Metallothionein-III protects against 6-hydroxydopamine-induced oxidative stress by increasing expression of heme oxygenase-1 in a PI3K and ERK/Nrf2-dependent manner

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hwang, Yong Pil; Kim, Hyung Gyun; Han, Eun Hee

2008-09-15

The zinc-binding protein metallothionein-III (MT-III) is associated with resistance to neuronal injury. However, the underlying mechanism for its effects is unclear. In this study, we demonstrate that MT-III prevents the accumulation of reactive oxygen species (ROS) in dopaminergic SH-SY5Y cells challenged with the Parkinson's disease-related neurotoxin 6-hydroxydopamine (6-OHDA) by a mechanism that involves phosphatidylinositol 3-kinase (PI3K) and ERK kinase/NF-E2-related factor 2 (Nrf2) dependent induction of the stress response protein heme oxygenase-1 (HO-1). Pretreatment of SH-SY5Y cells with MT-III significantly reduced 6-OHDA-induced generation of ROS, caspase-3 activation, and subsequent cell death. Also, MT-III up-regulates HO-1 expression and this expression confers neuroprotectionmore » against oxidative injury induced by 6-OHDA. Moreover, MT-III induces Nrf2 nuclear translocation, which is upstream of MT-III-induced HO-1 expression, and PI3K and ERK1/2 activation, a pathway that is involved in induced Nrf2 nuclear translocation, HO-1 expression and neuroprotection. Taken together, these results suggest that the PI3K and ERK/Nrf2 signaling pathway controls the intracellular levels of ROS by regulating the expression of the antioxidant enzyme HO-1.« less

Signal transduction involved in lipoxin A4-induced protection of tubular epithelial cells against hypoxia/reoxygenation injury

PubMed Central

Wu, Sheng-Hua; Wang, Ming-Jie; Lü, Jing; Chen, Xiao-Qing

2017-01-01

Previous studies have reported that lipoxin A4 (LXA4) may exert a renoprotective effect on ischemia/reperfusion injury in various animal models. The underlying mechanism of LXA4-induced renoprotection during ischemia/reperfusion injury remains to be elucidated. The present study investigated LXA4-induced protection on renal tubular cells subjected to hypoxia/reoxygenation (H/R) injury, and determined the effects of peroxisome proliferator-activated receptor-γ (PPARγ) and heme oxygenase-1 (HO-1) on LXA4 treatment. HK-2 human tubular epithelial cells exposed to H/R injury were pretreated with LXA4, signal molecule inhibitors or the HO-1 inhibitor zinc protoporphyrin-IX, or were transfected with PPARγ small interfering RNA (siRNA) or nuclear factor E2-related factor 2 (Nrf2) siRNA. The protein and mRNA expression levels of PPARγ and HO-1 were analyzed using western blotting and reverse transcription-quantitative polymerase chain reaction. Binding activity of Nrf2 to the HO-1 E1 enhancer was determined using chromatin immunoprecipitation. Nrf2 binding to the HO-1 antioxidant responsive element (ARE) was assessed using electrophoretic mobility shift assay. Preincubation of cells with LXA4 exposed to H/R injury led to a decreased production of inducible nitrogen oxide synthase, malondialdehyde, γ-glutamyl transpeptidase, leucine aminopeptidase and N-acetyl-β-glucosaminidase. In addition, LXA4 pretreatment increased cell viability, protein and mRNA expression levels of PPARγ and HO-1 and PPARγ and HO-1 promoter activity. SB20358 is a p38 mitogen-activated protein kinase (p38 MAPK) pathway inhibitor, which reduced LXA4-induced PPARγ expression levels. LXA4 treatment upregulated p38 MAPK activation, Nrf2 nuclear translocation and increased binding activity of Nrf2 to HO-1 ARE and E1 enhancer in cells exposed to H/R injury. Transfection of the cells with PPARγ siRNA reduced the LXA4-induced Nrf2 translocation. Transfection of the cells with PPARγ siRNA or Nrf2 siRNA also reduced the LXA4-induced increase in HO-1 expression. In conclusion, LXA4-induced protection of renal tubular cells against H/R injury was associated with the induction of PPARγ and HO-1, via activation of the p38 MAPK pathway, as well as Nrf2 nuclear translocation and binding to HO-1 ARE and E1 enhancer. Therefore, LXA4-induced renoprotection is associated with activation of the p38 MAPK/PPARγ/Nrf2-ARE/HO-1 pathway. PMID:28259922

Nuclear translocation of Nrf2 and expression of antioxidant defence genes in THP-1 cells exposed to carbon nanotubes.

PubMed

Brown, David M; Donaldson, Kenneth; Stone, Vicki

2010-06-01

Carbon nanotubes have a wide range of applications in various industries and their use is likely to rise in the future. Currently, a major concern is that with the increasing use and production of these materials, there may be increased health risks to exposed workers. Long (> 15 microm) straight nanotubes may undergo frustrated phagocytosis which is likely to result in reduced clearance. We examine here the effects of multiwalled carbon nanotubes of different sizes on monocytic THP-1 cells, with regard to their ability to stimulate increased expression of the HO-1 and GST genes and their ability to produce nuclear translocation of the transcription factor, Nrf2, as well as the release of several pro-inflammatory cytokines and mediators of inflammation. Our results suggest that long (50 microm) carbon nanotubes (62.5 microg/ml for 4 hours) produce increased nuclear translocation of Nrf2 and increased HO-1 gene expression compared with shorter entangled nanotubes. There was no increased gene expression for GST. The long nanotubes (NT1) caused increased release of the proinflammatory cytokine IL-1beta, an effect which was diminished by the antioxidant trolox, suggesting a role of oxidative stress in the upregulation of this cytokine. Tentatively, our study suggests that long carbon nanotubes may exert their effect in THP-1 cells in part via an oxidative stress mechanism.

Redox regulation of antioxidant enzymes: post-translational modulation of catalase and glutathione peroxidase activity by resveratrol in diabetic rat liver.

PubMed

Sadi, Gökhan; Bozan, Davut; Yildiz, Huseyin Bekir

2014-08-01

Resveratrol is a strong antioxidant that exhibits blood glucose-lowering effects, which might contribute to its usefulness in preventing complications associated with diabetes. The present study aimed to investigate resveratrol effects on catalase (CAT) and glutathione peroxidase (GPx) gene and protein expression, their phosphorylation states and activities in rat liver of STZ-induced diabetes. Diabetes increased the levels of total protein phosphorylation and p-CAT, while mRNA expression, protein levels, and activity were reduced. Although diabetes induced transcriptional repression over GPx, it did not affect the protein levels and activity. When resveratrol was administered to diabetic rats, an increase in activity was associated with an increase in p-GPx levels. Decrease in Sirtuin1 (SIRT1) and nuclear factor erythroid 2-related factor (Nrf2) and increase in nuclear factor kappa B (NFκB) gene expression in diabetes were associated with a decrease in CAT and GPx mRNA expression. A possible compensatory mechanism for reduced gene expression of antioxidant enzymes is proved to be nuclear translocation of redox-sensitive Nrf2 and NFκB in diabetes which is confirmed by the increase in nuclear and decrease in cytoplasmic protein levels of Nrf2 and NFκB. Taken together, these findings revealed that an increase in the oxidized state in diabetes intricately modified the cellular phosphorylation status and regulation of antioxidant enzymes. Gene regulation of antioxidant enzymes was accompanied by nuclear translocation of Nrf2 and NFκB. Resveratrol administration also activated a coordinated cytoprotective response against diabetes-induced changes in liver tissues.

Activation of KGFR-Akt-mTOR-Nrf2 signaling protects human retinal pigment epithelium cells from Ultra-violet.

PubMed

Hu, Haitao; Hao, Lanxiang; Tang, Chunzhou; Zhu, Yunxi; Jiang, Qin; Yao, Jin

2018-01-15

Ultra-violet (UV) radiation causes oxidative injuries to human retinal pigment epithelium (RPE) cells. We tested the potential effect of keratinocyte growth factor (KGF) against the process. KGF receptor (KGFR) is expressed in ARPE-19 cells and primary human RPE cells. Pre-treatment with KGF inhibited UV-induced reactive oxygen species (ROS) production and RPE cell death. KGF activated nuclear-factor-E2-related factor 2 (Nrf2) signaling in RPE cells, causing Nrf2 Ser-40 phosphorylation, stabilization and nuclear translocation as well as expression of Nrf2-dependent genes (HO1, NOQ1 and GCLC). Nrf2 knockdown (by targeted shRNAs) or S40T mutation almost reversed KGF-induced RPE cell protection against UV. Further studies demonstrated that KGF activated KGFR-Akt-mTORC1 signaling to mediate downstream Nrf2 activation. KGFR shRNA or Akt-mTORC1 inhibition not only blocked KGF-induced Nrf2 Ser-40 phosphorylation and activation, but also nullified KGF-mediated RPE cell protection against UV. We conclude that KGF-KGFR activates Akt-mTORC1 downstream Nrf2 signaling to protect RPE cells from UV radiation. Copyright © 2017 Elsevier Inc. All rights reserved.

Global mapping of binding sites for Nrf2 identifies novel targets in cell survival response through ChIP-Seq profiling and network analysis

PubMed Central

Malhotra, Deepti; Portales-Casamar, Elodie; Singh, Anju; Srivastava, Siddhartha; Arenillas, David; Happel, Christine; Shyr, Casper; Wakabayashi, Nobunao; Kensler, Thomas W.; Wasserman, Wyeth W.; Biswal, Shyam

2010-01-01

The Nrf2 (nuclear factor E2 p45-related factor 2) transcription factor responds to diverse oxidative and electrophilic environmental stresses by circumventing repression by Keap1, translocating to the nucleus, and activating cytoprotective genes. Nrf2 responses provide protection against chemical carcinogenesis, chronic inflammation, neurodegeneration, emphysema, asthma and sepsis in murine models. Nrf2 regulates the expression of a plethora of genes that detoxify oxidants and electrophiles and repair or remove damaged macromolecules, such as through proteasomal processing. However, many direct targets of Nrf2 remain undefined. Here, mouse embryonic fibroblasts (MEF) with either constitutive nuclear accumulation (Keap1−/−) or depletion (Nrf2−/−) of Nrf2 were utilized to perform chromatin-immunoprecipitation with parallel sequencing (ChIP-Seq) and global transcription profiling. This unique Nrf2 ChIP-Seq dataset is highly enriched for Nrf2-binding motifs. Integrating ChIP-Seq and microarray analyses, we identified 645 basal and 654 inducible direct targets of Nrf2, with 244 genes at the intersection. Modulated pathways in stress response and cell proliferation distinguish the inducible and basal programs. Results were confirmed in an in vivo stress model of cigarette smoke-exposed mice. This study reveals global circuitry of the Nrf2 stress response emphasizing Nrf2 as a central node in cell survival response. PMID:20460467

Piper betle induces phase I & II genes through Nrf2/ARE signaling pathway in mouse embryonic fibroblasts derived from wild type and Nrf2 knockout cells.

PubMed

Wan Hasan, Wan Nuraini; Kwak, Mi-Kyoung; Makpol, Suzana; Wan Ngah, Wan Zurinah; Mohd Yusof, Yasmin Anum

2014-02-23

Nuclear factor-erythroid 2 p45 related factor 2 (Nrf2) is a primary transcription factor, protecting cells from oxidative stress by regulating a number of antioxidants and phase II detoxifying enzymes. Dietary components such as sulforaphane in broccoli and quercetin in onions have been shown to be inducers of Nrf2. Piper betle (PB) grows well in tropical climate and the leaves are used in a number of traditional remedies for the treatment of stomach ailments and infections among Asians. The aim of this study was to elucidate the effect of Piper betle (PB) leaves extract in Nrf2 signaling pathway by using 2 types of cells; mouse embryonic fibroblasts (MEFs) derived from wild-type (WT) and Nrf2 knockout (N0) mice. WT and N0 cells were treated with 5 and 10 μg/ml of PB for 10 and 12-h for the determination of nuclear translocation of Nrf2 protein. Luciferase reporter gene activity was performed to evaluate the antioxidant response element (ARE)-induction by PB. Real-time PCR and Western blot were conducted on both WT and N0 cells after PB treatment for the determination of antioxidant enzymes [superoxide dismutase (SOD1) and heme-oxygenase (HO-1)], phase I oxidoreductase enzymes [ quinone oxidoreductase (NQO1)] and phase II detoxifying enzyme [glutathione S-transferase (GST)]. Nuclear translocation of Nrf2 by PB in WT cells was better after 10 h incubation compared to 12 h. Real time PCR and Western blot analysis showed increased expressions of Nrf2, NQO1 and GSTA1 genes with corresponding increases in glutathione, NQO1 and HO-1 proteins in WT cells. Reporter gene ARE was stimulated by PB as shown by ARE/luciferase assay. Interestingly, PB induced SOD1 gene and protein expressions in N0 cells but not in WT cells. The results of this study confirmed that PB activated Nrf2-ARE signaling pathway which subsequently induced some phase I oxidoreductase, phase II detoxifying and antioxidant genes expression via ARE reporter gene involved in the Nrf2 pathway with the exception of SOD1 which may not be dependent on this pathway.

Piper betle induces phase I & II genes through Nrf2/ARE signaling pathway in mouse embryonic fibroblasts derived from wild type and Nrf2 knockout cells

PubMed Central

2014-01-01

Background Nuclear factor-erythroid 2 p45 related factor 2 (Nrf2) is a primary transcription factor, protecting cells from oxidative stress by regulating a number of antioxidants and phase II detoxifying enzymes. Dietary components such as sulforaphane in broccoli and quercetin in onions have been shown to be inducers of Nrf2. Piper betle (PB) grows well in tropical climate and the leaves are used in a number of traditional remedies for the treatment of stomach ailments and infections among Asians. The aim of this study was to elucidate the effect of Piper betle (PB) leaves extract in Nrf2 signaling pathway by using 2 types of cells; mouse embryonic fibroblasts (MEFs) derived from wild-type (WT) and Nrf2 knockout (N0) mice. Methods WT and N0 cells were treated with 5 and 10 μg/ml of PB for 10 and 12-h for the determination of nuclear translocation of Nrf2 protein. Luciferase reporter gene activity was performed to evaluate the antioxidant response element (ARE)-induction by PB. Real-time PCR and Western blot were conducted on both WT and N0 cells after PB treatment for the determination of antioxidant enzymes [superoxide dismutase (SOD1) and heme-oxygenase (HO-1)], phase I oxidoreductase enzymes [NAD(P)H: quinone oxidoreductase (NQO1)] and phase II detoxifying enzyme [glutathione S-transferase (GST)]. Results Nuclear translocation of Nrf2 by PB in WT cells was better after 10 h incubation compared to 12 h. Real time PCR and Western blot analysis showed increased expressions of Nrf2, NQO1 and GSTA1 genes with corresponding increases in glutathione, NQO1 and HO-1 proteins in WT cells. Reporter gene ARE was stimulated by PB as shown by ARE/luciferase assay. Interestingly, PB induced SOD1 gene and protein expressions in N0 cells but not in WT cells. Conclusion The results of this study confirmed that PB activated Nrf2-ARE signaling pathway which subsequently induced some phase I oxidoreductase, phase II detoxifying and antioxidant genes expression via ARE reporter gene involved in the Nrf2 pathway with the exception of SOD1 which may not be dependent on this pathway. PMID:24559113

Lipoxin A4-Induced Heme Oxygenase-1 Protects Cardiomyocytes against Hypoxia/Reoxygenation Injury via p38 MAPK Activation and Nrf2/ARE Complex

PubMed Central

Chen, Xiao-Qing; Wu, Sheng-Hua; Zhou, Yu; Tang, Yan-Rong

2013-01-01

Objective To investigate whether lipoxin A4 (LXA4) increases expression of heme oxygenase-1(HO-1) in cardiomyocytes, whether LXA4-induced HO-1 protects cardiomyocytes against hypoxia/reoxygenation (H/R) injury, and what are the mechanisms involved in the LXA4-induced HO-1 induction. Methods Rat cardiomyocytes were exposed to H/R injury with or without preincubation with LXA4 or HO-1 inhibitor ZnPP-IX or various signal molecule inhibitors. Expressions of HO-1 protein and mRNA were analyzed by using Western blot and RT-PCR respectively. Activity of nuclear factor E2-related factor 2 (Nrf2) binding to the HO-1 E1 enhancer was assessed by chromatin immunoprecipitation. Nrf2 binding to the HO-1 antioxidant responsive element (ARE) were measured by using electrophoretic mobility shift assay. Results Pretreatment of the cells undergoing H/R lesion with LXA4 significantly reduced the lactate dehydrogenase and creatine kinase productions, increased the cell viability, and increased the expressions of HO-1 protein and mRNA and HO-1 promoter activity. HO-1 inhibition abolished the protective role of LXA4 on the cells undergoing H/R lesion. LXA4 increased p38 mitogen-activated protein kinase (p38 MAPK) activation, nuclear translocation of Nrf2, Nrf2 binding to the HO-1 ARE and E1 enhancer in cardiomyocytes with or without H/R exposure. Conclusion The protection role of LXA4 against H/R injury of cardiomyocytes is related to upregulation of HO-1, via activation of p38 MAPK pathway and nuclear translocation of Nrf2 and Nrf2 binding to the HO-1 ARE and E1 enhancer, but not via activation of phosphatidyinositol-3-kinase or extracellular signal-regulated kinase pathway. PMID:23826208

Active form of vitamin D ameliorates non-alcoholic fatty liver disease by alleviating oxidative stress in a high-fat diet rat model.

PubMed

Zhu, Chong-Gui; Liu, Ya-Xin; Wang, Hao; Wang, Bao-Ping; Qu, Hui-Qi; Wang, Bao-Li; Zhu, Mei

2017-07-28

The purpose of this study was to determine whether treatment using the active form of vitamin D (1,25(OH) 2 D 3 ) could protect against high-fat diet (HFD)-induced non-alcoholic fatty liver disease (NAFLD) in rats and ameliorate oxidative stress. Male Sprague-Dawley rats were divided into three groups and treated with standard chow, HFD, or HFD plus intraperitoneal injection of 1,25(OH) 2 D 3 (5 μg/kg body weight, twice per week), respectively, for 16 weeks. Serum lipid profiles, hepatic function, intrahepatic lipid, and calcium levels were determined. Hepatic histology was examined using hematoxylin/eosin, Masson's trichrome, and Oil Red O staining. Oxidative stress was assessed by measuring hepatic malondialdehyde (MDA) and F2α-isoprostane content. Expression of nuclear factor-erythroid-2-related factor 2 (Nrf2) and downstream target genes was analyzed using quantitative RT-PCR. 1,25(OH) 2 D 3 treatment improved the serum lipid profile, reduced intrahepatic lipid levels, and attenuated hepatic steatosis and inflammation in HFD rats. Furthermore, MDA and F2α-isoprostane levels in liver tissue were reduced by 1,25(OH) 2 D 3 administration. Although 1,25(OH) 2 D 3 did not regulate the expression of Nrf2 mRNA, it did induce Nrf2 nuclear translocation. The expression of Nrf2 target genes, including Gclc, Nqo1, Sod2, and Cat, was up-regulated by 1,25(OH) 2 D 3 . We conclude that 1,25(OH) 2 D 3 protects against HFD-induced NAFLD by attenuating oxidative stress, inducing NRF2 nuclear translocation, and up-regulating the expression of genes encoding antioxidant enzymes.

Ginger compound [6]-shogaol and its cysteine-conjugated metabolite (M2) activate Nrf2 in colon epithelial cells in vitro and in vivo.

PubMed

Chen, Huadong; Fu, Junsheng; Chen, Hao; Hu, Yuhui; Soroka, Dominique N; Prigge, Justin R; Schmidt, Edward E; Yan, Feng; Major, Michael B; Chen, Xiaoxin; Sang, Shengmin

2014-09-15

In this study, we identified Nrf2 as a molecular target of [6]-shogaol (6S), a bioactive compound isolated from ginger, in colon epithelial cells in vitro and in vivo. Following 6S treatment of HCT-116 cells, the intracellular GSH/GSSG ratio was initially diminished but was then elevated above the basal level. Intracellular reactive oxygen species (ROS) correlated inversely with the GSH/GSSG ratio. Further analysis using gene microarray showed that 6S upregulated the expression of Nrf2 target genes (AKR1B10, FTL, GGTLA4, and HMOX1) in HCT-116 cells. Western blotting confirmed upregulation, phosphorylation, and nuclear translocation of Nrf2 protein followed by Keap1 decrease and upregulation of Nrf2 target genes (AKR1B10, FTL, GGTLA4, HMOX1, and MT1) and glutathione synthesis genes (GCLC and GCLM). Pretreatment of cells with a specific inhibitor of p38 (SB202190), PI3K (LY294002), or MEK1 (PD098059) attenuated these effects of 6S. Using ultra-high-performance liquid chromatography-tandem mass spectrometry, we found that 6S modified multiple cysteine residues of Keap1 protein. In vivo 6S treatment induced Nrf2 nuclear translocation and significantly upregulated the expression of MT1, HMOX1, and GCLC in the colon of wild-type mice but not Nrf2(-/-) mice. Similar to 6S, a cysteine-conjugated metabolite of 6S (M2), which was previously found to be a carrier of 6S in vitro and in vivo, also activated Nrf2. Our data demonstrated that 6S and its cysteine-conjugated metabolite M2 activate Nrf2 in colon epithelial cells in vitro and in vivo through Keap1-dependent and -independent mechanisms.

Ginger Compound [6]-Shogaol and Its Cysteine-Conjugated Metabolite (M2) Activate Nrf2 in Colon Epithelial Cells in Vitro and in Vivo

PubMed Central

2015-01-01

In this study, we identified Nrf2 as a molecular target of [6]-shogaol (6S), a bioactive compound isolated from ginger, in colon epithelial cells in vitro and in vivo. Following 6S treatment of HCT-116 cells, the intracellular GSH/GSSG ratio was initially diminished but was then elevated above the basal level. Intracellular reactive oxygen species (ROS) correlated inversely with the GSH/GSSG ratio. Further analysis using gene microarray showed that 6S upregulated the expression of Nrf2 target genes (AKR1B10, FTL, GGTLA4, and HMOX1) in HCT-116 cells. Western blotting confirmed upregulation, phosphorylation, and nuclear translocation of Nrf2 protein followed by Keap1 decrease and upregulation of Nrf2 target genes (AKR1B10, FTL, GGTLA4, HMOX1, and MT1) and glutathione synthesis genes (GCLC and GCLM). Pretreatment of cells with a specific inhibitor of p38 (SB202190), PI3K (LY294002), or MEK1 (PD098059) attenuated these effects of 6S. Using ultra-high-performance liquid chromatography–tandem mass spectrometry, we found that 6S modified multiple cysteine residues of Keap1 protein. In vivo 6S treatment induced Nrf2 nuclear translocation and significantly upregulated the expression of MT1, HMOX1, and GCLC in the colon of wild-type mice but not Nrf2–/– mice. Similar to 6S, a cysteine-conjugated metabolite of 6S (M2), which was previously found to be a carrier of 6S in vitro and in vivo, also activated Nrf2. Our data demonstrated that 6S and its cysteine-conjugated metabolite M2 activate Nrf2 in colon epithelial cells in vitro and in vivo through Keap1-dependent and -independent mechanisms. PMID:25148906

A prenylated flavonoid, 10-oxomornigrol F, exhibits anti-inflammatory effects by activating the Nrf2/heme oxygenase-1 pathway in macrophage cells.

PubMed

Tran, Phi-Long; Tran, Phuong Thao; Tran, Huynh Nguyen Khanh; Lee, Suhyun; Kim, Okwha; Min, Buyng-Sun; Lee, Jeong-Hyung

2018-02-01

Prenylated flavonoids are a unique class of naturally occurring flavonoids that have various pharmacological activities. In the present study, we investigated the anti-inflammatory effect in murine macrophages of a prenylated flavonoid, 10-oxomornigrol F (OMF), which was isolated from the twigs of Morus alba (Moraceae). OMF inhibited the lipopolysaccharide (LPS)-induced production of nitric oxide (NO), tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6 in RAW264.7 cells, as well as in mouse bone marrow-derived macrophages (BMMs). OMF also rescued LPS-induced septic mortality in ICR mice. LPS-induced expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), TNF-α and IL-6 was also significantly suppressed by OMF treatment in RAW264.7 cells. Treatment of RAW264.7 cells with OMF induced heme oxygenase (HO)-1 mRNA and protein expression and increased the nuclear translocation of the nuclear factor-E2-related factor 2 (Nrf2) as well as the expression of Nrf2 target genes, such as NAD(P)H:quinone oxidoreductase 1 (NQO1). Treatment of RAW264.7 cells with OMF increased the intracellular level of reactive oxygen species (ROS) and the phosphorylation levels of p38 mitogen-activated protein kinase (MAPK); co-treatment with the antioxidant N-acetyl-cysteine (NAC) blocked this OMF-induced p38 MAPK phosphorylation. Moreover, NAC, or SB203580 (a p38 MAPK inhibitor), blocked the OMF-induced nuclear translocation of Nrf2 and HO-1 expression, suggesting that OMF induces HO-1 expression by activating Nrf2 through the p38 MAPK pathway. Consistent with the notion that the Nrf2/HO-1 pathway has anti-inflammatory properties, inhibiting HO-1 significantly abrogated the anti-inflammatory effects of OMF in LPS-stimulated RAW264.7 cells. Taken together, these findings suggest that OMF exerts its anti-inflammatory effect by activating the Nrf2/HO-1 pathway, and may be a potential Nrf2 activator to prevent or treat inflammatory diseases. Copyright © 2017 Elsevier B.V. All rights reserved.

Amomum tsao-ko suppresses lipopolysaccharide-induced inflammatory responses in RAW264.7 macrophages via Nrf2-dependent heme oxygenase-1 expression.

PubMed

Li, Bin; Choi, Hee-Jin; Lee, Dong-Sung; Oh, Hyuncheol; Kim, Youn-Chul; Moon, Jin-Young; Park, Won-Hwan; Park, Sun-Dong; Kim, Jai-Eun

2014-01-01

Amomum tsao-ko Crevost et Lemaire, used as a spice in Asia, is an important source of Chinese cuisine and traditional Chinese medicines. A. tsao-ko is reported to exert a variety of biological and pharmacological activities, including anti-proliferative, anti-oxidative and neuroprotective effects. In this study, NNMBS227, consisting of the ethanol extract of A. tsao-ko, exhibited potent anti-inflammatory activities in RAW264.7 macrophages. We investigated the effect of NNMBS227 in the suppression of pro-inflammatory mediators, including pro-inflammatory enzymes (inducible nitric oxide synthase and cyclooxygenase-2) and cytokines (tumor necrosis factor-α and interleukin-1β) in LPS stimulated macrophages. NNMBS227 also inhibited the phosphorylation and degradation of IκB-α, as well as the nuclear translocation of nuclear factor kappa B (NF-κB) p65 caused by stimulation with LPS. In addition, NNMBS227 induced heme oxygenase (HO)-1 expression through the nuclear translocation of nuclear factor E2-related factor 2 (Nrf2) in macrophages. Using tin protoporphyrin (SnPP), an HO activity inhibitor, we confirmed an association between the anti-inflammatory effects of NNMBS227 and the up-regulation of HO-1. These findings suggest that Nrf2-dependent increases in the expression of HO-1 induced by NNMBS227 conferred anti-inflammatory activities in LPS stimulated RAW264.7 macrophages.

Development of a cell-based high throughput luciferase enzyme fragment complementation assay to identify nuclear-factor-e2-related transcription factor 2 activators.

PubMed

Xie, Wensheng; Pao, Christina; Graham, Taylor; Dul, Ed; Lu, Quinn; Sweitzer, Thomas D; Ames, Robert S; Li, Hu

2012-12-01

Nuclear-factor-E2-related transcription factor 2 (Nrf2) regulates a large panel of Phase II genes and plays an important role in cell survival. Nrf2 activation has been shown as preventing cigarette smoke-induced alveolar enlargement in mice. Therefore, activation of the Nrf2 protein by small-molecule activators represents an attractive therapeutic strategy that is used for chronic obstructive pulmonary disease. In this article, we describe a cell-based luciferase enzyme fragment complementation assay that identifies Nrf2 activators. This assay is based on the interaction of Nrf2 with its nuclear partner MafK or runt-related transcription factor 2 (RunX2) and is dependent on the reconstitution of a "split" luciferase. Firefly luciferase is split into two fragments, which are genetically fused to Nrf2 and MafK or RunX2, respectively. BacMam technology was used to deliver the fusion constructs into cells for expression of the tagged proteins. When the BacMam-transduced cells were treated with Nrf2 activators, the Nrf2 protein was stabilized and translocated into the nucleus where it interacted with MafK or RunX2. The interaction of Nrf2 and MafK or RunX2 brought together the two luciferase fragments that form an active luciferase. The assay was developed in a 384-well format and was optimized by titrating the BacMam concentration, transduction time, cell density, and fetal bovine serum concentration. It was further validated with known Nrf2 activators. Our data show that this assay is robust, sensitive, and amenable to high throughput screening of a large compound collection for the identification of novel Nrf2 activators.

Lycopene inhibits ICAM-1 expression and NF-κB activation by Nrf2-regulated cell redox state in human retinal pigment epithelial cells.

PubMed

Yang, Po-Min; Wu, Zhi-Zhen; Zhang, Yu-Qi; Wung, Being-Sun

2016-06-15

Age-related macular degeneration (AMD) is one of the most common diseases leading to blindness in elderly people. The progression of AMD may be prevented through anti-inflammation and antioxidation in retinal pigment epithelium (RPE) cells. Lycopene, a carotenoid, has been shown to possess both antioxidative and anti-inflammatory properties. This research was conducted to detail the mechanisms of these effects of lycopene-treated RPE cells. We exposed ARPE-19 cells to TNFα after pretreatment with lycopene, and measured monocyte adhesion, ICAM-1 expression, NF-κB nuclear translocation, and transcriptional activity. Cell viability was assayed with Alamar Blue. The cell redox state was tested by glutathione (GSH) and reactive oxygen species (ROS) levels. The importance of the Nrf2 pathway was tested in nuclear translocation, promoter reporter assay, and siRNA. Lycopene could reduce TNF-α-induced monocyte adhesion and H2O2- induced cell damage in RPE cells. Furthermore, lycopene inhibits ICAM-1 expression and abolishes NF-κB activation for up to 12h in TNFα-treated RPE cells. Lycopene upregulates Nrf2 levels in nuclear extracts and increases the transactivity of antioxidant response elements. The use of Nrf2 siRNA blocks the inhibitory effect of lycopene in TNF-α-induced ICAM-1 expression and NF-κB activation. Glutamate-cysteine ligase (GCL) is the rate-limiting enzyme in the de novo synthesis of GSH. We found that lycopene increases intracellular GSH levels and GCL expression. Following lycopene treatment, TNF-α-induced ROS production was abolished. The Nrf2-regulated antioxidant property plays a pivotal role in the anti-inflammatory mechanism underlying the inhibition of NF-κB activation in lycopene-treated ARPE-19 cells. Copyright © 2016 Elsevier Inc. All rights reserved.

Melatonin attenuates memory impairment induced by Klotho gene deficiency via interactive signaling between MT2 receptor, ERK, and Nrf2-related antioxidant potential.

PubMed

Shin, Eun-Joo; Chung, Yoon Hee; Le, Hoang-Lan Thi; Jeong, Ji Hoon; Dang, Duy-Khanh; Nam, Yunsung; Wie, Myung Bok; Nah, Seung-Yeol; Nabeshima, Yo-Ichi; Nabeshima, Toshitaka; Kim, Hyoung-Chun

2014-12-30

We demonstrated that oxidative stress plays a crucial role in cognitive impairment in klotho mutant mice, a genetic model of aging. Since down-regulation of melatonin due to aging is well documented, we used this genetic model to determine whether the antioxidant property of melatonin affects memory impairment. First, we examined the effects of melatonin on hippocampal oxidative parameters and the glutathione/oxidized glutathione (GSH/GSSG) ratio and memory dysfunction of klotho mutant mice. Second, we investigated whether a specific melatonin receptor is involved in the melatonin-mediated pharmacological response by application with melatonin receptor antagonists. Third, we examined phospho-extracellular-signal-regulated kinase (ERK) expression, nuclear factor erythroid 2-related factor 2 (Nrf2) nuclear translocation, Nrf2 DNA binding activity, and glutamate-cysteine ligase (GCL) mRNA expression. Finally, we examined effects of the ERK inhibitor SL327 in response to antioxidant efficacy and memory enhancement mediated by melatonin. Treatment with melatonin resulted in significant attenuations of oxidative damage, a decrease in the GSH/GSSG ratio, and a significant amelioration of memory impairment in this aging model. These effects of melatonin were significantly counteracted by the selective MT2 receptor antagonist 4-P-PDOT. Importantly, 4-P-PDOT or SL327 also counteracted melatonin-mediated attenuation in response to the decreases in phospho-ERK expression, Nrf2 nuclear translocation, Nrf2 DNA-binding activity, and GCL mRNA expression in the hippocampi of klotho mutant mice. SL327 also counteracted the up-regulation of the GSH/GSSG ratio and the memory enhancement mediated by melatonin in klotho mutant mice. Melatonin attenuates oxidative stress and the associated memory impairment induced by klotho deficiency via signaling interaction between the MT2 receptor and ERK- and Nrf2-related antioxidant potential. © The Author 2015. Published by Oxford University Press on behalf of CINP.

Melatonin Attenuates Memory Impairment Induced by Klotho Gene Deficiency Via Interactive Signaling Between MT2 Receptor, ERK, and Nrf2-Related Antioxidant Potential

PubMed Central

Shin, Eun-Joo; Chung, Yoon Hee; Le, Hoang-Lan Thi; Jeong, Ji Hoon; Dang, Duy-Khanh; Nam, Yunsung; Wie, Myung Bok; Nah, Seung-Yeol; Nabeshima, Yo-Ichi; Nabeshima, Toshitaka; Kim, Hyoung-Chun

2015-01-01

Background: We demonstrated that oxidative stress plays a crucial role in cognitive impairment in klotho mutant mice, a genetic model of aging. Since down-regulation of melatonin due to aging is well documented, we used this genetic model to determine whether the antioxidant property of melatonin affects memory impairment. Methods: First, we examined the effects of melatonin on hippocampal oxidative parameters and the glutathione/oxidized glutathione (GSH/GSSG) ratio and memory dysfunction of klotho mutant mice. Second, we investigated whether a specific melatonin receptor is involved in the melatonin-mediated pharmacological response by application with melatonin receptor antagonists. Third, we examined phospho-extracellular-signal-regulated kinase (ERK) expression, nuclear factor erythroid 2-related factor 2 (Nrf2) nuclear translocation, Nrf2 DNA binding activity, and glutamate-cysteine ligase (GCL) mRNA expression. Finally, we examined effects of the ERK inhibitor SL327 in response to antioxidant efficacy and memory enhancement mediated by melatonin. Results: Treatment with melatonin resulted in significant attenuations of oxidative damage, a decrease in the GSH/GSSG ratio, and a significant amelioration of memory impairment in this aging model. These effects of melatonin were significantly counteracted by the selective MT2 receptor antagonist 4-P-PDOT. Importantly, 4-P-PDOT or SL327 also counteracted melatonin-mediated attenuation in response to the decreases in phospho-ERK expression, Nrf2 nuclear translocation, Nrf2 DNA-binding activity, and GCL mRNA expression in the hippocampi of klotho mutant mice. SL327 also counteracted the up-regulation of the GSH/GSSG ratio and the memory enhancement mediated by melatonin in klotho mutant mice. Conclusions: Melatonin attenuates oxidative stress and the associated memory impairment induced by klotho deficiency via signaling interaction between the MT2 receptor and ERK- and Nrf2-related antioxidant potential. PMID:25550330

Ginsenoside Rb1 improves postoperative fatigue syndrome by reducing skeletal muscle oxidative stress through activation of the PI3K/Akt/Nrf2 pathway in aged rats.

PubMed

Zhuang, Cheng-Le; Mao, Xiang-Yu; Liu, Shu; Chen, Wei-Zhe; Huang, Dong-Dong; Zhang, Chang-Jing; Chen, Bi-Cheng; Shen, Xian; Yu, Zhen

2014-10-05

Ginsenoside Rb1 is reported to possess anti-fatigue activity, but the mechanisms remain unknown. The aim of this study was to investigate the molecular mechanisms responsible for the anti-fatigue effect of ginsenoside Rb1 on postoperative fatigue syndrome induced by major small intestinal resection (MSIR) in aged rat. Aged rats with MSIR were administrated with ginsenoside Rb1 (15 mg/kg) once a day from 3 days before surgery to the day of sacrifice, or with saline as corresponding controls. Rats without MSIR but going through the same surgery procedure were administrated with saline as blank controls. Anti-fatigue effect was assessed by an open field test; superoxide dismutase, reactive oxygen species and malondialdehyde in skeletal muscle were determined. The mRNA levels of Akt2 and Nrf2 in skeletal muscle were measured by real-time quantitative PCR. The activation of Akt and Nrf2 was examined by western blot and immunohistofluorescence. Our results revealed that ginsenoside Rb1 significantly increased the journey and the rearing frequency, decreased the time of rest in aged rats with MSIR. In addition, ginsenoside Rb1 significantly reduced reactive oxygen species and malondialdehyde release and increased the superoxide dismutase activity of skeletal muscle in aged rats with MSIR. Ginsenoside Rb1 also increased the expression of Akt2 and Nrf2 mRNA, up-regulated Akt phosphorylation and Nrf2 nuclear translocation. These findings indicate that ginsenoside Rb1 has an anti-fatigue effect on postoperative fatigue syndrome in aged rat, and the mechanism possibly involves activation of the PI3K/Akt pathway with subsequent Nrf2 nuclear translocation and induction of antioxidant enzymes. Copyright © 2014 Elsevier B.V. All rights reserved.

Regulation of hemeoxygenase-1 gene expression by Nrf2 and c-Jun in tertiary butylhydroquinone-stimulated rat primary astrocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Jin-Sun; Kim, Hee-Sun, E-mail: hskimp@ewha.ac.kr

2014-05-16

Highlights: • tBHQ increased HO-1 mRNA and protein levels in rat primary astrocytes. • tBHQ enhanced HO-1 gene transcription in an ARE-dependent manner. • tBHQ increased the nuclear translocation and DNA binding of Nrf2 and c-Jun to ARE. • Nrf2 and c-Jun are involved in the differential modulation of HO-1 expression. • Nrf2 and c-Jun regulate HO-1 expression via their coordinated interaction. - Abstract: Hemeoxygenase-1 (HO-1) is a phase II antioxidant enzyme that is primarily involved in detoxification and cytoprotection in a variety of tissues. However, the mechanism underlying HO-1 gene expression remains unclear. In the present study, we investigatedmore » the regulation of HO-1 expression in primary cultured astrocytes by using the natural antioxidant compound tertiary butylhydroquinone (tBHQ). We found that tBHQ increased HO-1 mRNA and protein levels. Promoter analysis revealed that tBHQ enhanced HO-1 gene transcription in an antioxidant response element (ARE)-dependent manner. In addition, tBHQ increased the nuclear translocation and DNA binding of Nrf2 and c-Jun to ARE. Small interfering RNA (siRNA) experiments demonstrated that Nrf2 and c-Jun are involved in the differential modulation of HO-1 expression. Thus, Nrf2 knockdown reduced the basal level of HO-1 expression but did not affect the fold induction by tBHQ. On the other hand, knockdown of c-Jun diminished tBHQ-mediated induction of HO-1 without affecting basal expression. The data suggest that Nrf2 generally modulates the basal expression of HO-1, while c-Jun mediates HO-1 induction in response to tBHQ. The results of co-immunoprecipitation assays demonstrated a physical interaction between Nrf2 and c-Jun in tBHQ-treated astrocytes. The results suggest that Nrf2 and c-Jun regulate HO-1 expression via their coordinated interaction in tBHQ-treated rat primary astrocytes.« less

Non-covalent Small-Molecule Kelch-like ECH-Associated Protein 1-Nuclear Factor Erythroid 2-Related Factor 2 (Keap1-Nrf2) Inhibitors and Their Potential for Targeting Central Nervous System Diseases.

PubMed

Pallesen, Jakob S; Tran, Kim T; Bach, Anders

2018-05-29

The transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) has a protective effect against oxidative stress and plays a major role in inflammation and central nervous system (CNS) diseases. Inhibition of the protein-protein interaction (PPI) between Nrf2 and its repressor protein, Kelch-like ECH-associated protein 1 (Keap1), leads to translocation of Nrf2 from the cytosol to the nucleus and expression of detoxifying antioxidant enzymes. To date, several non-covalent small-molecule Keap1-Nrf2 inhibitors have been identified; however, many of them contain carboxylic acids and are rather large in size, which likely prevents or decreases CNS permeability. This Perspective describes current small-molecule Keap1-Nrf2 inhibitors with experimental evidence for the ability to inhibit the Keap1-Nrf2 interaction by binding to Keap1 in a non-covalent manner. Binding data, biostructural studies, and biological activity are summarized for the inhibitors, and their potential as CNS tool compounds is discussed by analyzing physicochemical properties, including CNS multiparameter optimization (MPO) scoring algorithms. Finally, several strategies for identifying CNS-targeting Keap1 inhibitors are described.

Erythropoietin and Nrf2: key factors in the neuroprotection provided by apo-lactoferrin.

PubMed

Zakharova, E T; Sokolov, A V; Pavlichenko, N N; Kostevich, V A; Abdurasulova, I N; Chechushkov, A V; Voynova, I V; Elizarova, A Yu; Kolmakov, N N; Bass, M G; Semak, I V; Budevich, A I; Kozhin, P M; Zenkov, N K; Klimenko, V M; Kirik, O V; Korzhevskii, D E; Menshchikova, E B; Vasilyev, V B

2018-05-10

Among the properties of lactoferrin (LF) are bactericidal, antianemic, immunomodulatory, antitumour, antiphlogistic effects. Previously we demonstrated its capacity to stabilize in vivo HIF-1-alpha and HIF-2-alpha, which are redox-sensitive multiaimed transcription factors. Various tissues of animals receiving recombinant human LF (rhLF) responded by expressing the HIF-1-alpha target genes, hence such proteins as erythropoietin (EPO), ceruloplasmin, etc. were synthesized in noticeable amounts. Among organs in which EPO synthesis occurred were brain, heart, spleen, liver, kidneys and lungs. Other researchers showed that EPO can act as a protectant against severe brain injury and status epilepticus in rats. Therefore, we tried rhLF as a protector against the severe neurologic disorders developed in rats, such as the rotenone-induced model of Parkinson's disease and experimental autoimmune encephalomyelitis as a model of multiple sclerosis, and observed its capacity to mitigate the grave symptoms. Moreover, an intraperitoneal injection of rhLF into mice 1 h after occlusion of the medial cerebral artery significantly diminished the necrosis area measured on the third day in the ischaemic brain. During this period EPO was synthesized in various murine tissues. It was known that EPO induces nuclear translocation of Nrf2, which, like HIF-1-alpha, is a transcription factor. In view that under conditions of hypoxia both factors demonstrate a synergistic protective effect, we suggested that LF activates the Keap1/Nrf2 signaling pathway, an important link in proliferation and differentiation of normal and malignant cells. J774 macrophages were cultured for 3 days without or in the presence of ferric and ferrous ions (RPMI-1640 and DMEM/F12, respectively). Then cells were incubated with rhLF or Deferiprone. Confocal microscopy revealed nuclear translocation of Nrf2 (the key event in Keap1/Nrf2 signaling) induced by apo-rhLF (iron-free, RPMI-1640). The reference compound Deferiprone (iron chelator) had the similar effect. Upon iron binding (in DMEM/F12) rhLF did not activate the Keap1/Nrf2 pathway. Added to J774, apo-rhLF enhanced transcription of Nrf2-dependent genes coding for glutathione S-transferase P and heme oxygenase-1. Western blotting revealed presence of Nrf2 in mice brain after 6 days of oral administration of apo-rhLF, but not Fe-rhLF or equivalent amount of PBS. Hence, apo-LF, but not holo-LF, induces the translocation of Nrf2 from cytoplasm to the nucleus, probably due to its capacity to induce EPO synthesis.

Friedelane-type triterpenoids as selective anti-inflammatory agents by regulation of differential signaling pathways in LPS-stimulated macrophages

DOE Office of Scientific and Technical Information (OSTI.GOV)

Villar-Lorenzo, Andrea, E-mail: avillar@iib.uam.es

A series of 31 pentacyclic triterpenoids isolated from the root barks of Celastrus vulcanicola and Maytenus jelskii were tested for cytotoxicity and inhibitory activity against lipopolysaccharide (LPS)-induced nitric oxide (NO) production in RAW 264.7 macrophages. Compounds 18 (C18) and 25 (C25) exhibited significant inhibition of LPS-induced NO release at 50 and 25 μM concentrations, respectively, and decreased mRNAs of pro-inflammatory cytokines. At the molecular level, C18 neither inhibited LPS-mediated phosphorylation of mitogen activated protein kinases (MAPKs) nor nuclear translocation of nuclear factor kappa beta (NFκB). Instead, C18 enhanced and prolonged nuclear translocation of nuclear factor-erythroid 2-related factor 2 (Nrf2) andmore » increased the expression of its target genes including hemeoxigenase 1 (HO1). C25 efficiently inhibited LPS-mediated phosphorylation of JNK, p38 and ERK, without affecting NFκB or Nrf2 signaling pathways. Both compounds reduced LPS-mediated processing of caspase-1 and the cleavage of interleukin 1β (IL1β) proform, reflecting their ability to target the inflammasome. C25 also counteracted LPS effects on iNOS expression and pro-inflammatory cytokines mRNA levels in Bv-2 microglial cells. The anti-inflammatory effect of both compounds was also assessed in human macrophages. Our results suggest that triterpenoids C18 and C25 possess anti-inflammatory effects, which may be therapeutically relevant for diseases linked to inflammation. - Highlights: • Compounds 18 (C18) and 25 (C25) exert anti-inflammatory effects in macrophages. • C18 enhanced nuclear translocation of Nrf2 and increased HO1 expression. • C25 inhibited the phosphorylation of JNK, p38 and ERK, members of the MAPKs family. • C25 reduced LPS-mediated processing of caspase-1 and the cleavage of interleukin 1β. • C18 and C25 may be therapeutic agents for diseases linked to inflammation.« less

Co-regulation of nuclear respiratory factor-1 by NFκB and CREB links LPS-induced inflammation to mitochondrial biogenesis

PubMed Central

Suliman, Hagir B.; Sweeney, Timothy E.; Withers, Crystal M.; Piantadosi, Claude A.

2010-01-01

The nuclear respiratory factor-1 (NRF1) gene is activated by lipopolysaccharide (LPS), which might reflect TLR4-mediated mitigation of cellular inflammatory damage via initiation of mitochondrial biogenesis. To test this hypothesis, we examined NRF1 promoter regulation by NFκB, and identified interspecies-conserved κB-responsive promoter and intronic elements in the NRF1 locus. In mice, activation of Nrf1 and its downstream target, Tfam, by Escherichia coli was contingent on NFκB, and in LPS-treated hepatocytes, NFκB served as an NRF1 enhancer element in conjunction with NFκB promoter binding. Unexpectedly, optimal NRF1 promoter activity after LPS also required binding by the energy-state-dependent transcription factor CREB. EMSA and ChIP assays confirmed p65 and CREB binding to the NRF1 promoter and p65 binding to intron 1. Functionality for both transcription factors was validated by gene-knockdown studies. LPS regulation of NRF1 led to mtDNA-encoded gene expression and expansion of mtDNA copy number. In cells expressing plasmid constructs containing the NRF-1 promoter and GFP, LPS-dependent reporter activity was abolished by cis-acting κB-element mutations, and nuclear accumulation of NFκB and CREB demonstrated dependence on mitochondrial H2O2. These findings indicate that TLR4-dependent NFκB and CREB activation co-regulate the NRF1 promoter with NFκB intronic enhancement and redox-regulated nuclear translocation, leading to downstream target-gene expression, and identify NRF-1 as an early-phase component of the host antibacterial defenses. PMID:20587593

EGCG evokes Nrf2 nuclear translocation and dampens PTP1B expression to ameliorate metabolic misalignment under insulin resistance condition.

PubMed

Mi, Yashi; Zhang, Wentong; Tian, Haoyu; Li, Runnan; Huang, Shuxian; Li, Xingyu; Qi, Guoyuan; Liu, Xuebo

2018-03-01

As a major nutraceutical component of green tea (-)-epigallocatechin-3-gallate (EGCG) has attracted interest from scientists due to its well-documented antioxidant and antiobesity bioactivities. In the current study, we aimed to investigate the protective effect of EGCG on metabolic misalignment and in balancing the redox status in mice liver and HepG2 cells under insulin resistance condition. Our results indicated that EGCG accelerates the glucose uptake and evokes IRS-1/Akt/GLUT2 signaling pathway via dampening the expression of protein tyrosine phosphatase 1B (PTP1B). Consistently, ectopic expression of PTP1B by Ad-PTP1B substantially impaired EGCG-elicited IRS-1/Akt/GLUT2 signaling pathway. Moreover, EGCG co-treatment stimulated nuclear translocation of Nrf2 by provoking P13K/AKT signaling pathway and thus modulated the downstream expressions of antioxidant enzymes such as HO-1 and NQO-1 in HepG2 cells. Furthermore, knockdown Nrf2 by small interfering RNA (siRNA) notably enhanced the expression of PTP1B and blunt EGCG-stimulated glucose uptake. Consistent with these results, in vivo study revealed that EGCG supplement significantly ameliorated high-fat and high-fructose diet (HFFD)-triggered insulin resistance and oxidative stress by up-regulating the IRS-1/AKT and Keap1/Nrf2 transcriptional pathways. Administration of an appropriate chemopreventive agent, such as EGCG, could potentially serve as an additional therapeutic intervention in the arsenal against obesity.

