Search

Non-transferrin bound iron (NTBI) is commonly detected in patients with systemic iron overload whose serum iron-binding capacity has been surpassed. It has been perceived as an indicator of iron overload, impending organ damage and a chelation target in poly-transfused thalassemia patients. However, NTBI is a heterogeneous entity comprising various iron complexes, including a significant redox-active and readily chelatable fraction, which we have designated as "labile plasma iron" (LPI). We found that LPI levels can be affected by plasma components such as citrate, uric acid, and albumin. However, the inclusion of a mild metal mobilizing agent in the LPI assay (designated here as "eLPI"), at concentrations that do not affect transferrin-bound iron, largely overcomes such effects and provides a measure of the full NTBI content. We analyzed three distinct groups of poly-transfused, iron overloaded thalassemia patients: non-chelated children (3-13 yrs, Gaza, Palestine), chelated adolescents-young adults (13-28 yrs, Israel), and chelated adults (27-61 yrs, Israel) for LPI and eLPI. The eLPI levels in all three groups were roughly commensurate (r(2) = 0.61-0.75) with deferrioxamine-detectable NTBI, i.e., DCI. In older chelated patients, eLPI levels approximated those of LPI, but in poly-transfused unchelated children eLPI was notably higher than LPI, a difference attributed to plasma properties affected by labile iron due to lack of chelation, possibly reflecting age-dependent attrition of plasma components. We propose that the two formats of NTBI measurement presented here are complementary and used together could provide more comprehensive information on the forms of NTBI in patients and their response to chelation., (Copyright © 2011 Wiley Periodicals, Inc.)

The purpose of this article is to set forth our approach to diagnosing and managing the thalassemias, including β-thalassemia intermedia and β-thalassemia major. The article begins by briefly describing recent advances in our understanding of the pathophysiology of thalassemia. In the discussion on diagnosing the condition, we cover the development of improved diagnostic tools, including the use of very small fetal DNA samples to detect single point mutations with great reliability for prenatal diagnosis of homozygous thalassemia. In our description of treatment strategies, we focus on how we deal with clinical manifestations and long-term complications using the most effective current treatment methods for β-thalassemia. The discussion of disease management focuses on our use of transfusion therapy and the newly developed oral iron chelators, deferiprone and deferasirox. We also deal with splenectomy and how we manage endocrinopathies and cardiac complications. In addition, we describe our use of hematopoietic stem cell transplantation, which has produced cure rates as high as 97%, and the use of cord blood transplantation. Finally, we briefly touch on therapies that might be effective in the near future, including new fetal hemoglobin inducers and gene therapy.

Many patients with myelodysplastic syndromes (MDS) become dependent on blood transfusions and develop transfusional iron overload, which is exacerbated by increased absorption of dietary iron in response to ineffective erythropoiesis. However, it is uncertain whether there is an association among iron accumulation, clinical complications, and decreased likelihood of survival in MDS patients. Here, we discuss our current understanding of the effects of transfusion dependency and iron overload in MDS, indicate our knowledge gaps, and suggest that particular emphasis should be placed on further characterizing the role of redox-active forms of labile iron, which may be as important as the total iron burden.

In beta-hemoglobinopathies, such as beta-thalassemia (thal) and sickle cell anemia, the primary defects are mutations in the beta-globin gene. However, many aspects of the pathophysiology are mediated by oxidative stress. Fermented papaya preparation (FPP), a natural health food product obtained by biofermentation of carica papaya, has been shown to limit oxidative stress both in vitro and in vivo. We studied the effect of FPP on two groups of beta-thal patients: beta-thal, major and intermedia, (in Israel) and E-beta-thal (in Singapore). The results indicated that in both groups FPP treatment increased the content of reduced glutathione (GSH) in red blood cells (RBC), and decreased their reactive oxygen species (ROS) generation, membrane lipid peroxidation, and externalization of phosphatidylserine (PS), indicating amelioration of their oxidative status, without a significant change in the hematological parameters. Since the turnover of the erythron is relatively slow, it is possible that longer duration of treatment, probably with the addition of an iron chelator, is required in order to achieve the latter goals., (Copyright 2010 John Wiley & Sons, Ltd.)

On the basis of all the presented data, one can conclude that oxidative stress plays a major role in the pathophysiology of thalassemia and other congenital and acquired hemolytic anemias. Free extracellular (labile plasma iron, LPI) and intracellular (labile iron pool, LIP) iron species that have been identified in thalassemic blood cells are responsible for generation of oxidative stress by catalyzing formation of oxygen radicals over the antioxidant capacity of the cell. Consequently, there is a rationale for iron chelation to eliminate the free-iron species, which in this respect, act like antioxidants. In addition, antioxidants such as vitamin E and polyphenols are also capable of ameliorating increased oxidative stress parameters and, given together with iron chelators, may provide a substantial improvement in the pathophysiology of hemolytic anemias and particularly in thalassemia.

