Showing papers in "Blood Cells Molecules and Diseases in 2009"

The global prevalence of glucose-6-phosphate dehydrogenase deficiency: a systematic review and meta-analysis.

Abstract: Glucose-6-phosphate deficiency is the most prevalent enzyme deficiency, with an estimated 400 million people affected worldwide. This inherited deficiency causes neonatal hyperbilirubinemia and chronic hemolytic anemia. Although most affected individuals are asymptomatic, exposure to oxidative stressors such as certain drugs or infection, can elicit acute hemolysis. To characterize the global prevalence of G6PD deficiency, we conducted a systematic review of the G6PD deficiency literature, drawing studies from various databases, including MEDLINE/Pubmed and Biosis. Selected studies included cross-sectional and longitudinal studies published between 1960 and 2008. Additionally, meta-analytic procedures were employed to assess the degree of heterogeneity amongst prevalence estimates and, where appropriate, pool them. The searches yielded a total of 280 prevalence estimates, corresponding to 88 countries. The highest prevalence rates were reported among Sub-Saharan African countries, even after adjusting for assessment method. Meta-analysis revealed a high degree of heterogeneity for regional and global prevalence estimates. This heterogeneity in reported estimates appeared to be due to differences in G6PD deficiency assessment and diagnostic procedures. The magnitude and variation in global, regional, and country-level prevalence rates of G6PD deficiency are of public health import, particularly in planning programs to improve neonatal health and in the distribution of various medications, especially antimalarial drugs, as G6PD deficiency is most prevalent in malaria-endemic areas.

Spatial analysis of erythrocyte membrane fluctuations by digital holographic microscopy

Abstract: Red blood cell (RBC) membrane fluctuations provide important insights into cell states We present a spatial analysis of red blood cell membrane fluctuations by using digital holographic microscopy (DHM) This interferometric and dye-free technique, possessing nanometric axial and microsecond temporal sensitivities enables to measure cell membrane fluctuations (CMF) on the whole cell surface DHM acquisition is combined with a model which allows extracting the membrane fluctuation amplitude, while taking into account cell membrane topology Uneven distribution of CMF amplitudes over the RBC surface is observed, showing maximal values in a ring corresponding to the highest points on the RBC torus as well as in some scattered areas in the inner region of the RBC CMF amplitudes of 359+/-89 nm and 47+/-05 nm (averaged over the cell surface) were determined for normal and ethanol-fixed RBCs, respectively

Improving survival with deferiprone treatment in patients with thalassemia major: a prospective multicenter randomised clinical trial under the auspices of the Italian Society for Thalassemia and Hemoglobinopathies.

Abstract: The prognosis for thalassemia major has dramatically improved in the last two decades. However, many transfusion-dependent patients continue to develop progressive accumulation of iron. This can lead to tissue damage and eventually death, particularly from cardiac disease. Previous studies that investigated iron chelation treatments, including retrospective and prospective non-randomised clinical trials, suggested that mortality, due mainly to cardiac damage, was reduced or completely absent in patients treated with deferiprone (DFP) alone or a combined deferiprone-deferoxamine (DFP-DFO) chelation treatment. However, no survival analysis has been reported for a long-term randomised control trial. Here, we performed a multicenter, long-term, randomised control trial that compared deferoxamine (DFO) versus DFP alone, sequential DFP-DFO, or combined DFP-DFO iron chelation treatments. The trial included 265 patients with thalassemia major, with 128 (48.3%) females and 137 (51.7%) males. No deaths occurred with the DFP-alone or the combined DFP-DFO treatments. One death occurred due to graft versus host disease (GVHD) in a patient that had undergone bone marrow transplantation; this patient was censored at the time of transplant. Only one death occurred with the DFP-DFO sequential treatment in a patient that had experienced an episode of heart failure one year earlier. Ten deaths occurred with the deferoxamine treatment. The main factors that correlated with an increase in the hazard ratio for death were: cirrhosis, arrhythmia, previous episode of heart failure, diabetes, hypogonadism, and hypothyroidism. In a Cox regression model, the interaction effect of sex and age was statistically significant (p-value<0.013). For each increasing year of age, the hazard ratio for males was 1.03 higher than that for females (p-value<0.013). In conclusion, the results of this study show that the risk factors for predicting mortality in patients with thalassemia major are deferoxamine-treatment, complications, and the interaction effect of sex and age.

BCL11A represses HBG transcription in K562 cells.

