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Hundreds of millions of people worldwide are exposed to unacceptable levels of carcinogenic inorganic arsenic. Animal models have shown that selenium and arsenic are mutually protective through the formation and elimination of the seleno-bis(S-glutathionyl) arsinium ion [(GS) 2 AsSe] - . Consistent with this, human selenium deficiency in arsenic-endemic regions is associated with arsenic-induced disease, leading to the initiation of human selenium supplementation trials. In contrast to the protective effect observed in vivo, in vitro studies have suggested that selenite increases arsenite cellular retention and toxicity. This difference might be explained by the rapid conversion of selenite to selenide in vivo. In the current study, selenite did not protect the human hepatoma (HepG2) cell line against the toxicity of arsenite at equimolar concentrations, however selenide increased the IC 50 by 2.3-fold. Cytotoxicity assays of arsenite + selenite and arsenite + selenide at different molar ratios revealed higher overall mutual antagonism of arsenite + selenide toxicity than arsenite + selenite. Despite this protective effect, in comparison to 75 Se-selenite, HepG2 cells in suspension were at least 3-fold more efficient at accumulating selenium from reduced 75 Se-selenide, and its accumulation was further increased by arsenite. X-ray fluorescence imaging of HepG2 cells also showed that arsenic accumulation, in the presence of selenide, was higher than in the presence of selenite. These results are consistent with a greater intracellular availability of selenide relative to selenite for protection against arsenite, and the formation and retention of a less toxic product, possibly [(GS) 2 AsSe] - ., (Copyright © 2020 Elsevier B.V. All rights reserved.)

(79)Se is among the most important long lived radionuclides in spent nuclear fuel and selenite, SeO3(2-), is its typical form in intermediate redox potential. The sorption behaviour of selenite and the bacterial impact on the selenite sorption in a 7-m-deep profile of a nutrient-poor boreal bog was studied using batch sorption experiments. The batch distribution coefficient (Kd) values of selenite decreased as a function of sampling depth and highest Kd values, 6600 L/kg dry weight (DW), were observed in the surface moss and the lowest in the bottom clay at 1700 L/kg DW. The overall maximum sorption was observed at pH between 3 and 4 and the Kd values were significantly higher in unsterilized compared to sterilized samples. The removal of selenite from solution by Pseudomonas sp., Burkholderia sp., Rhodococcus sp. and Paenibacillus sp. strains isolated from the bog was affected by incubation temperature and time. In addition, the incubation of sterilized surface moss, subsurface peat and gyttja samples with added bacteria effectively removed selenite from the solution and on average 65% of selenite was removed when Pseudomonas sp. or Burkholderia sp. strains were used. Our results demonstrate the important role of bacteria for the removal of selenite from the solution phase in the bog environment, having a high organic matter content and a low pH., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

This study (i) quantified the kinetics of selenate sorption and (ii) measured the influence of biotic processes in soil selenate stabilisation. Stirred flow-through reactor experiments were conducted on samples of a silty clay soil (pH = 8, Eh = 240-300 mV) from Bure (France) in both non-sterile and sterile conditions. Parameters of the proposed two-site sorption model (EK), adapted from van Genuchten and Wagenet (1989), were estimated by nonlinear regression. Fast selenate sorption on type-1 sites was moderate, with an equilibrium constant of 25.5 and 39.1 L/kg for non-sterile and sterile conditions. Rate-limited sorption on type-2 sites increased with time, and was predominant for longer periods of time in non-sterile conditions. At equilibrium, it would represent over 96% of the sorbed inventory, with mean sorption times of 17 h and 191 h for non-sterile and sterile conditions. Our results showed for Bure soil that (i) selenate sorption in flowing and mildly-oxidising conditions was strongly kinetically controlled, especially in non-sterile conditions, (ii) selenate desorption was much slower than sorption, which suggests its pseudo-irreversible stabilisation, and (iii) microbial activity increased the contribution of rate-limited sorption on type-2 sites, for which it increased sorption rate by a factor 7 but also facilitated its reversibility. This work stresses the limits of the Kd approach to represent selenate sorption in flowing conditions and supports an alternative formulation like the EK model, but also points out that biotic conditions are significant sources of variability for sorption parameters., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