Hyperoside attenuates hydrogen peroxide-induced L02 cell damage via MAPK-dependent Keap{sub 1}-Nrf{sub 2}-ARE signaling pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xing, Hai-Yan; Liu, Yao; Chen, Jian-Hong

Highlights: {yields} Hyperoside attenuated H{sub 2}O{sub 2}-induced L02 cell damage. {yields} Hyperoside up-regulated HO-1 expression at both mRNA and protein levels. {yields} Hyperoside activated both Nrf{sub 2} nuclear translocation and gene expression. {yields} Hyperoside may inhibit Keap{sub 1} mRNA translation or protein degradation. {yields} Phosphorylation of ERK and p38 is involved in hyperoside-mediated Nrf{sub 2} activation. -- Abstract: The flavonoid hyperoside has been reported to elicit cytoprotection against oxidative stress partly by increasing the activity of antioxidant enzymes, such as glutathione peroxidase, superoxide dismutase and catalase. However, the cellular and molecular mechanisms underlying this effect remain unclear. Here, hepatic L02more » cells exposed to H{sub 2}O{sub 2} (100 {mu}M) were used to demonstrate that hyperoside protected cells by significantly inhibiting overproduction of intracellular ROS, depletion of the mitochondrial membrane potential and leakage of lactate dehydrogenase. Hyperoside further enhanced the cellular antioxidant defense system through increasing the activity of heme oxygenase-1 (HO-1), and by up-regulating HO-1 expression. Meanwhile, real time PCR, western blot and immunofluorescence studies revealed that hyperoside stimulated nuclear translocation of the Nrf{sub 2} transcription factor in a dose-dependent manner, and this effect was significantly suppressed by pharmacological inhibition of the mitogen-activated protein kinases (MAPK) p38 and ERK. Collectively, our data provide the first description of the mechanism underlying hyperoside's ability to attenuate H{sub 2}O{sub 2}-induced cell damage, namely this compound interacts with the MAPK-dependent Keap{sub 1}-Nrf{sub 2}-ARE signaling pathway to up-regulate HO-1 expression and enhance intracellular antioxidant activity.« less

Oncogenic mutations in KEAP1 disturbing inhibitory Nrf2-Keap1 interaction: Activation of antioxidative pathway in papillary thyroid carcinoma.

PubMed

Danilovic, Debora Lucia Seguro; de Mello, Evandro Sobroza; Frazzato, Eliana Salgado Turri; Wakamatsu, Alda; de Lima Jorge, Alexander Augusto; Hoff, Ana Oliveira; Marui, Suemi

2018-06-01

Nuclear factor erythroid 2-like 2 (NFE2L2) encodes Nrf2, transcription factor of antioxidative genes. In the presence of reactive oxygen species, Keap1 (Kelch-ECH-associating protein-1) inhibitor complex undergoes conformational changes disrupting Keap1-Nrf2 binding and Nrf2 translocates into nucleus. We evaluated the presence of mutations in NFE2L2 and KEAP1 in papillary thyroid carcinomas (PTCs) and correlated them with clinical presentation. Coding regions of NFE2L2 and KEAP1 were sequenced in 131 patients with PTC. Clinical and histopathological features were analyzed. Immunohistochemical analysis of Nrf2 expression was performed in mutated carcinomas. Although no mutations were found in NFE2L2, missense mutations in KEAP1 were observed in 6 patients with PTC (4.6%). Immunohistochemistry showed increased Nrf2 expression in nuclei of all mutated carcinomas, which presented poor prognostic features in histopathology. We identified mutations in KEAP1 associated with Nrf2 overexpression in PTC. Mutations favored disruption of inhibitory interaction Nrf2-Keap1 to enable increased antioxidant Nrf2 activity, possibly with prognostic consequences. © 2018 Wiley Periodicals, Inc.

Rosmarinic acid counteracts activation of hepatic stellate cells via inhibiting the ROS-dependent MMP-2 activity: Involvement of Nrf2 antioxidant system

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lu, Changfang; Zou, Yu; Liu, Yuzhang

Recently, oxidative stress is involved in hepatofibrogenesis. Matrix metalloproteinase-2 (MMP-2) is required for activation of hepatic stellate cells (HSCs) in response to reactive oxygen species (ROS). This study was designed to explore the hypothesis that the inhibitory effect of rosmarinic acid (RA) on HSCs activation might mainly result from its antioxidant capability by increasing the synthesis of glutathione (GSH) involved in nuclear factor kappa B (NF-κB)-dependent inhibition of MMP-2 activity. Here, we demonstrate that RA reverses activated HSCs to quiescent cells. Concomitantly, RA inhibits MMP-2 activity. RNA interference-imposed knockdown of NF-κB abolished down-regulation of MMP-2 by RA. RA-mediated inactivation ofmore » NF-κB could be blocked by the diphenyleneiodonium chloride (DPI; a ROS inhibitor). Conversely, transfection of dominant-negative (DN) mutant of extracellular signal-regulated kinases 2 (ERK2), c-Jun N-terminal kinase 1 (JNK1), or p38α kinase had no such effect. Simultaneously, RA suppresses ROS generation and lipid peroxidation (LPO) whereas increases cellular GSH in HSC-T6 cells. Furthermore, RA significantly increased antioxidant response element (ARE)-mediated luciferase activity, nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) and catalytic subunits from glutamate cysteine ligase (GCLc) expression, but not modulatory subunits from GCL (GCLm). RA-mediated up-regulation of GClc is inhibited by the shRNA-induced Nrf2 knockdown. The knocking down of Nrf2 or buthionine sulfoximine (a GCL inhibitor) abolished RA-mediated inhibition of ROS. Collectively, these results provide novel insights into the mechanisms of RA as an antifibrogenic candidate in the prevention and treatment of liver fibrosis. - Highlights: • RA reverses activated HSCs to quiescent cells. • RA suppresses MMP-2 activity through a NF-κB-dependent pathway. • Inhibition of oxidative stress by RA is dependent on nuclear translocation of Nrf2. • RA-mediated down-regulation of MMP-2 was ROS-dependent.« less

Iron depletion in HCT116 cells diminishes the upregulatory effect of phenethyl isothiocyanate on heme oxygenase-1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bolloskis, Michael P.; Carvalho, Fabiana P.; Loo, George, E-mail: g_loo@uncg.edu

Some of the health-promoting properties of cruciferous vegetables are thought to be partly attributed to isothiocyanates. These phytochemicals can upregulate the expression of certain cytoprotective stress genes, but it is unknown if a particular nutrient is involved. Herein, the objective was to ascertain if adequate iron is needed for enabling HCT116 cells to optimally express heme oxygenase-1 (HO-1) when induced by phenethyl isothiocyanate (PEITC). PEITC increased HO-1 expression and also nuclear translocation of Nrf2, which is a transcription factor known to activate the HO-1 gene. However, in HCT116 cells that were made iron-deficient by depleting intracellular iron with deferoxamine (DFO),more » PEITC was less able to increase HO-1 expression and nuclear translocation of Nrf2. These suppressive effects of DFO were overcome by replenishing the iron-deficient cells with the missing iron. To elucidate these findings, it was found that PEITC-induced HO-1 upregulation can be inhibited with thiol antioxidants (glutathione and N-acetylcysteine). Furthermore, NADPH oxidase inhibitors (diphenyleneiodonium and apocynin) and a superoxide scavenger (Tiron) each inhibited PEITC-induced HO-1 upregulation. In doing so, diphenyleneiodonium was the most potent and also inhibited nuclear translocation of redox-sensitive Nrf2. Collectively, the results imply that the HO-1 upregulation by PEITC involves an iron-dependent, oxidant signaling pathway. Therefore, it is concluded that ample iron is required to enable PEITC to fully upregulate HO-1 expression in HCT116 cells. As such, it is conceivable that iron-deficient individuals may not reap the full health benefits of eating PEITC-containing cruciferous vegetables that via HO-1 may help protect against multiple chronic diseases. - Highlights: • PEITC increased HO-1 expression in HCT116 cells. • PEITC-induced HO-1 upregulation was impaired in iron-depleted HCT116 cells. • Impairment of PEITC-induced HO-1 upregulation was reversible with iron restoration. • PEITC increased nuclear expression of Nrf2 but not in iron-depleted cells. • NADPH oxidase inhibitors inhibited PEITC-induced HO-1 upregulation.« less

Astaxanthin Induces the Nrf2/HO-1 Antioxidant Pathway in Human Umbilical Vein Endothelial Cells by Generating Trace Amounts of ROS.

PubMed

Niu, Tingting; Xuan, Rongrong; Jiang, Ligang; Wu, Wei; Zhen, Zhanghe; Song, Yuling; Hong, Lili; Zheng, Kaiqin; Zhang, Jiaxing; Xu, Qingshan; Tan, Yinghong; Yan, Xiaojun; Chen, Haimin

2018-02-14

Astaxanthin is a powerful antioxidant that possesses potent protective effects against various human diseases and physiological disorders. However, the mechanisms underlying its antioxidant functions in cells are not fully understood. In the present study, the effects of astaxanthin on reactive oxygen species (ROS) production and antioxidant enzyme activity, as well as mitogen-activated protein kinases (MAPKs), phosphatidylinositol 3-kinase (PI3K)/Akt, and the nuclear factor erythroid 2-related factor 2 (Nrf-2)/heme oxygenase-1 (HO-1) pathways in human umbilical vein endothelial cells (HUVECs), were examined. It was shown that astaxanthin (0.1, 1, and 10 μM) induced ROS production by 9.35%, 14.8%, and 18.06% compared to control, respectively, in HUVECs. In addition, astaxanthin increased the mRNA levels of phase II enzymes HO-1 and also promoted GSH-Px enzyme activity. Furthermore, we observed ERK phosphorylation, nuclear translocation of Nrf-2, and activation of antioxidant response element-driven luciferase activity upon astaxanthin treatment. Knockdown of Nrf-2 by small interfering RNA inhibited HO-1 mRNA expression by 60%, indicating that the Nrf-2/ARE signaling pathway is activated by astaxanthin. Our results suggest that astaxanthin activates the Nrf-2/HO-1 antioxidant pathway by generating small amounts of ROS.

NRF2-regulation in brain health and disease: implication of cerebral inflammation

PubMed Central

Sandberg, Mats; Patil, Jaspal; D’Angelo, Barbara; Weber, Stephen G; Mallard, Carina

2014-01-01

The nuclear factor erythroid 2 related factor 2 (NRF2) is a key regulator of endogenous inducible defense systems in the body. Under physiological conditions NRF2 is mainly located in the cytoplasm. However, in response to oxidative stress, NRF2 translocates to the nucleus and binds to specific DNA sites termed “anti-oxidant response elements” or “electrophile response elements” to initiate transcription of cytoprotective genes. Acute oxidative stress to the brain, such as stroke and traumatic brain injury is increased in animals that are deficient in NRF2. Insufficient NRF2 activation in humans has been linked to chronic diseases such as Parkinson’s disease, Alzheimer’s disease and amyotrophic lateral sclerosis. New findings have also linked activation of the NRF2 system to anti-inflammatory effects via interactions with NF-κB. Here we review literature on cellular mechanisms of NRF2 regulation, how to maintain and restore NRF2 function and the relationship between NRF2 regulation and brain damage. We bring forward the hypothesis that inflammation via prolonged activation of key kinases (p38 and GSK-3β) and activation of histone deacetylases gives rise to dysregulation of the NRF2 system in the brain, which contributes to oxidative stress and injury. PMID:24262633

Dibenzoylmethane Protects Against CCl4-Induced Acute Liver Injury by Activating Nrf2 via JNK, AMPK, and Calcium Signaling.

PubMed

Cao, Mingnan; Wang, Huixia; Guo, Limei; Yang, Simin; Liu, Chun; Khor, Tin Oo; Yu, Siwang; Kong, Ah-Ng

2017-11-01

Oxidative stress is an important pathogenic factor in various hepatic diseases. Nuclear factor-erythroid 2-related factor-2 (Nrf2), which coordinates the expression of an array of antioxidant and detoxifying genes, has been proposed as a potential target for prevention and treatment of liver disease. Dibenzoylmethane (DBM) is a minor ingredient in licorice that activates Nrf2 and prevents various cancers and oxidative damage. In the present study, the mechanisms by which DBM activates Nrf2 signaling were delineated, and its protective effect against carbon tetrachloride (CCl 4 )-induced liver injury was examined. DBM potently induced the expression of HO-1 in cells and in the livers of mice, but this induction was diminished in Nrf2-deficient mice and cells. Overexpression of Nrf2 enhanced DBM-induced HO-1 expression, while overexpression of a dominant-negative fragment of Nrf2 inhibited this induction. DBM treatment resulted in dissociation from Keap1 and nuclear translocation of Nrf2. Moreover, DBM activated Akt/protein kinase B, mitogen-activated protein kinases, and AMP-activated protein kinase and increased intracellular calcium levels. Inhibition of JNK, AMPK, or intracellular calcium signaling significantly suppressed the induction of HO-1 expression by DBM. Finally, DBM treatment significantly inhibited CCl 4 -induced acute liver injury in wild-type but not in Nrf2-deficient mice. Taken together, our results revealed the mechanisms by which DBM activates Nrf2 and induces HO-1 expression, and provide molecular basis for the design and development of DBM and its derivatives for prevention or treatment of liver diseases by targeting Nrf2.

Mechanism of phytoestrogen puerarin-mediated cytoprotection following oxidative injury: Estrogen receptor-dependent up-regulation of PI3K/Akt and HO-1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hwang, Yong Pil; Jeong, Hye Gwang

2008-12-15

Phytoestrogens are polyphenolic non-steroidal plant compounds with estrogen-like biological activity. The phytoestrogen puerarin, the main isoflavone glycoside found in the root of Pueraria lobata, has been used for various medicinal purposes in traditional Chinese medicines for thousands of years. Recent studies have indicated that the estrogen receptor (ER), through interaction with p85, regulates phosphoinositide 3-kinase (PI3K) activity, revealing a physiologic, non-nuclear function of ER that may be relevant in cytoprotection. In this study, we demonstrate that the phytoestrogen puerarin inhibits tert-butyl hydroperoxide (t-BHP)-induced oxidative injury via an ER-dependent G{beta}1/PI3K/Akt and heme oxygenase-1 (HO-1) pathway. Pretreatment of Hepa1c1c7 and HepG2 cellsmore » with puerarin significantly reduced t-BHP-induced caspase-3 activation and subsequent cell death. Also, puerarin up-regulated HO-1 expression and this expression conferred cytoprotection against oxidative injury induced by t-BHP. Moreover, puerarin induced Nrf2 nuclear translocation, which is upstream of puerarin-induced HO-1 expression, and PI3K activation, a pathway that is involved in induced Nrf2 nuclear translocation, HO-1 expression and cytoprotection. Puerarin-induced up-regulation of HO-1 and cytoprotection against t-BHP were abolished by silencing Nrf2 expression with specific siRNA. Also, puerarin-mediated increases in PI3K activation and HO-1 induction were reversed by co-treatment with ICI 182,780 and pertussis toxin. Taken together, these results suggest that puerarin augments cellular antioxidant defense capacity through ER-dependent HO-1 induction via the G{beta}1/PI3K/Akt-Nrf2 signaling pathway, thereby protecting cells from oxidative stress.« less

Involvement of the p62/NRF2 signal transduction pathway on erythrophagocytosis.

PubMed

Santarino, Inês B; Viegas, Michelle S; Domingues, Neuza S; Ribeiro, Ana M; Soares, Miguel P; Vieira, Otília V

2017-07-19

Erythrophagocytosis, the phagocytic removal of damaged red blood cells (RBC), and subsequent phagolysosome biogenesis are important processes in iron/heme metabolism and homeostasis. Phagolysosome biogenesis implies the interaction of nascent phagosomes with endocytic compartments and also autophagy effectors. Here, we report that besides recruitment of microtubule-associated protein-1-light chain 3 (LC3), additional autophagy machinery such as sequestosome 1 (p62) is also acquired by single-membrane phagosomes at very early stages of the phagocytic process and that its acquisition is very important to the outcome of the process. In bone marrow-derived macrophages (BMDM) silenced for p62, RBC degradation is inhibited. P62, is also required for nuclear translocation and activation of the transcription factor Nuclear factor E2-related Factor 2 (NRF2) during erythrophagocytosis. Deletion of the Nrf2 allele reduces p62 expression and compromises RBC degradation. In conclusion, we reveal that erythrophagocytosis relies on an interplay between p62 and NRF2, potentially acting as protective mechanism to maintain reactive oxygen species at basal levels and preserve macrophage homeostasis.

Nrf2 Activation Induced by Sirt1 Ameliorates Acute Lung Injury After Intestinal Ischemia/Reperfusion Through NOX4-Mediated Gene Regulation.

PubMed

Chai, DongDong; Zhang, Lei; Xi, SiWei; Cheng, YanYong; Jiang, Hong; Hu, Rong

2018-01-01

Nuclear erythroid 2-related factor-2 (Nrf2) is a major stress-response transcription factor that has been implicated in regulating ischemic angiogenesis. We investigated the effects of Nrf2 in regulating revascularization and modulating acute lung injury. The expression of Nrf2 and sirtuin1 (Sirt1) was assessed in lung tissue by western blotting and immunofluorescence staining after intestinal ischemia/reperfusion (IIR) in Nrf2-/- and wild-type (WT) mice. The involvement of Nrf2 in angiogenesis, cell viability, and migration was investigated in human pulmonary microvascular endothelial cells (PMVECs). Additionally, the influence of Nrf2 expression on NOX pathway activation was measured in PMVECs after oxygen-glucose deprivation/reoxygenation. We found activation and nuclear accumulation of Nrf2 in lung tissue after IIR. Compared to IIR in WT mice, IIR in Nrf2-/- mice significantly enhanced leukocyte infiltration and collagen deposit, and inhibited endothelial cell marker CD31 expression. Nrf2 upregulation and translocation into the nucleus stimulated by Sirt1 overexpression exhibited remission of histopathologic changes and enhanced CD31 expression. Nrf2 knockdown repressed non-phagocytic cell oxidase 4 (NOX4), hypoxia-inducible factor (HIF-1α) and vascular endothelial growth factor (VEGF) expression after IIR. Nrf2 upregulation by Sirt1 enhances NOX4, HIF-1α and VEGF expression after IIR in WT mice. Furthermore, Nrf2 knockdown suppressed cell viability, capillary tube formation and cell migration in PMVECs after oxygen-glucose deprivation/reoxygenation and also inhibited NOX4, HIF-1 and VEGF expression. Moreover, NOX4 knockdown in PMVECs decreased the levels of VEGF, HIF-1α and angiogenesis. Nrf2 stimulation by Sirt1 plays an important role in sustaining angiogenic potential through NOX4-mediated gene regulation. © 2018 The Author(s). Published by S. Karger AG, Basel.

Intracellular degradation of functionalized carbon nanotube/iron oxide hybrids is modulated by iron via Nrf2 pathway

PubMed Central

Elgrabli, Dan; Dachraoui, Walid; Marmier, Hélène de; Ménard-Moyon, Cécilia; Bégin, Dominique; Bégin-Colin, Sylvie; Bianco, Alberto; Alloyeau, Damien; Gazeau, Florence

2017-01-01

The in vivo fate and biodegradability of carbon nanotubes is still a matter of debate despite tremendous applications. In this paper we describe a molecular pathway by which macrophages degrade functionalized multi-walled carbon nanotubes (CNTs) designed for biomedical applications and containing, or not, iron oxide nanoparticles in their inner cavity. Electron microscopy and Raman spectroscopy show that intracellularly-induced structural damages appear more rapidly for iron-free CNTs in comparison to iron-loaded ones, suggesting a role of iron in the degradation mechanism. By comparing the molecular responses of macrophages derived from THP1 monocytes to both types of CNTs, we highlight a molecular mechanism regulated by Nrf2/Bach1 signaling pathways to induce CNT degradation via NOX2 complex activation and O2•−, H2O2 and OH• production. CNT exposure activates an oxidative stress-dependent production of iron via Nrf2 nuclear translocation, Ferritin H and Heme oxygenase 1 translation. Conversely, Bach1 was translocated to the nucleus of cells exposed to iron-loaded CNTs to recycle embedded iron. Our results provide new information on the role of oxidative stress, iron metabolism and Nrf2-mediated host defence for regulating CNT fate in macrophages. PMID:28120861

EX4 stabilizes and activates Nrf2 via PKCδ, contributing to the prevention of oxidative stress-induced pancreatic beta cell damage

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Mi-Hwi; Kim, Eung-Hwi

Oxidative stress in pancreatic beta cells can inhibit insulin secretion and promote apoptotic cell death. Exendin-4 (EX4), a glucagon-like peptide-1 receptor agonist, can suppress beta cell apoptosis, improve beta cell function and protect against oxidative damage. In this study, we investigated the molecular mechanisms for antioxidative effects of EX4 in pancreatic beta cells. INS-1 cells, a rat insulinoma cell line, were pretreated with EX4 and exposed to palmitate or H{sub 2}O{sub 2}. Reactive oxygen species (ROS) production, and glutathione and insulin secretion were measured. The mRNA and protein expression levels of antioxidant genes were examined. The level of nuclear factormore » erythroid 2-related factor 2 (Nrf2), its binding to antioxidant response element (ARE), and its ubiquination in the presence of EX4 were determined. The Nrf2 signaling pathway was determined using rottlerin (protein kinase [PK]Cδ inhibitor), H89 (PKA inhibitor) and LY294002 (phosphatidylinositide 3-kinase [PI3K] inhibitor). EX4 treatment decreased ROS production, recovered cellular glutathione levels and insulin secretion in the presence of oxidative stress in INS-1 cells. The expression levels of glutamate-cysteine ligase catalytic subunit and heme oxygenase-1 were increased by EX4 treatment. EX4 promoted Nrf2 translocation, ARE binding activity and enhanced stabilization of Nrf2 by inhibition of ubiquitination. Knockdown of Nrf2 abolished the effect of EX4 on increased insulin secretion. Inhibition of PKCδ attenuated Nrf2 translocation and antioxidative gene expression by EX4 treatment. We suggest that EX4 activates and stabilizes Nrf2 through PKCδ activation, contributing to the increase of antioxidant gene expression and consequently improving beta cell function in the presence of oxidative stress. - Highlights: • EX4 protects against oxidative stress-induced pancreatic beta cell dysfunction. • EX4 increases antioxidant gene expression. • Antioxidative effect of EX4 is mediated by Nrf2. • EX4 increases Nrf2 level by stabilizing Nrf2 protein. • EX4 stabilizes Nrf2 by activation of PKCδ.« less

Activation of Nrf2 contributes to the protective effect of Exendin-4 against angiotensin II-induced vascular smooth muscle cell senescence.

PubMed

Zhou, Tengfei; Zhang, Mengqian; Zhao, Liang; Li, Aiqin; Qin, Xiaomei

2016-10-01

Oxidative stress and impaired antioxidant defense are believed to be contributors to the cardiovascular aging process. The transcription factor nuclear factor-E2-related factor 2 (Nrf2) plays a key role in orchestrating cellular antioxidant defenses and maintaining redox homeostasis. Our previous study showed that Exendin-4, a glucagon-like peptide-1 analog, alleviates angiotensin II (ANG II)-induced vascular smooth muscle cell (VSMC) senescence by inhibiting Rac1 activation via cAMP/PKA (Zhao L, Li AQ, Zhou TF, Zhang MQ, Qin XM. Am J Physiol Cell Physiol 307: C1130-C1141, 2014). The objective of this study is to investigate if Nrf2 mediates the antisenescent effect of Exendin-4 in ANG II-induced VSMCs. Here we report that Exendin-4 triggered Nrf2 nuclear translocation, a downstream target of cAMP-responsive element-binding protein (CREB) and expressions of antioxidant genes heme oxygenase-1 (HO-1) and NAD(P)H quinone oxidoreductase-1 (NQO-1) in a dose- and time-dependent manner. In addition, knock-down of Nrf2 attenuated the inhibitory effects of Exendin-4 on ANG II-induced superoxidant generation and VSMC senescence. PKA/CREB pathway participated in the upregulations of HO-1 and NQO-1 induced by Exendin-4. Notably, our study revealed that Exendin-4 dose-dependently increased the acetylation of Nrf2 and the recruitment of transcriptional coactivator CREB binding protein (CBP) to Nrf2. The Exendin-4-induced Nrf2 transactivation was diminished in the presence of CBP small interfering RNA. Microscope imaging of Nrf2, as well as immunoblotting for Nrf2, showed that the Exendin-4-evoked Nrf2 acetylation favored its nuclear retention. Importantly, CBP silencing attenuated the suppressing effects of Exendin-4 on ANG II-induced VSMC senescence and superoxidant production. In conclusion, these results provide a mechanistic insight into how Nrf2 signaling mediates the antisenescent and antioxidative effects induced by Exendin-4 in VSMCs. Copyright © 2016 the American Physiological Society.

Baicalin Ameliorates Liver Injury Induced by Chronic plus Binge Ethanol Feeding by Modulating Oxidative Stress and Inflammation via CYP2E1 and NRF2 in Mice

PubMed Central

He, Ping; Wu, Yafeng; Shun, Jianchao; Liang, Yaodong; Cheng, Mingliang

2017-01-01

Alcoholic liver injury leads to serious complication including death. The potential role of baicalin at the transcription level in mice model of alcohol injury is not known yet. In this study, we examined the effect of baicalin against chronic plus binge ethanol model in mice and understanding the mechanism of protection. Liver function, histology, steatosis, inflammation, NF-κB activity, oxidative stress sources, nuclear translocation of NRF2 transcription factor, and cell death were assessed. Treatment with baicalin ameliorated ethanol-induced oxidative stress, inflammation, and cell death. Baicalin attenuated ethanol-induced proinflammatory molecules such as TNF-α, IL-1β, MIP-2, and MCP-1 and reversed redox-sensitive transcription factor NF-κB activation. Baicalin also modulated Kupffer cell activation in vitro. Baicalin inhibited ethanol-induced expression of reactive oxygen species (ROS) generating enzymes NOX2, p67phox, xanthine oxidase, and iNOS in addition to CYP2E1 activities. Baicalin also enhanced ethanol-induced NRF2 nuclear translocation and increased downstream target gene HO-1 as antioxidant defense. Finally, baicalin reduced significant apoptotic and necrotic cell death. Our study suggests that baicalin ameliorates chronic plus binge ethanol-induced liver injury involving molecular crosstalk of multiple pathways at the transcriptional level and through upregulation of antioxidant defense mechanism. PMID:28951767

The role of Nrf2 transcription factor in viral infection.

PubMed

Ramezani, Ali; Nahad, Mehdi Parsa; Faghihloo, Ebrahim

2018-05-08

The nuclear factor erythroid 2 related factor 2 (Nrf2) is a major regulator of intracellular inducible defense systems against harmful endogenous and exogenous substances in the body. Under normal conditions Nrf2 is mainly binds to keap1 and located in the cytoplasm. However, in response to oxidative and electrophile stress, Nrf2 translocated to the nucleus and link to anti-oxidant response elements to induce the transcription of cytoprotective genes. Most viruses cause oxidative stress and increase the activity of radicals and reactive oxygen species (ROS), subsequently, the cellular protection system activates the Nrf2 and increases the expression of cytoprotective genes. However, in some cases, the activation of Nrf2 is not ROS-dependent, and is carried out directly via the ROS-independent pathway. Many viruses cause the activation of Nrf2, which is involved in the pathogenesis and the progression of the virus infection and even in its chronic form. However, some viruses inhibit the activation of Nrf2, in which case the virus also benefits of this mechanism to maintain the homeostasis of the cell. However, the challenge between the Nrf2/ARE signaling pathway of and viral infections is unknown in some cases, and in order to know more details in this regard, a more detailed seems necessary. © 2018 Wiley Periodicals, Inc.

MHY1485 ameliorates UV-induced skin cell damages via activating mTOR-Nrf2 signaling.

PubMed

Yang, Bo; Xu, Qiu-Yun; Guo, Chun-Yan; Huang, Jin-Wen; Wang, Shu-Mei; Li, Yong-Mei; Tu, Ying; He, Li; Bi, Zhi-Gang; Ji, Chao; Cheng, Bo

2017-02-21

Ultra Violet (UV)-caused skin cell damage is a main cause of skin cancer. Here, we studied the activity of MHY1485, a mTOR activator, in UV-treated skin cells. In primary human skin keratinocytes, HaCaT keratinocytes and human skin fibroblasts, MHY1485 ameliorated UV-induced cell death and apoptosis. mTOR activation is required for MHY1485-induced above cytoprotective actions. mTOR kinase inhibitors (OSI-027, AZD-8055 and AZD-2014) or mTOR shRNA knockdown almost abolished MHY1485-induced cytoprotection. Further, MHY1485 treatment in skin cells activated mTOR downstream NF-E2-related factor 2 (Nrf2) signaling, causing Nrf2 Ser-40 phosphorylation, stabilization/upregulation and nuclear translocation, as well as mRNA expression of Nrf2-dictated genes. Contrarily, Nrf2 knockdown or S40T mutation almost nullified MHY1485-induced cytoprotection. MHY1485 suppressed UV-induced reactive oxygen species production and DNA single strand breaks in skin keratinocytes and fibroblasts. Together, we conclude that MHY1485 inhibits UV-induced skin cell damages via activating mTOR-Nrf2 signaling.

Induction of the pi class of glutathione S-transferase by carnosic acid in rat Clone 9 cells via the p38/Nrf2 pathway.

PubMed

Lin, Chia-Yuan; Wu, Chi-Rei; Chang, Shu-Wei; Wang, Yu-Jung; Wu, Jia-Jiuan; Tsai, Chia-Wen

2015-06-01

Induction of phase II enzymes is important in cancer chemoprevention. We compared the effect of rosemary diterpenes on the expression of the pi class of glutathione S-transferase (GSTP) in rat liver Clone 9 cells and the signaling pathways involved. Culturing cells with 1, 5, 10, or 20 μM carnosic acid (CA) or carnosol (CS) for 24 h in a dose-dependent manner increased the GSTP expression. CA was more potent than CS. The RNA level and the enzyme activity of GSTP were also enhanced by CA treatment. Treatment with 10 μM CA highly induced the reporter activity of the enhancer element GPEI. Furthermore, CA markedly increased the translocation of nuclear factor erythroid-2 related factor 2 (Nrf2) from the cytosol to the nucleus after 30 to 60 min. CA the stimulated the protein induction of p38, nuclear Nrf2, and GSTP was diminished in the presence of SB203580 (a p38 inhibitor). In addition, SB203580 pretreatment or silencing of Nrf2 by siRNA suppressed the CA-induced GPEI-DNA binding activity and GSTP protein expression. Knockdown of p38 or Nrf2 by siRNA abolished the activation of p38 and Nrf2 as well as the protein induction and enzyme activity of GSTP by CA. These results suggest that CA up-regulates the expression and enzyme activity of GSTP via the p38/Nrf2/GPEI pathway.

Characterization of the Antioxidant Effects of γ-Oryzanol: Involvement of the Nrf2 Pathway

PubMed Central

Rungratanawanich, W.; Serafini, M. M.; Guarienti, M.; Catanzaro, M.; Marziano, M.; Memo, M.; Lanni, C.

2018-01-01

γ-Oryzanol (ORY) is well known for its antioxidant potential. However, the mechanism by which ORY exerts its antioxidant effect is still unclear. In this paper, the antioxidant properties of ORY were investigated for its potential effects as a reactive oxygen and nitrogen species (ROS/RNS) scavenger and in activating antioxidant-promoting intracellular pathways utilizing the human embryonic kidney cells (HEK-293). The 24 h ORY exposure significantly prevented hydrogen peroxide- (H2O2-) induced ROS/RNS production at 3 h, and this effect was sustained for at least 24 h. ORY pretreatment also enhanced the activity of antioxidant enzymes: superoxide dismutase (SOD) and glutathione peroxidase (GPX). Interestingly, ORY induced the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) nuclear translocation and upregulation of Nrf2-dependent defensive genes such as NAD(P)H quinone reductase (NQO1), heme oxygenase-1 (HO-1), and glutathione synthetase (GSS) at mRNA and protein levels in both basal condition and after H2O2 insult. Thus, this study suggested an intriguing effect of ORY in modulating the Nrf2 pathway, which is also involved in regulating longevity as well as age-related diseases. PMID:29725495

Characterization of the Antioxidant Effects of γ-Oryzanol: Involvement of the Nrf2 Pathway.

PubMed

Rungratanawanich, W; Abate, G; Serafini, M M; Guarienti, M; Catanzaro, M; Marziano, M; Memo, M; Lanni, C; Uberti, D

2018-01-01

γ -Oryzanol (ORY) is well known for its antioxidant potential. However, the mechanism by which ORY exerts its antioxidant effect is still unclear. In this paper, the antioxidant properties of ORY were investigated for its potential effects as a reactive oxygen and nitrogen species (ROS/RNS) scavenger and in activating antioxidant-promoting intracellular pathways utilizing the human embryonic kidney cells (HEK-293). The 24 h ORY exposure significantly prevented hydrogen peroxide- (H 2 O 2 -) induced ROS/RNS production at 3 h, and this effect was sustained for at least 24 h. ORY pretreatment also enhanced the activity of antioxidant enzymes: superoxide dismutase (SOD) and glutathione peroxidase (GPX). Interestingly, ORY induced the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) nuclear translocation and upregulation of Nrf2-dependent defensive genes such as NAD(P)H quinone reductase (NQO1), heme oxygenase-1 (HO-1), and glutathione synthetase (GSS) at mRNA and protein levels in both basal condition and after H 2 O 2 insult. Thus, this study suggested an intriguing effect of ORY in modulating the Nrf2 pathway, which is also involved in regulating longevity as well as age-related diseases.

Ginsenoside Rb1 protects against 6-hydroxydopamine-induced oxidative stress by increasing heme oxygenase-1 expression through an estrogen receptor-related PI3K/Akt/Nrf2-dependent pathway in human dopaminergic cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hwang, Yong Pil; College of Pharmacy, Chosun University, Gwangju; Jeong, Hye Gwang, E-mail: hgjeong@cnu.ac.k

Phytoestrogens are polyphenolic non-steroidal plant compounds with estrogen-like biological activity. Ginseng, the root of Panax ginseng C.A. Meyer (Araliaceae), is a popular traditional herbal medicine. Ginsenoside Rb1 (Rb1), an active component commonly found in ginseng root, is a phytoestrogen that exerts estrogen-like activity. In this study, we demonstrate that the phytoestrogen Rb1 inhibits 6-hydroxydopamine (6-OHDA)-induced oxidative injury via an ER-dependent Gbeta1/PI3K/Akt and heme oxygenase-1 (HO-1) pathway. Pretreatment of SH-SY5Y cells with Rb1 significantly reduced 6-OHDA-induced caspase-3 activation and subsequent cell death. Rb1 also up-regulated HO-1 expression, which conferred cytoprotection against 6-OHDA-induced oxidative injury. Moreover, Rb1 induced both Nrf2 nuclear translocation,more » which is upstream of HO-1 expression and PI3K activation, a pathway that is involved in induced Nrf2 nuclear translocation, HO-1 expression and cytoprotection. Also, Rb1-mediated increases in PI3K activation and HO-1 induction were reversed by co-treatment with ICI 182,780 and pertussis toxin. Taken together, these results suggest that Rb1 augments the cellular antioxidant defenses through ER-dependent HO-1 induction via the Gbeta1/PI3K/Akt-Nrf2 signaling pathway, thereby protecting cells from oxidative stress. Thus our study indicates that Rb1 has a partial cytoprotective role in dopaminergic cell culture systems.« less

Sulforaphane protects against cytokine- and streptozotocin-induced {beta}-cell damage by suppressing the NF-{kappa}B pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Song, Mi-Young; Kim, Eun-Kyung; Moon, Woo-Sung

2009-02-15

Sulforaphane (SFN) is an indirect antioxidant that protects animal tissues from chemical or biological insults by stimulating the expression of several NF-E2-related factor-2 (Nrf2)-regulated phase 2 enzymes. Treatment of RINm5F insulinoma cells with SFN increases Nrf2 nuclear translocation and expression of phase 2 enzymes. In this study, we investigated whether the activation of Nrf2 by SFN treatment or ectopic overexpression of Nrf2 inhibited cytokine-induced {beta}-cell damage. Treatment of RIN cells with IL-1{beta} and IFN-{gamma} induced {beta}-cell damage through a NF-{kappa}B-dependent signaling pathway. Activation of Nrf2 by treatment with SFN and induction of Nrf2 overexpression by transfection with Nrf2 prevented cytokinemore » toxicity. The mechanism by which Nrf2 activation inhibited NF-{kappa}B-dependent cell death signals appeared to involve the reduction of oxidative stress, as demonstrated by the inhibition of cytokine-induced H{sub 2}O{sub 2} production. The protective effect of SFN was further demonstrated by the restoration of normal insulin secreting responses to glucose in cytokine-treated rat pancreatic islets. Furthermore, pretreatment with SFN blocked the development of type 1 diabetes in streptozotocin-treated mice.« less

Role of nuclear factor-erythroid 2-related factor 2 (Nrf2) in the transcriptional regulation of brain ABC transporters during acute acetaminophen (APAP) intoxication in mice.

PubMed

Ghanem, Carolina I; Rudraiah, Swetha; Bataille, Amy M; Vigo, María B; Goedken, Michael J; Manautou, José E

2015-04-01

Changes in expression of liver ABC transporters have been described during acute APAP intoxication. However, the effect of APAP on brain ABC transporters is poorly understood. The aim of this study was to evaluate the effect of APAP on brain ABC transporters expression and the role of the oxidative stress sensor Nrf2. Male C57BL/6J mice were administered APAP (400mg/kg) for analysis of brain mRNA and protein expression of Mrp1-6, Bcrp and P-gp. The results show induction of P-gp, Mrp2 and Mrp4 proteins, with no changes in Bcrp, Mrp1 or Mrp5-6. The protein values were accompanied by corresponding changes in mRNA levels. Additionally, brain Nrf2 nuclear translocation and expression of two Nrf2 target genes, quinone oxidoreductase 1 (Nqo1) and Hemoxygenase 1 (Ho-1), was evaluated at 6, 12 and 24h after APAP treatment. Nrf2 nuclear content increased by 58% at 12h after APAP along with significant increments in mRNA and protein expression of Nqo1 and Ho-1. Furthermore, APAP treated Nrf2 knockout mice did not increase mRNA or protein expression of Mrp2 and Mrp4 as observed in wildtypes. In contrast, P-gp induction by APAP was observed in both genotypes. In conclusion, acute APAP intoxication induces protein expression of brain P-gp, Mrp2 and Mrp4. This study also suggests that brain changes in Mrp2 and Mrp4 expression may be due to in situ Nrf2 activation by APAP, while P-gp induction is independent of Nrf2 function. The functional consequences of these changes in brain ABC transporters by APAP deserve further attention. Copyright © 2015 Elsevier Inc. All rights reserved.

Neuroligin-3 protects retinal cells from H2O2-induced cell death via activation of Nrf2 signaling.

PubMed

Li, Xiu-Miao; Huang, Dan; Yu, Qing; Yang, Jian; Yao, Jin

2018-05-25

Intensified oxidative stress can cause severe damage to human retinal pigment epithelium (RPE) cells and retinal ganglion cells (RGCs). The potential effect of neuroligin-3 (NLGN3) against the process is studied here. Our results show that NLGN3 efficiently inhibited hydrogen peroxide (H 2 O 2 )-induced death and apoptosis in human RPE cells and RGCs. H 2 O 2 -induced reactive oxygen species (ROS) production, lipid peroxidation and DNA damage in retinal cells were alleviated by NLGN3. NLGN3 activated nuclear-factor-E2-related factor 2 (Nrf2) signaling, enabling Nrf2 protein stabilization, nuclear translocation and expression of key anti-oxidant enzymes (HO1, NOQ1 and GCLC) in RPE cells and RGCs. Further results demonstrate that NLGN3 activated Akt-mTORC1 signaling in retinal cells. Conversely, Akt-mTORC1 inhibitors (RAD001 and LY294002) reduced NLGN3-induced HO1, NOQ1 and GCLC mRNA expression. Significantly, Nrf2 silencing by targeted shRNAs reversed NLGN3-induced retinal cytoprotection against H 2 O 2 . We conclude that NLGN3 activates Nrf2 signaling to protect human retinal cells from H 2 O 2 . NLGN3 could be further tested as a valuable retinal protection agent. Copyright © 2018 Elsevier Inc. All rights reserved.

Lipoicmethylenedioxyphenol Reduces Experimental Atherosclerosis through Activation of Nrf2 Signaling

PubMed Central

Ying, Zhekang; Chen, Minjie; Xie, Xiaoyun; Wang, Xiaoke; Kherada, Nisharahmed; Desikan, Rajagopal; Mihai, Georgeta; Burns, Patrick; Sun, Qinghua; Rajagopalan, Sanjay

2016-01-01

Objective Oxidative stress is implicated in the pathogenesis of atherosclerosis, and Nrf2 is the transcriptional factor central in cellular antioxidant responses. In the present study, we investigate the effect of a dihydrolipoic acid derivative lipoicmethylenedioxyphenol (LMDP) on the progression of atherosclerosis and test whether its effect on atherosclerosis is mediated by Nrf2. Methods and Results Both magnetic resonance imaging (MRI) scanning and en face analysis reveal that 14 weeks of treatment with LMDP markedly reduced atherosclerotic burden in a rabbit balloon vascular injury model. Myograph analyses show decreased aortic contractile response to phenylephrine and increased aortic response to acetylcholine and insulin in LMDP-treated animals, suggesting that LMDP inhibits atherosclerosis through improving vascular function. A role of Nrf2 signaling in mediating the amelioration of vascular function by LMDP was supported by increased Nrf2 translocation into nuclear and increased expression of Nrf2 target genes. Furthermore, chemotaxis analysis with Boydem chamber shows that leukocytes isolated from LMDP-treated rabbits had reduced chemotaxis, and knock-down of Nrf2 significantly reduced the effect of LMDP on the chemotaxis of mouse macrophages. Conclusion Our results support that LMDP has an anti-atherosclerotic effect likely through activation of Nrf2 signaling and subsequent inhibition of macrophage chemotaxis. PMID:26859892

Induction of glutathione synthesis and heme oxygenase 1 by the flavonoids butein and phloretin is mediated through the ERK/Nrf2 pathway and protects against oxidative stress.

PubMed

Yang, Ya-Chen; Lii, Chong-Kuei; Lin, Ai-Hsuan; Yeh, Yu-Wen; Yao, Hsien-Tsung; Li, Chien-Chun; Liu, Kai-Li; Chen, Haw-Wen

2011-12-01

Butein and phloretin are chalcones that are members of the flavonoid family of polyphenols. Flavonoids have well-known antioxidant and anti-inflammatory activities. In rat primary hepatocytes, we examined whether butein and phloretin affect tert-butylhydroperoxide (tBHP)-induced oxidative damage and the possible mechanism(s) involved. Treatment with butein and phloretin markedly attenuated tBHP-induced peroxide formation, and this amelioration was reversed by l-buthionine-S-sulfoximine [a glutamate cysteine ligase (GCL) inhibitor] and zinc protoporphyrin [a heme oxygenase 1 (HO-1) inhibitor]. Butein and phloretin induced both HO-1 and GCL protein and mRNA expression and increased intracellular glutathione (GSH) and total GSH content. Butein treatment activated the ERK1/2 signaling pathway and increased Nrf2 nuclear translocation, Nrf2 nuclear protein-DNA binding activity, and ARE-luciferase reporter activity. The roles of the ERK signaling pathway and Nrf2 in butein-induced HO-1 and GCL catalytic subunit (GCLC) expression were determined by using RNA interference directed against ERK2 and Nrf2. Both siERK2 and siNrf2 abolished butein-induced HO-1 and GCLC protein expression. These results suggest the involvement of ERK2 and Nrf2 in the induction of HO-1 and GCLC by butein. In an animal study, phloretin was shown to increase GSH content and HO-1 expression in rat liver and decrease carbon tetrachloride-induced hepatotoxicity. In conclusion, we demonstrate that butein and phloretin up-regulate HO-1 and GCL expression through the ERK2/Nrf2 pathway and protect hepatocytes against oxidative stress. Copyright © 2011 Elsevier Inc. All rights reserved.

Anti-neuroinflammatory Effect of Emodin in LPS-Stimulated Microglia: Involvement of AMPK/Nrf2 Activation.

PubMed

Park, Sun Young; Jin, Mei Ling; Ko, Min Jung; Park, Geuntae; Choi, Young-Whan

2016-11-01

AMPK/Nrf2 signaling regulates multiple antioxidative factors and exerts neuroprotective effects. Emodin is one of the main bioactive components extracted from Polygonum multiflorum, a plant possessing important activities for human health and for treating a variety of diseases. This study examined whether emodin can activate AMPK/Nrf2 signaling and induce the expression of genes targeted by this pathway. In addition, the anti-neuroinflammatory properties of emodin in lipopolysaccharide (LPS)-stimulated microglia were examined. In microglia, the emodin treatment increased the levels of LKB1, CaMKII, and AMPK phosphorylation. Emodin increased the translocation and transactivity of Nrf2 and enhanced the levels of HO-1 and NQO1. In addition, the emodin-mediated expression of HO-1 and NQO1 was attenuated completely by an AMPK inhibitor (compound C). Moreover, emodin decreased dramatically the LPS-induced production of NO and PGE 2 as well as the protein expression and promoter activity of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). In addition, emodin effectively inhibited the production of pro-inflammatory cytokines, TNF-α and IL-6, and reduced the level of IκBα phosphorylation, leading to the suppression of the nuclear translocation, phosphorylation, and transactivity of NF-κB. Emodin also suppressed the LPS-stimulated activation of STATs, JNK, and p38 MAPK. The anti-inflammatory effects of emodin were reversed by transfection with Nrf-2 and HO-1 siRNA and by a co-treatment with an AMPK inhibitor. These results suggest that emodin isolated from P. multiflorum can be used as a natural anti-neuroinflammatory agent that exerts its effects by inducing HO-1 and NQO1 via AMPK/Nrf2 signaling in microglia.

Curcumin mitigates axonal injury and neuronal cell apoptosis through the PERK/Nrf2 signaling pathway following diffuse axonal injury.

PubMed

Huang, Tingqin; Zhao, Junjie; Guo, Dan; Pang, Honggang; Zhao, Yonglin; Song, Jinning

2018-05-23

Diffuse axonal injury (DAI) accounts for more than 50% of all traumatic brain injury. In response to the mechanical damage associated with DAI, the abnormal proteins produced in the neurons and axons, namely, β-APP and p-tau, induce endoplasmic reticulum (ER) stress. Curcumin, a major component extracted from the rhizome of Curcuma longa, has shown potent anti-inflammatory, antioxidant, anti-infection, and antitumor activity in previous studies. Moreover, curcumin is an activator of nuclear factor-erythroid 2-related factor 2 (Nrf2) and promotes its nuclear translocation. In this study, we evaluated the therapeutic potential of curcumin for the treatment of DAI and investigated the mechanisms underlying the protective effects of curcumin against neural cell death and axonal injury after DAI. Rats subjected to a model of DAI by head rotational acceleration were treated with vehicle or curcumin to evaluate the effect of curcumin on neuronal and axonal injury. We observed that curcumin (20 mg/kg intraperitoneal) administered 1 h after DAI induction alleviated the aggregation of p-tau and β-APP in neurons, reduced ER-stress-related cell apoptosis, and ameliorated neurological deficits. Further investigation showed that the protective effect of curcumin in DAI was mediated by the PERK/Nrf2 pathway. Curcumin promoted PERK phosphorylation, and then Nrf2 dissociated from Keap1 and was translocated to the nucleus, which activated ATF4, an important bZIP transcription factor that maintains intracellular homeostasis, but inhibited the CHOP, a hallmark of ER stress and ER-associated programmed cell death. In summary, we demonstrate for the first time that curcumin confers protection against abnormal proteins and neuronal apoptosis after DAI, that the process is mediated by strengthening of the unfolded protein response to overcome ER stress, and that the protective effect of curcumin against DAI is dependent on the activation of Nrf2.

IDH1 R132H mutation regulates glioma chemosensitivity through Nrf2 pathway.

PubMed

Li, Kaishu; Ouyang, Leping; He, Mingliang; Luo, Ming; Cai, Wangqing; Tu, Yalin; Pi, Rongbiao; Liu, Anmin

2017-04-25

Numerous studies have reported that glioma patients with isocitrate dehydrogenase 1(IDH1) R132H mutation are sensitive to temozolomide treatment. However, the mechanism of IDH1 mutations on the chemosensitivity of glioma remains unclear. In this study, we investigated the role and the potential mechanism of Nrf2 in IDH1 R132H-mediated drug resistance. Wild type IDH1 (R132H-WT) and mutant IDH1 (R132H) plasmids were constructed. Stable U87 cells and U251 cells overexpressing IDH1 were generated. Phenotypic differences between IDH1-WT and IDH1 R132H overexpressing cells were evaluated using MTT, cell colony formation assay, scratch test assay and flow cytometry. Expression of IDH1 and its associated targets, nuclear factor-erythroid 2-related factor 2 (Nrf2), NAD(P)H quinine oxidoreductase 1 (NQO1), multidrug resistant protein 1 (MRP1) and p53 were analyzed. The IDH1 R132H overexpressing cells were more sensitive to temozolomide than WT and the control, and Nrf2 was significantly decreased in IDH1 R132H overexpressing cells. We found that knocking down Nrf2 could decrease resistance to temozolomide. The nuclear translocation of Nrf2 in IDH1 R132H overexpressing cells was lower than the WT and the control groups after temozolomide treatment. When compared with WT cells, NQO1 expression was reduced in IDH1 R132H cells, especially after temozolomide treatment. P53 was involved in the resistance mechanism of temozolomide mediated by Nrf2 and NQO1. Nrf2 played an important role in IDH1 R132H-mediated drug resistance. The present study provides new insight for glioma chemotherapy with temozolomide.

