Selenium-Enriched E. coli Bacteria Mitigate the Age-Associated Degeneration of Cholinergic Neurons in C. elegans.

Selenium (Se) is an essential trace element for humans and animals, but high-dose supplementation with Se compounds, most notably selenite, may exert cytotoxic and other adverse effects. On the other hand, bacteria, including Escherichia coli (E. coli), are capable of reducing selenite to red elemental Se that may serve as a safer Se source. Here, we examined how a diet of Se-enriched E. coli bacteria affected vital parameters and age-associated neurodegeneration in the model organism Caenorhabditis elegans (C. elegans). The growth of E. coli OP50 for 48 h in medium supplemented with 1 mM sodium selenite resulted in reddening of the bacterial culture, accompanied by Se accumulation in the bacteria. Compared to nematodes supplied with the standard E. coli OP50 diet, the worms fed on Se-enriched bacteria were smaller and slimmer, even though their food intake was not diminished. Nevertheless, given the choice, the nematodes preferred the standard diet. The fecundity of the worms was not affected by the Se-enriched bacteria, even though the production of progeny was somewhat delayed. The levels of the Se-binding protein SEMO-1, which serves as a Se buffer in C. elegans, were elevated in the group fed on Se-enriched bacteria. The occurrence of knots and ruptures within the axons of cholinergic neurons was lowered in aged nematodes provided with Se-enriched bacteria. In conclusion, C. elegans fed on Se-enriched E. coli showed less age-associated neurodegeneration, as compared to nematodes supplied with the standard diet.

Long-term suboptimal dietary trace element supply does not affect trace element homeostasis in murine cerebellum.

The ageing process is associated with alterations of systemic trace element (TE) homeostasis increasing the risk, e.g. neurodegenerative diseases. Here, the impact of long-term modulation of dietary intake of copper, iron, selenium, and zinc was investigated in murine cerebellum. Four- and 40-wk-old mice of both sexes were supplied with different amounts of those TEs for 26 wk. In an adequate supply group, TE concentrations were in accordance with recommendations for laboratory mice while suboptimally supplied animals received only limited amounts of copper, iron, selenium, and zinc. An additional age-adjusted group was fed selenium and zinc in amounts exceeding recommendations. Cerebellar TE concentrations were measured by inductively coupled plasma-tandem mass spectrometry. Furthermore, the expression of genes involved in TE transport, DNA damage response, and DNA repair as well as selected markers of genomic stability [8-oxoguanine, incision efficiency toward 8-oxoguanine, 5-hydroxyuracil, and apurinic/apyrimidinic sites and global DNA (hydroxy)methylation] were analysed. Ageing resulted in a mild increase of iron and copper concentrations in the cerebellum, which was most pronounced in the suboptimally supplied groups. Thus, TE changes in the cerebellum were predominantly driven by age and less by nutritional intervention. Interestingly, deviation from adequate TE supply resulted in higher manganese concentrations of female mice even though the manganese supply itself was not modulated. Parameters of genomic stability were neither affected by age, sex, nor diet. Overall, this study revealed that suboptimal dietary TE supply does not substantially affect TE homeostasis in the murine cerebellum.

Selenium-binding protein 1 (SELENBP1) is a copper-dependent thiol oxidase.

Selenium-binding protein 1 (SELENBP1) was reported to act as a methanethiol oxidase (MTO) in humans, catalyzing the conversion of methanethiol to hydrogen peroxide, hydrogen sulfide and formaldehyde. Here, we identify copper ions as essential to this novel MTO activity. Site-directed mutagenesis of putative copper-binding sites in human SELENBP1 produced as recombinant protein in E. coli resulted in loss of its enzymatic function. On the other hand, the eponymous binding of selenium (as selenite) was no requirement for MTO activity and only moderately increased SELENBP1-catalyzed oxidation of methanethiol. Furthermore, SEMO-1, the SELENBP1 ortholog recently identified in the nematode C. elegans, also requires copper ions, and MTO activity was enhanced or abrogated, respectively, if worms were grown in the presence of cupric chloride or of a Cu chelator. In addition to methanethiol, we identified novel substrates of SELENBP1 from the group of volatile sulfur compounds, ranging from ethanethiol to 1-pentanethiol as well as 2-propene-1-thiol. Gut microbiome-derived methanethiol as well as food-derived volatile sulfur compounds (VSCs) account for malodors that may contribute to extraoral halitosis in humans, if not metabolized properly. As SELENBP1 is particularly abundant in tissues exposed to VSCs, such as colon, liver, and lung, it appears to contribute to copper-dependent VSC degradation.

