the presence of a de novo gene inversion mutation (a type 1, distal pattern inversion). Neither parent showed any evidence of the factor VIII inversion in their somatic DNA. The severe haemophilia A documented in this girl is therefore the result of two de novo mutations affecting the factor VIII gene, a maternally derived X chromosome deletion and a paternal factor VIII inversion mutation.
Differential effects of recombinant human interleukin‐13 on the in vitro growth of human haemopoietic progenitor cells
Authors: Xi XIAODONG , SCHLEGEL NICOLE , CAEN JACQUES P. , MINTY ADRIAN , FOURNIER SYLVIE , CAPUT DANIEL , FERRARA PASCUAL , HAN ZHONG C.
Published: March 2008

Summary. Effects of recombinant human interleukin (IL)‐13 on in vitro haemopoiesis from non‐adherent mononuclear cells (NAMC) or highly enriched CD34+ cells of human cord blood (CB) were studied. IL‐13 significantly increased megakaryocyte (MK) colony formation from either NAMC or CD34+ cells cultured in a plasma clot system supplemented with aplastic anaemia serum (AAS) and phytohaemag‐glutinin‐stimulated human peripheral blood leucocyte‐conditioned medium (PHA‐LCM) in a dose‐dependent manner. Experiments using a modified plasma clot culture, in which normal AB serum and various cytokines were added to replace AAS and PHA‐LCM, demonstrated an increased MK colony number in the presence of IL‐13, especially in combination with IL‐3. However, IL‐13 had no stimulatory effect, but rather a slight inhibitory effect in some cases on granulocyte‐macrophage (GM) colony formation in both plasma clot cultures. Furthermore, the growth of GM progenitor cells in a methylcellulose culture system in the presence of IL‐3, GM‐CSF, Epo, G‐CSF or in combination was significantly inhibited by the addition of IL‐13. On the other hand, high concentrations (lOOng/ml) of IL‐13 were needed to cause a slight inhibition on the growth of BFU‐E‐derived colonies under the same methylcellulose culture. These results indicate that IL‐13, alone and synergistically with the effect of IL‐3, promotes MK colony formation, but it inhibits the growth of GM and erythroid progenitor cells in vitro.
A linkage between hereditary hyperferritinaemia not related to iron overload and autosomal dominant congenital cataract
Authors: GIRELLI DOMENTCO , OUVIERI OLIVIERO , FRANCESCHI LUCIA DE , CORROCHER ROBERTO , BERGAMASCHI GAETANO , CAZZOLA MARK
Published: March 2008

Summary. The only genetic disorder with elevated serum ferritin levels so far described is hereditary HLA‐related haemochromatosis. On the other hand, hereditary cataract is both genotypically as well as phenotypically heterogenous, and no specific locus or any useful marker has been yet identified. We studied two Italian families in whom a combination of elevated serum ferritin not related to iron overload and congenital nuclear cataract is transmitted as an autosomal dominant trait. Affected individuals have normal serum iron and transferrin saturation, but high serum ferritin. Red cell counts are normal and venesection therapy rapidly produces iron‐deficiency anaemia.

This genetic disorder, which is characterized by hyperferritinaemia, differs from hereditary HLA‐related haemochromatosis mostly for the absence of iron overload. A gene responsible for the congenital nuclear cataract likely maps on chromosome 19q close to the ferritin L‐subunit gene. Within families with autosomal dominant congenital cataract, serum ferritin might be an early marker of disease.
Detection of c‐fms protooncogene in early mouse embryos by whole mount in situ hybridization indicates roles for macrophages in tissue remodelling
Authors: HUME DAVID A. , MONKLEY SUSAN J. , WAINWRIGHT BRANDON J.
Published: March 2008