Abstract: BCL11A on chromosome 2p16 was recently shown to be a major quantitative trait locus for Hb F level and F-cell number in several populations with or without beta-hemoglobinopathy. We now show that BCL11A isoforms are expressed in K562 cells. Butyrate induction of HBG globin production in K562 is associated with reduced BCL11A. Conversely, augmented expression of BCL11A in K562 cells through transfection of BCL11A expression vector results in more than 50% reduction of HBG promoter transcription activity. This transcription repression can be abrogated by sodium butyrate. BCL11A binds to GGCCGG motif in nucleotide -56 to -51 on the HBG proximal promoter. Together, these data are consistent with BCL11A being able to bind to a core motif in the HBG proximal promoter, recruit and interact with partners to form a repression complex, leading to deacetylation of histones and down-regulation of the HBG transcription.

Platelet function in hypertension.

Abstract: Platelets from hypertensive patients show increased sensitivity to agonists and have high intracellular free Ca2+ concentration. Furthermore, in hypertension, platelets show enhanced endogenous production of reactive oxygen species and a reduced antioxidant status which increases protein tyrosine phosphorylation, enhances Ca2+ mobilization and attenuates NO bioavailability. The study of the abnormalities in platelet function in hypertensive patients can lead to the development of new pharmacological strategies to prevent and/or palliate hypertension-derived complications associated to platelet hyperactivity.

Hereditary stomatocytosis and cation-leaky red cells--recent developments.

Abstract: The hereditary stomatocytoses (HSt) are a diverse group of conditions. Common features include hemolytic anemia, a red cell cation leak and morphological changes, but the severity of the condition can vary enormously. We have previously shown that one form of HSt (cryohydrocytosis), where the monovalent cation leak is increased at low temperature, results from amino acid substitutions in the membrane domain of band 3 (anion exchanger 1, SLC4A1). These substitutions appear to convert band 3 from an anion exchanger into a cation channel. More recently we found that over-hydrated hereditary stomatocytosis (OHSt) results from amino acid substitutions in Rh-associated glycoprotein (RhAG), a putative gas channel protein. Both band 3 and RhAG associate in the red cell membrane to form a macrocomplex that is thought to be involved in red cell gas exchange. In this paper I will review the data that has been published so far on the molecular basis of HSt. I will mention other similar conditions that cause either a cation leak or stomatocytosis or both, and consider the mechanisms of red cell shape change and permeability.

Molecular, hematological and clinical aspects of thalassemia major and thalassemia intermedia associated with Hb E-β-thalassemia in Northeast Thailand

Abstract: Hb E-beta-thalassemia is the most common form of beta-thalassemia found in Thailand. The disease exhibits a varied clinical expression ranging from severe transfusion dependence to relatively mild thalassemia intermedia. We evaluated the effects of primary and secondary genetic factors in modulating the hematological and clinical presentation of 148 northeast Thai patients including 103 severe thalassemia major (TM) and 45 thalassemia intermedia (TI). Among 148 cases examined, eleven different mutations including two novel ones; (beta(33/34 (-G)) and beta(IVS2#815 C-T)) were identified in trans to the beta(E) gene in two TM cases. The other 9 known mutations included beta(41/42), beta(17), beta(IVS2#654), beta(-28), beta(71/72), beta(35), beta(IVS1#5), beta(IVS1#1) and beta(41). Except for the beta(-28) mutation which was found only in the TI group, others mutations were identified in both TM and TI. Co-inheritance of alpha-thalassemia as a phenotype modulating factor was not evident in this study, nor was the presence of the -158 (G)gamma-globin Xmn I polymorphism. Further analysis of the polymorphic (TG)n(CG)m repeats within the IVS2 of the two gamma-globin genes revealed no different proportions of the polymorphic patterns among TM and TI groups of patients either. Our data reveals that in the majority of these Hb E-beta-thalassemia patients, it is very hard to predict the clinical phenotype of the patients from the beta-globin mutations and these secondary genetic modifiers.

Regional heterogeneity of β-thalassemia mutations in the multi ethnic Indian population

Abstract: To determine the frequencies of β-thalassemia mutations in different states of India and to compare this with the available data in Asian Indians for a comprehensive catalogue of molecular defects in the Indian population β-thalassemia mutations were characterized in 2456 heterozygotes using reverse dot blot hybridization, ARMS and DNA sequencing 36 β-thalassemia mutations were characterized from 18 different states in India Seven mutations were common, accounting for 958% of mutated alleles Marked regional diversity was seen in different parts of the country Among the tribal populations, only 2 mutations (IVS I-5 (G → C) and CD15 (G → A) accounted for over 90% of mutant alleles A compilation of all the studies in Asian Indians reported so far showed the presence of 63 mutations in the Indian population This large study adds to the existing data to give a detailed account of the molecular basis of β-thalassemia in India This information is important for establishing prenatal diagnosis programmes in different states in India as well as other countries in which there is a major influx of Indian immigrants

Hydroxyurea in sickle cell disease--a study of clinico-pharmacological efficacy in the Indian haplotype.

