bluuebunny/crossref_metadata_2025 huggingface dataset:
Stem cell factor enhances the growth of primitive erythroid progenitors to a greater extent than interleukin‐3 in patients with aplastic anaemia
Authors: Amano Yoshiro , Koike Kenichi , Nakahata Tatsutoshi
Published: March 2008

Summary We examined the combined effects of stem cell factor (SCF). or interleukin‐3 (IL‐3) with erythropoietin on the development of haemopoietic progenitors in 19 patients with aplastic anaemia (AA) and eight normal controls by using an in vitro clonal assay. SCF significantly enhanced the growth of total erythroid colonies (erythroid bursts, mixed colonies) in 11 patients and all normal controls, whereas IL‐3 did so in only three patients. The number of SCF‐ or IL‐3–dependent erythroid colonies was substantially lower in AA patients than in the controls. Comparison of the capacity of SCF and IL‐3 to increase total erythroid colony growth indicated that half of the AA patients responded more strongly to SCF than the normal controls, while few patients responded in such a manner to IL‐3. These findings suggest that SCF in vivo will have a more dramatic effect than IL‐3 in improving anaemia in patients with AA.
Cytokine mRNA expression of bone marrow stromal cells from patients with aplastic anaemia and myelodysplastic syndrome
Authors: Hirayama Yasuo , Kohgo Yutaka , Matsunaga Takuya , Ohi Shigeo , Sakamaki Sumio , Niitsu Yoshiro
Published: March 2008

Summary We studied mRNA expression of the cytokine granulocyte‐colony stimulating factor (G‐CSF), interleukin‐1β (IL‐1β), IL‐6, IL‐8 and stem cell factor of stromal cells derived from bone marrows of nine normal volunteers, eight patients with aplastic anaemia (AA) and seven patients with myelodysplastic syndrome (MDS). The proportion of endothelial cells. macrophages. fibroblast‐like cells and adipocytes in stromal cells showed no differences between normal volunteers and the patients. Levels of cytokine inKNA expression were determined by reverse transcription‐polymerase chain reaction. Spontaneous expression occurred and this was augmented by IPS stimulation in cells of all the normal volunteers and in most patients.

When stimulated by LPS. the mean G‐CSF and IL‐1β mRNA expressions in patients with AA were significantly higher than normal volunteers. but there was one patient showing lower IL‐1β. IL‐6 and 71,‐8 expression with no response to LPS. LPS‐induced IL‐6 and IL‐8 expression of two patients with MDS were significantly higher than normal.

The spontaneous and LPS‐induced protein concentration of G‐CSF, IL‐6 and IL‐8 in culture supernatants from 15, 10 and four patients, correlated well with the niKNA expression. The correlation coefficients were 0.92. 0.78 and 0.91. respectively.

In conclusion, there were a few patients whose aetiology appeared to be reduction of stromal cytokine expression in AA. but most patients with AA and MDS expressed normal or high levels of cytokine mKNA.
Transforming growth factor beta‐1 (TGF‐β1) released by an Epstein‐Barr virus (EBV) positive spontaneous lymphoblastoid cell line from a patient with Kostmann's congenital neutropenia inhibits the growth of normal committed haemopoietic progenitors in vitro
Authors: Corcione A. , Roncella S. , Cutrona G. , Mori P. G. , Ferrarini M. , Pistoia V.
Published: March 2008

Summary This study reports the characterization of a spontaneous lymphoblastoid cell line (LCL) raised from the peripheral blood of a patient with Kostmann's congenital neutropenia. The LCL was composed of EBV‐infected polyclonal B cells and displayed surface markers and pattern of growth in vitro typical of normal LCLs. The supernatant of the LCL contained a colony inhibiting activity (CIA) that decreased the cloning efficiency of normal committed haemopoietic progenitors and was identified as immunoreactive transforming growth factor β1 (TGF‐β1) by neutralization experiments with a specific antiserum. Control studies with a panel of LCLs spontaneously derived from the peripheral blood of patients seropositive for Epstein‐Barr virus (EBV) infections showed that 5/30 LCLs produced a CIA. This CIA was not identifiable as TGF‐β1 but rather was due to the combined effects of tumour necrosis factor α (TNFα). tumour necrosis factor β (TNFβ) and interferon α (IFNα), that were present in the LCL supernatants. The hypothesis that the B cells latently infected by EBV in vivo and possibly expanded as a consequence of the infection may have contributed to the inhibition of the patient granulopoiesis by releasing TGF‐β1 will be discussed.
The relationship between human megakaryocyte nuclear DNA content and gene expression
Authors: Hancock V. , Martin J. F. , Lelchuk R.
Published: March 2008

Summary Increased platelet reactivity has been implicated in various vascular diseases. Since the protein content of platelets is determined mainly by the megakaryocyte. alterations in megakaryocyte mKNA expression may influence platelet protein content and therefore activity. In order to determine whether DNA content (ploidy) of a megakaryocyte influences its mRNA expression, we have developed a method to investigate the relationship between megakaryocyte ploidy and gene expression. By measuring the ploidy and mRNA expression in individual cells, we have shown that in the physiological state there is an increase in mRNA expression for β‐actin, glycoprotein IIb and neuropeptide Y with increase in ploidy and that this increase levels off at high ploidy values. This may have relevance in the understanding of platelet reactivity in pathological events.
Procoagulant activity of the MC28 fibrosarcoma cell line in vitro and in vivo
Authors: Amirkhosravi Mohammadali , Francis John L.
Published: March 2008

