bluuebunny/crossref_metadata_2025 huggingface dataset:
Correlation of serological, quantitative and cell‐mediated functional assays of maternal alloantibodies with the severity of haemolytic disease of the newborn
Authors: Hadley A. G. , Kumpel B. M. , Leader K. A. , Poole G. D. , Fraser I. D.
Published: March 2008

Serum samples containing IgG red blood cell (RBC) antibodies were collected without reference to clinical information from 131 pregnant alloimmunized women. Anti‐D and anti‐K were present in sera from 75 and 20 patients respectively. Antibody titres were determined by indirect antiglobulin test (IAGT), anti‐D levels were measured by AutoAnalyzer, RBC‐binding IgG was quantified using an enzyme‐linked immunosorbent assay (SOL‐ELISA), and functional activities were measured using the monocyte chemiluminescence (CL) test, antibody‐dependent monocytemediated and K cell‐mediated cytotoxicity (ADCC) assays, and rosette formation with U937 cells.

Details of clinical outcomes were obtained retrospectively from 104 pregnancies. Forty‐one babies were ‘antigen‐negative’, and of the remainder, four required top‐up transfusions, 12 required exchange transfusions, three received intrauterine transfusions, and two died in utero. A comparison of test results with severity of haemolytic disease of the newborn (HDN) showed that, provided sera tested were collected within 8 weeks of the expected delivery date, the CL test and the monocyte‐mediated ADCC assay differentiated those D‐positive babies which required exchange transfusions from those unaffected or only mildly affected. The usefulness of results from the AutoAnalyzer and IAGT in predicting disease severity was compromised by the wide range of results from mothers of unaffected babies. This variability was less apparent in the SOL‐ELISA which predicted severe HDN with greater precision. Results from the U937 rosette assay and the K cell‐mediated ADCC assay failed to correlate with disease severity.
The erythrocytes in paroxysmal nocturnal haemoglobinuria of intermediate sensitivity to complement lysis
Authors: Rosse Wendell F. , Hoffman Sharon , Campbell Milton , Borowitz Michael , Moore Joseph O. , Parker Charles J.
Published: March 2008

The sensitivity to lysis by complement of the erythrocytes of 56 patients with paroxysmal nocturnal haemoglobinuria (PNH) was compared to the membrane expression of decay accelerating factor (DAF, CD55), membrane inhibitor of reactive lysis (MIRL, CD59) and acetylcholinesterase (AChE). Most patients (36/50 72% in whom the analysis could be made) appeared to have erythrocytes of intermediate sensitivity to complement in the blood. These cells appeared as a discrete population of cells (PNH II cells), as a ‘tail’ of cells slightly less sensitive than the predominant PNH III cells (previously called PNH IIIb cells), or as a continuous spectrum of cells sensitive to complement. The PNH III cells totally lacked all three proteins (DAF, MIRL, AChE) by flow cytometric analysis whereas PNH I cells appeared to have normal or nearly normal amounts of each. The cells of intermediate sensitivity (PNH II) had coordinately decreased expression of all three proteins; the level of expression of DAF and MIRL paralleled the sensitivity of the cells to the haemolytic action of complement.
Placental lactogen is a haemopoietic hormone
Authors: Berczi Istvan , Nagy Eva
Published: March 2008

The anaemia and thrombocytopenia of hypophysectomized (Hypox) rats could be corrected readily by daily treatment with human placental lactogen. Spontaneous DNA synthesis in the bone marrow of Hypox rats was grossly impaired, which was also normalized by placental lactogen. Human placental lactogen exerted a direct mitogenic effect on rat bone marrow cells in vitro. These results indicate that placental lactogen is a potent haemopoietic hormone.
Primary human acute myeloblastic leukaemia: an analysis of in vitro granulocytic maturation following stimulation with retinoic acid and G‐CSF
Authors: Colombat Ph. , Santini V. , Krefft Delwel J. , Bredmond J. L. , Löwenberg B.
Published: March 2008

Acute myeloid leukaemia (AML) is characterized by the inability of myeloid cells to reach terminal maturation. We examined to what degree granulocytic maturation could be achieved by stimulating AML blast cells in an in vitro serum‐free system with a combination of granulocyte‐colony stimulating factor (G‐CSF) and all‐trans‐retinoic acid (RA). Specimens from 41 AML patients were cultured for 7 d and then examined for cytochemistry (myeloperoxidase, Sudan Black, napthyl‐ASD‐chloracetate esterase, periodic acid Schiff) and nitroblue tetrazolium reduction. The expression of CD11a, CD11b. CD11c, CD15, CD18, CD10, CD24 and B13‐3 membrane antigens was also evaluated. Morphological and cytochemical studies were also performed after AML colony culture and culture of normal bone marrow cells (NBM). The comparative analysis of the panel of parameters was indicative of granulocytic maturation although to different degrees.