Abstract: There is clinical variability in the presentation of sickle cell disease among Indians. Vaso-occlusive crisis is common among non-tribal patients. Hydroxyurea, induces fetal hemoglobin (HbF) synthesis and reduces the clinical severity of sickle cell disease but individual patients have a variable response. This study was undertaken to investigate the efficacy and safety of hydroxyurea in Indians with severe manifestations where the beta(s) gene is linked to the Arab-Indian haplotype and is associated with higher HbF levels. Seventy-seven patients (29 adult sickle homozygous, 25 pediatric sickle homozygous, 23 adult sickle beta-thalassemia) selected for hydroxyurea therapy were evaluated for clinical, hematological, biochemical and genetic parameters and were followed for 24 months. Ninety-eight point seven percent of the sickle chromosomes were linked to the Arab-Indian haplotype, 27% of patients had associated alpha thalassemia and 65% were Xmn I +/+. Seventy-eight percent of the patients had no further crises after starting hydroxyurea. This effect was accompanied by a significant increase in HbF (p<0.001), but this increase was variable in individual cases. There was also an increase in gamma gene mRNA expression in the few cases so studied. Hemoglobin levels increased significantly (p<0.001) resulting in the cessation of blood transfusions. Leucopoenia was observed in one patient. Hydroxyurea was effective in reducing the clinical severity in Indian patients who initially had higher HbF levels and the presence of ameliorating factors, such as alpha-thalassemia and the Xmn I polymorphism. Hydroxyurea therapy with careful monitoring can thus change the quality of life of Indians with sickle cell disease.

Influence of CYP2C9 and VKORC1 on warfarin response during initiation of therapy.

Abstract: Background Although multiple reports have documented the influence of CYP2C9 and VKORC1 variants on warfarin dose, risk of over-anticoagulation and hemorrhage, their influence on anticoagulation maintenance and individual proportion of time spent in target INR range (PPTR) is limited. Moreover the potential benefit of genotype-guided dosing implemented after initiation of therapy in a racially diverse population has not been explored. Herein we present the influence of CYP2C9 and VKORC1 C1173T on warfarin response during the first 30 days of therapy. Methods Warfarin dose was empirically determined in 250 African Americans 271 European Americans. The influence of CYP2C9 and VKORC1 on rate of INR increase, anticoagulation maintenance, risk of over-anticoagulation, and change in dose over 30 days was evaluated after adjustment for socio-demographic, lifestyle and clinical factors. Possession of variant VKORC1 (± variant CYP2C9 ) genotype was associated with a more rapid attainment of target INR and higher frequency of dose adjustments. Patients possessing variant genotypes spent less time in target range. However adjustment for rate of INR increase rendered the association non-significant. European Americans (but not African Americans) possessing variant VKORC1 (± variant CYP2C9 ) genotype had a higher risk of over-anticoagulation. Neither CYP2C9 nor VKORC1 influenced the risk of minor hemorrhage. CYP2C9 and VKORC1 explained 6.3% of the variance in dose change over the first 30 days of therapy demonstrating that the usefulness of genotype-guided dosing may extend beyond first day of therapy. Conclusion The benefit of genotype-based dose prediction may extend beyond first few days of therapy. Whether genotype-guided dosing will decrease the risk of over-anticoagulation, improve anticoagulation control and most importantly improve outcomes for chronic warfarin users remains to be proven.

Infection and autoimmunity

Abstract: The development of some autoimmune diseases is increasing in the developed world faster than can be accounted for by genetic change. The development of these autoimmune diseases, such as Type 1 diabetes, is known to be influenced by both genetic and environmental factors. Environmental factors which have been considered to play a role include infectious agents such as viruses or bacteria. The search for a common initiating infection in the aetiology of Type 1 diabetes as proved thus far inconclusive. An alternative way of considering a role for infection is that infection may have historically prevented the development of autoimmune disease. In the developing world changes have occurred such that many chronic infections have been eliminated and this may have led to the emergence of autoimmune pathology. Evidence in support of this hypothesis is considered here and factors governing the development of autoimmunity compared with those which might have influenced the development of childhood leukaemia.

Alternative Runx1 promoter usage in mouse developmental hematopoiesis.

Abstract: The interest in stem cell based therapies has emphasized the importance of understanding the cellular and molecular mechanisms by which stem cells are generated in ontogeny and maintained throughout adult life. Hematopoietic stem cells (HSCs) are first found in clusters of hematopoietic cells budding from the luminal wall of the major arteries in the developing mammalian embryo. The transcription factor Runx1 is critical for their generation and is specifically expressed at sites of HSC generation, prior to their formation. To understand better the transcriptional hierarchies that converge on Runx1 during HSC emergence, we have initiated studies into its transcriptional regulation. Here we systematically analyzed Runx1 P1 and P2 alternative promoter usage in hematopoietic sites and in sorted cell populations during mouse hematopoietic development. Our results indicate that Runx1 expression in primitive erythrocytes is largely P2-derived, whilst in definitive hematopoietic stem and/or progenitor cells from the yolk sac or AGM and vitelline and umbilical arteries both the distal P1 and proximal P2 promoters are active. After cells have migrated to the fetal liver, the P1 gradually becomes the main hematopoietic promoter and remains this into adulthood. In addition, we identified a novel P2-derived Runx1 isoform.