In a context of environmental risk assessment of nuclear (79)Se radionuclide, the impact of low Se-selenite concentrations (0.008 and 8 mg kg(-1)) on bacterial communities of two soils, a silty clay loam and a sandy soil, was investigated over a 6-month incubation time. This Se-selenite was partially labelled with (75)Se. The state of the Se-impacted bacterial communities was analyzed through total bacterial counts, DNA fingerprints (ARISA profiles) and metabolic profiling (carbon substrate utilization patterns). Furthermore, the genetic diversity of bacterial populations involved in Se volatilization was evaluated by tpm (thiopurine methyltransferase gene) profiling. Emissions of (75)Se and CaCl(2)-extractable (75)Se were measured by γ-spectrometry and scintillation analysis. Se-selenite inputs changed transiently the substrate utilization patterns of bacterial communities but did not affect the other indicators. Se volatilization was at its highest level just after adding Se-selenite and for about 1 week. This volatilization was proportional to the added Se-selenite concentrations. It was 100-fold higher in silty clay loam, even though Se bioavailability was reduced in this soil. The soils were amended with crushed grass 3 months after the addition of Se-selenite. This organic amendment affected the organization of bacterial communities and increased the Se-volatilizing activities of both soils. Original soil organic carbon and bacterial diversity and activities seemed responsible for the different levels of Se emissions observed in soils. tpm lineages, encoding Se methyltransferases, were detected in both soils, confirming the broad distribution of tpm-harbouring bacteria and their probable role in the emissions of volatile Se. Five distinct groups of tpm were recorded per soil, with tpmI lineage being detected throughout the incubation period. This study demonstrates the ability of bacterial communities at volatilizing Se concentrations inferior to geochemical backgrounds and suggests that a probable transfer of nuclear Se will occur through volatilization after an environmental spill.

Selenite (SeO(3)(2-)) assimilation into a bacterial selenoprotein depends on thioredoxin (trx) reductase in Esherichia coli, but the molecular mechanism has not been elucidated. The mineral-oil overlay method made it possible to carry out anaerobic enzyme assay, which demonstrated an initial lag-phase followed by time-dependent steady NADPH consumption with a positive cooperativity toward selenite and trx. SDS-PAGE/autoradiography using (75)Se-labeled selenite as substrate revealed the formation of trx-bound selenium in the reaction mixture. The protein-bound selenium has metabolic significance in being stabilized in the divalent state, and it also produced the selenopersulfide (-S-SeH) form by the catalysis of E. coli trx reductase (TrxB).

The application of radionuclides for the localization of essential trace elements in vivo and the characterization of their binding proteins is a story of intermittently made improvements of the techniques used for their detection. In this study we present the use of neutron activation analysis and different autoradiographic imaging methods including real-time digital autoradiography to reveal new insights in the hierarchy of selenium homeostasis. Selenoproteins containing the essential trace element selenium play important roles in the CNS. Although the CNS does not show the highest selenium concentration in the case of selenium-sufficient supply in comparison with other organs, it shows a high priority for selenium uptake and retention in the case of dietary selenium deficiency. To characterize the hierarchy of selenium supply in the brain, in vivo radiotracer labeling with (75)Se in rats with different selenium status was combined with autoradiographic detection of (75)Se in brain tissue sections and (75)Se-labeled selenoproteins after protein separation by two-dimensional gel electrophoresis. This study demonstrates significant differences in the uptake of (75)Se into the brain of rats with different selenium status. A brain region-specific uptake pattern of the radiotracer (75)Se in selenium-deficient rats could be revealed and the CSF was identified as a key part of the brain selenium homeostasis.

The crystal structure of the DNA-damage checkpoint inhibitor of sporulation, Sda, from Bacillus subtilis, has been solved by the MAD technique using selenomethionine-substituted protein. The structure closely resembles that previously solved by NMR, as well as the structure of a homologue from Geobacillus stearothermophilus solved in complex with the histidine kinase KinB. The structure contains three molecules in the asymmetric unit. The unusual trimeric arrangement, which lacks simple internal symmetry, appears to be preserved in solution based on an essentially ideal fit to previously acquired scattering data for Sda in solution. This interpretation contradicts previous findings that Sda was monomeric or dimeric in solution. This study demonstrates the difficulties that can be associated with the characterization of small proteins and the value of combining multiple biophysical techniques. It also emphasizes the importance of understanding the physical principles behind these techniques and therefore their limitations.