IDH1 R132H mutation regulates glioma chemosensitivity through Nrf2 pathway

PubMed Central

Luo, Ming; Cai, Wangqing; Tu, Yalin; Pi, Rongbiao; Liu, Anmin

2017-01-01

Purpose Numerous studies have reported that glioma patients with isocitrate dehydrogenase 1(IDH1) R132H mutation are sensitive to temozolomide treatment. However, the mechanism of IDH1 mutations on the chemosensitivity of glioma remains unclear. In this study, we investigated the role and the potential mechanism of Nrf2 in IDH1 R132H-mediated drug resistance. Methods Wild type IDH1 (R132H-WT) and mutant IDH1 (R132H) plasmids were constructed. Stable U87 cells and U251 cells overexpressing IDH1 were generated. Phenotypic differences between IDH1-WT and IDH1 R132H overexpressing cells were evaluated using MTT, cell colony formation assay, scratch test assay and flow cytometry. Expression of IDH1 and its associated targets, nuclear factor-erythroid 2-related factor 2 (Nrf2), NAD(P)H quinine oxidoreductase 1 (NQO1), multidrug resistant protein 1 (MRP1) and p53 were analyzed. Results The IDH1 R132H overexpressing cells were more sensitive to temozolomide than WT and the control, and Nrf2 was significantly decreased in IDH1 R132H overexpressing cells. We found that knocking down Nrf2 could decrease resistance to temozolomide. The nuclear translocation of Nrf2 in IDH1 R132H overexpressing cells was lower than the WT and the control groups after temozolomide treatment. When compared with WT cells, NQO1 expression was reduced in IDH1 R132H cells, especially after temozolomide treatment. P53 was involved in the resistance mechanism of temozolomide mediated by Nrf2 and NQO1. Conclusions Nrf2 played an important role in IDH1 R132H-mediated drug resistance. The present study provides new insight for glioma chemotherapy with temozolomide. PMID:28427200

Inhibition of early T cell cytokine production by arsenic trioxide occurs independently of Nrf2.

PubMed

VanDenBerg, Kelly R; Freeborn, Robert A; Liu, Sheng; Kennedy, Rebekah C; Zagorski, Joseph W; Rockwell, Cheryl E

2017-01-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a stress-activated transcription factor that induces a variety of cytoprotective genes. Nrf2 also mediates immunosuppressive effects in multiple inflammatory models. Upon activation, Nrf2 dissociates from its repressor protein, Keap1, and translocates to the nucleus where it induces Nrf2 target genes. The Nrf2-Keap1 interaction is disrupted by the environmental toxicant and chemotherapeutic agent arsenic trioxide (ATO). The purpose of the present study was to determine the effects of ATO on early events of T cell activation and the role of Nrf2 in those effects. The Nrf2 target genes Hmox-1, Nqo-1, and Gclc were all upregulated by ATO (1-2 μM) in splenocytes derived from wild-type, but not Nrf2-null, mice, suggesting that Nrf2 is activated by ATO in splenocytes. ATO also inhibited IFNγ, IL-2, and GM-CSF mRNA and protein production in wild-type splenocytes activated with the T cell activator, anti-CD3/anti-CD28. However, ATO also decreased production of these cytokines in activated splenocytes from Nrf2-null mice, suggesting the inhibition is independent of Nrf2. Interestingly, ATO inhibited TNFα protein secretion, but not mRNA expression, in activated splenocytes suggesting the inhibition is due to post-transcriptional modification. In addition, c-Fos DNA binding was significantly diminished by ATO in wild-type and Nrf2-null splenocytes activated with anti-CD3/anti-CD28, consistent with the observed inhibition of cytokine production by ATO. Collectively, this study suggests that although ATO activates Nrf2 in splenocytes, inhibition of early T cell cytokine production by ATO occurs independently of Nrf2 and may instead be due to impaired AP-1 DNA binding.

MHY1485 ameliorates UV-induced skin cell damages via activating mTOR-Nrf2 signaling

PubMed Central

Yang, Bo; Xu, Qiu-Yun; Guo, Chun-Yan; Huang, Jin-Wen; Wang, Shu-Mei; Li, Yong-Mei; Tu, Ying; He, Li; Bi, Zhi-Gang; Ji, Chao; Cheng, Bo

2017-01-01

Ultra Violet (UV)-caused skin cell damage is a main cause of skin cancer. Here, we studied the activity of MHY1485, a mTOR activator, in UV-treated skin cells. In primary human skin keratinocytes, HaCaT keratinocytes and human skin fibroblasts, MHY1485 ameliorated UV-induced cell death and apoptosis. mTOR activation is required for MHY1485-induced above cytoprotective actions. mTOR kinase inhibitors (OSI-027, AZD-8055 and AZD-2014) or mTOR shRNA knockdown almost abolished MHY1485-induced cytoprotection. Further, MHY1485 treatment in skin cells activated mTOR downstream NF-E2-related factor 2 (Nrf2) signaling, causing Nrf2 Ser-40 phosphorylation, stabilization/upregulation and nuclear translocation, as well as mRNA expression of Nrf2-dictated genes. Contrarily, Nrf2 knockdown or S40T mutation almost nullified MHY1485-induced cytoprotection. MHY1485 suppressed UV-induced reactive oxygen species production and DNA single strand breaks in skin keratinocytes and fibroblasts. Together, we conclude that MHY1485 inhibits UV-induced skin cell damages via activating mTOR-Nrf2 signaling. PMID:28061443

Novel antioxidants' synthesis and their anti-oxidative activity through activating Nrf2 signaling pathway.

PubMed

Wu, Jianzhang; Ren, Jiye; Yao, Song; Wang, Jiabing; Huang, Lili; Zhou, Peng; Yun, Di; Xu, Qing; Wu, Shoubiao; Wang, Zhankun; Qiu, Peihong

2017-04-01

Novel structure compounds (WS) containing 3,4,5-trimethoxyphenyl and acyl pyrazole were designed and synthesized based combination principles. Among them, WS13 was screened out to possess desirable anti-oxidative activity in vitro. Cell survival assay and apoptosis experiment in H 2 O 2 induced PC12 cells injury model all showed that its cytoprotection exhibited a concentration-effect manner. WS13 at 10μM could remove ROS with equal effiency to edaravone. Further, it clearly activated Nrf2 nuclear translocation and upregulated GCLC mRNA transcription and protein expression in dose-dependent manner, and its cytoprotection was reversed by GCLC protein inhibitor. In total, WS13 with further promotion can serve as Nrf2-GCLC activator in anti-oxidative therapy. Copyright © 2017 Elsevier Ltd. All rights reserved.

L-cysteine protects intestinal integrity, attenuates intestinal inflammation and oxidant stress, and modulates NF-κB and Nrf2 pathways in weaned piglets after LPS challenge.

PubMed

Song, Ze he; Tong, Guo; Xiao, Kan; Jiao, Le fei; Ke, Ya lu; Hu, Cai hong

2016-04-01

In this study we investigated whetherL-cysteine (L-cys) could alleviate LPS-induced intestinal disruption and its underlying mechanism. Piglets fed with anL-cys-supplemented diet had higher average daily gain.L-cys alleviated LPS-induced structural and functional disruption of intestine in weanling piglets, as demonstrated by higher villus height, villus height (VH) to crypt depth (CD) ratio, and transepithelial electrical resistance (TER) and lower FITC-dextran 4 (FD4) kDa flux in jejunum and ileum. Supplementation withL-cys up-regulated occludin and claudin-1 expression, reduced caspase-3 activity and enhanced proliferating cell nuclear antigen expression of jejunum and ileum relative to LPS group. Additionally,L-cys suppressed the LPS-induced intestinal inflammation and oxidative stress, as demonstrated by down-regulated TNF-α, IL-6 and IL-8 mRNA levels, increased catalase, superoxide dismutase, glutathione peroxidase activity, glutathione (GSH) contents and the ratio of GSH and oxidized glutathione in jejunum and ileum. Finally, a diet supplemented withL-cys inhibited NF-κB(p65) nuclear translocation and elevated NF erythroid 2-related factor 2 (Nrf2) translocation compared with the LPS group. Collectively, our results indicated the protective function ofL-cys on intestinal mucosa barrier may closely associated with its anti-inflammation, antioxidant and regulating effect on the NF-κB and Nrf2 signaling pathways. © The Author(s) 2016.

Apigenin exhibits protective effects in a mouse model of d-galactose-induced aging via activating the Nrf2 pathway.

PubMed

Sang, Ying; Zhang, Fan; Wang, Heng; Yao, Jianqiao; Chen, Ruichuan; Zhou, Zhengdao; Yang, Kun; Xie, Yan; Wan, Tianfeng; Ding, Hong

2017-06-21

The aim of the present research was to study the protective effects and underlying mechanisms of apigenin on d-galactose-induced aging mice. Firstly, apigenin exhibited a potent antioxidant activity in vitro. Secondly, d-galactose was administered by subcutaneous injection once daily for 8 weeks to establish an aging mouse model to investigate the protective effect of apigenin. We found that apigenin supplementation significantly ameliorated aging-related changes such as behavioral impairment, decreased organic index, histopathological injury, increased senescence-associated β-galactosidase (SAβ-gal) activity and advanced glycation end product (AGE) level. Further data showed that apigenin facilitated Nrf2 nuclear translocation both in aging mice and normal young mice, and the Nrf2 expression of normal young mice was higher than that of natural senile mice. In addition, the expressions of Nrf2 downstream gene targets, including HO-1 and NQO1, were also promoted by apigenin administration. Moreover, apigenin also decreased the MDA level and elevated SOD and CAT activities. In conclusion, focusing on the Nrf2 pathway is a suitable strategy to delay the aging process, and apigenin may exert an anti-senescent effect process via activating the Nrf2 pathway.

Hericium erinaceus Inhibits TNF-α-Induced Angiogenesis and ROS Generation through Suppression of MMP-9/NF-κB Signaling and Activation of Nrf2-Mediated Antioxidant Genes in Human EA.hy926 Endothelial Cells.

PubMed

Chang, Hebron C; Yang, Hsin-Ling; Pan, Jih-Hao; Korivi, Mallikarjuna; Pan, Jian-You; Hsieh, Meng-Chang; Chao, Pei-Min; Huang, Pei-Jane; Tsai, Ching-Tsan; Hseu, You-Cheng

2016-01-01

Hericium erinaceus (HE) is an edible mushroom that has been shown to exhibit anticancer and anti-inflammatory activities. We investigated the antiangiogenic and antioxidant potentials of ethanol extracts of HE in human endothelial (EA.hy926) cells upon tumor necrosis factor-α- (TNF-α-) stimulation (10 ng/mL). The underlying molecular mechanisms behind the pharmacological efficacies were elucidated. We found that noncytotoxic concentrations of HE (50-200 μg/mL) significantly inhibited TNF-α-induced migration/invasion and capillary-like tube formation of endothelial cells. HE treatment suppressed TNF-α-induced activity and/or overexpression of matrix metalloproteinase-9 (MMP-9) and intercellular adhesion molecule-1 (ICAM-1). Furthermore, HE downregulated TNF-α-induced nuclear translocation and transcriptional activation of nuclear factor-κB (NF-κB) followed by suppression of I-κB (inhibitor-κB) degradation. Data from fluorescence microscopy illustrated that increased intracellular ROS production upon TNF-α-stimulation was remarkably inhibited by HE pretreatment in a dose-dependent manner. Notably, HE triggered antioxidant gene expressions of heme oxygenase-1 (HO-1), γ-glutamylcysteine synthetase (γ-GCLC), and glutathione levels, which may contribute to inhibition of ROS. Increased antioxidant status was associated with upregulated nuclear translocation and transcriptional activation of NF-E2 related factor-2 (Nrf2) in HE treated cells. Our findings conclude that antiangiogenic and anti-inflammatory activities of H. erinaceus may contribute to its anticancer property through modulation of MMP-9/NF-κB and Nrf2-antioxidant signaling pathways.

Hericium erinaceus Inhibits TNF-α-Induced Angiogenesis and ROS Generation through Suppression of MMP-9/NF-κB Signaling and Activation of Nrf2-Mediated Antioxidant Genes in Human EA.hy926 Endothelial Cells

PubMed Central

Chang, Hebron C.; Yang, Hsin-Ling; Pan, Jih-Hao; Korivi, Mallikarjuna; Pan, Jian-You; Hsieh, Meng-Chang; Chao, Pei-Min; Huang, Pei-Jane; Tsai, Ching-Tsan; Hseu, You-Cheng

2016-01-01

Hericium erinaceus (HE) is an edible mushroom that has been shown to exhibit anticancer and anti-inflammatory activities. We investigated the antiangiogenic and antioxidant potentials of ethanol extracts of HE in human endothelial (EA.hy926) cells upon tumor necrosis factor-α- (TNF-α-) stimulation (10 ng/mL). The underlying molecular mechanisms behind the pharmacological efficacies were elucidated. We found that noncytotoxic concentrations of HE (50–200 μg/mL) significantly inhibited TNF-α-induced migration/invasion and capillary-like tube formation of endothelial cells. HE treatment suppressed TNF-α-induced activity and/or overexpression of matrix metalloproteinase-9 (MMP-9) and intercellular adhesion molecule-1 (ICAM-1). Furthermore, HE downregulated TNF-α-induced nuclear translocation and transcriptional activation of nuclear factor-κB (NF-κB) followed by suppression of I-κB (inhibitor-κB) degradation. Data from fluorescence microscopy illustrated that increased intracellular ROS production upon TNF-α-stimulation was remarkably inhibited by HE pretreatment in a dose-dependent manner. Notably, HE triggered antioxidant gene expressions of heme oxygenase-1 (HO-1), γ-glutamylcysteine synthetase (γ-GCLC), and glutathione levels, which may contribute to inhibition of ROS. Increased antioxidant status was associated with upregulated nuclear translocation and transcriptional activation of NF-E2 related factor-2 (Nrf2) in HE treated cells. Our findings conclude that antiangiogenic and anti-inflammatory activities of H. erinaceus may contribute to its anticancer property through modulation of MMP-9/NF-κB and Nrf2-antioxidant signaling pathways. PMID:26823953

t-BHQ Provides Protection against Lead Neurotoxicity via Nrf2/HO-1 Pathway

PubMed Central

Ye, Fang; Li, Xiaoyi; Li, Lili; Yuan, Jing; Chen, Jun

2016-01-01

The neurotoxicity of lead has been well established, and oxidative stress is strongly associated with lead-induced neurotoxicity. Nrf2 is important for protection against oxidative stress in many disease models. We applied t-BHQ, which is an Nrf2 activator, to investigate the possible role of Nrf2 in the protection against lead neurotoxicity. t-BHQ significantly attenuated the oxidative stress in developmental rats by decreasing MDA level, as well as by increasing SOD activity and GSH content, in the hippocampus and frontal cortex. Furthermore, neuronal apoptosis was detected by Nissl staining, and Bax expression was inhibited in the t-BHQ-treated group. Results showed that t-BHQ suppressed ROS production and caspase 3/7 activity but increased intracellular GSH content, in SH-SY5Y cells under lead exposure. Moreover, in vivo and in vitro, t-BHQ enhanced the nuclear translocation of Nrf2 and binding to ARE areas but did not induce Nrf2 transcription. These phenomena were confirmed using RT-PCR, EMSA, Western blot, and immunofluorescence analyses. Subsequent upregulation of the expression of HO-1, NQO1, and GCLC was observed. However, knockdown of Nrf2 or HO-1 adversely affected the protective effects of t-BHQ against lead toxicity in SH-SY5Y cells. Thus, t-BHQ can protect against lead neurotoxicity, depending on the Nrf2/HO-1 pathway. PMID:26798413

Phase II enzyme induction by a carotenoid, lutein, in a PC12D neuronal cell line

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miyake, Seiji; Department of Ophthalmology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582; Wakasa Seikatsu Co., Ltd., 134 Chudoujiminami-cho, Shimogyo-ku, Kyoto 600-8813

Highlights: • Lutein reduced ROS levels in a PC12D neuronal cell line. • Lutein induced mRNAs of phase II antioxidative enzymes in PC12D neuronal cells. • Lutein increased protein levels of HO-1, SOD2, and NQO-1 in PC12D neuronal cells. • Lutein had no effect on intranuclear Nrf2 levels in PC12D neuronal cells. • Lutein did not activate potential upstream Nrf2 nuclear translocation pathways. - Abstract: The mechanism by which lutein, a carotenoid, acts as an antioxidant in retinal cells is still not fully understood. Here, lutein treatment of a neuronal cell line (PC12D) immediately resulted in reduced intracellular ROS levels,more » implying that it has a direct role in ROS scavenging. Significantly, lutein treatment also induced phase II antioxidative enzyme expression, probably via a nuclear factor-like 2 (Nrf2) independent pathway. This latter mechanism could explain why lutein acts diversely to protect against oxidative/cytotoxic stress, and why it is physiologically involved in the human neural tissue, such as the retina.« less

Agmatine Reduces Lipopolysaccharide-Mediated Oxidant Response via Activating PI3K/Akt Pathway and Up-Regulating Nrf2 and HO-1 Expression in Macrophages

PubMed Central

Chai, Jianshen; Luo, Li; Hou, Fengyan; Fan, Xia; Yu, Jing; Ma, Wei; Tang, Wangqi; Yang, Xue; Zhu, Junyu; Kang, Wenyuan; Yan, Jun; Liang, Huaping

2016-01-01

Macrophages are key responders of inflammation and are closely related with oxidative stress. Activated macrophages can enhance oxygen depletion, which causes an overproduction of reactive oxygen species (ROS) and leads to further excessive inflammatory response and tissue damage. Agmatine, an endogenous metabolite of L-arginine, has recently been shown to have neuroprotective effects based on its antioxidant properties. However, the antioxidant effects of agmatine in peripheral tissues and cells, especially macrophages, remain unclear. In this study we explored the role of agmatine in mediating antioxidant effects in RAW 264.7 cells and studied its antioxidant mechanism. Our data demonstrate that agmatine is an activator of Nrf2 signaling that markedly enhances Nrf2 nuclear translocation, increases nuclear Nrf2 protein level, up-regulates the expression of the Nrf2 downstream effector HO-1, and attenuates ROS generation induced by Lipopolysaccharide (LPS). We further demonstrated that the agmatine-induced activation of Nrf2 is likely through the PI3K/Akt pathway. LY294002, a specific PI3K/Akt inhibitor, abolished agmatine-induced HO-1 up-regulation and ROS suppression significantly. Inhibiting HO-1 pathway significantly attenuated the antioxidant effect of agmatine which the products of HO-1 enzymatic activity contributed to. Furthermore, the common membrane receptors of agmatine were evaluated, revealing that α2-adrenoceptor, I1-imidazoline receptor or I2-imidazoline receptor are not required by the antioxidant properties of agmatine. Taken together, our findings revealed that agmatine has antioxidant activity against LPS-induced ROS accumulation in RAW 264.7 cells involving HO-1 expression induced by Nrf2 via PI3K/Akt pathway activation. PMID:27685463

Mangiferin activates the Nrf2-ARE pathway and reduces etoposide-induced DNA damage in human umbilical cord mononuclear blood cells.

PubMed

Zhang, Benping; Zhao, Jie; Li, Shanshan; Zeng, Linglan; Chen, Yan; Fang, Jun

2015-04-01

Mangiferin (2-C-β-d-gluco-pyranosyl-1,3,6,7-tetrahydroxyxanthone) is a well-known natural antioxidant distributed in various plants of the Anacardiaceae and Gentianaceae families. Mangiferin can inhibit carcinogen-induced lung or colon tumor formation in experimental animals. However, the molecular mechanisms of its chemopreventive activity remain unexplored. This study aimed to investigate the effects of mangiferin on chemical carcinogen-induced DNA damage and Nrf2-ARE signaling in hematopoietic cells. Mononuclear cells (MNCs) were isolated from human umbilical cord blood (hUCB). DNA damage was evaluated by comet and micronucleus assays. The expression of Nrf2 and NQO1 was examined by immunofluorescence and western blotting. An electrophoretic mobility shift assay (EMSA) was used to detect the binding activity of Nrf2 with NQO1-ARE sequences. We found that mangiferin treatment significantly reduced DNA damage in etoposide-treated MNCs, which was verified by decreased olive tail moment (OTM) and micronucleus (MN) frequency. Mangiferin treatment significantly promoted Nrf2 translocation into the nucleus and increased nuclear Nrf2 expression. Moreover, NQO1, an Nrf2 signaling target, was significantly upregulated by mangiferin treatment, and the binding activity of Nrf2 with NQO1-ARE sequences was elevated after mangiferin treatment. Mangiferin activated Nrf2 signaling, upregulated NQO1 expression, and significantly reduced etoposide-induced DNA damage. Thus, mangiferin is a potential cytoprotective agent for hematopoietic cells.

Hydrogen sulfide (H2S) attenuates uranium-induced acute nephrotoxicity through oxidative stress and inflammatory response via Nrf2-NF-κB pathways.

PubMed

Zheng, Jifang; Zhao, Tingting; Yuan, Yan; Hu, Nan; Tang, Xiaoqing

2015-12-05

As an endogenous gaseous mediator, H2S exerts anti-oxidative, anti-inflammatory and cytoprotective effects in kidneys. This study was designed to investigate the protective effect of H2S against uranium-induced nephrotoxicity in adult SD male rats after in vivo effect of uranium on endogenous H2S formation was explored in kidneys. The levels of endogenous H2S and H2S-producing enzymes (CBS and CSE) were measured in renal homogenates from rats intoxicated by an intraperitoneally (i.p.) injection of uranyl acetate at a single dose of 2.5, 5 or 10 mg/kg. In rats injected i.p. with uranyl acetate (5 mg/kg) or NaHS (an H2S donor, 28 or 56 μmol/kg) alone or in combination, we determined biochemical parameters and histopathological alteration to assess kidney function, examined oxidative stress markers, and investigated Nrf2 and NF-κB pathways in kidney homogenates. The results suggest that uranium intoxication in rats decreased endogenous H2S generation as well as CBS and CSE protein expression. NaHS administration in uranium-intoxicated rats ameliorated the renal biochemical indices and histopathological effects, lowered MDA accumulation, and restored GSH level and anti-oxidative enzymes activities like SOD, CAT, GPx and GST. NaHS treatment in uranium-intoxicated rats activated uranium-inhibited protein expression and nuclear translocation of transcription factor Nrf2, which increased protein expression of downstream target-Nrf2 genes HO-1, NQO-1, GCLC, and TXNRD-1. NaHS administration in uranium-intoxicated rats inhibited uranium-induced nuclear translocation and phosphorylation of transcription factor κB/p65, which decreased protein expression of target-p65 inflammatory genes TNF-α, iNOS, and COX-2. Taken together, these data implicate that H2S can afford protection to rat kidneys against uranium-induced adverse effects through induction of antioxidant defense by activating Nrf2 pathway and reduction of inflammatory response by suppressing NF-κB pathway. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

The role of Nrf2 in oxidative stress-induced endothelial injuries.

PubMed

Chen, Bo; Lu, Yanrong; Chen, Younan; Cheng, Jingqiu

2015-06-01

Endothelial dysfunction is an important risk factor for cardiovascular disease, and it represents the initial step in the pathogenesis of atherosclerosis. Failure to protect against oxidative stress-induced cellular damage accounts for endothelial dysfunction in the majority of pathophysiological conditions. Numerous antioxidant pathways are involved in cellular redox homeostasis, among which the nuclear factor-E2-related factor 2 (Nrf2)/Kelch-like ECH-associated protein 1 (Keap1)-antioxidant response element (ARE) signaling pathway is perhaps the most prominent. Nrf2, a transcription factor with a high sensitivity to oxidative stress, binds to AREs in the nucleus and promotes the transcription of a wide variety of antioxidant genes. Nrf2 is located in the cytoskeleton, adjacent to Keap1. Keap1 acts as an adapter for cullin 3/ring-box 1-mediated ubiquitination and degradation of Nrf2, which decreases the activity of Nrf2 under physiological conditions. Oxidative stress causes Nrf2 to dissociate from Keap1 and to subsequently translocate into the nucleus, which results in its binding to ARE and the transcription of downstream target genes. Experimental evidence has established that Nrf2-driven free radical detoxification pathways are important endogenous homeostatic mechanisms that are associated with vasoprotection in the setting of aging, atherosclerosis, hypertension, ischemia, and cardiovascular diseases. The aim of the present review is to briefly summarize the mechanisms that regulate the Nrf2/Keap1-ARE signaling pathway and the latest advances in understanding how Nrf2 protects against oxidative stress-induced endothelial injuries. Further studies regarding the precise mechanisms by which Nrf2-regulated endothelial protection occurs are necessary for determining whether Nrf2 can serve as a therapeutic target in the treatment of cardiovascular diseases. © 2015 Society for Endocrinology.

α-Lipoic acid protects against the cytotoxicity and oxidative stress induced by cadmium in HepG2 cells through regeneration of glutathione by glutathione reductase via Nrf2/ARE signaling pathway.

PubMed

Shi, Chunli; Zhou, Xue; Zhang, Jiayu; Wang, Jiachun; Xie, Hong; Wu, Zhigang

2016-07-01

α-Lipoic acid (α-LA) is a potent natural antioxidant, which is capable of regenerating glutathione (GSH). However, the mechanisms by which α-LA regenerates reduced glutathione (rGSH) via the reduction of oxidized glutathione (GSSG) by glutathione reductase (GR) are still not well understood. In the present study, we investigated if α-LA replenished rGSH by GR via Nrf2/ARE signaling pathway in cadmium-treated HepG2 cells. We found that α-LA antagonized the oxidative damage and alleviated the cytotoxicity in cadmium-induced HepG2 cells by regeneration of rGSH. α-LA regenerated rGSH by activating Nrf2 signaling pathway via promoting the nuclear translocation of Nrf2, which upregulates the transcription of GR, and thus increased the activity of GR. Our results indicated that α-LA was an effective agent to antagonize the oxidative stress and alleviate the cytotoxicity in cadmium-treated HepG2 cells by regenerating rGSH through activating Nrf2 signaling pathway. Copyright © 2016. Published by Elsevier B.V.

Enhancement of the Effect of Methyl Pyropheophorbide-a-Mediated Photodynamic Therapy was Achieved by Increasing ROS through Inhibition of Nrf2-HO-1 or Nrf2-ABCG2 Signaling.

PubMed

Tian, Si; Yong, Min; Zhu, Jiang; Zhang, Li; Pan, Li; Chen, Qing; Li, Kai-Ting; Kong, Yu-Han; Jiang, Yuan; Yu, Ting-He; Yu, Le-Hua; Bai, Ding-Qun

2017-01-01

Emerging evidence indicates that the transcription factor nuclear factor-E2-related factor 2 (Nrf2) plays an essential role in cellular defense against oxidative stress; its activation has been related to cytoprotection. Here, we investigated the role of Nrf2 in improving the efficacy of methyl pyropheophorbide-amediated photodynamic therapy (Mppa-PDT) via the downregulation of Nrf2. Human ovarian cancer A2780 cells and SKOV3 cells were treated with Mppa-PDT and siRNA transfection was performed to inhibit Nrf2. After treated with siRNA and Mppa-PDT, the cell viability was examined with CCK-8 assay; cell apoptosis was detected tested by flow cytometry with Annexin V-FITC/PI; the celluar reactive oxygen species (ROS) and mitochondrial membrane potential were measured with DCFHDA and JC-1 staining; expression of protein was assessed by western blot analysis. We found that Nrf2 translocated from the cytoplasm to the nucleus in vitro and in vivo, and the expression of Nrf2 and P-Nrf2 increased through a possible mechanism regulated by mitogen-activated protein kinase (MAPK) after Mppa-PDT treatment. Furthermore, cytotoxicity and apoptosis induced by Mppa-PDT increased after Nrf2down-regulation. Nrf2 down -regulation increased reactive oxygen species (ROS) levels by attenuating antioxidants or pumping Mppa out of cells,which resulted from the inhibition of Nrf2-HO-1 or Nrf2- ABCG2 signaling. In addition, SKOV3 cells exhibited increased resistance to Mppa-PDT, and the expression levels of P-Nrf2 and ABCG2 were higher in SKOV3 cells than in A2780 cells, suggesting that Nrf2-ABCG2 signaling might be involved in the intrinsic resistanceto Mppa-PDT. These results provided evidence that Nrf2 down-regulation can enhance the effect of Mppa-PDT. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

5-Hydroxy-3,6,7,8,3'4'-hexamethoxyflavone inhibits nitric oxide production in lipopolysaccharide-stimulated BV2 microglia via NF-κB suppression and Nrf-2-dependent heme oxygenase-1 induction.

PubMed

Kang, Chang-Hee; Kim, Min Jeong; Seo, Min Jeong; Choi, Yung Hyun; Jo, Wol Soon; Lee, Kyung-Tae; Jeong, Yong Kee; Kim, Gi-Young

2013-07-01

In this study, we found that 5-hydroxy-3,6,7,8,3'4'-hexamethoxyflavone (5HHMF) from Hizikia fusiforme considerably inhibits lipopolysaccharide (LPS)-stimulated NO production by suppressing the expression of inducible NO synthase (iNOS) in BV2 microglia. In addition, 5HHMF blocked LPS-induced phosphorylation of IκB, resulting in suppression of the nuclear translocation of nuclear factor-κB (NF-κB) subunits, namely p65 and p50, which are important molecules involved in the regulation of iNOS expression. Pyrrolidine dithiocarbamate (PDTC), a specific NF-κB inhibitor, along with 20S proteasome inhibitor (PSI) significantly inhibited LPS-induced iNOS expression, which indirectly suggested that 5HHMF downregulated iNOS expression by suppressing NF-κB activity. Thus, we found that 5HHMF enhances heme oxygenase-1 (HO-1) expression via nuclear factor-erythroid 2-related factor 2 (Nrf2) activation. In addition, cobalt protoporphyrin (CoPP), a specific HO-1 inducer, predominantly suppressed LPS-induced NO production. In contrast, zinc protoporphyrin (ZnPP), a specific HO-1 inhibitor, showed a partial suppressive effect of 5HHMF on LPS-induced NO production. Further, 5HHMF increased specific DNA-binding activity of Nrf2, and transient knockdown with Nrf2 siRNA subsequently reversed 5HHMF-induced NO inhibition, which was followed by suppression of HO-1 activity. Taken together, our findings indicate that 5HHMF suppresses NO production through modulation of iNOS, consequently suppressing NF-κB activity and induction of Nrf2-dependent HO-1 activity. Copyright © 2013 Elsevier Ltd. All rights reserved.

Curcumin plays neuroprotective roles against traumatic brain injury partly via Nrf2 signaling.

PubMed

Dong, Wenwen; Yang, Bei; Wang, Linlin; Li, Bingxuan; Guo, Xiangshen; Zhang, Miao; Jiang, Zhenfei; Fu, Jingqi; Pi, Jingbo; Guan, Dawei; Zhao, Rui

2018-05-01

Traumatic brain injury (TBI), which leads to high mortality and morbidity, is a prominent public health problem worldwide with no effective treatment. Curcumin has been shown to be beneficial for neuroprotection in vivo and in vitro, but the underlying mechanism remains unclear. This study determined whether the neuroprotective role of curcumin in mouse TBI is dependent on the NF-E2-related factor (Nrf2) pathway. The Feeney weight-drop contusion model was used to mimic TBI. Curcumin was administered intraperitoneally 15 min after TBI induction, and brains were collected at 24 h after TBI. The levels of Nrf2 and its downstream genes (Hmox-1, Nqo1, Gclm, and Gclc) were detected by Western blot and qRT-PCR at 24 h after TBI. In addition, edema, oxidative damage, cell apoptosis and inflammatory reactions were evaluated in wild type (WT) and Nrf2-knockout (Nrf2-KO) mice to explore the role of Nrf2 signaling after curcumin treatment. In wild type mice, curcumin treatment resulted in reduced ipsilateral cortex injury, neutrophil infiltration, and microglia activation, improving neuron survival against TBI-induced apoptosis and degeneration. These effects were accompanied by increased expression and nuclear translocation of Nrf2, and enhanced expression of antioxidant enzymes. However, Nrf2 deletion attenuated the neuroprotective effects of curcumin in Nrf2-KO mice after TBI. These findings demonstrated that curcumin effects on TBI are associated with the activation the Nrf2 pathway, providing novel insights into the neuroprotective role of Nrf2 and the potential therapeutic use of curcumin for TBI. Copyright © 2018. Published by Elsevier Inc.

Piper betle Induced Cytoprotective Genes and Proteins via the Nrf2/ARE Pathway in Aging Mice.

PubMed

Aliahmat, Nor Syahida; Abdul Sani, Nur Fathiah; Wan Hasan, Wan Nuraini; Makpol, Suzana; Wan Ngah, Wan Zurinah; Mohd Yusof, Yasmin Anum

2016-01-01

The objective of this study was to elucidate the underlying antioxidant mechanism of aqueous extract of Piper betle (PB) in aging rats. The nuclear factor erythroid 2-related factor 2 (Nrf2)/ARE pathway involving phase II detoxifying and antioxidant enzymes plays an important role in the antioxidant system by reducing electrophiles and reactive oxygen species through induction of phase II enzymes and proteins. Genes and proteins of phase II detoxifying antioxidant enzymes were analyzed by QuantiGenePlex 2.0 Assay and Western blot analysis. PB significantly induced genes and proteins of phase II and antioxidant enzymes, NAD(P)H quinone oxidoreductase 1, and catalase in aging mice (p < 0.05). The expression of these enzymes were stimulated via translocation of Nrf2 into the nucleus, indicating the involvement of ARE, a cis-acting motif located in the promoter region of nearly all phase II genes. PB was testified for the first time to induce cytoprotective genes through the Nrf2/ARE signaling pathway, thus unraveling the antioxidant mechanism of PB during the aging process. © 2016 S. Karger AG, Basel.

Andrographolide stimulates p38 mitogen-activated protein kinase-nuclear factor erythroid-2-related factor 2-heme oxygenase 1 signaling in primary cerebral endothelial cells for definite protection against ischemic stroke in rats.

PubMed

Yen, Ting-Lin; Chen, Ray-Jade; Jayakumar, Thanasekaran; Lu, Wan-Jung; Hsieh, Cheng-Ying; Hsu, Ming-Jen; Yang, Chih-Hao; Chang, Chao-Chien; Lin, Yen-Kuang; Lin, Kuan-Hung; Sheu, Joen-Rong

2016-04-01

Stroke pathogenesis involves complex oxidative stress-related pathways. The nuclear factor erythroid-2-related factor 2 (Nrf2) and heme oxygenase 1 (HO-1) pathways have been considered molecular targets in pharmacologic intervention for ischemic diseases. Andrographolide, a labdane diterpene, has received increasing attention in recent years because of its various pharmacologic activities. We determined that andrographolide modulates the mitogen-activated protein kinase (MAPK)-Nrf2-HO-1 signaling cascade in primary cerebral endothelial cells (CECs) to provide positive protection against middle cerebral artery occlusion (MCAO)-induced ischemic stroke in rats. In the present study, andrographolide (10 μM) increased HO-1 protein and messenger RNA expressions, Nrf2 phosphorylation, and nuclear translocation in CECs, and these activities were disrupted by a p38 MAPK inhibitor, SB203580, but not by the extracellular signal-regulated kinase inhibitor PD98059 or c-Jun amino-terminal kinase inhibitor SP600125. Similar results were observed in confocal microscopy analysis. Moreover, andrographolide-induced Nrf2 and HO-1 protein expressions were significantly inhibited by Nrf2 small interfering RNA. Moreover, HO-1 knockdown attenuated the protective effect of andrographolide against oxygen-glucose deprivation-induced CEC death. Andrographolide (0.1 mg/kg) significantly suppressed free radical formation, blood-brain barrier disruption, and brain infarction in MCAO-insulted rats, and these effects were reversed by the HO-1 inhibitor zinc protoporphyrin IX. The mechanism is attributable to HO-1 activation, as directly evidenced by andrographolide-induced pronounced HO-1 expression in brain tissues, which was highly localized in the cerebral capillary. In conclusion, andrographolide increased Nrf2-HO-1 expression through p38 MAPK regulation, confirming that it provides protection against MCAO-induced brain injury. These findings provide strong evidence that andrographolide could be a therapeutic agent for treating ischemic stroke or neurodegenerative diseases. Copyright © 2016 Elsevier Inc. All rights reserved.

Anti-inflammatory effects of novel polygonum multiflorum compound via inhibiting NF-κB/MAPK and upregulating the Nrf2 pathways in LPS-stimulated microglia.

PubMed

Park, Sun Young; Jin, Mei Ling; Kang, Nam Jun; Park, Geuntae; Choi, Young-Whan

2017-06-09

The incorporation of Polygonum multiflorum into the diet can result in anti-aging effects owing to its wide range of biological and pharmaceutical properties. We investigated the anti-neuroinflammatory properties of CRPE56IGIH isolated from P. multiflorum by focusing on its role in the induction of phase II antioxidant enzymes and the modulation of upstream signaling pathways. In microglia, CRPE56IGIH significantly inhibited lipopolysaccharide (LPS)-stimulated nitric oxide and prostaglandin E 2 production with nonspecific cytotoxicity. CRPE56IGIH also markedly inhibited LPS-inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 protein and mRNA expression in the same manner as it inhibited nitric oxide and prostaglandin E 2 production. In the control cells, NF-κB transactivation and nuclear translocation occurred at a baseline level, which was significantly increased in response to LPS. However, pretreatment with CRPE56IGIH concentration-dependently inhibited the LPS-induced NF-κB transactivation and nuclear translocation. The phosphorylation of Janus kinase-signal transducers and activators of transcription and mitogen-activated protein kinases was markedly upregulated by LPS, but considerably and dose-dependently inhibited by pretreatment with CRPE56IGIH. Furthermore, CRPE56IGIH induced the expression of phase II antioxidant enzymes, including heme oxygenase-1 (HO-1) and NADPH dehydrogenase quinone-1 (NQO-1). The activation of upstream signaling pathways, such as the Nrf2 pathway, was significantly increased following CRPE56IGIH treatment. Furthermore, the anti-neuroinflammatory effect of CRPE56IGIH was reversed by transfection of Nrf2, HO-1, and NQO-1 siRNA. Our results indicated that CRPE56IGIH isolated from P. multiflorum could be used as a natural anti-neuroinflammatory agent that induces phase II antioxidant enzymes via Nrf2 signaling. Copyright © 2017 Elsevier B.V. All rights reserved.

Investigation of the effect of a panel of model hepatotoxins on the Nrf2-Keap1 defence response pathway in CD-1 mice.

PubMed

Randle, Laura E; Goldring, Chris E P; Benson, Craig A; Metcalfe, Peter N; Kitteringham, Neil R; Park, B Kevin; Williams, Dominic P

2008-01-20

The Keap1-Nrf2-ARE signalling pathway has emerged as an important regulator of the mammalian defence system to enable detoxification and clearance of foreign chemicals. Recent studies by our group using paracetamol (APAP), diethylmaleate and buthionine sulphoximine have shown that for a given xenobiotic molecule, Nrf2 induction in the murine liver is associated with protein reactivity and glutathione depletion. Here, we have investigated, in vivo, whether the ability of four murine hepatotoxins, paracetamol, bromobenzene (BB), carbon tetrachloride (CCl4) and furosemide (FS) to deplete hepatic glutathione (GSH) is related to induction of hepatic Nrf2 nuclear translocation and Nrf2-dependent gene expression. Additionally, we studied whether hepatic Nrf2 nuclear translocation is a general response during the early stages of acute hepatic chemical stress in vivo. Male CD-1 mice were administered APAP (3.5 mmol/kg), FS (1.21 mmol/kg), BB (4.8 mmol/kg) and CCl4 (1 mmol/kg) for 1, 5 and 24h. Each compound elicited significant serum ALT increases after 24h (ALT U/L: APAP, 3036+/-1462; BB, 5308+/-2210; CCl4, 5089+/-1665; FS, 2301+/-1053), accompanied by centrilobular damage as assessed by histopathology. Treatment with APAP also elicited toxicity at a much earlier time point (5h) than the other hepatotoxins (ALT U/L: APAP, 1780+/-661; BB, 161+/-15; CCl4, 90+/-23; FS, 136+/-27). Significant GSH depletion was seen with APAP (9.6+/-1.7% of control levels) and BB (52.8+/-6.2% of control levels) 1h after administration, but not with FS and CCl4. Western Blot analysis revealed an increase in nuclear Nrf2, 1h after administration of BB (209+/-10% control), CCl4 (146+/-3% control) and FS (254+/-41% control), however this was significantly lower than the levels observed in the APAP-treated mice (462+/-36% control). The levels of Nrf2-dependent gene induction were also analysed by quantitative real-time PCR and Western blotting. Treatment with APAP for 1h caused a significant increase in the levels of haem oxygenase-1 (HO-1; 2.85-fold) and glutamate cysteine ligase (GCLC; 1.62-fold) mRNA. BB and FS did not affect the mRNA levels of either gene after 1h of treatment; however CCl4 significantly increased HO-1 mRNA at this time point. After 24h treatment with the hepatotoxins, there was evidence for the initiation of a late defence response. BB significantly increased both HO-1 and GCLC protein at this time point, CCl4 increased GCLC protein alone, although FS did not alter either of these proteins. In summary, we have demonstrated that the hepatotoxins BB, CCl4 and FS can induce a small but significant increase in Nrf2 accumulation in hepatic nuclei. However, this was associated with modest changes in hepatic GSH, a delayed development of toxicity and was insufficient to activate an early functional adaptive response to these hepatotoxins.

Photoprotection by dietary phenolics against melanogenesis induced by UVA through Nrf2-dependent antioxidant responses

PubMed Central

Chaiprasongsuk, Anyamanee; Onkoksoong, Tasanee; Pluemsamran, Thanyawan; Limsaengurai, Saowalak; Panich, Uraiwan

2015-01-01

Dietary phenolics may play a protective role in UV-mediated skin pigmentation through their antioxidant and UV-absorbing actions. In this study, we investigated whether genetic silencing of Nrf2, regulating the transcription of antioxidant genes, affected melanogenesis in primary human epidermal melanocytes (HEMn) and B16F10 melanoma cells subjected to UVA (8 J/cm2) exposure. Then, we explored the antimelanogenic actions of phenolics; caffeic acid (CA) and ferulic acid (FA) providing partial UVA protection; quercetin (QU) and rutin (RU) providing strong UVA protection and; avobenzone (AV), an efficient UVA filter, in association with modulation of Nrf2-mediated antioxidant defenses in response to UVA insults in B16F10 cells. Upon oxidative insults, Nrf2 silencing promoted melanogenesis in both HEMn and B16F10 cells irradiated with UVA. Stimulation of melanogenesis by UVA correlated with increased ROS and oxidative DNA damage (8-OHdG), GSH depletion as well as a transient downregulation of Nrf2 nuclear translocation and of Nrf2-ARE signaling in B16F10 cells. All test compounds exerted antimelanogenic effects with respect to their abilities to reverse UVA-mediated oxidative damage as well as downregulation of Nrf2 activity and its target antioxidants (GCLC, GST and NQO1) in B16F10 cells. In conclusion, defective Nrf2 may promote melanogenesis under UVA irradiation through oxidative stress mechanisms. Compounds with antioxidant and/or UVA absorption properties could protect against UVA-induced melanogenesis through indirect regulatory effect on Nrf2-ARE pathway. PMID:26765101

Escin activates AKT-Nrf2 signaling to protect retinal pigment epithelium cells from oxidative stress

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Kaijun; Zhejiang Provincial Key Lab of Ophthalmology, Hangzhou; Jiang, Yiqian

Here we explored the anti-oxidative and cytoprotective potentials of escin, a natural triterpene-saponin, against hydrogen peroxide (H{sub 2}O{sub 2}) in retinal pigment epithelium (RPE) cells. We showed that escin remarkably attenuated H{sub 2}O{sub 2}-induced death and apoptosis of established (ARPE-19) and primary murine RPE cells. Meanwhile, ROS production and lipid peroxidation by H{sub 2}O{sub 2} were remarkably inhibited by escin. Escin treatment in RPE cells resulted in NF-E2-related factor 2 (Nrf2) signaling activation, evidenced by transcription of anti-oxidant-responsive element (ARE)-regulated genes, including HO-1, NQO-1 and SRXN-1. Knockdown of Nrf2 through targeted shRNAs/siRNAs alleviated escin-mediated ARE gene transcription, and almost abolishedmore » escin-mediated anti-oxidant activity and RPE cytoprotection against H{sub 2}O{sub 2}. Reversely, escin was more potent against H{sub 2}O{sub 2} damages in Nrf2-over-expressed ARPE-19 cells. Further studies showed that escin-induced Nrf2 activation in RPE cells required AKT signaling. AKT inhibitors (LY294002 and perifosine) blocked escin-induced AKT activation, and dramatically inhibited Nrf2 phosphorylation, its cytosol accumulation and nuclear translocation in RPE cells. Escin-induced RPE cytoprotection against H{sub 2}O{sub 2} was also alleviated by the AKT inhibitors. Together, these results demonstrate that escin protects RPE cells from oxidative stress possibly through activating AKT-Nrf2 signaling.« less

Diabetes-induced hepatic pathogenic damage, inflammation, oxidative stress, and insulin resistance was exacerbated in zinc deficient mouse model.

PubMed

Zhang, Chi; Lu, Xuemian; Tan, Yi; Li, Bing; Miao, Xiao; Jin, Litai; Shi, Xue; Zhang, Xiang; Miao, Lining; Li, Xiaokun; Cai, Lu

2012-01-01

Zinc (Zn) deficiency often occurs in the patients with diabetes. Effects of Zn deficiency on diabetes-induced hepatic injury were investigated. Type 1 diabetes was induced in FVB mice with multiple low-dose streptozotocin. Hyperglycemic and age-matched control mice were treated with and without Zn chelator, N,N,N',N'-tetrakis (2-pyridylemethyl) ethylenediamine (TPEN), at 5 mg/kg body-weight daily for 4 months. Hepatic injury was examined by serum alanine aminotransferase (ALT) level and liver histopathological and biochemical changes. Hepatic Zn deficiency (lower than control level, p<0.05) was seen in the mice with either diabetes or TPEN treatment and more evident in the mice with both diabetes and TPEN. Zn deficiency exacerbated hepatic injuries, shown by further increased serum ALT, hepatic lipid accumulation, inflammation, oxidative damage, and endoplasmic reticulum stress-related cell death in Diabetes/TPEN group compared to Diabetes alone. Diabetes/TPEN group also showed a significant decrease in nuclear factor-erythroid 2-related factor 2 (Nrf2) expression and transcription action along with significant increases in Akt negative regulators, decrease in Akt and GSK-3β phosphorylation, and increase in nuclear accumulation of Fyn (a Nrf2 negative regulator). In vitro study with HepG2 cells showed that apoptotic effect of TPEN at 0.5-1.0 µM could be completely prevented by simultaneous Zn supplementation at the dose range of 30-50 µM. Zn is required for maintaining Akt activation by inhibiting the expression of Akt negative regulators; Akt activation can inhibit Fyn nuclear translocation to export nuclear Nrf2 to cytoplasm for degradation. Zn deficiency significantly enhanced diabetes-induced hepatic injury likely through down-regulation of Nrf2 function.

Airborne nitro-PAHs induce Nrf2/ARE defense system against oxidative stress and promote inflammatory process by activating PI3K/Akt pathway in A549 cells.

PubMed

Shang, Yu; Zhou, Qian; Wang, Tiantian; Jiang, Yuting; Zhong, Yufang; Qian, Guangren; Zhu, Tong; Qiu, Xinghua; An, Jing

2017-10-01

Ambient particulate matter (PM) is a worldwide health issue of concern. However, limited information is available regarding the toxic contributions of the nitro-derivatives of polycyclic aromatic hydrocarbons (nitro-PAHs). This study intend to examine whether 1-nitropyrene (1-NP) and 3-nitrofluoranthene (3-NF) could activate the nuclear factor-erythroid 2-related factor 2/antioxidant response element (Nrf2/ARE) antioxidant defense system, and whether the phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) pathway participates in regulating pro-inflammatory responses in A549 cells. Firstly, 1-NP and 3-NF concentration-dependently induced cellular apoptosis, reactive oxygen species (ROS) generation, DNA damage, S phase cell cycle arrest and differential expression of related cytokine genes. Secondly, 1-NP and 3-NF activated the Nrf2/ARE defense system, as evidenced by increased protein expression levels and nuclear translocation of transcription factor Nrf2, elevated Nrf2/ARE binding activity, up-regulated expression of the target gene heme oxygenase-1 (HO-1). Significantly increased protein expression of DNA-dependent protein kinase catalytic subunit (DNA-PKcs) and phosphorylation level of Akt indicated that the PI3K/Akt pathway was activated during pro-inflammatory process. Further, both PI3K inhibitor (LY294002) and Akt inhibitor (MK-2206) reversed the elevated TNF-α expression to control level. Our results suggested that Nrf2/ARE pathway activation might cause an initiation step in cellular protection against oxidative stress caused by nitro-PAHs, and the PI3K/Akt pathway participated in regulating inflammatory responses. Copyright © 2017 Elsevier Ltd. All rights reserved.

Flavone-resistant Leishmania donovani Overexpresses LdMRP2 Transporter in the Parasite and Activates Host MRP2 on Macrophages to Circumvent the Flavone-mediated Cell Death*

PubMed Central

Chowdhury, Sayan; Mukhopadhyay, Rupkatha; Saha, Sourav; Mishra, Amartya; Sengupta, Souvik; Roy, Syamal; Majumder, Hemanta K.

