Robert C. Hider

Chemistry and biology of siderophores

Siderophores are compounds produced by bacteria, fungi and graminaceous plants for scavenging iron from the environment. They are low-molecular-weight compounds (500-1500 daltons) possessing a high affinity for iron(III) (Kf > 1030), the biosynthesis of which is regulated by iron levels and the function of which is to supply iron to the cell. This article briefly describes the classification and chemical properties of siderophores, before outlining research on siderophore biosynthesis and transport. Clinically important siderophores and the therapeutic potential of siderophore design are described. Appendix 1 provides a comprehensive list of siderophore structures.

Identification of an Intestinal Heme Transporter

Dietary heme iron is an important nutritional source of iron in carnivores and omnivores that is more readily absorbed than non-heme iron derived from vegetables and grain. Most heme is absorbed in the proximal intestine, with absorptive capacity decreasing distally. We utilized a subtractive hybridization approach to isolate a heme transporter from duodenum by taking advantage of the intestinal gradient for heme absorption. Here we show a membrane protein named HCP 1 (heme carrier protein 1), with homology to bacterial metal-tetracycline transporters, mediates heme uptake by cells in a temperature-dependent and saturable manner. HCP 1 mRNA was highly expressed in duodenum and regulated by hypoxia. HCP 1 protein was iron regulated and localized to the brush-border membrane of duodenal enterocytes in iron deficiency. Our data indicate that HCP 1 is the long-sought intestinal heme transporter.

Evaluating the Toxicity of Airborne Particulate Matter and Nanoparticles by Measuring Oxidative Stress Potential—A Workshop Report and Consensus Statement

Background: There is a strong need for laboratory in vitro test systems for the toxicity of airborne particulate matter and nanoparticles. The measurement of oxidative stress potential offers a promising way forward. Objectives:Aworkshop was convened involving leading workers from the field in order to review the available test methods and to generate a Consensus Statement. Discussions: Workshop participants summarised their own research activities as well as discussion the relative merits of different test methods. Conclusions: In vitro test methods have an important role to play in the screening of toxicity in airborne particulate matter and nanoparticles. In vitro cell challenges were preferable to in vitro acellular systems but both have a potential major role to play and offer large cost advantages relative to human or animal inhalation studies and animal in vivo installation experiments. There remains a need to compare tests one with another on standardised samples and also to establish a correlation with the results of population-based epidemiology.

The crucial role of metal ions in neurodegeneration: the basis for a promising therapeutic strategy

The variety of factors and events involved in neurodegeneration renders the subject a major challenge. Neurodegenerative disorders include a number of different pathological conditions, which share similar critical metabolic processes, such as protein aggregation and oxidative stress, both of which are associated with the involvement of metal ions. In this review, Alzheimers disease, Parkinsons disease and prion disease are discussed, with the aim of identifying common trends underlying these devastating neurological conditions. Chelation therapy could be a valuable therapeutic approach, since metals are considered to be a pharmacological target for the rationale design of new therapeutic agents directed towards the treatment of neurodegeneration.

The Relationship of Intracellular Iron Chelation to the Inhibition and Regeneration of Human Ribonucleotide Reductase

The depletion of cellular iron can lead to the inhibition of ribonucleotide reductase, preventing new DNA synthesis and hence inhibiting cell proliferation. Electron paramagnetic resonance (EPR) spectroscopy has been used to examine simultaneously for the first time the relationship between chelation of intracellular iron and the rate of removal and regeneration of the tyrosyl radical of ribonucleotide reductase within intact human leukemia K562 cells. The different physiochemical characteristics of relatively hydrophobic low molecular weight bidentate hydroxypyridinone chelators and the higher molecular weight hexadentate ferrioxamine have been exploited to elucidate these interactions further. The base-line concentration of EPR-detectable mononuclear nonheme iron complexes was 3.15 ± 1.05 μM, rising on incubation with chelators more rapidly with hydroxypyridinones than with desferrioxamine. Hydroxypyridinones also removed the tyrosyl radical more rapidly, apparently as a consequence of depletion of the intracellular iron pools necessary to regenerate the active enzyme and compatible with their reportedly greater cell toxicity. The radical decay rate is consistent with previous models, suggesting that iron is spontaneously removed from mammalian ribonucleotide reductase. Upon removal of extracellular chelator the regeneration of the tyrosyl radical was significantly faster for hydroxypyridinones than for desferrioxamine, consistent with their differential effects on cell cycle synchronization.