Protein 4.2: a complex linker.

Abstract: The peripheral membrane protein, protein 4.2, is one of the most abundant protein components of the erythrocyte membrane. Protein 4.2 has an important role in red cell membrane structure, its absence due to natural mutations in humans or gene knockout in mice has a detrimental effect on membrane stability and results in hereditary spherocytosis. It is known to be a point of connection between the band 3 complex and the Rhesus protein complex, through its associations with band 3 and CD47 and also via interactions with the cytoskeletal protein ankyrin. Considering its relatively high abundance and importance in stability of the erythrocyte membrane, protein 4.2 has proved a somewhat neglected protein in recent years. In this review we will summarize our current understanding of protein 4.2, discuss its known interactions and describe the effects and implications of protein 4.2 deficiency. Based on protein 4.2's close homology with transglutaminase family proteins, we propose a new speculative “open” homology structure for protein 4.2 that may represent the active, membrane associated protein 4.2 molecule in red blood cells and also explain the dependence of protein 4.2 on band 3 binding for stability.

Spartathlon, a 246 kilometer foot race: effects of acute inflammation induced by prolonged exercise on circulating progenitor reparative cells.

Abstract: Endothelial progenitor cells (EPCs) and the recently described circulating fibrocytes (CFs) are strongly associated with tissue repair. We investigated the kinetics of both "repair" progenitor cells in healthy athletes who participated in the "Spartahlon" ultradistance foot race (246 km continuous running exercise), which provides a unique model of inducing dramatic systemic inflammatory changes. Peripheral blood mononuclear cells (PBMCs) were isolated from 10 volunteer athletes, who completed successfully the race, before, at the end, and at 48 h post-race. EPCs and CFs were detected as endothelial colony-forming units (CFU-ECs) and as the number of adherent with a spindle-shaped morphology Collagen I(+) cells detected after 6-day culture of PBMCs, respectively. The marked increase of plasma levels of CRP, IL-6, SAA, MCP-1, IL-8, sVCAM-1, sICAM-1, thrombomodulin (sTM) and NT-pro-BNP at the end of race established acute inflammation and tissue injury. EPCs increased by nearly eleven-fold in peripheral blood at the end of the race from 44.5+/-2.5/ml to 494.6+/-27.9/ml and remained increased 428.5+/-31.5/ml at 48 h post-race (p<0.0001). The number of the fibrocytes cultured from PBMCs obtained before, at the end, and 48 h post-race did not reveal any significant difference. These findings indicate that bone marrow responses to acute inflammatory damage, induced by exhausting exercise, with a rapid release of EPCs but not CFs into circulation. Given the ability of EPCs to promote angiogenesis and vascular regeneration, we may suggest that this kind of cell mobilization may serve as a physiologic repair mechanism in acute inflammatory tissue injury.

Fibrin architecture in clots: a quantitative polarized light microscopy analysis.

Abstract: Fibrin plays a vital structural role in thrombus integrity. Thus, the ability to assess fibrin architecture has a potential to provide insight into thrombosis and thrombolysis. Fibrin has an anisotropic molecular structure, which enables it to be seen with polarized light. Therefore, we aimed to determine if automated polarized light microscopy methods of quantifying two structural parameters; fibrin fiber bundle orientation and fibrin's optical retardation (OR: a measure of molecular anisotropy) could be used to assess thrombi. To compare fibrin fiber bundle orientation we analyzed picrosirius red-stained sections obtained from clots formed: (A) in vitro, (B) in injured and stenotic coronary arteries, and (C) in surgically created aortic aneurysms (n=6 for each group). To assess potential changes in OR, we examined fibrin in picrosirius red-stained clots formed after ischemic preconditioning (10 min ischemia+10 min reflow; a circumstance shown to enhance lysability) and in control clots (n=8 each group). The degree of fibrin organization differed significantly according to the location of clot formation; fibrin was most aligned in the aneurysms and least aligned in vitro whereas fibrin in the coronary clots had an intermediate organization. The OR of fibrin in the clots formed after ischemic preconditioning was lower than that in controls (2.9+/-0.5 nm versus 5.4+/-1.0 nm, P<0.05). The automated polarized light analysis methods not only enabled fibrin architecture to be assessed, but also revealed structural differences in clots formed under different circumstances.