Clostridium difficile is a nosocomial pathogen whose incidence and importance are on the rise. Previous work in our laboratory characterized the central role of selenoenzyme-dependent Stickland reactions in C. difficile metabolism. In this work we have identified, using mass spectrometry, a stable complex formed upon reaction of auranofin (a gold-containing drug) with selenide in vitro. X-ray absorption spectroscopy supports the structure that we proposed on the basis of mass-spectrometric data. Auranofin potently inhibits the growth of C. difficile but does not similarly affect other clostridia that do not utilize selenoproteins to obtain energy. Moreover, auranofin inhibits the incorporation of radioisotope selenium ((75)Se) in selenoproteins in both Escherichia coli, the prokaryotic model for selenoprotein synthesis, and C. difficile without impacting total protein synthesis. Auranofin blocks the uptake of selenium and results in the accumulation of the auranofin-selenide adduct in the culture medium. Addition of selenium in the form of selenite or L-selenocysteine to the growth medium significantly reduces the inhibitory action of auranofin on the growth of C. difficile. On the basis of these results, we propose that formation of this complex and the subsequent deficiency in available selenium for selenoprotein synthesis is the mechanism by which auranofin inhibits C. difficile growth. This study demonstrates that targeting selenium metabolism provides a new avenue for antimicrobial development against C. difficile and other selenium-dependent pathogens.

Semaphore crabs (Heloecius cordiformis), soldier crabs (Mictyris platycheles), ghost shrimps (Trypaea australiensis), pygmy mussels (Xenostrobus securis), and polychaetes (Eunice sp.), key benthic prey items of predatory fish commonly found in estuaries throughout southeastern Australia, were exposed to dissolved (109)Cd and (75)Se for 385 h at 30 k Bq/l (uptake phase), followed by exposure to radionuclide-free water for 189 h (loss phase). The whole body uptake rates of (75)Se by pygmy mussels, semaphore crabs and soldier crabs were 1.9, 2.4 and 4.1 times higher than (109)Cd, respectively. There were no significant (P>0.05) differences between the uptake rates of (75)Se and (109)Cd for ghost shrimps and polychaetes. The uptake rates of (109)Cd and (75)Se were highest in pygmy mussels; about six times higher than in soldier crabs for (109)Cd and in polychaetes for (75)Se - the organisms with the lowest uptake rates. The loss rates of (109)Cd and (75)Se were highest in semaphore crabs; about four times higher than in polychaetes for (109)Cd and nine times higher than in ghost shrimps for (75)Se - the organisms with the lowest loss rates. The loss of (109)Cd and (75)Se in all organisms was best described by a two (i.e. short and a longer-lived) compartment model. In the short-lived, or rapidly exchanging, compartment, the biological half-lives of (75)Se (16-39 h) were about three times greater than those of (109)Cd (5-12h). In contrast, the biological half-lives of (109)Cd in the longer-lived, or slowly exchanging compartment(s), were typically greater (1370-5950 h) than those of (75)Se (161-1500 h). Semaphore crabs had the shortest biological half-lives of both radionuclides in the long-lived compartment, whereas polychaetes had the greatest biological half-life for (109)Cd (5950 h), and ghost shrimps had the greatest biological half-life for (75)Se (1500 h). This study provides the first reported data for the biological half-lives of Se in estuarine decapod crustaceans. Moreover, it emphasises the importance of determining metal(loid) accumulation and loss kinetics in keystone prey items, which consequently influences their trophic transfer potential to higher-order predators.

In the present article the radiotracer techniques have been combined with biochemical separation procedures to investigate the effects of changes in the selenium status on the expression of the selenium-containing proteins in the lung and their subcellular fractions. Subcellular separation of the lung has been achieved by differential ultracentrifugation. The selenium-containing proteins in these compartments have been investigated by labeling of rats in vivo with (75)Se, gel electrophoretic separation of the proteins, and autoradiographic detection of the tracer. In the lung of the selenium-deficient animals, the selenium administered was used predominantly to restore the levels of the selenoproteins, while in the lung of the selenium-sufficient animals most of the selenium retained was incorporated into the glutathione peroxidase. Also, higher activity of this enzyme has been found in the lung of the selenium-sufficient animals. The differences in the specific incorporation of the element in the selenium deficiency into different compounds suggested that there are different metabolic pathways for selenium, strongly dependent on its status.