The cells from 25/41 cases showed morphologic maturation (May‐Grünwald‐Giemsa). A positive correlation was evident between morphological maturation and CD11b expression (11/22 patients) as well as that of CD11c and CD15 (6 patients). Napthyl ASD chloroacetate esterase and PAS stainings also correlated with morphology (in 10/22 and 10/24 patients respectively). Nevertheless, the pattern of granulocytic maturation was remarkably variable among the 41 cases examined. The cells from only a few patients acquired the full spectrum of granulocytic markers. The comparison with normal bone marrow blasts indicates that serum‐free culture conditions can, per se, limit granulocytic maturation, but it also confirms the intrinsic inability of AML cells to attain complete maturation in response to two potent granulocytic inducers.
Prognostic importance of Sudan Black positivity: a study of bone marrow slides from 1386 patients with de novo acute myeloid leukaemia
Authors: Hoyle C. F. , Gray R. G. , Wheatley K. , Swirsky D. , Bastos M. de , Sherrington P. , Rees J. K. H. , Hayhoe F. G. J.
Published: March 2008

Analysis of bone marrow slides from 1386 patients entered into the Medical Research Council's 8th and 9th trials in Acute Myeloid Leukaemia confirmed that features associated with differentiation in blast cells, in particular increasing Sudan Black (SB) positivity, were the most important morphological features for predicting remission achievement (P=0·002) and hence survival (P<0·0001). SB positivity was also weakly predictive of remission duration (P=0·05). A low complement of maturing granulocytes was associated with early induction death and a high percentage of blasts with shorter remissions. The few patients with acute promyelocytic leukaemia (FAB M3) had a high haemorrhagic death rate during induction and a low relapse rate. Apart from this, lineage involvement was not predictive of outcome. Multiple lineage leukaemias, in particular those with megakaryocytic and/or erythroid involvement, which had been reported previously to have a poor prognosis, did not have any worse remission rates in this series. When more than one cell line was involved, no combination with particularly good or poor prognosis could be identified. Multivariate analysis suggested that percentage SB positivity was adequate on its own to divide granulocytic leukaemias into poorly differentiated (<50% SB +ve) and well‐differentiated groups (50% or more SB + ve) without the need for further measurements. This simple and reproducible test was strongly predictive of resistant disease but not of induction deaths. It was of considerably greater prognostic value—and was less open to inter‐observer disagreement—than the FAB criteria which are usually used to classify granulocytic lineage leukaemias into the M1 and M2 subgroups. It is proposed that ≥ 50% of blasts with SB positivity should replace blasts > 10% of maturing myeloid cells for this sub‐categorization between Ml and M2.
A cathepsin‐like cysteine proteinase proaggregating activity in thrombotic thrombocytopenic purpura
Authors: Falanga A. , Consonni R. , Ruggenenti P. , Barbui T.
Published: March 2008

The platelet aggregating activity (PAA) of serum and plasma from thrombotic thrombocytopenic purpura (TTP, eight patients) and haemolytic uraemic syndrome (HUS, three patients) was investigated, Sample‐induced aggregation of washed platelets and the parallel 3H‐serotonin release were studied both in the presence and the absence of purified von Willebrand factor (vWF) or defibrinated cryoprecipitate, according to Kelton et al (1987). All the samples but two were positive in the test, vWF being indispensable for aggregation in all but two of the active cases. Samples from five patients during remission were ineffective under the same conditions, as were 14 controls from healthy donors.

An inhibition study using four cysteine proteinase inhibitors (1 mM iodoacetamide, 0·5 mM E‐64, 0·1 mM antipain and 0·1 mM leupeptin) and two serine proteinase inhibitors (200 U/ml aprotinin and 0·2 U/ml hirudin) confirmed that TTP/HUS's PAA behaved enzymatically as a cysteine proteinase, as described (Murphy et al, 1987a). These results were confirmed on a protein preparation obtained from pooled plasmas after AI(OH)3 adsorption, ammonium sulphate precipitation, binding to and elution from a p‐(chloromercuri)benzoate (PCMB) affinity resin.

To further characterize this activity we investigated whether it was a calcium‐dependent neutral protease or a lysosomal cathepsin by testing the sensitivity to a series of peptidyl diazomethyl ketones, synthetic inhibitors specifically designed for cysteine proteinases. We confirm here the presence of a circulating platelet aggregating cysteine proteinase in TTP/HUS and provide evidence for the first time that this activity behaves like a lysosomal cathepsin.
Diagnosis of paroxysmal nocturnal haemoglobinuria using immunophenotyping of peripheral blood cells
Authors: Schubert Jörg , Alvarado Marco , Uciechowski Peter , Zielinska‐Skowronek Margot , Freund Matthias , Vogt Hans , Schmidt Reinhold E.
Published: March 2008

Paroxysmal nocturnal haemoglobinuria (PNH) is now generally accepted as a disease in which bone marrow derived cells are deficient in phosphatidylinositolglycan (PIG)‐anchored surface molecules. A series of new monoclonal antibodies detecting PIG‐anchored surface structures on human leucocytes (CD48, CD55, CD59) has recently been described. In the present study 12 patients with the diagnosis PNH and a positive Ham test were examined for PIG‐anchored surface antigen expression on various cell lineages using immunofluorescence. In all patients deficient cells were detected in erythrocyte, granulocyte and monocyte analysis. A deficient lymphocyte subset was also observed in all but one of these patients. Using two‐colour analysis, all lymphocyte subpopulations such as T, B and NK cells were found to be affected. In addition, peripheral blood cells of 22 patients with severe aplastic anaemia (SAA) were tested for the PIG‐anchoring defect. In five of these patients the defect was detected, and in four of the five the lack of PIG‐anchored molecules was confined to the granulocyte and monocyte lineages apparently without affecting the erythrocytes. The results of these studies demonstrate that cytofluorographic testing of peripheral blood cells provides a simple and reliable method for establishing the diagnosis of PNH. Furthermore, especially in the case of aplastic anaemia patients, the sensitivity of immunophenotyping might be superior to conventional laboratory tests.
Lewis antigen expression in benign and malignant tissues from RBC Le(a—b—) cancer patients
Authors: Langkilde Niels C. , Wolf Hans , ØRntoft Torben F.
Published: March 2008

