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1.
A method for the determination of desferrioxamine-available iron in tissue fractions is described which involves incubation with desferrioxamine, extraction of desferrioxamine and its iron-bound form, ferrioxamine, and quantitation of these two forms of the drug by reversed-phase hplc analysis. Chelatable iron levels in the 1-10µMolar region could be accurately and reproducibly measured using this technique.

The desferrioxamine-available iron levels in both the cortex and medulla of rabbit kidneys were significantly elevated (up to 2-fold) after the organs had been subjected to 2 hours warm ischaemia or 24 hours cold storage at 0°C In hypertonic citrate solution. There was no change in the total iron content of the tissues under these circumstances and thus a redistribution of intracellular iron to more available pools had presumably taken place as a result of ischaemia. This redistribution of iron may be an important factor in the initiation of peroxidative damage to cell membranes upon reperfusion of the organ with oxygen.  相似文献   

2.
Intracellular iron loosely bound to proteins such as ferritin or in the form of low molecular weight chelates is available to catalyze adverse reactions such as the formation of reactive free radicals. A method to measure this small but important iron pool by utilizing the highly specific iron-chelator desferrioxamine is described. Following incubation of tissue fractions with desferrioxamine, the parent compound and its iron-bound form, ferrioxamine, are extracted using solid-phase cartridges and quantitated by reversed-phase HPLC using uv detection. Calculation of the ferrioxamine:desferrioxamine ratio and comparison with a standard curve constructed using a series of known iron concentrations allow the determination of micromolar amounts of desferrioxamine-available iron in biological samples.  相似文献   

3.
We have studied the facilitation of iron transfer from transferrin to desferrioxamine by various anions. Most of the anions which can substitute for HCO3? in the ternary complex of transferrin · Fe · HCO3 do not facilitate iron transfer; anions which do facilitate iron transfer do not necessarily form stable ternary complexes. Combinations of anions effective in transfer have a less-than-additive effect, suggesting a common reaction pathway. We suggest that the transfer of iron from transferrin to desferrioxamine involves a substitution step and a subsequent chelation step, and that the efficiency of the overall reaction is a function of both these attributes of the anion.  相似文献   

4.
《Free radical research》2013,47(6):317-320
The normal brain contains regions with high concentrations of iron, part of which appears to be in a low molecular mass chelatable form. Iron complexes with a molecular mass of below 10,000, were measured in ultrafiltrates of homogenized gerbil brains using the bleomycin assay, and were found to average 20.5 ± 3.5 μM (n = 8). As expected, no bleomycin detectable iron was found in the plasma of these animals.

No obvious difference in the tissue levels of bleomycin-detectable iron was recorded following ischaemia and reperfusion. This is probably due to the already abundant presence of iron in the brain and the likely release of iron from protected sites due to structural damage inherent in the preparative procedures used.  相似文献   

5.
Iron metabolism in K562 erythroleukemic cells   总被引:7,自引:0,他引:7  
Iron delivery to K562 cells is enhanced by desferrioxamine through induction of transferrin receptors. Experiments were performed to further characterize this event with respect to iron metabolism and heme synthesis. In control cells, up to 85% of the iron taken up from iron-transferrin was incorporated into ferritin, 7% into heme, and the remainder into compartments not yet identified. In cells grown with desferrioxamine, net accumulation of intracellular desferrioxamine (14-fold) was observed and iron incorporation into ferritin and heme was inhibited by 86% and 75%, respectively. In contrast, complete inhibition of heme synthesis in cells grown with succinylacetone had no effect on transferrin binding or iron uptake. Exogenous hemin (30 microM) inhibited transferrin binding and iron uptake by 70% and heme synthesis by 90%. These effects were already evident after 2 h. Thus, although heme production could be reduced by desferrioxamine, succinylacetone, and hemin, cell iron uptake was enhanced only by the intracellular iron chelator. The effects of exogenous heme are probably unphysiologic and the greater inhibition of iron flow into heme can be explained by effects on early steps of heme synthesis. We conclude that in this cell model a chelatable intracellular iron pool rather than heme synthesis mediates regulation of iron uptake.  相似文献   

6.
The effect of changes in iron availability and induction of differentiation on transferrin receptor expression and ferritin levels has been examined in the promonocytic cell line U937. Addition of iron (as 200 micrograms/ml saturated transferrin) or retinoic acid (1 microM) both caused approx. 70% reduction in the average number of surface transferrin receptors, while the iron chelator desferrioxamine caused an 84% increase. Comparable changes also occurred in the levels of transferrin receptor mRNA. Neither iron nor retinoic acid significantly altered the half-life of transferrin receptor mRNA in the presence of actinomycin D (approx. 75 min) but a 10-fold increase in stability occurred in the presence of desferrioxamine. Iron and retinoic acid both caused an increase in intracellular ferritin levels (approx. 4-and 3-fold, respectively), while desferrioxamine reduced ferritin levels by approx. two-thirds. The effect of iron and retinoic acid added together did not differ greatly from that of each agent alone. None of the treatments greatly affected levels of L-ferritin mRNA. Virtually no H-ferritin mRNA was detected in U937 cells. These results show that changes in ferritin and transferrin receptor caused by treatment with retinoic acid are similar to those induced by excess iron, and suggest that changes in these proteins during cell differentiation are due to redistribution of intracellular iron into the regulatory pool(s), rather than to iron-independent mechanisms.  相似文献   