Summary. The c‐fms protooncogene which encodes the receptor for macrophage colony‐stimulating factor (CSF‐1) was localized in the developing mouse embryo by whole in situ hybridization, c‐fms was expressed first in placental trophoblasts. Around 9‐5 dpc, isolated c‐/ms‐positive cells became detectable in the yolk sac and by 10‐5 dpc large numbers were detectable throughout the embryo. The localization of c‐fms expression was consistent with its restriction to macrophages, and with the location of those macrophages in sites of tissue turnover and extensive cell death.
Hepatosplenic gamma‐delta T‐cell malignant lymphoma: report of the first case in childhood, including molecular minimal residual disease follow‐up
Authors: GARCIA‐SANCHEZ F. , MENÁRGUEZ J. , CRISTOBAL E. , CANTAI.EJO A. , GIL J. , ALGARA P. , VICARIO J. L.
Published: March 2008

Summary. We report the first case of T‐cell 7.5+ hepatosplenic malignant lymphoma in childhood. Tumour‐specific oligoprobes were developed against the single Vl‐Jl rearrangement of the δ T‐cell receptor (TCR) gene in order to perform minimal residual disease (MRD) studies. Molecular analysis in serial bone marrow samples proved to be of predictive value concerning the clinical outcome. Clonotypic DNA was not detected in peripheral blood during the course of the disease until a refractory terminal leukaemic phase took place 18 months after the diagnosis. This case demonstrates the usefulness of MRU studies to monitor the course of disease in at least some subsets of peripheral T‐cell lymphomas.
Acute myeloblasts leukaemia without Philadelphia chromosome developing after interferon therapy for chronic myelocytic leukaemia with Philadelphia chromosome
Authors: OHTSUKA EIICHI , KIKUCHI HIROSHI , ABE YUSEI , MORTYAMA KYOKO , OHNO EIJI , HIROTA KIYOSHI , TEZONO KAZUHITO , NASU MASARU
Published: March 2008

Summary. We report a patient who developed Philadelphia chromosome negative acute myeloblastic leukaemia with trisomy 8 and trisomy 11 after receiving treatment with alkylating agents and interferon for chronic myelocytic leukaemia positive for Philadelphia chromosome. Leukaemic cells were positive for myeloperoxidase and expressed CD13, CD33 and DR; some expressed CD2, CD4 and CD34. The fluorescence in situ hybridization method revealed that bcr‐abl fusion genes were absent from > 90% of the bone marrow cells. The major bcr rearrangement was not detected by Southern blot analysis. We conclude that the leukaemic cells negative for Philadelphia chromosome may have developed as a result of treatment with alkylating agents and interferon in the present case.
A novel amber mutation in a β°‐thalassaemia gene (β37TGG→TAG), with direct detection by mapping the restriction fragments in amplified genomic DNA
Authors: Xu XIANGMIN , LIAO CAN , LIU ZHONGYING , LI JIAN , PENG ZHAOHUI , QIU LUO‐LIN , ZHANG JIZENG
Published: March 2008

Summary. A novel amber mutation, a G to A substitution at the second position of codon 3 7 in the β‐globin gene that changes the tryptophan coding triplet (TGG) to a termination codon (TAG), was found in a Chinese β‐thalassaemia carrier. The mutant gene creates an additional Dde I recognition site and eliminates the Ava II site, so this point mutation can be directly identified by restriction enzyme analysis.
GM‐CSF‐induced acute eosinophilic pneumonia
Authors: SEEBACH JORG , SPEICH RUDOLF , FEHR JORG , TUCHSCHMID PETER , RUSSI ERICH
Published: March 2008

Summary. There is increasing evidence that haemopoietic growth factors are effective in reversal of neutropenia associated with large granular lymphocytes (LGLs) proliferation. A 19‐year‐old woman with CD3+/TCRγδ+, CD4−, CD8− LGL proliferation and severe neutropenia repeatedly developed blood eosinophilia, fever and dyspnoea after administration of GM‐CSF. Acute eosinophilic pneumonia with a lobar lung infiltrate pleural effusion, and marked bronchoalveolar lavage fluid eosinophilia was diagnosed. Treatment with corticosteroids and discontinuation of GM‐CSF lead to rapid improvement. In addition, haematological analysis revealed that H*1 Technicon, a widely‐used automated cell counter, may misinterpret eosinophils erroneously as neutrophils, therefore examination of blood smears to prevent eosinophil‐medlated toxicity during GM‐CSF treatment is recommended.
Molecular analysis of PML‐RARa fusion mRNA detected by reverse transcription‐polymerase chain reaction assay in long‐term disease‐free acute promyelocytic leukaemia patients
Authors: MARTINELLI G. , REMIDDI C. , VISANI G. , FARABEGOLI P. , TESTONI N. , ZACCARIA A. , MANFROI S. , CENACCHI A. , Russo D. , BANDINI G. , AMABILE M. , DE VIVO A. , TURA S.
Published: March 2008