Measurement of trace elements in murine liver tissue samples: Comparison between ICP-MS/MS and TXRF.

BACKGROUND: Trace elements exhibit essential functions in many physiological processes. Thus, for research focusing on trace element homeostasis and metabolism analytical methods allowing for multi-element analyses are fundamental. Small sample amounts may be a big challenge in trace element analyses especially if also other end points want to be addressed in the same sample. Therefore, the aim of the present study was to examine trace elements (iron, copper, zinc, and selenium) in murine liver tissue prepared by a RIPA buffer-based lyses method. METHODS AND RESULTS: After centrifugation, lysates and pellets were obtained and trace elements were analyzed with TXRF in liver lysates. The results were compared to that obtained by a standard microwave-assisted acidic digestion with subsequent ICP-MS/MS analysis of the same liver tissue, liver lysates, and remaining pellets. In addition, trace element concentrations, determined in murine serum with both methods, were compared. For serum samples, both TXRF and ICP-MS/MS provide similar and highly correlating results. Furthermore, in liver lysate samples prepared with RIPA buffer, comparable trace element concentrations were measured by TXRF as with the standard digestion technique and ICP-MS/MS. Only marginal amounts of trace elements were detected in the pellets. CONCLUSION: Taken together, the results obtained by the present study indicate that the RIPA buffer-based method is suitable for sample preparation for trace element analyses via TXRF, at least for the here investigated murine liver samples.

Selenium homeostasis in human brain cells: Effects of copper (II) and Se species.

BACKGROUND: Both essential trace elements selenium (Se) and copper (Cu) play an important role in maintaining brain function. Homeostasis of Cu, which is tightly regulated under physiological conditions, seems to be disturbed in Alzheimer´s (AD) and Parkinson´s disease (PD) patients. Excess Cu promotes the formation of oxidative stress, which is thought to be a major cause for development and progression of neurological diseases (NDs). Most selenoproteins exhibit antioxidative properties and may counteract oxidative stress. However, expression of selenoproteins is altered under conditions of Se deficiency. Serum Se levels are decreased in AD and PD patients suggesting Se as an important factor in the development and progression of NDs. The aim of this study was to elucidate the interactions between Cu and Se in human brain cells particularly with respect to Se homeostasis. METHODS: Firstly, modulation of Se status by selenite or SeMet were assessed in human astrocytes and human differentiated neurons. Therefore, cellular total Se content, intra- and extracellular selenoprotein P (SELENOP) content, and glutathione peroxidase (GPX) activity were quantified. Secondly, to investigate the impact of Cu on these markers, cells were exposed to copper(II)sulphate (CuSO4) for 48 h. In addition, putative protective effects of Se on Cu-induced toxicity, as measured by cell viability, DNA damage, and neurodegeneration were investigated. RESULTS: Modulation of cellular Se status was strongly dependent on Se species. In detail, SeMet increased total cellular Se and SELENOP content, whereas selenite led to increased GPX activity and SELENOP excretion. Cu treatment resulted in 133-fold higher cellular Cu concentration with a concomitant decrease in Se content. Additionally, SELENOP excretion was suppressed in both cell lines, while GPX activity was diminished only in astrocytes. These effects of Cu could be partially prevented by the addition of Se depending on the cell line and Se species used. While Cu-induced oxidative DNA damage could not be prevented by addition of Se regardless of chemical species, SeMet protected against neurite network degeneration triggered by Cu. CONCLUSION: Cu appears to negatively affect Se status in astrocytes and neurons. Especially with regard to an altered homeostasis of those trace elements during aging, this interaction is of high physiological relevance. Increasing Cu concentrations associated with decreased selenoprotein expression or functionality might be a promoting factor for the development of NDs.