Unlabelled: One complication in patients with beta-thalassaemia who had prolonged survival is chronic hypercoagulable state, which results in thromboembolic events involving major organs including the brain. We determined the prevalence of microthrombosis in the brain in an asymptomatic subgroup of patients with b-thalassaemia intermedia (TI) who had undergone splenectomy. This retrospective review included 200 patients with TI diagnosed on the basis of blood count, haemoglobin (Hb) electrophoresis and clinical data. Their ages ranged from 18 to 34 years, 19 (63.3%) were women and 11 (36.7%) were men. We selected 30 patients at random who fulfilled the inclusion criteria: Hb concentration >7 g/dl), splenectomy and platelet count >500,000/ml. Their mean Hb concentration was 8.4 g/dl and their mean ferritin concentration was 519 ng/ml. Magnetic resonance imaging (MRI) was done in every patient, and the findings were interpreted by an expert neuroradiologist. Imaging studies showed pathological findings in 28% of the patients. Six had changes in the white matter suggestive of ischaemia and two had evidence of small infarctions., Conclusion: 1) In this small subset of patients diagnostic magnetic resonance imaging to monitor early asymptomatic or subclinical vascular damage in the brain can be considered when they reach the age of 20 years, and repeated every 3-5 years. 2) Treatment with antiplatelet aggregants is suggested in patients with documented asymptomatic brain ischaemia. 3) These results require confirmation in a larger group of similar patients with other types of thalassaemia who are multitransfused or have an intact spleen.

Objectives: To evaluate the hematological profile, oxidative stress, and antioxidant parameters in beta-thalassemia/Hb E patients treated with curcuminoids for 12 months., Design and Methods: Twenty-one beta-thalassemia/Hb E patients were given 2 capsules of 250 mg each of curcuminoids (a total of 500 mg) daily for 12 months. Blood was collected every 2 months during treatment and 3 months after withdrawal and was determined for complete blood count, malonyldialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), reduced glutathione (GSH) in red blood cells (RBC), and non-transferrin bound iron (NTBI) in serum., Results: The increased oxidative stress in beta-thalassemia/Hb E patients was shown by higher levels of MDA, SOD, GSH-Px in RBC, serum NTBI, and lower level of RBC GSH. Curcuminoids administration resulted in improvement of all the measured parameters as long as they were administered. After 3 months withdrawal of treatment, all parameters returned close to baseline levels., Conclusion: Curcuminoids may be used to ameliorate oxidative damage in patients with beta-thalassemia/Hb E disease., (Copyright 2009 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.)

Iron overload (IO) in the heart is a life-threatening complication in transfusion-dependent patients with thalassaemia major (TM) and to a lesser extent in sickle cell disease (SCD), while no data are available in patients with sickle/beta(0)-thalassaemia. Iron deposition in the heart, liver and pancreas was assessed using T2* MRI sequences, as well as free iron species assays - non-transferrin bound iron (NTBI) and labile plasma iron (LPI), in addition to serum ferritin, percentage transferrin saturation and serum hepcidin, in 10 multitransfused patients (>30 yr) with sickle/beta(0)-thalassaemia. None of the patients had iron deposition in the heart. Three patients had mild, one had moderate, and two had severe liver IO. Two patients had mild iron deposition in the pancreas. In all the patients, serum hepcidin levels were normal - NTBI and LPI were not detected. Possible explanations of these findings are discussed.

β-thalassemia is an inherited disorder due to mutations found in the β-globin gene, leading to anemia and requiring sporadic or chronic blood transfusions for survival. Without proper chelation, β-thalassemia results in iron overload. Ineffective erythropoiesis can lead to iron overload even in untransfused patients who are affected by β-thalassemia intermedia. Better understanding of the molecular biologic aspects of this disorder has led to improvements in population screening and prenatal diagnosis, which, in turn, have led to dramatic reductions in the number of children born with β-thalassemia major in the Mediterranean littoral. However, as a consequence of decreases in neonatal and childhood mortality in other geographical areas, β-thalassemia has become a worldwide clinical problem. A number of unsolved pathophysiological issues remain, such as ineffective erythropoieis, abnormal iron absorption, oxidative stress, splenomegaly and thrombosis. In the last few years, novel studies have the potential to introduce new therapeutic approaches that might reduce these problems and limit the need for blood transfusion.