Summary Experimental evidence suggests that many tumours can activate blood coagulation and that such interaction is part of the pathology of metastatic tumour growth. This study aimed to study the procoagulant activity of the methylcholanthrene‐induced (MC28) fibrosarcoma to determine whether coagulation activation by these cells could explain the previously reported effects of oral anticoagulants on lung seeding in this model. MC28 cells shortened the recalcification times of normal and factor VII‐deficient plasma and directly activated factor X in a chromogenic assay, but did not aggregate platelets iri vitro in either whole blood or platelet‐rich plasma. Cellular coagulant activity was calcium‐dependent, blocked by DFP and concanavalin A but not inhibited by iodoacetamide, E‐64 or antibodies to human tissue factor or factor VII. Injection of viable MC28 cells into hooded Lister rats induced a decrease in platelet count (P<0.001), plasma factor X (P<0.001) and fibrinogen (P<0.05) and a marked increase in plasma haemoglobin (P< 0.001). These effects were either not observed or were considerably less marked in heparinized or warfarinized animals. Injection of MC28 cells treated with concanavalin A in vitro completely abolished the clotting changes observed with untreated cells. In conclusion, MC28 cells possessed a potent factor X‐activating serine proteinase procoagulant in vitro, which had some of the characteristics of a tissue factor/ factor VIIa complex. In vivo, MC28 cells caused clotting activation and intravascular fibrin generation. Since thrombocytopenia was abolished by heparin and the cells lacked platelet aggregating activity in vitro, thrombocytopenia was probably secondary to intravascular coagulation and thrombin generation. The trigger for intravascular clotting activation appeared to be the cellular procoagulant activity since it was abolished by prior in vitro blockade of the latter with concanavalin A.
The ‘euthyroid sick syndrome’: incidence, risk factors and prognostic value soon after allogeneic bone marrow transplantation
Authors: Vexiau Patrick , Perez‐Castiglioni Patricia , Socié Gèrard , Devergie Agnès , Toubert Marie Elisabeth , Aractingi Sélim , Gluckman Eliane
Published: March 2008

Summary We studied the incidence of thyroid function abnormalities observed soon after allogeneic bone marrow transplantations (BMT) and their predictive value on the overall prognosis. Free serum thyroxine, free serum triiodothyronine, total serum reverse triiodothyronine and serum thyrotropin levels were systematically measured in 78 patients before and 3 months after BMT. 41 (52%) had normal hormone levels and 37 (48%) had abnormal ones, among whom four (5%) had peripheral compensated hypothyroidism and 33 (43%) were described as having ‘euthyroid sick syndrome’ (low thyroxine state, or low T3 syndrome). Two factors strongly influenced the appearance of thyroid abnormalities: steroid dose at the time of thyroid function testing, and age (≤16 years/>16 years). Among the younger patients, 21 had no thyroid abnormalities, while five did. Among the older patients. 20 had no thyroid abnormalities, while 32 did (P<0.001). The occurrence of thyroid abnormalities seemed to influence survival strongly, since the 30‐month projected survival time was 83% for patients without abnormalities whereas it was 49% for patients with an abnormal profile (P < 0.001). In conclusion, evidence obtained among our population reveals that euthyroid sick syndrome indicates a poor prognosis and that it is very important to monitor thyroid hormone levels (particnlady free hormones) soon after allogeneic BMT and regularly thereafter.
Retinoic acid‐induced apoptosis and regression of a refractory Epstein‐Barr virus‐containing T cell lymphoma expressing multidrug‐resistance phenotypes
Authors: Su Ih‐Jen , Cheng Ann‐Lii , Tsai Tsern‐Fan , Lay Jong‐Ding
Published: March 2008

Summary The virus‐associated T cell leukaemias/lymphomas are characterized by a poor prognosis primarily because of the rapid emergence of drug resistance which may lead to failure of subsequent chemotherapy. We report here a case of Epstein‐Barr virus‐associated T cell lymphoma which relapsed soon after chemotherapy and radiotherapy. The neoplastic cells of the relapsed tumour expressed high levels of multi‐drug resistance gene (mdr 1)‐related P‐glycoprotein and glutathione‐S‐transferase‐π, both of which were absent in the pre‐chemotherapy tumour tissues. Empirical treatment with oral 13‐cis‐retinoic acid (RA) was then given with subsequent complete disappearance of the tumour. The therapeutic effect of KA appears to act through an apoptotic process. In accordance with our previous report of a successful salvage of a refractory Ki‐1 large cell lymphoma. RA appears to be a potentially useful drug for some specific type T‐cell lymphomas.
The effects of saline solutions on red cell shape: a scanning‐electron‐microscope‐based study
Authors: Simpson L. O.
Published: March 2008

Summary Red cells appear to change shape in response to alterations in their environment both in vitro and in viva To investigate the qualitative aspects of this phenomenon, five drops of a venous blood sample were fixed in buffered glutaraldehyde for baseline observations and 0.5 ml of blood added to 4 ml of four different saline solutions. Triplicated 10 drop samples from the suspensions were fixed in the glutaraldehyde solution after 2.5, 5, 10, 20 and 40 min and prepared for scanning electron microscopy.

