Copper availability contributes to iron perturbations in human nonalcoholic fatty liver disease

BACKGROUND ; AIMS: Iron perturbations are frequently observed in nonalcoholic fatty liver disease (NAFLD). We aimed to investigate a potential association of copper status with disturbances of iron homeostasis in NAFLD. METHODS: We retrospectively studied 140 NAFLD patients and 25 control subjects. Biochemical and hepatic iron and copper parameters were analyzed. Hepatic expression of iron regulatory molecules was investigated in liver biopsy specimens by reverse-transcription polymerase chain reaction and Western blot analysis. RESULTS: NAFLD patients had lower hepatic copper concentrations than control subjects (21.9 +/- 9.8 vs 29.6 +/- 5.1 microg/g; P = .002). NAFLD patients with low serum and liver copper concentrations presented with higher serum ferritin levels (606.7 +/- 265.8 vs 224.2 +/- 176.0 mg/L; P < .001), increased prevalence of siderosis in liver biopsy specimens (36/46 vs 10/47 patients; P < .001), and with elevated hepatic iron concentrations (1184.4 +/- 842.7 vs 319.9 +/- 451.3 microg/g; P = .020). Lower serum concentrations of the copper-dependent ferroxidase ceruloplasmin (21.7 +/- 4.1 vs 30.4 +/- 6.4 mg/dL; P < .001) and decreased liver ferroportin (FP-1; P = .009) messenger RNA expression were found in these patients compared with NAFLD patients with high liver or serum copper concentrations. Accordingly, in rats, a reduced dietary copper intake was paralleled by a decreased hepatic FP-1 protein expression. CONCLUSIONS: A significant proportion of NAFLD patients should be considered copper deficient. Our results indicate that copper status is linked to iron homeostasis in NAFLD, suggesting that low copper bioavailability causes increased hepatic iron stores via decreased FP-1 expression and ceruloplasmin ferroxidase activity thus blocking liver iron export in copper-deficient subjects.