Eight red blood cell (RBC) Le(a—b—) individuals were selected from a series of patients with bladder or colon cancer. Defined by the presence or absence of α1‐4‐l‐fucosyltransferase activity in saliva, four of these patients were characterized as non‐genuine Lewis negative [RBC Le(a—b—) with α1‐4‐l‐fucosyltransferase activity in saliva], and four as genuine Lewis negative [RBC Le(a—b—) with no α1‐4‐l‐fucosyltransferase activity in saliva]. Stainings of paraffin embedded formalin fixed tissue sections for Lea and Led antigens were performed by means of an indirect immunohistochemical method on all malignant and benign tissue previously removed from these eight patients. Leb antigens were always expressed independently of both the Lewis and the secretor status of the individual. Lea antigens, on the other hand, showed a different staining pattern. Although primarily expressed in non‐genuine Le(a—b—) individuals, Lea antigens were expressed in genuine Le(a—b—) individuals as well—to a limited extent, but still detectable. Thus, these findings seem to show that the Lewis antigen expression is tissue dependent, and it is not possible to predict tissue Lewis antigen expression by merely examining erythrocytes or saliva.
Comparisons of human, rat and mouse erythropoietins by isoelectric focusing: differences between serum and urinary erythropoietins
Authors: Tam R. C. , Coleman S. L. , Tiplady R. J. , Storring P. L. , Cotes P. Mary
Published: March 2008

Isoelectric focusing (IEF) in the pH range 2·5‐5·0 has been used to compare the immunoreactive (ir) erythropoietin (Epo) in paired samples of serum and urine from three patients, two with idiopathic aplastic anaemia and one with paroxysmal nocturnal haemoglobinuria and also from three anaemic rats. Serum samples only were also examined from two further patients with aplastic anaemia and from three mice, made anaemic (like the rats) by irradiation and phenylhydrazine treatment. Most of the ir‐Epo recovered after IEF was found in the pH range 2·5–3·9. For the sera, the proportion of more acidic ir‐Epo with pl < 3·0 recovered after IEF increased from human to rat to mouse. Human sera contained a greater proportion of ir‐Epo with pl > 3·4 than rat or mouse sera. For the urines, the distribution of ir‐Epo by IEF was similar between human and rat. For both species, the proportion of ir‐Epo with pl< 3·0 recovered after IEF was greater in urine than in the paired serum samples. The Second International Reference Preparation of Human Urinary Epo differed from the Epo in unextracted human urine in that there was a lower proportion of ir‐Epo with pI < 3·0. The differences observed between serum and urinary Epo are of particular interest because only the urinary form of native human Epo has ever been purified, and because this was used to compare native with rDNA‐derived Epo.
Non‐radioactive detection of immunoglobulin and T cell receptor gene rearrangement in acute lymphoblastic leukaemia
Authors: Martin Gerald , Lawlor Emer
Published: March 2008

Rearrangements of the heavy chain immunoglobulin gene and T cell receptor β gene were investigated in 25 patients suffering from precursor B cell acute leukaemia and six patients suffering from T cell acute leukaemia using biotinylated DNA probes. All precursor B acute leukaemia patients had IgH gene rearrangements and 63% of those studied also had TCRβ gene rearrangements. All T cell acute leukaemia patients had TCRβ gene rearrangements and germline IgH configuration. Dilution experiments indicated that DNA from leukaemic cells representing 1–2% of a tested sample could be detected using this technique which compares favourably to radioactive DNA probes.
Book Review
Authors: Ludlam C. A.
Published: March 2008

Platelet Heterogeneity. Edited by John Martin and Anthony Trowbridge.
Homotypic cell aggregations of human myeloma cells with ICAM‐1 and LFA‐1 molecules
Authors: Kawano Michio M. , Huang Naihui , Tanaka Hideo , Ishikawa Hideaki , Sakai Akira , Tanabe Osamu , Nobuyoshi Masaharu , Kuramoto Atsushi
Published: March 2008

Some myeloma cells freshly isolated from bone marrow aspirates in human myelomas and some myeloma cell lines formed spontaneous cell aggregations in vitro (homotypic cell aggregations). In order to clarify the surface molecules involved in homotypic cell aggregations and physiological roles of these cell aggregations, we investigated the expressions of intercellular adhesion molecule 1 (ICAM‐1) and lymphocyte function‐associated antigen 1 (LFA‐1) on 20 samples of freshly isolated myeloma cells and three myeloma cell lines and the effect of anti‐ICAM‐1 and anti‐LFA‐1α antibodies on myeloma cell proliferation in vitro. All myeloma cells that we tested expressed ICAM‐1 on their surface. Among them, myeloma cells that strongly coexpressed LFA‐1α, formed homotypic cell aggregates in vitro. These spontaneous cell aggregations were completely released by adding either anti‐ICAM‐1 or anti‐LFA‐1α antibody. During short‐term culture, spontaneous proliferation of myeloma cells in vitro and their proliferative responses to recombinant interleukin‐6 (rIL‐6) were not affected by pretreatment of myeloma cells with anti‐ICAM‐1 or anti‐LFA‐1α antibody. Therefore these data suggest that homotypic cell aggregation of myeloma cells is mediated by ICAM‐1 and LFA‐1 molecules, but myeloma cell proliferation may not be modulated by these adhesion molecules during shortterm cultures.
Recurring mutations at CpG dinucleotides in the region of the von Willebrand factor gene encoding the glycoprotein Ib binding domain, in patients with type IIB von Willebrand's disease
Authors: Lillicrap David , Murray Elizabeth W. , Benford Kerry , Blanchette Victor S. , Rivard George E. , Wensley Richard , Giles Alan R.
Published: March 2008