Purpose: Combat soldiers have a higher prevalence of anemia than age- and gender-matched civilians. This may be caused by hemodilution, which is typical among athletes, or by reduced body iron stores. The aim of this study was to investigate the incidence of iron-deficiency anemia in recruits to the Israel Defense Force after 6 months of training., Methods: Blood was collected from recruits before training. After 6 months of follow-up, 153 paired blood samples were collected from the initial cohort. Total blood count and serum iron, transferrin, and ferritin were measured at both time points. Soluble transferrin receptor (sTfR) was measured in 119 of the paired samples and the sTfR/log ferritin ratio was calculated., Results: At recruitment, mean hemoglobin concentration was 14.7 +/- .9 g/dl. Iron-transferrin saturation was 34.1% +/- 13.6%, and mean ferritin concentration was 53.6 +/- 33.2 ng/ml. Anemia prevalence (Hb <14 g/dl) was 17.6%, and 14.9% of participants were iron-deficient (ferritin <22 mg/dl). At 6 months, 50.3% of the cohort was anemic, and 27.3% demonstrated iron-store depletion. Paired analysis showed an average reduction of .83 g/dl in hemoglobin (p < .001), and of 9.8 mg/dl in ferritin (p < .001). sTfR increased from 1.9 to 2.1 mg/dl (p < .003) among recruits who became anemic. Half of the recruits experienced mild anemia after 6 months of training. Iron store depletion was observed among 24.5% of the cohort after training, as opposed to 15% at recruitment. Overall, these changes were not accompanied by a significant increase in sTfR, but among the subset of anemic subjects, there was a slight increase in index value., Conclusions: In half of the cases, new-onset anemia was attributable to iron deficiency, and in the remainder, to hemodilution.

Many patients with chronic lymphocytic leukemia (CLL) develop progressive treatment-resistant disease. Rituximab (RTX), a monoclonal antibody targeting CD20 on B lymphocytes and widely used in other indolent B-cell neoplasms is less efficacious in CLL, possibly because of associated complement deficiencies. Initial in vitro and in vivo observations support the central role of complement in rituximab-mediated loss of CD20(+) cells in CLL. In an open trial conducted in outpatient hematology clinics in Israel and Greece, we examined whether providing complement by concurrent administration of fresh frozen plasma (FFP) would enhance the effect of RTX in CLL. Five patients with severe treatment-resistant CLL were included in the trial. All had been previously treated with fludarabine, and three also failed treatment with RTX. Each patient was treated with two units of FFP followed with RTX 375 mg/m(2) as a single agent, repeated every 1-2 weeks as needed. A rapid and dramatic clinical and laboratory response was achieved in all patients. Lymphocyte counts dropped markedly followed by shrinkage of lymph nodes and spleen and improvement of the anemia and thrombocytopenia. This could be maintained over 8 months (median) with additional cycles if necessary. Treatment was well tolerated in all cases. In conclusion, adding FFP to RTX may provide a useful therapeutic option in patients with advanced CLL resistant to treatment. Further studies are needed to confirm and study the efficacy of the FFP/RTX combination in advanced CLL, establish the mechanisms, and possibly extend its use to other B-cell-dependent pathologies, such as treatment-refractory autoimmune diseases.

The oxidative status of cells is determined by the balance between pro-oxidants and antioxidants. Pro-oxidants, referred to as reactive oxygen species (ROS), are classified into radicals and nonradicals. The radicals are highly reactive due to their tendency to accept or donate an electron and attain stability. When cells experience oxidative stress, ROS, which are generated in excess, may oxidize proteins, lipids and DNA - leading to cell death and organ damage. Oxidative stress is believed to aggravate the symptoms of many diseases, including hemolytic anemias. Oxidative stress was found in the beta-hemoglobinopathies (sickle cell anemia and thalassemia), glucose-6-phosphate dehydrogenase deficiency, hereditary spherocytosis, congenital dyserythropoietic anaemias and Paroxysmal Nocturnal Hemoglobinuria. Although oxidative stress is not the primary etiology of these diseases, oxidative damage to their erythroid cells plays a crucial role in hemolysis due to ineffective erythropoiesis in the bone marrow and short survival of red blood cells (RBC) in the circulation. Moreover, platelets and polymorphonuclear (PMN) white cells are also exposed to oxidative stress. As a result some patients develop thromboembolic phenomena and recurrent bacterial infections in addition to the chronic anemia. In this review we describe the role of oxidative stress and the potential therapeutic potential of anti-oxidants in various hemolytic anemias.