The properties of (79)Se make it of likely potential importance in safety studies for geological disposal of radioactive wastes. Despite a substantial literature on toxic and nutritional aspects of selenium in the environment little consideration has been given to the behaviour of radioactive selenium and its potential transfer from a radioactive waste repository to the biosphere. Column experiments (15 x 50 cm), using a sandy loam soil, indicated that the upwards migration of (75)Se (as a surrogate for (79)Se) from a contaminated water table was dependent upon the redox status of the soil. Low redox conditions within the water table strongly limited upwards (75)Se soil migration, presumably due to the immobilisation of reduced Se species. Under natural conditions, (79)Se from a radioactive waste repository is therefore likely to accumulate at considerable depth. As a consequence, its absence from the rooting zone is likely to limit its transfer into plants. Nevertheless, the column experiments indicated that when an overlap between roots and soil contamination occurs, uptake into the plant is observed. Quantification of (75)Se volatilisation from the column surfaces suggested that this is a significant pathway by which (79)Se may move either directly from soil to the atmosphere, or from soil to plants and then to the atmosphere.

The intensity of (75)Se transport in the predominant direction after intraperitoneal injection of [(75)Se]selenate was compared in 1- and 3-month-old rats receiving common vivarium ration or sucrose diet. The incorporation percent, blood/liver relative radioactivity, and relative radioactivity difference coefficient were evaluated in the blood and liver. The dynamics of label incorporation in the blood of rats fed common diets has two peaks (at 1-3 h and 12-24 h) and a drop at 6 h. Coefficient of difference in 1-month-old rats was characterized by a greater amplitude of fluctuations than in 3-month-old animals.

Stable isotope tracers are safe and nutritionally relevant tools for the investigation of mineral metabolism in man. Increased research into the functional role of selenium has resulted in a need for well-characterised, isotopically enriched solutions of the element in order to determine the nutritional relevance of selenium fortification of foods. A simple method for the conversion of isotopically enriched elemental selenium (2.5-10 mg) into selenite and selenate, and their accurate characterisation and quantification is described. Analysis of selenite and selenate tracers using continuous-flow hydride generation-atomic absorption spectrometry technique was based on the specificity of the selenium hydride reaction and allowed their precise (RSD<2.5%) and accurate determination in aqueous solutions. The detection and determination limits were at 0.13 and 0.36 microg Se/l, respectively. Isotopically enriched elemental selenium was converted into selenite and selenate by a nitric acid and a combined nitric acid/hydrogen peroxide oxidation, respectively. The conversion was quantitative (>95%) and specific for both inorganic selenocompounds. Selenite and selenate labels were stable in 0.1 mol/l nitric acid for at least 18 months, i.e. making them ideally suitable for use in long-term metabolic studies. An overview of data relating to the absorption and retention of selenium by humans obtained using the two, well-characterised, tracers is presented and indicates that selenite and selenate are equally well retained in adult men and infants, despite differences in their absorption and urinary excretion characteristics.

Background: Whether ileal absorption of bile acid is up or downregulated in chronic cholestasis is still debated, and most evidence has come from animal studies., Aims: To compare ileal bile acid absorption in patients with primary biliary cirrhosis (PBC) and in healthy control subjects, and to assess the effect of ursodeoxycholic acid (UDCA)., Patients: We studied 14 PBC patients before and during (n=11) UDCA administration, 14 healthy control subjects, and 14 Crohn's disease patients (as disease controls)., Methods: We used cholescintigraphy to measure retention in the enterohepatic circulation over five successive days of the bile acid analogue (75)Se-homocholic acid-taurine ((75)SeHCAT) as an index of ileal bile acid absorption. Results were expressed as (75)SeHCAT fractional turnover rate (FTR) and t(1/2)12., Results: (75)SeHCAT FTR was 0.19 (0.11)/day, 0.34 (0.11)/day (p<0.001), and 0.83 (0.32)/day in PBC patients, healthy controls (p<0.0001), and Crohn's patients (p<0.001), respectively, which increased to 0.36 (0.16)/day in PBC patients during UDCA treatment (p<0.005). (75)SeHCAT t(1/2)12 was 4.8 (2.1) days in PBC patients, 2.2 (0.5) days (p<0.001) in healthy controls, and 1.0 (0.5) days (p<0.001) in Crohn's disease patients. (75)SeHCAT t(1/2)12 decreased to 2.2 (0.93) days (p< 0.001) in PBC patients during UDCA treatment., Conclusions: Our results support the concept that ileal bile acid absorption is upregulated in PBC patients, and that this effect may contribute towards damaging the cholestatic liver. This upregulation of bile acid absorption is abolished by UDCA.