The mutant von Willebrand factor (vWf) molecule in type IIB von Willebrand's disease (vWd) has an increased binding affinity for the platelet receptor glycoprotein Ib (GpIb). In previous studies we have confirmed genetic linkage of this phenotype to the vWf gene and in this report we document three recurring missense mutations in the region of the gene that encodes the GpIb binding domain. Two families with type IIB vWd were found to have an arginine to tryptophan substitution at residue 543, three families had a valine to methionine substitution at residue 553, and one kindred had an arginine to glutamine change at amino acid 578. None of these sequence changes were found in 200 normal vWf genes and within each of the six families the mutations were only found in affected subjects. This is strong circumstantial evidence in support of these substitutions representing the disease causing mutations in these families. All three of these substitutions have occurred at CpG dinucleotide sequences, and their polymorphic associations indicate that they represent recurring new mutations. Missense mutations at these sites may represent the underlying genetic pathology in a large number of type IIB vWd families.
Colony‐cluster ratio and cluster number in cultures of circulating myeloid progenitors as indicators of high‐risk myelodysplasia
Authors: Tennant G. B. , Bowen D. T. , Jacobs A.
Published: March 2008

Circulating myeloid progenitors were assayed in 172 normal subjects and 147 patients with myelodysplastic syndrome (MDS). Patients whose cultures had colony/cluster ratios (CCR) < 0·3 had significantly shorter survival periods than comparable patients with CCR > 0·3. A second prognostic indicator, which complemented CCR, was identified in patients with < 5% blasts. Median survival was significantly reduced in patients with > 15 clusters/ml blood despite colony and cluster numbers being predominantly within the normal range. Characteristic differences were found in three FAB groups large enough to allow statistical analysis. Survival amongst patients with refractory anaemia with excess of blasts (RAEB) was related to CCR and was independent of cluster number. Amongst sideroblastic patients (SA) survival related only to cluster number. Refractory anaemia (RA) patients included individuals in both high‐risk groups with only three patients out of 64 showing both features. Amongst all the MDS patients, those with CCR > 0·3 and < 15 clusters/ml blood formed a low‐risk group (n= 60) with a relatively good prognosis of whom 85% survived the study period (median duration 938 d) including 94% of those in this group with < 5% marrow blast cells.
Primary thrombocythaemia in pregnancy
Authors: Beard J. , Hillmen P. , Anderson C. C. , Lewis S. M. , Pearson T. C.
Published: March 2008

Nine pregnancies in six patients with primary thrombocythaemia are reported. Eight pregnancies resulted in the delivery of normal infants. One pregnancy ended in spontaneous abortion at 7 weeks gestation. One pregnancy was complicated by superficial thrombophlebitis and a postpartum haemorrhage. We suggest that pregnancy in patients with primary thrombocythaemia can have a favourable outcome, but requires close monitoring. Administration of aspirin during pregnancy may be of benefit.
A comparison of sickle cell syndromes in Northern Greece
Authors: Christakis J. , Vavatsi N. , Hassapopoulou H. , Angeloudi M. , Papadopoulou M. , Loukopoulos D. , Morris J. s. , Serjeant B. E. , Serjeant G. R.
Published: March 2008

Haematological and clinical characteristics have been examined in 30 patients with homozygous sickle cell (SS) disease, 28 with sickle cell‐beta° thalassaemia, and 21 with sickle cell‐beta+ thalassaemia. The latter could be divided into three groups on their molecular basis and HbA levels, four subjects with an IVS‐2 nt 745 mutation having 34% HbA (designated Sbeta+ thalassaemia type I), 14 subjects with an IVS‐1 nt 110 mutation having 8–15% HbA (designated Sbeta+ thalassaemia type 11). and three subjects with an IVS–1 nt 6 mutation having 20–25% HbA (designated Sbeta+ thalassaemia type III). Comparisons were conducted between SS disease, Sbeta° thalassaemia, and Sbeta+ thalassaemia type II. Compared to SS disease, both thalassaemia syndromes had higher HbAr levels and red cell counts and lower mean cell haemoglobin content (MCHC), mean cell volume (MCV) and MCH, and Sbeta° thalassaemia had higher HbF and reticulocyte counts. Compared to Sbeta° thalassaemia. Sbeta+ thalassaemia had a higher haemoglobin and MCHC. Clinically, persistence of splenomegaly was more common in Sbeta° and Sbeta+ thalassaemia type II compared to SS disease. Few significant differences occurred between SS disease, Sbeta° and Sbeta+ thalassaemia type II in Northern Greece suggesting that the 8–15% HbA in the latter condition was insufficient to modify the clinical course.
Combination therapy with interferon alpha‐2b plus low‐dose interferon gamma in pretreated patients with Ph‐positive chronic myelogenous leukaemia
Authors: Wandl Ursula B. , Kloke Otto , Nagel‐Hiemke Marianne , Moritz Thomas , Becher Reinhard , Opalka Bertram , Holtkamp Wilhelm , Bartels Heinrich , Seeber Siegfried , Niederle Norbert
Published: March 2008

Between March 1988 and July 1990, 28 adults with chronic myelogenous leukaemia (CML) were treated with a combination of recombinant human interferon (IFN) alpha‐2b s.c. (initial dose 4 x 106 U/m2) and recombinant human IFN gamma s.c. (50 μg totally) daily. All patients were in chronic phase disease and had been treated previously with chemotherapy or bone marrow transplantation. A complete haematologic remission was achieved in three patients (11%), a haematologic remission in 12 patients (43%), and a partial haematologic remission in seven patients (25%). Six patients did not respond to this schedule. Acute side‐effects were flu‐like symptoms, fever and chills. During long‐term treatment six patients developed poly‐arthralgia. Haematotoxicity WHO grade III occurred in three patients, and WHO grade IV in two patients. One patient developed psychosis, and in another patient an exacerbation of a pre‐existing sarcoidosis was observed. We conclude that this combination is tolerable and effective in inducing haematological remissions in pretreated CML patients.
Shedding of CD9 antigen by bone marrow cells from patients with acute lymphoblastic leukaemia
Authors: Komada Yoshihiro , Zhang Shao‐Li , Zhou Yan‐Wen , Shibata Takeo , Azuma Eiichi , Sakurai Minoru
Published: March 2008