Thromboembolic complications are not uncommon in thalassemia. Previous studies suggest increased platelet aggregation and a potential role of pathological changes in the red blood cell (RBC) lipid membrane, induced by oxidative stress. In the present study, platelet adhesion and the effect of thalassemic RBC on platelet adhesion under flow conditions were evaluated, using the Cone and Plate (let) Analyzer(CPA). Twenty-two beta-thalassemia patients and 22 blood type-matched healthy controls were studied. An increased platelet adhesion (% surface coverage, SC), was observed in patients as compared to controls (p < 0.05). When platelet count and haematocrit were normalized by autologous reconstitution, a significant increase in platelet aggregation (average size, AS) was observed (p < 0.05). Increased platelet adhesion (SC and AS), was demonstrated in six patients with a history of thrombosis as compared to 16 patients without any history of thrombosis (p < or = 0.007) and in 17 splenectomized patients as compared to five non-splenectomized patients (p = 0.003). In reconstitution studies, thalassemic RBC mixed with normal platelet-rich plasma significantly increased platelet adhesion compared to normal RBC (SC p < 0.03, AS p < 0.02). Thalassemic platelets reconstituted with normal RBC, had increased aggregation (AS, p < 0.004) in comparison with normal platelets. The results indicate that increased platelet adhesion in beta-thalassemia is induced by both platelets and RBC. Increased platelet adhesion correlated with clinical thrombotic events and thus may suggest a mechanism of thrombosis in thalassemic patients. The potential application of the CPA in identifying thalassemic patients with high risk for thrombosis should be studied prospectively in a larger cohort of patients.

In beta-thalassemia, the mechanism driving ineffective erythropoiesis (IE) is insufficiently understood. We analyzed mice affected by beta-thalassemia and observed, unexpectedly, a relatively small increase in apoptosis of their erythroid cells compared with healthy mice. Therefore, we sought to determine whether IE could also be characterized by limited erythroid cell differentiation. In thalassemic mice, we observed that a greater than normal percentage of erythroid cells was in S-phase, exhibiting an erythroblast-like morphology. Thalassemic cells were associated with expression of cell cycle-promoting genes such as EpoR, Jak2, Cyclin-A, Cdk2, and Ki-67 and the antiapoptotic protein Bcl-X(L). The cells also differentiated less than normal erythroid ones in vitro. To investigate whether Jak2 could be responsible for the limited cell differentiation, we administered a Jak2 inhibitor, TG101209, to healthy and thalassemic mice. Exposure to TG101209 dramatically decreased the spleen size but also affected anemia. Although our data do not exclude a role for apoptosis in IE, we propose that expansion of the erythroid pool followed by limited cell differentiation exacerbates IE in thalassemia. In addition, these results suggest that use of Jak2 inhibitors has the potential to profoundly change the management of this disorder.

Many aspects of the pathology in beta-hemoglobinopathies (beta-thalassemia and sickle cell anemia) are mediated by oxidative stress. Fermented papaya preparation (FPP) was tested for its antioxidant effects: the scavenging effect was determined spectrofluorometrically in a cell-free system using 2'-7'-dichlorofluorescin-diacetate (DCF). Both spontaneous and H(2)O(2)-induced DCF oxidations were decreased by FPP in a dose-dependent fashion. Using flow cytometry, it was shown that in vitro treatment of blood cells from beta-thalassemic patients with FPP increased the glutathione content of red blood cells (RBC), platelets and polymorphonuclear (PMN) leukocytes, and reduced their reactive oxygen species, membrane lipid peroxidation and externalization of phosphatidylserine. These effects result in (a) reduced thalassemic RBC sensitivity to hemolysis and phagocytosis by macrophages; (b) improved PMN ability to generate oxidative burst - an intracellular mechanism of bacteriolysis, and (c) reduced platelet tendency to undergo activation, as reflected by fewer platelets carrying external phosphatidylserine. Oral administration of FPP to beta-thalassemic mice (50 mg/mouse/day for 3 months) and to patients (3 g x 3 times/day for 3 months), reduced all the above mentioned parameters of oxidative stress (p < 0.001 in mice and p < 0.005 in patients). These results suggest that FPP, as a potent antioxidant, might alleviate symptoms associated with oxidative stress in severe forms of thalassemia., (Copyright (c) 2008 John Wiley & Sons, Ltd.)