Red cell shape analysis of the resulting micrographs showed that the cells had changed shape, although no two patterns of change were the same.
Factors influencing haematological recovery in 53 patients with acute myeloid leukaemia in first remission after autologous bone marrow transplantation
Authors: Pendry K. , Alcorn M. J. , Burnett A. K.
Published: March 2008

SummaryThe kinetics of haematological recovery were retrospectively analysed in 53 patients with acute myeloid leukaemia in first remission after myeloablative chemoradiotherapy followed by autologous bone marrow transplantation. The median time to achieve a neutrophil count of 1 × 109/1 was 46d (22–196 d) and median time to achieve unsupported platelet counts of 20 × 109/1 and 50 × 109/1 was 70 d (24–310 d) and 126 d (29–497 d) respectively. Multivariate analysis revealed two factors that were significantly associated with delayed neutrophil and platelet recovery: (1) use of high dose fractionated TBI and mononuclear cell cryopreservation, and (2) low platelet count at the time of bone marrow harvest. There was no correlation with: number of courses of chemotherapy, remission to ABMT interval, CMV status, indices of autograft quality or the development of elevated platelet associated immunoglobulin. Delayed haematological recovery did not predict for relapse or death. Delayed platelet recovery did, however, present significant problems with increased blood and platelet requirements and lengthening of hospital stay.
Erythrocyte haemolysate interacts with ATP–Fe to form a complex containing iron, ATP and 13 800 MW polypeptide
Authors: Weaver Janet , Zhan Hui , Pollack Simeon
Published: March 2008

Summary Iron first entering the reticulocyte is bound to ATP in the low MW cytosolic pool; some is also ‘loosely bound’ to haemoglobin, coeluting with haemoglobin from a molecular sieve column though not incorporated into haem. When haemolysate is mixed with ATP‐Fe in vitro a similar high MW iron‐containing complex is formed: the ATP‐Fe interacts with a non‐haemoglobin constituent of the haemolysate to form a high MW ATP‐Fe complex in which the ratio of ATP:Fe (originally 6:1) is reversed, so that the complex contains more iron than ATP. The high MW ATP‐Fe complex is formed even when ATP is in 150‐fold molar excess and is formed without detectable hydrolysis of the ATP. The activity of haemolysate in forming the high MW ATP‐Fe complex is not diminished by dialysis; all of the activity is recovered in the haemoglobin‐containing fraction obtained from an Ultrogel AcA 44 column. The activity does not derive from haemoglobin since 85% of the activity is removed when haemoglobin is purified from haemolysate with DEAE‐Sephadex. The chelatable iron pool of the cell probably includes both the high MW ATP‐Fe complex and low MW ATP‐Fe. Shunting of ATP‐Fe to a high MW aggregate reduces the amount of iron present in the highly reactive low MW form and thus probably serves to limit the formation of cell damaging radicals.
Clinical, haematological and cytogenetic features in 24 patients with structural rearrangements of the Q arm of chromosome 3
Authors: Grigg A. P. , Gascoyne R. D. , Phillips G. L. , Horsman D. E.
Published: March 2008

Summary To determine the frequency and clinical significance of acquired abnormalities of chromosome 3 at q21 and q25–26 in haematological malignancy, we reviewed the haematological and cytogenetic features of 24 patients with a 3q rearrangement identified amongst 1200 cases with clonal cytogenetic abnormalities detected at our institutions during a 12‐year period. Thirteen patients presented with de novo acute myeloid leukaemia (AML), 10 with myelodysplasia (MDS) and one in blast phase of chronic myelogenous leukaemia. Twenty patients (83%) had megakaryocytic dysplasia and 14 (58%) had normal or increased numbers of megakaryocyte, but only four patients (16%) had absolute thrombocytosis > 500 × 109/l (three with AML, one with MDS transforming to AML).

A review of 205 cases of AML investigated in our institutions between 1985 and 1990 for whom cytogenetic results were obtained revealed that a platelet count > 500 × 109/l at presentation was highly suggestive of an underlying 3q abnormality. A limited review of the literature on this subject confirmed that 15‐20% of patients with a 3q abnormality will have thrombocytosis. The response of these patients to conventional antileukaemic therapy is uniformly poor, despite haematological and clinical differences between the subtypes of 3q rearrangements.
Immunohistological patterns of myeloid antigens: tissue distribution of CD13, CD14, CD16, CD31, CD36, CD65, CD66 and CD67
Authors: Bordessoule D. , Jones M. , Gatter K. C. , Mason D. Y.
Published: March 2008

Summary. A number of differentiation antigens on myeloid cells have been defined on the CD classification system by the four International Workshops on Human Leucocyte Differentiation Antigens. The distribution of eight of these antigens (CD13, CD14, CD16, CD31, CD36, CD65, CD66, CD67) have been studied in human tissues, with the aim of documenting their immunohistological patterns and their degree of myeloid restriction. CD13, the most widely distributed antigen, was found in skin, bile canaliculi, kidney and pancreas. CD14 was not restricted to monocytes and tissue macrophages, being also strongly expressed on dendritic reticulum cells. CD 16 was expressed on granulocytes and tissue macrophages (alveolar and Kupffer cells) and in the red pulp of the spleen. CD31 and CD36 gave a characteristic staining of vascular endothelium, corresponding to their identification as the platelet glycoproteins gp IIa and gp IV. Antibodies against the most recently defined myeloid antigens (CD65, CD66 and CD67) appeared to be more specific for myeloid differentiation than previously described ‘myeloid antigens’.
Decreased levels of myeloid progenitor cells associated with long‐term administration of recombinant human granulocyte colony‐stimulating factor in patients with autoimmune neutropenia
Authors: Taniguchi Shuichi , Shibuya Tsunefumi , Harada Mine , Niho Yoshiyuki
Published: March 2008