The levels of soluble CD9 antigen released into spent medium from bone marrow (BM) cells were assayed using a unique enzyme‐linked immunosorbent assay. We demonstrated that a considerable amount of soluble CD9 antigen was consistently detected in the spent medium from CD9+ leukaemic blasts, but little from normal or regenerating BM cells. The sensitivity and specificity of CD9 antigen assay were tested. First, BM cells taken from patients in complete morphologic remission (CR) were studied, and the distinct level of CD9 antigen was detected in 16·3% of the cases. Second, the outcome of leukaemic patients with significant shedding of CD9 antigen from BM cells while in CR was investigated: all patients have developed systemic relapse within 5–24 (median 15·4) weeks. In contrast, 24/34 patients without the increase of CD9 antigen levels are still in CR (follow‐up 19–149 weeks, median 56·2 weeks). Only 10 patients in this group have relapsed so far. No false‐positive results were detected with this sensitive assay for soluble CD9 antigen, and the introduction of appropriately planned prospective studies is justified on the basis of these observations.
PCR analysis of HIV‐1 sequences and differential immunological features in seronegative and seropositive haemophiliacs
Authors: Chelucci C. , Hassan H. J. , Gringeri A. , Macioce G. , Mariani G. , Santagostino E. , Testa U. , Vulcano F. , Mannucci P. M. , Peschle C.
Published: March 2008

We have compared the immunological features of two matched groups of seronegative and seropositive haemophilia A individuals. Both groups were exposed from 1981 to 1985 to comparable amounts and batches of FVIII concentrates not subjected to virus inactivation procedures, and had therefore a 100% probability of receiving HIV‐contaminated material. The presence of proviral HIV‐1 sequences was evaluated by PCR in the DNA from peripheral blood lymphocytes and/or monocytes. After hybridization with specific probes, DNA from all seropositive haemophiliacs revealed HIV sequences; no HIV sequences were observed from the DNA of seronegative patients, even after two rounds of amplification, thus suggesting that these patients were not affected by a latent HIV infection.

Seronegative/PCR‐ and seropositive/PCR+ patients showed a normal and reduced number of CD4+ lymphocytes, and a slight and marked increase of CD8+ cells respectively. Activated T cells expressing the HLA‐DR antigen were elevated in both groups. Interestingly, a significant reduction of circulating CD56+/CD3‐ NK lymphocytes was observed only in seropositive haemophiliacs, whereas NK lymphocytes with CD56+/CD3+ phenotype were within normal levels in both groups. In seropositive patients no correlation was found between the number of CD4+ and CD56+/CD3‐ lymphocytes. The marked reduction of CD56+/CD3‐ lymphocytes observed in seropositive haemophiliacs in addition to the CD4+ cell depletion may represent a key pathogenetic factor which facilitates the onset and/or the progression of HIV‐1 infection in haemophiliacs, and is related to the capacity of HIV to infect NK cells.
A prospective study to determine the safety of omitting the antiglobulin crossmatch from pretransfusion testing
Authors: Heddle Nancy M. , O'Hoski Pamela , Singer Joel , McBride John A. , Ali Mahmoud A. M. , Kelton John G.
Published: March 2008

Transfusion medicine laboratories routinely perform a series of pretransfusion serological tests including: ABO grouping, Rh typing, and investigation of the recipient's serum to detect antibodies against blood group antigens (antibody screen). As a final check, most laboratories also perform a crossmatch in which the recipient's serum is incubated with the donor's red cells followed by the addition of an antiglobulin reagent (antiglobulin crossmatch). The need for the antiglobulin crossmatch when the antibody screen is negative has been questioned because there are few antibodies that are detected by this test. Such antibodies are usually directed against low incidence antigens that are not expressed on the screening cells and in many cases the clinical importance of these antibodies is uncertain. For these reasons, we performed a prospective study in which patients requiring red cell transfusion had a group and screen performed. If the antibody screen was negative the antiglobulin crossmatch was omitted. Following the transfusion of the blood, the antiglobulin crossmatch was performed to look for any potential incompatibility. All patients were monitored both serologically and clinically.

Over the 2‐year interval of the study 9128 patients were entered. There were 8936 patients (97·9%) with a negative antibody screen and 26·9% (2404 patients) were transfused a total of 10899 red cell concentrates. The antiglobulin crossmatch performed after the transfusion indicated that 168 red cell concentrates (1·5%) would have been incompatible if the antiglobulin crossmatch had been performed pretransfusion. These 168 red cell concentrates were transfused to 119 patients during 130 transfusion episodes (defined as all transfusions administered within 24 h). Of the 130 transfusion episodes, 79·2% (103/130) were false positive laboratory results. There were 27 transfusion episodes where the antiglobulin crossmatch on blood transfused was positive due to an IgG antibody. Even though these transfused red cell concentrates were designated incompatible by the antiglobulin crossmatch, none of the patients receiving this blood had clinical or serological evidence of haemolysis. We concluded that the antiglobulin phase of the crossmatch can be omitted from pretransfusion testing without putting patients at risk.
Serum erythropoietin in the diagnosis of polycythaemia and after phlebotomy treatment
Authors: Birgegård Gunnar , Wide Leif
Published: March 2008