Background: beta-thalassemia is the most common inherited single gene disorder worldwide, and glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common human enzyme deficiency. The goal of this study was to compare the frequency of beta-thalassemia trait and G6PD among the Moslem and Jewish populations in Shiraz, southern Iran., Methods: We examined 201 Moslems and 187 Jewish subjects who were selected by random sampling. For diagnosis of thalassemia, complete blood count and hemoglobin electrophoresis were carried out and for G6PD deficiency, fluorescent spot test methods were used as a screening test., Results: Among Moslem subjects, 14 cases (7.0%) were diagnosed as carriers of beta-thalassemia minor, whereas no carriers were detected among Jewish subjects. Seven Moslems (7%) and eight Jewish subjects (7.5%) were found to have G6PD deficiency. Among both groups the most common mutation was the Mediterranean type (563 C>T). In one Moslem subject, the detected mutation was 1003 (G>A) and in two Jewish subjects the mutations were 1376 (G>T) and G6PD A-., Conclusions: Whereas the frequency of beta-thalassemia minor among Moslems is higher than in the Jews in Shiraz, the frequency of G6PD deficiency was not significantly different in the two populations. These findings suggest that obligatory premarital beta-thalassemia screening for Jews in the community is not necessary, whereas neonatal screening for G6PD could be useful for both Jews and Moslems.

Chronic hypercoagulable state expressed clinically by thromboembolic events has been described in thalassemia. One of the affected organs is the brain where symptomatic and asymptomatic damage has been reported. The present report describes seven cases who presented with the signs of cerebrovascular accident (CVA), five ischemic and two with hemorrhage. Two of them died. All patients were splenectomized, five received regular blood transfusions, and their ferritin levels were between 1,200 and 3,000 mg %. In addition, four patients had congestive heart failure and atrial fibrillation, and three had "Bronze diabetes," The recommendation on the basis of the results is that well-designed clinical trials are indicated to monitor asymptomatic brain damage by magnetic resonance imaging in splenectomized patients over the age of 20 years, who are not regularly transfused and have a high risk to develop thromboembolic events. In this subset of patients, anticoagulant and/or antiplatelet therapy should be considered. Moreover, treatment of additional complications resulting from iron overload, which may contribute to the etiology of CVA such as cardiac failure and arrhythmia with or without "bronze diabetes," is mandatory.

Low-risk myelodysplastic syndrome (MDS) is characterized by cytopenia, mainly anemia, because of ineffective hematopoiesis. Some of the patients with ineffective erythropoiesis, with or without ring sideroblasts in their bone marrow, develop severe anemia requiring frequent blood transfusions and consequently develop iron overload. Excess free iron in cells catalyses the generation of reactive oxygen species (ROS) that cause cell and tissue damage. Using flow cytometry techniques, we compared the oxidative status of red blood cells (RBC), platelets and neutrophils in 14 MDS patients with those of normal donors. The results show that ROS were higher while reduced glutathione (GSH) was lower in their RBC and platelets compared with normal cells. In neutrophils, no difference was found in ROS, while the GSH levels were lower. A correlation (r = 0.6) was found between serum ferritin levels of the patients and the ROS in their RBC and platelets. The oxidative stress was ameliorated by a short incubation with the iron-chelators, the deferrioxamine and deferiprone or with antioxidants such as N-acetylcysteine, suggesting that MDS patients might benefit from treatment with iron-chelators and antioxidants.

The method of cardiovascular T2 magnetic resonance imaging (MRI) allows in vivo estimation of iron in the heart and liver and was used to measure the degree of iron overload in 10 transfused MDS patients (average 90 blood units) and in 3 patients with congenital hemolytic anemia. In all MDS patients iron overload was found in the liver but not in the heart. Patients with congenital anemias had iron in both organs despite iron chelation. It is possible that in MDS more time and more transfusions are required to induce iron accumulation in the myocardium. Therefore, cardiac MRI may serve as a diagnostic tool to assess if and when iron chelation is indicated., ((c) 2007 Wiley-Liss, Inc.)

Thalassaemia and sickle cell disease (SCD) represent the most common forms of hereditary haemolytic anaemia and result from a partial or complete lack of synthesis of one of the major alpha- or beta-globin chains of haemoglobin A or from a single amino acid mutation (beta(6Glu-->Val)) of the beta-globin chain respectively. Although they have different pathophysiologies, patients with these conditions manifest both biochemical and clinical evidence of hypercoagulability. While the frequency of various thrombotic complications may vary in beta-thalassaemia and homozygous SCD [sickle cell anaemia (SCA)], patients with both diseases manifest decreased levels of natural anticoagulant proteins, as well as increased markers of thrombin generation and platelet activation. The abnormal phospholipid membrane assymetry present in the red blood cells of beta-thalassaemia and SCA patients, with resultant phosphatidylserine exposure appears to play a significant role in the aetiology of the observed hypercoagulable state. This review presents the available data on the aetiology and clinical manifestations of the coagulation and platelet activation that exist in both beta-thalassaemia and SCA, as well as the potential therapeutic implications resulting from this hypercoagulability.