The Nutritional Supply of Iodine and Selenium Affects Thyroid Hormone Axis Related Endpoints in Mice.

Selenium and iodine are the two central trace elements for the homeostasis of thyroid hormones but additional trace elements such as iron, zinc, and copper are also involved. To compare the primary effects of inadequate intake of selenium and iodine on the thyroid gland, as well as the target organs of thyroid hormones such as liver and kidney, mice were subjected to an eight-week dietary intervention with low versus adequate selenium and iodine supply. Analysis of trace element levels in serum, liver, and kidney demonstrated a successful intervention. Markers of the selenium status were unaffected by the iodine supply. The thyroid gland was able to maintain serum thyroxine levels even under selenium-deficient conditions, despite reduced selenoprotein expression in liver and kidney, including deiodinase type 1. Thyroid hormone target genes responded to the altered selenium and iodine supply, whereas the iron, zinc, and copper homeostasis remained unaffected. There was a notable interaction between thyroid hormones and copper, which requires further clarification. Overall, the effects of an altered selenium and iodine supply were pronounced in thyroid hormone target tissues, but not in the thyroid gland.

Copper interferes with selenoprotein synthesis and activity.

Selenium and copper are essential trace elements for humans, needed for the biosynthesis of enzymes contributing to redox homeostasis and redox-dependent signaling pathways. Selenium is incorporated as selenocysteine into the active site of redox-relevant selenoproteins including glutathione peroxidases (GPX) and thioredoxin reductases (TXNRD). Copper-dependent enzymes mediate electron transfer and other redox reactions. As selenoprotein expression can be modulated e.g. by H2O2, we tested the hypothesis that copper status affects selenoprotein expression. To this end, hepatocarcinoma HepG2 cells and mice were exposed to a variable copper and selenium supply in a physiologically relevant concentration range, and transcript and protein expression as well as GPX and TXNRD activities were compared. Copper suppressed selenoprotein mRNA levels of GPX1 and SELENOW, downregulated GPX and TXNRD activities and decreased UGA recoding efficiency in reporter cells. The interfering effects were successfully suppressed by applying the copper chelators bathocuproinedisulfonic acid or tetrathiomolybdate. In mice, a decreased copper supply moderately decreased the copper status and negatively affected hepatic TXNRD activity. We conclude that there is a hitherto unknown interrelationship between copper and selenium status, and that copper negatively affects selenoprotein expression and activity most probably via limiting UGA recoding. This interference may be of physiological relevance during aging, where a particular shift in the selenium to copper ratio has been reported. An increased concentration of copper in face of a downregulated selenoprotein expression may synergize and negatively affect the cellular redox homeostasis contributing to disease processes.

A quick and simple method for the determination of six trace elements in mammalian serum samples using ICP-MS/MS.

In order to assess the individual trace element status of humans for either medical or scientific purposes, amongst others, blood serum levels are determined. Furthermore, animal models are used to study interactions of trace elements. Most published methods require larger amounts (500-1000 µL) of serum to achieve a reliable determination of multiple trace elements. However, oftentimes, these amounts of serum cannot be dedicated to a single analysis and the amount available for TE-determination is much lower. Therefore, a published ICP-MS/MS method for trace element determination in serum was miniaturized, optimized and validated for the measurement of Mn, Fe, Cu Zn, I and Se in as little as 50 µL of human and murine serum and is presented in this work. For validation, recoveries of multiple LOTs and levels from commercially available human reference serum samples were determined, intra- and inter-day variations were assessed and limits of detection and quantification determined. It is shown, that the method is capable of giving accurate and reproducible results for all six elements within the relevant concentration ranges for samples from humans living in central Europe as well as from laboratory mice. As a highlight, the achieved limits of detection and quantification for Mn were found to be at 0.02 µg/L serum and 0.05 µg/L serum, respectively, while using an alkaline diluent for the parallel determination of iodine.

