Chelation therapy with desferrioxamine does not normalize ferritin level but attenuates oxidative damage and improves total antioxidant level in Malaysian Chinese beta-thalassaemia major patients.

Beta-thalassaemia major causes severe anaemia and patients with it may be transfusion-dependent for life. Regular blood transfusions cause iron-overload that leads to oxidative damage which can hasten mortality. The objective of this research was to study the oxidant-antioxidant indices in beta-thalassaemia major patients at the University of Malaya Medical Centre (UMMC) who were on desferrioxamine-chelation or without chelation therapy. Blood was collected from 39 Chinese patients and 20 controls. Plasma and peripheral blood mononuclear cell lysates (PBMC) were extracted and biochemical tests to evaluate oxidative stress were performed. Oxidative stress was evident in these patients as advanced oxidized protein products (AOPP) and lipid hydroperoxides were elevated, whereas glutathione peroxidase activity and the ferric reducing antioxidant power (FRAP) were reduced. The catalase activity in the patients' PBMC was elevated, possibly as a compensatory mechanism for the reduced glutathione peroxidase activity in both red blood cells and PBMC. The lower FRAP and higher AOPP levels in the non-chelated patients compared with the chelated patients were indicative of a lower oxidative stress level in the chelated patients. The ferritin levels in the chelated and non-chelated patients were high and the mean levels of liver enzyme activities in the majority of patients were elevated regardless of chelation therapy. In conclusion, this study indicates that desferrioxamine chelation therapy does not normalize ferritin level but attenuates oxidative damage and improves total antioxidant level in Malaysian Chinese beta-thalassaemia major patients.

Chelation therapy with desferrioxamine does not normalize ferritin level but attenuates oxidative damage and improves total antioxidant level in malaysian Chinese β-thalassaemia major patients / La terapia de quelación con deferoxamina no normaliza los niveles de ferritinaa pero atenúa el daño oxidativo y mejora el nivel antioxidante total en los pacientes sinomalayos que padecen de talasemia β

Beta-thalassaemia major causes severe anaemia and patients with it may be transfusion-dependent for life. Regular blood transfusions cause iron-overload that leads to oxidative damage which can hasten mortality. The objective of this research was to study the oxidant-antioxidant indices in β-thalassaemia major patients at the University of Malaya Medical Centre (UMMC) who were on desferrioxaminechelation or without chelation therapy. Blood was collected from 39 Chinese patients and 20 controls. Plasma and peripheral blood mononuclear cell lysates (PBMC) were extracted and biochemical tests to evaluate oxidative stress were performed. Oxidative stress was evident in these patients as advanced oxidized protein products (AOPP) and lipid hydroperoxides were elevated, whereas glutathione peroxidase activity and the ferric reducing antioxidant power (FRAP) were reduced. The catalase activity in the patients' PBMC was elevated, possibly as a compensatory mechanism for the reduced glutathione peroxidase activity in both red blood cells and PBMC. The lower FRAP and higher AOPP levels in the non-chelated patients compared with the chelated patients were indicative of a lower oxidative stress level in the chelated patients. The ferritin levels in the chelated and non-chelated patients were high and the mean levels of liver enzyme activities in the majority of patients were elevated regardless of chelation therapy. In conclusion, this study indicates that desferrioxamine chelation therapy does not normalize ferritin level but attenuates oxidative damage and improves total antioxidant level in Malaysian Chinese β-thalassaemia major patients.

La beta-talasemia mayor causa anemia severa, y los pacientes con este padecimiento pueden hacerse dependientes de las transfusiones de sangre por el resto de sus vidas. Las transfusiones regulares de sangre dan lugar a una sobrecarga de hierro que conduce al dano oxidativo, el cual a su vez puede acelerar la mortalidad. El objetivo de esta investigación fue estudiar las tasas de oxidantesantioxidantes en pacientes de beta-talasemia mayor en el Centro Médico de la Universidad de Malaya, tanto aquellos bajo tratamiento de quelación con deferoxamina, como aquellos sin terapia de quelación alguna. Se recogieron muestras de sangre de 39 pacientes chinos y 20 controles. Se extrajeron plasma y lisados de celulas mononucleares perifericas (CMSP), y se realizaron pruebas bioquimicas para evaluar el estrés oxidativo. El estrés oxidativo era evidente en estos pacientes en forma de productos avanzados de oxidación de proteinas (PAOP), y los hidroperoxidos de lipidos eran elevados, en tanto que la actividad de glutatión peroxidasa y el poder reductor ferrico/antioxidante (FRAP) era reducida. La actividad de la catalasa en los pacientes de CMSP era elevada, posiblemente como un mecanismo compensatorio frente a la actividad de glutatión peroxidasa reducida tanto en los globulos rojos como en las CMSP. Los niveles más bajos de FRAP y los más altos de PAOP en los pacientes no quelados en comparación con los pacientes quelados, indicaban un bajo nivel de estrés oxidativo en los pacientes quelados. Los niveles de ferritina tanto en los pacientes quelados como en los no quelados, eran altos, y los niveles promedio de actividades enzimaticas del higado fueron elevados en la mayoria de los pacientes, independientemente de la terapia de quelación. En conclusión, este estudio indica que la terapia de quelación con deferoxamina no normaliza el nivel de ferritina, pero en cambio atenua el daño oxidativo, y mejora el nivel antioxidante total en los pacientes sinomalayos afectados por la betatalasemia mayor.

