Identification of the NF-E2-related Factor-2-dependent Genes Conferring Protection against Oxidative Stress in Primary Cortical Astrocytes Using Oligonucleotide Microarray Analysis

The antioxidant responsive element (ARE) mediates transcriptional regulation of phase II detoxification enzymes and antioxidant proteins such as NAD(P)H:quinone oxidoreductase (NQO1), glutathione S-transferases, and glutamate-cysteine ligase. In this study, we demonstrate that NF-E2-related factor-2 (Nrf2) plays a major role in transcriptional activation of ARE-driven genes and identify Nrf2-dependent genes by oligonucleotide microarray analysis using primary cortical astrocytes from Nrf2(+/+) and Nrf2(-/-) mice. Nrf2(-/-) astrocytes had decreased basal NQO1 activity and no induction by tert-butylhydroquinone compared with Nrf2(+/+) astrocytes. Similarly, both basal and induced levels of human NQO1-ARE-luciferase expression in Nrf2(-/-) astrocytes were significantly lower than in Nrf2(+/+) astrocytes. Furthermore, human NQO1-ARE-luciferase expression in Nrf2(-/-) astrocytes was restored by overexpression of Nrf2, whereas ARE activation in Nrf2(+/+) astrocytes was completely blocked by dominant-negative Nrf2. In addition, we observed that Nrf2-dependent genes protected primary astrocytes from H(2)O(2)- or platelet-activating factor-induced apoptosis. In support of these observations, we identified Nrf2-dependent genes encoding detoxification enzymes, glutathione-related proteins, antioxidant proteins, NADPH-producing enzymes, and anti-inflammatory genes using oligonucleotide microarrays. Proteins within these functional categories are vital to the maintenance and responsiveness of a cell defense system, suggesting that an orchestrated change in gene expression via Nrf2 and the ARE gives a synergistic protective effect against oxidative stress.