Summary. We studied the long‐term in vivo effect of recombinant human granulocyte colony stimulating factor (rhG‐CSF) on in vitro growth of granulocyte/macrophage colony forming cells (GM‐CFC) in bone marrow and peripheral blood obtained from two patients with autoimmune neutropenia, who received rhG‐CSF. Along with rhG‐CSF treatment for more than 40 d, numbers of GM‐CFC‐derived colonies from both bone marrow and peripheral blood gradually decreased to a significant level though white blood cells in peripheral blood and nucleated cells in bone marow were increased in number. This observation suggests that long‐term administration of rhG‐CSF may preferentially activate a differentiation pathway for granulopoiesis while proliferation of GM‐CFC is not induced as expected in response to rhG‐CSF.
Impact of interferon alpha‐induced cytogenetic improvement on survival in chronic myelogenous leukaemia
Authors: Kloke O. , Niederle N. , Qiu J. Y. , Wandl U. , Moritz T. , Nagel‐Hiemke M. , Hawig I. , Opalka B. , Seeber S. , Becher R.
Published: March 2008

Summary. The objective of this study was to investigate the prognostic impact of the reduction of Philadelphia chromosome (Ph) positive metaphases by treatment of chronic myelogenous leukaemia (CML) with interferon (IFN) alpha. Therefore, we evaluated the outcome of patients with previously untreated chronic phase Ph‐positive CML, enrolled from 1984 to 1990 into two consecutive IFN trials at our institution. Of a total of 71 patients, 62 (87%) were evaluable for cytogenetic response. No cytogenetic improvement was seen in 16 patients (23%), 28 patients (38%) had a decrease in Ph‐positive bone marrow metaphases to levels ranging from 35% to 95%, and nine patients (13%) to levels between 5% and 34%. In nine patients (13%), Ph‐positive metaphases were no longer detectable. After a median follow‐up period of 33 months, the projected 5‐year survival is 55% for the 62 patients evaluable for cytogenetic response. In this patient population there was no significant difference in the survival probability according to patients’risk status as defined by the Sokal score. Categorization according to the extent of Ph reduction, however, allowed three groups with significantly different prognoses to be identified. Patients achieving a Ph reduction to less than 35% were found to constitute a low risk group with a median survival not yet known and a projected 5‐year survival of 90%. The 5‐year survival rate was 55% for patients with a Ph reduction to levels between 35% and 95%, and less than 10% for those without any cytogenetic improvement. Thus, this study demonstrates that cytogenetic improvement on IFN treatment is an important prognostic factor for survival.
Altered expression of the human retinoblastoma gene in monocytic leukaemias
Authors: Weide Rudolf , Parviz Behnoush , Pflüger Karl‐Heinz , Havemann Klaus
Published: March 2008

Summary. Inactivation of the retinoblastoma growth suppressor gene (RB) is responsible for the development of retinoblastomas and occurs frequently in osteosarcomas and small cell lung carcinoma. Knowledge about the involvement of RB in the pathogenesis of myeloid leukaemias is still scarce. In this study we looked at the expression of the retinoblastoma gene product (p105) in 20 primary myelomonocytic and monoblastic leukaemias by Western blotting and immunocytochemistry using the anti‐p105‐monoclonal antibody PMG3‐245. We found absence of or barely detectable levels of p105 in 11 patients (55%). Absence of or low levels of p105 were correlated with a higher leucocyte count at presentation (133 × 109/l v 83 × 109/l) and with the occurrence of extramedullary leukaemia (8/10 v 2/10). We conclude that abnormal expression of RB with absence of p105 or strongly reduced p105 levels occurs frequently in myelomonocytic and monoblastic leukaemias and that this may be correlated with a more malignant course of the disease.
Dimethyl sulphoxide and haemin induce ferrochelatase mRNA by different mechanisms in murine erythroleukaemia cells
Authors: Fukuda Yoshiaki , Fujita Hiroyoshi , Taketani Shigeru , Sassa Shigeru
Published: March 2008

The level of mRNA encoding ferrochelatase (FeC), the terminal enzyme of the haem biosynthetic pathway, was examined in murine erythroleukaemia (MEL) cells when they were induced to undergo erythroid cell differentiation by treatment with dimethyl sulphoxide (DMSO), or haemin. FeC mRNA increased within 12 h after DMSO or haemin treatment of MEL cells, and its level continued to increase for 48 h. Treatment of cells with succinylacetone (SA), a potent inhibitor of haem synthesis, suppressed a DMSO‐mediated increase in FeC mRNA, and haemin treatment reversed a SA‐mediated decrease in FeC mRNA. Nuclear runoff analyses showed that, while DMSO increased the rate of transcription of FeC mRNA, haemin did not. These results indicate that the induction of FeC mRNA by DMSO is largely transcriptional, while that by haemin is post‐transcriptional.
Acute hepatitis C infection in patients undergoing therapy for haematological malignancies: a clinical and virological study
Authors: Brink N. S. , Chopra R. , Perrons C. J. , Ring C. J. A. , Garson J. A. , Briggs E. M. , Goldstone A. H. , Linch D. C. , Tedder R. S.
Published: March 2008

Summary. Patients receiving multiple transfusions are at risk of acquiring hepatitis C (HCV) infection from a donor population which is unscreened for hepatitis C antibodies (anti‐HCV). Prior to the introduction of blood donor screening for anti‐HCV in the U.K., a group of patients undergoing therapy for haematological malignancies, with repeatedly abnormal liver function tests, were investigated for acute HCV infection. Thirty‐two patients had repeatedly raised serum transaminases, and eight of these (25%) had evidence of an acute HCV infection. The diagnosis was made by the detection of HCV‐RNA in the patients' serum using a complementary DNA/polymerase chain reaction (cDNA/PCR) procedure. All eight patients had received myeloablative chemotherapy and three had undergone bone marrow transplantation.

