Selenium (Se), a micronutrient essential for human health, is incorporated into at least 25 selenoproteins including selenoprotein P (SePP), which transports Se within the body. This research identified two single nucleotide polymorphisms (SNPs) in the SePP gene, one in the coding region (position 24731, causing an Ala to Thr change) and one in the 3'untranslated region (position 25191). Their frequency was similar in Caucasian, Chinese, and South Asian populations. Prospectively genotyped volunteers were supplemented for 6 wk with 100 microg sodium selenite/day. Blood samples were analyzed for plasma Se and selenoprotein biomarkers at baseline, after supplementation, and during a washout period. Plasma Se, SePP, and glutathione peroxidase 3 (GPx3) levels increased with supplementation. Baseline plasma Se content depended on both SePP genotypes and body mass index. Presupplementation SePP concentration was associated with gender and genotype at SNP 24731 and postsupplementation concentration with SNP 25191. Both SNPs and gender were associated with differences in GPx3 activity, plasma, and erythrocyte thioredoxin reductase 1 concentrations and lymphocyte glutathione peroxidase 1 and 4 activities and concentrations. In conclusion, the data reveal two common functional SNPs within the human SePP gene that may predict behavior of biomarkers of Se status and response to supplementation and thus susceptibility to disease.

Glutathione S-transferases play a central role in drug detoxification and have been implicated in the sensitivity of tumour cells to anticancer drugs. In this study, glutathione S-transferase (GST) isozyme expression in normal and tumour tissue from human lung, colon, stomach, breast, kidney and liver tissue has been quantified using sensitive and subunit specific radioimmunoassays (RIA), together with Western blot analysis and measurement of substrate metabolism. Glutathione S-transferase pi was the predominant GST in the majority of the tumours examined. The concentration of this enzyme was increased significantly in tumour tissue relative to normal lung, colon, and stomach tissue. A strong correlation was observed (r = 0.77, P less than 0.01) between GST activity and GST pi levels in those tumour samples. The concentrations of the alpha class GST, the predominant isoenzymes in normal stomach, kidney and liver, decreased dramatically in tumour tissue from these organs. Western blot analysis revealed the presence of novel polypeptides that cross-reacted with antisera raised against alpha and mu class GST. Our data demonstrates that although GST pi is the predominant GST isoenzyme in many tumours, significant levels of the other GST subunits are also present and collectively can represent a significant proportion of the GST content. Therefore the properties of all the GST isoenzymes need consideration when assessing the role of these proteins in drug resistance. Selenium-dependent glutathione peroxidase, an enzyme activity also implicated in the mode of action of certain antitumour agents, was also studied and shown to be the predominant glutathione-dependent peroxidase in all tumours except the hepatoma.

In mammals, the cytosolic glutathione S-transferases (GSTs; EC 2.5.1.18) are a supergene family comprised of four multigene families, named alpha, mu, pi and theta. In man, within the mu class gene family there is a gene (the GSTmu 1 locus) that is polymorphic and is only expressed in 50-55% of individuals. It has previously been reported, using trans-stilbene oxide (tSBO) as a specific substrate for the expressed phenotype, that smokers with the null phenotype had a greater susceptibility to lung cancer. In a subsequent study, it was shown that on Southern blot analyses of human DNAs using a GSTmu 1 cDNA probe a DNA fragment was absent in certain individuals. The absence of this band correlated with the tSBO null phenotype. In the present work, DNA clones derived from GST mu class genomic sequences were used as probes in Southern blot analyses and confirmed the correlation between the lack of a DNA fragment and the null phenotype; moreover in this case, using radioimmunoassay for the GST mu protein, these probes were then used in a genotyping assay to investigate further the association of GSTmu 1 polymorphism with susceptibility to lung cancer. It was found that in a control group of 225 individuals, of unknown smoking history, 42% lacked the restriction fragment and were homozygous null, and therefore 58% were either heterozygous or were homozygous normal. Among 228 lung cancer patients, which included all tumour types, a similar distribution occurred, namely 43% were homozygous and 57% were heterozygous or homozygous normal. If, however, the tumours were analysed by tumour type a small but significant positive correlation with the homozygous null genotype was seen in squamous carcinoma of the lung, and an apparently negative correlation with adenocarcinoma of the lung.

Discovery of genetic mechanisms for resistance to obesity and diabetes may illuminate new therapeutic strategies for the treatment of this global health challenge. We used the polygenic Lean mouse model, selected for low adiposity over 60 generations, to identify thiosulfate sulfurtransferase (Tst, Rhodanese) as a candidate obesity-resistance gene with selectively increased adipocyte expression. Elevated adipose Tst expression correlated with indices of metabolic health across diverse mouse strains. Transgenic overexpression of Tst in adipocytes protected mice from diet-induced obesity and insulin-resistant diabetes. Tst gene deficiency markedly exacerbated diabetes whereas pharmacological TST activation ameliorated diabetes in mice in vivo. Mechanistically, TST selectively augmented mitochondrial function combined with degradation of reactive oxygen species and sulfide. In humans, adipose TST mRNA correlated positively with adipose insulin sensitivity and negatively with fat mass. Genetic identification of Tst as a beneficial regulator of adipocyte mitochondrial function may have therapeutic significance for type 2 diabetes.