The female gaucher patient: the impact of enzyme replacement therapy around key reproductive events (menstruation, pregnancy and menopause)

Abstract: Background The principal manifestations of type 1 Gaucher disease (GD) (increased risk of bleeding, anaemia, splenomegaly, hepatomegaly and bone disease) are likely to affect females during reproductive events such as menarche and menstruation; fertility, pregnancy, parity, delivery and lactation; and menopause. In order to determine the optimal management of female Gaucher patients based on available data, we examine reproductive events and GD in untreated and alglucerase and/or imiglucerase-treated females. Methods A panel of international clinicians experienced in the management of GD reviewed and presented evidence from peer-reviewed literature, a pharmacovigilance database on imiglucerase, and their own clinical experience to support discussions and recommendations. Nine panel members completed a 130-item-questionnaire on the outcomes of the management of female patients in their clinical practice. Results, covering menarche (137 females), menstruation (261 reports), fertility (295 females), pregnancy (416 pregnancies in 247 women) and menopause (45 women) were analysed. Data from a recent Canadian survey on 50 patients with 39 pregnancies, the imiglucerase pharmacovigilance database (100 pregnancies), and relevant literature (56 items covering 398 pregnancies in 205 women) were also reviewed. Key results Menarche : May be delayed in girls with GD. Menorrhagia : Appears to be more common in GD than in the non-Gaucher population and may be ameliorated by alglucerase and/or imiglucerase treatment (menorrhagia in 67/133 (50.4%) untreated females compared with 37/128 (28.9%) treated; Mann–Whitney U test: p = 0.001). Fertility : There is no evidence of decreased fertility in GD. Pregnancy : Pregnancy in GD may be complicated by haematological disease, organomegaly and bone involvement. GD diagnosis occurs frequently during pregnancy. Questionnaire results demonstrate: a reduced risk of spontaneous abortion in women treated with alglucerase and/or imiglucerase (untreated: 26/189 (13.8%); treated 1/58 (1.7%) χ 2 p = 0.010); reduced risk of Gaucher-related complications during delivery (untreated 43/109 (39.4%); treated 3/46 (6.5%) χ 2 p χ 2 p = 0.014). There is no evidence to date of any untoward effect of alglucerase and/or imiglucerase on the fetus, or on infants breast fed by mothers receiving alglucerase and/or imiglucerase. Menopause : The impact of GD on menopause requires further study especially in relation to bone pathology. Conclusions On the basis of this review, GD may have an impact on reproductive events in affected women. Enzyme therapy may have benefits in reducing menorrhagia, spontaneous abortions and complications associated with delivery and the postpartum period.

Enhanced expression of STIM1/Orai1 and TRPC3 in platelets from patients with type 2 diabetes mellitus.

Abstract: Type 2 diabetes mellitus (DM2) is a metabolic syndrome that contributes to both macrovascular and microvascular disorders, where platelet hyperaggregability, associated to abnormal intracellular Ca(2+) homeostasis, plays an important role. We have now investigated the expression of different proteins associated to Ca(2+) entry, a major Ca(2+) signalling event. DM2 donors were randomly selected from normotensive patients with glycosylated Hb levels (HbA1c) over 6%. Control subjects were normal age- and gender-matched healthy people with HbA1c levels in the normal range (3.5-5%). Expression of TRPC1, 3 and 6, STIM1 and Orai1 was analyzed by Western blotting in DM2 patients and controls. Expression of TRPC1 in platelets from DM2 donors and controls was similar; however, expression of TRPC6 is reduced in platelets from DM2 patients as compared to healthy controls. We have found that expression of TRPC3, Orai1 and STIM1 is enhanced in DM2 subjects as compared to controls. Our findings provide an explanation to the enhanced Ca(2+) entry induced by physiological agonists in platelets from DM2 patients.

Olive tree wood phenolic compounds with human platelet antiaggregant properties

Abstract: Oleuropein and (+)-cycloolivil are natural polyphenolic compounds with a significant radical scavenging activity present in olive tree. We have investigated the antiaggregant effects of oleuropein and (+)-cycloolivil isolated from an ethyl acetate extract of olive tree wood. Oleuropein and (+)-cycloolivil reduced the ability of thrombin to stimulate platelet aggregation. Both compounds reduced thrombin-evoked Ca 2+ release and entry to a similar extent to hydroxytyrosol. This effect was greater in platelets from patients with type 2 diabetes mellitus than in controls. Thrombin-, thapsigargin- and 2,5-di-(tert-butyl)-1,4-hydroquinone (TBHQ)-evoked protein tyrosine phosphorylation, which is involved in Ca 2+ signalling and platelet aggregation, is inhibited by oleuropein and (+)-cycloolivil. oleuropein and (+)-cycloolivil are natural oxygen radical scavengers that reduce thrombin-induced protein tyrosine phosphorylation, Ca 2+ signalling and platelet aggregation. These observations suggest that oleuropein and (+)-cycloolivil may prevent thrombotic complications associated to platelet hyperaggregability and be the base for the development of antiaggregant therapeutic strategies.