HCV infection contributed significantly to the morbidity of this group of patients in the short term whilst they were undergoing treatment for their underlying haematological condition. The long‐term effects have yet to be evaluated. In an attempt to decrease hepatic damage due to HCV. three patients were placed on interferon therapy. None showed a sustained reduction in serum transaminases or HCV viraemia. It is hoped that the introduction of anti‐HCV screening of blood donors, will reduce the frequency of transfusion‐acquired HCV infections. Early observations suggest that this is the case, as we have seen no new cases of HCV infection in our unit since the introduction of donor screening in September 1991.
The peanut‐agglutinin (PNA)‐binding surface components of malignant plasma cells
Authors: Slupsky Joseph R. , Duggan‐Keen Margaret , Booth Laurence A. , Karpas Abraham , Rhodes Elizabeth G. H. , Cawley John C. , Zuzel Mirko
Published: March 2008

Summary. Plasma cells within bone marrow aspirates from multiple myeloma patients have been shown to be reactive with the lectin peanut agglutinin (PNA). This has been recently exploited by using PNA for purging bone marrow of malignant cells in autotransplantation therapy of the disease. The purpose of this investigation was to isolate and characterize the PNA‐binding proteins of myeloma cells. We used the malignant plasma cell‐derived line Karpas‐620 (K620) as a model, and showed by affinity chromatography, SDS‐PAGE, and immunoprecipitation that, among several PNA‐binding proteins, a major one is an incompletely sialylated form of CD44. CD44 is a well‐known homing receptor protein which is rich in carbohydrate and usually completely sialylated so that it does not react with PNA. We have then examined the PNA reactivity of myeloma cells from different patients and showed a clear difference in the profile of PNA‐binding proteins from case to case. Moreover, in contrast to K620 cells, some of the patient plasma cells tested did not have a PNA‐binding form of CD44. In conclusion, therefore, we have shown that a number of different proteins participate in PNA binding by malignant plasma cells. Moreover, we have demonstrated a novel, incompletely sialylated form of CD44 on a myeloma cell line. It is known that the level of glycosylation of CD44 and other proteins may affect their function, but how this relates to the malignant behaviour of plasma cells remains to be determined.
Acute phase proteins and prognosis in multiple myeloma
Authors: Merlini Giampaolo , Perfetti Vittorio , Gobbi Paolo G. , Quaglini Silvana , Franciotta Diego M. , Marinone Gabriella , Ascari Edoardo
Published: March 2008

Serum IL‐6 levels have been shown to correlate with disease severity and prognosis in patients with plasma cell dyscrasias. Among its pleiotropic actions, IL‐6 is also the major regulator of the acute phase response in humans. The possible impact on survival of the major serum acute phase proteins (s.APP) [C‐reactive protein (s.CRP), alpha‐1‐anti‐trypsin (s.AAT), haptoglobin, acid alpha‐1‐glycoprotein and alpha‐2‐macroglobulin (used as control)] was assessed on a population of 103 consecutive, previously untreated myeloma patients. Univariate analysis showed that among the acute phase proteins only s.AAT (P= 0.015) and s.CRP (P= 0.027) were significantly correlated with survival. The multivariate Cox proportional hazard model applied to s.APP and other common parameters showed that s.β‐2‐microglobulin (s.b2M), s.calcium, s.creatinine, BM plasma cell percentage, age and s.AAT correlated significantly with survival. Combining s.b2M and s.AAT allowed stratification of myeloma patients: those with low levels of s.b2M (3 mg/l) and of s.AAT ( 3 g/l) presented an excellent prognosis (median survival exceeding 10 years) while those presenting higher values of the two parameters presented a median survival of 2.5 years (P= 0.002).
A comparison of the antithrombotic effects of heparin and of low molecular weight heparins with increasing antifactor Xa/antifactor IIa ratio in the rabbit
Authors: Carrie D. , Caranobe C. , Boneu B.
Published: March 2008

Summary. This study compares the ability of unfractionated heparin (UH) and of three low molecular weight heparins (LMWHs) to inhibit venous thrombosis growth in the rabbit. Logiparin® (LHN‐1), Fraxiparin® (CY216) and CY222 were selected because they present very different antifactor Xa/ antifactor IIa ratios: 1.7, 3.8 and 6.8 respectively. Heparins were delivered under continuous intravenous infusion for 4 h at increasing doses from 10 to 250 antifactor Xa U kg−1 h−1. The minimum dose providing the maximum inhibitory effect was 50 antifactor Xa U kg−1 h−1. On the basis of this system of units the four heparins were equipotent antithrombotic agents. Due to the highest antifactor Xa/antifactor IIa ratio, CY222 became the most potent antithrombotic agent when the doses were expressed in antithrombin units. Because UH is cleared faster than LMWHs at low dose regimen, the antifactor Xa steady state concentrations generated by the continuous infusion of any of the LMWHs were higher than those generated by UH. In addition the ex vivo antifactor Xa/ antifactor IIa ratio of each of the LMWH was superior to the in vitro ratio. These results indicate that the antithrombotic activity of LMWH is not directly related to its subfraction catalysing thrombin inhibition.
The β+ IVS, I‐NT no. 6 (T→C) thalassaemia in heterozygotes with an associated Hb Valletta or Hb S heterozygosity in homozygotes from Malta
Authors: Scerri C. A. , Abela W. , Galdies R. , Pizzuto M. , Grech J. L. , Felice A. E.
Published: March 2008

