Fluvastatin protects vascular smooth muscle cells against oxidative stress through the Nrf2-dependent antioxidant pathway.
ABSTRACT HMG-CoA reductase inhibitors (statins) have pleiotropic actions, including the ability to reduce vascular oxidative stress. Transcription factor nuclear factor-erythroid 2-related factor 2 (Nrf2) is an important regulator of cellular oxidative stress. This study examined the role of Nrf2 in statin-mediated antioxidant effects in vascular smooth muscle cells.
In cultured human coronary artery smooth muscle cells (hCASMCs), fluvastatin activated the nuclear translocation of Nrf2, as evaluated by Western blotting and immunocytochemical analyses. Nrf2-antioxidant response element (ARE) activity was measured with a luciferase assay after transfection of reporter plasmids containing AREs. Fluvastatin significantly increased the transcriptional activity of the ARE. Electromobility shift assays using an ARE probe detected a complex that was significantly increased in intensity by fluvastatin. Western blotting and luciferase assay revealed fluvastatin activated Nrf2 via the PI3K/Akt pathway. Statins upregulated the Nrf2-related antioxidant genes heme oxygenase-1, NAD(P)H quinone oxidoreductase-1, and glutamate-cysteine ligase modifier subunits. Inhibition of Nrf2 by siRNA reduced statin-induced upregulation of these antioxidant genes. Moreover, Nrf2 siRNA markedly reduced the cytoprotective effects of fluvastatin against H(2)O(2) administration in hCASMCs.
Fluvastatin exerts cytoprotective effects against oxidative stress, inducing antioxidant genes through Nrf2/ARE in hCASMCs. These results suggest that the Nrf2/ARE pathway plays an important role in the regulation of statin-mediated antioxidant effects in vascular smooth muscle cells.
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ABSTRACT: Nrf2/ARE pathway plays an important role in adapt to oxidative stress caused by pro-oxidants and electrophiles through up-regulating phase II detoxifying enzymes. Our previous study has demonstrated that PCB quinone exposure causes severe cellular oxidative stress (Toxicology in Vitro 26 (2012) 841-848). There are no reports describing the ability of PCB quinone on Nrf2/ARE activation. In the present study, we found that exposure to PCB29-pQ resulted in a significant increase in Nrf2 and Keap1 expression in total protein, as well as the Nrf2 targeting genes, including NQO1 and HO-1. Next, immunocytochemistry analysis identified the accumulation of Nrf2 in nucleus subsequent to PCB29-pQ treatment. The increased Nrf2 and constant Keap1 expression in nucleus suggested the dissociation of Nrf1/Keap1 complex. Similarly, mRNA level of Nrf2 was elevated significantly with PCB29-pQ treatment, but not Keap1. Additionally, PCB29-pQ treatment led to significant up-regulation of the mRNA level of antioxidant enzymes, NQO1 and HO-1, in a concentration-dependent manner. Electrophoretic mobility shift assay and luciferase reporter assay further confirmed the formation of Nrf2-ARE complex. PCB29-pQ treatment has no effect on mitogen-activated protein kinase signaling, however, phospho-AKT was up-regulated and GSK-3β was down-regulated. Pretreatment with LY294002, a specific inhibitor of phosphatidylinositol 3-kinase (PI3K), suppressed the phosphorylation of AKT and inhibited PCB29-pQ induced Nrf2/HO-1 activation, meanwhile, GSK-3β expression was increased accordingly. At last, reactive oxygen species (ROS) scavengers inhibited PCB29-pQ induced Nrf2 activation partly. These results suggested that Nrf2 activation by PCB29-pQ in HepG2 cells is associated with ROS and AKT pathway but not MAPK signaling, the activation of Nrf2/ARE may be an adaptive response to oxidative stress.Chemico-biological interactions 12/2013; · 2.46 Impact Factor
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ABSTRACT: Anti-oxidant capacity is crucial defence against environmental or endogenous oxidative stress. Nuclear factor erythroid 2-related factor 2 (Nrf2) is a redox-sensitive transcription factor that plays a key defensive role against oxidative and cytotoxic stress and cellular senescence. However, Nrf2 signalling is impaired in several aging-related diseases, such as chronic pulmonary obstructive disease (COPD), cancer, and neurodegenerative diseases. Thus, novel therapeutics that enhance Nrf2 signalling are an attractive approach to treat these diseases. Nrf2 was stabilized by SKI-II (2-(p-hydroxyanilino)-4-(p-chlorophenyl) thiazole), which is a known sphingosine kinase inhibitor, in human bronchial epithelial cell line, BEAS2B, and in primary human bronchial epithelial cells, leading to enhancement of anti-oxidant proteins, such as HO-1, NQO1 and GCLM. The activation of Nrf2 was achieved by the generation of inactive dimerized form of Keap1, a negative regulator of Nrf2 expression, which was independent of sphingosine kinase inhibition. Using mice that were exposed to cigarette smoke, SKI-II induced Nrf2 expression together with HO-1 in their lungs. In addition, SKI-II reduced cigarette smoke mediated oxidative stress, macrophages and neutrophil infiltration and markers of inflammation in mice. SKI-II appears to be a novel activator of Nrf2 signalling via the inactivation of Keap1.PLoS ONE 02/2014; 9(2):e88168. · 3.53 Impact Factor
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ABSTRACT: Oxidative stress, a pathogenetic factor in many conditions, including chronic obstructive pulmonary disease, arises due to accumulation of reactive oxygen species and defective antioxidant defenses in the lungs. The latter is due, at least in part, to impaired activation of NF-E2-related factor 2 (Nrf2), a transcription factor involved in the activation of antioxidant and cytoprotective genes. The bromodomain and extraterminal (BET) proteins, Brd2, Brd3, Brd4, and BrdT, bind to acetylated lysine residues on histone or nonhistone proteins recruiting transcriptional regulators and thus activating or repressing gene transcription. We investigated whether BET proteins modulate the regulation of Nrf2-dependent gene expression in primary human airway smooth muscle cells and the human monocytic cell line, THP-1. Inhibition of BET protein bromodomains using the inhibitor JQ1+ or attenuation of Brd2 and Brd4 expression using small interfering RNA led to activation of Nrf2-dependent transcription and expression of the antioxidant proteins heme oxygenase-1, NADPH quinone oxidoreductase 1, and glutamate-cysteine ligase catalytic subunit. Also, JQ1+ prevented H2O2-induced intracellular reactive oxygen species production. By coimmunoprecipitation, BET proteins were found to be complexed with Nrf2, whereas chromatin-immunoprecipitation studies indicated recruitment of Brd2 and Brd4 to Nrf2-binding sites on the promoters of heme oxygenase-1 and NADPH quinone oxidoreductase 1. BET proteins, particularly Brd2 and Brd4, may play a key role in the regulation of Nrf2-dependent antioxidant gene transcription and are hence an important target for augmenting antioxidant responses in oxidative stress-mediated diseases.The Journal of Immunology 04/2014; · 5.36 Impact Factor