The objective of this study was to determine whether simian immunodeficiency virus (SIV) infection of macaques could be used as a model system to assess the role of selenium in AIDS. Plasma and serum selenium levels were determined by standard assays in monkeys before and after inoculation of SIV. SIV-infected cells or cells expressing the HIV Tat protein were labeled with 75Se, and protein extracts were prepared and electrophoresed to analyze selenoprotein expression. Total tRNA was isolated from CEMx174 cells infected with SIV or from KK1 cells infected with HIV, and selenocysteine tRNA isoforms were characterized by reverse phase chromatography. SIV-infected monkeys show a decrease in blood selenium levels similar to that observed in AIDS with development of SAIDS. Cells infected with SIV in vitro exhibit reduced selenoprotein levels and an accumulation of small molecular weight selenium compounds relative to uninfected cells. Examination of the selenocysteine tRNA isoforms in HIV-infected KK1 cells or SIV-infected CEMx174 cells reveals an isoform distribution characteristic of selenium-deficient cells. Furthermore, transfection of Jurkat E6 cells with the Tat gene selectively altered selenoprotein synthesis, with GPX4 and Sep15 being the most inhibited and TR1 the most enhanced. Taken together, the data show that monkeys infected with SIV in vivo and cells infected with SIV in vitro will provide appropriate models for investigating the mechanism(s) responsible for reduced selenium levels that accompany the progression of AIDS in HIV disease.

In selenoproteins, incorporation of the amino acid selenocysteine is specified by the UGA codon, usually a stop signal. The alternative decoding of UGA is conferred by an mRNA structure, the SECIS element, located in the 3'-untranslated region of the selenoprotein mRNA. Because of the non-standard use of the UGA codon, current computational gene prediction methods are unable to identify selenoproteins in the sequence of the eukaryotic genomes. Here we describe a method to predict selenoproteins in genomic sequences, which relies on the prediction of SECIS elements in coordination with the prediction of genes in which the strong codon bias characteristic of protein coding regions extends beyond a TGA codon interrupting the open reading frame. We applied the method to the Drosophila melanogaster genome, and predicted four potential selenoprotein genes. One of them belongs to a known family of selenoproteins, and we have tested experimentally two other predictions with positive results. Finally, we have characterized the expression pattern of these two novel selenoprotein genes.

In this work, we report on a relationship between urinary selenium and the development of cervical uterine cancer. A simple chemical method was developed to concentrate trace amounts of selenium from relatively large urine samples by use of small activated carbon filters. When these filters are irradiated with thermal neutrons, selenium can be determined either by 77mSe (t1/2 = 17.5 s) or 75Se (t1/2 = 120 d). In this article, we report the results for 82 urine samples from women with cervical uterine cancer in several stages of development and from healthy controls. These results show a statistically significant increase of selenium excretion in cancer patients as compared to controls. Urinary selenium excretion is highest for patients in the intermediate stages of the disease.

The substrate-specific selenoprotein B of glycine reductase (PBglycine) from Eubacterium acidaminophilum was purified and characterized. The enzyme consisted of three different subunits with molecular masses of about 22 (alpha), 25 (beta) and 47 kDa (gamma), probably in an alpha 2 beta 2 gamma 2 composition. PBglycine purified from cells grown in the presence of [75Se]selenite was labeled in the 47-kDa subunit. The 22-kDa and 47-kDa subunits both reacted with fluorescein thiosemicarbazide, indicating the presence of a carbonyl compound. This carbonyl residue prevented N-terminal sequencing of the 22-kDa (alpha) subunit, but it could be removed for Edman degradation by incubation with o-phenylenediamine. A DNA fragment was isolated and sequenced which encoded beta and alpha subunits of PBglycine (grdE), followed by a gene encoding selenoprotein A (grdA2) and the gamma subunit of PBglycine (grdB2). The cloned DNA fragment represented a second GrdB-encoding gene slightly different from a previously identified partial grdBl-containing fragment. Both grdB genes contained an in-frame UGA codon which confirmed the observed selenium content of the 47-kDa (gamma) subunit. Peptide sequence analyses suggest that grdE encodes a proprotein which is cleaved into the previously sequenced N-terminal 25-kDa (beta) subunit and a 22-kDa (alpha) subunit of PBglycine. Cleavage most probably occurred at an -Asn-Cys- site concomitantly with the generation of the blocking carbonyl moiety from cysteine at the alpha subunit.