Summary. In vitro DNA amplification and dot blot analysis with synthetic allele specific oligonucleotides (ASO) identified the β IVS, I‐6 (T→C) thalassaemia in 78% of 32 chromosomes from 16 β‐thalassaemia homozygotes in Malta. The preponderance of a single thalassaemia mutation in one population is unusual. The β+ IVS, I‐6C thalassaemia mutation was also found in three carriers who had an associated β globin heterozygosity, i.e. Hb Valletta (or α2β287PRO) or Hb S (or α2β26VAL). The proportion of Hb A in these cases (av. = 29.7%) provided objective documentation of the relatively mild effect of this mutation on in vivo globin gene expression. However, the expression of homozygous disease was more severe in developing children compared to adults. The β+ IVS, I‐6C mutation complicates population testing because heterozygotes can have Hb A2 levels below those classically associated with β thalassaemia.
Abnormal neutrophil chemotaxis in Gaucher disease
Authors: Aker M. , Zimran A. , Abrahamov A. , Horowitz M. , Matzner Y.
Published: March 2008

The tendency towards infection described in Gaucher disease patients has been attributed to their post‐splenectomy state. We noticed that certain patients with intact spleen have also suffered from recurrent pyogenic infections, thus an attempt to study their neutrophil functionhas been made. Nine of 29 patients studied expressed significant decrease in neutrophil chemotaxis directed towards zymosan activated serum or N‐formyl‐methionylleucyl‐phenylalanine. Random migration was significantly impaired in five of those nine patients. Adherence of neutrophils to nylon fibres and O2‐ production were intact. the patients with impaired chemotaxis were significantly affected by their disease (early onset of symptoms and severity score index > 10) and most of them had genotypes associated with severe disease (1448/1448 and 1226/84GG). No correlation was found with the spleen status. Three of the patients with impaired chemotaxis, and none of the patients with normal neutrophil function, suffered from recurrent pyogenic infections. It is suggested that the described neutrophil migration impairment may contribute to the tendency towards infection in certain patients with advanced Gaucher disease.
The CD11/CD18 granulocyte adhesion molecules in myelodysplastic syndromes
Authors: Mazzone Antonino , Ricevuti Giovanni , Pasotti Davide , Fossati Gianluca , Mazzucchelli Iolanda , Cavigliano Paola , Notario Antonia
Published: March 2008

We have evaluated the function of granulocytes in 14 patients suffering from myelodysplastic syndrome (MDS). We also evaluated the functional and immunochemical activities of five monoclonal antibodies (MoAbs) reactive with the CD11/CD18 leucocyte adhesion molecules of granulocytes. Granulocytes showed a decrease in chemotaxis (P < 0.001) and in aggregation (P < 0.01) using various agents as a stimulus. Cytofluorimetric and immunoenzymatic assays with alkaline phosphatase (APAAP) analysis showed decreased expression of the CD11b/CD18 receptor detected by OKM1 (P < 0.001). Despite LFA‐1 and‐CD11a/CD18 was expressed in normal amounts. The studies of upregulation of granulocytes CD11b/CD18 and image analysis of immunochemical preparation (APAAP) demonstrated decreased expression of CD11b/CD18 in granulocytes from MDS compared to controls (P < 0.001). We conclude that granulocyte dysfunction in MDS may be correlated with decreased expression of surface CD11b/CD18 leucocyte adhesion molecules or their structural modification.
UVB radiation exposes fibrinogen binding sites on platelets by activating protein kinase C via reactive oxygen species
Authors: van Marwijk Kooy M. , Akkerman J. W. N. , Asbeck S. van , Borghuis Lizette , van Prooijen H. C.
Published: March 2008

Previous studies have shown that ultraviolet B (UVB) radiation causes platelet aggregation by exposing fibrinogen binding sites via activation of an intracellular mechanism. In the present study we have further investigated the routes of platelet activation following UVB exposure. Evidence is provided that UVB radiation does not activate the platelets via the classical Phospholipase A2 and Phospholipase C routes. Despite this observation, UVB‐induced fibrinogen binding was found to be correlated with a 40% increase in phosphorylated 47 kD protein. Both findings could be completely inhibited in the presence of staurosporine, a potent inhibitor of protein kinase C (PK‐C). In efforts to explain the mechanism of PK‐C activation by UV radiation we found that both UV‐induced PK‐C activation and platelet aggregation were significantly reduced in the presence of specific scavengers for reactive oxygen species including superoxide dismutase and catalase.

We conclude that exposure of platelets to UVB radiation can activate PK‐C via oxygen radicals, resulting in exposure of fibrinogen binding sites and subsequent platelet aggregation.
Gender and response to antilymphocyte globulin (ALG) for severe aplastic anaemia
Authors: Nissen Catherine , Gratwohl Alois , Tichelli André , Stebler Christine , Würsch Andreas , Moser Yolanda , Carbonare Verena Dalle , Signer Erich , Buser Mauro , Ritz Rudolf , Speck Bruno
Published: March 2008