Glutathione S-transferase M1 gene polymorphisms are associated with cardiac iron deposition in patients with beta-thalassemia major.

Patients with beta-thalassemia (thal) major are subject to peroxidative tissue injury by iron overload. Glutathione S-transferases work as antioxidants, and their activity is determined genetically. In this study, we used multiplex polymerase chain reaction (m-PCR) to analyze polymorphisms of two endogenous antioxidant agents, glutathione S-transferase M1 (GSTM1) and glutathione S-transferase T1 (GSTT1), and to determine their roles in 41 patients with beta-thal major. Our results showed that the GSTM1 and GSTT1 null genotypes were not associated with any incidence of endocrine dysfunction (including diabetes mellitus, hypogonadism, hypothyroidism, and growth hormone deficiency), liver function, or impaired left ventricular ejection fraction (LVEF). The GSTM1 null genotype, but not the GSTT1 null genotype, was associated with a decreased signal intensity ratio on cardiac magnetic resonance imaging (MRI). Our results suggest that genetic variations of the GSTM1 enzyme are associated with cardiac iron deposition in patients with beta-thal major.

Elevated plasma chemokine CCL18/PARC in beta-thalassemia.

Plasma CCL18/PARC, a member of the CC chemokine family, has been found to be several ten-fold increased in symptomatic Gaucher type I patients. Elevated plasma chitotriosidase levels are a well-known abnormality in Gaucher patients, however, its diagnostic use is limited by the frequent genetic deficiency in the protein. Like the situation in Gaucher disease, lipids accumulate in macrophages of patients suffering from beta-thalassemia, and, in both conditions, increased chitotriosidase levels occur. We here report that plasma CCL18/PARC is also significantly increased in patients with beta-thalassemia major (range 76.8-4977.8, median=650.8 ng/ml, n=36 and control range 10-72, median=33 ng/ml n=36 respectively, P<0.001). The CCL18/PARC levels are lower than in Gaucher patients (range 174.8-10798.7, median 2538.2 ng/ml, n=28, P<0.001). In our cohort of beta-thalassemic patients, CCL18/PARC showed a significant negative correlation to iron chelation therapy and a significant positive correlation to ferritin and chitotriosidase levels, the latter only in the patients with the wild type genotype for the enzyme. Our study demonstrates that beta-thalassemic patients have increased CCL18/PARC levels that could be of value in monitoring iron overload and compliance to therapy.

HCV and HGV infection, iron overload and liver disease in multitransfused patients with thalassaemia and persistently normal or abnormal transaminase levels.

Prevalence and influence on liver disease of HCV and HGV infections, and HCV genotypes were studied in 28 HCV-Ab positive multitransfused thalassaemia patients with persistently normal ALT levels (group A) matched by sex and age with 28 patients with increased ALT levels (group B). Laboratory and virologic tests (all patients), liver biopsy (28 patients) and LIC by SQUID (30 patients) were performed. In group A, HCV-RNA was positive in 39%, genotype 2a was detected in 91%. In Group B, HCV-RNA was positive in 89%, prevalence of genotype 1b and 2a was 52% and 48% respectively; compared with group A, they had significantly increased values of gammaGT, AF, BA, TP, IgG, IgA, LIC (group B: 2,142 -/+ 1,524 microg/g liver; group A: 1,084 -/+ 610 microg/g liver). Overall prevalence of HGV-RNA was low (12.5%) and not significantly different between groups. Liver biopsies revealed no cirrhosis and severe fibrosis was found in 3 HCV viremic patients in group B. In 14 viremic patients examined both for LIC and liver histology, mild fibrosis was observed in 71%, in which iron overload was below 5 times the normal value. In conclusion, in patients with normal ALT levels, active HCV infection must be excluded by evaluation of HCV-RNA. Liver biopsy is indicated in HCV viremic patients, independent of ALT levels; in non-viremic patients, increased ALT levels may be due to iron overload and LIC measurement is indicated. Our data emphasise the crucial role of chelation therapy to maintain low LIC levels in order to prevent progression of fibrosis to cirrhosis in patients with HCV chronic hepatitis.