Serum erythropoietin (S‐Epo) was measured with a radiommunoassay method in 36 patients with polycythaemia vera, 17 patients with secondary polycythaemia and 14 patients with relative polycythaemia. The diagnoses were made without the aid of the S‐Epo values. It was found that S‐Epo was below the reference range in 34/36 patients with polycythaemia vera (mean 2·1 ± 1·0 U/1), elevated in all cases of secondary polycythaemia (mean 121·7 ± 242 U/l) and normal in all but one of the cases with relative polycythaemia (mean 7·0 ± 2·5 U/l). Previous studies of S‐Epo levels in the differential diagnosis of polycythaemia have shown significant differences between the means of the groups but a considerable overlap. After phlebotomy treatment to normal haematocrit levels, the S‐Epo levels remained subnormal in most of the polycythaemia vera patients even after 18 months at a haematocrit around 45%. Two patients who had been kept at normal haematocrit for 6 and 7 years both had subnormal S‐Epo.

We conclude that with an Epo assay method of high sensitivity and specificity it is possible to differentiate between different forms of polycythaemia with a high degree of certainty, even between patients with relative polycythaemia and polycythaemia vera patients.

The reason why S‐Epo remains low in spite of a normal haematocrit in treated polycythaemia vera patients is not known.
Monocytes enhance endothelial von Willebrand factor release and prostacyclin production with different kinetics and dependency on intercellular contact between these two cell types
Authors: Hakkert Betty C. , Rentenaar Jeannette M. , Van Mourik Jan A.
Published: March 2008

Summary Human umbilical vein endothelial cells cultured on a collagen lattice were used to study the effects of the interaction between human monocytes and endothelial cells on endothelial von Willebrand Factor (vWF) release and prostacyclin (PGI2) production by these cells. The effects of monocytes were compared with those of other leucocytes, conditioned media from monocytes, and agonists such as interleukin 1 (IL‐1) and the phorbol ester PMA. Because the cell culture system used allows simultaneous analysis of the lumenal and ablumenal compartment of endothelial cell monolayers, we also studied into which direction these products were released by endothelial cells. Under quiescent conditions the concentration of vWF in the ablumenal compartment was about three‐fold higher than that in the lumenal compartment, whereas PGI2 was equally distributed between the two compartments. Direct cell‐cell contact between purified monocytes and endothelial cells strongly enhanced both vWF release and PGI2 synthesis, in a dose‐dependent and monocyte‐specific manner. The monocyte‐induced enhancement of PGI2 production, however, was much earlier in onset than that of vWF. Secretory products from monocytes also enhanced endothelial PGI2 synthesis, although to a lesser extent than with monocytes that were in direct contact with endothelial cells. In contrast, the mono‐cyte‐induced enhancement of endothelial vWF release was completely dependent on the direct interaction between monocytes and endothelial cells.
In vivo targeting of Hodgkin and Reed‐Sternberg cells of Hodgkin's disease with monoclonal antibody Ber‐H2 (CD30): immimohistological evidence
Authors: Falini B. , Flenghi L. , Fedeli L. , Broe M. K. , Bonino C. , Stein H. , Düurkop H. , Bigerna B. , Barbabietola G. , Venturi S. , Aversa F. , Pizzolo G. , Bartoli A. , Pileri S. , Sabattini E. , And R. Palumbo , Martelli M. F.
Published: March 2008

Summary The ability of the Ber‐H2 (CD30) monoclonal antibody (mAb) to target in vivo Hodgkin (H) and Reed‐Sternberg (R‐S) cells was investigated in six patients with advanced Hodgkin's disease (HD). The patients were injected with scaled‐up quantities of‘cold’Ber‐H2 mixed‐up to a small dose of 131I‐labelled Ber‐H2. and in vivo binding of the antibody to H and R‐S cells was assessed by immunohistologi‐cal analysis of tumour biopsies and immunoscintigraphy. Only 50% of tumour sites were imaged at scintigraphy by the 131I‐labelled Ber‐H2. In contrast, immunohistological studies on tissue biopsies, taken 24–72 h following the mAb injection, showed that H and R‐S cells in all tumour sites, including those that were not imaged by immunoscintigraphy, were specifically and strongly labelled in vivo by the injected Ber‐H2, at a dose as low as 30–50 mg of antibody. In vivo binding of a single dose of Ber‐H2 mAb to H and R‐S cells did not result in any anti‐tumour effect. The excellent in vivo targeting of H and R‐S cells with the Ber‐H2 mAb may have been the result of multiple favourable factors, including: (a) the restricted expression of the CD30 antigen in normal human tissues; (b) the low level of soluble CD30 in the serum of our patients; and (c) the high affinity of the Ber‐H2 mAb for the CD30 molecule. The immunohistological results presented in this study provide a strong argument for using the Ber‐H2 mAb as a carrier for delivering cytotoxic agents (isotopes or toxins) to neoplastic cells of HD refractory to conventional therapy.
Patterns of development of tachyphylaxis in patients with haemophilia and von Willebrand disease after repeated doses of desmopressin (DDAVP)
Authors: Mannucci Pier Mannuccio , And Donato Bettega , Cattaneo Marco
Published: March 2008