Strategies of "Pseudomonas aeruginosa" to overcome copper limitation

Summary Copper is an important trace element and micronutrient for living organisms as it is the cofactor of several enzymes involved in diverse biological redox processes such as aerobic respiration, denitrification and photosynthesis. Despite its importance, copper may be poorly bioavailable in soils and aquatic environments, as well as in the human body, especially at physiological or alkaline pH. In this work, we have investigated the strategies that the versatile bacterium and opportunistic pathogen Pseudomonas aeruginosa has evolved to face and overcome copper limitation. The global response of the P. aeruginosa to copper limitation was assessed under aerobic conditions. Numerous iron uptake functions (including the siderophores pyoverdine and pyochelin) were down-regulated whereas expression of cioAB (encoding an alternative, copper-independent, cyanide-resistant ubiquinol oxidase) was up-regulated. Wild type P. aeruginosa was able to grow aerobically in a defined glucose medium depleted of copper by a copper chelator, whereas a cioAB mutant did not grow. Thus, P. aeruginosa relies on the CioAB enzyme to cope with severe copper deprivation. A quadruple cyo cco1 cco2 cox mutant, which was deleted for all known heme-copper terminal oxidases of P. aeruginosa, grew aerobically, albeit more slowly than did the wild type, indicating that the CioAB enzyme is capable of energy conservation. However, the expression of a cioA'-'lacZ fusion was less dependent on the copper status in the quadruple mutant than in the wild type, suggesting that copper availability might affect cioAB expression indirectly, via the function of the heme-copper oxidases. These results suggest that the CioAB enzyme can be used as a by-pass strategy to overcome severe copper limitation and perform aerobic respiration even if virtually no copper is available. The PA0114 gene, which encodes a protein of the SCOT/SenC family, was found to be important for copper acquisition and aerobic respiration in low copper conditions. A PA0114 (sent) mutant grew poorly in low copper media and had low terminal oxidase activity with TMPD (N,N,N',N'-tetramethyl-p-phenylenediamine), but expressed the CioAB enzyme at elevated levels. Addition of copper reversed these phenotypes, suggesting that periplasmic copper capture by the SenC protein is another strategy that helps P. aeruginosa to adapt to copper deprivation. RESUME Le cuivre est un micronutriment important pour les organismes vivants. Il représente le cofacteur de plusieurs enzymes impliquées dans une multitude de processus biologiques tels que la respiration aérobie, la dénitrification et la photosynthèse. Malgré son importance, le cuivre peut être peu disponible dans les sols, les environnements aquatiques et le corps humain, spécialement à pH physiologique ou alcalin. Dans ce travail nous avons étudié les stratégies développées par la bactérie pathogène opportuniste Pseudomonas aeruginosa PAO1 afm de faire face et de surmonter le manque de cuivre. La réponse globale de P. aeruginosa à la carence de cuivre a été analysée dans des conditions aérobie. Les résultats obtenus ont montré que plusieurs gènes impliqués dans l'acquisition du fer, tels que les gènes codant pour les sidérophores (pyoverdine et pyochéline), étaient réprimés, tandis que l'expression de l'opéron cioAB, codant pour l'oxydase terminale insensible au cyanure (CIO), était augmentée. La souche sauvage P. aeruginosa est capable de croître dans un milieu où la concentration en cuivre est limitée, due à la présence d'un chélateur spéciftque de cuivre, tandis que le mutant cioAB ne croît pas dans ces conditions. Nous avons conclu que P. aeruginosa nécessite l'oxydase terminale CIO pour faire face à la carence en cuivre. Un quadruple mutant affecté dans toutes les oxydases dépendantes du cuivre (cyo ccol cco2 cox) et appartenant aux oxydases de type hème-cuivre, peut croître en aérobie, néanmoins plus lentement que la souche sauvage, ce qui montre que l'enzyme CIO est capable de conserver l'énergie. L'expression de la fusion rapportrice cioA'-'IacZ chez le quadruple mutant est moins dépendante de la disponibilité de cuivre que chez la souche sauvage. Ces résultats suggèrent que la disponibilité de cuivre influence l'expression de cioAB d'une façon indirecte, par le biais des oxydases terminales de type héme-cuivre. Il est donc possible qu'en cas de carence de cuivre, P. aeruginosa utilise l'enzyme CIO comme stratégie afin de surmonter ce manque et de réaliser la respiration aérobie. Nous avons démontré que le gène PA0114, codant pour une protéine appartenant à la famille SCO1/SenC, est important dans l'acquisition et dans la respiration aérobie dans des environnements où le cuivre est présent en faible concentration. En ces conditions, la croissance du mutant senC est faible; de plus, l'activité des oxydases terminales en présence du donneur d'électrons TMPD (N,N,N,N'-tetraméthyl-p-phénylenediamine) est basse. Toutefois, l'addition de cuivre au milieu de culture permet de restaurer le phénotype du type sauvage. Ces résultats montrent que la protéine SenC est capable d'acquérir le cuivre et représente donc une autre stratégie chez P. aeruginosa pour s'adapter à un manque de cuivre.

Adaptation of aerobically growing Pseudomonas aeruginosa to copper starvation.

Restricted bioavailability of copper in certain environments can interfere with cellular respiration because copper is an essential cofactor of most terminal oxidases. The global response of the metabolically versatile bacterium and opportunistic pathogen Pseudomonas aeruginosa to copper limitation was assessed under aerobic conditions. Expression of cioAB (encoding an alternative, copper-independent, cyanide-resistant ubiquinol oxidase) was upregulated, whereas numerous iron uptake functions (including the siderophores pyoverdine and pyochelin) were expressed at reduced levels, presumably reflecting a lower demand for iron by respiratory enzymes. Wild-type P. aeruginosa was able to grow aerobically in a defined glucose medium depleted of copper, whereas a cioAB mutant did not grow. Thus, P. aeruginosa relies on the CioAB enzyme to cope with severe copper deprivation. A quadruple cyo cco1 cco2 cox mutant, which was deleted for all known heme-copper terminal oxidases of P. aeruginosa, grew aerobically, albeit more slowly than did the wild type, indicating that the CioAB enzyme is capable of energy conservation. However, the expression of a cioA'-'lacZ fusion was less dependent on the copper status in the quadruple mutant than in the wild type, suggesting that copper availability might affect cioAB expression indirectly, via the function of the heme-copper oxidases.

