Glucosinolates from pak choi and broccoli induce enzymes and inhibit inflammation and colon cancer differently.

High consumption of Brassica vegetables is considered to prevent especially colon carcinogenesis. The content and pattern of glucosinolates (GSLs) can highly vary among different Brassica vegetables and may, thus, affect the outcome of Brassica intervention studies. Therefore, we aimed to feed mice with diets containing plant materials of the Brassica vegetables broccoli and pak choi. Further enrichment of the diets by adding GSL extracts allowed us to analyze the impact of different amounts (GSL-poor versus GSL-rich) and different patterns (broccoli versus pak choi) of GSLs on inflammation and tumor development in a model of inflammation-triggered colon carcinogenesis (AOM/DSS model). Serum albumin adducts were analyzed to confirm the up-take and bioactivation of GSLs after feeding the Brassica diets for four weeks. In agreement with their high glucoraphanin content, broccoli diets induced the formation of sulforaphane-lysine adducts. Levels of 1-methoxyindolyl-3-methyl-histidine adducts derived from neoglucobrassicin were the highest in the GSL-rich pak choi group. In the colon, the GSL-rich broccoli and the GSL-rich pak choi diet up-regulated the expression of different sets of typical Nrf2 target genes like Nqo1, Gstm1, Srxn1, and GPx2. GSL-rich pak choi induced the AhR target gene Cyp1a1 but did not affect Ugt1a1 expression. Both colitis and tumor number were drastically reduced after feeding the GSL-rich pak choi diet while the other three diets had no effect. GSLs can act anti-inflammatory and anti-carcinogenic but both effects depend on the specific amount and pattern of GSLs within a vegetable. Thus, a high Brassica consumption cannot be generally considered to be cancer-preventive.

Feeding of selenium alone or in combination with glucoraphanin differentially affects intestinal and hepatic antioxidant and phase II enzymes in growing rats.

The anti-carcinogenic effects of sulforaphane (SFN) are based on the up-regulation of antioxidant enzymes (AE) and phase II enzymes (PIIE) through the transcription factor Nrf2. Current knowledge on the roles of the SFN precursor glucoraphanin (GRA) on these processes is limited. Anti-carcinogenic effects of Se depending on glutathione peroxidase (GPx) activity have also been reported. We studied effects and possible synergisms of Se and GRA on the expression and activity of a broad spectrum of AE and PIIE in jejunum, colon and the liver of rats fed diets differing in Se and GRA concentration. In all organs, GPx1 mRNA expression was 70 % to 90 % lower in Se deficiency than in Se sufficiency. GPx2 expression increased in jejunum and liver under Se deficiency and decreased in the colon. Se deficiency increased most colonic AE and PIIE compared to Se adequacy. Adequate and in particular supranutritive Se combined with GRA increased colonic AE and PIIE expression up to 3.72-fold. In the liver Se deficiency raised the expression of AE and PIIE up to 4.49-fold. GRA attenuated liver AE and PIIE response in Se deficiency. Expression- and correlation analyses revealed that Keap1 mRNA better reflects AE and PIIE gene expression than Nrf2 mRNA. We conclude that: (1) GPx1 sensitively indicates Se deficiency; (2) the influence of Se and Nrf2/Keap1 on GPx2 expression depends on the organ; (3) GRA combined with supranutritive Se may effectively protect against inflammation and colon cancer; (4) future investigations on AE and PIIE expression should consider the role of Keap1 to a higher extent.

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.

Glucoraphanin does not reduce plasma homocysteine in rats with sufficient Se supply via the induction of liver ARE-regulated glutathione biosynthesis enzymes.

Data from human and animal trials have revealed contradictory results regarding the influence of selenium (Se) status on homocysteine (HCys) metabolism. It was hypothesised that sufficient Se reduces the flux of HCys through the transsulphuration pathway by decreasing the expression of glutathione (GSH) synthesising enzymes. Glucoraphanin (GRA) is a potent inducer of genes regulated via an antioxidant response element (ARE), including those of GSH biosynthesis. We tested the hypothesis that GRA supplementation to rat diets lowers plasma HCys levels by increasing GSH synthesis. Therefore 96 weaned albino rats were assigned to 8 groups of 12 and fed diets containing four different Se levels (15, 50, 150 and 450 µg kg(diet)(-1)), either without GRA (groups: C15, C50, C150 and C450) or in combination with 700 µmol GRA kg(diet)(-1) (groups G15, G50, G150 and G450). Rats fed the low Se diets C15 and G15 showed an impressive decrease of plasma HCys. Se supplementation increased plasma HCys and lowered GSH significantly by reducing the expression of GSH biosynthesis enzymes. As new molecular targets explaining these results, we found a significant down-regulation of the hepatic GSH exporter MRP4 and an up-regulation of the HCys exporter Slco1a4. In contrast to our hypothesis, GRA feeding did not reduce plasma HCys levels in Se supplemented rats (G50, G150 and 450) through inducing GSH biosynthesis enzymes and MRP4, but reduced their mRNA in some cases to a higher extent than Se alone. We conclude: 1. That the long-term supplementation of moderate GRA doses reduces ARE-driven gene expression in the liver by increasing the intestinal barrier against oxidative stress. 2. That the up-regulation of ARE-regulated genes in the liver largely depends on GRA cleavage to free sulforaphane and glucose by plant-derived myrosinase or bacterial ß-glucosidases. As a consequence, higher dietary GRA concentrations should be used in future experiments to test if GRA or sulforaphane can be established as HCys lowering compounds.

Breakdown products of neoglucobrassicin inhibit activation of Nrf2 target genes mediated by myrosinase-derived glucoraphanin hydrolysis products.

Glucosinolates (GLSs) present in Brassica vegetables serve as precursors for biologically active metabolites, which are released by myrosinase and induce phase 2 enzymes via the activation of Nrf2. Thus, GLSs are generally considered beneficial. The pattern of GLSs in plants is various, and contents of individual GLSs change with growth phase and culture conditions. Whereas some GLSs, for example, glucoraphanin (GRA), the precursor of sulforaphane (SFN), are intensively studied, functions of others such as the indole GLS neoglucobrassicin (nGBS) are rather unknown as are functions of combinations thereof. We therefore investigated myrosinase-treated GRA, nGBS and synthetic SFN for their ability to induce NAD(P)H:quinone oxidoreductase 1 (NQO1) as typical phase 2 enzyme, and glutathione peroxidase 2 (GPx2) as novel Nrf2 target in HepG2 cells. Breakdown products of nGBS potently inhibit both GRA-mediated stimulation of NQO1 enzyme and Gpx2 promoter activity. Inhibition of promoter activity depends on the presence of an intact xenobiotic responsive element (XRE) and is also observed with benzo[a]pyrene, a typical ligand of the aryl hydrocarbon receptor (AhR), suggesting that suppressive effects of nGBS are mediated via AhR/XRE pathway. Thus, the AhR/XRE pathway can negatively interfere with the Nrf2/ARE pathway which has consequences for dietary recommendations and, therefore, needs further investigation.