Summary When patients with mild haemophilia or von Willebrand disease (vWD) are repeatedly treated with desmopressin (DDAVP) at relatively short time intervals, some of them may become less responsive or unresponsive. The development of tachyphylaxis would limit the usefulness of DDAVP in clinical management of these patients. On the other hand, tachyphylaxis is not consistent, and its patterns of development are unknown. The aim of this study was to evaluate in controlled conditions the occurrence of tachyphylaxis by giving intravenous DDAVP (0.3 μg/kg) on four consecutive days to a selected group of patients with mild haemophilia A (n = 22) and type I vWD (n = 15). After each dose, we measured parameters known to change after DDAVP, i.e. factor VIII coagulant activity, bleeding time, von Willebrand factor antigen, ristocetin cofactor and tissue‐type plasminogen activator antigen. We found that on average the responses obtained after the second dose of DDAVP were approximately 30% less than those obtained after the first, but were not further reduced after the third and fourth dose. At all time intervals after DDAVP, patients with vWD responded relatively better than patients with haemophilia, and there were fewer vWD patients who responded poorly or became unresponsive. In vWD patients there were no significant changes in the bleeding time responses and in blood pressure and heart rate. The clinical implications of these findings are that repeated doses of DDAVP can be given efficaciously to many patients (particularly to those with vWD), even though responses lower than those seen after the first dose should be expected.
Identification of autoantigens in autoimmune haemolytic anaemia by a non‐radioisotope immunoprecipitation method
Authors: Barker R. N. , Casswell K. M. , Reid M. E. , Sokol R. J. , Elson C. J.
Published: March 2008

Summary In human autoimmune anaemia (AIHA), warm autoantibodies frequently appear to have serological specificity for the Rh complex, but to date, immunochemical techniques have failed to demonstrate that the antibodies react with Rh‐associated polypeptides. We describe the immunoprecipitation of red blood cell (RBC) autoantigens. using biotin labelling and a luminescent detection method. In three cases of warm AIHA, a band of 32 kD and a diffuse zone of 38–51 kD or 40–51 kD were specifically precipitated by eluted RBC autoantibody. This pattern corresponds closely with that precipitated by two Rh‐specific monoclonal antibodies, BRIC 207 and AB5. Antibody from the three eluates also showed serological specificity for the Rh complex in a haemagglutination assay against a panel of RBC with a range of Rh phenotypes, including rare ‐D‐/‐D‐and Rh null cells. Eluted autoantibody from another warm AIHA patient immunoprecipitated a peptide of 67 kD that did not correspond in apparent molecular mass either with Rh‐associated bands, or with major RBC membrane proteins or sialoglyco‐proteins (SGP). The haemagglutination assay showed that this eluate contained both Rh‐specific and Rh‐unrelated antibody. Warm autoantibody eluted from the RBC of a clinically normal, but direct antiglobulin test positive, blood donor was serologically unreactive with the Rh complex, and immunoprecipitated unknown peptides of 26, 29, 35,48 and 51 kD, together with a band of 90 kD that comigrated with SGPα2 (glycophorin A). In six further warm AIHA cases, no antigens were precipitated by autoantibody‐containing RBC eluates. Overall, the results demonstrate that autoantibodies bind to Rh polypeptides in some, but not all, patients with warm AIHA, suggesting that the aetiology of the disease may vary.
Hereditary stomatocytosis: phenotypical expression of sodium transport and band 7 peptides in 44 cases
Authors: And A. Kanzaki , Yawata Y.
Published: March 2008

Summary The clinical heterogeneity and the role of red cell membrane protein band 7 in membrane transport were studied in 44 patients with hereditary stomatocytosis with normal red cell membrane lipids. These patients were arbitrarily categorized into three phenotypes, based on the extent of sodium influx: hereditary stomatocytosis Type I: with markedly increased Na influx (8.90±3.39 mmol/lRBC/h); Type II: with moderately increased Na influx (2.10±0.79) and Type III with normal Na influx (1.31±0.13). The three groups of patients were compared with normal controls (1.29±0.14). The extent of anaemia and jaundice was almost identical in the three groups in the presence of nearly the same degree of stomatocytosis (I: 54.8±10.7%, II: 38.8±12.8, and III: 40.2±10.8). Approximately one third of the cases (14/44) with hereditary stomatocytosis showed no overt haemolysis even with marked stomatocytosis. Cell hydration was abnormal in Type I (MCV 119.6±8.5 fl, MCHC 29.3±1.8%) but normal in Types II and III (MCV 98.2±11.7, 94.1±8.5; MCHC 34.4 ± 2.1, 34.5 ± 2.2). These results indicate that there was no correlation between the extent of Na influx and either the degree of stomatocytosis or the extent of overt haemolysis.

The role of band 7 in membrane transport was also studied. Three components (30 kD, 28 kD and 26 kD polypeptides) of band 7 were analysed by SDS‐PAGE and NEPHGE/SDS‐PAGE, and the content of these polypeptides were expressed as the ratio to band 5. The 30 kDa polypeptide in the three groups was nearly identical to that in normal controls (12.3 ±4.0), except for non‐haemolysing patients in Type II. The 28 kD peptide was also decreased in five out of nine cases of Type II (25.7 ± 5.6) as compared with normal controls (32.9 ± 3.6) and cases of Type I (35.8±2.8) and Type III (32 ± 7 ± 2.9). No deficiency of this peptide was noted in Type I patients. No correlation was observed between the content of the 28 kD polypeptide and Na influx (r=0.416), but the 26 kD polypeptide tended to be elevated in cases with overt haemolysis. These results suggest that band 7 may not be essentially involved in the formation of stomatocytic changes, although the presence of subtle defects in band 7 structure and function may not be ruled out. The present findings provide an important starting point to initiate further extensive investigations.
A large erythroid spectrin β‐chain variant
Authors: Johnson Robert M. , Ravindranath Y. , Brohn Fred , Hussain Mukarram
Published: March 2008