The influence of mineral solubility and soil solution concentration on the toxicity of copper to Eisenia fetida Savigny

The OECD 14 d earthworm acute toxicity test was used to determine the toxicity of copper added as copper nitrate (Cu(NO3)(2)), copper sulphate (CuSO4) and malachite (Cu-2(OH)(2)(CO3)) to Eisenia fetida Savigny. Cu(NO3)(2), and CuSO4 were applied in both an aqueous (aq) and solid (s) form, Cu-2(OH)(2)(CO3) was added as a solid. Soil solution was extracted by centrifugation, and analysed for copper. Two extractants [0.01 M CaCl2 and 0.005 M diethylenetriminpentaacetic acid (DTPA)] were used as a proxy of the bioavailable copper fraction in the soil. For bulk soil copper content the calculated copper toxicity decreased in the order nitrate > sulphide > carbonate, the same order as decreasing solubility of the metal compounds. For Cu(NO3)(2) and CuSO4, the LC50s obtained were not significantly different when the compound was added in solution or solid form. There was a significant correlation between the soil solution copper concentration and the percentage earthworm mortality for all 3 copper compounds (P less than or equal to 0.05) indicating that the soil pore water copper concentration is important for determining copper availability and toxicity to E. fetida. In soil avoidance tests the earthworms avoided the soils treated with Cu(NO3)(2) (aq and s) and CuSO4 (aq and s), at all concentrations used (110-8750 mug Cu g(-1), and 600-8750 mug Cu g(-1) respectively). In soils treated with Cu-2(OH2)CO3, avoidance behaviour was exhibited at all concentrations greater than or equal to3500 mug Cu g(-1). There was no significant correlation between the copper extracted by either CaCl2 or DTPA and percentage mortality. These two extractants are therefore not useful indicators of copper availability and toxicity to E. fetida.

A role for low hepatic copper concentrations in nonalcoholic Fatty liver disease

Copper has a role in antioxidant defense, lipid peroxidation, and mitochondrial function, and copper deficiency has been linked to atherogenic dyslipidemia. We aimed to investigate the potential role of copper availability in the pathogenesis of nonalcoholic fatty liver disease (NAFLD).

Distribution and availability of copper, iron, manganese and zinc in the archaeological black earth profile from the Amazon region

Os solos de terra preta arqueológica são ricos em matéria orgânica, contêm fragmentos cerâmicos e artefatos líticos e apresentam nutrientes em concentrações mais elevadas do que outros tipos de solos. Com o intuito de contribuir com informações sobre concentrações de micronutrientes disponíveis em solos de terra preta, foram avaliadas através de extrações químicas sequenciais, a distribuição e disponibilidade de Cu, Fe, Mn e Zn em um perfil de terra preta arqueológica no município de Juruti, estado do Pará. As maiores concentrações de Cu, Fe, Mn e Zn no perfil são encontradas na fração residual. Este estudo mostrou que ferro, manganês e zinco são preferencialmente disponíveis a partir das frações, associados aos óxidos de Fe-Mn, variando de 1265,39 a 1818,12 mg kg^-1, 0,83 a 48,51 mg kg^-1 e 1,92 a 12,05 mg kg^-1, respectivamente, e o cobre a partir da matéria orgânica, variando de 0,13 a 0,45 mg kg^-1.