7.
The main iron chelator used for transfusional iron overload is desferrioxamine, which is expensive, has toxic side effects, and has to be given subcutaneously. An orally active iron chelator is therefore required. The effects of oral 1,2-dimethyl-3-hydroxypyrid-4-one on urinary iron excretion were studied in eight patients who had received multiple transfusions: four had myelodysplasia and four beta thalassaemia major. Different daily doses of the drug up to 100 mg/kg/day, alone or in combination with ascorbic acid, were used. In three patients with thalassaemia the effect of the drug was compared with that of subcutaneous desferrioxamine at the same daily dose. In all eight patients a single dose of oral 1,2-dimethyl-3-hydroxypyrid-4-one resulted in substantial urinary iron excretion, mainly in the first 12 hours. Urinary iron excretion increased with the dose and with the degree of iron loading of the patient. Giving two or three divided doses over 24 hours resulted in higher urinary iron excretion than a single dose of the same amount over the same time. In most patients coadministration of oral ascorbic acid further increased urinary iron excretion. 1,2-Dimethyl-3-hydroxypyrid-4-one caused similar iron excretion to that achieved with subcutaneous desferrioxamine at a comparable dose. In some cases the iron excretion was sufficiently high (maximum 99 mg/day) to suggest that a negative iron balance could be easily achieved with these protocols in patients receiving regular transfusions. No evidence of toxicity was observed on thorough clinical examination or haematological and biochemical testing in any of the patients. None of the patients had any symptoms that could be ascribed to the drug. These results suggest that the oral chelator 1,2-dimethyl-3-hydroxypyrid-4-one is as effective as subcutaneous desferrioxamine in increasing urinary iron excretion in patients loaded with iron. Its cheap synthesis, oral activity, and lack of obvious toxicity at effective doses suggest that it should be developed quickly and thoroughly tested for the management of transfusional iron overload.  相似文献   

8.
The release of iron from horse spleen ferritin by the chelating agents desferrioxamine B, rhodotorulic acid, 2,3-dihydroxybenzoate, 2,2′-bipyridyl and pyridine-2-aldehyde-2-pyridyl hydrazone (Paphy) has been studied in vitro. Ferritin prepared by classical procedures involving thermal denaturation releases its iron less effectively than ferritin isolated by a modified procedure that avoids this step. Desferrioxamine B and rhodotorulic acid are the most effective in releasing iron from both preparations of ferritin. When FMN is added, iron release by desferrioxamine B, rhodotorulic acid, and 2,3-dihydroxybenzoate was effectively blocked, whereas both bipyridyl and Paphy showed a marked simulation. A substantial increase in iron release was also observed for bipyridyl and Paphy with ascorbate; a less important increase was noted for rhodotorulic acid. EDTA exerted a marked inhibition of iron release from ferritin with rhodotorulic acid, 2,3-dihydroxybenzoate, bipyridyl, and Paphy. The effects of citrate and oxalate on iron release by the chelators was small. The effect of the concentration of flavin on iron release from ferritin by bipyridyl and desferrioxamine B have been studied. Desferrioxamine is unable to mobilize FeII from ferritin following reduction by reduced FMN, whereas bipyridyl can rapidly complex the ferrous iron. The results are discussed in the context of our current concepts of storage iron mobilization in the treatment of iron overload.  相似文献   

9.
In this work we have investigated the effects of iron-induced free radical formation in normal human erythrocytes in vitro, as a model system for studying iron damage, and in erythrocytes from patients with β-thalassaemia major. The resulting oxidative effects were measured in terms of methaemoglobin formation and reduced glutathione loss. The effects of desferrioxamine, an iron-chelating agent, were also investigated.