Progressive iron overload is the most salient and ultimately fatal complication of beta-thalassemia. However, little is known about the relationship among ineffective erythropoiesis (IE), the role of iron-regulatory genes, and tissue iron distribution in beta-thalassemia. We analyzed tissue iron content and iron-regulatory gene expression in the liver, duodenum, spleen, bone marrow, kidney, and heart of mice up to 1 year old that exhibit levels of iron overload and anemia consistent with both beta-thalassemia intermedia (th3/+) and major (th3/th3). Here we show, for the first time, that tissue and cellular iron distribution are abnormal and different in th3/+ and th3/th3 mice, and that transfusion therapy can rescue mice affected by beta-thalassemia major and modify both the absorption and distribution of iron. Our study reveals that the degree of IE dictates tissue iron distribution and that IE and iron content regulate hepcidin (Hamp1) and other iron-regulatory genes such as Hfe and Cebpa. In young th3/+ and th3/th3 mice, low Hamp1 levels are responsible for increased iron absorption. However, in 1-year-old th3/+ animals, Hamp1 levels rise and it is rather the increase of ferroportin (Fpn1) that sustains iron accumulation, thus revealing a fundamental role of this iron transporter in the iron overload of beta-thalassemia.

Beta-thalassaemia is a congenital haemolytic anaemia characterized by partial (intermedia, TI) or complete (major, TM) deficiency in the production of beta-globin chains. The primary aim of this study was to determine the prevalence of thromboembolic events in patients with beta-thalassaemia. To achieve this, a multiple choice questionnaire was sent to 56 tertiary referral centres in eight countries (Lebanon, Italy, Israel, Greece, Egypt, Jordan, Saudi Arabia and Iran), requesting specific information on patients who had experienced a thromboembolic event. The study demonstrated that thromboembolic events occurred in a clinically relevant proportion (1.65%) of 8,860 thalassaemia patients (TI - 24.7% or TM - 75.3%) from the Mediterranean and Iran. Thromboembolism occurred 4.38 times more frequently in TI than TM (p < 0.001), with more venous events occurring in TI and more arterial events occurring in TM. Thrombosis in thalassaemia was also more common in females, splenectomized patients and those with profound anaemia (haemoglobin <9 g/dl). Due to the increased risk of thromboembolic events, the rationale for splenectomy should perhaps be re-assessed and the role of transfusion therapy for the prophylaxis of thrombosis, among other complications, be evaluated prospectively.

Beta-thalassaemia represents a group of diseases, in which ineffective erythropoiesis is accompanied by iron overload. In a mouse model of beta-thalassaemia, we observed that the liver expressed relatively low levels of hepcidin, which is a key factor in the regulation of iron absorption by the gut and of iron recycling by the reticuloendothelial system. It was hypothesised that, despite the overt iron overload, a putative plasma factor found in beta-thalassaemia might suppress liver hepcidin expression. Sera from beta-thalassaemia and haemochromatosis (C282Y mutation) patients were compared with those of healthy individuals regarding their capacity to induce changes the expression of key genes of iron metabolism in human HepG2 hepatoma cells. Sera from beta-thalassaemia major patients induced a major decrease in hepcidin (HAMP) and lipocalin2 (oncogene 24p3) (LCN2) expression, as well as a moderate decrease in haemojuvelin (HFE2) expression, compared with sera from healthy individuals. A significant correlation was found between the degree of downregulation of HAMP and HFE2 induced by beta-thalassaemia major sera (r = 0.852, P < 0.0009). Decreased HAMP expression was also found in HepG2 cells treated with sera from beta-thalassaemia intermedia patients. In contrast, the majority of sera from hereditary haemochromatosis patients induced an increase in HAMP expression, which correlated with transferrin (Tf) saturation (r = 0.765, P < 0.0099). Our results suggest that, in beta-thalassaemia, serum factors might override the potential effect of iron overload on HAMP expression, thereby providing an explanation for the failure to arrest excessive intestinal iron absorption in these patients.