Copper, zinc and manganese in soils of two watersheds in Santa Catarina with intensive use of pig slurry

Systematic pig slurry application to crop soils may lead to the accumulation of heavy metals in regions with intensive pig raising. The aim of this study was to evaluate the accumulation of Cu, Zn and Mn in soils under systematic pig slurry application. For this purpose, soil samples were collected from two of the most representative watersheds of Santa Catarina where the predominant activity is pig raising. In each watershed, 12 properties were chosen to evaluate the different systems of pig husbandry (complete cycle (CC), farrowing (FaU) and finishing units (FiU)). Based on information of the producers, soil samples were collected in areas with and without systematic manure application. To determine the total Cu, Zn and Mn content in soils and manure, a methodology proposed by the Environmental Protection Agency of the United States (USEPA), method nº 3050B, was used. For the available heavy metal content, Cu and Zn was extracted with HCl 0.1 mol L-1 and Mn with KCl 1 mol L-1. Data were subjected to multivariate analysis, using the canonical discriminant analysis to identify the metals that best differentiate the soils studied within each swine housing system. Successive pig slurry applications cause an increase in Cu, Zn and Mn availability in the soil and this indicates the need for monitoring of the metal concentrations over time. The critical values of Cu in the soil can be reached and exceeded more rapidly than Zn. The results showed that the soil type may be one of the attribute underlying the determination of public policies in pig raising and waste management because soils such as Inceptisols were shown to be more prone to possible contamination since they may more rapidly reach total critical Cu levels.

PHYTOAVAILABILITY OF COPPER IN INDUSTRIAL BY-PRODUCTS AND MINERAL FERTILIZERS

Alternative copper (Cu) sources could be used in fertilizer production, although the bioavailability of copper in these materials is unknown. The objective of this study was to evaluate the extractants neutral ammonium citrate (NAC), 2 % citric acid, 1 % acetic acid, 10 % HCl, 10 % H2SO4, buffer solution pH 6.0, DTPA, EDTA, water, and hot water in the quantification of available Cu content in several sources, relating them to the relative agronomic efficiency (RAE) of wheat grown in a clayey Latossolo Vermelho eutrófico (Oxisol) and Neossolo Quartzarênico (Typic Quartzipsamment). Copper was applied at the rate of 1.5 mg kg-1 as scrap slag, brass slag, Cu ore, granulated copper, and copper sulfate. The extractants 10 % HCl, 10 % H2SO4, and NAC extracted higher Cu concentrations. The RAE values of brass slag and Cu ore were similar to or higher than those of Cu sulfate and granulated Cu. Solubility in the 2nd NAC extractant, officially required for mineral fertilizers with Cu, was lower than 60 % for the scrap slag, Cu ore, and granulated copper sources. This fact indicates that adoption of the NAC extractant may be ineffective for industrial by-products, although no extractant was more efficient in predicting Cu availability for wheat fertilized with the Cu sources tested.

Development of hypogene and supergene alteration and copper mineralization patterns, Sar Cheshmeh porphyry copper deposits, Iran /

The formation of the Sar Cheshmeh porphyry Cu-Mo deposit is related to the culmination of calc-alkaline igneous activity in the Kerman region. The deposit comprises a suite of Late Cenozoic intrusive sub-volcanic and extrusive rocks emplaced into a folded series of Eocene andesitic lavas and pyroclastic sediments. The earliest stage of magmatism was emplacement of a large granodiorite stock about 29 m.y.b.p. This was followed by intrusion of two separate porphyritic bodies at 15 (Sar Cheshrneh porphyry) and 12 m.y.b.p. (Late porphyry) and a series of sub-volcanic dikes between 12 and 9 m.y.b.p. Magmatic activity terminated with multi-phase extrusion of a Pelean dacitic dome complex between 10 and 2.8 m.y.b.p. The country rocks and the earlier porphyritic intrusions are pervasively altered to biotite-rich potassium silicate (metasomatic and hydrothermal) sericite-clay, phyllic and chlorite-clay, argillic assemblages. These grade outwards to an extensive propylitic zone. Within the ore body, the later intra-. and post-mineral dikes only reach the propylitic grade. At least three different sets of quartz veins are present, including a sericite-chlorite-quartz set which locally retrogrades pervasive secondary biotite to sericite. In the hypogene zone, metasomatic and hydrothermal alteration is related to all stages of magmatism but copper mineralization and veining are restricted to a period of 15 to 9 m.y.b.p.related to the early intrusive phases. The copper mineralization and silicate alteration do not fit a simple annular ring model but have been greatly modified by, 1. The existence of an ititial, outer ring, of metasomatic alteration overprinted by an inner.ring of hydrothermal alteration and, 2. later extensive dilating effects of intra- and post-mineral dikes. The hydrothermal clay mineral assemblage in the hypogene zone is illite-chlorite-kaolinite-smectite (beidellite). Preliminary studies indicate that the amount of each of these clays varies vertically and that hydrothermal zonation of clay minerals is possible. However, these minerals alter to illite-kaolinite assemblages in the supergene sulfide zone and to more kaolinite-rich assemblages in the supergene leached zone. Hydrothermal biotite breaks down readily in the supergene zone and is not well preserved in surface outcrops. The distribution of copper minerals in the supergene sulfide enrichment zone is only partly related to rock type being more dependent on topography and the availability of fractures.

