Regular articleInduction of hepatic thioredoxin reductase activity by sulforaphane, both in Hepa1c1c7 cells and in male Fisher 344 rats☆
Introduction
Mammalian TR is a 110 kDa homodimeric selenoprotein that catalyzes the NADPH-dependent reduction of thioredoxin. Electrons are transferred from NADPH via FAD to a conserved redox site. Each monomer is comprised of a prosthetic FAD group, a redox active disulfide site and a pentultimate selenocysteine amino acid residue [1].
Mammalian TR is an antioxidant that it is able to reduce various substrates including lipoic acid, lipid hydroperoxides, NK-lysin, vitamin K3, dehydroascorbic acid, ascorbyl free radical, and the tumor suppressor protein p53 [for review see 2]. It is hypothesized that the presence of the selenocysteine redox site in mammalian TR allows for this broad reducing substrate specificity [2].
Thioredoxin provides reducing equivalents for ribonucleotide reductase, which is essential for DNA synthesis [3]. Also, it regulates the action of several gene transcription factors including NF-κB [4], AP-1 [5] and the glucocorticoid receptor [6] through a redox mechanism by providing reducing equivalents to facilitate binding to DNA [7]. Thioredoxin must be in a reduced form to be active and TR is the primary means of reduction. Ultimately, all of the functions of thioredoxin are tied to the presence of TR.
In animals, TR activity is regulated by dietary Se, and activity drops to less than 5% of normal values in selenium deficient rat livers [8], [9]. In cell culture, TR activity is increased in a dose dependent manner by 1-10 μM supplemental Se, and falls to below 5% of controls when cultures are depleted of Se [10].
Both TR activity and protein levels are induced by t-butylhydroquinone (BHQ) in cultured cortical astrocytes [11]. Because BHQ is known to upregulate a number of detoxification enzymes via the ARE, these researchers suggested that TR may be regulated in part by the ARE which is known to mediate the induction of QR and other enzymes such as γ-glutamylcysteine synthetase and glutathione S-transferase [for review see 12]. The objective of the present study was to determine if the antioxidant selenoprotein TR can be induced by SF, a compound from broccoli known to induce detoxification enzymes via the ARE [12].
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Materials
Solvents used for extraction and purification were of reagent-grade, and solvents used for gas chromatography (GC) or high performance liquid chromatography (HPLC) analysis were of HPLC-grade. All solvents were purchased from Fisher Scientific (Fair Lawn, NJ). Sulforaphane was purified from broccoli seed by the method of Matusheski et al., [13]. Purity was determined to be >99% by GC analysis. Unless otherwise noted, all other materials used in this study were obtained from Sigma (St. Louis,
Animal experiment
Food intake for the highest treatment group, [20% broccoli + 5mmol SF/kg diet] was significantly lower than intake for the group receiving control diet. However, it was not different from any other diet containing 20% broccoli. Weight gain was not different among groups (data not shown).
Diets containing both 20% broccoli and purified SF increased hepatic TR activity compared to the control diet (P < 0.05, Fig. 1). In contrast, hepatic GSH-Px activity was significantly depressed in animals fed
Discussion
The present study demonstrates that in livers of rats fed adequate Se, TR activity and protein can be increased by dietary SF + broccoli. Conversely, we found that hepatic GSH-Px activity and protein were decreased in rats by dietary SF + broccoli. The animals employed in this study were also used in a study relating dietary SF to the induction of hepatic and colonic QR.2 In that study, hepatic QR induction was found to correlate closely with urinary SF mercapturate (r = 0.73, P = 0.0006).
Notes
References (36)
- et al.
Isolation and characterization of thioredoxin, the hydrogen donor from Escherichia coli
B J Biol Chem
(1964) - et al.
Evidence that the endogenous heat-stable glucocorticoid receptor-activating factor is thioredoxin
J Biol Chem
(1983) - et al.
Effect of selenium on rat thioredoxin reductase activity
Biochemical Pharmacology
(1999) - et al.
Determination of thioredoxin reductase activity in rat liver supernatant
AnalBiochem
(1997) - et al.
Meth Enzymol
(1967) - et al.
Thioredoxin and thioredoxin reductase
Methods Enzymol
(1995) - et al.
Selenium content of foods purchased in North Dakota
Nutr Res
(1996) - et al.
Selenium regulation of thioredoxin reductase activity and mRNA levels in rat liver
J Nutr Biochem
(2001) - et al.
Selenium from high selenium broccoli protects rats from colon cancer
J Nutr
(2000) - et al.
The core promoter of human thioredoxin reductase 1cloning, transcriptional activity, and Oct-1, Sp1, and Sp3 binding reveal a housekeeping-type promoter for the AU-rich element-regulated gene
J Biol Chem
(2001)
The antioxidant responsive element. Activation by oxidative stress and identification of the DNA consensus sequence required for functional activity
J Biol Chem
Transcriptional regulation of the antioxidant response element. Activation by Nrf2 and repression by MafK
J Biol Chem
Transcriptional regualtion of a rat liver glutathione-S-transferase Ya subunit gene
J Biol Chem
Hemin-induced activation of the thioredoxin gene by Nrf2. A differential regulation of the antioxidant responsive element by a switch of its binding factors
J Biol Chem
The chemical form of selenium influences 3,2′-dimethyl-4-aminobiphenyl- DNA adduct formation in rat colon
J Nutr
Dietary selenium reduces the formation of aberrant crypts in rats administered 3,2′-dimethyl-4-aminobiphenyl
Toxicol Appl Pharmacol
Selenocysteine, identified as the penultimate C-terminal residue in human T-cell thioredoxin reductase, corresponds to TGA in the human placental gene
Proc Natl Acad Sci USA
Thioredoxin reductase
Biochem J
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