Promising results of the chaperone effect caused by iminosugars and aminocyclitol derivatives on mutant glucocerebrosidases causing Gaucher disease

Abstract: Gaucher disease is an autosomal recessive disorder. It is characterized by the accumulation of glucosylceramide in lysosomes of mononuclear phagocyte system, attributable to acid beta-glucosidase deficiency. The main consequences of this disease are hepatosplenomegaly, skeletal lesions and, sometimes, neurological manifestations. At sub-inhibitory concentrations, several competitive inhibitors act as chemical chaperones by inducing protein stabilization and increasing enzymatic activity. Here we tested two iminosugars (NB-DNJ and NN-DNJ) and four aminocyclitols with distinct degrees of lipophilicity as pharmacological chaperones for glucocerebrosidase (GBA). We report an increase in the activity of GBA using NN-DNJ, NB-DNJ and aminocyclitol 1 in stably transfected cell lines, and an increment with NN-DNJ and aminocyclitol 4 in patient fibroblasts. These results on specific mutations validate the use of chemical chaperones as a therapeutic approach for Gaucher disease. However, the development and analysis of new compounds is required in order to find more effective therapeutic agents that are active on a broader range of mutations.

Runx1 promotes B-cell survival and lymphoma development.

Abstract: Runx1 is essential for the homeostatic control of normal hematopoiesis and is required for lymphoid development. Translocations or point mutations that result in RUNX1 loss or disrupted function predispose to leukemia but data derived from model systems suggests that Runx genes can also be pro-oncogenic. Here we investigate the effects of enforced Runx1 expression in lymphoid lineages both in vivo and in vitro and show that transgene expression enhanced cell survival in the thymus and bone marrow but strongly inhibited the expansion of hematopoietic and B cell progenitors in vitro. Despite this, modestly enhanced levels of Runx1 accelerated Myc-induced lymphomagenesis in both the B cell and T cell lineages. Together these data provide formal proof that wild type Runx1 can promote oncogenesis in lymphoid tissues and that, in addition to loss of function, gain of function may have an aetiological role in leukemia.

HLA-haploidentical blood and bone marrow transplantation with anti-thymocyte globulin: long-term comparison with HLA-identical sibling transplantation.

Abstract: We present an update of our results regarding related HLA-haploidentical and HLA-identical sibling hematopoietic stem cell transplantation (HSCT) in patients with leukemia. We compared the outcomes of 107 patients with leukemia undergoing HLA-identical sibling (n=51) or related HLA-haploidentical (n=56) HSCT performed during the same time period. Patients received BU-CY/CY-TBI in HLA-identical sibling HSCT or TBI+Ara-C+CY+ATG/CCNU+Ara-C+Bu+CY+ATG in haploidentical HSCT as conditioning regimens, followed by unmanipulated marrow and/or peripheral blood (PB) transplantation. All patients achieved full engraftment. The cumulative incidence of grades II through IV acute graft-versus-host disease (aGvHD) in the matched and haploidentical cohorts was 13.7% and 26.8% (P 0.05). The two-year incidence of treatment-related mortality for matched and haploidentical patients was 7.8% and 12.5%, respectively, with P>0.05, and the incidence of relapse was 17% and 22%, respectively, with P>0.05. The two-year adjusted leukemia-free survival for matched versus haploidentical patients was 76% and 68%, respectively, with P>0.05, and the overall survival for matched versus haploidentical patients was 80% and 70%, respectively, with P>0.05. Multivariate analyses showed that only advanced disease stage and a diagnosis of acute leukemia were related to increased risk of relapse, treatment failure, and overall mortality. In summary, HSCT performed with related HLA-haploidentical donors is a feasible approach with acceptable outcomes.

Splenic marginal zone lymphoma: Clinical clustering of immunoglobulin heavy chain repertoires.

Abstract: Immunoglobulin gene usage and somatic mutation patterns were studied in 59 patients with splenic marginal zone lymphoma and were correlated with clinical characteristics Fifty-nine IGHV rearrangements were amplified IGHV1, IGHV3, and IGHV4 subgroups accounted for 30%, 56%, and 14% of sequences, respectively IGHV genes most frequently used were IGHV1-2 (n=12), IGHV3-23 (n=15), IGHV3-30 (n=7) and IGHV4-34 (n=5) IGHV was unmutated in 25% Villous lymphocytes >10% were detected in 50% of patients belonging to the IGHV1-2 group, in 21% of the IGHV3-23 group, and in no patient of the IGHV3-30 group (p=005) Liver involvement was present in 50% of the IGHV3-30 group, in 9% of the IGHV3-23 group, and in no patient of the IGHV1-2 group (p=004) HCV-serology was positive in 50% of the IGHV3-30 group, in 7% of the IGHV3-23 group, and in 17% of the IGHV1-2 group (p=004) The proportion of intermediate and high risk patients according to the SMZL score was higher in the unmutated respect to the mutated group (69% vs 32%, p=005) In conclusion, IGHV rearrangement analysis in splenic marginal zone B-cell lymphoma reveals a non-random preference for use of IGHV1-2, IGHV3-23 and IGHV3-30 genes, whose presence differs according to clinical features and prognostic category

RUNX factors in development: lessons from invertebrate model systems.