2014-01-01

In parasites, ATP-binding cassette (ABC) transporters represent an important family of proteins related to drug resistance and other biological activities. Resistance of leishmanial parasites to therapeutic drugs continues to escalate in developing countries, and in many instances, it is due to overexpressed ABC efflux pumps. Progressively adapted baicalein (BLN)-resistant parasites (pB25R) show overexpression of a novel ABC transporter, which was classified as ABCC2 or Leishmania donovani multidrug resistance protein 2 (LdMRP2). The protein is primarily localized in the flagellar pocket region and in internal vesicles. Overexpressed LdABCC2 confers substantial BLN resistance to the parasites by rapid drug efflux. The BLN-resistant promastigotes when transformed into amastigotes in macrophage cells cannot be cured by treatment of macrophages with BLN. Amastigote resistance is concomitant with the overexpression of macrophage MRP2 transporter. Reporter analysis and site-directed mutagenesis assays demonstrated that antioxidant response element 1 is activated upon infection. The expression of this phase II detoxifying gene is regulated by NFE2-related factor 2 (Nrf2)-mediated antioxidant response element activation. In view of the fact that the signaling pathway of phosphoinositol 3-kinase controls microfilament rearrangement and translocation of actin-associated proteins, the current study correlates with the intricate pathway of phosphoinositol 3-kinase-mediated nuclear translocation of Nrf2, which activates MRP2 expression in macrophages upon infection by the parasites. In contrast, phalloidin, an agent that prevents depolymerization of actin filaments, inhibits Nrf2 translocation and Mrp2 gene activation by pB25R infection. Taken together, these results provide insight into the mechanisms by which resistant clinical isolates of L. donovani induce intracellular events relevant to drug resistance. PMID:24706751

Chemopreventive effects of Furan-2-yl-3-pyridin-2-yl-propenone against 7,12-dimethylbenz[a]anthracene-inducible genotoxicity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hwang, Yong Pil; Han, Eun Hee; Choi, Jae Ho

2008-05-01

1-Furan-2-yl-3-pyridin-2-yl-propenone (FPP-3) is an anti-inflammatory agent with a propenone moiety and chemically synthesized recently. In this study, we examined the chemopreventive effect of FPP-3 on 7,12-dimethylbenz[a]anthracene (DMBA)-induced genotoxicity in MCF-7 cells. FPP-3 reduced the formation of the DMBA-DNA adduct. DMBA-induced CYP1A1 and CYP1B1 gene expression and enzyme activity were inhibited by FPP-3. It inhibited DMBA-induced aryl hydrocarbon receptor (AhR) transactivation and DMBA-inducible nuclear localization of the AhR. Induction of detoxifying phase II genes by chemopreventive agents represents a coordinated protective response against oxidative stress and neoplastic effects of carcinogens. Transcription factor NF-E2 related factor 2 (Nrf2) regulates antioxidant response elementmore » (ARE) of phase II detoxifying and antioxidant enzymes, such as glutathione S-transferase (GST) and NAD(P)H:quinone oxidoreductase (QR). FPP-3 increased the expression and enzymatic activity of GST and QR. Moreover, FPP-3 increased transcriptional activity of GST and QR. GST and QR induction and Nrf2 translocation by FPP-3 were blocked by the PKC inhibitor Goe6983, and the p38 inhibitor SB203580. These results reflected a partial role of PKC{delta} and p38 signaling in FPP-3-mediated GSTA and QR induction through nuclear translocation of Nrf2. Classically, chemopreventive agents either inhibit CYP metabolizing enzyme or induce phase II detoxifying enzymes. These results suggest that FPP-3 has a potent protective effect against DMBA-induced genotoxicity through modulating phase I and II enzymes and that it has potential as a chemopreventive agent.« less

Anti-apoptotic effect of phloretin on cisplatin-induced apoptosis in HEI-OC1 auditory cells.

PubMed

Choi, Byung-Min; Chen, Xiao Yan; Gao, Shang Shang; Zhu, Rizhe; Kim, Bok-Ryang

2011-01-01

Cisplatin is a highly effective chemotherapeutic agent, but it has significant ototoxic side effects. Apoptosis is an important mechanism of cochlear hair cell loss following exposure to cisplatin. The present study examined the effects of phloretin, a natural polyphenolic compound found in apples and pears, on cisplatin-induced apoptosis. We found that phloretin induced the expression of heme oxygenase-1 (HO-1) protein in a concentration- and time-dependent manner. Phloretin induced nuclear factor-E2-related factor 2 (Nrf2) nuclear translocation, and dominant-negative Nrf2 attenuated phloretin-induced expression of HO-1. Phloretin activated the JNK, ERK and p38 mitogen-activated protein kinase (MAPK) pathways, and the JNK pathway played an important role in phloretin-induced HO-1 expression. Phloretin protected the cells against cisplatin-induced apoptosis. The protective effect of phloretin was abrogated by zinc protoporphyrin IX (ZnPP IX), a HO inhibitor. Furthermore, phloretin pretreatment inhibited mitochondrial dysfunction and the activation of caspases. These results demonstrate that the expression of HO-1 induced by phloretin is mediated by both the JNK pathway and Nrf2; the expression inhibits cisplatin-induced apoptosis in HEI-OC1 cells.

Nrf2 protects photoreceptor cells from photo-oxidative stress induced by blue light.

PubMed

Chen, Wan-Ju; Wu, Caiying; Xu, Zhenhua; Kuse, Yoshiki; Hara, Hideaki; Duh, Elia J

2017-01-01

Oxidative stress plays a key role in age-related macular degeneration and hereditary retinal degenerations. Light damage in rodents has been used extensively to model oxidative stress-induced photoreceptor degeneration, and photo-oxidative injury from blue light is particularly damaging to photoreceptors. The endogenous factors protecting photoreceptors from oxidative stress, including photo-oxidative stress, are continuing to be elucidated. In this study, we evaluated the effect of blue light exposure on photoreceptors and its relationship to Nrf2 using cultured murine photoreceptor (661W) cells. 661W cells were exposed to blue light at 2500 lux. Exposure to blue light for 6-24 h resulted in a significant increase in intracellular reactive oxygen species (ROS) and death of 661W cells in a time-dependent fashion. Blue light exposure resulted in activation of Nrf2, as indicated by an increase in nuclear translocation of Nrf2. This was associated with a significant induction of expression of Nrf2 as well as an array of Nrf2 target genes, including antioxidant genes, as indicated by quantitative reverse transcription PCR (qRT-PCR). In order to determine the functional role of Nrf2, siRNA-mediated knockdown studies were performed. Nrf2-knockdown in 661W cells resulted in significant exacerbation of blue light-induced reactive oxygen species levels as well as cell death. Taken together, these findings indicate that Nrf2 is an important endogenous protective factor against oxidative stress in photoreceptor cells. This suggests that drugs targeting Nrf2 could be considered as a neuroprotective strategy for photoreceptors in AMD and other retinal conditions. Copyright © 2016 Elsevier Ltd. All rights reserved.

Astrocyte NMDA receptors' activity sustains neuronal survival through a Cdk5–Nrf2 pathway

PubMed Central

Jimenez-Blasco, D; Santofimia-Castaño, P; Gonzalez, A; Almeida, A; Bolaños, J P

2015-01-01

Neurotransmission unavoidably increases mitochondrial reactive oxygen species. However, the intrinsic antioxidant defense of neurons is weak and hence the mechanism whereby these cells are physiologically protected against oxidative damage is unknown. Here we found that the antioxidant defense of neurons is repressed owing to the continuous protein destabilization of the master antioxidant transcriptional activator, nuclear factor-erythroid 2-related factor-2 (Nrf2). By contrast, Nrf2 is highly stable in neighbor astrocytes explaining their robust antioxidant defense and resistance against oxidative stress. We also show that subtle and persistent stimulation of N-methyl-d-aspartate receptors (NMDAR) in astrocytes, through a mechanism not requiring extracellular Ca2+ influx, upregulates a signal transduction pathway involving phospholipase C-mediated endoplasmic reticulum release of Ca2+ and protein kinase Cδ activation. Active protein kinase Cδ promotes, by phosphorylation, the stabilization of p35, a cyclin-dependent kinase-5 (Cdk5) cofactor. Active p35/Cdk5 complex in the cytosol phosphorylates Nrf2 at Thr395, Ser433 and Thr439 that is sufficient to promote Nrf2 translocation to the nucleus and induce the expression of antioxidant genes. Furthermore, this Cdk5–Nrf2 transduction pathway boosts glutathione metabolism in astrocytes efficiently protecting closely spaced neurons against oxidative damage. Thus, intercellular communication through NMDAR couples neurotransmission with neuronal survival. PMID:25909891

Akt and ERK/Nrf2 activation by PUFA oxidation-derived aldehydes upregulates FABP4 expression in human macrophages.

PubMed

Lázaro, Iolanda; Ferré, Raimon; Masana, Lluís; Cabré, Anna

2013-10-01

In macrophages, adipocyte fatty acid-binding protein (FABP4) coordinates key events in oxidized LDL-induced foam cell formation, such as cholesterol trafficking and inflammatory responses. Nrf2 is a redox-sensitive transcription factor with antioxidant and anti-inflammatory properties. We investigated the involvement of the Nrf2 signaling pathway in FABP4-upregulation in response to aldehydes that are derived from polyunsaturated fatty acid (PUFA) oxidation. Using RT-PCR and western blotting, we found that the aldehyde 2,4-decadienal (2,4-DDE) produced a marked increase in FABP4 mRNA and protein levels. 2,4-DDE acts at the transcriptional level of FABP4 by promoting mRNA synthesis and prolonging the half-life of the de novo synthesized mRNA. 2,4-DDE consistently enhanced nuclear translocation of phosphorylated Nrf2, which was mediated by the activation of the Akt and ERK signaling pathways. A chromatin immunoprecipitation assay showed the in vivo binding of activated Nrf2 to a newly identified ARE site in the human FABP4 promoter. We propose an Akt and ERK/Nrf2-dependent FABP4 upregulation pathway in response to PUFA oxidation end-products in human macrophages. These results open a new avenue for putative therapeutic targets addressed to control atherogenesis. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Lycopene inhibits NF-κB activation and adhesion molecule expression through Nrf2-mediated heme oxygenase-1 in endothelial cells.

PubMed

Yang, Po-Min; Chen, Huang-Zhi; Huang, Yu-Ting; Hsieh, Chia-Wen; Wung, Being-Sun

2017-06-01

The endothelial expression of cell adhesion molecules plays a leading role in atherosclerosis. Lycopene, a carotenoid with 11 conjugated double bonds, has been shown to have anti-inflammatory properties. In the present study, we demonstrate a putative mechanism for the anti-inflammatory effects of lycopene. We demonstrate that lycopene inhibits the adhesion of tumor necrosis factor α (TNFα)-stimulated monocytes to endothelial cells and suppresses the expression of intercellular cell adhesion molecule-1 (ICAM-1) at the transcriptional level. Moreover, lycopene was found to exert its inhibitory effects by blocking the degradation of the inhibitory protein, IκBα, following 6 h of pre-treatment. In TNFα-stimulated endothelial cells, nuclear factor-κB (NF-κB) nuclear translocation and transcriptional activity were abolished by up to 12 h of lycopene pre-treatment. We also found that lycopene increased the intracellular glutathione (GSH) level and glutamate-cysteine ligase expression. Subsequently, lycopene induced nuclear factor-erythroid 2 related factor 2 (Nrf2) activation, leading to the increased expression of downstream of heme oxygenase-1 (HO-1). The use of siRNA targeting HO-1 blocked the inhibitory effects of lycopene on IκB degradation and ICAM-1 expression. The inhibitory effects of lycopene thus appear to be mediated through its induction of Nrf2-mediated HO-1 expression. Therefore, the findings of the present study indicate that lycopene suppresses the activation of TNFα-induced signaling pathways through the upregulation of Nrf2-mediated HO-1 expression.

Pterostilbene protects against UVB-induced photo-damage through a phosphatidylinositol-3-kinase-dependent Nrf2/ARE pathway in human keratinocytes.

PubMed

Li, Huaping; Jiang, Na; Liang, Bihua; Liu, Qing; Zhang, Erting; Peng, Liqian; Deng, Huiyan; Li, Runxiang; Li, Zhenjie; Zhu, Huilan

2017-11-01

Ultraviolet B (UVB) irradiation is the initial etiological factor for various skin disorders, including erythema, sunburn, photoaging, and photocarcinogenesis. Pterostilbene (Pter) displayed remarkable antioxidant, anti-inflammatory, and anticarcinogenic activities. This study aimed to investigate the effective mechanism of Pter against UVB-induced photodamage in immortalized human keratinocytes. Human keratinocytes were pretreated with Pter (5 and 10 μM) for 24 h prior to UVB irradiation (300 mJ/cm 2 ). Harvested cells were analyzed by MTT, DCFH-DA, comet, western blotting, luciferase promoter, small interference RNA transfection, and quantitative real-time polymerase chain reaction assay. Pter significantly attenuated UVB-induced cell death and reactive oxygen species (ROS) generation, and effectively increased nuclear translocation of NF-E2-related factor-2 (Nrf2), expression of Nrf2-dependent antioxidant enzymes, and DNA repair activity. Moreover, the protective effects of Pter were abolished by small interference RNA-mediated Nrf2 silencing. Furthermore, Pter was also found to induce the phosphorylation of Nrf2 and the known phosphatidylinositol-3-kinase (PI3K) phosphorylated kinase, Akt. The specific inhibitor of PI3K, LY294002, successfully abrogated Pter-induced Nrf2 phosphorylation, activation of Nrf2-antioxidant response element pathway, ROS scavenging ability, and DNA repair activity. The present study indicated that Pter effectively protected against UVB-induced photodamage by increasing endogenous defense mechanisms, scavenging UVB-induced ROS, and aiding in damaged DNA repair through a PI3K-dependent activation of Nrf2/ARE pathway.

Liquiritigenin and liquiritin alleviated MCT-induced HSOS by activating Nrf2 antioxidative defense system.

PubMed

Huang, Zhenlin; Sheng, Yuchen; Chen, Minwei; Hao, Zhanxia; Hu, Feifei; Ji, Lili

2018-06-14

Hepatic sinusoidal obstruction syndrome (HSOS) is a serious and life-threatening liver disease. Liquiritigenin (LG) and liquiritin (LQ) are natural flavonoids distributed in Glycyrrhizae Radix et Rhizoma (Gan-cao). This study aims to investigate the protective effect and mechanism of LG and LQ against monocrotaline (MCT)-induced HSOS. Results of serum alanine/aspartate aminotransferases (ALT/AST) activities, liver histological evaluation and scanning electron microscope observation, and hepatic metalloproteinase-9 (MMP-9) expression demonstrated that LG and LQ both alleviated HSOS induced by MCT in rats. Results of hepatic reactive oxygen species (ROS), malondialdehyde (MDA), 4-hydroxynonenal (4-HNE), oxidized glutathione (GSSG) and reduced glutathione (GSH) contents, glutathione reductase (GR) and superoxide dismutase (SOD) activities showed that LG and LQ attenuated MCT-induced liver oxidative stress injury. Furthermore, LG and LQ were found to promote Nrf2 nuclear translocation and lead to the increased expression of Nrf2 downstream antioxidative genes. Molecule docking analysis indicated the potential interaction of LG and LQ with Nrf2 binding site in the kelch-like ECH-associated protein-1 (Keap1) protein. Finally, Nrf2 knock-out mice were used. The results showed that LG and LQ both alleviated MCT-induced HSOS in wild-type mice, but such protection was totally diminished in Nrf2 knock-out mice. In conclusion, our study revealed that LG and LQ alleviated MCT-induced HSOS by inducing the activation of hepatic Nrf2 antioxidative defense system. Copyright © 2018. Published by Elsevier Inc.

Tartary buckwheat flavonoids ameliorate high fructose-induced insulin resistance and oxidative stress associated with the insulin signaling and Nrf2/HO-1 pathways in mice.

PubMed

Hu, Yuanyuan; Hou, Zuoxu; Yi, Ruokun; Wang, Zhongming; Sun, Peng; Li, Guijie; Zhao, Xin; Wang, Qiang

2017-08-01

The present study was conducted to explore the effects of a purified tartary buckwheat flavonoid fraction (TBF) on insulin resistance and hepatic oxidative stress in mice fed high fructose in drinking water (20%) for 8 weeks. The results indicated that continuous administration of TBF dose-dependently improved the insulin sensitivity and glucose intolerance in high fructose-fed mice. TBF treatment also reversed the reduced level of insulin action on the phosphorylation of insulin receptor substrate-1 (IRS-1), protein kinase B (Akt) and phosphatidylinositol 3-kinase (PI3K), as well as the translocation of glucose transporter type 4 (GLUT4) in the insulin-resistant liver. Furthermore, TBF was found to exert high antioxidant capacity as it acts as a shield against oxidative stress induced by high fructose by restoring the antioxidant status, and modulating nuclear factor E2 related factor 2 (Nrf2) translocation to the nucleus with subsequently up-regulated antioxidative enzyme protein expression. Histopathological examinations revealed that impaired pancreatic/hepatic tissues were effectively restored in high fructose-fed mice following TBF treatment. Our results show that TBF intake is effective in preventing the conversion of high fructose-induced insulin resistance and hepatic oxidative stress in mice by improving the insulin signaling molecules and the Nrf2 signal pathway in the liver.

Procyanidin B2 ameliorates carrageenan-induced chronic nonbacterial prostatitis in rats via anti-inflammatory and activation of the Nrf2 pathway.

PubMed

Wang, Wei; Chen, Renzong; Wang, Jiye

2017-11-04

Prostatitis is one of the most prevalent problems in andriatry and urinary surgery. In the present study, we evaluated the effect of procyanidin B2 (PB) on carrageenan-induced chronic nonbacterial prostatitis in rats. Results showed that PB significantly decreased the prostatic index and enhanced the body weight inhibited by carrageenan. Biochemical results revealed that PB significantly lowered the prostatic specific antigen (PSA) and alleviated oxidative stress in serum. The levels of TNF-α, IL-6, and IL-10 in prostatic homogenate were also significantly decreased after PB treatment. We also found evidence that PB treatment reversed the suppression of Nrf2 nuclear translocation, and increased the expressions of NQO1 and HO-1 in the prostate glands. In conclusion, treatment with PB attenuates carrageenan-induced chronic nonbacterial prostatitis via anti-inflammatory and activation of the Nrf2 pathway. Copyright © 2017 Elsevier Inc. All rights reserved.

Triterpenic acids-enriched fraction from Cyclocarya paliurus attenuates non-alcoholic fatty liver disease via improving oxidative stress and mitochondrial dysfunction.

PubMed

Zhao, Meng-Ge; Sheng, Xue-Ping; Huang, Ya-Ping; Wang, Yi-Ting; Jiang, Cui-Hua; Zhang, Jian; Yin, Zhi-Qi

2018-08-01

The effects of triterpenic acids-enriched fraction from Cyclocarya paliurus (CPT) on nonalcoholic fatty liver disease (NAFLD) were investigated using in vivo and in vitro models. In high fat diet-induced Wister rats, CPT significantly increased superoxide dismutase (SOD) activity and glutathione/oxidized glutathione (GSH/GSSG) ratio, reduced malondialdehyde (MDA) and protein carbonyl (PCO) levels. Moreover, CPT restored mitochondrial membrane potential dysfunction, decreased cytochrome P450 enzyme 2E1 (CYP2E1) activity, improved nuclear factor erythroid 2-related factor 2 (Nrf2) and Nrf2-mediated antioxidant enzyme heme oxygenase1 (HO-1) expression. In free fatty acids-induced HepG2 cells, CPT dramatically decreased ROS content, increased mitochondrial NADH dehydrogenase (Complex I) and mitochondrial cytochrome C oxidase (Complex IV) levels. Furthermore, CPT could upregulate HO-1, quinine oxidoreductase 1 (NQO1) expression, and increase Nrf2 translocation from cytoplasm-to-nucleus. The results indicated CPT could protect mitochondria function and improve oxidative stress by activating Nrf2. Therefore, it can be inferred that CPT may be a potential agent against NAFLD. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Nitro-oleic acid ameliorates oxygen and glucose deprivation/re-oxygenation triggered oxidative stress in renal tubular cells via activation of Nrf2 and suppression of NADPH oxidase.

PubMed

Nie, Huibin; Xue, Xia; Liu, Gang; Guan, Guangju; Liu, Haiying; Sun, Lina; Zhao, Long; Wang, Xueling; Chen, Zhixin

2016-01-01

Nitroalkene derivative of oleic acid (OA-NO 2 ), due to its ability to mediate revisable Michael addition, has been demonstrated to have various biological properties and become a therapeutic agent in various diseases. Though its antioxidant properties have been reported in different models of acute kidney injury (AKI), the mechanism by which OA-NO 2 attenuates intracellular oxidative stress is not well investigated. Here, we elucidated the anti-oxidative mechanism of OA-NO 2 in an in vitro model of renal ischemia/reperfusion (I/R) injury. Human tubular epithelial cells were subjected to oxygen and glucose deprivation/re-oxygenation (OGD/R) injury. Pretreatment with OA-NO 2 (1.25 μM, 45 min) attenuated OGD/R triggered reactive oxygen species (ROS) generation and subsequent mitochondrial membrane potential disruption. This action was mediated via up-regulating endogenous antioxidant defense components including superoxide dismutase (SOD1), heme oxygenase 1 (HO-1), and γ-glutamyl cysteine ligase modulatory subunits (GCLM). Moreover, subcellular fractionation analyses demonstrated that OA-NO 2 promoted nuclear translocation of nuclear factor-E2- related factor-2 (Nrf2) and Nrf2 siRNA partially abrogated these protective effects. In addition, OA-NO 2 inhibited NADPH oxidase activation and NADPH oxidase 4 (NOX4), NADPH oxidase 2 (NOX2) and p22 phox up-regulation after OGD/R injury, which was not relevant to Nrf2. These results contribute to clarify that the mechanism of OA-NO 2 reno-protection involves both inhibition of NADPH oxidase activity and induction of SOD1, Nrf2-dependent HO-1, and GCLM.

Quercetin Reduces Tumor Necrosis Factor Alpha-Induced Muscle Atrophy by Upregulation of Heme Oxygenase-1.

PubMed

Kim, Yeji; Kim, Chu-Sook; Joe, Yeonsoo; Chung, Hun Taeg; Ha, Tae Youl; Yu, Rina

2018-06-01

The inflammatory cytokine tumor necrosis factor α (TNFα), upregulated in the obese condition, promotes protein degradation and is implicated in obesity-related skeletal muscle atrophy and age-related sarcopenia. Quercetin, a flavonoid, elicits antioxidative and anti-inflammatory activities. In this study, we investigated the effect of quercetin on TNFα-induced skeletal muscle atrophy as well as its potential mechanism of action. In this study, we observed that quercetin suppressed expression of TNFα-induced atrophic factors such as MAFbx/atrogin-1 and MuRF1 in myotubes, and it enhanced heme oxygenase-1 (HO-1) protein level accompanied by increased nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) in myotubes. The HO-1 inhibitor ZnPP suppressed the inhibitory actions of quercetin on TNFα-induced atrophic responses and degradation of IκB-α in myotubes. Moreover, quercetin supplementation to high-fat diet-fed obese mice inhibited obesity-induced atrophic responses in skeletal muscle, accompanied by upregulation of HO-1 and inactivation of nuclear factor-kappa B (NF-κB), and the quercetin actions were attenuated in Nrf2-deficient mice. These findings suggest that quercetin protects against TNFα-induced muscle atrophy under obese conditions through Nrf2-mediated HO-1 induction accompanied by inactivation of NF-κB. Quercetin may be used as a dietary supplement to protect against obesity-induced skeletal muscle atrophy.

The Protective Effect of Glycyrrhetinic Acid on Carbon Tetrachloride-Induced Chronic Liver Fibrosis in Mice via Upregulation of Nrf2

PubMed Central

Chen, Shaoru; Zou, Liyi; Li, Li; Wu, Tie

2013-01-01

This study was designed to investigate the potentially protective effects of glycyrrhetinic acid (GA) and the role of transcription factor nuclear factor-erythroid 2(NF-E2)-related factor 2 (Nrf2) signaling in the regulation of Carbon Tetrachloride (CCl4)-induced chronic liver fibrosis in mice. The potentially protective effects of GA on CCl4-induced chronic liver fibrosis in mice were depicted histologically and biochemically. Firstly, histopathological changes including regenerative nodules, inflammatory cell infiltration and fibrosis were induced by CCl4.Then, CCl4 administration caused a marked increase in the levels of serum aminotransferases (GOT, GPT), serum monoamine oxidase (MAO) and lipid peroxidation (MDA) as well as MAO in the mice liver homogenates. Also, decreased nuclear Nrf2 expression, mRNA levels of its target genes such as superoxide dismutase 3 (SOD3), catalase (CAT), glutathione peroxidase 2 (GPX2), and activity of cellular antioxidant enzymes were found after CCl4 exposure. All of these phenotypes were markedly reversed by the treatment of the mice with GA. In addition, GA exhibited the antioxidant effects in vitro by on FeCl2-ascorbate induced lipid peroxidation in mouse liver homogenates, and on DPPH scavenging activity. Taken together, these results suggested that GA can protect the liver from oxidative stress in mice, presumably through activating the nuclear translocation of Nrf2, enhancing the expression of its target genes and increasing the activity of the antioxidant enzymes. Therefore, GA may be an effective hepatoprotective agent and viable candidate for treating liver fibrosis and other oxidative stress-related diseases. PMID:23341968

Berberine activates Nrf2 nuclear translocation and inhibits apoptosis induced by high glucose in renal tubular epithelial cells through a phosphatidylinositol 3-kinase/Akt-dependent mechanism.

PubMed

Zhang, Xiuli; Liang, Dan; Lian, Xu; Jiang, Yan; He, Hui; Liang, Wei; Zhao, Yue; Chi, Zhi-Hong

2016-06-01

Apoptosis of tubular epithelial cells is a major feature of diabetic kidney disease, and hyperglycemia triggers the generation of free radicals and oxidant stress in tubular cells. Berberine (BBR) is identified as a potential anti-diabetic herbal medicine due to its beneficial effects on insulin sensitivity, glucose metabolism and glycolysis. In this study, the underlying mechanisms involved in the protective effects of BBR on high glucose-induced apoptosis were explored using cultured renal tubular epithelial cells (NRK-52E cells) and human kidney proximal tubular cell line (HK-2 cells). We identified the pivotal role of phosphatidylinositol 3-kinase (PI3K)/Akt in BBR cellular defense mechanisms and revealed the novel effect of BBR on nuclear factor (erythroid-derived 2)-related factor-2 (Nrf2) and heme oxygenase (HO)-1 in NRK-52E and HK-2 cells. BBR attenuated reactive oxygen species production, antioxidant defense (GSH and SOD) and oxidant-sensitive proteins (Nrf2 and HO-1), which also were blocked by LY294002 (an inhibitor of PI3K) in HG-treated NRK-52E and HK-2 cells. Furthermore, BBR improved mitochondrial function by increasing mitochondrial membrane potential. BBR-induced anti-apoptotic function was demonstrated by decreasing apoptotic proteins (cytochrome c, Bax, caspase3 and caspase9). All these findings suggest that BBR exerts the anti-apoptosis effects through activation of PI3K/Akt signal pathways and leads to activation of Nrf2 and induction of Nrf2 target genes, and consequently protecting the renal tubular epithelial cells from HG-induced apoptosis.

The Flavones Apigenin and Luteolin Induce FOXO1 Translocation but Inhibit Gluconeogenic and Lipogenic Gene Expression in Human Cells

PubMed Central

Bumke-Vogt, Christiane; Osterhoff, Martin A.; Borchert, Andrea; Guzman-Perez, Valentina; Sarem, Zeinab; Birkenfeld, Andreas L.; Bähr, Volker; Pfeiffer, Andreas F. H.

2014-01-01

The flavones apigenin (4′,5,7,-trihydroxyflavone) and luteolin (3′,4′,5,7,-tetrahydroxyflavone) are plant secondary metabolites with antioxidant, antiinflammatory, and anticancer activities. We evaluated their impact on cell signaling pathways related to insulin-resistance and type 2 diabetes. Apigenin and luteolin were identified in our U-2 OS (human osteosarcoma) cell screening assay for micronutrients triggering rapid intracellular translocation of the forkhead box transcription factor O1 (FOXO1), an important mediator of insulin signal transduction. Insulin reversed the translocation of FOXO1 as shown by live cell imaging. The impact on the expression of target genes was evaluated in HepG2 (human hepatoma) cells. The mRNA-expression of the gluconeogenic enzymes phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pc), the lipogenic enzymes fatty-acid synthase (FASN) and acetyl-CoA-carboxylase (ACC) were down-regulated by both flavones with smaller effective dosages of apigenin than for luteolin. PKB/AKT-, PRAS40-, p70S6K-, and S6-phosphorylation was reduced by apigenin and luteolin but not that of the insulin-like growth factor receptor IGF-1R by apigenin indicating a direct inhibition of the PKB/AKT-signaling pathway distal to the IGF-1 receptor. N-acetyl-L-cysteine did not prevent FOXO1 nuclear translocation induced by apigenin and luteolin, suggesting that these flavones do not act via oxidative stress. The roles of FOXO1, FOXO3a, AKT, sirtuin1 (SIRT1), and nuclear factor (erythroid-derived2)-like2 (NRF2), investigated by siRNA knockdown, showed differential patterns of signal pathways involved and a role of NRF2 in the inhibition of gluconeogenic enzyme expression. We conclude that these flavones show an antidiabetic potential due to reduction of gluconeogenic and lipogenic capacity despite inhibition of the PKB/AKT pathway which justifies detailed investigation in vivo. PMID:25136826

Effects of monascin on anti-inflammation mediated by Nrf2 activation in advanced glycation end product-treated THP-1 monocytes and methylglyoxal-treated wistar rats.

PubMed

Lee, Bao-Hong; Hsu, Wei-Hsuan; Huang, Tao; Chang, Yu-Ying; Hsu, Ya-Wen; Pan, Tzu-Ming

2013-02-13

Hyperglycemia is associated with advanced glycation end products (AGEs). This study was designed to evaluate the inhibitory effects of monascin on receptor for advanced glycation end product (RAGE) signal and THP-1 monocyte inflammation after treatment with S100b, a specific ligand of RAGE. Monascin inhibited cytokine production by S100b-treated THP-1 monocytes via up-regulation of nuclear factor-erythroid 2-related factor-2 (Nrf2) and alleviated p47phox translocation to the membrane. Methylglyoxal (MG, 600 mg/kg bw) was used to induce diabetes in Wistar rats. Inhibitions of RAGE and p47phox by monascin were confirmed by peripheral blood mononuclear cells (PBMCs) of MG-induced rats. Silymarin (SM) was used as a positive control group. It was found that monascin promoted heme oxygenase-1 (HO-1) expression mediated by Nrf2. Suppressions of AGEs, tumor necrosis factor-α (TNF-α), and interleukin-1β (IL-β) in serum of MG-induced rats were attenuated in the monascin administration group treated with retinoic acid (RA). RA treatment resulted in Nrf2 inactivation by increasing RA receptor-α (RARα) activity, suggesting that RA acts as an inhibitor of Nrf2. The results showed that monascin exerted anti-inflammatory and antioxidative effects mediated by Nrf2 to prevent the development of diseases such as type 2 diabetes caused by inflammation.

Hemistepsin A ameliorates acute inflammation in macrophages via inhibition of nuclear factor-κB and activation of nuclear factor erythroid 2-related factor 2.

PubMed

Kim, Jae Kwang; Lee, Ji Eun; Jung, Eun Hye; Jung, Ji Yun; Jung, Dae Hwa; Ku, Sae Kwang; Cho, Il Je; Kim, Sang Chan

2018-01-01

Hemistepsin A (HsA) is a sesquiterpene lactone isolated from Hemistepta lyrata (Bunge) Bunge. We investigated the anti-inflammatory effects of HsA and sought to determine its mechanisms of action in macrophages. HsA pretreatment inhibited nitric oxide production, and reduced the expression of iNOS and COX-2 in Toll-like receptor ligand-stimulated RAW 264.7 cells. Additionally, HsA decreased the secretion of proinflammatory cytokines in lipopolysaccharide (LPS)-stimulated Kupffer cells as well as in RAW 264.7 cells. HsA inhibited phosphorylation of IKKα/β and degradation of IκBα, resulting in decreased nuclear translocation of nuclear factor-κB (NF-κB) and its transcriptional activity. Moreover, HsA phosphorylated nuclear factor erythroid 2-related factor 2 (Nrf2), increased expression levels of antioxidant genes, and attenuated LPS-stimulated H 2 O 2 production. Phosphorylation of p38 and c-Jun N-terminal kinase was required for HsA-mediated Nrf2 phosphorylation. In a D-galactosamine/LPS-induced liver injury model, HsA ameliorated D-galactosamine/LPS-induced hepatocyte degeneration and inflammatory cells infiltration. Moreover, immunohistochemical analyses using nitrotyrosine, 4-hydroxynonenal, and cleaved poly (ADP-ribose) polymerase antibodies revealed that HsA protected the liver from oxidative stress. Furthermore, HsA reduced the numbers of proinflammatory cytokine-positive cells in hepatic tissues. Thus, these results suggest HsA may be a promising natural product to manage inflammation-mediated tissue injuries through inhibition of NF-κB and activation of Nrf2. Copyright © 2017 Elsevier Ltd. All rights reserved.

Dual-energy precursor and nuclear erythroid-related factor 2 activator treatment additively improve redox glutathione levels and neuron survival in aging and Alzheimer mouse neurons upstream of reactive oxygen species.

PubMed

Ghosh, Debolina; LeVault, Kelsey R; Brewer, Gregory J

2014-01-01

To determine whether glutathione (GSH) loss or increased reactive oxygen species (ROS) are more important to neuron loss, aging, and Alzheimer's disease (AD), we stressed or boosted GSH levels in neurons isolated from aging 3xTg-AD neurons compared with those from age-matched nontransgenic (non-Tg) neurons. Here, using titrating with buthionine sulfoximine, an inhibitor of γ-glutamyl cysteine synthetase (GCL), we observed that GSH depletion increased neuronal death of 3xTg-AD cultured neurons at increasing rates across the age span, whereas non-Tg neurons were resistant to GSH depletion until old age. Remarkably, the rate of neuron loss with ROS did not increase in old age and was the same for both genotypes, which indicates that cognitive deficits in the AD model were not caused by ROS. Therefore, we targeted for neuroprotection activation of the redox sensitive transcription factor, nuclear erythroid-related factor 2 (Nrf2) by 18 alpha glycyrrhetinic acid to stimulate GSH synthesis through GCL. This balanced stimulation of a number of redox enzymes restored the lower levels of Nrf2 and GCL seen in 3xTg-AD neurons compared with those of non-Tg neurons and promoted translocation of Nrf2 to the nucleus. By combining the Nrf2 activator together with the NADH precursor, nicotinamide, we increased neuron survival against amyloid beta stress in an additive manner. These stress tests and neuroprotective treatments suggest that the redox environment is more important for neuron survival than ROS. The dual neuroprotective treatment with nicotinamide and an Nrf2 inducer indicates that these age-related and AD-related changes are reversible. Copyright © 2014 Elsevier Inc. All rights reserved.

Withaferin A induces heme oxygenase (HO-1) expression in endothelial cells via activation of the Keap1/Nrf2 pathway.

PubMed

Heyninck, Karen; Sabbe, Linde; Chirumamilla, Chandra Sekhar; Szarc Vel Szic, Katarzyna; Vander Veken, Pieter; Lemmens, Kristien J A; Lahtela-Kakkonen, Maija; Naulaerts, Stefan; Op de Beeck, Ken; Laukens, Kris; Van Camp, Guy; Weseler, Antje R; Bast, Aalt; Haenen, Guido R M M; Haegeman, Guy; Vanden Berghe, Wim

2016-06-01

Withaferin A (WA), a natural phytochemical derived from the plant Withania somnifera, is a well-studied bioactive compound exerting a broad spectrum of health promoting effects. To gain better insight in the potential therapeutic capacity of WA, we evaluated the transcriptional effects of WA on primary human umbilical vein endothelial cells (HUVECs) and an endothelial cell line (EA.hy926). RNA microarray analysis of WA treated HUVEC cells demonstrated increased expression of the antioxidant gene heme oxygenase (HO-1). Transcriptional regulation of this gene is strongly dependent on the transcription factor NF-E2-related factor 2 (Nrf2), which senses chemical changes in the cell and coordinates transcriptional responses to maintain chemical homeostasis via expression of antioxidant genes and cytoprotective Phase II detoxifying enzymes. Under normal conditions, Nrf2 is kept in the cytoplasm by Kelch-like ECH-associated protein 1 (Keap1), an adaptor protein controlling the half-life of Nrf2 via constant proteasomal degradation. In this study we demonstrate that WA time- and concentration-dependently induces HO-1 expression in endothelial cells via upregulation and increased nuclear translocation of Nrf2. According to the crucial negative regulatory role of Keap1 in Nrf2 expression levels, a direct interaction of WA with Keap1 could be demonstrated. In vitro and in silico evaluations suggest that specific cysteine residues in Keap1 might be involved in the interaction with WA. Copyright © 2016 Elsevier Inc. All rights reserved.

Visualization of Oxidative Stress Induced by Experimental Periodontitis in Keap1-Dependent Oxidative Stress Detector-Luciferase Mice.

PubMed

Kataoka, Kota; Ekuni, Daisuke; Tomofuji, Takaaki; Irie, Koichiro; Kunitomo, Muneyoshi; Uchida, Yoko; Fukuhara, Daiki; Morita, Manabu

2016-11-16

The aim of this study was to investigate whether a Keap1-dependent oxidative stress detector-luciferase (OKD-LUC) mouse model would be useful for the visualization of oxidative stress induced by experimental periodontitis. A ligature was placed around the mandibular first molars for seven days to induce periodontitis. Luciferase activity was measured with an intraperitoneal injection of d-luciferin on days 0, 1, and 7. The luciferase activity in the periodontitis group was significantly greater than that in the control group at seven days. The expressions of heme oxygenase-1 (HO-1) and malondialdehyde in periodontal tissue were significantly higher in the periodontitis group than in the control group. Immunofluorescent analysis confirmed that the nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) occurred more frequently in the periodontitis group than in the control group. This study found that under oxidative stress induced by experimental periodontitis, the Nrf2/antioxidant defense pathway was activated and could be visualized from the luciferase activity in the OKD-LUC model. Thus, the OKD-LUC mouse model may be useful for exploring the mechanism underlying the relationship between the Nrf2/antioxidant defense pathway and periodontitis by enabling the visualization of oxidative stress over time.

Mechanism of Mechanical Trauma-Induced Extracellular Matrix Remodeling of Fibroblasts in Association with Nrf2/ARE Signaling Suppression Mediating TGF-β1/Smad3 Signaling Inhibition

PubMed Central

Liu, Cheng; Li, Qiannan; Wang, Linlin; Min, Jie; Hu, Ming; Hong, Shasha

2017-01-01

Stress urinary incontinence (SUI) is a common hygienic problem affecting the quality of women's life worldwide. In this research, we revealed the involvement and regulation of extracellular matrix (ECM) remodeling, oxidative damage, and TGF-β1 signaling in the pathological mechanisms of mechanical trauma-induced SUI. We found that excessive mechanical strain significantly increased apoptosis rate, decreased cell viability and ECM production, and broke the balance of MMPs/TIMPs compared with the nonstrain control (NC) group. The expression levels of TGFβ1, p-Smad3, Nrf2, GPx1, and CAT were downregulated, the production of ROS, 8-OHdG, 4-HNE, and MDA was increased, and the nuclear translocation of Smad2/3 was suppressed after 5333 μstrain's treatment. Both mTGF-β1 pretreatment and Nrf2 overexpression could reverse mechanical injury-induced TGFβ1/Smad3 signaling inhibition and ECM remodeling, whereas mTGF-β1 had no effect on Nrf2 expression. Nrf2 overexpression significantly alleviated mechanical injury-induced ROS accumulation and oxidative damage; in contrast, Nrf2 silencing exhibited opposite effects. Besides, vaginal distention- (VD-) induced in vivo SUI model was used to confirm the in vitro results; Nrf2 knockout aggravates mechanical trauma-induced LPP reduction, ECM remodeling, oxidative damage, and TGF-β1/Smad3 suppression in mice. Therefore, we deduce that mechanical injury-induced ECM remodeling might be associated with Nrf2/ARE signaling suppression mediating TGF-β1/Smad3 signaling inhibition. This might reflect a new molecular target for SUI researches. PMID:29109834

Lycium barbarum polysaccharide protects human keratinocytes against UVB-induced photo-damage.

PubMed

Li, Huaping; Li, Zhenjie; Peng, Liqian; Jiang, Na; Liu, Qing; Zhang, Erting; Liang, Bihua; Li, Runxiang; Zhu, Huilan

2017-02-01

Ultraviolet B (UVB) irradiation plays a key role in skin damage, which induces oxidative and inflammatory damages, thereby causing photoaging or photocarcinogenesis. Lycium barbarum polysaccharide (LBP), the most biologically active fraction of wolfberry, possesses significant antioxidative and anti-inflammatory effects on multiple tissues. In the present study, the photoprotective effects and potential underlying molecular mechanisms of LBP against UVB-induced photo-damage were investigated in immortalized human keratinocytes (HaCaT cells). The data indicated that pretreatment with LBP significantly attenuated UVB-induced decrease in cell viability, increase in ROS production and DNA damage. LBP also significantly suppressed UVB-induced p38 MAPK activation, and subsequently reversed caspase-3 activation and MMP-9 expression. Notably, LBP was found to induce Nrf2 nuclear translocation and increase the expression of Nrf2-dependent ARE target genes. Furthermore, the protective effects of LBP were abolished by siRNA-mediated Nrf2 silencing. These results showed that the antioxidant LBP could partially protect against UVB irradiation-induced photo-damage through activation of Nrf2/ARE pathway, thereby scavenging ROS and reducing DNA damage, and subsequently suppressing UVB-induced p38 MAP pathway. Thus, LBP can be potentially used for skincare against oxidative damage from environmental insults.

TOP 1 and 2, polysaccharides from Taraxacum officinale, inhibit NFκB-mediated inflammation and accelerate Nrf2-induced antioxidative potential through the modulation of PI3K-Akt signaling pathway in RAW 264.7 cells.

PubMed

Park, Chung Mu; Cho, Chung Won; Song, Young Sun

2014-04-01

Anti-inflammatory and anti-oxidative activities of polysaccharides from Taraxacum officinale (TOP 1 and 2) were analyzed in RAW 264.7 cells. First, lipopolysaccharide (LPS) was applied to identify anti-inflammatory activity of TOPs, which reduced expression of inducible nitric oxide synthase (iNOS) and tumor necrosis factor (TNF)-α. TOPs treatment inhibited phosphorylation of inflammatory transcription factor, nuclear factor (NF)κB, and its upstream signaling molecule, PI3K/Akt. Second, cytoprotective potential of TOPs against oxidative stress was investigated via heme oxygenase (HO)-1 induction. HO-1, one of phase II enzymes shows antioxidative activity, was potently induced by TOPs treatment, which was in accordance with the nuclear translocation of nuclear factor-erythroid 2 p45-related factor 2 (Nrf2). In addition, TOPs treatment phosphorylated PI3K/Akt with slight activation of c-Jun NH2-terminal kinase (JNK). TOPs-mediated HO-1 induction protected macrophage cells from oxidative stress-induced cell death, which was confirmed by SnPP and CoPP (HO-1 inhibitor and inducer, respectively). Consequently, TOPs potently inhibited NFκB-mediated inflammation and accelerated Nrf2-mediated antioxidative potential through the modulation of PI3K/Akt pathway, which would contribute to their promising strategy for novel anti-inflammatory and anti-oxidative agents. Copyright © 2014. Published by Elsevier Ltd.

RS9, a novel Nrf2 activator, attenuates light-induced death of cells of photoreceptor cells and Müller glia cells.

PubMed

Inoue, Yuki; Shimazawa, Masamitsu; Noda, Yasuhiro; Nagano, Ryota; Otsuka, Tomohiro; Kuse, Yoshiki; Nakano, Yukimichi; Tsuruma, Kazuhiro; Nakagami, Yasuhiro; Hara, Hideaki

2017-06-01

The retina is highly sensitive to oxidative stress because of its high consumption of oxygen associated with the phototransductional processes. Recent findings have suggested that oxidative stress is involved in the pathology of age-related macular degeneration, a progressive degeneration of the central retina. A well-known environmental risk factor is light exposure, as excessive and continuous light exposure can damage photoreceptors. Nuclear factor-erythroid 2-related factor 2 (Nrf2) is a transcriptional factor that controls antioxidative responses and phase 2 enzymes. Thus, we hypothesized that RS9, a specific activator of Nrf2, decreases light-induced retinal cell death in vivo and in vitro. Nrf2 was detected in the nucleus of the 661W cells exposed to RS9 and also after light exposure, and the Nrf2-antioxidant response element binding was increased in 661W cells after exposure to RS9. Consequentially, the expression of the phase 2 enzyme's mRNAs of Ho-1, Nqo-1, and Gclm genes was increased in 661W cells after exposure to RS9. Furthermore, RS9 decreased the light-induced death of 661W cells (2500 lux, 24 h), and also reduced the functional damages and the histological degeneration of the nuclei in the outer nuclear layer or the retina in the in vivo studies (8000 lux, 3 h). Heme oxygenase-1 was increased after light exposure, and Nrf2 was translocated into the nucleus after light exposure in vivo. Silencing of Ho-1 reduced the protective effects of RS9 against light-induced death of 661W cells. These findings indicate that RS9 has therapeutic potential for retinal diseases that are aggravated by light exposure. © 2017 International Society for Neurochemistry.

The chalcone compound isosalipurposide (ISPP) exerts a cytoprotective effect against oxidative injury via Nrf2 activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Han, Jae Yun; Cho, Seung Sik; Yang, Ji Hye

2015-08-15

The chalcone compound isosalipurposide (ISPP) has been successfully isolated from the native Korean plant species Corylopsis coreana Uyeki (Korean winter hazel). However, the therapeutic efficacy of ISPP remains poorly understood. This study investigated whether ISPP has the capacity to activate NF-E2-related factor (Nrf2)-antioxidant response element (ARE) signaling and induce its target gene expression, and to determined the protective role of ISPP against oxidative injury of hepatocytes. In HepG2 cells, nuclear translocation of Nrf2 is augmented by ISPP treatment. Consistently, ISPP increased ARE reporter gene activity and the protein levels of glutamate cysteine ligase (GCL) and hemeoxygenase (HO-1), resulting in increasedmore » intracellular glutathione levels. Cells pretreated with ISPP were rescued from tert-butylhydroperoxide-induced reactive oxygen species (ROS) production and glutathione depletion and consequently, apoptotic cell death. Moreover, ISPP ameliorated the mitochondrial dysfunction and apoptosis induced by rotenone which is an inhibitor of complex 1 of the mitochondrial respiratory chain. The specific role of Nrf2 activation by ISPP was demonstrated using an ARE-deletion mutant plasmid and Nrf2-knockout cells. Finally, we observed that extracellular signal-regulated kinase (ERK) and AMP-activated protein kinase (AMPK), but not protein kinase C (PKC)-δ or other mitogen-activated protein kinases (MAPKs), are involved in the activation of Nrf2 by ISPP. Taken together, our results demonstrate that ISPP has a cytoprotective effect against oxidative damage mediated through Nrf2 activation and induction of its target gene expression in hepatocytes. - Highlights: • We investigated the effect of ISPP on Nrf2 activation. • ISPP increased Nrf2 activity and its target gene expression. • ISPP inhibited the mitochondrial dysfunction and ROS production. • Nrf2 activation by ISPP is dependent on ERK1/2 and AMPK phosphorylation. • ISPP may be a promising therapeutics for the oxidative stress-mediated liver disease.« less

2,4,6-Trichlorophenol cytotoxicity involves oxidative stress, endoplasmic reticulum stress, and apoptosis.

PubMed

Zhang, Xiaoning; Zhang, Xiaona; Niu, Zhidan; Qi, Yongmei; Huang, Dejun; Zhang, Yingmei

2014-01-01

This study aims to evaluate the cytotoxicity and potential mechanisms of 2,4,6-trichlorophenol (2,4,6-TCP) in mouse embryonic fibroblasts. Our results show that 2,4,6-TCP causes morphological changes and reduces cell viability. The overproduction of reactive oxygen species, the upregulation of nuclear factor-E2-related factor 2 (Nrf2) and heme oxygenase 1 (HMOX1) messenger RNA (mRNA) expressions, and the nuclear translocation of Nrf2 protein demonstrate that 2,4,6-TCP induces oxidative stress, and the Nrf2/HMOX1 pathway might be involved in 2,4,6-TCP-induced antioxidative response. Simultaneously, our data also demonstrate that 2,4,6-TCP upregulates the expressions of binding immunoglobulin protein, inositol-requiring enzyme/endonuclease 1α, and C/EBP homologous protein; stimulates α subunit of eukaryotic translation initiation factor 2 phosphorylation; and induces the splicing of Xbp1 mRNA, suggesting that endoplasmic reticulum (ER) stress is triggered. Moreover, 2,4,6-TCP alters the mitochondrial membrane potential and increases the apoptosis rate, the caspase 3 activity, and the Bax/Bcl-2 ratio, demonstrating that the mitochondrial pathway is involved in the 2,4,6-TCP-induced apoptosis. Thus, these results show that 2,4,6-TCP induces oxidative stress, ER stress, and apoptosis, which together contribute to its cytotoxicity in vitro. © The Author(s) 2014.

Sulforaphane Inhibits Lipopolysaccharide-Induced Inflammation, Cytotoxicity, Oxidative Stress, and miR-155 Expression and Switches to Mox Phenotype through Activating Extracellular Signal-Regulated Kinase 1/2-Nuclear Factor Erythroid 2-Related Factor 2/Antioxidant Response Element Pathway in Murine Microglial Cells.

PubMed

Eren, Erden; Tufekci, Kemal Ugur; Isci, Kamer Burak; Tastan, Bora; Genc, Kursad; Genc, Sermin

2018-01-01

Sulforaphane (SFN) is a natural product with cytoprotective, anti-inflammatory, and antioxidant effects. In this study, we evaluated the mechanisms of its effects on lipopolysaccharide (LPS)-induced cell death, inflammation, oxidative stress, and polarization in murine microglia. We found that SFN protects N9 microglial cells upon LPS-induced cell death and suppresses LPS-induced levels of secreted pro-inflammatory cytokines, tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6. SFN is also a potent inducer of redox sensitive transcription factor, nuclear factor erythroid 2-related factor 2 (Nrf2), which is responsible for the transcription of antioxidant, cytoprotective, and anti-inflammatory genes. SFN induced translocation of Nrf2 to the nucleus via extracellular signal-regulated kinase 1/2 (ERK1/2) pathway activation. siRNA-mediated knockdown study showed that the effects of SFN on LPS-induced reactive oxygen species, reactive nitrogen species, and pro-inflammatory cytokine production and cell death are partly Nrf2 dependent. Mox phenotype is a novel microglial phenotype that has roles in oxidative stress responses. Our results suggested that SFN induced the Mox phenotype in murine microglia through Nrf2 pathway. SFN also alleviated LPS-induced expression of inflammatory microRNA, miR-155. Finally, SFN inhibits microglia-mediated neurotoxicity as demonstrated by conditioned medium and co-culture experiments. In conclusion, SFN exerts protective effects on microglia and modulates the microglial activation state.