"Effects of some heavy metals copper. zinc and lead on certain tissues of £153 Qarsia (Hamilton and Buchanan) in different environments

The research investigations on pollution, particularly in coastal/ estuarine environments are recent ones and started only in 1970s. Hence the informations available are fragmentary and scattered. They throw some light only on either the concentration of heavy metals in water or in sediment or in organisms. No concerted efforts have been made to consolidate and correlate the results between the environment and biota. Literature on the level of concentration of heavy metals in different tissues of organisms with regard to their availability in the living media, their ratio, their inter—relationship, tolerance limit of organisms, etc. are very few or rather nil. in view of the importance enumerated above, the candidate has selected the topic "Effects of some heavy metals copper, zinc and lead on certain tissues of E E (Hamilton and Buchanan) in different environments" for detailed studies and to understand systematically (i) the source of effluents and wastes, (ii) the concentration of heavy metals copper, zinc and lead in water, in sediments and in tissues of the test animal, (iii) their effects, (iv) capacity of tolerance and accumulation in different tissues of the animal, and (V) the "Bioaccumulation Factor", etc.

EVALUATION of HEAVY METAL AVAILABILITY IN CONTAMINATED SEDIMENTS FROM THE ILHA SOLTEIRA HYDROELECTRIC DAM on THE PARANA RIVER AT ILHA SOLTEIRA, SP, BRAZIL

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Beneficial effects of dietary copper supplementation on serum lipids and antioxidant defenses in rats

Background: A nutrition experiment was utilized to investigate the effects of two levels of dietary copper (Cu) supplementation on lipid profile and antioxidant defenses in serum of rats. Methods: Male Wistar rats (180-200 g; n = 10) were divided into three groups: control group (A), fed a basal diet with 6 mu g Cu/g, and rats fed a basal diet with Cu (CuSO4) supplementation from aqueous solutions, for 4 weeks at the final concentrations of 2 mg Cu/rat (B) and 3 mg Cu/rat (C). Results: No significant changes were observed in final body weight, body weight gain, food consumption, total serum protein and high-density lipoprotein. Cu supplementation reduced the triacylglycerol (TG), total cholesterol and low-density lipoprotein (LDL-C). The LDL-C/TG ratio and total antioxidant substances (TAS) were higher in (B) and (C) groups than in (A) group. There was a positive correlation between Cu supplementation and ceruloplasmin levels. The markers of oxidative stress, lipid hydroperoxide and lipoperoxide were decreased with Cu supplementation. No alterations were observed in superoxide dismutase, indicating saturation of Cu enzyme site. The glutathione peroxidase activities (GSH-Px) were increased in both Cu-supplemented groups. Considering that a copper-selenium interaction can affect mineral availability of both elements, the effects of Cu on TAS and GSH-Px activities were associated with increased selenium disposal. Conclusions: Dietary Cu supplementation had beneficial effects on lipid profile by improving endogenous antioxidant defenses and decreasing the oxidative stress in vivo. Copyright (C) 2005 S. Karger AG, Basel.