Summary. By use of RT‐PCR of PML/RARa, we evaluated bone marrow aspirates in 10 patients with APL in long‐term disease‐free status after induction chemotherapy and consolidation (median 54 months; range 33‐101 months from complete remission). All patients were in clinical and cytogenetic remission at the time of molecular evaluation (range 32‐96 months from CR). All patients but one were found to be RT‐PCR negative at the molecular level for the expression of PML‐RARa transcript, confirming that the majority of the patients with long‐term survival of APL are characterized by the eradication of the neoplastic clone.
Molecular genetics of glucose‐6‐phosphate dehydrogenase (G6PD) deficiency in Spain: identification of two new point mutations in the G6PD gene
Authors: Rovira A. , Vulliamy T. , Pujades M. A. , Luzzatto L. , Vives J. Ll.
Published: March 2008

Summary .In order to explore the nature of glucose‐6‐phosphate dehydrogenase (G6PD) deficiency in Spain, we have analysed the G6PD gene in 11 unrelated Spanish G6PD‐deficient males and their relatives by using the polymerase chain reaction and single‐strand conformation polymorphism (PCR‐SSCP) analysis combined with a direct PCR‐sequencing procedure and PCR‐restriction enzyme (RE) analysis. We have identified eight different missense mutations, six of which have been reported in previously described G6PD variants. In nine patients who had presented with acute favism we found the following mutations: G6PD A_376G–202A (four cases)| G6PD Union1360T (two cases), G6PD Mediterranean563T, (one case) and G6PD Aures143c (one case). In the remaining patient a novel A to G transition was found at nucleotide position 209 which has not been reported in any other ethnic group. This mutation results in a (70) Tyr to Cys substitution and the resulting G6PD variant was biochemically characterized and designated as G6PD Murcia. This new mutation creates a Bsp 12861 recognition site which enabled us to rapidly detect it by PCRRE analysis. In two patients with chronic non‐spherocytic haemolytic anaemia (CNSHA) we found the underlying genetic defects, as had been noted previously, to be located within a cluster of mutations in exon 10. One of them had the T to C transition at nucleotide 1153, causing a (3 8 5) Cys to Arg substitution, previously described in G6PD Tomah. The other, previously reported as having a variant called G6PD Clinic, has a G to A transition at nucleotide 1215 that produces a (405) Met to He substitution, thus confirming that G6PD Clinic is a new class I variant.
Glucosamine 6‐phosphate deaminase in normal human erythrocytes
Authors: Weidanz Jon A. , Campbell Patrick , DeLucas Lawrence J. , Jin Jin , Moore Dwight , Roden Lennart , Yu Hao , Heilmann Eric , Vezza Anne C.
Published: March 2008

Summary In the course of an investigation of hexosamine catabolism in the human malaria parasite, Plasmodium falciparum, it became apparent that a basic understanding of the relevant enzymatic reactions in the host erythrocyte is lacking. To acquire the necessary basic knowledge, we have determined the activities of several enzymes involved in hexosamine metabolism in normal human red blood cells. In the present communication we report the results of studies of glucosamine 6‐phosphate deaminase (GlcN6‐P) using a newly developed sensitive radiometric assay. The mean specific activity in extracts of fresh erythrocytes assayed within 4h of collection was 14–7nmol/h/mg protein, whereas preparations from older erythrocytes that had been stored at 4°C for up to 4 weeks had a mean specific activity of 6–2 nmol/h/mg. Characterization of the deaminase by chromatofocusing gave a pi of 8–55. The enzyme was optimally active at pH 9–0 and had a Km of 41 MUM. The metal chelators EDTA and EGTA were non‐inhibitory; however, inhibition was observed in the presence of metal ions, especially Cu2+, Ni2+ and Zn2+. In addition, the deaminase was also inhibited by several sugar phosphates including the reaction product, fructose 6‐phosphate.
Management of persistent B19 parvovirus infection in AIDS
Authors: Ramratnam Bharat , Gollerkeri Ashwin , Schiffman Fred J. , Rintels Peter , Flanigan Timothy P.
Published: March 2008