Sulforaphane Inhibits Lipopolysaccharide-Induced Inflammation, Cytotoxicity, Oxidative Stress, and miR-155 Expression and Switches to Mox Phenotype through Activating Extracellular Signal-Regulated Kinase 1/2–Nuclear Factor Erythroid 2-Related Factor 2/Antioxidant Response Element Pathway in Murine Microglial Cells

PubMed Central

Eren, Erden; Tufekci, Kemal Ugur; Isci, Kamer Burak; Tastan, Bora; Genc, Kursad; Genc, Sermin

2018-01-01

Sulforaphane (SFN) is a natural product with cytoprotective, anti-inflammatory, and antioxidant effects. In this study, we evaluated the mechanisms of its effects on lipopolysaccharide (LPS)-induced cell death, inflammation, oxidative stress, and polarization in murine microglia. We found that SFN protects N9 microglial cells upon LPS-induced cell death and suppresses LPS-induced levels of secreted pro-inflammatory cytokines, tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6. SFN is also a potent inducer of redox sensitive transcription factor, nuclear factor erythroid 2-related factor 2 (Nrf2), which is responsible for the transcription of antioxidant, cytoprotective, and anti-inflammatory genes. SFN induced translocation of Nrf2 to the nucleus via extracellular signal-regulated kinase 1/2 (ERK1/2) pathway activation. siRNA-mediated knockdown study showed that the effects of SFN on LPS-induced reactive oxygen species, reactive nitrogen species, and pro-inflammatory cytokine production and cell death are partly Nrf2 dependent. Mox phenotype is a novel microglial phenotype that has roles in oxidative stress responses. Our results suggested that SFN induced the Mox phenotype in murine microglia through Nrf2 pathway. SFN also alleviated LPS-induced expression of inflammatory microRNA, miR-155. Finally, SFN inhibits microglia-mediated neurotoxicity as demonstrated by conditioned medium and co-culture experiments. In conclusion, SFN exerts protective effects on microglia and modulates the microglial activation state. PMID:29410668

An essential role of Nrf2 in American ginseng-mediated anti-oxidative actions in cardiomyocytes.

PubMed

Li, Jinqing; Ichikawa, Tomonaga; Jin, Yu; Hofseth, Lorne J; Nagarkatti, Prakash; Nagarkatti, Mitzi; Windust, Anthony; Cui, Taixing

2010-07-20

Ginseng has been used as a folk medicine for thousands of years in Asia, and has become a popular herbal medicine world-wide. Recent studies have revealed that ginseng, including American ginseng, exerts antioxidant effects in the cardiovascular system; however, the underlying mechanisms are not fully understood. Thus, we investigated role of Nrf2, a master transcription factor of endogenous anti-oxidative defense systems, in the regulation of American ginseng-mediated anti-oxidative actions in cardiomyocytes. A standardized crude extract of American ginseng was supplied by the National Research Council of Canada, Institute for National Measurement Standards. H9C2 cells, a rat cardiomyocyte cell line, were exposed to angiotensin II (Ang II) or tumor necrosis factor alpha (TNFalpha) to induce oxidative stress that was examined by measuring formation of reactive oxygen and nitrogen species. Oxidative stress-induced cell death was induced by exogenous addition of hydrogen peroxide (H(2)O(2)). Proteins were measured by Western blot and mRNA expression was determined by quantitative real time PCR. Nrf2-driven transcriptional activity was assessed by antioxidant response element (ARE)-luciferase reporter assay. Direct Nrf2 binding to its target gene promoters was determined by chromatin immunoprecipitation assay. Adenoviral over-expression of Nrf2 shRNA was utilized to knock down Nrf2 in H9C2 cells. Immunochemical staining was applied for Nrf2 expression in the heart. American ginseng induced dramatic increases in Nrf2 protein expression, Nrf2 nuclear translocation, Nrf2 transcriptional activity, direct Nrf2 binding to its target gene promoters, and expression of a group of anti-oxidative genes driven by Nrf2 in H9C2 cells. In addition, American ginseng inhibited Ang II- or TNFalpha-induced free radical formation and H(2)O(2)-induced cell death in H9C2 cells over-expressed with control shRNA but not in the cells over-expressed with Nrf2 shRNA. Finally, oral administration of American ginseng markedly increased Nrf2 activity in murine hearts. These results demonstrate that American ginseng suppresses oxidative stress and oxidative stress-induced cell death in cardiomyocytes through activating the Nrf2 pathway, thereby providing cardioprotection against pathological cardiac remodeling. Copyright 2010 Elsevier Ireland Ltd. All rights reserved.

Activation of Nrf2-Antioxidant Signaling by 1,25-Dihydroxycholecalciferol Prevents Leptin-Induced Oxidative Stress and Inflammation in Human Endothelial Cells.

PubMed

Teixeira, Thaisa M; da Costa, Danielly C; Resende, Angela C; Soulage, Christophe O; Bezerra, Flavia F; Daleprane, Julio B

2017-04-01

Background: Obesity is associated with hyperleptinemia and endothelial dysfunction. Hyperleptinemia has been reported to induce both oxidative stress and inflammation by increasing reactive oxygen species production. Objective: The objective of this study was to determine the effects of 1,25-dihydroxycholecalciferol [1,25(OH) 2 D 3 ] against leptin-induced oxidative stress and inflammation in human endothelial cells. Methods: Small interfering RNA (siRNA) were used to knock down the expression of vitamin D receptor (VDR) in human umbilical vein endothelial cells (HUVECs). HUVECs were pretreated for 4 h with physiologic (10 -10 M) or supraphysiologic (10 -7 M) concentrations of 1,25(OH) 2 D 3 and exposed to leptin (10 ng/mL). Superoxide anion production and translocation of nuclear factor (erythroid-derived 2)-like 2 (NRF2) and nuclear transcription factor κB (NF-κB) subunit p65 to the nucleus and the activation of their target genes were quantified. Results: Pretreatment of HUVECs with 1,25(OH) 2 D 3 prevented the leptin-induced increase in superoxide anion production ( P < 0.05). Pretreatment with 1,25(OH) 2 D 3 further increased NRF2 translocation to the nucleus (by 3-fold; P < 0.05) and increased mRNA expression of superoxide dismutase 2 ( SOD2 ; by 2-fold), glutathione peroxidase ( GPX ; by 3-fold), NAD(P)H dehydrogenase (quinone) 1 ( NQO1 ; by 4-fold), and heme oxygenase 1 ( HMOX1 ; by 2-fold) ( P < 0.05). Leptin doubled the translocation of NF-κB ( P < 0.05) to the nucleus and increased ( P < 0.05) the upregulation of vascular inflammatory mediators such as monocyte chemoattractant protein 1 ( MCP1 ; by 1-fold), transforming growth factor β ( TGF β by 1-fold), and vascular cell adhesion molecule 1 ( VCAM1 ; by 4-fold) ( P < 0.05), which were prevented ( P < 0.05) by pretreatment with 1,25(OH) 2 D 3 Protective effects of 1,25(OH) 2 D 3 were confirmed to be VDR dependent by using VDR siRNA. Conclusion: Pretreatment with 1,25(OH) 2 D 3 in the presence of a high concentration of leptin has a beneficial effect on HUVECs through the regulation of mediators of antioxidant activity and inflammation. © 2017 American Society for Nutrition.

Dioscin alleviates BDL- and DMN-induced hepatic fibrosis via Sirt1/Nrf2-mediated inhibition of p38 MAPK pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gu, Lina; Tao, Xufeng; Xu, Youwei

Oxidative stress is involved in hepatic stellate cells (HSCs) activation and extracellular matrix overproduction. We previously reported the promising effects of dioscin against CCl{sub 4}-induced liver fibrosis, but its effects and mechanisms on BDL- and DMN-induced liver fibrosis remain unknown. The results in the present study indicated that dioscin significantly inhibited HSCs activation and attenuated hepatic fibrosis in rats. Furthermore, dioscin markedly up-regulated the levels of sirtuin 1 (Sirt1), HO-1, GST, GCLC and GCLM via increasing the nuclear translocation of nuclear erythroid factor 2-related factor 2 (Nrf2), which in turn inhibited mitogen-activated protein kinase 14 (p38 MAPK) phosphorylation and reducedmore » the levels of COL1A1, COL3A1, α-SMA and fibronectin. These results were further validated by knockdown of Sirt1 and Nrf2 using siRNAs silencing, and abrogation of p38 MAPK using SB-203580 (a p38 MAPK inhibitor) in HSC-T6 and LX-2 cells. Collectively, our findings confirmed the potent effects of dioscin against liver fibrosis and also provided novel insights into the mechanisms of this compound as a candidate for the prevention of liver fibrosis in the future. - Highlights: • Dioscin showed potent effects against BDL- and DMN-induced liver fibrosis in rats. • Dioscin significantly suppressed oxidative stress. • Dioscin triggered Sirt1/Nrf2-mediated inhibition of p38 MAPK pathway. • Dioscin should be developed as a novel candidate to treat liver fibrosis.« less

Eriodictyol protects against H(2)O(2)-induced neuron-like PC12 cell death through activation of Nrf2/ARE signaling pathway.

PubMed

Lou, Haiyan; Jing, Xu; Ren, Dongmei; Wei, Xinbing; Zhang, Xiumei

2012-07-01

Eriodictyol, a flavonoid isolated from the Chinese herb Dracocephalum rupestre has long been established as an antioxidant. The present study was designed to explore the protective effects of eriodictyol against hydrogen peroxide (H(2)O(2))-induced neurotoxicity with cultured rat pheochromocytoma cells (PC12 cells) and the possible mechanisms involved. For this purpose, differentiated PC12 cells were cultured and exposed to 200 μM H(2)O(2) in the absence or presence of eriodictyol (20, 40 and 80 μM). In addition, the potential contribution of the Nrf2/ARE neuroprotective pathway in eriodictyol-mediated protection against H(2)O(2)-induced neurotoxicity was also investigated. The results showed that H(2)O(2)-induced cell death can be inhibited in the presence of eriodictyol as measured by assays for MTT and apoptosis. Further study revealed that eriodictyol induced the nuclear translocation of Nrf2, enhanced the expression of heme oxygenase (HO-1) and γ-glutamylcysteine synthetase (γ-GCS), and increased the levels of intracellular glutathione. Treatment of PC12 cells with Nrf2 small interference RNA abolished eriodictyol-induced HO-1 and γ-GCS expression and its protective effects. In conclusion, these results suggest that eriodictyol upregulates HO-1 and γ-GCS expression through the activation of Nrf2/ARE pathway and protects PC12 cells against H(2)O(2)-induced oxidative stress. Copyright © 2012 Elsevier Ltd. All rights reserved.

A novel role of the NRF2 transcription factor in the regulation of arsenite-mediated keratin 16 gene expression in human keratinocytes.

PubMed

Endo, Hitoshi; Sugioka, Yoshihiko; Nakagi, Yoshihiko; Saijo, Yasuaki; Yoshida, Takahiko

2008-07-01

Inorganic sodium arsenite (iAs) is a ubiquitous environmental contaminant and is associated with an increased risk of skin hyperkeratosis and cancer. We investigated the molecular mechanisms underlying the regulation of the keratin 16 (K16) gene by iAs in the human keratinocyte cell line HaCaT. We performed reverse transcriptase polymerase chain reaction, luciferase assays, Western blots, and electrophoretic mobility shift assays to determine the transcriptional regulation of the K16 gene by iAs. We used gene overexpression approaches to elucidate the nuclear factor erythroid-derived 2 related factor 2 (NRF2) involved in the K16 induction. iAs induced the mRNA and protein expression of K16. We also found that the expression of K16 was transcriptionally induced by iAs through activator protein-1-like sites and an antioxidant response element (ARE) in its gene promoter region. Treatment with iAs also enhanced the production and translocation of the NRF2 transcription factor, an ARE-binding protein, into the nucleus without modification of its mRNA expression. In addition, iAs elongated the half-life of the NRF2 protein. When overexpressed in HaCaT cells, NRF2 was also directly involved in not only the up-regulation of the detoxification gene thioredoxin but also K16 gene expression. Our data clearly indicate that the K16 gene is a novel target of NRF2. Furthermore, our findings also suggest that NRF2 has opposing roles in the cell--in the activation of detoxification pathways and in promoting the development of skin disorders.

Indoxyl Sulfate Affects Glial Function Increasing Oxidative Stress and Neuroinflammation in Chronic Kidney Disease: Interaction between Astrocytes and Microglia.

PubMed

Adesso, Simona; Magnus, Tim; Cuzzocrea, Salvatore; Campolo, Michela; Rissiek, Björn; Paciello, Orlando; Autore, Giuseppina; Pinto, Aldo; Marzocco, Stefania

2017-01-01

Indoxyl sulfate (IS) is a protein-bound uremic toxin resulting from the metabolism of dietary tryptophan which accumulates in patients with impaired renal function, such as chronic kidney disease (CKD). IS is a well-known nephrovascular toxin but little is known about its effects on central nervous system (CNS) cells. Considering the growing interest in the field of CNS comorbidities in CKD, we studied the effect of IS on CNS cells. IS (15-60 μM) treatment in C6 astrocyte cells increased reactive oxygen species release and decreased nuclear factor (erythroid-derived 2)-like 2 (Nrf2) activation, and heme oxygenase-1 (HO-1) and NAD(P)H dehydrogenase quinone 1 expression. Moreover, IS increased Aryl hydrocarbon Receptor (AhR) and Nuclear Factor-kB (NF-kB) activation in these cells. Similiar observations were made in primary mouse astrocytes and mixed glial cells. Inducible nitric oxide synthase and cyclooxygenase-2 (COX-2) expression, tumor necrosis factor-α and interleukin-6 release and nitrotyrosine formation were increased by IS (15-60 μM) in primary mouse astrocytes and mixed glial cells. IS increased AhR and NF-kB nuclear translocation and reduced Nrf2 translocation and HO-1 expression in primary glial cells. In addition, IS induced cell death in neurons in a dose dependent fashion. Injection of IS (800 mg/kg, i.p.) into mice induced histological changes and increased COX-2 expression and nitrotyrosine formation in thebrain tissue. Taken together, our results show a significant contribution of IS in generating a neurotoxic enviroment and it could also have a potential role in neurodegeneration. IS could be considered also a potential therapeutical target for CKD-associated neurodegenerative complications.

Mangiferin Upregulates Glyoxalase 1 Through Activation of Nrf2/ARE Signaling in Central Neurons Cultured with High Glucose.

PubMed

Liu, Yao-Wu; Cheng, Ya-Qin; Liu, Xiao-Li; Hao, Yun-Chao; Li, Yu; Zhu, Xia; Zhang, Fan; Yin, Xiao-Xing

2017-08-01

Mangiferin, a natural C-glucoside xanthone, has anti-inflammatory, anti-oxidative, neuroprotective actions. Our previous study showed that mangiferin could attenuate diabetes-associated cognitive impairment of rats by enhancing the function of glyoxalase 1 (Glo-1) in brain. The aim of this study was to investigate whether Glo-1 upregulation by mangiferin in central neurons exposed to chronic high glucose may be related to activation of Nrf2/ARE pathway. Compared with normal glucose (25 mmol/L) culture, Glo-1 protein, mRNA, and activity levels were markedly decreased in primary hippocampal and cerebral cortical neurons cultured with high glucose (50 mmol/L) for 72 h, accompanied by the declined Nrf2 nuclear translocation and protein expression of Nrf2 in cell nucleus, as well as protein expression and mRNA level of γ-glutamylcysteine synthetase (γ-GCS) and superoxide dismutase activity, target genes of Nrf2/ARE signaling. Nonetheless, high glucose cotreating with mangiferin or sulforaphane, a typical inducer of Nrf2 activation, attenuated the above changes in both central neurons. In addition, mangiferin and sulforaphane significantly prevented the formation of advanced glycation end-products (AGEs) reflecting Glo-1 activity, while elevated the level of glutathione, a cofactor of Glo-1 activity and production of γ-GCS, in high glucose cultured central neurons. These findings demonstrated that Glo-1 was greatly downregulated in central neurons exposed to chronic high glucose, which is expected to lead the formation of AGEs and oxidative stress damages. We also proved that mangiferin enhanced the function of Glo-1 under high glucose condition by inducing activation of Nrf2/ARE signaling pathway.

PML-Nuclear Bodies Regulate the Stability of the Fusion Protein Dendra2-Nrf2 in the Nucleus.

PubMed

Burroughs, Andrea Flores; Eluhu, Sylvia; Whalen, Diva; Goodwin, J Shawn; Sakwe, Amos M; Arinze, Ifeanyi J

2018-05-22

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a basic leucine-zipper transcription factor essential for cellular responses to oxidative stress. Degradation of Nrf2 in the cytoplasm, mediated by Keap1-Cullin3/RING box1 (Cul3-Rbx1) E3 ubiquitin ligase and the proteasome, is considered the primary pathway controlling the cellular abundance of Nrf2. Although the nucleus has been implicated in the degradation of Nrf2, little information is available on how this compartment participates in degrading Nrf2. Here, we fused the photoconvertible fluorescent protein Dendra2 to Nrf2 and capitalized on the irreversible change in color (green to red) that occurs when Dendra2 undergoes photoconversion to study degradation of Dendra2-Nrf2 in single live cells. Using this approach, we show that the half-life (t1/2) of Dendra2-Nrf2 in the whole cell, under homeostatic conditions, is 35 min. Inhibition of the proteasome with MG-132 or induction of oxidative stress with tert-butylhydroquinone (tBHQ) extended the half-life of Dendra2-Nrf2 by 6- and 28-fold, respectively. By inhibiting nuclear export using Leptomycin B, we provide direct evidence that degradation of Nrf2 also occurs in the nucleus and involves PML-NBs (Promyelocytic Leukemia-nuclear bodies). We further demonstrate that co-expression of Dendra2-Nrf2 and Crimson-PML-I lacking two PML-I sumoylation sites (K65R and K490R) changed the decay rate of Dendra2-Nrf2 in the nucleus and stabilized the nuclear derived Nrf2 levels in whole cells. Altogether, our findings provide direct evidence for degradation of Nrf2 in the nucleus and suggest that modification of Nrf2 in PML nuclear bodies contributes to its degradation in intact cells. © 2018 The Author(s). Published by S. Karger AG, Basel.

Endothelial HO-1 induction by model TG-rich lipoproteins is regulated through a NOX4-Nrf2 pathway1[S

PubMed Central

Latham Birt, Sally H.; Purcell, Robert; Botham, Kathleen M.; Wheeler-Jones, Caroline P. D.

2016-01-01

Circulating levels of chylomicron remnants (CMRs) increase postprandially and their composition directly reflects dietary lipid intake. These TG-rich lipoproteins likely contribute to the development of endothelial dysfunction, albeit via unknown mechanisms. Here, we investigated how the FA composition of CMRs influences their actions on human aortic endothelial cells (HAECs) by comparing the effects of model CMRs—artificial TG-rich CMR-like particles (A-CRLPs)—containing TGs extracted from fish, DHA-rich algal, corn, or palm oils. HAECs responded with distinct transcriptional programs according to A-CRLP TG content and oxidation status, with genes involved in antioxidant defense and cytoprotection most prominently affected by n-3 PUFA-containing A-CRLPs. These particles were significantly more efficacious inducers of heme oxygenase-1 (HO-1) than n-6 PUFA corn or saturated FA-rich palm CRLPs. Mechanistically, HO-1 induction by all CRLPs requires NADPH oxidase 4, with PUFA-containing particles additionally dependent upon mitochondrial reactive oxygen species. Activation of both p38 MAPK and PPARβ/δ culminates in increased nuclear factor erythroid 2-related factor 2 (Nrf2) expression/nuclear translocation and HO-1 induction. These studies define new molecular pathways coupling endothelial cell activation by model CMRs with adaptive regulation of Nrf2-dependent HO-1 expression and may represent key mechanisms through which dietary FAs differentially impact progression of endothelial dysfunction. PMID:27185859

Isodeoxyelephantopin induces protective autophagy in lung cancer cells via Nrf2-p62-keap1 feedback loop

PubMed Central

Wang, Yang; Zhang, Jing; Huang, Zhi-Hao; Huang, Xiao-Hui; Zheng, Wei-Bin; Yin, Xing-Feng; Li, Yao-Lan; Li, Bin; He, Qing-Yu

2017-01-01

Isodeoxyelephantopin (ESI), isolated from Elephantopus scaber L. has been reported to exert anticancer effects. In this study, we aimed to investigate whether and how cancer cells exert protective responses against ESI treatment. Confocal fluorescence microscopy showed that ESI significantly induced autophagy flux in the lung cancer cells expressing mCherry-EGFP-LC3 reporter. Treatment of the cells with ESI increased the expression levels of the autophagy markers including LC3-II, ATG3 and Beclin1 in a dose-dependent manner. Pretreatment with autophagy inhibitor 3-methyladenine (3-MA) not only attenuated the effects of ESI on autophagy, but also enhanced the effects of ESI on cell viability and apoptosis. Mechanistically, the SILAC quantitative proteomics coupled with bioinformatics analysis revealed that the ESI-regulated proteins were mainly involved in Nrf2-mediated oxidative stress response. We found that ESI induced the nuclear translocation of Nrf2 for activating the downstream target genes including HO-1 and p62 (SQSTM1). More importantly, ESI-induced p62 could competitively bind with Keap1, and releases Nrf2 to activate downstream target gene p62 as a positive feedback loop, therefore promoting autophagy. Furthermore, knockdown of Nrf2 or p62 could abrogate the ESI-induced autophagy and significantly enhanced the anticancer effect of ESI. Taken together, we demonstrated that ESI can sustain cell survival by activating protective autophagy through Nrf2-p62-keap1 feedback loop, whereas targeting this regulatory axis combined with ESI treatment may be a promising strategy for anticancer therapy. PMID:28617433

Isodeoxyelephantopin induces protective autophagy in lung cancer cells via Nrf2-p62-keap1 feedback loop.

PubMed

Wang, Yang; Zhang, Jing; Huang, Zhi-Hao; Huang, Xiao-Hui; Zheng, Wei-Bin; Yin, Xing-Feng; Li, Yao-Lan; Li, Bin; He, Qing-Yu

2017-06-15

Isodeoxyelephantopin (ESI), isolated from Elephantopus scaber L. has been reported to exert anticancer effects. In this study, we aimed to investigate whether and how cancer cells exert protective responses against ESI treatment. Confocal fluorescence microscopy showed that ESI significantly induced autophagy flux in the lung cancer cells expressing mCherry-EGFP-LC3 reporter. Treatment of the cells with ESI increased the expression levels of the autophagy markers including LC3-II, ATG3 and Beclin1 in a dose-dependent manner. Pretreatment with autophagy inhibitor 3-methyladenine (3-MA) not only attenuated the effects of ESI on autophagy, but also enhanced the effects of ESI on cell viability and apoptosis. Mechanistically, the SILAC quantitative proteomics coupled with bioinformatics analysis revealed that the ESI-regulated proteins were mainly involved in Nrf2-mediated oxidative stress response. We found that ESI induced the nuclear translocation of Nrf2 for activating the downstream target genes including HO-1 and p62 (SQSTM1). More importantly, ESI-induced p62 could competitively bind with Keap1, and releases Nrf2 to activate downstream target gene p62 as a positive feedback loop, therefore promoting autophagy. Furthermore, knockdown of Nrf2 or p62 could abrogate the ESI-induced autophagy and significantly enhanced the anticancer effect of ESI. Taken together, we demonstrated that ESI can sustain cell survival by activating protective autophagy through Nrf2-p62-keap1 feedback loop, whereas targeting this regulatory axis combined with ESI treatment may be a promising strategy for anticancer therapy.

Alkaloids from Piper nigrum Exhibit Antiinflammatory Activity via Activating the Nrf2/HO-1 Pathway.

PubMed

Ngo, Quynh Mai Thi; Tran, Phuong Thao; Tran, Manh Hung; Kim, Jeong Ah; Rho, Seong Soo; Lim, Chi-Hwan; Kim, Jin-Cheol; Woo, Mi Hee; Choi, Jae Sui; Lee, Jeong-Hyung; Min, Byung Sun

2017-04-01

In the present study, ten alkaloids, namely chabamide (1), pellitorine (2), retrofractamide A (3), pyrroperine (4), isopiperolein B (5), piperamide C9:1 (8E) (6), 6,7-dehydrobrachyamide B (7), 4,5-dihydropiperine (8), dehydropipernonaline (9), and piperine (10), were isolated from the fruits of Piper nigrum. Among these, chabamide (1), pellitorine (2), retrofractamide A (3), isopiperolein B (5), and 6,7-dehydrobrachyamide B (7) exhibited significant inhibitory activity on lipopolysaccharide-induced nitric oxide (NO) production in RAW264.7 cells, with IC 50 values of 6.8, 14.5, 30.2, 23.7, and 38.5 μM, respectively. Furthermore, compound 1 inhibited lipopolysaccharide-induced NO production in bone marrow-derived macrophages with IC 50 value of 9.5 μM. Consistent with NO inhibition, treatment of RAW264.7 cells with chabamide (1), pellitorine (2), and 6,7-dehydrobrachyamide B (7) suppressed expression of inducible NO synthase and cyclooxygenase-2. Chabamide (1), pellitorine (2), and 6,7-dehydrobrachyamide B (7) induced heme-oxygenase-1 expression at the transcriptional level. In addition, compound 1 induced the nuclear translocation of nuclear factor-E2-related factor 2 (Nrf2) and upregulated the expression of Nrf2 target genes, NAD(P)H:quinone oxidoreductase 1 and γ-glutamyl cysteine synthetase catalytic subunit, in a concentration-dependent manner in RAW264.7 cells. These findings suggest that chabamide (1) from P. nigrum exert antiinflammatory effects via the activation of the Nrf2/heme-oxygenase-1 pathway; hence, it might be a promising candidate for the treatment of inflammatory diseases. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

DMF, but not other fumarates, inhibits NF-κB activity in vitro in an Nrf2-independent manner.

PubMed

Gillard, Geoffrey O; Collette, Brian; Anderson, John; Chao, Jianhua; Scannevin, Robert H; Huss, David J; Fontenot, Jason D

2015-06-15

Fumarate-containing pharmaceuticals are potent therapeutic agents that influence multiple cellular pathways. Despite proven clinical efficacy, there is a significant lack of data that directly defines the molecular mechanisms of action of related, yet distinct fumarate compounds. We systematically compared the impact of dimethyl fumarate (DMF), monomethyl fumarate (MMF) and a mixture of monoethyl fumarate salts (Ca(++), Mg(++), Zn(++); MEF) on defined cellular responses. We demonstrate that DMF inhibited NF-κB-driven cytokine production and nuclear translocation of p65 and p52 in an Nrf2-independent manner. Equivalent doses of MMF and MEF did not affect NF-κB signaling. These results highlight a key difference in the biological impact of related, yet distinct fumarate compounds. Copyright © 2015. Published by Elsevier B.V.

Eicosapentaenoic acid prevents TCDD-induced oxidative stress and inflammatory response by modulating MAP kinases and redox-sensitive transcription factors

PubMed Central

Palanisamy, Kalaiselvi; Krishnaswamy, Rajashree; Paramasivan, Poornima; Chih-Yang, Huang; Vishwanadha, Vijaya Padma

2015-01-01

Background and Purpose Oxidative stress and subsequent activation of inflammatory responses is a widely accepted consequence of exposure to environmental toxins. TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin), a well-known environmental toxin, exerts its toxicity through many signalling mechanisms, with liver being the principal organ affected. However, an effective antidote to TCDD-induced toxicity is unknown. The present study evaluated the effect of eicosapentaenoic acid (EPA), an n3 fatty acid, on TCDD-induced toxicity. Experimental Approach In cultures of HepG2 cells, the EPA/AA ratio was determined using gas chromatography, oxidative stress and inflammatory responses through reactive oxygen species (ROS) levels, antioxidant status, [Ca2+]i, nuclear migration of two redox-sensitive transcription factors, NF-κB p65 and Nrf-2, expression of MAP kinase (p-Erk, p-p38), NF-κB p65, COX-2 and Nrf-2. Cellular changes in ΔΨm, acidic vesicular organelle formation, cell cycle analysis and scanning electron microscopy analysis were performed. Key Results EPA offered significant cytoprotection by increasing EPA/AA ratios in cell membranes, inhibiting ROS generation, enhancing antioxidant status and modulating nuclear translocation of redox-sensitive transcription factors (NF-κB p65 and Nrf-2) and expression of NF-κB p65, COX-2 and Nrf-2. Furthermore, TCDD-induced upstream events of MAPK phosphorylation, the increase in [Ca2+]i levels and cell surface changes in microvilli were significantly inhibited by EPA. EPA treatment maintained ΔΨm and prevented formation of acidic vesicular organelles. Conclusion and Implications The present study demonstrates for the first time some underlying molecular mechanisms of cytoprotection exerted by EPA against TCDD-induced oxidative stress and inflammatory responses. PMID:26177858

Sofalcone, a gastric mucosa protective agent, increases vascular endothelial growth factor via the Nrf2-heme-oxygenase-1 dependent pathway in gastric epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shibuya, Akiko; Onda, Kenji, E-mail: knjond@toyaku.ac.jp; Kawahara, Hirofumi

2010-07-30

Research highlights: {yields} Sofalcone increases HO-1 in gastric epithelial cells. {yields} The induction of HO-1 by sofalcone treatment follows the activation of Nrf2. {yields} The production of VEGF by sofalcone treatment is mediated by HO-1 induction. -- Abstract: Sofalcone, 2'-carboxymethoxy-4,4-bis(3-methyl-2-butenyloxy)chalcone, is an anti-ulcer agent that is classified as a gastric mucosa protective agent. Recent studies indicate heat shock proteins such as HSP32, also known as heme-oxygenase-1(HO-1), play important roles in protecting gastrointestinal tissues from several stresses. We have previously reported that sofalcone increases the expression of HO-1 in adipocytes and pre-adipocytes, although the effect of sofalcone on HO-1 induction inmore » gastrointestinal tissues is not clear. In the current study, we investigated the effects of sofalcone on the expression of HO-1 and its functional role in rat gastric epithelial (RGM-1) cells. We found that sofalcone increased HO-1 expression in RGM-1 cells in both time- and concentration-dependent manners. The HO-1 induction was associated with the nuclear translocation of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) in RGM-1 cells. We also observed that sofalcone increased vascular endothelial growth factor (VEGF) production in the culture medium. Treatment of RGM-1 cells with an HO-1 inhibitor (tin-protoporphyrin), or HO-1 siRNA inhibited sofalcone-induced VEGF production, suggesting that the effect of sofalcone on VEGF expression is mediated by the HO-1 pathway. These results suggest that the gastroprotective effects of sofalcone are partly exerted via Nrf2-HO-1 activation followed by VEGF production.« less

Modulation of NRF2 signaling pathway by nuclear receptors: implications for cancer.

PubMed

Namani, Akhileshwar; Li, Yulong; Wang, Xiu Jun; Tang, Xiuwen

2014-09-01

Nuclear factor-erythroid 2 p45-related factor 2 (NRF2, also known as Nfe2l2) plays a critical role in regulating cellular defense against electrophilic and oxidative stress by activating the expression of an array of antioxidant response element-dependent genes. On one hand, NRF2 activators have been used in clinical trials for cancer prevention and the treatment of diseases associated with oxidative stress; on the other hand, constitutive activation of NRF2 in many types of tumors contributes to the survival and growth of cancer cells, as well as resistance to anticancer therapy. In this review, we provide an overview of the NRF2 signaling pathway and discuss its role in carcinogenesis. We also introduce the inhibition of NRF2 by nuclear receptors. Further, we address the biological significance of regulation of the NRF2 signaling pathway by nuclear receptors in health and disease. Finally, we discuss the possible impact of NRF2 inhibition by nuclear receptors on cancer therapy. Copyright © 2014 Elsevier B.V. All rights reserved.

4-hydroxy-2-nonenal protects against cardiac ischemia-reperfusion injury via the Nrf2-dependent pathway.

PubMed

Zhang, Yan; Sano, Motoaki; Shinmura, Ken; Tamaki, Kayoko; Katsumata, Yoshinori; Matsuhashi, Tomohiro; Morizane, Shintaro; Ito, Hideyuki; Hishiki, Takako; Endo, Jin; Zhou, Heping; Yuasa, Shinsuke; Kaneda, Ruri; Suematsu, Makoto; Fukuda, Keiichi

2010-10-01

Reactive oxygen species (ROS) attack polyunsaturated fatty acids of the membrane and trigger lipid peroxidation, which results in the generation of alpha,beta-unsaturated aldehydes, such as 4-hydroxy-2-nonenal (4-HNE). There is compelling evidence that high concentrations of aldehydes are responsible for much of the damage elicited by cardiac ischemia-reperfusion injury, while sublethal concentrations of aldehydes stimulate stress resistance pathways, to achieve cardioprotection. We investigated the mechanism of cardioprotection mediated by 4-HNE. For cultured cardiomyocytes, 4-HNE was cytotoxic at higher concentrations (>or=20 microM) but had no appreciable cytotoxicity at lower concentrations. Notably, a sublethal concentration (5muM) of 4-HNE primed cardiomyocytes to become resistant to cytotoxic concentrations of 4-HNE. 4-HNE induced nuclear translocation of transcription factor NF-E2-related factor 2 (Nrf2), and enhanced the expression of gamma-glutamylcysteine ligase (GCL) and the core subunit of the Xc(-) high-affinity cystine transporter (xCT), thereby increasing 1.45-fold the intracellular GSH levels. Cardiomyocytes treated with either Nrf2-specific siRNA or the GCL inhibitor l-buthionine sulfoximine (BSO) were less tolerant to 4-HNE. Moreover, the cardioprotective effect of 4-HNE pretreatment against subsequent glucose-free anoxia followed by reoxygenation was completely abolished in these cells. Intravenous administration of 4-HNE (4 mg/kg) activated Nrf2 in the heart and increased the intramyocardial GSH content, and consequently improved the functional recovery of the left ventricle following ischemia-reperfusion in Langendorff-perfused hearts. This cardioprotective effect of 4-HNE was not observed for Nrf2-knockout mice. In summary, 4-HNE activates Nrf2-mediated gene expression and stimulates GSH biosynthesis, thereby conferring on cardiomyocytes protection against ischemia-reperfusion injury. Copyright 2010 Elsevier Ltd. All rights reserved.

Antioxidant axis Nrf2-keap1-ARE in inhibition of alcoholic liver fibrosis by IL-22

PubMed Central

Ni, Ya-Hui; Huo, Li-Juan; Li, Ting-Ting

2017-01-01

AIM To explore the effect of interleukin (IL)-22 on in vitro model of alcoholic liver fibrosis hepatic stellate cells (HSCs), and whether this is related to regulation of Nrf2-keap1-ARE. METHODS HSC-T6 cells were incubated with 25, 50, 100, 200 and 400 μmol/L acetaldehyde. After 24 and 48 h, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to detect proliferation of HSCs to choose the best concentration and action time. We used the optimal concentration of acetaldehyde (200 μmol/L) to stimulate HSCs for 24 h, and treated the cells with a final concentration of 10, 20 or 50 ng/mL IL-22. The cell proliferation rate was detected by MTT assay. The cell cycle was analyzed by flow cytometry. The expression of nuclear factor-related factor (Nrf)2 and α-smooth muscle antigen was detected by western blotting and immunocytochemistry. The levels of malondialdehyde (MDA) and glutathione (GSH) were measured by spectrophotometry. RESULTS In the MTT assay, when HSCs were incubated with acetaldehyde, activity and proliferation were higher than in the control group, and were most obvious after 48 h treatment with 200 μmol/L acetaldehyde. The number of cells in G0/G1 phases was decreased and the number in S phase was increased in comparison with the control group. When treated with different concentrations of IL-22, HSC-T6 cell activity and proliferation rate were markedly decreased in a dose-dependent manner, and cell cycle progression was arrested from G1 to S phase. Western blotting and immunocytochemistry demonstrated that expression of Nrf2 total protein was not significantly affected. Expression of Nrf2 nuclear protein was low in the control group, increased slightly in the model group (or acetaldehyde-stimulated group), and increased more obviously in the IL-22 intervention groups. The levels of MDA and GSH in the model group were significantly enhanced in comparison with those in the control group. In cells treated with IL-22, the MDA level was attenuated but the GSH level was further increased. These changes were dose-dependent. CONCLUSION IL-22 inhibits acetaldehyde-induced HSC activation and proliferation, which may be related to nuclear translocation of Nrf2 and increased activity of the antioxidant axis Nrf2-keap1-ARE. PMID:28373766

Modulation of NF-κB and Nrf2 control of inflammatory responses in FHs 74 Int cell line is tocopherol isoform-specific

PubMed Central

Elisia, Ingrid

2013-01-01

The present study investigates the relative ability of α-, γ-, and δ-tocopherol (Toc) to modulate cell signaling events that are associated with inflammatory responses in fetal-derived intestinal (FHs 74 Int) cells. Secretion of the proinflammatory cytokine IL-8 in FHs 74 Int cells was stimulated in the following order: α-Toc < γ-Toc < δ-Toc. A similar proinflammatory response was observed when inflammation was induced in FHs 74 Int cells. Modulation of IL-8 expression by Toc corresponded to an isoform-specific modulation of NF-κB and nuclear factor-erythroid 2-related factor 2 (Nrf2) cell signaling pathways involved in expression of proinflammatory cytokines and antioxidant enzymes, respectively. δ-Toc and, to a lesser extent, γ-Toc activated NF-κB and Nrf2 signaling, as indicated by the greater nuclear translocation of transcription factors. Activation of NF-κB signaling by γ- and δ-Toc was accompanied by upregulation of NF-κB target genes, such as IL-8 and prostaglandin-endoperoxide synthase 2, with and without a prior IFNγ-PMA challenge. Nevertheless, γ- and δ-Toc, particularly δ-Toc, concurrently downregulated glutamate-cysteine ligase, a Nrf2 target gene that encodes for glutathione biosynthesis. This observation was substantiated by confirmation that γ- and δ-Toc were effective at decreasing glutamate-cysteine ligase protein expression and cellular glutathione content. Downregulation of glutathione content in fetal intestinal cells corresponded to induction of apoptosis-mediated cytotoxicity. In conclusion, γ- and δ-Toc are biologically active isoforms of vitamin E and show superior bioactivity to α-Toc in modulating cell signaling events that contribute to a proinflammatory response in fetal-derived intestinal cells. PMID:24136788

Evaluation of the rotenone-induced activation of the Nrf2 pathway in a neuronal model derived from human induced pluripotent stem cells.

PubMed

Zagoura, Dimitra; Canovas-Jorda, David; Pistollato, Francesca; Bremer-Hoffmann, Susanne; Bal-Price, Anna

2017-06-01

Human induced pluripotent stem cells (hiPSCs) are considered as a powerful tool for drug and chemical screening and development of new in vitro testing strategies in the field of toxicology, including neurotoxicity evaluation. These cells are able to expand and efficiently differentiate into different types of neuronal and glial cells as well as peripheral neurons. These human cells-based neuronal models serve as test systems for mechanistic studies on different pathways involved in neurotoxicity. One of the well-known mechanisms that are activated by chemically-induced oxidative stress is the Nrf2 signaling pathway. Therefore, in the current study, we evaluated whether Nrf2 signaling machinery is expressed in human induced pluripotent stem cells (hiPSCs)-derived mixed neuronal/glial culture and if so whether it becomes activated by rotenone-induced oxidative stress mediated by complex I inhibition of mitochondrial respiration. Rotenone was found to induce the activation of Nrf2 signaling particularly at the highest tested concentration (100 nM), as shown by Nrf2 nuclear translocation and the up-regulation of the Nrf2-downstream antioxidant enzymes, NQO1 and SRXN1. Interestingly, exposure to rotenone also increased the number of astroglial cells in which Nrf2 activation may play an important role in neuroprotection. Moreover, rotenone caused cell death of dopaminergic neurons since a decreased percentage of tyrosine hydroxylase (TH + ) cells was observed. The obtained results suggest that hiPSC-derived mixed neuronal/glial culture could be a valuable in vitro human model for the establishment of neuronal specific assays in order to link Nrf2 pathway activation (biomarker of oxidative stress) with additional neuronal specific readouts that could be applied to in vitro neurotoxicity evaluation. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Sulforaphane exerts its anti-inflammatory effect against amyloid-β peptide via STAT-1 dephosphorylation and activation of Nrf2/HO-1 cascade in human THP-1 macrophages.

PubMed

An, Ye Won; Jhang, Kyoung A; Woo, So-Youn; Kang, Jihee Lee; Chong, Young Hae

2016-02-01

Alzheimer's disease (AD) is the most common neurodegenerative disorder worldwide, accounting for most cases of dementia in elderly individuals, and effective therapies are still lacking. This study was designed to investigate the anti-inflammatory properties of sulforaphane against Aβ1-42 monomers in human THP-1 microglia-like cells. The results showed that sulforaphane preferentially inhibited cathepsin B- and caspase-1-dependent NLRP3 inflammasome activation induced by mostly Aβ1-42 monomers, an effect that potently reduced excessive secretion of the proinflammatory cytokine interleukin-1β (IL-1β). Subsequent mechanistic studies revealed that sulforaphane mitigated the activation of signal transducer and activator of transcription-1 induced by Aβ1-42 monomers. Sulforaphane also increased nuclear factor erythroid 2-related factor 2 (Nrf2) nuclear translocation, which was followed by upregulation of heme-oxygenase 1 (HO-1). The anti-inflammatory effect of sulforaphane on Aβ1-42-induced IL-1β production was diminished by small interfering RNA-mediated knockdown of Nrf2 or HO-1. Moreover, sulforaphane significantly attenuated the levels of microRNA-146a, which is selectively upregulated in the temporal cortex and hippocampus of AD brains. The aforementioned effects of sulforaphane were replicated by the tyrosine kinase inhibitor, herbimycin A, and Nrf2 activator. These results indicate that signal transducer and activator of transcription-1 dephosphorylation, HO-1 and its upstream effector, Nrf2, play a pivotal role in triggering an anti-inflammatory signaling cascade of sulforaphane that results in decreases of IL-1β release and microRNA-146a production in Aβ1-42-stimulated human microglia-like cells. These findings suggest that the phytochemical sulforaphane has a potential application in AD therapeutics. Copyright © 2016 Elsevier Inc. All rights reserved.

Salvianolic acid B protects against paraquat-induced pulmonary injury by mediating Nrf2/Nox4 redox balance and TGF-β1/Smad3 signaling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Bin, E-mail: iamicehe@163.com

The present study was aimed at exploring the protective effects of Salvianolic acid B (SalB) against paraquat (PQ)-induced lung injury in mice. Lung fibrotic injuries were induced in mice by a single intragastrical administration of 300 mg/kg PQ, then the mice were administrated with 200 mg/kg, 400 mg/kg SalB, 100 mg/kg vitamin C (Vit C) and dexamethasone (DXM) for 14 days. PQ-triggered structure distortion, collagen overproduction, excessive inflammatory infiltration, pro-inflammatory cytokine release, and oxidative stress damages in lung tissues and mortality of mice were attenuated by SalB in a dose-dependent manner. Furthermore, SalB was noted to enhance the expression andmore » nuclear translocation of nuclear factor erythroid 2–related factor 2 (Nrf2) and reduce expression of the reactive oxygen species-generating enzyme Nox4 [NADPH (reduced form of nicotinamide adenine dinucleotide phosphate) oxidase-4]. SalB also inhibited the increasing expression of transforming growth factor (TGF)-β1 and the phosphorylation of its downstream target Smad3 which were enhanced by PQ. These results suggest that SalB may exert protective effects against PQ-induced lung injury and pulmonary fibrosis. Its mechanisms involve the mediation of Nrf2/Nox4 redox balance and TGF-β1/Smad3 signaling. - Highlights: • Salvianolic acid B (SalB) reduced Paraquat-induced mortality and pulmonary injury in mice. • SalB has anti-oxidation, anti-inflammatory and anti-fibrogenic effects simultaneously. • Its mechanisms were targeting Nrf2-Nox4 redox balance and TGF-β1/Smad3 signaling.« less

Dimerumic acid attenuates receptor for advanced glycation endproducts signal to inhibit inflammation and diabetes mediated by Nrf2 activation and promotes methylglyoxal metabolism into d-lactic acid.

PubMed

Lee, Bao-Hong; Hsu, Wei-Hsuan; Hsu, Ya-Wen; Pan, Tzu-Ming

2013-07-01

This study was designed to evaluate the effects of dimerumic acid (DMA) on receptor for advanced glycation endproducts (RAGE) signal activation and THP-1 monocyte inflammation treated with S100b, a specific ligand of RAGE. We found that DMA inhibited inflammatory cytokine production via upregulation of nuclear factor-erythroid 2-related factor 2 (Nrf2) and alleviated oxidative stress through attenuation of p47phox translocation to the membrane of S100b-treated THP-1 monocytes. We found that DMA activated Nrf2 mediated by the p38 kinase pathway in THP-1 monocytes. However, anti-inflammatory activity of DMA was attenuated by Nrf2 siRNA treatment. In an animal model, methylglyoxal (MG; 200mg/kg bw) was chosen to induce diabetes in Balb/C mice (6 weeks) in this work. The in vivo verification of anti-inflammation in peripheral blood mononuclear cells by DMA treatment was confirmed by tumor necrosis factor-α and interleukin-1β measurements. Oral glucose tolerance test, insulin tolerance test, hyperinsulinemia, and hyperglycemia were improved in MG-treated mice by DMA treatment and these effects were greater than those of silymarin and N-acetylcysteine. Furthermore, DMA increased hepatic glyoxalase mRNA and glutathione mediated by Nrf2 activation to metabolize MG into d-lactic acid, thereby reducing serum and hepatic AGE levels and suppressing inflammatory factor generation in MG-treated mice. However, DMA did not exert the antiglycation activity in MG-bovine serum albumin incubation. Taken together, the results indicate that DMA is a novel antioxidant and Nrf2 activator that lowers AGE levels and may prove to be an effective treatment for diabetes. Copyright © 2013 Elsevier Inc. All rights reserved.

New hits as phase II enzymes inducers from a focused library with heteroatom-heteroatom and Michael-acceptor motives.

PubMed

Cabrera, Mauricio; de Ovalle, Stefani; Bollati-Fogolín, Mariela; Nascimento, Fabiana; Corbelini, Patrícia; Janarelli, Fernanda; Kawano, Daniel; Eifler-Lima, Vera Lucia; González, Mercedes; Cerecetto, Hugo

2015-11-01

The increased activity of phase-II-detoxification enzymes, such as quinone reductase (QR) and glutation S -transferase (GST), correlates with protection against chemically induced carcinogenesis. Herein we studied 11 different chemotypes, pyrazole, 1,2,4-oxadiazole, 1,2,5-oxadiazole, 1,2,3-thiadiazole, 1,2,4-thiazole, 1,3,4-oxathiazole, thienyl hydrazone, α,β-unsaturated-oxime, α,β-unsaturated- N -oxide, coumarin and α,β-unsaturated-carbonyl, as phase-II enzymes inducers in order to identify new pharmacophores with chemopreventive activity. Fifty-four compounds were analyzed on wild-type mouse-hepatoma Hepa-1c1c7 and on the aryl-hydrocarbon-nuclear-translocator (Arnt)-defective mutant BpRc1 cells. New monofunctional inducers of QR and GST were identified, the 1,2,5-oxadiazol-2-oxide (3) , the 1,2,4-triazine-4-oxides (23) and (32) and the tetrahydropyrimidinones (28) and (49) . It was confirmed that Nrf2 nuclear translocation is the operative molecular mechanism that allows compound (3) to exert protective effects via expression of downstream phase-II enzymes.

New hits as phase II enzymes inducers from a focused library with heteroatom–heteroatom and Michael-acceptor motives

PubMed Central

Cabrera, Mauricio; de Ovalle, Stefani; Bollati-Fogolín, Mariela; Nascimento, Fabiana; Corbelini, Patrícia; Janarelli, Fernanda; Kawano, Daniel; Eifler-Lima, Vera Lucia; González, Mercedes; Cerecetto, Hugo

2015-01-01

The increased activity of phase-II-detoxification enzymes, such as quinone reductase (QR) and glutation S-transferase (GST), correlates with protection against chemically induced carcinogenesis. Herein we studied 11 different chemotypes, pyrazole, 1,2,4-oxadiazole, 1,2,5-oxadiazole, 1,2,3-thiadiazole, 1,2,4-thiazole, 1,3,4-oxathiazole, thienyl hydrazone, α,β-unsaturated-oxime, α,β-unsaturated-N-oxide, coumarin and α,β-unsaturated-carbonyl, as phase-II enzymes inducers in order to identify new pharmacophores with chemopreventive activity. Fifty-four compounds were analyzed on wild-type mouse-hepatoma Hepa-1c1c7 and on the aryl-hydrocarbon-nuclear-translocator (Arnt)-defective mutant BpRc1 cells. New monofunctional inducers of QR and GST were identified, the 1,2,5-oxadiazol-2-oxide (3), the 1,2,4-triazine-4-oxides (23) and (32) and the tetrahydropyrimidinones (28) and (49). It was confirmed that Nrf2 nuclear translocation is the operative molecular mechanism that allows compound (3) to exert protective effects via expression of downstream phase-II enzymes. PMID:28031894

The synthetic triterpenoid RTA dh404 (CDDO-dhTFEA) restores endothelial function impaired by reduced Nrf2 activity in chronic kidney disease.