Results of the first international round robin for the quantification of urinary and plasma hepcidin assays: need for standardization

This study indicates that hepcidin levels reported by various methods vary considerably but analytical variance is generally low and similar for all methods. See related perspective article on page 1631. The recently discovered iron regulatory peptide hormone hepcidin holds promise as a novel biomarker in iron metabolism disorders. To date, various mass spectrometry and immunochemical methods have been developed for its quantification in plasma and urine. Differences in methodology and analytical performance hinder the comparability of data. As a first step towards method harmonization, several hepcidin assays were compared. Worldwide eight laboratories participated in a urinary and plasma round robin in which hepcidin was analyzed. For both urine and plasma: (i) the absolute hepcidin concentrations differed widely between methods, (ii) the between-sample variation and the analytical variation of the methods are similar. Importantly, the analytical variation as percentage of the total variance is low for all methods, indicating their suitability to distinguish hepcidin levels of different samples. Spearman correlations between methods were generally high. The round robin results inform the scientific and medical community on the status and agreement of the current hepcidin methods. Ongoing initiatives should facilitate standardization by exchanging calibrators and representative samples.

Metals ions and neurodegeneration.

Neurodegenerative disorders include a variety of pathological conditions, which share similar critical metabolic processes such as protein aggregation and oxidative stress, both of which are associated with the involvement of metal ions. In this review Alzheimer’s disease and Parkinson’s disease are mainly discussed, with the aim of identifying common trends underlying these neurological conditions. Chelation therapy could be a valuable therapeutic approach, since metals are considered to be a pharmacological target for the rationale design of new therapeutic agents directed towards the treatment of neurodegeneration.

Ferric saccharate induces oxygen radical stress and endothelial dysfunction in vivo

Background  Intravenous iron supplementation is used widely in haemodialysis patients. However, nontransferrin‐bound iron (NTBI), which increases after intravenous supplementation of ferric saccharate, has been suggested to act as a catalytic agent in oxygen radical formation in vitro and may thus contribute to endothelial impairment in vivo.

A direct method for quantification of non-transferrin-bound iron

A direct method for quantification of non-transferrin-bound iron has been developed. This assay relies on the use of a large excess of a low affinity ligand (nitrilotriacetic acid, NTA) which removes and complexes all low molecular weight iron and iron nonspecifically bound to serum proteins. Iron bound to transferrin, ferritin, desferrioxamine, and its metabolites is unaffected. The Fe-NTA complex present in the serum ultrafiltrate is then quantified using an automated HPLC procedure where on-column derivatization with a high affinity iron chelator (3-hydroxy-1-propyl-2-methyl-pyridin-4-one) takes place. The iron complexes of desferrioxamine and its metabolites are unaffected by the above-derivatization procedure. With minor modifications, this method is equally applicable for the quantification of low molecular weight iron in other biological fluids.

Fenton chemistry and oxidative stress mediate the toxicity of the β-amyloid peptide in a Drosophila model of Alzheimer’s disease

The mechanism by which aggregates of the β‐amyloid peptide (Aβ) mediate their toxicity is uncertain. We show here that the expression of the 42‐amino‐acid isoform of Aβ (Aβ1–42) changes the expression of genes involved in oxidative stress in a Drosophila model of Alzheimer’s disease. A subsequent genetic screen confirmed the importance of oxidative stress and a molecular dissection of the steps in the cellular metabolism of reactive oxygen species revealed that the iron‐binding protein ferritin and the H2O2 scavenger catalase are the most potent suppressors of the toxicity of wild‐type and Arctic (E22G) Aβ1–42. Likewise, treatment with the iron‐binding compound clioquinol increased the lifespan of flies expressing Arctic Aβ1–42. The effect of iron appears to be mediated by oxidative stress as ferritin heavy chain co‐expression reduced carbonyl levels in Aβ1–42 flies by 65% and restored the survival and locomotion function to normal. This was achieved despite the presence of elevated levels of the Aβ1–42. Taken together, our data show that oxidative stress, probably mediated by the hydroxyl radical and generated by the Fenton reaction, is essential for Aβ1–42 toxicity in vivo and provide strong support for Alzheimer’s disease therapies based on metal chelation.