Summary An HIV+ 26‐year‐old white man with a CD4 count of 006 × 109/1 was found to have red blood cell aplasia secondary to B19 parvovirus infection. Regular infusions of intravenous immunoglobulin (IVIG) were begun and resulted in marked reticulocytosis and correction of anaemia. The patient has been followed for over 4 years and has become anaemic and reticulocytopenic whenever IVIG was interrupted. Serial dot blot analysis of the patient's sera for B19 parvovirus DNA showed absence of DNA immediately following IVIG treatments but reappearance within 3–6 weeks. Regular IVIG was effective in controlling but not eradicating B19 parvovirus infection in this HIV+ patient.
Corticosteroid resistance is increased in lymphoblasts from adults compared with children: preliminary results of in vitro drug sensitivity study in adults with acute lymphoblastic leukaemia
Authors: Maung Z. T. , Reid M. M. , Matheson E. , Taylor P. R. A. , Proctor S. J. , Hall A. G.
Published: March 2008

Summary The prognosis of acute lymphoblastic leukaemia (ALL) in adults is poor compared with children in terms of complete remission (CR) and leukaemia‐free survival. In children in vitro resistance of leukaemic cells to various cytotoxic agents is an independent poor prognostic marker, but the relevance of in vitro drug resistance in adults to poor prognosis has not been described. Lymphoblasts from 16 adults and 32 children with ALL at initial presentation were assayed for in vitro drug sensitivity in a short‐term culture system using the reduction of 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyl tetrazolium bromide (MTT) as an indicator of cell viability. The following drugs were tested: prednisolone, daunorubicin, mitozantrone, etoposide, melphalan and 6‐thioguanine. At initial presentation, lymphoblasts from adults demonstrated a significantly higher degree of in vitro resistance to prednisolone than those from children (P < 001). Glucocorticoid resistance may be a fundamental difference between adult and childhood ALL which may underlie different biological aspects and also explain the difference in prognosis. Lymphoblasts from adults who achieved CR were more sensitive in vitro to prednisolone (P = 007), daunorubicin (P < 005), mitozantrone (P < 001) and melphalan (P < 005) than cells from those who did not. The MTT assay can predict response to induction chemotherapy in adults and therefore discriminate between standard‐ and high‐risk patients. The assay, however, is not suitable for selection of the most effective agent for treatment because of in vitro cross‐resistance of lymphoblasts to various drugs tested.
Heparin‐induced thrombocytopenia: an improved method of detection based on lumi‐aggregometry
Authors: Stewart Michael W. , Etches Wai S. , Boshkov Lynn K. , Gordon Philip A.
Published: March 2008

Summary Heparin‐induced thrombocytopenia (HIT) is a recognized complication of heparin administration. Early detection of this syndrome is essential in the prevention of immune‐mediated thromboembolic sequelae. The 14C‐sero‐tonin release assay (SRA) has been used in reference laboratories to identify sera from patients on heparin therapy capable of inducing platelet dense granule release. In an attempt to improve existing methodologies, we employed luminographic detection of platelet‐dense granule ATP release as an endpoint of HIT antibody‐mediated platelet activation. Sera tested included 10 SRA confirmed positive and five SRA confirmed negative samples (to establish the assay), five samples from patients with thrombocytopenia not on heparin therapy and 34 patients suspected of HIT syndrome. All SRA confirmed positive sera (n= 19) were positive by the luminographic procedure. 24/26 SRA confirmed negative sera and five sera from thrombocytopenic patients not on heparin therapy were negative using luminography. Two of four sera yielding equivocal SRA results were found to be positive by the luminographic technique. The data suggest that the use of a lumi‐aggregometer in the coagulation laboratory to detect HIT antibody‐induced platelet activation is a reliable alternative to the SRA. The luminographic procedure is both rapid and sensitive, and does not require the use of biohazardous radio‐isotopes.
Evidence for the aetiological role of blood‐borne virus infections in causing reduced lectin‐induced T‐cell proliferation in haemophilic boys
Authors: Evans J. A. , Pasi K. J. , Hill F. G. H.
Published: March 2008