PubMed

Aminzadeh, Mohammad A; Reisman, Scott A; Vaziri, Nosratola D; Shelkovnikov, Stan; Farzaneh, Seyed H; Khazaeli, Mahyar; Meyer, Colin J

2013-01-01

Chronic kidney disease (CKD) is associated with endothelial dysfunction and accelerated cardiovascular disease, which are largely driven by systemic oxidative stress and inflammation. Oxidative stress and inflammation in CKD are associated with and, in part, due to impaired activity of the cytoprotective transcription factor Nrf2. RTA dh404 is a synthetic oleanane triterpenoid compound which potently activates Nrf2 and inhibits the pro-inflammatory transcription factor NF-κB. This study was designed to test the effects of RTA dh404 on endothelial function, inflammation, and the Nrf2-mediated antioxidative system in the aorta of rats with CKD induced by 5/6 nephrectomy. Sham-operated rats served as controls. Subgroups of CKD rats were treated orally with RTA dh404 (2 mg/kg/day) or vehicle for 12 weeks. The aortic rings from untreated CKD rats exhibited a significant reduction in the acetylcholine-induced relaxation response which was restored by RTA dh404 administration. Impaired endothelial function in the untreated CKD rats was accompanied by significant reduction of Nrf2 activity (nuclear translocation) and expression of its cytoprotective target genes, as well as accumulation of nitrotyrosine and upregulation of NAD(P)H oxidases, 12-lipoxygenase, MCP-1, and angiotensin II receptors in the aorta. These abnormalities were ameliorated by RTA dh404 administration, as demonstrated by the full or partial restoration of the expression of all the above analytes to sham control levels. Collectively, the data demonstrate that endothelial dysfunction in rats with CKD induced by 5/6 nephrectomy is associated with impaired Nrf2 activity in arterial tissue, which can be reversed with long term administration of RTA dh404.

Emodin Inhibition of Influenza A Virus Replication and Influenza Viral Pneumonia via the Nrf2, TLR4, p38/JNK and NF-kappaB Pathways.

PubMed

Dai, Jian-Ping; Wang, Qian-Wen; Su, Yun; Gu, Li-Ming; Zhao, Ying; Chen, Xiao-Xua; Chen, Cheng; Li, Wei-Zhong; Wang, Ge-Fei; Li, Kang-Sheng

2017-10-18

Lasting activations of toll-like receptors (TLRs), MAPK and NF-κB pathways can support influenza A virus (IAV) infection and promote pneumonia. In this study, we have investigated the effect and mechanism of action of emodin on IAV infection using qRT-PCR, western blotting, ELISA, Nrf2 luciferase reporter, siRNA and plaque inhibition assays. The results showed that emodin could significantly inhibit IAV (ST169, H1N1) replication, reduce IAV-induced expressions of TLR2/3/4/7, MyD88 and TRAF6, decrease IAV-induced phosphorylations of p38/JNK MAPK and nuclear translocation of NF-κB p65. Emodin also activated the Nrf2 pathway, decreased ROS levels, increased GSH levelss and GSH/GSSG ratio, and upregulated the activities of SOD, GR, CAT and GSH-Px after IAV infection. Suppression of Nrf2 via siRNA markedly blocked the inhibitory effects of emodin on IAV-induced activations of TLR4, p38/JNK, and NF-κB pathways and on IAV-induced production of IL-1β, IL-6 and expression of IAV M2 protein. Emodin also dramatically increased the survival rate of mice, reduced lung edema, pulmonary viral titer and inflammatory cytokines, and improved lung histopathological changes. In conclusion, emodin can inhibit IAV replication and influenza viral pneumonia, at least in part, by activating Nrf2 signaling and inhibiting IAV-induced activations of the TLR4, p38/JNK MAPK and NF-κB pathways.

Bisphenol A increases BeWo trophoblast survival in stress-induced paradigms through regulation of oxidative stress and apoptosis.

PubMed

Ponniah, Muralitharan; Billett, E Ellen; De Girolamo, Luigi A

2015-09-21

Bisphenol A (BPA) is ubiquitous in the environment and is reported to be present at high concentrations in placental tissue, where its presence raises concerns over its potential to disrupt placental function. This report investigates how BPA interferes with the survival of human choriocarcinoma BeWo cells (a model of placental trophoblasts) under stress-induced paradigms reminiscent of pathways activated in placental development. These include conditions that promote oxidative stress (glutathione depletion) and apoptosis (serum withdrawal) or mimic hypoxia (HIF-1α accumulation via dimethyloxalylglycine treatment). Treatment of BeWo cells with BPA during stress-induced paradigms led to a consistent and significant increase in cell viability, with a concomitant increase in glutathione levels and a reduction in apoptosis. Assessment of the antioxidant capacity of BPA revealed its ability to quench reactive oxygen species and reduce the levels generated during glutathione and serum depletion. BPA was also able to reduce the activation of the antioxidant response element (ARE) through mediation of its activators, nuclear factor erythroid related factor family members (Nrf's). Indeed, the expression and nuclear translocation of Nrf2 (an important ARE activator) were impaired by BPA, while Nrf1 and Nrf3 expression levels were increased. Furthermore, BPA increased the levels of the anti-apoptotic proteins (Bcl-2 and Hsp70) and decreased HIF-1α levels during stress-induced conditions. Together, these results indicate that BPA inhibits trophoblast cell death under conditions of cellular stress. This could have implications on placental trophoblasts during development.

Mitochondrial Effects of PGC-1alpha Silencing in MPP+ Treated Human SH-SY5Y Neuroblastoma Cells

PubMed Central

Ye, Qinyong; Chen, Chun; Si, Erwang; Cai, Yousheng; Wang, Juhua; Huang, Wanling; Li, Dongzhu; Wang, Yingqing; Chen, Xiaochun

2017-01-01

The dopaminergic neuron degeneration and loss that occurs in Parkinson’s disease (PD) has been tightly linked to mitochondrial dysfunction. Although the aged-related cause of the mitochondrial defect observed in PD patients remains unclear, nuclear genes are of potential importance to mitochondrial function. Human peroxisome proliferator-activated receptor γ coactivator-1alpha (PGC-1α) is a multi-functional transcription factor that tightly regulates mitochondrial biogenesis and oxidative capacity. The goal of the present study was to explore the potential pathogenic effects of interference by the PGC-1α gene on N-methyl-4-phenylpyridinium ion (MPP+)-induced SH-SY5Y cells. We utilized RNA interference (RNAi) technology to probe the pathogenic consequences of inhibiting PGC-1α in the SH-SY5Y cell line. Remarkably, a reduction in PGC-1α resulted in the reduction of mitochondrial membrane potential, intracellular ATP content and intracellular H2O2 generation, leading to the translocation of cytochrome c (cyt c) to the cytoplasm in the MPP+-induced PD cell model. The expression of related proteins in the signaling pathway (e.g., estrogen-related receptor α (ERRα), nuclear respiratory factor 1 (NRF-1), NRF-2 and Peroxisome proliferator-activated receptor γ (PPARγ)) also decreased. Our finding indicates that small interfering RNA (siRNA) interference targeting the PGC-1α gene could inhibit the function of mitochondria in several capacities and that the PGC-1α gene may modulate mitochondrial function by regulating the expression of ERRα, NRF-1, NRF-2 and PPARγ. Thus, PGC-1α can be considered a potential therapeutic target for PD. PMID:28611589

Shikonin inhibits oxidized LDL-induced monocyte adhesion by suppressing NFκB activation via up-regulation of PI3K/Akt/Nrf2-dependent antioxidation in EA.hy926 endothelial cells.

PubMed

Huang, Chin-Shiu; Lin, Ai-Hsuan; Yang, Ting-Chun; Liu, Kai-Li; Chen, Haw-Wen; Lii, Chong-Kuei

2015-02-01

Oxidized low-density lipoprotein (oxLDL) is a key contributor to atherogenesis through multiple mechanisms, including the reactive oxygen species (ROS)-mediated nuclear factor-kappaB (NFκB) signaling pathway. Although shikonin, one of the main active components isolated from the Chinese herb Lithospermum erythrorhizon, has been shown to possess cardioprotective, antioxidative, and anti-inflammatory effects, the mechanisms underlying these actions are not well understood. In this study, we used EA.hy926 endothelial-like cells to examine the anti-atherogenic activity of shikonin. Shikonin (0-1 μM) concentration-dependently induced heme oxygenase-1, glutamate cysteine ligase modifier subunit, catalase, superoxide dismutase 1, glutathione peroxidase 1, and glutathione reductase protein and mRNA expression and glutathione content via activation of the phosphatidylinositol 3-kinase (PI3K)/Akt/Nrf2 signaling pathway. In the presence of oxLDL (40 μg/ml), shikonin pretreatment reversed oxLDL-induced ROS production, antioxidant response element reporter activity, NFκB nuclear translocation, and intercellular adhesion molecule (ICAM)-1 and E-selectin expression and suppressed the increase of monocyte adhesion to endothelial cells. Nrf2 knockdown by using RNA interference attenuated the ability of shikonin to inhibit oxLDL-induced NFκB DNA binding activity, adhesion molecule expression, and monocyte adhesion. Taken together, these results suggest that shikonin protects against oxLDL-induced endothelial damage by suppressing ROS/NFκB-mediated ICAM-1 and E-selectin expression via up-regulation of PI3K/Akt/Nrf2-dependent antioxidant enzyme expression. Copyright © 2014 Elsevier Inc. All rights reserved.

Anti-leukaemic effects induced by APR-246 are dependent on induction of oxidative stress and the NFE2L2/HMOX1 axis that can be targeted by PI3K and mTOR inhibitors in acute myeloid leukaemia cells.

PubMed

Ali, Dina; Mohammad, Dara K; Mujahed, Huthayfa; Jonson-Videsäter, Kerstin; Nore, Beston; Paul, Christer; Lehmann, Sören

2016-07-01

The small molecule APR-246 (PRIMA-1(MET) ) is a novel drug that restores the activity of mutated and unfolded TP53 protein. However, the mechanisms of action and potential off-target effects are not fully understood. Gene expression profiling in TP53 mutant KMB3 acute myeloid leukaemia (AML) cells showed that genes which protected cells from oxidative stress to be the most up-regulated. APR-246 exposure also induced reactive oxygen species (ROS) formation and depleted glutathione in AML cells. The genes most up-regulated by APR-246, confirmed by quantitative real time polymerase chain reaction, were heme oxygenase-1 (HMOX1, also termed HO-1), SLC7A11 and RIT1. Up-regulation of HMOX1, a key regulator of cellular response to ROS, was independent of TP53 mutational status. NFE2L2 (also termed Nrf2), a master regulator of HMOX1 expression, showed transcriptional up-regulation and nuclear translocation by APR-246. Down-regulation of NFE2L2 by siRNA in AML cells significantly increased the antitumoural effects of APR-246. The PI3K inhibitor wortmannin and the mTOR inhibitor rapamycin inhibited APR-246-induced nuclear translocation of NFE2L2 and counteracted the protective cellular responses to APR-246, resulting in synergistic cell killing together with APR-246. In conclusion, ROS induction is important for antileukaemic activities of APR-246 and inhibiting the protective response of the Nrf-2/HMOX1 axis using PI3K inhibitors, enhances the antileukaemic effects. © 2016 John Wiley & Sons Ltd.

Topical application of ointment containing 0.5% green tea catechins suppresses tongue oxidative stress in 5-fluorouracil administered rats.

PubMed

Miyai, Hisataka; Maruyama, Takayuki; Tomofuji, Takaaki; Yoneda, Toshiki; Azuma, Tetsuji; Mizuno, Hirofumi; Sugiura, Yoshio; Kobayashi, Terumasa; Ekuni, Daisuke; Morita, Manabu

2017-10-01

The purpose of this study was to investigate the preventive effects of topical application of green tea catechins on tongue oxidative stress induced by 5-fluorouracil (5-FU) administration in rats. Male Wistar rats (n=28, 8 weeks old) were divided into four groups of seven rats each: a negative control group (saline administration and application of ointment without green tea catechins), a positive control group (5-FU administration and application of ointment without green tea catechins), and two experimental groups (5-FU administration and application of ointment containing 0.1% or 0.5% green tea catechins). Topical application of each ointment to the ventral surface of the tongue was performed once a day for 5days. The level of 8-hydroxydeoxyguanosine (8-OHdG) was determined to evaluate oxidative stress. Fluorescence staining was also performed to confirm nuclear factor erythroid 2-related factor 2 (Nrf2) translocation to the nucleus. After the experimental period, the ratios of 8-OHdG-positive cells in the ventral tongue tissue were higher in the positive control group than in the negative control group (P<0.05). On the other hand, those in the 0.5% green tea catechin group, but not in the 0.1% green tea catechin group, were lower than the positive control group (P<0.05). In addition, Nrf2 translocation to the nucleus was greater in the 0.5% green tea catechin group than in the positive control group (P<0.05). Topical application of ointment containing 0.5% green tea catechins could prevent tongue oxidative stress in 5-FU administered rats, via up-regulation of the Nrf2 signaling pathway. Copyright © 2017 Elsevier Ltd. All rights reserved.

Arctigenin Increases Hemeoxygenase-1 Gene Expression by Modulating PI3K/AKT Signaling Pathway in Rat Primary Astrocytes.

PubMed

Jeong, Yeon-Hui; Park, Jin-Sun; Kim, Dong-Hyun; Kim, Hee-Sun

2014-11-01

In the present study, we found that the natural compound arctigenin inhibited hydrogen peroxide-induced reactive oxygen species (ROS) production in rat primary astrocytes. Since hemeoxygenase-1 (HO-1) plays a critical role as an antioxidant defense factor in the brain, we examined the effect of arctigenin on HO-1 expression in rat primary astrocytes. We found that arctigenin increased HO-1 mRNA and protein levels. Arctigenin also increases the nuclear translocation and DNA binding of Nrf2/c-Jun to the antioxidant response element (ARE) on HO-1 promoter. In addition, arctigenin increased ARE-mediated transcriptional activities in rat primary astrocytes. Further mechanistic studies revealed that arctigenin increased the phosphorylation of AKT, a downstream substrate of phosphatidylinositol 3-kinase (PI3K). Treatment of cells with a PI3K-specific inhibitor, LY294002, suppressed the HO-1 expression, Nrf2 DNA binding and ARE-mediated transcriptional activities in arctigenin-treated astrocyte cells. The results collectively suggest that PI3K/AKT signaling pathway is at least partly involved in HO-1 expression by arctigenin via modulation of Nrf2/ARE axis in rat primary astrocytes.

Arctigenin Increases Hemeoxygenase-1 Gene Expression by Modulating PI3K/AKT Signaling Pathway in Rat Primary Astrocytes

PubMed Central

Jeong, Yeon-Hui; Park, Jin-Sun; Kim, Dong-Hyun; Kim, Hee-Sun

2014-01-01

In the present study, we found that the natural compound arctigenin inhibited hydrogen peroxide-induced reactive oxygen species (ROS) production in rat primary astrocytes. Since hemeoxygenase-1 (HO-1) plays a critical role as an antioxidant defense factor in the brain, we examined the effect of arctigenin on HO-1 expression in rat primary astrocytes. We found that arctigenin increased HO-1 mRNA and protein levels. Arctigenin also increases the nuclear translocation and DNA binding of Nrf2/c-Jun to the antioxidant response element (ARE) on HO-1 promoter. In addition, arctigenin increased ARE-mediated transcriptional activities in rat primary astrocytes. Further mechanistic studies revealed that arctigenin increased the phosphorylation of AKT, a downstream substrate of phosphatidylinositol 3-kinase (PI3K). Treatment of cells with a PI3K-specific inhibitor, LY294002, suppressed the HO-1 expression, Nrf2 DNA binding and ARE-mediated transcriptional activities in arctigenin-treated astrocyte cells. The results collectively suggest that PI3K/AKT signaling pathway is at least partly involved in HO-1 expression by arctigenin via modulation of Nrf2/ARE axis in rat primary astrocytes. PMID:25489416

Z-ligustilide ameliorated ultraviolet B-induced oxidative stress and inflammatory cytokine production in human keratinocytes through upregulation of Nrf2/HO-1 and suppression of NF-κB pathway.

PubMed

Wu, Zhouwei; Uchi, Hiroshi; Morino-Koga, Saori; Shi, Weimin; Furue, Masutaka

2015-09-01

Ultraviolet B (UVB), a harmful environmental factor, is responsible for a variety of skin disorders including skin inflammation through reactive oxygen species (ROS) and inflammatory mediator production. Here, we investigated the effect of Z-ligustilide (Z-lig), an active ingredient isolated from the medicinal plants Cnidium officinale and Angelica acutiloba, on UVB-induced ROS generation and inflammatory mediator production in normal human epidermal keratinocytes (NHEKs) as well as its underlying mechanisms. Z-lig significantly rescued UVB-induced NHEKs damage in a dosage-dependent manner. Pretreatment of NHEKs with Z-lig inhibited UVB-induced ROS production in NHEKs. Both silencing the nuclear factor E2-related factor 2 (Nrf2) and the supplement of tin protoporphyrin IX (SnPP), a haeme oxygenase-1 (HO-1) inhibitor, cancelled the inhibitory effect of Z-lig on UVB-induced ROS upregulation in NHEKs. Moreover, pretreatment of NHEKs with Z-lig reduced UVB-induced nuclear factor kappa B (NF-κB)-dependent inflammatory mediators (IL-6, IL-8 and MCP-1) production at both mRNA and protein level. In the presence of Z-lig, UVB-induced NF-κB subunit p65 nuclear translocation was abolished, and the IκBα degradation was suppressed. Taken together, these findings suggest that Z-lig can suppress UVB-induced ROS generation through Nrf2/HO-1 upregulation and inflammation by suppressing the NF-κB pathway, suggesting that Z-lig may be beneficial in protecting skin from UVB exposure. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Protective Effects of Maillard Reaction Products of Whey Protein Concentrate against Oxidative Stress through an Nrf2-Dependent Pathway in HepG2 Cells.

PubMed

Pyo, Min Cheol; Yang, Sung-Yong; Chun, Su-Hyun; Oh, Nam Su; Lee, Kwang-Won

2016-09-01

Whey protein concentrate (WPC), which contains α-lactalbumin and β-lactoglobulin, is utilized widely in the food industry. The Maillard reaction is a complex reaction that produces Maillard reaction products (MRPs), which are associated with the formation of antioxidant compounds. In this study, the hepatoprotection activity of MRPs of WPC against oxidative stress through the nuclear factor-E2-related factor 2 (Nrf2)-dependent antioxidant pathway in HepG2 cells was examined. Glucose-whey protein concentrate conjugate (Glc-WPC) was obtained from Maillard reaction between WPC and glucose. The fluorescence intensity of Glc-WPC increased after 7 d compared to native WPC, and resulted in loss of 48% of the free amino groups of WPC. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns of Glc-WPC showed the presence of a high-molecular-weight portion. Treatment of HepG2 cells with Glc-WPC increased cell viability in the presence of oxidative stress, inhibited the generation of intracellular reactive oxygen species by tert-butyl hydroperoxide (t-BHP), and increased the glutathione level. Nrf2 translocation and Nrf2, reduced nicotinamide adenine dinucleotide phosphate (NAD(P)H)-quinone oxidoreductase 1 (NOQ1), heme oxygenase-1 (HO-1), glutamate-L-cysteine ligase (GCL)M and GCLC mRNA levels were increased by Glc-WPC. Also, Glc-WPC increased the phosphorylation of extracellular signal-regulated kinase (ERK) 1/2 and c-Jun N-terminal kinase (JNK). The results of this study demonstrate that Glc-WPC activates the Nrf2-dependent pathway through the phosphorylation of ERK1/2 and JNK in HepG2 cells, and induces production of antioxidant enzymes and phase II enzymes.

Docosahexaenoic Acid (DHA) Provides Neuroprotection in Traumatic Brain Injury Models via Activating Nrf2-ARE Signaling.

PubMed

Zhu, Wei; Ding, Yuexia; Kong, Wei; Li, Tuo; Chen, Hongguang

2018-04-16

In this study, we explored the neuroprotective effects of docosahexaenoic acid (DHA) in traumatic brain injury (TBI) models. In this study, we first confirmed that DHA was neuroprotective against TBI via the NSS test and Morris water maze experiment. Western blot was conducted to identify the expression of Bax, caspase-3, and Bcl-2. And the cell apoptosis of the TBI models was validated by TUNEL staining. Relationships between nuclear factor erythroid 2-related factor 2-antioxidant response element (Nrf2-ARE) pathway-related genes and DHA were explored by RT-PCR and Western blot. Rats of the DHA group performed remarkably better than those of the TBI group in both NSS test and water maze experiment. DHA conspicuously promoted the expression of Bcl-2 and diminished that of cleaved caspase-3 and Bax, indicating the anti-apoptotic role of DHA. Superoxide dismutase (SOD) activity and cortical malondialdehyde content, glutathione peroxidase (GPx) activity were renovated in rats receiving DHA treatment, implying that the neuroprotective influence of DHA was derived from lightening the oxidative stress caused by TBI. Moreover, immunofluorescence and Western blot experiments revealed that DHA facilitated the translocation of Nrf2 to the nucleus. DHA administration also notably increased the expression of the downstream factors NAD(P)H:quinone oxidoreductase (NQO-1) and heme oxygenase 1(HO-1). DHA exerted neuroprotective influence on the TBI models, potentially through activating the Nrf2- ARE pathway.

Penicillinolide A: a new anti-inflammatory metabolite from the marine fungus Penicillium sp. SF-5292.

PubMed

Lee, Dong-Sung; Ko, Wonmin; Quang, Tran Hong; Kim, Kyoung-Su; Sohn, Jae Hak; Jang, Jae-Hyuk; Ahn, Jong Seog; Kim, Youn-Chul; Oh, Hyuncheol

2013-11-12

In the course of studies on bioactive metabolites from marine fungi, a new 10-membered lactone, named penicillinolide A (1) was isolated from the organic extract of Penicillium sp. SF-5292 as a potential anti-inflammatory compound. The structure of penicillinolide A (1) was mainly determined by analysis of NMR and MS data and Mosher's method. Penicillinolide A (1) inhibited the production of NO and PGE2 due to inhibition of the expression of iNOS and COX-2. Penicillinolide A (1) also reduced TNF-α, IL-1β and IL-6 production, and these anti-inflammatory effects were shown to be correlated with the suppression of the phosphorylation and degradation of IκB-α, NF-κB nuclear translocation, and NF-κB DNA binding activity. In addition, using inhibitor tin protoporphyrin (SnPP), a competitive inhibitor of HO activity, it was verified that the inhibitory effects of compound 1 on the production of pro-inflammatory mediators and NF-κB DNA binding activity were partially associated with HO-1 expression through Nrf2 nuclear translocation.

Penicillinolide A: A New Anti-Inflammatory Metabolite from the Marine Fungus Penicillium sp. SF-5292

PubMed Central

Lee, Dong-Sung; Ko, Wonmin; Quang, Tran Hong; Kim, Kyoung-Su; Sohn, Jae Hak; Jang, Jae-Hyuk; Ahn, Jong Seog; Kim, Youn-Chul; Oh, Hyuncheol

2013-01-01

In the course of studies on bioactive metabolites from marine fungi, a new 10-membered lactone, named penicillinolide A (1) was isolated from the organic extract of Penicillium sp. SF-5292 as a potential anti-inflammatory compound. The structure of penicillinolide A (1) was mainly determined by analysis of NMR and MS data and Mosher’s method. Penicillinolide A (1) inhibited the production of NO and PGE2 due to inhibition of the expression of iNOS and COX-2. Penicillinolide A (1) also reduced TNF-α, IL-1β and IL-6 production, and these anti-inflammatory effects were shown to be correlated with the suppression of the phosphorylation and degradation of IκB-α, NF-κB nuclear translocation, and NF-κB DNA binding activity. In addition, using inhibitor tin protoporphyrin (SnPP), a competitive inhibitor of HO activity, it was verified that the inhibitory effects of compound 1 on the production of pro-inflammatory mediators and NF-κB DNA binding activity were partially associated with HO-1 expression through Nrf2 nuclear translocation. PMID:24225730

UV-A Irradiation Activates Nrf2-Regulated Antioxidant Defense and Induces p53/Caspase3-Dependent Apoptosis in Corneal Endothelial Cells.

PubMed

Liu, Cailing; Vojnovic, Dijana; Kochevar, Irene E; Jurkunas, Ula V

2016-04-01

To examine whether Nrf2-regulated antioxidant defense and p53 are activated in human corneal endothelial cells (CEnCs) by environmental levels of ultraviolet A (UV-A), a known stimulator of oxidative stress. Immortalized human CEnCs (HCEnCi) were exposed to UV-A fluences of 2.5, 5, 10, or 25 J/cm2, then allowed to recover for 3 to 24 hours. Control HCEnCi did not receive UV-A. Reactive oxygen species (ROS) were measured using H2DCFDA. Cell cytotoxicity was evaluated by lactate dehydrogenase (LDH) release. Levels of Nrf2, HO-1, NQO-1, p53, and caspase3 were detected by immunnoblotting or real-time PCR. Activated caspase3 was measured by immunoblotting and a fluorescence assay. Exposure of HCEnCi to 5, 10, and 25 J/cm2 UV-A increased ROS levels compared with controls. Nrf2, HO-1, and NQO-1 mRNA increased 1.7- to 3.2-fold at 3 and 6 hours after irradiation with 2.5 and 5 J/cm2 UV-A. At 6 hours post irradiation, UV-A (5 J/cm2) enhanced nuclear Nrf2 translocation. At 24 hours post treatment, UV-A (5, 10, and 25 J/cm2) produced a 1.8- to 2.8-fold increase in phospho-p53 and a 2.6- to 6.0-fold increase in activated caspase3 compared with controls, resulting in 20% to 42% cell death. Lower fluences of UV-A induce Nrf2-regulated antioxidant defense and higher fluences activate p53 and caspase3, indicating that even near-environmental levels of UV-A may affect normal CEnCs. This data suggest that UV-A may especially damage cells deficient in antioxidant defense, and thus may be involved in the etiology of Fuchs' endothelial corneal dystrophy (FECD).

UV-A Irradiation Activates Nrf2-Regulated Antioxidant Defense and Induces p53/Caspase3-Dependent Apoptosis in Corneal Endothelial Cells

PubMed Central

Liu, Cailing; Vojnovic, Dijana; Kochevar, Irene E.; Jurkunas, Ula V.

2016-01-01

Purpose To examine whether Nrf2-regulated antioxidant defense and p53 are activated in human corneal endothelial cells (CEnCs) by environmental levels of ultraviolet A (UV-A), a known stimulator of oxidative stress. Methods Immortalized human CEnCs (HCEnCi) were exposed to UV-A fluences of 2.5, 5, 10, or 25 J/cm2, then allowed to recover for 3 to 24 hours. Control HCEnCi did not receive UV-A. Reactive oxygen species (ROS) were measured using H2DCFDA. Cell cytotoxicity was evaluated by lactate dehydrogenase (LDH) release. Levels of Nrf2, HO-1, NQO-1, p53, and caspase3 were detected by immunnoblotting or real-time PCR. Activated caspase3 was measured by immunoblotting and a fluorescence assay. Results Exposure of HCEnCi to 5, 10, and 25 J/cm2 UV-A increased ROS levels compared with controls. Nrf2, HO-1, and NQO-1 mRNA increased 1.7- to 3.2-fold at 3 and 6 hours after irradiation with 2.5 and 5 J/cm2 UV-A. At 6 hours post irradiation, UV-A (5 J/cm2) enhanced nuclear Nrf2 translocation. At 24 hours post treatment, UV-A (5, 10, and 25 J/cm2) produced a 1.8- to 2.8-fold increase in phospho-p53 and a 2.6- to 6.0-fold increase in activated caspase3 compared with controls, resulting in 20% to 42% cell death. Conclusions Lower fluences of UV-A induce Nrf2-regulated antioxidant defense and higher fluences activate p53 and caspase3, indicating that even near-environmental levels of UV-A may affect normal CEnCs. This data suggest that UV-A may especially damage cells deficient in antioxidant defense, and thus may be involved in the etiology of Fuchs' endothelial corneal dystrophy (FECD). PMID:27127932

Activation of Nrf2/HO-1signaling pathway involves the anti-inflammatory activity of magnolol in Porphyromonas gingivalis lipopolysaccharide-stimulated mouse RAW 264.7 macrophages.

PubMed

Lu, Sheng-Hua; Hsu, Wen-Lin; Chen, Tso-Hsiao; Chou, Tz-Chong

2015-12-01

Magnolol isolated from Magnolia officinalis, a Chinese medical herb, exhibits an anti-inflammatory activity and a protective effect against periodontitis. The inflammation caused by lipopolysaccharide (LPS) from Porphyromonas gingivalis (P. gingivalis) has been considered a key inducer in the development of periodontitis. In this study, we investigated whether magnolol inhibits P. gingivalis LPS-evoked inflammatory responses in RAW 264.7 macrophages and the involvement of heme oxygenase-1 (HO-1). Magnolol significantly activated p38 MAPK, Nrf-2/HO-1 cascade and reactive oxygen species (ROS) formation. Notably, the Nrf-2 activation and HO-1 induction by magnolol were greatly diminished by blocking p38 MAPK activity and ROS production. Furthermore, in P. gingivalis LPS-stimulated macrophages, magnolol treatment remarkably inhibited the inflammatory responses evidenced by suppression of pro-inflammatory cytokine, prostaglandin E2, nitrite formation, and the expression of inducible nitric oxide synthase and cyclooxygenase-2, as well as NF-κB activation accompanied by a significant elevation of Nrf-2 nuclear translocation and HO-1 expression/activity. However, inhibiting HO-1 activity with tin protoporphyrin IX markedly reversed the anti-inflammatory effects of magnolol. Collectively, these findings provide a novel mechanism by which magnolol inhibits P. gingivalis LPS-induced inflammation in macrophages is at least partly mediated by HO-1 activation, and thereby promoting its clinical use in periodontitis. Copyright © 2015 Elsevier B.V. All rights reserved.

Modulation of Nrf2/Keap1 system by Wasabi 6-methylthiohexyl isothiocyanate in ARE-mediated NQO1 expression.

PubMed

Korenori, Yoshimi; Tanigawa, Shunsuke; Kumamoto, Takuma; Qin, Si; Daikoku, Yosuke; Miyamori, Koji; Nagai, Masashi; Hou, De-Xing

2013-05-01

6-Methylthiohexyl isothiocyanate (6-MTITC), one of the major bioactive ingredients in Japanese Wasabi, has revealed cytoprotective and cancer chemopreventive effects. This study aims to clarify the molecular mechanisms how 6-MTITC modulates nuclear factor E2-related factor 2 (Nrf2)/Kelchlike ECH-associating protein 1 (Keap1) system in antioxidant-responsive element (ARE)-mediated nicotinamide adenine dinucleotide phosphate (NADP): quinone oxidoreductase 1 (NQO1) expression. HepG2 cells were treated with 6-MTITC with varying time and dose. NQO1, Nrf2, and Keap1 proteins were detected by Western blotting. ARE transactivation was detected by electrophilic mobility shift assay and reporter gene assay. Nuclear localization of Nrf2 was determined by immunocytochemistry assay. Ubiquitination of Nrf2 and Keap1 was detected using immunoprecipitation after treatment with MG132. Small interfering RNA was used to knockdown Nrf2 or Keap1. The results revealed that 6-MTITC modulated Nrf2/ARE pathway by stimulating Keap1 modification, and inhibiting Nrf2 ubiquitination and protein turnover. These actions finally increased nuclear Nrf2 accumulation and ARE-binding activity. Moreover, silencing Nrf2 markedly reduced ARE-driven activity induced by 6-MTITC. 6-MTITC modulated ARE-driven NQO1 expression by stabilizing Nrf2 with enhanced Keap1 modification and decreased Nrf2 degradation. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Dual mechanisms of NF-kappaB inhibition in carnosol-treated endothelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lian, K.-C.; Chuang, J.-J.; Hsieh, C.-W.

2010-05-15

The increased adhesion of monocytes to injured endothelial layers is a critical early event in atherogenesis. Under inflammatory conditions, there is increased expression of specific cell adhesion molecules on activated vascular endothelial cells, which increases monocyte adhesion. In our current study, we demonstrate a putative mechanism for the anti-inflammatory effects of carnosol, a diterpene derived from the herb rosemary. Our results show that both carnosol and rosemary essential oils inhibit the adhesion of TNFalpha-induced monocytes to endothelial cells and suppress the expression of ICAM-1 at the transcriptional level. Moreover, carnosol was found to exert its inhibitory effects by blocking themore » degradation of the inhibitory protein IkappaBalpha in short term pretreatments but not in 12 h pretreatments. Our data show that carnosol reduces IKK-beta phosphorylation in pretreatments of less than 3 h. In TNFalpha-treated ECs, NF-kappaB nuclear translocation and transcriptional activity was abolished by up to 12 h of carnosol pretreatment and this was blocked by Nrf-2 siRNA. The long-term inhibitory effects of carnosol thus appear to be mediated through its induction of Nrf-2-related genes. The inhibition of ICAM-1 expression and p65 translocation is reversed by HO-1 siRNA. Carnosol also upregulates the Nrf-2-related glutathione synthase gene and thereby increases the GSH levels after 9 h of exposure. Treating ECs with a GSH synthesis inhibitor, BSO, blocks the inhibitory effects of carnosol. In addition, carnosol increases p65 glutathionylation. Hence, our present findings indicate that carnosol suppresses TNFalpha-induced singling pathways through the inhibition of IKK-beta activity or the upregulation of HO-1 expression. The resulting GSH levels are dependent, however, on the length of the carnosol pretreatment period.« less

Schisandra chinensis regulates drug metabolizing enzymes and drug transporters via activation of Nrf2-mediated signaling pathway

PubMed Central

He, Jin-Lian; Zhou, Zhi-Wei; Yin, Juan-Juan; He, Chang-Qiang; Zhou, Shu-Feng; Yu, Yang

2015-01-01

Drug metabolizing enzymes (DMEs) and drug transporters are regulated via epigenetic, transcriptional, posttranscriptional, and translational and posttranslational modifications. Phase I and II DMEs and drug transporters play an important role in the disposition and detoxification of a large number of endogenous and exogenous compounds. The nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is a critical regulator of a variety of important cytoprotective genes that are involved in disposition and detoxification of xenobiotics. Schisandra chinensis (SC) is a commonly used traditional Chinese herbal medicine that has been primarily used to protect the liver because of its potent antioxidative and anti-inflammatory activities. SC can modulate some DMEs and drug transporters, but the underlying mechanisms are unclear. In this study, we aimed to explore the role of Nrf2 in the regulatory effect of SC extract (SCE) on selected DMEs and drug transporters in human hepatocellular liver carcinoma cell line (HepG2) cells. The results showed that SCE, schisandrin A, and schisandrin B significantly increased the expression of NAD(P)H: Nicotinamide Adenine Dinucleotide Phosphate-oxidase or:quinone oxidoreductase 1, heme oxygenase-1, glutamate–cysteine ligase, and glutathione S-transferase A4 at both transcriptional and posttranscriptional levels. Incubation of HepG2 cells with SCE resulted in a significant increase in the intracellular level of glutathione and total glutathione S-transferase content. SCE significantly elevated the messenger ribonucleic acid and protein levels of P-glycoprotein and multidrug resistance-associated protein 2 and 4, whereas the expression of organic anion transporting peptide 1A2 and 1B1 was significantly downregulated by SCE. Knockdown of Nrf2 by small interfering ribonucleic acid attenuated the regulatory effect of SCE on these DMEs and drug transporters. SCE significantly upregulated Nrf2 and promoted the translocation of Nrf2 from cytoplasm to the nuclei. Additionally, SCE significantly suppressed the expression of cytosolic Kelch-like ECH-associated protein 1 (the repressor of Nrf2) and remarkably increased Nrf2 stability in HepG2 cells. Taken together, our findings suggest that the hepatoprotective effects of SCE may be partially ascribed to the modulation of DMEs and drug transporters via Nrf2-mediated signaling pathway. SCE may alter the pharmacokinetics of other coadministered drugs that are substrates of these DMEs and transporters and thus cause unfavorable herb–drug interactions. PMID:25552902

Berberine, a natural antidiabetes drug, attenuates glucose neurotoxicity and promotes Nrf2-related neurite outgrowth

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hsu, Ya-Yun; Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung 80708, Taiwan; Tseng, Yu-Ting

Reactive oxygen intermediates production and apoptotic damage induced by high glucose are major causes of neuronal damage in diabetic neuropathy. Berberine (BBR), a natural antidiabetes drug with PI3K-activating activity, holds promise for diabetes because of its dual antioxidant and anti-apoptotic activities. We have previously reported that BBR attenuated H{sub 2}O{sub 2} neurotoxicity via activating the PI3K/Akt/Nrf2-dependent pathway. In this study, we further explored the novel protective mechanism of BBR on high glucose-induced apoptotic death and neurite damage of SH-SY5Y cells. Results indicated BBR (0.1–10 nM) significantly attenuated reactive oxygen species (ROS) production, nucleus condensation, and apoptotic death in high glucose-treatedmore » cells. However, AG1024, an inhibitor of insulin growth factor-1 (IGF-1) receptor, significantly abolished BBR protection against high glucose-induced neuronal death. BBR also increased Bcl-2 expression and decreased cytochrome c release. High glucose down-regulated IGF-1 receptor and phosphorylation of Akt and GSK-3β, the effects of which were attenuated by BBR treatment. BBR also activated nuclear erythroid 2-related factor 2 (Nrf2), the key antioxidative transcription factor, which is accompanied with up-regulation of hemeoxygenase-1 (HO-1). Furthermore, BBR markedly enhanced nerve growth factor (NGF) expression and promoted neurite outgrowth in high glucose-treated cells. To further determine the role of the Nrf2 in BBR neuroprotection, RNA interference directed against Nrf2 was used. Results indicated Nrf2 siRNA abolished BBR-induced HO-1, NGF, neurite outgrowth and ROS decrease. In conclusion, BBR attenuated high glucose-induced neurotoxicity, and we are the first to reveal this novel mechanism of BBR as an Nrf2 activator against glucose neurotoxicity, providing another potential therapeutic use of BBR on the treatment of diabetic complications. - Highlights: • BBR attenuates high glucose-induced ROS production and neuronal cell death. • BBR activates IGF-1/Akt/GSK-3β signaling under normal and high glucose conditions. • BBR enhances HO-1 and NGF expression through stimulating Nrf2 translocation. • BBR promotes neurite outgrowth through Nrf2-dependent pathway.« less

Mild Hypothermia Attenuates Mitochondrial Oxidative Stress by Protecting Respiratory Enzymes and Upregulating MnSOD in a Pig Model of Cardiac Arrest

PubMed Central

Gong, Ping; Li, Chun-Sheng; Hua, Rong; Zhao, Hong; Tang, Zi-Ren; Mei, Xue; Zhang, Ming-Yue; Cui, Juan

2012-01-01

Mild hypothermia is the only effective treatment confirmed clinically to improve neurological outcomes for comatose patients with cardiac arrest. However, the underlying mechanism is not fully elucidated. In this study, our aim was to determine the effect of mild hypothermia on mitochondrial oxidative stress in the cerebral cortex. We intravascularly induced mild hypothermia (33°C), maintained this temperature for 12 h, and actively rewarmed in the inbred Chinese Wuzhishan minipigs successfully resuscitated after 8 min of untreated ventricular fibrillation. Cerebral samples were collected at 24 and 72 h following return of spontaneous circulation (ROSC). We found that mitochondrial malondialdehyde and protein carbonyl levels were significantly increased in the cerebral cortex in normothermic pigs even at 24 h after ROSC, whereas mild hypothermia attenuated this increase. Moreover, mild hypothermia attenuated the decrease in Complex I and Complex III (i.e., major sites of reactive oxygen species production) activities of the mitochondrial respiratory chain and increased antioxidant enzyme manganese superoxide dismutase (MnSOD) activity. This increase in MnSOD activity was consistent with the upregulation of nuclear factor erythroid 2-related factor 2 (Nrf2) mRNA and protein expressions, and with the increase of Nrf2 nuclear translocation in normothermic pigs at 24 and 72 h following ROSC, whereas mild hypothermia enhanced these tendencies. Thus, our findings indicate that mild hypothermia attenuates mitochondrial oxidative stress in the cerebral cortex, which may be associated with reduced impairment of mitochondrial respiratory chain enzymes, and enhancement of MnSOD activity and expression via Nrf2 activation. PMID:22532848

Taraxacum coreanum protects against glutamate-induced neurotoxicity through heme oxygenase-1 expression in mouse hippocampal HT22 cells.

PubMed

Yoon, Chi-Su; Ko, Wonmin; Lee, Dong-Sung; Kim, Dong-Cheol; Kim, Jongsu; Choi, Moonbum; Beom, Jin Seon; An, Ren-Bo; Oh, Hyuncheol; Kim, Youn-Chul

2017-04-01

Taraxacum coreanum Nakai is a dandelion that is native to Korea, and is widely used as an edible and medicinal herb. The present study revealed the neuroprotective effect of this plant against glutamate-induced oxidative stress in HT22 murine hippocampal neuronal cells. Ethanolic extracts from the aerial (TCAE) and the root parts (TCRE) of T. coreanum were prepared. Both extracts were demonstrated, by high performance liquid chromatography, to contain caffeic acid and ferulic acid as representative constituents. TCAE and TCRE significantly increased cell viability against glutamate-induced oxidative stress in mouse hippocampal HT22 cells. Western blot analysis revealed that treatment of HT22 cells with the extracts induced increased expression of the enzyme heme oxygenase-1 (HO-1), compared with untreated cells, in a concentration-dependent manner. Increased HO-1 enzymatic activity, compared with untreated cells, was also demonstrated following treatment with TCAE and TCRE. In addition, western blot analysis of the nuclear fractions of both TCAE and TCRE-treated HT22 cells revealed increased levels of nuclear factor erythroid 2 like 2 (Nrf2) compared with untreated cells, and decreased Nrf2 levels in the cytoplasmic fraction compared with untreated cells. The present study suggested that the neuroprotective effect of T. coreanum is associated with induction of HO-1 expression and Nrf2 translocation to the nucleus. Therefore, T. coreanum exhibits a promising function in prevention of neurodegeneration. Further studies will be required for the isolation and the full characterization of its active substances.

Insulin Inhibits Nrf2 Gene Expression via Heterogeneous Nuclear Ribonucleoprotein F/K in Diabetic Mice

PubMed Central

Ghosh, Anindya; Abdo, Shaaban; Zhao, Shuiling; Wu, Chin-Han; Shi, Yixuan; Lo, Chao-Sheng; Chenier, Isabelle; Alquier, Thierry; Filep, Janos G.; Ingelfinger, Julie R.; Zhang, Shao-Ling

2017-01-01

Oxidative stress induces endogenous antioxidants via nuclear factor erythroid 2–related factor 2 (Nrf2), potentially preventing tissue injury. We investigated whether insulin affects renal Nrf2 expression in type 1 diabetes (T1D) and studied its underlying mechanism. Insulin normalized hyperglycemia, hypertension, oxidative stress, and renal injury; inhibited renal Nrf2 and angiotensinogen (Agt) gene expression; and upregulated heterogeneous nuclear ribonucleoprotein F and K (hnRNP F and hnRNP K) expression in Akita mice with T1D. In immortalized rat renal proximal tubular cells, insulin suppressed Nrf2 and Agt but stimulated hnRNP F and hnRNP K gene transcription in high glucose via p44/42 mitogen-activated protein kinase signaling. Transfection with small interfering RNAs of p44/42 MAPK, hnRNP F, or hnRNP K blocked insulin inhibition of Nrf2 gene transcription. Insulin curbed Nrf2 promoter activity via a specific DNA-responsive element that binds hnRNP F/K, and hnRNP F/K overexpression curtailed Nrf2 promoter activity. In hyperinsulinemic-euglycemic mice, renal Nrf2 and Agt expression was downregulated, whereas hnRNP F/K expression was upregulated. Thus, the beneficial actions of insulin in diabetic nephropathy appear to be mediated, in part, by suppressing renal Nrf2 and Agt gene transcription and preventing Nrf2 stimulation of Agt expression via hnRNP F/K. These findings identify hnRNP F/K and Nrf2 as potential therapeutic targets in diabetes. PMID:28324005

5-Aminolevulinic acid with sodium ferrous citrate induces autophagy and protects cardiomyocytes from hypoxia-induced cellular injury through MAPK-Nrf-2-HO-1 signaling cascade

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhao, Mingyi; Guangdong Cardiovascular Institute, Guangdong General Hospital, Guangdong Academy of Medical Sciences, Guangzhou; Department of Pediatrics, The Third Xiangya Hospital, Central South University, Changsha

Background: Hypoxia causes cardiac disease via oxidative stress and mitochondrial dysfunction. 5-Aminolevulinic acid in combination with sodium ferrous citrate (ALA/SFC) has been shown to up-regulate heme oxygenase-1 (HO-1) and decrease macrophage infiltration and renal cell apoptosis in renal ischemia injury mice. However, its underlying mechanism remains largely unknown. The aim of this study was to investigate whether ALA/SFC could protect cardiomyocytes from hypoxia-induced apoptosis by autophagy via HO-1 signaling. Materials & methods: Murine atrial cardiomyocyte HL-1 cells were pretreated with ALA/SFC and then exposed to hypoxia. Results: ALA/SFC pretreatment significantly attenuated hypoxia-induced cardiomyocyte apoptosis, reactive oxygen species production, and mitochondrial injury,more » while it increased cell viability and autophagy levels. HO-1 expression by ALA/SFC was associated with up-regulation and nuclear translocation of Nrf-2, whereas Nrf-2 siRNA dramatically reduced HO-1 expression. ERK1/2, p38, and SAPK/JNK pathways were activated by ALA/SFC and their specific inhibitors significantly reduced ALA/SFC-mediated HO-1 upregulation. Silencing of either Nrf-2 or HO-1and LY294002, inhibitor of autophagy, abolished the protective ability of ALA/AFC against hypoxia-induced injury and reduced ALA/SFC-induced autophagy. Conclusion: Taken together, our data suggest that ALA/SFC induces autophagy via activation of MAPK/Nrf-2/HO-1 signaling pathway to protect cardiomyocytes from hypoxia-induced apoptosis. - Highlights: • ALA/SFC attenuates hypoxia-induced cardiomyocyte apoptosis, reactive oxygen species production, and mitochondrial injury. • ALA/SFC increases the heme oxygenase-1 expression via Nrf-2 and ERK1/2, p38, and SAPK/JNK pathways. • ALA/SFC induces autophagy and inhibition of autophagy prevent ALA/SFC-mediated suppression of hypoxia-induced injury.« less

Hydrogen sulphide induces HIF-1α and Nrf2 in THP-1 macrophages.

PubMed

Lohninger, Lilian; Tomasova, Lenka; Praschberger, Monika; Hintersteininger, Michael; Erker, Thomas; Gmeiner, Bernhard M K; Laggner, Hilde

2015-05-01

The transcription factor HIF-1α regulates the adaptive response of cells to hypoxia and oxidative stress. In addition, an important regulatory role for HIF-1α in immune reactions and inflammation is suggested. The present study attempts to investigate the effect of the gaseous signalling molecule hydrogen sulphide (H2S) on HIF-1α in THP-1 macrophages using the slow H2S releasing donor GYY4137. We found that H2S induced HIF-1α protein accumulation in THP-1 macrophages in a concentration-dependent manner. Western blot analysis of cell fractions showed that HIF-1α protein translocates into the nucleus and leads to an increase of its target protein glucose transporter-1 (GLUT-1). Activation of nuclear factor-κB (NF-κB), as well as secretion of the pro-inflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), were reduced in the presence of H2S. These findings indicate that HIF-1α accumulation due to H2S was not triggered by the NF-κB pathway. The antioxidant pathway Nrf2/HO-1 (nuclear factor erythroid 2-related factor 2/heme oxygenase-1) was activated by H2S. Inhibition of the p38 mitogen-activated protein kinase (MAPK) reversed H2S mediated effects, suggesting that the p38 MAPK pathway may be involved in H2S induced HIF-1α/Nrf2 signalling pathways. Copyright © 2015 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

Celecoxib exerts protective effects in the vascular endothelium via COX-2-independent activation of AMPK-CREB-Nrf2 signalling.

PubMed

Al-Rashed, Fahad; Calay, Damien; Lang, Marie; Thornton, Clare C; Bauer, Andrea; Kiprianos, Allan; Haskard, Dorian O; Seneviratne, Anusha; Boyle, Joseph J; Schönthal, Alex H; Wheeler-Jones, Caroline P; Mason, Justin C

2018-04-19

Although concern remains about the athero-thrombotic risk posed by cyclo-oxygenase (COX)-2-selective inhibitors, recent data implicates rofecoxib, while celecoxib appears equivalent to NSAIDs naproxen and ibuprofen. We investigated the hypothesis that celecoxib activates AMP kinase (AMPK) signalling to enhance vascular endothelial protection. In human arterial and venous endothelial cells (EC), and in contrast to ibuprofen and naproxen, celecoxib induced the protective protein heme oxygenase-1 (HO-1). Celecoxib derivative 2,5-dimethyl-celecoxib (DMC) which lacks COX-2 inhibition also upregulated HO-1, implicating a COX-2-independent mechanism. Celecoxib activated AMPKα (Thr172) and CREB-1 (Ser133) phosphorylation leading to Nrf2 nuclear translocation. Importantly, these responses were not reproduced by ibuprofen or naproxen, while AMPKα silencing abrogated celecoxib-mediated CREB and Nrf2 activation. Moreover, celecoxib induced H-ferritin via the same pathway, and increased HO-1 and H-ferritin in the aortic endothelium of mice fed celecoxib (1000 ppm) or control chow. Functionally, celecoxib inhibited TNF-α-induced NF-κB p65 (Ser536) phosphorylation by activating AMPK. This attenuated VCAM-1 upregulation via induction of HO-1, a response reproduced by DMC but not ibuprofen or naproxen. Similarly, celecoxib prevented IL-1β-mediated induction of IL-6. Celecoxib enhances vascular protection via AMPK-CREB-Nrf2 signalling, a mechanism which may mitigate cardiovascular risk in patients prescribed celecoxib. Understanding NSAID heterogeneity and COX-2-independent signalling will ultimately lead to safer anti-inflammatory drugs.