Importance of propionate for the repression of hepatic lipogenesis and improvement of insulin sensitivity in high-fat diet-induced obesity.

SCOPE: The SCFA acetate (Ac) and propionate (Pr) are major fermentation products of dietary fibers and provide additional energy to the host. We investigated short- and long-term effects of dietary Ac and Pr supplementation on diet-induced obesity and hepatic lipid metabolism. METHODS AND RESULTS: C3H/HeOuJ mice received high-fat (HF) diets supplemented with 5% SCFA in different Ac:Pr ratios, a high acetate (HF-HAc; 2.5:1 Ac:Pr) or high Pr ratio (HF-HPr; 1:2.5 Ac:Pr) for 6 or 22 weeks. Control diets (low-fat (LF), HF) contained no SCFA. SCFA did not affect body composition but reduced hepatic gene and protein expression of lipogenic enzymes leading to a reduced hepatic triglyceride concentration after 22 weeks in HF-HPr mice. Analysis of long-chain fatty acid composition (liver and plasma phospholipids) showed that supplementation of both ratios led to a lower ω6:ω3 ratio. Pr directly led to increased odd-chain fatty acid (C15:0, C17:0) formation as confirmed in vitro using HepG2 cells. Remarkably, plasma C15:0 was correlated with the attenuation of HF diet-induced insulin resistance. CONCLUSION: Dependent on the Ac:Pr ratio, especially odd-chain fatty acid formation and insulin sensitivity are differentially affected, indicating the importance of Pr.

Differential acute effects of selenomethionine and sodium selenite on the severity of colitis.

The European population is only suboptimally supplied with the essential trace element selenium. Such a selenium status is supposed to worsen colitis while colitis-suppressive effects were observed with adequate or supplemented amounts of both organic selenomethionine (SeMet) and inorganic sodium selenite. In order to better understand the effect of these selenocompounds on colitis development we examined colonic phenotypes of mice fed supplemented diets before the onset of colitis or during the acute phase. Colitis was induced by treating mice with 1% dextran sulfate sodium (DSS) for seven days. The selenium-enriched diets were either provided directly after weaning (long-term) or were given to mice with a suboptimal selenium status after DSS withdrawal (short-term). While long-term selenium supplementation had no effect on colitis development, short-term selenite supplementation, however, resulted in a more severe colitis. Colonic selenoprotein expression was maximized in all selenium-supplemented groups independent of the selenocompound or intervention time. This indicates that the short-term selenite effect appears to be independent from colonic selenoprotein expression. In conclusion, a selenite supplementation during acute colitis has no health benefits but may even aggravate the course of disease.

Selenium in the redox regulation of the Nrf2 and the Wnt pathway.

Selenium deficiency is known to increase cancer risk by so far unclear mechanisms. Selenium exerts its biological effects via selenocysteine as an integral part of selenoproteins. Certain selenoproteins have redox properties, thereby providing a tool to regulate hydroperoxide-mediated signaling. Selenium deficiency does not only reduce synthesis of selenoproteins but also affects the expression of other proteins and even pathways. A moderate Se deficiency activates the Nrf2 and the Wnt pathways. The link between both pathways appears to be GSK3ß which in the active state prepares Nrf2 as well as ß-catenin, the key player in Wnt signaling, for ubiquitination and proteasomal degradation, thus silencing their transcriptional activity. Upon stimulation by Wnt signals, GSK3ß becomes inactivated and transcription factors are stabilized. Many intermediate steps in both pathways can be modulated by hydroperoxides, making them predestined to be regulated by selenoproteins. Oxidation sensors are (i) Keap1 which keeps Nrf2 in the cytosol unless it is modified by hydroperoxides/electrophiles and (ii) nucleoredoxin (Nrx) which is associated with disheveled (Dvl). NOX1-derived H2O2 oxidizes Nrx leading to the liberation of Dvl and the activation of Wnt signaling. Selenium deficiency can support oxidation of both sensors and activate both pathways. The consequences are dual: while the Keap1/Nrf2 system is generally believed to protect against oxidative stress, diverse xenobiotics, inflammation, and carcinogenesis, the Wnt response is considered rather a risky one in these respects. However, not only healthy cells but also malignant ones benefit from intact Keap1/Nrf2 signaling, making a dysregulated hydroperoxide signaling a plausible explanation for the increased cancer risk in selenium deficiency.