We have evaluated the speed of haematological recovery in 103 severe aplastic anaemia (SAA) patients treated with antilymphocyte globulin (ALG) and followed at our institution for 3–15 years. We found that haemopoietic recovery was significantly delayed in six girls under the age of 10 years. This slow recovery in girls might be explained by their relative inability to release haemopoietic growth factors, granulocyte colony stimulating activity and burst promoting activity, compared to all other sex and age groups. This defect is not explained by disease severity at presentation and thus indicates a functional abnormality of monocytes/macro‐phages and T‐lymphocytes in addition to the deficiency of haemopoietic stem cells. In a multivariate analysis, low factor production and low pretreatment reticulocyte counts turned out to be strong predictors of slow haemopoietic recovery. We conclude that young girls have a particular form of SAA characterized by low haemopoietic factor production and delayed recovery after ALG. They are preferential candidates for early bone marrow transplantation or, if they are not eligible, for treatment with recombinant human haemopoietic growth factors.
Detection of BCR/ABL transcripts in chronic myeloid leukaemia by polymerase chain reaction and DNA enzyme immunoassay: a DNA probe assay without DNA labelling
Authors: Magalini A. R. , Primi D. , Albertini A. , Capucci A. , Rossi G. , Mantero G.
Published: March 2008

The detection of the t(9;22) translocation, typical of chronic myeloid leukaemia (CML), can be accomplished by cytogenetical detection of Philadelphia (Ph1) chromosome or by molecular analysis of the bcr/abl fusion gene with nucleic acid probes after amplification by polymerase chain reaction (PCR). PCR‐based approaches are now widely used for follow up of CML patients during therapy or after bone marrow transplantation (BMT). We describe here a microtitre, colorimetric assay (DNA Enzyme Immunoassay, DEIA) for analysis of t(9;22) translocation after enzymatical amplification of RNA from CML patients. This assay is based on the use of a monoclonal antibody specifically reacting with double stranded DNA, i.e. with hybridized DNA. The assay represents a nonisotopic alternative to other current hybridization assays and requires no modifications of primers, probe or target DNA.
B‐cell lymphoma of recipient origin 9 years after allogeneic bone marrow transplantation for T‐cell acute lymphoblastic leukaemia
Authors: Trimble Malcolm S. , Waye John S. , Walker Irwin R. , Brain Michael C. , Leber Brian F.
Published: March 2008

Summary. A 25‐year‐old woman developed an immuno‐blastic lymphoma 9 years after HLA‐identical allogeneic bone marrow transplantation for T‐cell acute lymphoblastic leukaemia in second remission. The B‐cell origin of the second malignancy was confirmed by gene rearrangement studies. Despite continued donor engraftment, two separate genotypic analyses identified the lymphoma to be of recipient origin. This is the longest latency of a post‐transplant recipient lymphoma yet reported and illustrates that recipient B‐cells may survive the transplant conditioning regimen and undergo malignant transformation in the presence of donor haemopoiesis.
Cytostatic drug resistance: parallel assessment of glutathione‐based detoxifying enzymes, O6‐alkylguanine‐DNA‐alkyltransferase and P‐glycoprotein in adult patients with leukaemia
Authors: Joncourt Franziska , Oberli Aurelia , Redmond Shelagh M. S. , Fey Martin F. , Tobler Andreas , Margison Geoffrey P. , Gratwohl Alois , Buser Kathrin , Cerny Thomas
Published: March 2008

Summary. The levels of several potential indicators of resistance to cytostatic drugs were measured in leukaemic cells of a total of 64 adult patients with acute or chronic Ieukaemias before and during treatment and at relapse or recurrence of disease and compared with those of mono‐nuclear cells from the bone marrow of healthy donors. The resistance factors included glutathione (GSH) and its associated enzymes glutathione‐S‐transferase (GST) and glutathione peroxidase (GPx) as well as O6‐alkyguanine‐DNA‐alkyltransferase (ATase) and P‐glycoprotein. Median values for most parameters were significantly higher in leukaemic cells than in those of normal donors although wide interindividual variation in the values of the various parameters, particularly GST, were seen. P‐glycoprotein was measurable in 12.5% of untreated leukaemias but in none of the normal donors. The values of the parameters in untreated leukaemic patients were not statistically different from those at relapse or during disease progression. However, the median values for GSH, GST and GPx but not ATase in samples from untreated patients were significantly higher than those in samples taken during drug treatment. Patient response, disease‐free survival or duration of remission did not correlate with the values of any of the parameters studied.
Biological activity of recombinant factor VIII variants lacking the central B‐domain and the heavy‐chain sequence Lys713‐Arg740: discordant in vitro and in vivo activity
Authors: Mertens K. , Donath M. J. S. H. , Leen R. W. van , Keyzer‐Nellen M. J. M. de , Verbeet M. Ph. , Bos J. M. Klaasse , Leyte A. , Mourik J. A. van
Published: March 2008

Summary. Recombinant factor VIII variants with overlapping deletions spanning the region Lys713‐Ile1668 have been expressed in mammalian cells, and analysed for biological activity both in vitro and in vivo. Two distinct assay systems were used to measure the activity in vitro. The one‐stage coagulation assay served to assess factor VIII procoagulant activity while a spectrophotometric assay was used for the quantification of factor VIII cofactor activity in factor IXa‐dependent factor X activation. Deletion of the entire B‐domain (Ser741‐Arg1648) resulted in a protein with similar procoagulant and cofactor activity. In contrast, factor VIII‐del(713–163 7), which has a deletion that also comprises the heavy‐chain sequence Lys713‐Arg740, had lost factor VIII procoagulant activity while factor VIII cofactor activity was retained. This functional inconsistency was further addressed by comparing purified factor VIII‐del(713–1637) with factor VIII‐del(868–1562). a mutant with normal in vitro activity.