Neutrophil elastase in patients with homozygous beta-thalassemia and pseudoxanthoma elasticum-like syndrome.

In this study we investigated the possible role of neutrophil (PMN) elastase and its natural inhibitor, alpha1-proteinase inhibitor (alpha1-PI) in the pathogenesis of the pseudoxanthoma elasticum (PXE)-like syndrome which is found in patients with homozygous beta-thalassemia. We studied 30 beta-thalassemia homozygotes with the PXE-like syndrome [PXE(+) group], 20 beta-thalassemia homozygotes without this syndrome [PXE(-) group] and 15 healthy controls. Plasma PMN elastase concentration in the PXE(+) and in the PXE(-) group was 136.4 +/- 89 and 163.8 +/- 126 microg/L, respectively (P > 0.05). In the control group, the concentration was 42.9 +/- 16.8 microg/L (P < 0.01 for the comparison with both patients' groups). The plasma alpha1-PI concentration in the PXE(+) and in the PXE(-) group was 2.28 +/- 0.75 and 2.6 +/- 0.96 g/L, respectively (P > 0.05). Using logistic regression, we studied the prognostic value for PXE of the following independent variables: number of transfusions, chelation therapy, mean hemoglobin concentration, PMN elastase concentration, alpha1-PI concentration, chronic transaminase elevation, and positivity for anti-HCV. None of the above variables was found to have significant prognostic value for the PXE. Plasma PMN elastase concentration is elevated in all beta-thalassemia homozygotes; its role in the pathogenesis of the PXE-like syndrome in beta-thalassemia can not be established, but our findings suggest that neutrophils of beta-thalassemia patients are activated, since PMN elastase is a marker of neutrophil activation.

Iron overload and urinary lysosomal enzyme levels in beta-thalassaemia major.

UNLABELLED: The urinary levels of the lysosomal enzymes N-acetyl-beta-D-glucosaminidase (NAG) (EC 3.2.1.52) and alpha-mannosidase (EC 3.2.1.24) were evaluated in patients with beta-thalassaemia major and normal control subjects. Two groups of patients with different degrees of iron overload, as judged by their serum ferritin levels, were investigated. Renal disease was not present in any of the patients. A statistically significant increase in the levels of NAG was observed in the high ferritin (> 3,000 mg/dl) group compared to the low ferritin (< 3,000 mg/dl) and the control groups. No difference was observed in the urinary alpha-mannosidase levels between the groups examined. The finding of increased NAG levels in the patients with the increased iron load suggests that kidney lysosomes are a target of iron toxicity. The different behaviour of the two lysosomal enzymes may reflect the intra- and inter-lysosomal heterogeneity in kidney. CONCLUSION: Iron overload resulted in increased urinary levels of the lysosomal enzyme NAG which has been proposed as an early marker of kidney damage. Reduction of iron load, achieved by regular desferrioxamine infusion, resulted in normalisation of the urinary enzyme levels. Thus kidney lysosomes appear to be a target and possibly a mediator of iron toxicity in this tissue.

Effects of deferoximine on chondrocyte alkaline phosphatase activity: proxidant role of deferoximine in thalassemia.

The homozygous form of beta-thalassemia, the most common single gene disorder, is treated by red cell transfusion therapy. Following transfusion, the chelator, deferoximine, is administered to patients to remove excess iron. However, when this drug is given to young children, metaphyseal dysplasia and abnormalities of linear growth are frequently observed. To explore the notion that deferoximine interferes with endochondral growth by chelating zinc, we examined the effect of the drug on chondrocytes maintained in long-term culture. We found that deferoximine caused a dose-dependent inhibition of a wide range of functions including cell proliferation, protein synthesis (and possibly under-hydroxylation of type X collagen), and mineral deposition. Directly relevant to the mineralization process was the observation that the drug dramatically lowered the activity of alkaline phosphatase, a zinc-requiring enzyme. To test the hypothesis that enzyme inhibition was due to chelation of zinc by deferoximine, the cell culture medium was supplemented with excess zinc. However, this treatment did not overcome the deferoximine-dependent change in enzyme activity. We next examined the possibility that deferoximine, in the presence of ascorbate, could form a free radical system that would serve to inactivate the enzyme. Using alkaline phosphatase extracted from chick cartilage, we noted that the activity of the phosphatase was markedly reduced in the presence of deferoximine and ascorbate. These effects were consistant with the notion that deferoximine and ascorbate can act as a prooxidant couple. This conclusion was confirmed when we measured the oxidative activities of the system using nitrobule tetrazolium and cytochrome c. Indeed, we noted that deferoximine markedly activates the autocatalytic oxidation of ascorbate.(ABSTRACT TRUNCATED AT 250 WORDS)