Summary. A large variant of erythrocyte β‐spectrin was found in a child presenting with hereditary elliptocytosis and anaemia. This polypeptide was phosphorylated, cross‐reacted with normal β‐spectrin in immunoblotting and formed a dimer with α‐spectrin that co‐purified with normal xβ dimer. The molecular weight was estimated to be 330 kD by SDS gel electrophoresis, which is 84 kD (35%) larger than the normal β‐chain. This variant has been tentatively named spectrin Detroit (βDetroit). Tryptic digests demonstrated a coexisting α‐spectrin variant SpαI/65 in the propositus, his father and a paternal uncle. Anaemia and elliptocytosis was associated with SpαI/65 rather than βDetroit, since other family members with βDetroit in whom α‐spectrin was normal had no morphological or clinical abnormalities. Family members were identified who had normal α‐spectrin but were heterozygotic for the large β‐spectrin. Their erythrocyte membranes were more rigid and fragile than normal. The fragility is probably a consequence of both weaker dimer association and spectrin deficiency. Variant spectrin dimers (αβDetroit) had a reduced self‐association constants. Binding to ankyrin was normal. Instability of βDetroit during erythropoiesis is suggested by the fact that it comprises only 25% of the β‐spectrin in βDetroit heterozygote erythrocytes, and total spectrin was reduced by 20%. Although βDetroit has some functional defects, this 84 kDa insert in erythrocyte spectrin is compatible with nearly normal function.
Distinct patterns of chromosome abnormalities characterize childhood non‐Hodgkin's lymphoma
Authors: Mikraki Vasiliki , Jhanwar Suresh C. , Filippa Daniel A. , Wollner Norma , Chaganti R. S. K.
Published: March 2008

Summary. We report here cytogenetic studies of a series of 23 childhood non‐Hodgkin's lymphomas (NHL), a group that has previously not been subjected to detailed cytogenetic analysis. Combining our results with data from 25 tumours in the published literature, we have performed the first cytogenetic analysis of a large series of childhood NHL. Our results show that the cytogenetic changes encountered in NHL of children are distinct and may be different from those seen in NHL of adults reflecting the previously recognized differences in histological presentation and clinical behaviour of the two entities. Thus, the most frequently occurring translocation in B‐cell lesions in children was t(8;14)(q24;1q32). Other translocations frequently seen in adults such as t(14;18)(q32;q21),t(11;14)(q13;q32) and t(3;22)(q27;q11) were either rare or so far not seen in children, although reciprocal translocations appeared to be generally prevalent in childhood NHL. Combining our data with those in the published literature, we have identified two new recurring translocations [t(1;17)(p36;q21) and t(1;14)(p36;q22)], and a recurring duplication [dup(11)(q13;q23)] in this group of lymphomas. In addition, our literature survey identified a third recurring translocation [t(5;14)(q23;q32)] which was previously reported in two cases of childhood NHL. Our analysis also showed differences in the types of nonrandom translocations between childhood NHL and acute lymphoblastic leukaemia (ALL) in children suggesting that biologically these entities are different from one another. This study thus uncovers patterns of chromosome change associated with childhood lymphoma thus providing new opportunities for investigation of their clinical significance by correlation analysis and biological significance by molecular analysis.
Antibodies to plasma proteins: an association with platelet transfusion refractoriness
Authors: Heal J. M. , Cowles J. , Masel D. , Rowe J. M. , Blumberg N.
Published: March 2008

Summary. We hypothesized that antibodies to HLA‐linked polymorphic plasma proteins could be involved in platelet refractoriness by an‘innocent bystander’or immune complex mechanism. Employing a kinetic enzyme‐linked immunosorbant assay (ELISA) technique the ability of IgG from the plasma of refractory patients to bind to albumin, fibrinogen, complement components C2 and C4 was measured. As compared with controls a high percentage of refractory patients had increased IgG capable of binding to all four plasma proteins: C2 (83%), C4 (83%), albumin (75%), fibrinogen (34%). In the presence of exogenous plasma proteins these antibodies mediated increased deposition of IgG onto normal donor platelets. The plasma protein binding IgG consisted both of monomeric IgG and a broad range of high molecular weight complexes. IgG anti‐plasma protein antibody could be eluted from platelets of refractory patients.

The development of anti‐plasma protein IgG was studied during the course of platelet transfusion therapy and found to increase progressively so that by the 20th transfusion > 90% of samples were positive. The presence of plasma protein binding activity correlated with the development of increased levels of platelet bound IgG and refractoriness. Multiple platelet transfusions lead to sensitization to polymorphic determinants on C2 and C4 as well as the formation of high molecular weight complexes. These antibodies and complexes contribute to the deposition of IgG on platelets and may contribute to refractoriness.
Treatment of advanced stage T cell lymphoblastic lymphoma: results of the United Kingdom Children's Cancer Study Group (UKCCSG) protocol 8503
Authors: Eden O. B. , Hann I. , Imeson J. , Cotterill S. , Gerrard M. , Pinkerton C. R.
Published: March 2008

Summary. Ninety‐five unselected patients with stage III and IV T cell lymphoblastic lymphoma were treated according to the United Kingdom Childrens Cancer Study Group protocol 8503. This was a continuous, intensive leukaemia type regimen including cranial irradiation (18 Gy in 10 fractions) and continuing chemotherapy for 2 years identical to the concurrent Medical Research Council ALL protocol. Four‐year event‐free survival was 65% (95% CI 5040%) with no significant difference between stage I11 and stage IV cases. 4.2% of patients died of infection or non‐tumour related events. Following relapse salvage was unlikely without high dose chemotherapy and bone marrow rescue. These results show an improvement over previous U.K. studies but we need to continue to search for subsets of patients with resistant disease for whom even more intensive therapy possibly combined with bone marrow rescue is required.