Cytochrome P450 2A5 and bilirubin: Mechanisms of gene regulation and cytoprotection

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Sangsoo Daniel; Antenos, Monica; Squires, E. James

2013-07-15

Bilirubin (BR) has recently been identified as the first endogenous substrate for cytochrome P450 2A5 (CYP2A5) and it has been suggested that CYP2A5 plays a major role in BR clearance as an alternative mechanism to BR conjugation by uridine-diphosphate glucuronyltransferase 1A1. This study investigated the mechanisms of Cyp2a5 gene regulation by BR and the cytoprotective role of CYP2A5 in BR hepatotoxicity. BR induced CYP2A5 expression at the mRNA and protein levels in a dose-dependent manner in primary mouse hepatocytes. BR treatment also caused nuclear translocation of Nuclear factor-E2 p45-related factor 2 (Nrf2) in hepatocytes. In reporter assays, BR treatment ofmore » primary hepatocytes transfected with a Cyp2a5 promoter-luciferase reporter construct resulted in a 2-fold induction of Cyp2a5 reporter activity. Furthermore, cotransfection of the hepatocytes with a Nrf2 expression vector without BR treatment resulted in an increase in Cyp2a5 reporter activity of approximately 2-fold and BR treatment of Nrf2 cotransfectants further increased reporter activity by 4-fold. In addition, site-directed mutation of the ARE in the reporter construct completely abolished both the BR- and Nrf2-mediated increases in reporter activity. The cytoprotective role of CYP2A5 against BR-mediated apoptosis was also examined in Hepa 1–6 cells that lack endogenous CYP2A5. Transient overexpression of CYP2A5 partially blocked BR-induced caspase-3 cleavage in Hepa 1–6 cells. Furthermore, in vitro degradation of BR was increased by microsomes from Hepa 1–6 cells overexpressing CYP2A5 compared to control cells transfected with an empty vector. Collectively, these results suggest that Nrf2-mediated CYP2A5 transactivation in response to BR may provide an additional mechanism for adaptive cytoprotection against BR hepatotoxicity. - Highlights: • The mechanism of Cyp2a5 gene regulation by BR was investigated. • The cytoprotective role of CYP2A5 in BR hepatotoxicity was determined. • BR induces CYP2A5 mRNA and protein expression. • BR increases CYP2A5 transcription via Nrf2 activation. • CYP2A5 overexpression increases BR clearance and reduces caspase-3 activation.« less

3,4-Dihydroxybenzalactone Suppresses Human Non-Small Cell Lung Carcinoma Cells Metastasis via Suppression of Epithelial to Mesenchymal Transition, ROS-Mediated PI3K/AKT/MAPK/MMP and NFκB Signaling Pathways.

PubMed

Chao, Wei; Deng, Jeng-Shyan; Li, Pei-Ying; Liang, Yu-Chia; Huang, Guan-Jhong

2017-03-28

3,4-Dihydroxybenzalactone (DBL) was isolated from Phellinus linteus (PL), which is a folk medicine possessing various physiological effects. In this study, we used highly metastatic A549 cells to investigate efficacy of DBL inhibition of cancer metastasis and possible mechanisms. The results revealed DBL inhibited migratory and invasive abilities of cancer cells at noncytotoxic concentrations. We found DBL suppressed enzymatic activities, protein expression, and RNA levels of matrix metalloproteinase (MMP)-2 and MMP-9. Western blot results showed DBL decreased phosphoinositide 3-kinase (PI3K)/AKT, phosphorylation status of mitogen-activated protein kinases (MAPKs), and focal adhesion kinase (FAK)/paxillin, which correlated with cell migratory ability. DBL also affected epithelial to mesenchymal transition (EMT)-related biomarkers. In addition, DBL enhanced cytoprotective effects through elevated antioxidant enzymes including heme oxygenase 1 (HO-1), catalase, glutathione peroxidase (GPx), and superoxide dismutase (SOD). Moreover, DBL influenced the nuclear translocation of nuclear factor κB (NFκB), nuclear factor erythroid 2-related factor 2 (Nrf2), Snail, and Slug in A549 cells. Taken together, these results suggested that treatment with DBL may act as a potential candidate to inhibit lung cancer metastasis by inhibiting MMP-2 and -9 via affecting PI3K/AKT, MAPKs, FAK/paxillin, EMT/Snail and Slug, Nrf2/antioxidant enzymes, and NFκB signaling pathways.

PKCδ Knockout Mice Are Protected from Dextromethorphan-Induced Serotonergic Behaviors in Mice: Involvements of Downregulation of 5-HT1A Receptor and Upregulation of Nrf2-Dependent GSH Synthesis.

PubMed

Tran, Hai-Quyen; Lee, Youngho; Shin, Eun-Joo; Jang, Choon-Gon; Jeong, Ji Hoon; Mouri, Akihiro; Saito, Kuniaki; Nabeshima, Toshitaka; Kim, Hyoung-Chun

2018-02-22

We investigated whether a specific serotonin (5-HT) receptor-mediated mechanism was involved in dextromethorphan (DM)-induced serotonergic behaviors. We firstly observed that the activation of 5-HT 1A receptor, but not 5-HT 2A receptor, contributed to DM-induced serotonergic behaviors in mice. We aimed to determine whether the upregulation of 5-HT 1A receptor induced by DM facilitates the specific induction of certain PKC isoform, because previous reports suggested that 5-HT 1A receptor activates protein kinase C (PKC). A high dose of DM (80 mg/kg, i.p.) induced a selective induction of PKCδ out of PKCα, PKCβI, PKCβII, PKCξ, and PKCδ in the hypothalamus of wild-type (WT) mice. More importantly, 5-HT 1A receptor co-immunoprecipitated PKCδ in the presence of DM. Consistently, rottlerin, a pharmacological inhibitor of PKCδ, or PKCδ knockout significantly protected against increases in 5-HT 1A receptor gene expression, 5-HT turnover rate, and serotonergic behaviors induced by DM. Treatment with DM resulted in an initial increase in nuclear factor erythroid-2-related factor 2 (Nrf2) nuclear translocation and DNA-binding activity, γ-glutamylcysteine (GCL) mRNA expression, and glutathione (GSH) level. This compensative induction was further potentiated by rottlerin or PKCδ knockout. However, GCL mRNA and GSH/GSSG levels were decreased 6 and 12 h post-DM. These decreases were attenuated by PKCδ inhibition. Our results suggest that interaction between 5-HT 1A receptor and PKCδ is critical for inducing DM-induced serotonergic behaviors and that inhibition of PKCδ attenuates the serotonergic behaviors via downregulation of 5-HT 1A receptor and upregulation of Nrf2-dependent GSH synthesis.

CDDO-Im protects from acetaminophen hepatotoxicity through induction of Nrf2-dependent genes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Reisman, Scott A.; Buckley, David B.; Tanaka, Yuji

CDDO-Im is a synthetic triterpenoid recently shown to induce cytoprotective genes through the Nrf2-Keap1 pathway, an important mechanism for the induction of cytoprotective genes in response to oxidative stress. Upon oxidative or electrophilic insult, the transcription factor Nrf2 translocates to the nucleus, heterodimerizes with small Maf proteins, and binds to antioxidant response elements (AREs) in the upstream promoter regions of various cytoprotective genes. To further elucidate the hepatoprotective effects of CDDO-Im, wild-type and Nrf2-null mice were pretreated with CDDO-Im (1 mg/kg, i.p.) or vehicle (DMSO), and then administered acetaminophen (500 mg/kg, i.p.). Pretreatment of wild-type mice with CDDO-Im reduced livermore » injury caused by acetaminophen. In contrast, hepatoprotection by CDDO-Im was not observed in Nrf2-null mice. CDDO-Im increased Nrf2 protein expression and Nrf2-ARE binding in wild-type, but not Nrf2-null mice. Furthermore, CDDO-Im increased the mRNA expression of the Nrf2 target genes NAD(P)H: quinone oxidoreductase-1 (Nqo1); glutamate-cysteine ligase, catalytic subunit (Gclc); and heme-oxygenase-1 (Ho-1), in both a dose- and time-dependent manner. Conversely, CDDO-Im did not induce Nqo1, Gclc, and Ho-1 mRNA expression in Nrf2-null mice. Collectively, the present study shows that CDDO-Im pretreatment induces Nrf2-dependent cytoprotective genes and protects the liver from acetaminophen-induced hepatic injury.« less

Baicalin Ameliorates Experimental Liver Cholestasis in Mice by Modulation of Oxidative Stress, Inflammation, and NRF2 Transcription Factor

PubMed Central

Feng, Xiaowen; Zhang, Feng; Xie, Haiyang

2017-01-01

Experimental cholestatic liver fibrosis was performed by bile duct ligation (BDL) in mice, and significant liver injury was observed in 15 days. Administration of baicalin in mice significantly ameliorates liver fibrosis. Experimental cholestatic liver fibrosis was associated with induced gene expression of fibrotic markers such as collagen I, fibronectin, alpha smooth muscle actin (SMA), and connective tissue growth factor (CTGF); increased inflammatory cytokines (TNFα, MIP1α, IL1β, and MIP2); increased oxidative stress and reactive oxygen species- (ROS-) inducing enzymes (NOX2 and iNOS); dysfunctional mitochondrial electron chain complexes; and apoptotic/necrotic cell death markers (DNA fragmentation, caspase 3 activity, and PARP activity). Baicalin administration on alternate day reduced fibrosis along with profibrotic gene expression, proinflammatory cytokines, oxidative stress, and cell death whereas improving the function of mitochondrial electron transport chain. We observed baicalin enhanced NRF2 activation by nuclear translocation and induced its target genes HO-1 and GCLM, thus enhancing antioxidant defense. Interplay of oxidative stress/inflammation and NRF2 were key players for baicalin-mediated protection. Stellate cell activation is crucial for initiation of fibrosis. Baicalin alleviated stellate cell activation and modulated TIMP1, SMA, collagen 1, and fibronectin in vitro. This study indicates that baicalin might be beneficial for reducing inflammation and fibrosis in liver injury models. PMID:28757911

Protective Effect of Flavonoids from Ziziphus jujuba cv. Jinsixiaozao against Acetaminophen-Induced Liver Injury by Inhibiting Oxidative Stress and Inflammation in Mice.

PubMed

Huang, Weizhen; Wang, Yongjie; Jiang, Xiaoyan; Sun, Yueyue; Zhao, Zhongxi; Li, Siying

2017-10-20

This study was aimed to investigate the chemical composition, antioxidant activities and hepatoprotective effect of flavonoids from Ziziphus jujub a cv. Jinsixiaozao (ZJF). The composition of ZJF was analyzed by high performance liquid chromatography (HPLC) and Liquid chromatography-mass spectrometry (LC-MS), and antioxidant properties were investigated by biological assays in vitro. The hepatoprotective activity of ZJF was evaluated in acetaminophen (APAP)-treated BALB/c mice. Results indicate that ZJF displayed significant antioxidant capacity. Pretreatment with ZJF significantly decreased APAP-elevated serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP) and total bilirubin (TB). Activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) were enhanced with ZJF administration, while malondialdehyde (MDA) level and glutathione (GSH) depletion were reduced. Meanwhile, ZJF reversed the suppression of nuclear factor erythroid 2-related factor 2 (Nrf2) nuclear translocation, and up-regulated the protein expression of NAD(P)H: quinone oxidoreductase 1(NQO1) in liver damage mice. Furthermore, ZJF attenuated APAP-induced inflammatory mediator production, such as nitric oxide (NO), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β). Expression of p65 showed that ZJF dampened nuclear factor-κB (NF-κB) activation. The results strongly indicate that the hepatoprotective role of ZJF in APAP-induced hepatotoxicity might result from its induction of antioxidant defense via activation of Nrf2 and reduction of inflammation via inhibition of NF-κB.

Decursin Isolated from Angelica gigas Nakai Rescues PC12 Cells from Amyloid β-Protein-Induced Neurotoxicity through Nrf2-Mediated Upregulation of Heme Oxygenase-1: Potential Roles of MAPK.

PubMed

Li, Li; Du, Ji-Kun; Zou, Li-Yi; Wu, Tie; Lee, Yong-Woo; Kim, Yong-Ho

2013-01-01

Decursin (D), purified from Angelica gigas Nakai, has been proven to exert neuroprotective property. Previous study revealed that D reduced A β 25 ‒ 35-induced cytotoxicity in PC12 cells. Our study explored the underlying mechanisms by which D mediates its therapeutic effects in vitro. Pretreatment of cells with D diminished intracellular generation of ROS in response to A β 25 ‒ 35. Western blot revealed that D significantly increased the expression and activity of HO-1, which was correlated with its protection against A β 25 ‒ 35-induced injury. Addition of ZnPP, an HO-1 competitive inhibitor, significantly attenuated its protective effect in A β 25 ‒ 35-treated cells, indicating the vital role of HO-1 resistance to oxidative injury. Moreover, D induced Nrf2 nuclear translocation, the upstream of HO-1 expression. While investigating the signaling pathways responsible for HO-1 induction, D activated ERK and dephosphorylated p38 in PC12 cells. Addition of U0126, a selective inhibitor of ERK, blocked D-induced Nrf2 activation and HO-1 induction and meanwhile reversed the protection of D against A β 25 ‒ 35-induced cell death. These findings suggest D augments cellular antioxidant defense capacity through both intrinsic free radical scavenging activity and activation of MAPK signal pathways that leads to Nrf2 activation, and subsequently HO-1 induction, thereby protecting the PC12 cells from A β 25 ‒ 35-induced oxidative cytotoxicity.

Decursin Isolated from Angelica gigas Nakai Rescues PC12 Cells from Amyloid β-Protein-Induced Neurotoxicity through Nrf2-Mediated Upregulation of Heme Oxygenase-1: Potential Roles of MAPK

PubMed Central

Li, Li; Du, Ji-kun; Zou, Li-yi; Wu, Tie; Lee, Yong-woo; Kim, Yong-ho

2013-01-01

Decursin (D), purified from Angelica gigas Nakai, has been proven to exert neuroprotective property. Previous study revealed that D reduced Aβ 25‒35-induced cytotoxicity in PC12 cells. Our study explored the underlying mechanisms by which D mediates its therapeutic effects in vitro. Pretreatment of cells with D diminished intracellular generation of ROS in response to Aβ 25‒35. Western blot revealed that D significantly increased the expression and activity of HO-1, which was correlated with its protection against Aβ 25‒35-induced injury. Addition of ZnPP, an HO-1 competitive inhibitor, significantly attenuated its protective effect in Aβ 25‒35-treated cells, indicating the vital role of HO-1 resistance to oxidative injury. Moreover, D induced Nrf2 nuclear translocation, the upstream of HO-1 expression. While investigating the signaling pathways responsible for HO-1 induction, D activated ERK and dephosphorylated p38 in PC12 cells. Addition of U0126, a selective inhibitor of ERK, blocked D-induced Nrf2 activation and HO-1 induction and meanwhile reversed the protection of D against Aβ 25‒35-induced cell death. These findings suggest D augments cellular antioxidant defense capacity through both intrinsic free radical scavenging activity and activation of MAPK signal pathways that leads to Nrf2 activation, and subsequently HO-1 induction, thereby protecting the PC12 cells from Aβ 25‒35-induced oxidative cytotoxicity. PMID:23762139

Pomegranate protects against arsenic-induced p53-dependent ROS-mediated inflammation and apoptosis in liver cells.

PubMed

Choudhury, Sreetama; Ghosh, Sayan; Mukherjee, Sudeshna; Gupta, Payal; Bhattacharya, Saurav; Adhikary, Arghya; Chattopadhyay, Sreya

2016-12-01

Molecular mechanisms involved in arsenic-induced toxicity are complex and elusive. Liver is one of the most favored organs for arsenic toxicity as methylation of arsenic occurs mostly in the liver. In this study, we have selected a range of environmentally relevant doses of arsenic to examine the basis of arsenic toxicity and the role of pomegranate fruit extract (PFE) in combating it. Male Swiss albino mice exposed to different doses of arsenic presented marked hepatic injury as evident from histological and electron microscopic studies. Increased activities of enzymes alanine aminotransferase, aspartate aminotransferase, lactate dehydrogenase and alkaline phosphatase corroborated extensive liver damage. It was further noted that arsenic exposure initiated reactive oxygen species (ROS)-dependent apoptosis in the hepatocytes involving loss of mitochondrial membrane potential. Arsenic significantly increased nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) and nuclear factor-κB (NF-κB), coupled with increase in phosphorylated Iκ-B, possibly as adaptive cellular survival strategies. Arsenic-induced oxidative DNA damage to liver cells culminated in p53 activation and increased expression of p53 targets like miR-34a and Bax. Pomegranate polyphenols are known to possess remarkable antioxidant properties and are capable of protecting normal cells from various stimuli-induced oxidative stress and toxicities. We explored the protective role of PFE in ameliorating arsenic-induced hepatic damage. PFE was shown to reduce ROS generation in hepatocytes, thereby reducing arsenic-induced Nrf2 activation. PFE also inhibited arsenic-induced NF-κB-inflammatory pathway. Data revealed that PFE reversed arsenic-induced hepatotoxicity and apoptosis by modulating the ROS/Nrf2/p53-miR-34a axis. For the first time, we have mapped the possible signaling pathways associated with arsenic-induced hepatotoxicity and its rescue by pomegranate polyphenols. Copyright © 2016 Elsevier Inc. All rights reserved.

Gankyrin has an antioxidative role through the feedback regulation of Nrf2 in hepatocellular carcinoma

PubMed Central

Yang, Chun; Tan, Ye-xiong; Yang, Guang-zhen; Zhang, Jian; Pan, Yu-fei; Liu, Chen; Fu, Jing; Chen, Yao; Ding, Zhi-wen

2016-01-01

Oxidative stress status has a key role in hepatocellular carcinoma (HCC) development and progression. Normally, reactive oxygen species (ROS) levels are tightly controlled by an inducible antioxidant program that responds to cellular stressors. How HCC cells respond to excessive oxidative stress remains elusive. Here, we identified a feedback loop between gankyrin, an oncoprotein overexpressed in human HCC, and Nrf2 maintaining the homeostasis in HCC cells. Mechanistically, gankyrin was found to interact with the Kelch domain of Keap1 and effectively competed with Nrf2 for Keap1 binding. Increased expression of gankyrin in HCC cells blocked the binding between Nrf2 and Keap1, inhibiting the degradation of Nrf2 by proteasome. Interestingly, accumulation and translocation of Nrf2 increased the transcription of gankyrin through binding to the ARE elements in the promoter of gankyrin. The positive feedback regulation involving gankyrin and Nrf2 modulates a series of antioxidant enzymes, thereby lowering intracellular ROS and conferring a steadier intracellular environment, which prevents mitochondrial damage and cell death induced by excessive oxidative stress. Our results indicate that gankyrin is a regulator of cellular redox homeostasis and provide a link between oxidative stress and the development of HCC. PMID:27091842

(-)-7(S)-hydroxymatairesinol protects against tumor necrosis factor-α-mediated inflammation response in endothelial cells by blocking the MAPK/NF-κB and activating Nrf2/HO-1.

PubMed

Yang, Di; Xiao, Chen-Xi; Su, Zheng-Hua; Huang, Meng-Wei; Qin, Ming; Wu, Wei-Jun; Jia, Wan-Wan; Zhu, Yi-Zhun; Hu, Jin-Feng; Liu, Xin-Hua

2017-08-15

Endothelial inflammation is an increasingly prevalent condition in the pathogenesis of many cardiovascular diseases. (-)-7(S)-hydroxymatairesinol (7-HMR), a naturally occurring plant lignan, possesses both antioxidant and anti-cancer properties and therefore would be a good strategy to suppress tumor necrosis factor-α (TNF-α)-mediated inflammation in vascular endothelial cells (VECs). The objective of this study is to evaluate for its anti-inflammatory effect on TNF-α-stimulated VECs and underling mechanisms. The effect of the 7-HMR on suppression of TNF-α-induced inflammation mediators in VECs were determined by qRT-PCR and Western blot. MAPKs and phosphorylation of Akt, HO-1 and NF-κB p65 were examined using Western blot. Nuclear localisation of NF-κB was also examined using Western blot and immunofluorescence. Here we found that 7-HMR could suppress TNF-α-induced inflammatory mediators, such as vascularcelladhesion molecule-1, interleukin-6 and inducible nitric oxide synthase expression both in mRNA and protein levels, and concentration-dependently attenuated reactive oxidase species generation. We further identified that 7-HMR remarkably induced superoxide dismutase and heme oxygenase-1 expression associated with degradation of Kelch-like ECH-associated protein 1 (keap1) and up-regulated nuclear factor erythroid 2-related factor 2 (Nrf2). In addition, 7-HMR time- and concentration-dependently attenuated TNF-α-induced phosphorylation of extracellular signal-regulated kinase 1/2 (ERK) and Akt, but not p38, or c-Jun N-terminal kinase 1/2. Moreover, 7-HMR significantly suppressed TNF-α-mediated nuclear factor-κB (NF-κB) activation by inhibiting phosphorylation and nuclear translocation of NF-κB p65. Our results demonstrated that 7-HMR inhibited TNF-α-stimulated endothelial inflammation, at least in part, through inhibition of NF-κB activation and upregulation of Nrf2-antioxidant response element signaling pathway, suggesting 7-HMR might be used as a promising vascular protective drug. Copyright © 2017. Published by Elsevier GmbH.

Possible involvement of nuclear factor erythroid 2-related factor 2 in the gene expression of Cyp2b10 and Cyp2a5.

PubMed

Ashino, Takashi; Ohkubo-Morita, Haruyo; Yamamoto, Masayuki; Yoshida, Takemi; Numazawa, Satoshi

2014-01-01

Cytochrome P450 gene expression is altered by various chemical compounds. In this study, we used nuclear factor erythroid 2-related factor 2 (Nrf2)-deficient (Nrf2(-⧸-)) mice to investigate the involvement of Nrf2 in Cyp2b10 and Cyp2a5 gene expression. Phorone, an Nrf2 activator, strongly increased Cyp2b10 and Cyp2a5 mRNA as well as Nrf2 target genes, including NAD(P)H-quinone oxidoreductase-1 and heme oxygenase-1, in wild-type mouse livers 8 h after treatment. The phorone-induced mRNA levels in Nrf2(-⧸-) mouse livers were lower than that in wild-type mouse livers. Nrf2(-⧸-) mice showed attenuated Cyp2b10 and Cyp2a5 induction by phenobarbital, a classical Cyp2b inducer. These findings suggest that the Nrf2 pathway is involved in Cyp2b10 and Cyp2a5 gene expression.

Possible involvement of nuclear factor erythroid 2-related factor 2 in the gene expression of Cyp2b10 and Cyp2a5☆

PubMed Central

Ashino, Takashi; Ohkubo-Morita, Haruyo; Yamamoto, Masayuki; Yoshida, Takemi; Numazawa, Satoshi

2014-01-01

Cytochrome P450 gene expression is altered by various chemical compounds. In this study, we used nuclear factor erythroid 2-related factor 2 (Nrf2)–deficient (Nrf2−⧸−) mice to investigate the involvement of Nrf2 in Cyp2b10 and Cyp2a5 gene expression. Phorone, an Nrf2 activator, strongly increased Cyp2b10 and Cyp2a5 mRNA as well as Nrf2 target genes, including NAD(P)H-quinone oxidoreductase-1 and heme oxygenase-1, in wild-type mouse livers 8 h after treatment. The phorone-induced mRNA levels in Nrf2−⧸− mouse livers were lower than that in wild-type mouse livers. Nrf2−⧸− mice showed attenuated Cyp2b10 and Cyp2a5 induction by phenobarbital, a classical Cyp2b inducer. These findings suggest that the Nrf2 pathway is involved in Cyp2b10 and Cyp2a5 gene expression. PMID:24494203

Induction of heme oxygenase-1 by chamomile protects murine macrophages against oxidative stress.

PubMed

Bhaskaran, Natarajan; Shukla, Sanjeev; Kanwal, Rajnee; Srivastava, Janmejai K; Gupta, Sanjay

2012-06-27

Protection of cells from oxidative insult may be possible through direct scavenging of reactive oxygen species, or through stimulation of intracellular antioxidant defense mechanisms by induction of antioxidant gene expression. In this study we investigated the cytoprotective effect of chamomile and elucidated the underlying mechanisms. The cytoprotective effect of chamomile was examined on H(2)O(2)-induced cellular stress in RAW 264.7 murine macrophages. RAW 264.7 murine macrophages treated with chamomile were protected from cell death caused by H(2)O(2). Treatment with 50μM H(2)O(2) for 6h caused significant increase in cellular stress accompanied by cell death in RAW 264.7 macrophages. Pretreatment with chamomile at 10-20μg/mL for 16h followed by H(2)O(2) treatment protected the macrophages against cell death. Chamomile exposure significantly increased the expression of antioxidant enzymes viz. heme oxygenase-1 (HO-1), peroxiredoxin-1 (Prx-1), and thioredoxin-1 (Trx-1) in a dose-dependent manner, compared with their respective controls. Chamomile increased nuclear translocation of Nrf2 with increased phosphorylated Nrf2 levels, and binding to the antioxidant response element in the nucleus. These molecular findings for the first time provide insights into the mechanisms underlying the induction of phase 2 enzymes through the Keap1-Nrf2 signaling pathway by chamomile, and provide evidence that chamomile possesses antioxidant and cytoprotective properties. Copyright © 2012 Elsevier Inc. All rights reserved.

Induction of heme oxygenase-1 by chamomile protects murine macrophages against oxidative stress

PubMed Central

Bhaskaran, Natarajan; Shukla, Sanjeev; Kanwal, Rajnee; Srivastava, Janmejai K; Gupta, Sanjay

2012-01-01

Aims Protection of cells from oxidative insult may be possible through direct scavenging of reactive oxygen species, or through stimulation of intracellular antioxidant defense mechanisms by induction of antioxidant gene expression. In this study we investigated the cytoprotective effect of chamomile and elucidated the underlying mechanisms. Main Methods The cytoprotective effect of chamomile was examined on H2O2-induced cellular stress in RAW 264.7 murine macrophages. Key Findings RAW 264.7 murine macrophages treated with chamomile were protected from cell death caused by H2O2. Treatment with 50 μM H2O2 for 6 h caused significant increase in cellular stress accompanied by cell death in RAW 264.7 macrophages. Pretreatment with chamomile at 10-20 μg/mL for 16 h followed by H2O2 treatment protected the macrophages against cell death. Chamomile exposure significantly increased the expression of antioxidant enzymes viz. heme oxygenase-1 (HO-1), peroxiredoxin-1 (Prx-1), and thioredoxin-1 (Trx-1) in a dose-dependent manner, compared with their respective controls. Chamomile increased nuclear translocation of Nrf2 with increased phosphorylated Nrf2 levels, and binding to the antioxidant response element in the nucleus. Significance These molecular findings for the first time provide insights into the mechanisms underlying the induction of phase 2 enzymes through the Keap1-Nrf2 signaling pathway by chamomile, and provide evidence that chamomile possesses antioxidant and cytoprotective properties. PMID:22683429

Melatonin successfully rescues hippocampal bioenergetics and improves cognitive function following drug intoxication by promoting Nrf2-ARE signaling activity.

PubMed

Chen, Li-You; Renn, Ting-Yi; Liao, Wen-Chieh; Mai, Fu-Der; Ho, Ying-Jui; Hsiao, George; Lee, Ai-Wei; Chang, Hung-Ming

2017-09-01

Prolonged exposure to gamma-hydroxybutyric acid (GHB) would cause drug intoxication in which impaired cognitive function results from enhanced hippocampal oxidative stress may serve as a major symptom in this deficiency. Considering melatonin possesses significant anti-oxidative efficacy, this study aimed to determine whether melatonin would successfully promote the nuclear factor erythroid 2-related factor 2 and antioxidant responsive element (Nrf2-ARE) signaling, depress oxidative stress, and rescue hippocampal bioenergetics and cognitive function following drug intoxication injury. Adolescent rats subjected to 10 days of GHB were received melatonin at doses of either 10 or 100 mg/kg. Time-of-flight secondary ion mass spectrometry, biochemical assay, quantitative histochemistry, [ 14 C]-2-deoxyglucose analysis, together with Morris water maze were employed to detect the molecular signaling, oxidative status, bioenergetic level, as well as the cognitive performances, respectively. Results indicated that in GHB-intoxicated rats, enhanced oxidative stress, increased cholesterol level, and decreased anti-oxidative enzymes activities were detected in hippocampal regions. Intense oxidative stress paralleled well with reduced bioenergetics and poor performance in behavioral testing. However, in rats treated with melatonin following GHB intoxication, all above parameters and cognitive function were gradually returned to nearly normal levels. Melatonin also remarkably promoted the translocation of Nrf2 from cytoplasm to nucleus in a dose-dependent manner, thereby increased the Nrf2-ARE signaling-related downstream anti-oxidative enzymes activities. As melatonin effectively rescues hippocampal bioenergetics through depressing the oxidative stress by promoting Nrf2-ARE molecular machinery, this study thus highlights for the first time that clinical use of melatonin may serve as a therapeutic strategy to improve the cognitive function in unsuspecting victims suffered from GHB intoxication injury. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Natural antioxidants exhibit chemopreventive characteristics through the regulation of CNC b-Zip transcription factors in estrogen-induced breast carcinogenesis.

PubMed

Chatterjee, Anwesha; Ronghe, Amruta; Singh, Bhupendra; Bhat, Nimee K; Chen, Jie; Bhat, Hari K

2014-12-01

The objective of the present study was to characterize the role of resveratrol (Res) and vitamin C (VC) in prevention of estrogen-induced breast cancer through regulation of cap "n"collar (CNC) b-zip transcription factors. Human breast epithelial cell line MCF-10A was treated with 17β-estradiol (E2) and VC or Res with or without E2. mRNA and protein expression levels of CNC b-zip transcription factors nuclear factor erythroid 2-related factor 1 (Nrf1), nuclear factor erythroid 2 related factor 2 (Nrf2), nuclear factor erythroid 2 related factor 3 (Nrf3), and Nrf2-regulated antioxidant enzymes superoxide dismutase 3 (SOD3) and quinone oxidoreductase 1 (NQO1) were quantified. The treatment with E2 suppressed, whereas VC and Res prevented E2-mediated decrease in the expression levels of SOD3, NQO1, Nrf2 mRNA, and protein in MCF-10A cells. The treatment with E2, Res, or VC significantly increased mRNA and protein expression levels of Nrf1. 17β-Estradiol treatment significantly increased but VC or Res decreased Nrf3 mRNA and protein expression levels. Our studies demonstrate that estrogen-induced breast cancer might be prevented through upregulation of antioxidant enzymes via Nrf-dependent pathways. © 2014 Wiley Periodicals, Inc.

Protective effects of nuclear factor erythroid 2-related factor 2 on whole body heat stress-induced oxidative damage in the mouse testis.

PubMed

Li, Yansen; Huang, Yi; Piao, Yuanguo; Nagaoka, Kentaro; Watanabe, Gen; Taya, Kazuyoshi; Li, ChunMei

2013-03-21

Whole body heat stress had detrimental effect on male reproductive function. It's known that the nuclear factor erythroid 2-related factor 2 (Nrf2) activates expression of cytoprotective genes to enable cell adaptation to protect against oxidative stress. However, it's still unclear about the exactly effects of Nrf2 on the testis. Here, we investigate the protective effect of Nrf2 on whole body heat stress-induced oxidative damage in mouse testis. Male mice were exposed to the elevated ambient temperature (42°C) daily for 2 h. During the period of twelve consecutive days, mice were sacrificed on days 1, 2, 4, 8 and 12 immediately following heat exposure. Testes weight, enzymatic antioxidant activities and concentrations of malondialdehyde (MDA) and glutathione (GSH) in the testes were determined and immunohistochemical detection of Nrf2 protein and mRNA expression of Nrf2-regulated genes were analyzed to assess the status of Nrf2-antioxidant system. Heat-exposed mice presented significant increases in rectal, scrotal surface and body surface temperature. The concentrations of cortisol and testosterone in serum fluctuated with the number of exposed days. There were significant decrease in testes weight and relative testes weight on day 12 compared with those on other days, but significant increases in catalase (CAT) activity on day 1 and GSH level on day 4 compared with control group. The activities of total superoxide dismutase (T-SOD) and copper-zinc SOD (CuZn-SOD) increased significantly on days 8 and 12. Moreover, prominent nuclear accumulation of Nrf2 protein was observed in Leydig cells on day 2, accompanying with up-regulated mRNA levels of Nrf2-regulated genes such as Nrf2, heme oxygenase 1 (HO-1), γ-Glutamylcysteine synthetase (GCLC) and NAD (P) H: quinone oxidoreductase 1 (NQO1)) in heat-treated groups. These results suggest that Nrf2 displayed nuclear accumulation and protective activity in the process of heat treated-induced oxidative stress in mouse testes, indicating that Nrf2 might be a potential target for new drugs designed to protect germ cell and Leydig cell from oxidative stress.

Bioactive compounds isolated from apple, tea, and ginger protect against dicarbonyl induced stress in cultured human retinal epithelial cells.

PubMed

Sampath, Chethan; Zhu, Yingdong; Sang, Shengmin; Ahmedna, Mohamed

2016-02-15

Methylglyoxal (MGO) is known to be a major precursor of advanced glycation end products (AGEs) which are linked to diabetes and its related complications. Naturally occurring bioactive compounds could play an important role in countering AGEs thereby minimizing the risk associated with their formation. In this study, eight specific bioactive compounds isolated from apple, tea and ginger were evaluated for their AGEs scavenging activity using Human Retinal Pigment Epithelial (H-RPE) cells treated with MGO. Among the eight specific compounds evaluated, (-)-epigallocatechin 3-gallate (EGCG) from tea, phloretin in apple, and [6]-shogaol and [6]-gingerol from ginger were found to be most effective in preventing MGO-induced cytotoxicity in the epithelial cells. Investigation of possible underlying mechanisms suggests that that these compounds could act by modulating key regulative detoxifying enzymes via modifying nuclear factor-erythroid 2-related factor 2 (Nrf2) function. MGO-induced cytotoxicity led to increased levels of AGEs causing increase in Nε-(Carboxymethyl) lysine (CML) and glutathione (GSH) levels and over expression of receptor for advanced glycation end products (RAGE). Data also showed that translocation of Nrf2 from cytosol to nucleus was inhibited, which decreased the expression of detoxifying enzyme like heme oxygenase-1 (HO-1). The most potent bioactive compounds scavenged dicarbonyl compounds, inhibited AGEs formation and significantly reduced carbonyl stress by Nrf2 related pathway and restoration of HO-1 expression. These findings demonstrated the protective effect of bioactive compounds derived from food sources against MGO-induced carbonyl stress through activation of the Nrf2 related defense pathway, which is of significant importance for therapeutic interventions in complementary treatment/management of diabetes-related complications. Copyright © 2016. Published by Elsevier GmbH.

DNA Protecting Activities of Nymphaea nouchali (Burm. f) Flower Extract Attenuate t-BHP-Induced Oxidative Stress Cell Death through Nrf2-Mediated Induction of Heme Oxygenase-1 Expression by Activating MAP-Kinases

PubMed Central

Ju, Mi-Kyoung

2017-01-01

This study was performed to investigate the antioxidant activities of Nymphaea nouchali flower (NNF) extract and the underlying mechanism using RAW 264.7 cells. The presence of gallic acid, catechin, epicatechin, epigallocatechin, epicatechin gallate, caffeic acid, quercetin, and apigenin in the NNF was confirmed by high-performance liquid chromatography (HPLC). The extract had a very potent capacity to scavenge numerous free radicals. NNF extract was also able to prevent DNA damage and quench cellular reactive oxygen species (ROS) generation induced by tert-Butyl hydroperoxide (t-BHP) with no signs of toxicity. The NNF extract was able to augment the expression of both primary and phase II detoxifying enzyme, resulting in combat the oxidative stress. This is accomplished by phosphorylation of mitogen-activated protein kinase (MAP kinase) (p38 kinase and extracellular signal-regulated kinase (ERK)) followed by enhancing the nuclear translocation of the nuclear factor erythroid 2-related factor 2 (Nrf2). This attenuates cellular ROS generation and confers protection from cell death. Altogether, the results of current study revealed that Nymphaea nouchali flower could be a source of natural phytochemicals that could lead to the development of new therapeutic agents for preventing oxidative stress associated diseases and attenuating disease progression. PMID:28956831

Nrf2-mediated mucoprotective and anti-inflammatory actions of Artemisia extracts led to attenuate stress related mucosal damages

PubMed Central

Park, Jong-Min; Han, Young-Min; Lee, Jin-Seok; Ko, Kwang Hyun; Hong, Sung-Pyo; Kim, Eun-Hee; Hahm, Ki-Baik

2015-01-01

The aim of this study was to compare biological actions between isopropanol and ethanol extracts of Artemisia including antioxidant, anti-inflammatory, and cytoprotective actions. Antioxidant activities were evaluated using 2,2-diphenyl-1-picrylhydrazyl (DPPH) method and confocal microscopy on lipopolysaccharide-induced RGM1 cells, cytoprotection effects evaluated by detecting heme oxygenase-1 (HO-1), Nf-E2 related factor2 (Nrf2) and heat shock protein 70 (HSP70), and anti-inflammatory effects investigated by measuring inflammatory mediators. Water immersion restraint stress was imposed to provoke stress related mucosal damages (SRMD) in rats. Isopropanol extracts of Artemisia showed the higher DPPH radical scavenging activity and lesser LPS-induced reactive oxygen species productions and increased HO-1 expression through increased nuclear translocation of Nrf2 transcription factor compared to ethanol extracts. The increased expression of HSP70 and decreased expression of endothelin-1 were only increased with isopropanol extracts. A concentration-dependent inhibition of LPS-induced COX-2 and iNOS even at a rather lower concentration than ethanol extract was achieved with isopropanol extracts. Cytokine protein array revealed Artemisia extracts significantly attenuated the levels of CXCL-1, CXCL-16, and MCP-1. These orchestrated actions led to significant rescue from SRMD. Conclusively, Artemisia extracts imposed significant antioxidant and anti-inflammatory activity against SRMD and isopropanol extracts were superior to ethanol extracts in these beneficiary actions of Artemisia. PMID:25759519

Ferulic Acid Regulates the Nrf2/Heme Oxygenase-1 System and Counteracts Trimethyltin-Induced Neuronal Damage in the Human Neuroblastoma Cell Line SH-SY5Y

PubMed Central

Catino, Stefania; Paciello, Fabiola; Miceli, Fiorella; Rolesi, Rolando; Troiani, Diana; Calabrese, Vittorio; Santangelo, Rosaria; Mancuso, Cesare

2016-01-01

Over the past years, several lines of evidence have pointed out the efficacy of ferulic acid (FA) in counteracting oxidative stress elicited by β-amyloid or free radical initiators, based on the ability of this natural antioxidant to up-regulate the heme oxygenase-1 (HO-1) and biliverdin reductase (BVR) system. However, scarce results can be found in literature regarding the cytoprotective effects of FA in case of damage caused by neurotoxicants. The aim of this work is to investigate the mechanisms through which FA exerts neuroprotection in SH-SY5Y neuroblastoma cells exposed to the neurotoxin trimethyltin (TMT). FA (1–10 μM for 6 h) dose-dependently increased both basal and TMT (10 μM for 24 h)-induced HO-1 expression in SH-SY5Y cells by fostering the nuclear translocation of the transcriptional activator Nrf2. In particular, the co-treatment of FA (10 μM) with TMT was also responsible for the nuclear translocation of HO-1 in an attempt to further increase cell stress response in SH-SY5Y cells. In addition to HO-1, FA (1–10 μM for 6 h) dose-dependently increased the basal expression of BVR. The antioxidant and neuroprotective features of FA, through the increase of HO activity, were supported by the evidence that FA inhibited TMT (10 μM)-induced lipid peroxidation (evaluated by detecting 4-hydroxy-nonenal) and DNA fragmentation in SH-SY5Y cells and that this antioxidant effect was reversed by the HO inhibitor Zinc-protoporphyrin-IX (5 μM). Among the by-products of the HO/BVR system, carbon monoxide (CORM-2, 50 nM) and bilirubin (BR, 50 nM) significantly inhibited TMT-induced superoxide anion formation in SH-SY5Y cells. All together, these results corroborate the neuroprotective effect of FA through the up-regulation of the HO-1/BVR system, via carbon monoxide and BR formation, and provide the first evidence on the role of HO-1/Nrf2 axis in FA-related enhancement of cell stress response in human neurons. PMID:26779023

Proton pump inhibitors suppress iNOS-dependent DNA damage in Barrett's esophagus by increasing Mn-SOD expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Thanan, Raynoo; Department of Environmental and Molecular Medicine, Mie University Graduate School of Medicine, Tsu, Mie 514-8507; Ma, Ning

2012-05-04

Highlights: Black-Right-Pointing-Pointer Inflammation by Barrett's esophagus (BE) is a risk factor of its adenocarcinoma (BEA). Black-Right-Pointing-Pointer 8-Nitroguanine and 8-oxodG are inflammation-related DNA lesions. Black-Right-Pointing-Pointer DNA lesions and iNOS expression were higher in the order, BEA > BE > normal tissues. Black-Right-Pointing-Pointer Proton pump inhibitors suppress DNA damage by increasing Mn-SOD via Nrf2 activation. Black-Right-Pointing-Pointer DNA lesions can be useful biomarkers to predict risk of BEA in BE patients. -- Abstract: Barrett's esophagus (BE), an inflammatory disease, is a risk factor for Barrett's esophageal adenocarcinoma (BEA). Treatment of BE patients with proton pump inhibitors (PPIs) is expected to reduce the riskmore » of BEA. We performed an immunohistochemical study to examine the formation of nitrative and oxidative DNA lesions, 8-nitroguanine and 8-oxo-7,8-dihydro-2 Prime -deoxygaunosine (8-oxodG), in normal esophageal, BE with pre- and post-treatment by PPIs and BEA tissues. We also observed the expression of an oxidant-generating enzyme (iNOS) and its transcription factor NF-{kappa}B, an antioxidant enzyme (Mn-SOD), its transcription factor (Nrf2) and an Nrf2 inhibitor (Keap1). The immunoreactivity of DNA lesions was significantly higher in the order of BEA > BE > normal tissues. iNOS expression was significantly higher in the order of BEA > BE > normal tissues, while Mn-SOD expression was significantly lower in the order of BEA < BE < normal tissues. Interestingly, Mn-SOD expression and the nuclear localization of Nrf2 were significantly increased, and the formation of DNA lesions was significantly decreased in BE tissues after PPIs treatment for 3-6 months. Keap1 and iNOS expression was not significantly changed by the PPIs treatment in BE tissues. These results indicate that 8-nitroguanine and 8-oxodG play a role in BE-derived BEA. Additionally, PPIs treatment may trigger the activation and nuclear translocation of Nrf2 resulting in the expression of antioxidant genes, leading to DNA damage suppression. These DNA lesions can be useful biomarkers to predict both the risk of BEA and the efficacy of PPIs treatment to prevent BEA in BE patients.« less

Trafficking of the transcription factor Nrf2 to promyelocytic leukemia-nuclear bodies: implications for degradation of NRF2 in the nucleus.

PubMed

Malloy, Melanie Theodore; McIntosh, Deneshia J; Walters, Treniqka S; Flores, Andrea; Goodwin, J Shawn; Arinze, Ifeanyi J

2013-05-17

Ubiquitylation of Nrf2 by the Keap1-Cullin3/RING box1 (Cul3-Rbx1) E3 ubiquitin ligase complex targets Nrf2 for proteasomal degradation in the cytoplasm and is an extensively studied mechanism for regulating the cellular level of Nrf2. Although mechanistic details are lacking, reports abound that Nrf2 can also be degraded in the nucleus. Here, we demonstrate that Nrf2 is a target for sumoylation by both SUMO-1 and SUMO-2. HepG2 cells treated with As2O3, which enhances attachment of SUMO-2/3 to target proteins, increased SUMO-2/3-modification (polysumoylation) of Nrf2. We show that Nrf2 traffics, in part, to promyelocytic leukemia-nuclear bodies (PML-NBs). Cell fractions harboring key components of PML-NBs did not contain biologically active Keap1 but contained modified Nrf2 as well as RING finger protein 4 (RNF4), a poly-SUMO-specific E3 ubiquitin ligase. Overexpression of wild-type RNF4, but not the catalytically inactive mutant, decreased the steady-state levels of Nrf2, measured in the PML-NB-enriched cell fraction. The proteasome inhibitor MG-132 interfered with this decrease, resulting in elevated levels of polysumoylated Nrf2 that was also ubiquitylated. Wild-type RNF4 accelerated the half-life (t½) of Nrf2, measured in PML-NB-enriched cell fractions. These results suggest that RNF4 mediates polyubiquitylation of polysumoylated Nrf2, leading to its subsequent degradation in PML-NBs. Overall, this work identifies Nrf2 as a target for sumoylation and provides a novel mechanism for its degradation in the nucleus, independent of Keap1.

Antiphotoaging Effect of Prunus yeonesis Blossom Extract via Inhibition of MAPK/AP-1 and Regulation of the TGF-βI/Smad and Nrf2/ARE Signaling Pathways.

PubMed

Li, Lu; Hwang, Eunson; Ngo, Hien T T; Lin, Pei; Gao, Wei; Liu, Ying; Yi, Tae-Hoo

2018-02-08

Cherry blossoms have attracted attention as an ingredient with potential for use in skincare products. However, no skin photoaging-related research has been performed with this plant. In this study, cherry blossom extract (CBE) at 1, 10 and 100 μg mL -1 was investigated for its skin antiphotoaging effects in UVB-irradiated normal human dermal fibroblasts (NHDF) cells in vitro. Our results showed that CBE markedly increased type-I procollagen during UVB exposure via two pathways. Firstly, transcription activator protein-1 expression and MAP kinases were downregulated, consequently reducing the production of matrix metalloproteinase (MMP)-1 and MMP-3. Secondly, transforming growth factor TGF-βI secretion was upregulated by Smads. Application of CBE facilitated the nuclear translocation of Nrf2 against reactive oxygen species (ROS)-induced damage, which is essential for the coordinated induction of cytoprotective enzymes. Together, our findings suggest that CBE may be a promising ingredient for skin aging therapy and provide a novel approach for alleviating cutaneous aging. © 2018 The American Society of Photobiology.

LAS0811: from combinatorial chemistry to activation of antioxidant response element.

PubMed

Zhu, Ming; Baek, Hyounggee; Liu, Ruiwu; Song, Aimin; Lam, Kit; Lau, Derick

2009-01-01

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.

LAS0811: From Combinatorial Chemistry to Activation of Antioxidant Response Element

PubMed Central

Zhu, Ming; Baek, Hyounggee; Liu, Ruiwu; Song, Aimin; Lam, Kit; Lau, Derick

2009-01-01

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. PMID:19794825

Induction of Lung GSH and Glutamate Cysteine Ligase by 1,4-phenylenebis(methylene)selenocyanate and its Glutathione Conjugate: Role of Nuclear factor-erythroid 2-Related Factor 2

PubMed Central

Emmert, Sans W.; El-Bayoumy, Karam; Das, Arunangshu; Sun, Yuan-Wan; Amin, Shantu; Desai, Dhimant; Aliaga, Cesar; Richie, John P.

2012-01-01

The synthetic organoselenium agent 1,4- phenylenebis(methylene)selenocyanate (p-XSC) and its glutathione (GSH) conjugate (p-XSeSG), are potent chemopreventive agents in several preclinical models. p-XSC is also an effective inducer of GSH in mouse lung. Our objectives were to test the hypothesis that GSH induction by p-XSC occurs through upregulation of the rate-limiting GSH biosynthetic enzyme glutamate cysteine ligase (GCL), through activation of antioxidant response elements (ARE) in GCL genes via activation of nuclear factor-erythroid 2-related factor 2 (Nrf2). p-XSC feeding (10 ppm Se) increased GSH (230%) and upregulated the catalytic subunit of GCL (GCLc) (55%), extracellular related kinase (ERK) (220%) and nuclear Nrf2 (610%) in lung but not liver after 14 days in the rat (P<0.05). Similarly, p-XSeSG feeding (10 ppm) induced lung GCLc (88%) and GSH (200%) (P<0.05), while the naturally-occurring selenomethionine had no effect. Both p-XSC and p-XSeSG activated a luciferase reporter in HepG2 ARE reporter cells up to 3-fold for p-XSC and ≥5-fold for p-XSeSG. Luciferase activation by p-XSeSG was associated with enhanced levels of GSH, GCLc and nuclear Nrf2, which were significantly reduced by co-incubation with short interfering RNA targeting Nrf2 (siNrf2). The dependence of GCL induction on Nrf2 was confirmed in Nrf2 deficient mouse embryonic fibroblasts (MEF) where p-XSeSG induced GCL subunits in wildtype, but not Nrf2 deficient cells (p<0.05). These results indicate that p-XSC may act through the Nrf2 pathway in vivo, and that p-XSeSG is the putative metabolite responsible for such activation, thus offering p-XSeSG as a less toxic, yet highly efficacious inducer of GSH. PMID:22542796

Role of nuclear factor-κB and heme oxygenase-1 in the mechanism of action of an anti-inflammatory chalcone derivative in RAW 264.7 cells

PubMed Central

Alcaraz, María José; Vicente, Ana María; Araico, Amparo; Dominguez, José N; Terencio, María Carmen; Ferrándiz, María Luisa

2004-01-01

The synthetic chalcone 3′,4′,5′,3,4,5-hexamethoxy-chalcone (CH) is an anti-inflammatory compound able to reduce nitric oxide (NO) production by inhibition of inducible NO synthase protein synthesis. In this work, we have studied the mechanisms of action of this compound. CH (10–30 μM) prevents the overproduction of NO in RAW 264.7 macrophages stimulated with lipopolysaccharide (1 μg ml−1) due to the inhibition of nuclear factor κB (NF-κB) activation. We have shown that treatment of cells with CH results in diminished degradation of the NF-κB–IκB complex leading to inhibition of NF-κB translocation into the nucleus, DNA binding and transcriptional activity. We also demonstrate the ability of this compound to activate NfE2-related factor (Nrf2) and induce heme oxygenase-1 (HO-1). Our results indicate that CH determines a rapid but nontoxic increase of intracellular oxidative species, which could be responsible for Nrf2 activation and HO-1 induction by this chalcone derivative.  This novel anti-inflammatory agent simultaneously induces a cytoprotective response (HO-1) and downregulates an inflammatory pathway (NF-κB) with a mechanism of action different from antioxidant chalcones. PMID:15249426

Competition of nuclear factor-erythroid 2 factors related transcription factor isoforms, Nrf1 and Nrf2, in antioxidant enzyme induction☆

PubMed Central

Chepelev, Nikolai L.; Zhang, Hongqiao; Liu, Honglei; McBride, Skye; Seal, Andrew J.; Morgan, Todd E.; Finch, Caleb E.; Willmore, William G.; Davies, Kelvin J.A.; Forman, Henry Jay

2013-01-01

Although the Nrf2 (nuclear factor-erythroid 2 p45 subunit-related factor 2) regulated expression of multiple antioxidant and cytoprotective genes through the electrophile responsive element (EpRE) is well established, interaction of Nrf2/EpRE with Nrf1, a closely-related transcription factor, is less well understood. Due to either proteolysis or alternative translation, Nrf1 has been found as proteins of varying size, p120, p95, and p65, which have been described as either activators of EpRE or competitive inhibitors of Nrf2. We investigated the effect of Nrf1 on EpRE-regulated gene expression using the catalytic and modifier subunits of glutamate cysteine ligase (GCLC and GCLM) as models and explored the potential role of Nrf1 in altering their expression in aging and upon chronic exposure to airborne nano-sized particulate matter (nPM). Nrf1 knockout resulted in the increased expression of GCLC and GCLM in human bronchial epithelial (HBE1) cells. Overexpression Nrf2 in combination with either p120 or p65 diminished or failed to further increase the GCLC- and GLCM-EpRE luciferase activity. All known forms of Nrf1 protein, remained unchanged in the lungs of mice with age or in response to nPM. Our study shows that Nrf1 could inhibit EpRE activity in vitro, whereas the precise role of Nrf1 in vivo requires further investigations. We conclude that Nrf1 may not be directly responsible for the loss of Nrf2-dependent inducibility of antioxidant and cytoprotective genes observed in aged animals. PMID:24024152

Mangiferin exerts hepatoprotective activity against D-galactosamine induced acute toxicity and oxidative/nitrosative stress via Nrf2–NFκB pathways

DOE Office of Scientific and Technical Information (OSTI.GOV)

Das, Joydeep; Ghosh, Jyotirmoy; Roy, Anandita

Mangiferin, a xanthone glucoside, is well known to exhibit antioxidant, antiviral, antitumor, anti-inflammatory and gene-regulatory effects. In the present study, we isolated mangiferin from the bark of Mangifera indica and assessed its beneficial role in galactosamine (GAL) induced hepatic pathophysiology. GAL (400 mg/kg body weight) exposed hepatotoxic rats showed elevation in the activities of serum ALP, ALT, levels of triglycerides, total cholesterol, lipid-peroxidation and reduction in the levels of serum total proteins, albumin and cellular GSH. Besides, GAL exposure (5 mM) in hepatocytes induced apoptosis and necrosis, increased ROS and NO production. Signal transduction studies showed that GAL exposure significantlymore » increased the nuclear translocation of NFκB and elevated iNOS protein expression. The same exposure also elevated TNF-α, IFN-γ, IL-1β, IL-6, IL-12, IL-18 and decreased IL-10 mRNA expressions. Furthermore, GAL also decreased the protein expression of Nrf2, NADPH:quinine oxidoreductase-1, heme oxygenase-1 and GSTα. However, mangiferin administration in GAL intoxicated rats or coincubation of hepatocytes with mangiferin significantly altered all these GAL-induced adverse effects. In conclusion, the hepatoprotective role of mangiferin was due to induction of antioxidant defense via the Nrf2 pathway and reduction of inflammation via NFκB inhibition. Highlights: ►Galactosamine induces hepatocytes death via oxidative and nitrosative stress. ►Mangiferin exerts hepatoprotective effect/antioxidant defense via Nrf2 pathway. ►Mangiferin exerts anti-inflammatory responses by inhibiting NF-κB. ►Mangiferin suppresses galactosamine-induced repression of IL-10 mRNA.« less

Nuclear respiratory factor 2 regulates the expression of the same NMDA receptor subunit genes as NRF-1: both factors act by a concurrent and parallel mechanism to couple energy metabolism and synaptic transmission.