The results show that the increased methaemoglobin formation after iron-induced oxidative stress is consistent with a decline in the intracellular glutathione levels and that this process is inhibited by desferrioxamine. Similar treatment of red cell haemolysates produces less methaemoglobin. This suggests that, on exposure of intact erythrocytes to iron-induced free radical effects, the red cell membrane exacerbates the breakdown of the antioxidant defences of the cell and the oxidation of haemoglobin.  相似文献   

10.
《Free radical research》2013,47(3-6):255-264
An overview of a series of experiments attempting to link iron and calcium redistribution and release of free fatty acids with falls in pH and adenine nucleotide levels during cold storage of rabbit kidneys is presented. The data reviewed strongly suggest that these events are inextricably linked to subsequent reperfusion injury. Circumstantial evidence incriminating iron was provided by experiments showing that iron chelation decreased reperfusion injury after warm (WI) and cold ischaemia (CI) in rat skin flap and rabbit kidney models. Evidence for a role for calcium was provided when it was found that a calcium channel blocking agent added to the saline flush solution before storage inhibited lipid peroxidation, whereas chemicals which caused release or influx of calcium into the cell exacerbated oxidative damage. Additional involvement of breakdown products of adenine nucleotides was suggested by the protection from lipid peroxidation afforded by allopurinol. Involvement of calcium-activated phospholipase A, was strongly suggested by increases In free fatty acids during cold storage and both this increase and lipid peroxidation were inhibited by addition of dibucaine to the storage solution.  相似文献   

11.
The iron chelators desferrioxamine and 1,2-dimethyl-3-hydroxypyrid-4-one (L1) inhibited human platelet aggregation in vitro as well as thromboxane A2 synthesis and conversion of arachidonate to lipoxygenase-derived products. Non-chelating compounds related to L1 were without effect on cyclooxygenase or lipoxygenase activity. Since both cyclooxygenase and lipoxygenase are iron-containing enzymes, it is suggested that the inhibition of platelet function by these iron chelators may be related to the removal or binding of iron associated with these enzymes. These iron chelators may therefore be of potential therapeutic value as platelet antiaggregatory agents and of possible use in the treatment of atherosclerotic and inflammatory joint diseases.  相似文献   

12.
The elemental content of the iron cores of haemosiderins isolated from animal and human tissues has been determined to ascertain whether changes in composition are correlated with structural differences previously identified in these mineralisation products. Significant differences were observed in the elemental composition of haemosiderins isolated from patients subjected to desferrioxamine-chelation therapy compared to patients who had been venesected. The P/Fe molar ratio was considerably higher in haemosiderin isolated from treated primary haemochromatosis (0.83), compared to untreated primary haemochromatosis (0.10) and treated secondary haemochromatosis (0.25), and this could account for the amorphous nature of these iron cores. The levels of M/Fe (M = Ca, Cu, Zn) were reduced in the haemosiderins derived from treated secondary haemochromatosis patients, possibly due to the chelation of these ions by desferrioxamine therapy. In an experimentally iron-loaded rat, receiving either desferrioxamine or 1,2-diethyl-3-hydroxypyrid-4-one, selective decreases in these three elements were also observed after two weeks of desferrioxamine therapy. Such changes may be important determinants in the modification of biomineralisation of the iron cores.  相似文献   

13.
《Free radical research》2013,47(1-2):125-127
The carrageenin rat paw edema was dose-dependently inhibited by i.v. and i.m. administration of desferrioxamine, a specific iron chelating agent. Therefore, iron-catalyzed formation of free radicals might be involved in this acute inflammatory reaction. In contrast, no antiinflammatory activity of desferrioxamine could be seen in rat adjuvant arthritis, a model of subacute and chronic inflammation.  相似文献   

14.
Abstract. Iron chelating agents have been demonstrated to inhibit tumour cell growth. However, in vitro and in vivo results using desferrioxamine a hexadentate iron chelating agent, for anti-cancer treatment are not always in agreement. Therefore, we have studied the response of three human tumour cell lines (HL-60 promyelocytic leukaemia, MCF-7 breast cancer and HepG2 hepatoma), grown in culture medium supplemented with either human pooled (HPS) or fet al bovine serum (FBS), to desferrioxamine. Desferrioxamine, at micromolar concentrations, induced severe cytotoxicity in all tumour cell lines grown in FBS medium. When grown in HPS medium, comparable desferrioxamine cytotoxicity was observed in the millimolar range. The addition of 50% saturated human transferrin to FBS medium resulted in protection against desferrioxamine cytotoxicity. HL-60 cells were further studied for iron metabolism characteristics. HL-60 cells, grown in medium with FBS, were found to have an 8.4 fold increase in surface transferrin receptor (TfR) expression ( P < 0.001) as compared with HL-60 cells grown in medium with HPS. However, iron uptake of HPS cultured HL-60 cells, after incubation with saturated human transferrin, was higher, resulting in a higher concentration of iron in HPS cultured HL-60 cells as compared with FBS cultured cells (1.72 ± 0.02 μmol/g protein v. 1.32 ± 0.14 μmol/g protein; P < 0.001). Using desferrioxamine it was shown that TfR expression is dependent on the biological availability of iron in the cell. Consistent with the lower iron content in FBS cultured cells, we conclude that the cytotoxicity of desferrioxamine is dependent on the ability of cells to replenish cellular iron stores from the culture medium. Cells grown in FBS medium lack this ability and are therefore more susceptible to desferrioxamine.  相似文献   