beta-Thalassemia is an inherited anemia in which synthesis of the hemoglobin beta-chain is decreased. The excess unmatched alpha-globin chains accumulate in the growing erythroid precursors, causing their premature death (ineffective erythropoiesis). Clinical features of beta-thalassemia include variably severe anemia and iron accumulation due to increased intestinal iron absorption. The most anemic patients require regular blood transfusions, which exacerbate their iron overload and result in damage to vital organs. The hepatic peptide hepcidin, a key regulator of iron metabolism in mammals, was recently found to be low in the urine of beta-thalassemia patients, compared with healthy controls, despite their iron overload. In our work, we measured by RQ-PCR the liver mRNA expression of hepcidin and other iron regulatory genes in beta-thalassemia major mouse model (C57Bl/6 Hbb(th3/th3)), and compared it with beta-thalassemia intermedia mouse model (C57Bl/6 Hbb(th3/+)) and control mice. We found decreased expression of hepcidin and TfR2 and increased expression of TfR1 and NGAL in the beta-thalassemia mouse models, compared with the control mice. Significant down-regulation of hepcidin expression in beta-thalassemia major, despite iron overload, might explain the increased iron absorption typically observed in thalassemia.

The incidence of infections among patients with thalassaemia and the role of risk factors for infection are uncertain. We studied the occurrence of infections necessitating hospitalisation in 92 homozygous beta-thalassaemia patients who had been followed longitudinally for decades, and investigated the role of potential risk factors for these infections. Pneumonia accounted for 26% of the infections and fever of unknown origin for 14%. Staphylococcus aureus was the major pathogen possibly related to injections associated with intensive chelation with deferoxamine. There was a significant increase in the rate of infection over time, notably after 15 years. Splenectomy correlated with the incidence of infection (P < 0.001) without being confounded by other variables and with highest frequencies of infections present after 10 years. A direct correlation between iron overload and infection was evident only before the initiation of iron-chelating treatment (P < 0.01). Following initiation of deferoxamine, paradoxically, the infection rate increased (P = 0.046). The combination of splenectomy and deferoxamine treatment was associated with the highest adjusted infection rate. Parathyroid dysfunction and glucose-6-phosphate dehydrogenase deficiency were significantly associated with infection (P = 0.02 and P = 0.04 respectively). The infection rate in thalassaemia is affected mainly by the duration of the disease and is increased by splenectomy and, in the long term, by treatment with deferoxamine.

Sickle cell disease (SCD) is basically a red blood cell (RBC) disorder characterised by sickling and haemolysis, but platelets and polymorphonuclear neutrophils (PMN) are also involved. Oxidative damage may play a role in the pathogenesis of SCD. Using flow cytometry, we measured oxidative-state markers simultaneously in RBC, platelets and PMN obtained from 25 normal donors, nine homozygous (SS) patients and six SS/beta-thalassaemia patients. Reactive oxygen species (ROS) and reduced glutathione (GSH) were measured following staining of blood samples with fluorescence probes and gating on specific subpopulations based on size and granularity. Ten- to 30-fold higher ROS production and 20-50% lower GSH content were found in RBC, platelets and PMN from SCD patients versus those of their normal counterparts. This could in part account for the clinical manifestations, such as haemolysis, a hypercoagulable state, recurrent bacterial infections and vaso-occlusive incidences, in SCD. We further showed that exposure of SCD samples to antioxidants, such as N-acetyl-cysteine, vitamin C and vitamin E, decreased their oxidative stress. These results suggest that antioxidant treatment of patients with SCD could reduce oxidative damage to RBC, PMN and platelets, thereby alleviating symptoms associated with their pathology. The flow cytometry techniques presented herein could assist in monitoring the efficacy of such treatment.

Purpose: There is a lack of awareness among physicians, dieticians, and public health planners as to the prevalence of iron deficiency and anemia among adolescents undergoing strenuous physical training. The aim of this study was to estimate the prevalence of iron deficiency and anemia among male adolescents undergoing such activity., Methods: We studied 292 male adolescents on the day of entry into a volunteer military unit. Hemoglobin (Hb), mean corpuscular volume, ferritin, iron, iron-transferrin saturation, and soluble transferrin receptor (TfR) were measured, and TfR-F index was calculated., Results: The mean Hb level (+/-SD) in the study population was 14.7 +/- .9 g/dL (range, 10.8-16.8 g/dL), mean ferritin level was 50.6 +/- 32.6 ng/mL (range, 5.4-162.5 ng/mL), and mean iron level was 97.1 +/- 39.9 microg/dL (range, 24-267 microg/dL). The prevalence of anemia (Hb <14 g/dL) was 18.5%, and 3.4% had Hb concentrations less than 13 g/dL. Iron deficiency (ferritin <22 ng/mL) was present in 18% of the subjects, and 11.3% had ferritin levels less than 17 ng/mL. The mean soluble transferrin receptor concentration was 1.9 +/- .8 mg/L, and the mean TfR-F index was 1.21 +/- .57., Conclusions: Nearly 19% of the study subjects had mild anemia at recruitment, and depletion of iron stores was observed among 18%. Overall, these changes were not accompanied by a significant increase in soluble TfR. This high prevalence is most likely the result of "sports anemia" due to the intense physical training regimen adopted prior to their recruitment.

