Sustained Activation of Nuclear Erythroid 2-Related Factor 2/Antioxidant Response Element Signaling Promotes Reductive Stress in the Human Mutant Protein Aggregation Cardiomyopathy in Mice

Oxidative stress can damage intestinal epithelial cell integrity and function, causing gastrointestinal disorders. Astragaloside IV (ASIV) exhibits a variety of biological and pharmacological properties, including anti-inflammatory and antioxidant effects. The purpose of this research was to investigate the cytoprotective action of ASIV and its mechanisms in calf small intestine epithelial cells with hydrogen peroxide (H2O2)-induced oxidative stress. ASIV pretreatment not only increased cell survival, but it also decreased reactive oxygen species generation and apoptosis, enhanced superoxide dismutase, catalase, and glutathione peroxidase levels, and it reduced malondialdehyde formation. Furthermore, pretreatment with ASIV elevated the mRNA and protein levels of nuclear factor erythroid 2-related factor 2 (NFE2L2), heme oxygenase-1 (HMOX1), and NAD(P)H quinone dehydrogenase 1 (NQO1). The NFE2L2 inhibitor ML385 inhibited NFE2L2 expression and then blocked HMOX1 and NQO1 expression. These results demonstrate that ASIV treatment effectively protects against H2O2-induced oxidative damage in calf small intestine epithelial cells through the activation of the NFE2L2-antioxidant response element signaling pathway.

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LAS0811: From Combinatorial Chemistry to Activation of Antioxidant Response Element

Journal of Biomedicine and Biotechnology ◽

10.1155/2009/420194 ◽

2009 ◽

Vol 2009 ◽

pp. 1-8 ◽

Cited By ~ 6

Author(s):

Ming Zhu ◽

Hyounggee Baek ◽

Ruiwu Liu ◽

Aimin Song ◽

Kit Lam ◽

...

Keyword(s):

Small Molecules ◽

Combinatorial Chemistry ◽

High Throughput Screening ◽

Cancer Chemoprevention ◽

Antioxidant Response ◽

Heme Oxygenase 1 ◽

Related Factor ◽

Antioxidant Response Element ◽

Response Element ◽

Enhanced Production

The antioxidant response element (ARE) and its transcription factor, nuclear factor-erythroid 2 p45-related factor 2 (Nrf2), are potential targets for cancer chemoprevention. We sought to screen small molecules synthesized with combinatorial chemistry for activation of ARE. By high-throughput screening of 9400 small molecules from 10 combinatorial chemical libraries using HepG2 cells with an ARE-driven reporter, we have identified a novel small molecule, 1,2-dimethoxy-4,5-dinitrobenzene (LAS0811), as an activator of the ARE. LAS0811 upregulated the activity of NAD(P)H:quinone oxidoreductase 1 (NQO1), a representative antioxidative enzyme regulated by ARE. It enhanced production of an endogenous reducing agent, glutathione (GSH). In addition, LAS0811 induced expression of heme oxygenase 1 (HO1), which is an ARE-regulated enzyme with anti-inflammatory activity. Furthermore, LAS0811 reduced cell death due to the cytotoxic stress of a strong oxidant, t-butyl hydroperoxide (t-BOOH). Mechanistically, LAS0811 upregulated the expression of Nrf2 and promoted its translocation into the nuclei leading to subsequent ARE activation. Taken together, LAS0811 is a novel activator of the ARE and its associated detoxifying genes and, thus, a potential agent for cancer chemoprevention.

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Identification of a novel Nrf2-regulated antioxidant response element (ARE) in the mouse NAD(P)H:quinone oxidoreductase 1 gene: reassessment of the ARE consensus sequence

Biochemical Journal ◽

10.1042/bj20030754 ◽

2003 ◽

Vol 374 (2) ◽

pp. 337-348 ◽

Cited By ~ 331

Author(s):

Paul NIOI ◽

Michael McMAHON ◽

Ken ITOH ◽

Masayuki YAMAMOTO ◽

John D. HAYES

Keyword(s):

Consensus Sequence ◽

Constitutive Expression ◽

Antioxidant Response ◽

Upstream Region ◽

Virus Protein ◽

Related Factor ◽

Antioxidant Response Element ◽

Mobility Shift ◽

Enhancer Activity ◽

Response Element

NQO1 [NAD(P)H:quinone oxidoreductase 1] has an integral role in cellular responses to oxidative stress. The expression of NQO1 is up-regulated in the mouse following challenge with electrophilic chemicals, in an Nrf2 (NF-E2 p45-related factor 2)-dependent fashion, but the molecular basis for this observation remains unexplained. Through characterization of the murine nqo1 5′-upstream region, we now show that Nrf2 regulates this gene directly via an ARE (antioxidant response element) that lies within a 24 bp region spanning nt −444 to −421. A comprehensive mutation study of this ARE revealed that it does not conform to the currently accepted ARE consensus sequence [(5′-TMAnnRTGAYnnnGCRwwww-3′, with essential nucleotides shown in capitals); two cytosine residues (shown in bold in the following sequence) that have been designated ‘n’ previously because they were thought to be redundant (5′-gagTcACaGTgAGtCggCAaaatt-3′) have now been found to be essential for enhancer activity; two guanines (also shown in bold) previously regarded as essential for ARE function (5′-gagTcACaGTgAGtCggCAaaatt-3′) have proven to be dispensable]. Examination of wild-type and nrf2−/− mouse embryonic fibroblasts demonstrated that Nrf2 is essential for both constitutive expression of NQO1 and its induction by sulphoraphane. Electrophoretic mobility-shift and chromatin immunoprecipitation assays revealed that Nrf2 associates, in low amounts, with the nqo1 ARE under constitutive conditions, and following sulphoraphane challenge of cells, Nrf2 is recruited to the ARE in substantially greater quantities, as a heterodimer with the small Maf (musculoaponeurotic fibrosarcoma virus) protein, MafK. Also, MafK was found to bind the nqo1 ARE in an Nrf2-independent fashion, and may contribute to transcriptional repression of the oxidoreductase gene. These findings allow a model for transcriptional control of nqo1 through the ARE to be proposed. Furthermore, our results indicate that distinct AREs have differential sequence requirements, and a universally applicable consensus sequence cannot be derived.

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