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.

The ability of selenium to protect cultured human coronary artery endothelial cells (HCAEC), human umbilical vein endothelial cells (HUVEC) and bovine aortic endothelial cells (BAEC) from oxidative damage induced by 100 microM t-butyl hydroperoxide (t-BuOOH) was compared. Preincubation of human endothelial cells for 24 h with sodium selenite at concentrations as low as 5 nM provided significant protection against the harmful effects of 100 microM t-BuOOH, with complete protection being achieved with 40 nM selenite. The preincubation period was required for selenite to exert this protective effect on endothelial cells. When compared with selenium-deficient cells, the activities of cytoplasmic glutathione peroxidase (GPX-1), phospholipid hydroperoxide glutathione peroxidase (GPX-4) and thioredoxin reductase (TR) were each induced approx. 3--4-fold by 40 nM selenite. HCAEC and HUVEC showed great similarity in their relative abilities to resist oxidative damage in the presence and absence of selenite, and the activities of TR and the GPXs were also similar in these cell types. BAEC were more susceptible to damage by 100 microM t-BuOOH than were human endothelial cells, and could not be protected completely by incubation with selenite at concentrations up to 160 nM. The activity of TR in human endothelial cells was approx. 25-fold greater than that in BAEC of a similar selenium status, but GPX-1 and GPX-4 activities were not significantly different between the human and bovine cells. These studies, although performed with a small number of cultures, show for the first time that selenium at low doses can provide significant protection of the human coronary artery endothelium against damage by oxidative stress. TR may be an important antioxidant selenoprotein in this regard, in addition to the GPXs. The data also suggest that HUVEC, but not BAEC, represent a suitable model system in which to study the effects of selenium on the endothelium of human coronary arteries.

Isothiocyanates (ITCs) found in cruciferous vegetables are potentially important anticarcinogenic phytochemicals for many types of cancers including breast cancer. In this study, we have shown that three isothiocyanates, sulforaphane, erucin, and iberin, are potent inducers of thioredoxin reductase 1 (TrxR1) in human breast cancer MCF-7 cells. Sulforaphane, erucin, and iberin at 1 microM induce TrxR1 mRNA 2-3-fold within 8 h of treatment, and induce mRNA 5-7-fold with 12 microM ITC treatments. Selenium did not affect sulforaphane-induced TrxR1 mRNA levels, but significantly enhanced both TrxR1 protein expression (up to 9-fold in erucin treatment) and corresponding activities. These results suggest that dietary ITCs are important factors in the regulation of redox status through the induction of the selenoprotein thioredoxin reductase.

The ability of selenium to protect cultured human coronary artery endothelial cells (HCAEC), human umbilical vein endothelial cells (HUVEC) and bovine aortic endothelial cells (BAEC) from oxidative damage induced by 100 microM t-butyl hydroperoxide (t-BuOOH) was compared. Preincubation of human endothelial cells for 24 h with sodium selenite at concentrations as low as 5 nM provided significant protection against the harmful effects of 100 microM t-BuOOH, with complete protection being achieved with 40 nM selenite. The preincubation period was required for selenite to exert this protective effect on endothelial cells. When compared with selenium-deficient cells, the activities of cytoplasmic glutathione peroxidase (GPX-1), phospholipid hydroperoxide glutathione peroxidase (GPX-4) and thioredoxin reductase (TR) were each induced approx. 3--4-fold by 40 nM selenite. HCAEC and HUVEC showed great similarity in their relative abilities to resist oxidative damage in the presence and absence of selenite, and the activities of TR and the GPXs were also similar in these cell types. BAEC were more susceptible to damage by 100 microM t-BuOOH than were human endothelial cells, and could not be protected completely by incubation with selenite at concentrations up to 160 nM. The activity of TR in human endothelial cells was approx. 25-fold greater than that in BAEC of a similar selenium status, but GPX-1 and GPX-4 activities were not significantly different between the human and bovine cells. These studies, although performed with a small number of cultures, show for the first time that selenium at low doses can provide significant protection of the human coronary artery endothelium against damage by oxidative stress. TR may be an important antioxidant selenoprotein in this regard, in addition to the GPXs. The data also suggest that HUVEC, but not BAEC, represent a suitable model system in which to study the effects of selenium on the endothelium of human coronary arteries.