Selenoprotein W as biomarker for the efficacy of selenium compounds to act as source for selenoprotein biosynthesis.

Selenium is an essential trace element and, like all elements, present in many different compounds with unequivocal functions. This fact is only sporadically mentioned when recommended intake or supplementation is indicated just as "selenium." In mammals, selenium is an integral part of selenoproteins as selenocysteine. Selenocysteine is formed from serine at the respective tRNA((ser)sec), a reaction that requires selenophosphate formed from selenide and ATP. Thus, only compounds that can be metabolized into selenide can serve as sources for selenoprotein biosynthesis. We therefore tested the ability of selenium compounds such as sodium selenite, methylseleninic acid (MeSeA), Se-methyl selenocysteine, and selenomethionine to increase the activity, protein, or mRNA levels of commonly used biomarkers of the selenium status, glutathione peroxidase-1 (GPx1) and thioredoxin reductase, and of putatively new biomarkers, selenoprotein W1 (SepW1), selenoprotein H, and selenoprotein 15 in three different cell lines. Selenite and MeSeA were most efficient in increasing all markers tested, whereas the other compounds had only marginal effects. Effects were higher in the noncancerous young adult mouse colon cells than in the cancer cell lines HepG2 and HT-29. At the protein level, SepW1 responded as well as GPx1 and at the mRNA level, even better. Thus, the outcome of selenium treatment strongly depends on the chemical form, the cell type, and the biomarker used for testing efficacy.

Physiological functions of GPx2 and its role in inflammation-triggered carcinogenesis.

Mammalian glutathione peroxidases (GPxs) are reviewed with emphasis on the role of the gastrointestinal GPx2 in tumorigenesis. GPx2 ranks high in the hierarchy of selenoproteins, corroborating its importance. Colocalization of GPx2 with the Wnt pathway in crypt bases of the intestine and its induction by Wnt signals point to a role in mucosal homeostasis, but GPx2 might also support tumor growth when increased by a dysregulated Wnt pathway. In contrast, the induction of GPx2 by Nrf2 activators and the upregulation of COX2 in cells with a GPx2 knockdown reveal inhibition of inflammation and suggest prevention of inflammation-mediated carcinogenesis. The Janus-faced role of GPx2 has been confirmed in a mouse model of inflammation-associated colon carcinogenesis (AOM/DSS), where GPx2 deletion increased inflammation and consequently tumor development, but decreased tumor size. The model further revealed a GPx2-independent decrease in tumor development by selenium (Se) and detrimental effects of the Nrf2-activator sulforaphane in moderate Se deficiency.

The yin and yang of nrf2-regulated selenoproteins in carcinogenesis.

The NF-E2-related factor-2 (Nrf2) is a transcription factor which regulates the major cellular defense systems and thereby contributes to the prevention of many diseases including cancer. Selenium deficiency is associated with a higher cancer risk making also this essential trace element a promising candidate for cancer prevention. Two selenoproteins, thioredoxin reductase-1 (TrxR1) and glutathione peroxidase-2 (GPx2), are targets for Nrf2. Selenium deficiency activates Nrf2 as does a TrxR1 knockout making a synergism between both systems plausible. Although this might hold true for healthy cells, the interplay may turn into the opposite in cancer cells. The induction of the detoxifying and antioxidant enzymes by Nrf2 will make cancer cells chemoresistant and will protect them against oxidative damage. The essential role of TrxR1 in maintaining proliferation makes its upregulation in cancer cells detrimental. The anti-inflammatory potential of GPx2 will help to inhibit cancer initiation and inflammation-triggered promotion, but its growth supporting potential will also support tumor growth. This paper considers beneficial and adverse consequences of the activation of Nrf2 and the selenoproteins which appear to depend on the cancer stage.