Summary T‐lymphocyte function, as expressed by polyclonal proliferation to lectin mitogens phytohaemagglutinin and concanavalin A, has been studied in a normal control population and three groups of haemophilic boys: group 1, HIV and HCV seronegative and treated only with a single heat‐treated factor VILT (FVITI) concentrate; group 2, HIV seronegative but HCV seropositive; group 3, all HIV and HCV seropositive. Groups 2 and 3 have been previously treated with unheated and heated FVIII concentrate. Group 1 boys (HIV/HCV uninfected) showed no significant reduction in lymphocyte proliferation when compared with a control population. Group 2 and 3 boys showed an impaired response to these mitogens compared to group 1 boys and the control group. There was no relationship between FVIII concentrate received and proliferative response. The absence of immune modulation in haemophilic boys who have not acquired HIV and HCV infection implicates chronic blood‐borne virus infections as the major contributory factors to impaired lymphocyte proliferative responses seen in haemophiliacs treated with large‐pool concentrates. The presence of virus infections, such as HCV, may account for similar lymphocyte function abnormalities observed in previously described cohorts.
Recombinant PDGF enhances megakaryocytopoiesis in vitro
Authors: Yang Mo , Chesterman Colin N. , Chong Beng H.
Published: March 2008

Summary. The effect of recombinant platelet‐derived growth factor (PDGF) on both murine and human megakaryocyte colony formation was studied in the plasma clot culture system. PDGF significantly stimulates megakaryocyte colony formation in a dose‐dependent manner. The minimum concentration which had a significant stimulating effect on colony forming unit megakaryocyte (CFU‐MK) was lOng/ml and maximum stimulation occurred at 50 ng/ml. The effect of PDGF was compared with that of interleukin (IL)‐3, IL‐6, granulocyte‐macrophage colony stimulating factor (GM‐CSF), erythropoietin (EPO) and acid fibroblast growth factor (aFGF) on megakaryocyte colony formation. The results showed that megakaryocyte colony stimulating activity of PDGF was slightly higher than those of GM‐CSF and aFGF, but lower than those of IL‐3, IL‐6 and EPO. The effect of PDGF in combination with IL‐3 or IL‐6 on megakaryocyte colony formation was also investigated. No synergistic action was found between PDGF and IL‐3 or IL‐6, but an additive effect was observed with IL‐3 plus IL‐6. We also studied the effects of PDGF in combination with anti‐IL6, anti‐IL‐3 or anti‐GM‐CSF antibody. The increase of megakaryocyte colony formation induced by PDGF was partially inhibited by anti‐IL‐6 or anti‐GM‐CSF antibody but not by anti‐IL‐3 antibody. These results indicate that PDGF is a positive regulator for megakaryocytopoiesis in vitro and IL‐6 and GM‐CSF may play a role in the mechanism whereby PDGF stimulates megakaryocytopoiesis.
Three unrelated Gaucher's disease patients with three novel point mutations in the glucocerebrosidase gene (P266R, D315H and A318D)
Authors: Walley A. J. , Ellis I. , Harris A.
Published: March 2008