Abstract: Runt-related (RUNX) transcription factors are evolutionarily conserved regulators of cell proliferation, differentiation and stem cell maintenance. They are critical for the correct development and function of a variety of human tissues, including during haematopoiesis. RUNX genes regulate various aspects of proliferation control, stem cell maintenance, lineage commitment and regulation of differentiation; disruptions in the correct function of RUNX genes have been associated with human pathologies, most prominently cancer. Because of the high context dependency and partial redundancy of vertebrate RUNX genes, invertebrate model systems have been studied in the hope of finding an ancestral function. Here we review the progress of these studies in three invertebrate systems, the fruit fly Drosophila melanogaster, the sea urchin Strongylocentrotus purpuratus and the nematode Caenorhabditis elegans. All essential aspects of RUNX function in vertebrates have counterparts in invertebrates, confirming the usefulness of these studies in simpler organisms. The fact that not all RUNX functions are conserved in all systems, though, underscores the importance of choosing the right model to ask specific questions.

Development and perspective of current Brazilian studies on the epidemiology of childhood leukemia.

Abstract: In this concise report, we describe the history and evolution of childhood acute leukemia studies in Brazil, and the application if key biomarkers for clinical trials and epidemiological studies over the past 8 years. Highlights of each ongoing study are summarized. A Brazilian network integrating hospitals and scientific institutions from all country regions has been established. This organization is made possible through informatics and computer networking, and the standardization of pathological reviews including immunophenotyping and molecular characterization of childhood leukemias. The unique characteristics of the Brazilian population combined with a large clinical and epidemiologic framework for patient ascertainment has enabled large-scale epidemiological studies on childhood leukemia in Brazil.

Expression of enzymatically active L-DOPA decarboxylase in human peripheral leukocytes.

Abstract: L-DOPA decarboxylase (DDC) is a pyridoxal 5-phosphate (PLP)-dependent enzyme that catalyses the decarboxylation of L-DOPA to dopamine. Dopamine (DA) has been found to be a regulating factor of the proliferation and differentiation of different leukocyte subtypes. In the present study, we report the expression of the gene that codes for the L-DOPA decarboxylase in human peripheral leukocytes and in T-lymphocytes, as well as the simultaneous detection of both neural and non-neural type DDC mRNA in the cellular components of this specialized connective tissue type. Furthermore, we have detected the neural type DDC transcript which lacks exon 3 and the alternative 37 kD alt-DDC protein isoform which lacks exons 10–15 but includes an alternative exon 10 in human peripheral leukocytes. Treatment of white blood cells with Triton X-114 resulted in the recovery of DDC in the detergent enriched and highly hydrophobic phases, suggesting association of DDC molecules with membranes in the studied cells. Enzymatic activity experiments revealed that DDC is active towards the decarboxylation of L-DOPA. The expression of enzymatically active DDC in human leukocytes could indicate a cross-talk between the nervous and the immune systems and raises new questions about the regulatory role of DDC in immune responses.

Interethnic differences of CYP2C9 alleles in healthy Hungarian and Roma population samples: Relationship to worldwide allelic frequencies

Abstract: CYP2C9 gene polymorphisms are widely studied in several ethnic groups, however they are less known in the Roma population. The aim of this work was to study the ethnic differences of the CYP2C9 allele distribution in a healthy Roma population in order to compare them with a healthy Hungarian population. A total of 535 Hungarian and 465 Roma volunteers were genotyped for the CYP2C9*2 (Arg144Cys) and CYP2C9*3 (Ile359Leu) allelic variants by PCR-RFLP assay. The frequencies of the CYP2C9*1, *2 and *3 alleles in the Hungarian population were 0.787, 0.125, and 0.088 and in Roma 0.727, 0.118, and 0.155, respectively. We found a significant difference in CYP2C9*3 prevalence between the Hungarian and Roma populations, which have therapeutic consequences (p<0.005). The distribution of *1/*1, *1/*2, *1/*3, *2/*2, *2/*3, and *3/*3 genotypes in Hungarians were 0.620, 0.195, 0.139, 0.021, 0.015, and 0.011, while in Roma were 0.533, 0.168, 0.219, 0.011, 0.047, and 0.022, respectively. A significant difference was found between the Hungarian and Roma populations regarding the *1/*1, *1/*3 and the *2/*3 (p<0.005) genotypes. This is the first study to investigate the polymorphisms of CYP2C9 gene in the two largest populations in Hungary, healthy Hungarians and Roma. The prevalence of variant CYP2C9 alleles in the Hungarian population is similar to that observed in other European populations. In contrast, the Roma population differs from Hungarians, from most of other Caucasian groups, and from Indians in the incidence of CYP2C9 common variants. The difference in allele distribution patterns between the two populations studied has therapeutic implications as it influences the optimization of therapies.