The pathophysiology of thalassemia is, to a certain extent, associated with the generation of labile iron in the pathological red blood cell (RBC). The appearance of such forms of iron at the inner and outer cell surfaces exposes the cell to conditions whereby the labile metal promotes the formation of reactive oxygen species (ROS) leading to cumulative cell damage. Another source of iron accumulation results from increased absorption due to decreased expression of hepcidin. The presence of labile plasma iron (LPI) was carried out using fluorescent probes in the FACS. RNA expression of hepcidin was measured in two models of thalassemic mice. Hepcidin expression was also measured in human hepatoma HepG2 cells following incubation with thalassemic sera. LPI was identified and could be quantitatively measured and correlated with other parameters of iron overload. Hepcidin expression was downregulated in the livers of thalassemic mice, in major more than in intermedia. Thalassemic sera down regulated hepcidin expression in HepG2 liver cells. A possible way to decrease iron absorption could be by modulating hepcidin expression pharmacologically, by gene therapy or by its administration. Treatment with combination of antioxidants such as N-acetylcysteine for proteins and vitamin E for lipids in addition to iron chelators could neutralize the deleterious effects of ROS and monitored by quantitation of LPI.

To develop new treatments for beta-thalassemia, it is essential to identify the genes involved in the relevant pathophysiological processes. Iron metabolism in thalassemia mice being investigated, focusing on the expression of a gene called hepcidin (Hamp), which is expressed in the liver and whose product (Hamp) is secreted into the bloodstream. In mice, iron overload leads to overexpression of Hamp, while Hamp-knockout mice suffer from hemochromatosis. The aim of this study is to investigate Hamp in the mouse model of beta-thalassemia and to address the potential gene transfer of Hamp to prevent abnormal iron absorption.

Unlabelled: beta thalassemia is an hereditary disease caused by mutations in the beta globin gene. The clinical course is characterized by chronic hemolytic anemia that required regular blood transfusions. Secondary to the blood transfusions, patients developed severe hemosiderosis that can cause death in the early twenties unless appropriate iron chelator therapy is given. Due to the severity of the disease and the expensive treatment, a prevention program should be instituted. We report our experience in a prevention program among pregnant women in the Jezreel and Eiron Valleys during a period of fifteen years., Results: A total of women were screened for beta thalassemia. Within this group, 928 were found to be carriers and 180 couples were found to be at risk to deliver an affected baby. Two hundred and fifty nine prenatal diagnoses were performed and in 45 cases the parents decided to abort the affected child. In ten cases the parents choose to deliver an affected baby. Fifteen different beta globin mutations were detected in the area covered by the program., Conclusions: A prevention program among pregnant women is feasible and prevents most of the new cases of hemoglobinopathies in the covered area., Recommendations: Based on our experience, and the relative low cost of this program, we recommend instituting a national prevention program for beta thalassemia throughout the country.

Thalassemia is a congenital hemolytic disorder caused by a partial or complete deficiency of alpha- or beta-globin chain synthesis. Homozygous carriers of beta-globin gene defects suffer from severe anemia and other serious complications from early childhood. The disease is treated by chronic blood transfusion. However, this can cause severe iron overload resulting in progressive organ failure. Some forms of alpha thalassemia are also associated with a similar clinical picture. Despite the difficulties associated with treatment, standards of care for thalassemic patients have improved in recent years, resulting in almost doubling of the average life expectancy. As a consequence, additional previously undescribed, complications are now being recognized. In particular, profound hemostatic changes have been observed in patients with beta-thalassemia major (beta-TM) and beta-thalassemia intermedia (beta-TI) and also in patients with alpha thalassemia (hemoglobin H disease). The presence of a higher than normal incidence of thromboembolic events, mainly in beta-TI, and the existence of prothrombotic hemostatic anomalies in the majority of the patients, even from a very young age, have led to the recognition of the existence of a chronic hypercoagulable state in thalassemic patients. Despite the appearance of numerous publications on the frequent occurrence of thromboembolic complications in thalassemia, this complication has not been emphasized or comprehensively reviewed. This review summarizes the current literature and discusses possible mechanisms of the lifelong hypercoagulable state that exists in thalassemia.