Summary. Three novel point mutations were detected in the glucocerebrosidase gene of three unrelated Gaucher's disease patients by direct sequencing of PCR products. The first is a C to G change at position 4263 in the genomic sequence (exon 7) which results in a proline to arginine change at position 266 in the mature enzyme (P266R). The second is a G to C change at position 5276 in the genomic sequence (exon 8) which results in an aspartic acid to histidine change at position 315 (D315H), The third is a C to A change at position 5286 in the genomic sequence (exon 8) which results in an alanine to aspartic acid change at position 318 (A318D). The first mutation destroys an AvaII restriction endonuclease site, the second creates a BspM I site and the third creates a BamH I site.
Polyclonal hypergammaglobulinemia in a case of B‐cell chronic lymphocytic leukaemia: the result of IL‐2 production by the proliferating monoclonal B cells?
Authors: Mouzaki Athanasia , Matthes Thomas , Miescher Peter A. , Beris Photis
Published: March 2008

Summary. CLL is typically characterized by acquired hypogammaglobulinemia. We report the case of a female patient suffering from B‐CLL who developed polyclonal hypergammaglobulinaemia: 38‐3 g/I polyclonal IgG, 0.97g/1 IgA and 0.33 g/1 IgM. Immunophenotyping showed a monoclonal lymphocytic population CD19+ CD5+ CD40+ CD23+, low slg+ (95%), K type in the great majority (96%). RT‐PCR of immunoglobulin genes gave evidence of monoclonal rearrangement of the IgM type. Our tests showed that IL‐2 was produced when leukaemic B cells were stimulated with phorbol myristate acetate, ionomycin and lipopoly‐saccharide. In addition, transfections with the full IL‐2 promoter or elements thereof revealed that IL‐2 expression is inducible and mediated through the NF‐kB‐promoter element. Finally, the amount of IL‐2 secreted by these cells is about 39ng/ml/106 cells, which is remarkably high for non‐T cells. These results suggest that the large amounts of polyclonal IgG seen in this case of B‐CLL are secreted by normal B cells which are in turn stimulated by IL‐2 produced by proliferating monoclonal (leukaemic) B cells. Under cyclosporin A treatment, immunoglobulin secretion and B cell count remained low.
Homozygous loss of the MTSl/pl6 and MTS2/pl5 genes in lymphoma and lymphoblastic leukaemia cell lines
Authors: Siebert Reiner , Willers Christoph P. , Schramm Alexander , Fossa Alexander , Dresen Irina M. Gana , Uppenkamp Michael , Nowrousian Mohammad R. , Seeber Siegfried , Opalka Bertram
Published: March 2008

Summary. The genes MTS1/p16 and MTS2/p15 located in 9p21 encoding cyclin‐dependent kinase‐4 inhibitors are homozygously deleted in a number of different tumour cell lines. By PCR analysis of 30 cell lines, including 10 acute lymphoblastic leukaemia (ALL) and 20 lymphoma cell lines, we found homozygous deletions of at least one locus in 11 (37%) cell lines. MTS1‐speciflc sequences were deleted in 70% of ALL (reaching 86% in T‐cell ALL) but in none of the non‐Hodgkin's lymphoma (NHL) cell lines. MTS2‐specific sequences were deleted in 40% of ALL and 17% of NHL cell lines. We observed a higher frequency of MTS1 deletions in ALL than in NHL (P< 0.001) and in T‐cell neoplasms compared to B‐cell neoplasms (67%v 6%; P= 0.001). In ALL‐derived cell lines deletions of the MTS2 gene only occurred in cases with MTS1 deletions but in NHL only in cases without MTS1 deletions.
The expression of mRNA for fibrinogen in megakaryocytes isolated from patients with T‐cell lymphoma
Authors: Podolak‐Dawidziak M. , Hancock V. , Lelchuk R. , Kotlarek‐Haus S. , Martin J. F.
Published: March 2008

Summary. The expression of fibrinogen mRNA was studied by in situ hybridization in freshly isolated megakaryocytes in 14 newly‐diagnosed patients: seven with non‐Hodgkin's lymphoma (NHL), three with immune thrombocytopenic purpura (ITP) and four haematologically normal patients prior to coronary artery bypass surgery. Fibrinogen mRNA in megakaryocytes was not detected in ITP, B‐cell lymphomas or in healthy donors. However, it was present in all patients with the high‐grade T‐cell lymphomas, both with and without thrombocytopenia.