The metallic group XVIa elements selenium and tellurium possess remarkably similar chemical properties. However, unlike selenium, tellurium is not an essential micronutrient and, indeed, induces both acute and chronic toxicity in a variety of species. Despite this, very little is known of the molecular mechanisms of toxicity of tellurium, particularly with respect to potential chemical interactions with selenium-containing components in the cell. In this work we describe a novel interaction of inorganic tellurite with hepatocellular selenoproteins, particularly with selenium-dependent glutathione peroxidase. The accumulation of (121Te)-tellurite into cultured primary rat liver hepatocytes was shown to be much more rapid than that of (75Se)-selenite on a molar basis. Neither the uptake of (121Te)-tellurite nor of (75Se)-selenite was affected by a large molar excess of the unlabelled counterpart, respectively. Interestingly, separation of the hepatocellular proteins on continuous pH denaturing gels demonstrated clear binding of radiolabelled tellurium to a number of protein bands, including one at 23 and one at 58 kDa, which corresponded to proteins readily labelled in cells treated with (75Se)-selenite. The binding of (121Te) to these proteins was insensitive to reduction with mercaptoethanol and not affected by pre-treatment of the cells with cycloheximide. When purified selenium-dependent glutathione peroxidase was treated directly with (121Te)-tellurite, the protein became labelled in an analogous manner to that achieved in intact cells. This was not affected by coincubation of the enzyme with (121Te)-tellurite and one or both of its substrates. Additionally, incubation of the peroxidase with tellurite effectively inhibited its ability to catalyse glutathione-dependent reduction of hydrogen peroxide. These data suggest that inorganic tellurite delivers tellurium to the intracellular milieu in a form capable of binding to some intracellular selenoproteins and at least in the case of glutathione peroxidase, cause inhibition of catalytic activity. The nature of the binding seems not to be due to the insertion of tellurocysteine into the protein and the insensitivity to reductive cleavage with mercaptoethanol seems to preclude the formation of stable telluro-selenides in the proteins. These data may offer alternative explanations for the established toxicity of tellurium via disruption of selenoprotein function, particularly by the induction of intracellular oxidative stress by the inhibition of Se-dependent glutathione peroxidase.

The compound Se-75 bis[beta-(N,N,N-trimethylamino-)ethyl]selenide diiodide (Se-75 BISTAES) has been synthesized and its biodistribution in rabbits studied. A high uptake of radioactivity is found in the knee cartilage. Good scans of the knee are obtained by nuclear scintigraphy at 15 minutes after the injection of Se-75 BISTAES. The results of an equilibrium dialysis study show that Se-75 BISTAES binds to chondroitin sulfate and the binding is directly proportional to the chondroitin concentration. It appears that Se-75 BISTAES or its derivative should have potential as an articular imaging agent.

The generation of reactive oxygen species has been implicated as part of the mechanism responsible for UVB-radiation-induced skin damage. In mice, evidence suggests that increased dietary selenium intake may protect skin from many of the harmful effects of UVB radiation. We sought to determine the selenoprotein profile of cultured human skin cells and whether selenium supplementation could protect keratinocytes and melanocytes from the lethal effects of UVB radiation. Labelling experiments using [75Se]selenite showed qualitative and quantitative differences in selenoprotein expression by human fibroblasts, keratinocytes and melanocytes. This was most noticeable for thioredoxin reductase (60 kDa) and phospholipid glutathione peroxidase (21 kDa); these proteins were identified by Western blotting. Despite these differences, we found that a 24 h preincubation with sodium selenite or selenomethionine protected both cultured human keratinocytes and melanocytes from UVB-induced cell death. With primary keratinocytes, the greatest reduction in cell death was found with 10 nM sodium selenite (79% cell death reduced to 21.7%; P<0.01) and with 50 nM selenomethionine (79% cell death reduced to 13.2%; P<0.01). Protection could be obtained with concentrations as low as 1 nM with sodium selenite and 10 nM with selenomethionine. When selenium was added after UVB radiation, little protection could be achieved, with cell death only being reduced from 88.5% to about 50% with both compounds. In all of the experiments sodium selenite was more potent than selenomethionine at providing protection from UVB radiation.