15.
Cell surface expression of the dopamine transporter (DAT) is determined by the relative rates of its internalization and recycling. Changes in the cellular labile iron pool (LIP) affect many cellular mechanisms including those that regulate DAT trafficking. In this study, we analyzed DAT expression and posttranslational modifications in response to changes in cellular iron in transfected neuroblastoma cells (N2a). Iron chelation by desferrioxamine (DFO) altered DAT protein levels by decreasing the stability of DAT mRNA. Increased phosphorylation and ubiquitination of this transporter protein following DFO treatment were also observed. Cellular iron depletion elevated protein levels of the early endosomal marker Rab5. Moreover, confocal microscopy studies showed increased localization of DAT into the endosomal compartment in DFO-treated cells compared to control. Together, these findings suggest that cellular iron depletion regulates DAT expression through reducing mRNA stability as well as an increasing in endocytosis.  相似文献   

16.
Desferrioxamine, an iron chelator with “hypoxia-mimetic” activity, promotes bone mineralization when used in aluminum-overloaded dialysis patients. However, the effect of desferrioxamine on osteoblastic differentiation from pluripotent mesenchymal stem cells (MSCs) has not been reported. In this study, pluripotent human MSCs and murine mesenchymal C3H10T1/2 cells were simultaneously treated with desferrioxamine and bone morphogenetic protein-2 (BMP2). In BMP2-treated MSCs, desferrioxamine levels of 15 μΜ were found to increase alkaline phosphatase (ALP) activity and calcium deposition, which were the markers of osteoblastic differentiation. These effects of desferrioxamine were accompanied by promoted phosphorylation of glycogen synthase kinase 3β (GSK-3β) and increased β-catenin protein content, a direct GSK-3β substrate. Knockdown of β-catenin by RNA interference eliminates this positive effect of desferrioxamine on ALP activity. Taken together, these data demonstrate that desferrioxamine plays a direct role in the differentiation of mesenchymal stem cells by activating β-catenin signaling cascades.  相似文献   

17.
18.
Iron is released in a desferrioxamine (DFO)-chelatable form (DCI) when erythrocytes are challenged by an oxidative stress. In &#103 -thalassemic erythrocytes, both DCI content and release (after aerobic incubation for 24 h) are increased and correlated with the fetal hemoglobin (HbF) levels. Since erythrocytes from newborns have an extremely high content of HbF and are exposed to conditions of oxidative stress, the release of iron in these erythrocytes was investigated. The erythrocyte DCI content was increased in preterm but not in term newborns as compared to adults, while the release was increased in both preterm and term erythrocytes. The level of plasma non protein-bound iron (NPBI), which was not detectable in adults, was much higher in preterm than in term newborns. When term plus preterm newborns were divided in two groups, normoxic and hypoxic, according to cord blood pH, it was found that both iron release and NBPI were markedly higher in the hypoxic newborns compared to normoxic ones. Similar results were also obtained when the preterm and term infants were considered separately on the basis of cord blood pH. Therefore, iron release and NPBI are higher when conditions of hypoxia occur. In fact, when the values for iron release and NPBI were separately plotted against cord blood pH values, significant negative correlations were seen in both cases. NPBI was correlated with iron release seen in all the newborns and a significant part of the released iron could be recovered into the incubation medium at the end of the incubation.  相似文献   

19.
J M Gutteridge 《FEBS letters》1986,201(2):291-295
Hydrogen peroxide and organic hydroperoxides react with haemoglobin to release iron which can be complexed to apotransferrin, bleomycin and desferrioxamine. This released iron promotes deoxyribose degradation by a Fenton reaction, DNA degradation in the presence of bleomycin and stimulates lipid peroxidation. It is likely that iron released from haemoglobin is the true generator of hydroxyl radicals in the Fenton reaction.  相似文献   

20.
A 32 year old woman with severe aplastic anaemia required frequent transfusions and consequently developed hyperferrioxaemia (54 microMol/l) and hyperferritinaemia (1,700 ng/ml). For the treatment of transfusion siderosis she was given 18 high dose courses each comprising 35 g of desferrioxamine. Because of pre-existing thrombocytopenia (platelet count 5 X 10(9)/l) the iron chelating agent was given by continuous intravenous infusion over 3 1/2 days. High dose desferrioxamine had to be abandoned because of severe bone pain. The desferrioxamine infusions achieved a negative iron balance, iron loss after each infusion being 100 to 200 mg in the urine and 400 mg in the faeces. Serum iron and ferritin concentrations fell almost to normal. This report shows that faecal iron excretion must be taken into account in assessing the balance of iron input and output during desferrioxamine treatment.  相似文献   

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