The homeostasis of iron, copper, and zinc in paracoccidioides brasiliensis, cryptococcus neoformans var. Grubii, and cryptococcus gattii: a comparative analysis

Iron, copper, and zinc are essential for all living organisms. Moreover, the homeostasis of these metals is vital to microorganisms during pathogenic interactions with a host. Most pathogens have developed specific mechanisms for the uptake of micronutrients from their hosts in order to counteract the low availability of essential ions in infected tissues. We report here an analysis of genes potentially involved in iron, copper, and zinc uptake and homeostasis in the fungal pathogens Paracoccidioides brasiliensis, Cryptococcus neoformans var. grubii, and Cryptococcus gattii. Although prior studies have identified certain aspects of metal regulation in Cryptococcus species, little is known regarding the regulation of these elements in P. brasiliensis. We also present amino acid sequences analyses of deduced proteins in order to examine possible conserved domains. The genomic data reveals, for the first time, genes associated to iron, copper, and zinc assimilation and homeostasis in P. brasiliensis. Furthermore, analyses of the three fungal species identified homologs to genes associated with high-affinity uptake systems, vacuolar and mitochondrial iron storage, copper uptake and reduction, and zinc assimilation. However, homologs to genes involved in siderophore production were only found in P. brasiliensis. Interestingly, in silico analysis of the genomes of P. brasiliensis Pb01, Pb03, and Pb18 revealed significant differences in the presence and/or number of genes involved in metal homeostasis, such as in genes related to iron reduction and oxidation. The broad analyses of the genomes of P. brasiliensis, C. neoformans var. grubii, and C. gattii for genes involved in metal homeostasis provide important groundwork for numerous interesting future areas of investigation that are required in order to validate and explore the function of the identified genes and gene pathways.

Cylindrospermopsis raciborskii exudate-Cu complexes: impact on copper dynamics and bioavailability in an aquatic food chain

The increasing contamination of aquatic environments motivates studies on the interactions among natural dissolved organic matter, metals, and the biota. This investigation focused on the organic exudates of the toxic cyanobacteria Cylindrospermopsis raciborskii as a Cu carrier through a three-level aquatic trophic chain (bacteria, protozoa, and copepod). The effects of bacteria activity and growth on the metal-organic complexes were evaluated through changes in free Cu2+ ions, total dissolved, and total particulate Cu. To be sure that the added copper would be complexed to the exudates, its complexing properties were previously determined. The cyanobacteria exudate-Cu complexes were furnished to bacteria that were further used as a food source to the protozoan Paramercium caudatum. This was then furnished as food to the copepod Mesocyclops sp. The results showed that, in general, the cyanobacterial exudates decreased Cu bioavailability and toxicity to the first trophic level (bacteria), but because the heterotrophic bacteria accumulated Cu, they were responsible for the transference for the otherwise low availability metal form. Both the bacteria and protozoan organisms accumulated Cu, but no metal accumulation was detected in the copepods.

Computer aided process planning (CAPP) for flat rolling of copper alloys

The manufacture of copper alloy flat rolled metals involves hot and cold rolling operations, together with annealing and other secondary processes, to transform castings (mainly slabs and cakes) into such shapes as strip, plate, sheet, etc. Production is mainly to customer orders in a wide range of specifications for dimensions and properties. However, order quantities are often small and so process planning plays an important role in this industry. Much research work has been done in the past in relation to the technology of flat rolling and the details of the operations, however, there is little or no evidence of any research in the planning of processes for this type of manufacture. Practical observation in a number of rolling mills has established the type of manual process planning traditionally used in this industry. This manual approach, however, has inherent drawbacks, being particularly dependent on the individual planners who gain their knowledge over a long span of practical experience. The introduction of the retrieval CAPP approach to this industry was a first step to reduce these problems. But this could not provide a long-term answer because of the need for an experienced planner to supervise generation of any plan. It also fails to take account of the dynamic nature of the parameters involved in the planning, such as the availability of resources, operation conditions and variations in the costs. The other alternative is the use of a generative approach to planning in the rolling mill context. In this thesis, generative methods are developed for the selection of optimal routes for single orders and then for batches of orders, bearing in mind equipment restrictions, production costs and material yield. The batch order process planning involves the use of a special cluster analysis algorithm for optimal grouping of the orders. This research concentrates on cold-rolling operations. A prototype model of the proposed CAPP system, including both single order and batch order planning options, has been developed and tested on real order data in the industry. The results were satisfactory and compared very favourably with the existing manual and retrieval methods.