Selenoproteins are involved in mechanisms of cell differentiation and defense. We investigated the expression of glutathione peroxidases, as well as other selenoproteins, in fetal human osteoblasts (hFOB-cells). Using 75-selenium metabolic labelling of viable hFOB-cells, we identified several selenoproteins in cell lysates of about 45-80 kDa and in the migration range of 14 kDa to 24 kDa. Cells expressed low mRNA levels of both cellular glutathione peroxidase and plasma glutathione peroxidase mRNA as analysed by Southern analysis of RT-PCR products. Basal cellular glutathione peroxidase enzyme activity in hFOB-cells (19.7 nmol NADPH oxidised per min and microg protein) was further increased 2.5-fold by the addition of 100 nM sodium selenite to the culture medium for 3 days. Furthermore, expression of selenoprotein P mRNA was demonstrated by RT-PCR. hFOB-cells did not show activities of the selenoproteins type I or type II 5'-deiodinase. In summary, we identified cellular glutathione peroxidase, plasma glutathione peroxidase and selenoprotein P among of a panel of several 75-selenium labelled proteins in human fetal osteoblasts. The expression of selenoproteins like glutathione peroxidases in hFOB-cells represents a new system of osteoblast antioxidative defense that may be relevant for the protection against hydrogen peroxide produced by osteoclasts during bone remodelling., (Copyright 1998 Academic Press.)

The blood-brain transfer, protein incorporation and metabolism of L-[75Se]selenomethionine (SeMet) of relatively high specific activity (> 400 GBq mmol) were studied in male Wistar rats. The highest uptake was found in the pancreas, followed by the tumor, kidney, liver, brain and muscle. In addition, plasma and brain samples of rats were analyzed for labeled fractions of free SeMet, metabolites, and SeMet bound to t-RNA and proteins. For example, free SeMet represented more than 80% of brain radioactivity at 1.5 min while it was less than 15% at 360 min. A concomitant increase was observed for protein bound SeMet in brain. A three-compartment model was applied to calculate the blood-brain transfer constant (K1 (0.15 +/- 0.070 mL g-1 min-1) and the rate constant of SeMet incorporation into proteins (k3 = 0.026 +/- 0.008 min-1). The apparent incorporation of methionine into proteins was estimated to be about 0.73 nmol g-1 min-1. From the studies it is concluded that the use of L-[75Se]selenomethionine may be appropriate to measure brain protein incorporation in humans with PET.

It is unclear whether bile acid absorption is affected by ileoanal pouch construction. Bile acid absorption was measured in the abdomen of 16 patients with pouches (nine with good and seven with poor pouch function based on a clinical score) and in six patients with an end ileostomy using a radiolabelled synthetic bile acid (75SeHCAT) enema and dynamic scintigraphy. The median (interquartile range) 75SeHCAT absorption was 81 (79-87) per cent in patients with ileostomy, 46 (43-53) per cent in patients having well functioning pouches, and 24 (18-38) per cent in the group with poor pouch function (P < 0.01). Log transformation of the absorption curves revealed a two-component uptake (fast component t1/2 = 1.4-7.2 min; slow component t1/2 = 16-144 min) in all patients with ileostomy, in eight of nine patients with pouches with good function, and in one of seven patients with a poorly functioning pouch (P < 0.05); in the other six patients with poorly functioning pouches, only the slow component of absorption was present. This test showed significantly reduced bile acid absorption in patients with an ileoanal pouch and gave an objective discrimination between well and poorly functioning pouches.

The effects of selenium compounds such as sodium selenite, sodium selenate, seleno-DL-cystine and seleno-DL-methionine (100 microM and 10 microM) on B16 and pigmented cloned pB16 murine melanoma cells were investigated in vitro. At the tested concentrations, B16 cells showed a greater sensitivity to the toxic effects of sodium selenite and seleno-DL-cystine than pB16 cells, whereas no decrease of B16 and pB16 cell number was observed after incubation with sodium selenate or seleno-DL-methionine. Glutathione (GSH) percentages were strongly decreased only by selenite and seleno-DL-cystine; it was marked more in B16 than in pB16 cells. The pretreatment of B16 cells with a GSH depleting agent (10 microM buthionine-[S,R]-sulfoximine) did not significantly influence the cytotoxic effects of selenite and seleno-DL-cystine. On both cell populations, GSH preincubation (50 microM) enhanced the cytotoxicity of selenite whereas the survival of seleno-DL-cystine treated cells was increased. Glutathione peroxidase (GSH-Px) activity in B16 cells was more sensitive than in pB16 cells to the activating effect of selenite, and particularly of seleno-DL-cystine: however, cell-free controls indicated that activation was mainly due to glutathione reductase. The rate of 75Se (as sodium selenite) uptake in both cell populations was maximal within the first hour of incubation, with a preferential accumulation in the cytosol; after 24 h of incubation, the amount of 75Se in cytosol and pellet was approximately the same.(ABSTRACT TRUNCATED AT 250 WORDS)