Kinetic studies of factor Xa formation revealed that higher concentrations of thrombin were required to develop the cofactor activity from factor VIII‐del(713–1637) than needed for factor VIII‐del(868–1562) or plasma factor VIII. The physiological significance of this finding was assessed in dogs with haemophilia A. Both deletion mutants were similar to plasma factor VIII with regard to binding to von Willebrand factor and half‐life and recovery. Employing the cuticle bleeding time model, factor VIII‐del(868–1562) was found to be indistinguishable from plasma factor VIII, whereas factor VIII‐del(713–1637) was less effective. The increased throm‐bin‐resistance of factor VIII‐del(713–1637) thus limits both procoagulant activity and haemostatic efficacy in cuticle bleeding. These observations suggest that the heavy‐chain sequence Lys713‐Arg740, although dispensable for factor VIII cofactor function per se, is involved in the proteolytic activation of factor VIII both in vitro and in vivo.
A polymerase chain reaction‐based method for the detection of trisomy 12 in B chronic lymphocytic leukaemia
Authors: Baker B. W. , Heighway J. , Browett P. J. , Norton J. D.
Published: March 2008

Summary. We report a polymerase chain reaction (PCR)‐based method for detection of clonal trisomy 12 cell populations in B cell chronic lymphocytic leukaemia which is particulary applicable to retrospective analysis or archival material. The method utilizes three restriction enzyme site polymorphisms linked to the KRAS2 locus on chromosome 12 to monitor allele imbalance resulting from duplication of this chromosome. 21/24 unselected chronic lymphocytic leukaemia cases (88%) were informative for at least one polymorphism and of these, eight (33% of total cases) displayed allelic imbalance consistent with trisomy 12.
Recombinant granulocyte‐macrophage colony stimulating factor followed by immunosuppressive therapy for aplastic anaemia
Authors: Doney Kristine , Storb Rainer , Appelbaum Frederick R. , Buckner C. Dean , Sanders Jean , Singer Jack , Hansen John A.
Published: March 2008

Summary. Seventeen patients with aplastic anaemia were treated with recombinant human granulocyte‐macrophage colony stimulating factor (rhGM‐CSF) for 14 d. Nonrespond‐ing patients were then treated with anti‐human thymocyte globulin (ATG), methylprednisolone and oxymetholone. Side‐effects of rhGM‐CSF included fever, nausea and vomiting, diarrhoea, bone pain, headache and chills. Two patients had sustained trilineage haemopoietic recovery after receiving only rhGM‐CSF. Of 11 patients who received immunosuppressive therapy, there was one complete response, two partial responses, one minimal response, and seven nonres‐ponses. Actuarial survival at 2 years is 64%. Early administration of rhGM‐CSF had no apparent effect on subsequent response to immunosuppressive therapy.
An HTLV‐I carrier who developed CD4 + T‐CLL expressed the IL‐2 receptor β chain alone without expressing the α chain
Authors: Mori Naoki , Murakami Shuichi , Oda Susumu , Eto Sumiya
Published: March 2008

Summary. We describe a case of T‐chronic lymphocytic leukaemia (T‐CLL) with monoclonal proliferation of CD3 + 4 + 8 — T cell expressing the interleukin 2 receptor (IL‐2R) β chain without expressing the α chain. Southern blot analysis of T‐cell receptor β chain gene revealed rearranged bands. The serum antibody was positive against human T‐cell lymphotropic virus type I (HTLV‐1)‐associated antigens, but monoclonal integration of proviral DNA was not detected in the leukaemic cells. In accordance with the expression of IL‐2R β chain, the leukaemic cells proliferated in response to exogenous IL‐2 without prior stimulation. However, culture supernatants of these cells did not show IL‐2 activity. Since the role of the IL‐2R β chain in freshly isolated cells from T‐CLL patients is not well understood, detailed analysis of this case would give us valuable information on the role of IL‐2/ IL‐2R system in the proliferation of the leukaemic T cells.
Hydroxyurea treatment for chronic myeloid leukaemia during pregnancy
Authors: Jackson Nicholas , Shukri Ahmad , Ali Kamaruzaman
Published: March 2008

Summary. A patient being treated for chronic myeloid leukaemia with hydroxyurea became pregnant. Despite an increase in the dose of hydroxyurea (to 3 g per day) during the pregnancy, her white blood cell count could only be controlled at about 150 × 109/l. A healthy baby girl was born at 3 7 weeks with normal blood counts and no evidence of congenital abnormality. There are now five reports of the use of hydroxyurea in pregnancy, and where leucapheresis is not available it may be the treatment of choice.
Aberrant CD8 antigen expression in a patient with B chronic lyrnphocytic leukaemia showing unusual disease progression
Authors: Ghosh K. , Sivakumaran M. , Wood J. K.
Published: March 2008

Summary. Co‐expression of T cell antigens in B‐CLL has been well recognized. The commonest T cell antigen known to be expressed in B‐CLL is CD 5, and recent reports suggest that CU 5 expression is associated with good prognosis. Expression of CD8 antigen, however, is much less common with uncertain prognostic significance. We report a case of B‐CLL with aberrant CD8 expression, who had unusual disease progression with a fatal outcome.
Detection of chimaerism after bone marrow transplantation using the double amplification refractory mutation system
Authors: Lo Y‐M. D. , Roux E. , Jeannet M. , Chapuis B. , Fleming K. A. , Wainscoat J. S.
Published: March 2008

Summary. We have developed a highly sensitive double amplification refractory mutation system (double ARMS) for the detection of extremely low levels of mixed chimaerism after bone marrow transplantation. The system we chose for double ARMS analysis is a highly polymorphic region 5′ to the human δ‐globin gene. Double ARMS analysis was found to correlate well with an established minisatellite system and was shown to demonstrate a very low level of mixed chimaerism in cases undetectable by the latter method.