Post-translational modifications of Runx1 regulate its activity in the cell

Abstract: In this report we review the current knowledge of the interaction of RUNX1(AML1) with serine/threonine kinases, lysine and arginine methyltransferases, lysine acetyltransferases, and histone deacetylases. We also discuss the effect of RUNX1-ETO fusion gene on DNA methylation. RUNX1 post-transcriptional modification can affect its role in influencing differentiation and self-renewal of hematopoietic cells. The goal of these studies is to develop targets for improved leukemia therapy.

Protein 4.1 and the control of ion channels.

Abstract: The classical function of 4.1R in red blood cells is to contribute to the mechanochemical properties of the membrane by promoting the interaction between spectrin and actin. More recently, it has been recognized that 4.1R is required for the stable cell surface accumulation of a number of erythrocyte membrane proteins. 4.1R is one member of the mammalian 4.1 family - the others being 4.1N, 4.1G and 4.1B - and is expressed in many cell types other than erythrocytes. Recently we have examined the phenotype of hearts from 4.1R knockout mice. Although they had a generally normal morphology, these hearts exhibited bradycardia, and prolongation of both action potentials and QT intervals. Electrophysiological analysis revealed anomalies in a range of ion channel activities. In addition, the immunoreactivity of voltage-gated Na(+) channel NaV1.5 was reduced, indicating a role for 4.1R in the cellular accumulation of this ion channel. 4.1 proteins also have roles in the accumulation of at least two other classes of ion channel. In epithelia, 4.1 interacts with the store-operated channel TRPC4. In neurons, the ligand-gated channels GluR1 and GluR4 require 4.1 proteins for cell surface accumulation. The spectrum of transmembrane proteins that bind to 4.1 proteins overlaps with that of ankyrin. A hypothesis to investigate in the future is that differential regulation of 4.1 and ankyrins (e.g. by PIP(2)) allows highly selective control of cell surface accumulation and transport activity of a specific range of ion channels.

Not all DMT1 mutations lead to iron overload

Abstract: DMT1 is a membrane-bound divalent metal transporter, which co-transports protons and (Fe(2+)) from an acidic microenvironment (endosome, duodenal lumen) to the cell cytosol. Results from animal models and from patients have shown that DMT1 is required for intestinal iron absorption and iron acquisition by erythrocytes. Only three human patients with DMT1 mutations have been described so far. They presented with hypochromic microcytic anemia and heavy liver iron overload, even at a very young age. Here, we report the fourth human case, a 7-year old boy with a new homozygous DMT1 mutation, microcytic anemia but no liver iron overload. The mutation introduces a Glycine to Arginine (p.G75R) amino acid substitution. Glycine75 is a highly conserved amino acid present in the first transmembrane domain of the protein and we hypothesize that this mutation fully impairs ferrous iron uptake from the diet and prevents the onset of liver iron overload.

Linkage of cytosolic peroxiredoxin 2 to erythrocyte membrane imposed by hydrogen peroxide-induced oxidative stress.

Abstract: Human erythrocyte peroxiredoxin 2 (Prx2) is a typical 2-cys cytosolic peroxiredoxin with thiol-dependent hydrogen peroxide scavenger activity. In a previous work, we reported Prx2 erythrocyte membrane linkage in some Hereditary Spherocytosis patients and that it seemed to be related to oxidative stress. The aim of the present work was to determine if Prx2 linkage to erythrocyte membrane could be induced by oxidative stress mediated by H 2 O 2 and to further understand how and why this process occurs. We performed in vitro assays in which catalase or both Hb autoxidation and catalase were inhibited, under H 2 O 2 -induced oxidative stress conditions. Erythrocyte membrane linked Prx2 was detected by immunoblotting and quantified by densitometry. As oxidative stress markers, we determined membrane bound hemoglobin and lipid peroxidation, and we found that their values increased with H 2 O 2 concentration. Prx2 linkage to the membrane also rose with increasing H 2 O 2 concentration, and was only observed when the oxidized form of the enzyme was present in the cytosol. Oxidized Hb and Prx2 membrane linkages appear to be independent processes, although, both result from oxidative stress and may be useful as oxidative stress and/or erythrocyte damage/senescence markers.