A 58 kDa selenium-labeled protein purified from mouse mammary epithelial cells (MMEC) was used to examine whether selenium modulates protein synthesis or is just a marker for cellular selenium status. The protein was isolated using Sephadex G150 gel filtration and DEAE-Sephadex A50 ion-exchange chromatography. It was further analysed using 2-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and was found as a single spot with a pI of 4.6. The immunoreactivity with anti-58 kDa antiserum and the 75Se signal co-localized on a single 58 kDa protein band on both 1D- and 2D-PAGE. Partial amino acid analysis of the peptide showed homology with the thiol protein disulfide oxidoreductase (TPDO). Varying the selenium concentration in culture medium did not affect the protein content or the immunoreactivity of the 58 kDa protein. Additionally, selenium did not seem to regulate the activity of TPDO in TM6 cells. The glutathione peroxidase activity of TM6 cells, taken as the internal positive control, was enhanced with the increase in selenium concentration in the medium. The results suggest that selenium is attached to the 58 kDa protein, but does not regulate either its protein synthesis or its functional activity. We conclude that selenium labeling of the 58 kDa protein reflects the cellular selenium status but probably is not involved in its chemopreventive ability.

Seventeen patients were operated on with intestinal shunts for morbid obesity, in eight a biliointestinal bypass (BI) was constructed and in the rest a conventional jejunoileal (JI)-shunt. The reduction in weight was similar in both groups, and so was malabsorption of fat, but the BI-group had significantly less bowel motions with less watery diarrhoea. Bile acid malabsorption was measured both chemically by estimating the total amount of faecal bile acids excreted, as well as indirectly by using a 75Se-labelled synthetic bile acid (SeHCAT). Both techniques revealed a substantial loss of bile acid after both types of operation, but patients with BI bypass surgery had significantly lower elimination time of the bile acid than those with JI-shunts. There was a significant negative correlation between SeHCAT retention and total faecal bile acids. However, some patients with low SeHCAT retention had normal or even reduced output of faecal bile acids. Estimation of faecal bile acids may display false negative results when the bile acid pool is decreased. The SeHCAT-test seems to be a better technique for measuring bile acid losses. The study suggests that BI bypass surgery for obesity seems to be advantageous over the JI shunt in reducing the postoperative loss of bile acids and choleretic diarrhoea, without influencing the weight loss.

The stability of tauro-23-[75Se]selena-25-homocholic acid (SeHCAT) towards deconjugation by the enzyme cholylglycine hydrolase was compared with that of taurocholate: whereas taurocholate underwent 58% deconjugation within 2 hr, SeHCAT suffered only 8% deconjugation plus 5% conversion to an unknown product within 24 hr. Incubation of SeHCAT under anaerobic conditions for 48 hr at 37 degrees C with human fecal organisms resulted in considerable deconjugation, 7 alpha-dehydroxylation, and dehydrogenation. Twenty-four hours after the simultaneous administration of SeHCAT and tauro-[24-14C]cholate to a rabbit the recovery of 75Se in bile was 90% of that of 14C. Forty-eight hours following administration of SeHCAT to a second rabbit residual bile radioactivity revealed 80% deconjugation and dehydroxylation and 60% reconjugation with glycine. Although SeHCAT is more resistant than taurocholate towards modification by fecal bacterial enzymes, within the rabbit it follows the principal metabolic pathways of the natural bile acids.

Pregnant rhesus monkeys (Macaca mulatta) were fed either selenium (Se) deficient or Se supplemented diets with adequate vitamin E. Except for some cardiac irregularities in the first babies born to these females, no physiological disorders due to Se deficiency were seen in a subsequent offspring. Plasma and erythrocyte glutathione peroxidase activities and blood Se levels increased in the Se supplemented monkeys but decreased in the deficient ones. The data indicated that hair Se levels reflect long term exposure to this element. In a very preliminary experiment, evidence was obtained to indicate that dietary protein deficiency along with Se deficiency will generate cardiomyopathic lesions characteristic of Se deficiency. It is hypothesized that, in addition to Se deficiency, another dietary deficiency (or abnormality) is necessary to produce Se deficiency lesions in higher primates. Higher glutathione transferase (or non-Se glutathione peroxidase) activity in tissues of rhesus monkeys may account for this resistance.