PubMed

Priya, Anusha; Johar, Kaid; Wong-Riley, Margaret T T

2013-01-01

Neuronal activity and energy metabolism are tightly coupled processes. Previously, we found that nuclear respiratory factor 1 (NRF-1) transcriptionally co-regulates energy metabolism and neuronal activity by regulating all 13 subunits of the critical energy generating enzyme, cytochrome c oxidase (COX), as well as N-methyl-d-aspartate (NMDA) receptor subunits 1 and 2B, GluN1 (Grin1) and GluN2B (Grin2b). We also found that another transcription factor, nuclear respiratory factor 2 (NRF-2 or GA-binding protein) regulates all subunits of COX as well. The goal of the present study was to test our hypothesis that NRF-2 also regulates specific subunits of NMDA receptors, and that it functions with NRF-1 via one of three mechanisms: complementary, concurrent and parallel, or a combination of complementary and concurrent/parallel. By means of multiple approaches, including in silico analysis, electrophoretic mobility shift and supershift assays, in vivo chromatin immunoprecipitation of mouse neuroblastoma cells and rat visual cortical tissue, promoter mutations, real-time quantitative PCR, and western blot analysis, NRF-2 was found to functionally regulate Grin1 and Grin2b genes, but not any other NMDA subunit genes. Grin1 and Grin2b transcripts were up-regulated by depolarizing KCl, but silencing of NRF-2 prevented this up-regulation. On the other hand, over-expression of NRF-2 rescued the down-regulation of these subunits by the impulse blocker TTX. NRF-2 binding sites on Grin1 and Grin2b are conserved among species. Our data indicate that NRF-2 and NRF-1 operate in a concurrent and parallel manner in mediating the tight coupling between energy metabolism and neuronal activity at the molecular level. Copyright © 2012 Elsevier B.V. All rights reserved.

Synergy between sulforaphane and selenium in the up-regulation of thioredoxin reductase and protection against hydrogen peroxide-induced cell death in human hepatocytes.

PubMed

Li, Dan; Wang, Wei; Shan, Yujuan; Barrera, Lawrence N; Howie, Alexander F; Beckett, Geoffrey J; Wu, Kun; Bao, Yongping

2012-07-15

Dietary isothiocyanates and selenium are chemopreventive agents and potent inducers of antioxidant enzymes. It has been previously shown that sulforaphane and selenium have a synergistic effect on the upregulation of thioredoxin reductase-1 (TrxR-1) in human hepatoma HepG2 cells. In this paper, further evidence is presented to show that sulforaphane and selenium synergistically induce TrxR-1 expression in immortalised human hepatocytes. Sulforaphane was found to be more toxic toward hepatocytes than HepG2 cells with IC50=25.1 and 56.4 μM, respectively. Sulforaphane can protect against hydrogen peroxide-induced cell death and this protection was enhanced by co-treatment with selenium. Using siRNA to knock down TrxR-1 or Nrf2, sulforaphane (5 μM)-protected cell viability was reduced from 73% to 46% and 34%, respectively, suggesting that TrxR-1 is an important enzyme in protection against hydrogen peroxide-induced cell death. Sulforaphane-induced TrxR-1 expression was positively associated with significant levels of Nrf2 translocation into the nucleus, but co-treatment with selenium showed no significant increase in Nrf2 translocation. Moreover, MAPK (ERK, JNK and p38) and PI3K/Akt signalling pathways were found to play no significant role in sulforaphane-induced Nrf2 translocation into the nucleus. However, blocking ERK and JNK signalling pathways decreased sulforaphane-induced TrxR-1 mRNA by about 20%; whereas blocking p38 and PI3K/AKT increased TrxR-1 transcription. In summary, a combination of sulforaphane and selenium resulted in a synergistic upregulation of TrxR-1 that contributed to the enhanced protection against free radical-mediated oxidative damage in human hepatocytes. Copyright © 2012 Elsevier Ltd. All rights reserved.

Mining a human transcriptome database for Nrf2 modulators

EPA Science Inventory

Nuclear factor erythroid-2 related factor 2 (Nrf2) is a key transcription factor important in the protection against oxidative stress. We developed computational procedures to enable the identification of chemical, genetic and environmental modulators of Nrf2 in a large database ...

Protective effect of coconut water concentrate and its active component shikimic acid against hydroperoxide mediated oxidative stress through suppression of NF-κB and activation of Nrf2 pathway.

PubMed

Manna, Krishnendu; Khan, Amitava; Kr Das, Dipesh; Bandhu Kesh, Swaraj; Das, Ujjal; Ghosh, Sayan; Sharma Dey, Rakhi; Das Saha, Krishna; Chakraborty, Anindita; Chattopadhyay, Sreya; Dey, Sanjit; Chattopadhyay, Debprasad

2014-08-08

Conventionally coconut water has been used as an 'excellent hydrating' drink that maintain the electrolyte balance and help in treating diverse ailments related to oxidative stress including liver function. The present study was aimed to elucidate whether and how the coconut water concentrate (CWC) and its major active phytoconstituent shikimic acid (SA) can effectively protect murine hepatocytes from the deleterious effect of hydroperoxide-mediated oxidative stress. Bioactivity guided fractionation of CWC resulted in the isolation of a couple of known compounds. Freshly isolated murine hepatocytes were exposed to hydrogen peroxide (H2O2) (1 and 3mM) in the presence or absence of CWC (200 and 400 μg/ml) and SA (40 μM) for the determination of antioxidative, DNA protective, cellular ROS level by modern methods, including immunoblot and flowcytometry to find out the possible mechanism of action. Pre-treatment of hepatocyte with CWC and SA showed significant prevention of H2O2-induced intracellular ROS generation, nuclear DNA damage along with the formation of hepatic TBARS and cellular nitrite. Further, the H2O2 induced cell death was arrested in the presence of CWC through the inhibition of CDC42 mediated SAPK/JNK pathways and activation of other molecules of apoptotic pathways, including Bax and caspase3. Moreover, CWC and SA help in maintaining the GSH level and endogenous antioxidants like Mn-SOD, to support intracellular defense mechanisms, probably through the transcriptional activation of Nrf2; and inhibition of nuclear translocation of NF-κB. CWC and its active components SA reversed the H2O2 induced oxidative damage in hepatocytes, probably through the inhibition of NF-κB, with the activation of PI3K/Akt/Nrf2 pathway and reduction of apoptosis by interfering the SAPK/JNK/Bax pathway. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

An exploration of the antioxidant effects of garlic saponins in mouse-derived C2C12 myoblasts.

PubMed

Kang, Ji Sook; Kim, Sung Ok; Kim, Gi-Young; Hwang, Hye Jin; Kim, Byung Woo; Chang, Young-Chae; Kim, Wun-Jae; Kim, Cheol Min; Yoo, Young Hyun; Choi, Yung Hyun

2016-01-01

In this study, we aimed to confirm the protective effects of garlic saponins against oxidative stress-induced cellular damage and to further elucidate the underlying mechanisms in mouse-derived C2C12 myoblasts. Relative cell viability was determined by 3-(4.5-dimethylthiazol-2-yl)-2.5 diphenyltetrazolium bromide assay. Comet assay was used to measure DNA damage and oxidative stress was determined using 2',7'-dichlorofluorescein diacetate to measure intracellular reactive oxygen species (ROS) generation. Western blot analysis and small interfering RNA (siRNA)-based knockdown were used in order to investigate the possible molecular mechanisms. Our results revealed that garlic saponins prevented hydrogen peroxide (H2O2)-induced growth inhibition and exhibited scavenging activity against intracellular ROS. We also observed that garlic saponins prevented H2O2-induced comet tail formation and decreased the phosphorylation levels of γH2AX expression, suggesting that they can prevent H2O2-induced DNA damage. In addition, garlic saponins increased the levels of heme oxygenase-1 (HO-1), a potent antioxidant enzyme associated with the induction and phosphorylation of nuclear factor erythroid 2-related factor 2 (Nrf2) and the translocation of Nrf2 from the cytosol into the nucleus. However, the protective effects of garlic saponins on H2O2-induced ROS generation and growth inhibition were significantly reduced by zinc protoporphyrin Ⅸ, an HO-1 competitive inhibitor. In addition, the potential of garlic saponins to mediate HO-1 induction and protect against H2O2‑mediated growth inhibition was adversely affected by transient transfection with Nrf2-specific siRNA. Garlic saponins activated extracellular signal‑regulated kinase (ERK) signaling, whereas a specific ERK inhibitor was able to inhibit HO-1 upregulation, as well as Nrf2 induction and phosphorylation. Taken together, the findings of our study suggest that garlic saponins activate the Nrf2/HO-1 pathway by enabling ERK to contribute to the induction of phase Ⅱ antioxidant and detoxifying enzymes, including HO-1 in C2C12 cells.

Protective effects of nuclear factor erythroid 2-related factor 2 on whole body heat stress-induced oxidative damage in the mouse testis

PubMed Central

2013-01-01

Background Whole body heat stress had detrimental effect on male reproductive function. It's known that the nuclear factor erythroid 2-related factor 2 (Nrf2) activates expression of cytoprotective genes to enable cell adaptation to protect against oxidative stress. However, it’s still unclear about the exactly effects of Nrf2 on the testis. Here, we investigate the protective effect of Nrf2 on whole body heat stress-induced oxidative damage in mouse testis. Methods Male mice were exposed to the elevated ambient temperature (42°C) daily for 2 h. During the period of twelve consecutive days, mice were sacrificed on days 1, 2, 4, 8 and 12 immediately following heat exposure. Testes weight, enzymatic antioxidant activities and concentrations of malondialdehyde (MDA) and glutathione (GSH) in the testes were determined and immunohistochemical detection of Nrf2 protein and mRNA expression of Nrf2-regulated genes were analyzed to assess the status of Nrf2-antioxidant system. Results Heat-exposed mice presented significant increases in rectal, scrotal surface and body surface temperature. The concentrations of cortisol and testosterone in serum fluctuated with the number of exposed days. There were significant decrease in testes weight and relative testes weight on day 12 compared with those on other days, but significant increases in catalase (CAT) activity on day 1 and GSH level on day 4 compared with control group. The activities of total superoxide dismutase (T-SOD) and copper-zinc SOD (CuZn-SOD) increased significantly on days 8 and 12. Moreover, prominent nuclear accumulation of Nrf2 protein was observed in Leydig cells on day 2, accompanying with up-regulated mRNA levels of Nrf2-regulated genes such as Nrf2, heme oxygenase 1 (HO-1), γ-Glutamylcysteine synthetase (GCLC) and NAD (P) H: quinone oxidoreductase 1 (NQO1)) in heat-treated groups. Conclusions These results suggest that Nrf2 displayed nuclear accumulation and protective activity in the process of heat treated-induced oxidative stress in mouse testes, indicating that Nrf2 might be a potential target for new drugs designed to protect germ cell and Leydig cell from oxidative stress. PMID:23514035

The C-terminal domain of Nrf1 negatively regulates the full-length CNC-bZIP factor and its shorter isoform LCR-F1/Nrf1β; both are also inhibited by the small dominant-negative Nrf1γ/δ isoforms that down-regulate ARE-battery gene expression.

PubMed

Zhang, Yiguo; Qiu, Lu; Li, Shaojun; Xiang, Yuancai; Chen, Jiayu; Ren, Yonggang

2014-01-01

The C-terminal domain (CTD, aa 686-741) of nuclear factor-erythroid 2 p45-related factor 1 (Nrf1) shares 53% amino acid sequence identity with the equivalent Neh3 domain of Nrf2, a homologous transcription factor. The Neh3 positively regulates Nrf2, but whether the Neh3-like (Neh3L) CTD of Nrf1 has a similar role in regulating Nrf1-target gene expression is unknown. Herein, we report that CTD negatively regulates the full-length Nrf1 (i.e. 120-kDa glycoprotein and 95-kDa deglycoprotein) and its shorter isoform LCR-F1/Nrf1β (55-kDa). Attachment of its CTD-adjoining 112-aa to the C-terminus of Nrf2 yields the chimaeric Nrf2-C112Nrf1 factor with a markedly decreased activity. Live-cell imaging of GFP-CTD reveals that the extra-nuclear portion of the fusion protein is allowed to associate with the endoplasmic reticulum (ER) membrane through the amphipathic Neh3L region of Nrf1 and its basic c-tail. Thus removal of either the entire CTD or the essential Neh3L portion within CTD from Nrf1, LCR-F1/Nrf1β and Nrf2-C112Nrf1, results in an increase in their transcriptional ability to regulate antioxidant response element (ARE)-driven reporter genes. Further examinations unravel that two smaller isoforms, 36-kDa Nrf1γ and 25-kDa Nrf1δ, act as dominant-negative inhibitors to compete against Nrf1, LCR-F1/Nrf1β and Nrf2. Relative to Nrf1, LCR-F1/Nrf1β is a weak activator, that is positively regulated by its Asn/Ser/Thr-rich (NST) domain and acidic domain 2 (AD2). Like AD1 of Nrf1, both AD2 and NST domain of LCR-F1/Nrf1β fused within two different chimaeric contexts to yield Gal4D:Nrf1β607 and Nrf1β:C270Nrf2, positively regulate their transactivation activity of cognate Gal4- and Nrf2-target reporter genes. More importantly, differential expression of endogenous ARE-battery genes is attributable to up-regulation by Nrf1 and LCR-F1/Nrf1β and down-regulation by Nrf1γ and Nrf1δ.

[Induction of uridine 5'-diphosphate-glucuronosyltransferase gene expression by sulforaphane and its mechanism: experimental study in human colon cancel cells].

PubMed

Wang, Min; Li, Yan-Qing; Zhong, Ning; Chen, Jian; Xu, Xiao-Qun; Yuan, Meng-Biao

2005-03-30

To study the induction of expression of uridine 5'-diphosphate (UDP)-glucuronosyltransferase (UGT) 1A in colon cancer cells by sulforaphane (SFN) and its possible mechanism. Human colon cancer cells of the line Caco-2 were cultured and added with SFN of different terminal concentrations, all below the concentration of IC(50). RT-PCR was used to examine the expression of UGT1A mRNA induced by SFN. Western blotting was used to detect the expression of UGT1A protein. The glucuronidation rate of N-hydroxy-PhIP was measured by high performance liquid chromatography (HPLC). The nuclear localization of transcription factor Nrf2 was observed by confocal laser microscopy. (1) Expression of UGT1A mRNA was observed in the Cac0-2 cells induced by SFN of the concentrations of 10 micromol/L approximately 35 micromol/L in a dose-independent manner (P < 0.05). Sulforaphane of the concentration of 25 micromol/L induced the UGT1A mRNA expression time-dependently. The levels of UGT1A1, UGT1A8, and UGT1A10 mRNA expression were significantly increased in the cells treated with 25 micromol/L sulforaphane compared to that in the controls (P = 0.006, P = 0.017, and P = 0.008 respectively). (2) The UGT1A protein band intensity increased significantly in the Coco-2 cells treated with sulforaphane of the concentrations 10 micromol/L approximately 30 micromol/L for 24 h in comparison with the control cells. (3) When the microsomes from the untreated Caco-2 cells were incubated with N-hydroxy-PhIP there was a minor HPLC peak at the expected retention time for N-hydroxy-PhIP-N2-glucuronide. This peak was dramatically increased in the sulforaphane-treated cells, suggesting higher activities of glucuronidation of N-hydroxy-PhIP. (4) Cytoplasmic labeling of NF-E2-related factor 2 (Nrf2), a transcription factor, with no nuclear staining was observed in the non-stimulated cells, whereas an intense nuclear labeling was observed in the sulforaphane-treated cells, indicating the induction of nuclear translocation of Nrf2 by sulforaphane. (1) Low dose sulforaphane induces the expression of UGT1A, UGT1A1, UGT1A A8, and UGT1A A10 mRNA significantly. These changes are accompanied by an increase in UGT1A1 protein and increase in heterocyclic aromatic amine glucuronidation. (2) The induction of the phase II enzyme activity by SFN occurs at the transcriptional level and is regulated by Nrf2.

Nuclear factor erythroid 2-related factor 2 deletion impairs glucose tolerance and exacerbates hyperglycemia in type 1 diabetic mice.

PubMed

Aleksunes, Lauren M; Reisman, Scott A; Yeager, Ronnie L; Goedken, Michael J; Klaassen, Curtis D

2010-04-01

The transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) induces a battery of cytoprotective genes after oxidative stress. Nrf2 aids in liver regeneration by altering insulin signaling; however, whether Nrf2 participates in hepatic glucose homeostasis is unknown. Compared with wild-type mice, mice lacking Nrf2 (Nrf2-null) have lower basal serum insulin and prolonged hyperglycemia in response to an intraperitoneal glucose challenge. In the present study, blood glucose, serum insulin, urine flow rate, and hepatic expression of glucose-related genes were quantified in male diabetic wild-type and Nrf2-null mice. Type 1 diabetes was induced with a single intraperitoneal dose (200 mg/kg) of streptozotocin (STZ). Histopathology and serum insulin levels confirmed depleted pancreatic beta-cells in STZ-treated mice of both genotypes. Five days after STZ, Nrf2-null mice had higher blood glucose levels than wild-type mice. Nine days after STZ, polyuria occurred in both genotypes with more urine output from Nrf2-null mice (11-fold) than wild-type mice (7-fold). Moreover, STZ-treated Nrf2-null mice had higher levels of serum beta-hydroxybutyrate, triglycerides, and fatty acids 10 days after STZ compared with wild-type mice. STZ reduced hepatic glycogen in both genotypes, with less observed in Nrf2-null mice. Increased urine output and blood glucose in STZ-treated Nrf2-null mice corresponded with enhanced gluconeogenesis (glucose-6-phosphatase and phosphoenolpyruvate carboxykinase)- and reduced glycolysis (pyruvate kinase)-related mRNA expression in their livers. Furthermore, the Nrf2 activator oltipraz lowered blood glucose in wild-type but not Nrf2-null mice administered STZ. Collectively, these data indicate that the absence of Nrf2 worsens hyperglycemia in type I diabetic mice and Nrf2 may represent a therapeutic target for reducing circulating glucose levels.

Glycogen synthase kinase 3 regulates expression of nuclear factor-erythroid-2 related transcription factor-1 (Nrf1) and inhibits pro-survival function of Nrf1

PubMed Central

Biswas, Madhurima; Kwong, Erick K.; Park, Eujean; Nagra, Parminder; Chan, Jefferson Y.

2013-01-01

Nuclear factor E2-related factor-1 (Nrf1) is a basic leucine zipper transcription factor that is known to regulate antioxidant and cytoprotective gene expression. It was recently shown that Nrf1 is regulated by SCF-Fbw7 ubiquitin ligase. However our knowledge of upstream signals that targets Nrf1 for degradation by the UPS is not known. We report here that Nrf1 expression is negatively regulated by glycogen synthase kinase 3 (GSK3) in Fbw7-dependent manner. We show that GSK3 interacts with Nrf1 and phosphorylates the Cdc4 phosphodegron domain (CPD) in Nrf1. Mutation of serine residue in the CPD of Nrf1 to alanine (S350A), blocks Nrf1 from phosphorylation by GSK3, and stabilizes Nrf1. Knockdown of Nrf1 and expression of a constitutively active form of GSK3 results in increased apoptosis in neuronal cells in response to ER stress, while expression of the GSK3 phosphorylation resistant S350A–Nrfl attenuates apoptotic cell death. Together these data suggest that GSK3 regulates Nrf1 expression and cell survival function in response to stress activation. PMID:23623971

Anti-inflammatory and heme oxygenase-1 inducing activities of lanostane triterpenes isolated from mushroom Ganoderma lucidum in RAW264.7 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Choi, Solip; Nguyen, Van Thu; Tae, Nara

Ganoderma lucidum is a popular medicinal mushroom used in traditional medicine for preventing or treating a variety of diseases. In the present study, we investigated the anti-inflammatory and heme oxygenase (HO)-1 inducing effects of 12 lanostane triterpenes from G. lucidum in RAW264.7 cells. Of these, seven triterpenes, butyl lucidenateE{sub 2}, butyl lucidenateD{sub 2} (GT-2), butyl lucidenate P, butyl lucidenateQ, Ganoderiol F, methyl ganodenate J and butyl lucidenate N induced HO-1 expression and suppressed lipopolysaccharide (LPS)-induced nitric oxide (NO) production. Inhibiting HO-1 activity abrogated the inhibitory effects of these triterpenes on the production of NO in LPS-stimulated RAW264.7 cells, suggesting themore » involvement of HO-1 in the anti-inflammatory effects of these triterpenes. We further studied the anti-inflammatory and HO-1 inducing effects of GT-2. Mitogen-activated protein kinase inhibitors or N-acetylcysteine, an antioxidant, did not suppress GT-2-mediated HO-1 induction; however, LY294002, a phosphoinositide 3-kinase (PI3K) inhibitor, blocked GT-2-induced HO-1 mRNA and protein expression. GT-2 increased nuclear translocation of nuclear factor-E2-related factor 2 (Nrf2) and knockdown of Nrf2 by small interfering RNA blocked GT-2-mediated HO-1 induction, suggesting that GT-2 induced HO-1 expression via the PI3K/AKT-Nrf2 pathway. Consistent with the notion that HO-1 has anti-inflammatory properties, GT-2 inhibited the production of tumor necrosis factor-α and interleukin-6, as well as inducible nitric oxide synthase and cyclooxygenase-2 expression. These findings suggest that HO-1 inducing activities of these lanostane triterpenes may be important in the understanding of a novel mechanism for the anti-inflammatory activity of G. lucidum. - Highlights: • The anti-inflammatory effects of selected triterpenes from Ganoderma lucidum are demonstrated. • Heme oxygenase-1 induction is attributable to the anti-inflammatory properties of these triterpenes. • The triterpenes induce heme oxygenase-1 expression via the AKT-Nrf2 pathway. • The mechanism explains the anti-inflammatory effect of triterpenes from G. lucidum.« less

Activation of the nuclear factor E2-related factor 2/anitioxidant response element alleviates the nitroglycerin-induced hyperalgesia in rats.

PubMed

Di, Wei; Shi, Xiaolei; Lv, Hua; Liu, Jun; Zhang, Hong; Li, Zhiwei; Fang, Yannan

2016-12-01

Antioxidants have been proven to weaken hyperalgesia in neuropathic pain. Endogenous antioxidant defense system may have a role in the prevention of hyperalgesia in migraine. In this study, we aimed to evaluate the role of nuclear factor E2-related factor 2/antioxidant response element (Nrf2/ARE) pathway in regulating the activation of the trigeminovascular system (TGVS) and hypersensitivity in nitroglycerin (NTG)-induced hyperalgesia rats. The expression levels of Nrf2, HO, HO1, and NQO1 in the trigeminal nucleus caudalis (TNC) were detected by western blot. Immunofluorescence was used to demonstrate the cell-specific localization of Nrf2 in TNC. Sulforaphane, a Nrf2 activator, was administered to NTG-induced rats. Then, the number of c-Fos- and nNOS-immunoreactive neurons in TNC was evaluated using immunofluorescence, and c-Fos and nNOS protein levels were quantified using western blot. Von Frey hair testing was used to evaluate the tactile thresholds of rats at different time points in different groups. Total cellular and nuclear levels of the proteins Nrf2, HO1, and NQO1 were elevated in TNC after NTG injection, and Nrf2 was found to be located in the nucleus and cytoplasm of the neurons. Sulforaphane pretreatment significantly increased the nuclear Nrf2, HO1, and NQO1 levels in TNC. In addition, sulforaphane exposure effectively inhibited the expression of nNOS and c-Fos, reduced the number of nNOS and c-Fos immunoreactive neurons in TNC, and attenuated the tactile thresholds induced by NTG injection. Oxidative stress was involved in nitroglycerin-induced hyperalgesia. Activation of the Nrf2/ARE pathway inhibited the activation of TGVS and prevented the induction of hyperalgesia. Sulforaphane might therefore be an effective agent for hyperalgesia. Further studies are needed to discover the underlying mechanisms of the process.

The NRF2 Activation and Antioxidative Response Are Not Impaired Overall during Hyperoxia-Induced Lung Epithelial Cell Death

PubMed Central

Potteti, Haranatha R.; Reddy, Narsa M.; Hei, Tom K.; Kalvakolanu, Dhananjaya V.; Reddy, Sekhar P.

2013-01-01

Lung epithelial and endothelial cell death caused by pro-oxidant insults is a cardinal feature of acute lung injury/acute respiratory distress syndrome (ALI/ARDS) patients. The NF-E2-related factor 2 (NRF2) activation in response to oxidant exposure is crucial to the induction of several antioxidative and cytoprotective enzymes that mitigate cellular stress. Since prolonged exposure to hyperoxia causes cell death, we hypothesized that chronic hyperoxia impairs NRF2 activation, resulting in cell death. To test this hypothesis, we exposed nonmalignant small airway epithelial cells (AECs) to acute (1–12 h) and chronic (36–48 h) hyperoxia and evaluated cell death, NRF2 nuclear accumulation and target gene expression, and NRF2 recruitment to the endogenous HMOX1 and NQO1 promoters. As expected, hyperoxia gradually induced death in AECs, noticeably and significantly by 36 h; ~60% of cells were dead by 48 h. However, we unexpectedly found increased expression levels of NRF2-regulated antioxidative genes and nuclear NRF2 in AECs exposed to chronic hyperoxia as compared to acute hyperoxia. Chromatin Immunoprecipitation (ChIP) assays revealed an increased recruitment of NRF2 to the endogenous HMOX1 and NQO1 promoters in AECs exposed to acute or chronic hyperoxia. Thus, our findings demonstrate that NRF2 activation and antioxidant gene expression are functional during hyperoxia-induced lung epithelial cell death and that chronic hyperoxia does not impair NRF2 signaling overall. PMID:23738042

Protective Effects of Hydrogen Sulfide in the Ageing Kidney.

PubMed

Hou, Cui-Lan; Wang, Ming-Jie; Sun, Chen; Huang, Yong; Jin, Sheng; Mu, Xue-Pan; Chen, Ying; Zhu, Yi-Chun

2016-01-01

Aims . The study aimed to examine whether hydrogen sulfide (H 2 S) generation changed in the kidney of the ageing mouse and its relationship with impaired kidney function. Results . H 2 S levels in the plasma, urine, and kidney decreased significantly in ageing mice. The expression of two known H 2 S-producing enzymes in kidney, cystathionine γ -lyase (CSE) and cystathionine- β -synthase (CBS), decreased significantly during ageing. Chronic H 2 S donor (NaHS, 50  μ mol/kg/day, 10 weeks) treatment could alleviate oxidative stress levels and renal tubular interstitial collagen deposition. These protective effects may relate to transcription factor Nrf2 activation and antioxidant proteins such as HO-1, SIRT1, SOD1, and SOD2 expression upregulation in the ageing kidney after NaHS treatment. Furthermore, the expression of H 2 S-producing enzymes changed with exogenous H 2 S administration and contributed to elevated H 2 S levels in the ageing kidney. Conclusions . Endogenous hydrogen sulfide production in the ageing kidney is insufficient. Exogenous H 2 S can partially rescue ageing-related kidney dysfunction by reducing oxidative stress, decreasing collagen deposition, and enhancing Nrf2 nuclear translocation. Recovery of endogenous hydrogen sulfide production may also contribute to the beneficial effects of NaHS treatment.

Nuclear factor erythroid 2-related factor-2 activity controls 4-hydroxynonenal metabolism and activity in prostate cancer cells.

PubMed

Pettazzoni, Piergiorgio; Ciamporcero, Eric; Medana, Claudio; Pizzimenti, Stefania; Dal Bello, Federica; Minero, Valerio Giacomo; Toaldo, Cristina; Minelli, Rosalba; Uchida, Koji; Dianzani, Mario Umberto; Pili, Roberto; Barrera, Giuseppina

2011-10-15

4-Hydroxynonenal (HNE) is an end product of lipoperoxidation with antiproliferative and proapoptotic properties in various tumors. Here we report a greater sensitivity to HNE in PC3 and LNCaP cells compared to DU145 cells. In contrast to PC3 and LNCaP cells, HNE-treated DU145 cells showed a smaller reduction in growth and did not undergo apoptosis. In DU145 cells, HNE did not induce ROS production and DNA damage and generated a lower amount of HNE-protein adducts. DU145 cells had a greater GSH and GST A4 content and GSH/GST-mediated HNE detoxification. Nuclear factor erythroid 2-related factor-2 (Nrf2) is a regulator of the antioxidant response. Nrf2 protein content and nuclear accumulation were higher in DU145 cells compared to PC3 and LNCaP cells, whereas the expression of KEAP1, the main negative regulator of Nrf2 activity, was lower. Inhibition of Nrf2 expression with specific siRNA resulted in a reduction in GST A4 expression and GS-HNE formation, indicating that Nrf2 controls HNE metabolism. In addition, Nrf2 knockdown sensitized DU145 cells to HNE-mediated antiproliferative and proapoptotic activity. In conclusion, we demonstrated that increased Nrf2 activity resulted in a reduction in HNE sensitivity in prostate cancer cells, suggesting a potential mechanism of resistance to pro-oxidant therapy. Copyright © 2011 Elsevier Inc. All rights reserved.

Curcumin Pretreatment Induces Nrf2 and an Antioxidant Response and Prevents Hemin-Induced Toxicity in Primary Cultures of Cerebellar Granule Neurons of Rats

PubMed Central

González-Reyes, Susana; Guzmán-Beltrán, Silvia; Medina-Campos, Omar Noel; Pedraza-Chaverri, José

2013-01-01

Curcumin is a bifunctional antioxidant derived from Curcuma longa. This study identifies curcumin as a neuroprotectant against hemin-induced damage in primary cultures of cerebellar granule neurons (CGNs) of rats. Hemin, the oxidized form of heme, is a highly reactive compound that induces cellular injury. Pretreatment of CGNs with 5–30 μM curcumin effectively increased by 2.3–4.9 fold heme oxygenase-1 (HO-1) expression and by 5.6–14.3-fold glutathione (GSH) levels. Moreover, 15 μM curcumin attenuated by 55% the increase in reactive oxygen species (ROS) production, by 94% the reduction of GSH/glutathione disulfide (GSSG) ratio, and by 49% the cell death induced by hemin. The inhibition of heme oxygenase system or GSH synthesis with tin mesoporphyrin and buthionine sulfoximine, respectively, suppressed the protective effect of curcumin against hemin-induced toxicity. These data strongly suggest that HO-1 and GSH play a major role in the protective effect of curcumin. Furthermore, it was found that 24 h of incubation with curcumin increases by 1.4-, 2.3-, and 5.2-fold the activity of glutathione reductase, glutathione S-transferase and superoxide dismutase, respectively. Additionally, it was found that curcumin was capable of inducing nuclear factor (erythroid-derived 2)-like 2 (Nrf2) translocation into the nucleus. These data suggest that the pretreatment with curcumin induces Nrf2 and an antioxidant response that may play an important role in the protective effect of this antioxidant against hemin-induced neuronal death. PMID:24454990

Eriodictyol-7-O-glucoside activates Nrf2 and protects against cerebral ischemic injury

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jing, Xu; Ren, Dongmei; Wei, Xinbing

Stroke is a complex disease that may involve oxidative stress-related pathways in its pathogenesis. The nuclear factor erythroid-2-related factor 2/antioxidant response element (Nrf2/ARE) pathway plays an important role in inducing phase II detoxifying enzymes and antioxidant proteins and thus has been considered a potential target for neuroprotection in stroke. The aim of the present study was to determine whether eriodictyol-7-O-glucoside (E7G), a novel Nrf2 activator, can protect against cerebral ischemic injury and to understand the role of the Nrf2/ARE pathway in neuroprotection. In primary cultured astrocytes, E7G increased the nuclear localization of Nrf2 and induced the expression of the Nrf2/ARE-dependentmore » genes. Exposure of astrocytes to E7G provided protection against oxygen and glucose deprivation (OGD)-induced oxidative insult. The protective effect of E7G was abolished by RNA interference-mediated knockdown of Nrf2 expression. In vivo administration of E7G in a rat model of focal cerebral ischemia significantly reduced the amount of brain damage and ameliorated neurological deficits. These data demonstrate that activation of Nrf2/ARE signaling by E7G is directly associated with its neuroprotection against oxidative stress-induced ischemic injury and suggest that targeting the Nrf2/ARE pathway may be a promising approach for therapeutic intervention in stroke. - Highlights: • E7G activates Nrf2 in astrocytes. • E7G stimulates expression of Nrf2-mediated cytoprotective proteins in astrocytes. • E7G protects astrocytes against OGD-induced cell death and apoptosis. • The neuroprotective effect of E7G involves the Nrf2/ARE pathway. • E7G protects rats against cerebral ischemic injury.« less

The C-Terminal Domain of Nrf1 Negatively Regulates the Full-Length CNC-bZIP Factor and Its Shorter Isoform LCR-F1/Nrf1β; Both Are Also Inhibited by the Small Dominant-Negative Nrf1γ/δ Isoforms that Down-Regulate ARE-Battery Gene Expression

PubMed Central

Zhang, Yiguo; Qiu, Lu; Li, Shaojun; Xiang, Yuancai; Chen, Jiayu; Ren, Yonggang

2014-01-01

The C-terminal domain (CTD, aa 686–741) of nuclear factor-erythroid 2 p45-related factor 1 (Nrf1) shares 53% amino acid sequence identity with the equivalent Neh3 domain of Nrf2, a homologous transcription factor. The Neh3 positively regulates Nrf2, but whether the Neh3-like (Neh3L) CTD of Nrf1 has a similar role in regulating Nrf1-target gene expression is unknown. Herein, we report that CTD negatively regulates the full-length Nrf1 (i.e. 120-kDa glycoprotein and 95-kDa deglycoprotein) and its shorter isoform LCR-F1/Nrf1β (55-kDa). Attachment of its CTD-adjoining 112-aa to the C-terminus of Nrf2 yields the chimaeric Nrf2-C112Nrf1 factor with a markedly decreased activity. Live-cell imaging of GFP-CTD reveals that the extra-nuclear portion of the fusion protein is allowed to associate with the endoplasmic reticulum (ER) membrane through the amphipathic Neh3L region of Nrf1 and its basic c-tail. Thus removal of either the entire CTD or the essential Neh3L portion within CTD from Nrf1, LCR-F1/Nrf1β and Nrf2-C112Nrf1, results in an increase in their transcriptional ability to regulate antioxidant response element (ARE)-driven reporter genes. Further examinations unravel that two smaller isoforms, 36-kDa Nrf1γ and 25-kDa Nrf1δ, act as dominant-negative inhibitors to compete against Nrf1, LCR-F1/Nrf1β and Nrf2. Relative to Nrf1, LCR-F1/Nrf1β is a weak activator, that is positively regulated by its Asn/Ser/Thr-rich (NST) domain and acidic domain 2 (AD2). Like AD1 of Nrf1, both AD2 and NST domain of LCR-F1/Nrf1β fused within two different chimaeric contexts to yield Gal4D:Nrf1β607 and Nrf1β:C270Nrf2, positively regulate their transactivation activity of cognate Gal4- and Nrf2-target reporter genes. More importantly, differential expression of endogenous ARE-battery genes is attributable to up-regulation by Nrf1 and LCR-F1/Nrf1β and down-regulation by Nrf1γ and Nrf1δ. PMID:25290918

Benznidazole, the trypanocidal drug used for Chagas disease, induces hepatic NRF2 activation and attenuates the inflammatory response in a murine model of sepsis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lambertucci, Flavia

Molecular mechanisms on sepsis progression are linked to the imbalance between reactive oxygen species (ROS) production and cellular antioxidant capacity. Previous studies demonstrated that benznidazole (BZL), known for its antiparasitic action on Trypanosoma cruzi, has immunomodulatory effects, increasing survival in C57BL/6 mice in a model of polymicrobial sepsis induced by cecal ligation and puncture (CLP). The mechanism by which BZL inhibits inflammatory response in sepsis is poorly understood. Also, our group recently reported that BZL is able to activate the nuclear factor erytroide-derived 2-Like 2 (NRF2) in vitro. The aim of the present work was to delineate the beneficial rolemore » of BZL during sepsis, analyzing its effects on the cellular redox status and the possible link to the innate immunity receptor TLR4. Specifically, we analyzed the effect of BZL on Nrf2 regulation and TLR4 expression in liver of mice 24 hours post-CLP. BZL was able to induce NRF2 nuclear protein localization in CLP mice. Also, we found that protein kinase C (PKC) is involved in the NRF2 nuclear accumulation and induction of its target genes. In addition, BZL prompted a reduction in hepatic CLP-induced TLR4 protein membrane localization, evidencing its immunomodulatory effects. Together, our results demonstrate that BZL induces hepatic NRF2 activation with the concomitant increase in the antioxidant defenses, and the attenuation of inflammatory response, in part, by inhibiting TLR4 expression in a murine model of sepsis. - Highlights: • BZL improves survival rate after polymicrobial sepsis • BZL enhances hepatic NRF2 nuclear accumulation in a model of sepsis, in part, by a mechanism dependent on PKC activation • BZL-enhanced NRF2 induction regulates antioxidant enzymes and increases antioxidant cellular defenses in sepsis • BZL blocks liver ROS production and ROS-induced TLR4 plasma membrane expression in septic mice.« less

The role of nuclear factor E2-Related factor 2 and uncoupling protein 2 in glutathione metabolism: Evidence from an in vivo gene knockout study

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Yanyan; The Hamner Institutes for Health Sciences, Research Triangle Park, NC; Xu, Yuanyuan, E-mail: yyxu@cmu.edu.cn

Nuclear factor E2-related factor 2 (NRF2) and uncoupling protein 2 (UCP2) are indicated to protect from oxidative stress. They also play roles in the homeostasis of glutathione. However, the detailed mechanisms are not well understood. In the present study, we found Nrf2-knockout (Nrf2-KO) mice exhibited altered glutathione homeostasis and reduced expression of various genes involved in GSH biosynthesis, regeneration, utilization and transport in the liver. Ucp2-knockout (Ucp2-KO) mice exhibited altered glutathione homeostasis in the liver, spleen and blood, as well as increased transcript of cystic fibrosis transmembrane conductance regulator in the liver, a protein capable of mediating glutathione efflux. Nrf2-Ucp2-doublemore » knockout (DKO) mice showed characteristics of both Nrf2-KO and Ucp2-KO mice. But no significant difference was observed in DKO mice when compared with Nrf2-KO or Ucp2-KO mice, except in blood glutathione levels. These data suggest that ablation of Nrf2 and Ucp2 leads to disrupted GSH balance, which could result from altered expression of genes involved in GSH metabolism. DKO may not evoke more severe oxidative stress than the single gene knockout. - Highlights: • Nrf2/Ucp2 deficiency leads to alteration of glutathione homeostasis. • Nrf2 regulates expression of genes in glutathione generation and utilization. • Ucp2 affects glutathione metabolism by regulating hepatic efflux of glutathione. • Nrf2 deficiency may not aggravate oxidative stress in Ucp2-deficient mice.« less

Discovery and Development of Kelch-like ECH-Associated Protein 1. Nuclear Factor Erythroid 2-Related Factor 2 (KEAP1:NRF2) Protein-Protein Interaction Inhibitors: Achievements, Challenges, and Future Directions.

PubMed

Jiang, Zheng-Yu; Lu, Meng-Chen; You, Qi-Dong

2016-12-22

The transcription factor Nrf2 is the primary regulator of the cellular defense system, and enhancing Nrf2 activity has potential usages in various diseases, especially chronic age-related and inflammatory diseases. Recently, directly targeting Keap1-Nrf2 protein-protein interaction (PPI) has been an emerging strategy to selectively and effectively activate Nrf2. This Perspective summarizes the progress in the discovery and development of Keap1-Nrf2 PPI inhibitors, including the Keap1-Nrf2 regulatory mechanisms, biochemical techniques for inhibitor identification, and approaches for identifying peptide and small-molecule inhibitors, as well as discusses privileged structures and future directions for further development of Keap1-Nrf2 PPI inhibitors.

The novel compound Sul-121 inhibits airway inflammation and hyperresponsiveness in experimental models of chronic obstructive pulmonary disease

PubMed Central

Han, Bing; Poppinga, Wilfred J.; Zuo, Haoxiao; Zuidhof, Annet B.; Bos, I. Sophie T.; Smit, Marieke; Vogelaar, Pieter; Krenning, Guido; Henning, Robert H.; Maarsingh, Harm; Halayko, Andrew J.; van Vliet, Bernard; Stienstra, Stef; Graaf, Adrianus Cornelis van der; Meurs, Herman; Schmidt, Martina

2016-01-01

COPD is characterized by persistent airflow limitation, neutrophilia and oxidative stress from endogenous and exogenous insults. Current COPD therapy involving anticholinergics, β2-adrenoceptor agonists and/or corticosteroids, do not specifically target oxidative stress, nor do they reduce chronic pulmonary inflammation and disease progression in all patients. Here, we explore the effects of Sul-121, a novel compound with anti-oxidative capacity, on hyperresponsiveness (AHR) and inflammation in experimental models of COPD. Using a guinea pig model of lipopolysaccharide (LPS)-induced neutrophilia, we demonstrated that Sul-121 inhalation dose-dependently prevented LPS-induced airway neutrophilia (up to ~60%) and AHR (up to ~90%). Non-cartilaginous airways neutrophilia was inversely correlated with blood H2S, and LPS-induced attenuation of blood H2S (~60%) was prevented by Sul-121. Concomitantly, Sul-121 prevented LPS-induced production of the oxidative stress marker, malondialdehyde by ~80%. In immortalized human airway smooth muscle (ASM) cells, Sul-121 dose-dependently prevented cigarette smoke extract-induced IL-8 release parallel with inhibition of nuclear translocation of the NF-κB subunit, p65 (each ~90%). Sul-121 also diminished cellular reactive oxygen species production in ASM cells, and inhibited nuclear translocation of the anti-oxidative response regulator, Nrf2. Our data show that Sul-121 effectively inhibits airway inflammation and AHR in experimental COPD models, prospectively through inhibition of oxidative stress. PMID:27229886

The role of nuclear factor E2-Related factor 2 and uncoupling protein 2 in glutathione metabolism: Evidence from an in vivo gene knockout study.

PubMed

Chen, Yanyan; Xu, Yuanyuan; Zheng, Hongzhi; Fu, Jingqi; Hou, Yongyong; Wang, Huihui; Zhang, Qiang; Yamamoto, Masayuki; Pi, Jingbo

2016-09-09

Nuclear factor E2-related factor 2 (NRF2) and uncoupling protein 2 (UCP2) are indicated to protect from oxidative stress. They also play roles in the homeostasis of glutathione. However, the detailed mechanisms are not well understood. In the present study, we found Nrf2-knockout (Nrf2-KO) mice exhibited altered glutathione homeostasis and reduced expression of various genes involved in GSH biosynthesis, regeneration, utilization and transport in the liver. Ucp2-knockout (Ucp2-KO) mice exhibited altered glutathione homeostasis in the liver, spleen and blood, as well as increased transcript of cystic fibrosis transmembrane conductance regulator in the liver, a protein capable of mediating glutathione efflux. Nrf2-Ucp2-double knockout (DKO) mice showed characteristics of both Nrf2-KO and Ucp2-KO mice. But no significant difference was observed in DKO mice when compared with Nrf2-KO or Ucp2-KO mice, except in blood glutathione levels. These data suggest that ablation of Nrf2 and Ucp2 leads to disrupted GSH balance, which could result from altered expression of genes involved in GSH metabolism. DKO may not evoke more severe oxidative stress than the single gene knockout. Copyright © 2016 Elsevier Inc. All rights reserved.

Glycogen synthase kinase 3 regulates expression of nuclear factor-erythroid-2 related transcription factor-1 (Nrf1) and inhibits pro-survival function of Nrf1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Biswas, Madhurima; Kwong, Erick K.; Park, Eujean

2013-08-01

Nuclear factor E2-related factor-1 (Nrf1) is a basic leucine zipper transcription factor that is known to regulate antioxidant and cytoprotective gene expression. It was recently shown that Nrf1 is regulated by SCF–Fbw7 ubiquitin ligase. However our knowledge of upstream signals that targets Nrf1 for degradation by the UPS is not known. We report here that Nrf1 expression is negatively regulated by glycogen synthase kinase 3 (GSK3) in Fbw7-dependent manner. We show that GSK3 interacts with Nrf1 and phosphorylates the Cdc4 phosphodegron domain (CPD) in Nrf1. Mutation of serine residue in the CPD of Nrf1 to alanine (S350A), blocks Nrf1 frommore » phosphorylation by GSK3, and stabilizes Nrf1. Knockdown of Nrf1 and expression of a constitutively active form of GSK3 results in increased apoptosis in neuronal cells in response to ER stress, while expression of the GSK3 phosphorylation resistant S350A–Nrf1 attenuates apoptotic cell death. Together these data suggest that GSK3 regulates Nrf1 expression and cell survival function in response to stress activation. Highlights: • The effect of GSK3 on Nrf1 expression was examined. • GSK3 destabilizes Nrf1 protein via Fbw7 ubiquitin ligase. • GSK3 binds and phosphorylates Nrf1. • Protection from stress-induced apoptosis by Nrf1 is inhibited by GSK3.« less

Fasting Induces Nuclear Factor E2-Related Factor 2 and ATP-Binding Cassette Transporters via Protein Kinase A and Sirtuin-1 in Mouse and Human

PubMed Central

Kulkarni, Supriya R.; Donepudi, Ajay C.; Xu, Jialin; Wei, Wei; Cheng, Qiuqiong C.; Driscoll, Maureen V.; Johnson, Delinda A.; Johnson, Jeffrey A.; Li, Xiaoling

2014-01-01

Abstract Aims: The purpose of this study was to determine whether 3′-5′-cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) and Sirtuin-1 (SIRT1) dependent mechanisms modulate ATP-binding Cassette (ABC) transport protein expression. ABC transport proteins (ABCC2–4) are essential for chemical elimination from hepatocytes and biliary excretion. Nuclear factor-E2 related-factor 2 (NRF2) is a transcription factor that mediates ABCC induction in response to chemical inducers and liver injury. However, a role for NRF2 in the regulation of transporter expression in nonchemical models of liver perturbation is largely undescribed. Results: Here we show that fasting increased NRF2 target gene expression through NRF2- and SIRT1–dependent mechanisms. In intact mouse liver, fasting induces NRF2 target gene expression by at least 1.5 to 5-fold. In mouse and human hepatocytes, treatment with 8-Bromoadenosine-cAMP, a cAMP analogue, increased NRF2 target gene expression and antioxidant response element activity, which was decreased by the PKA inhibitor, H-89. Moreover, fasting induced NRF2 target gene expression was decreased in liver and hepatocytes of SIRT1 liver-specific null mice and NRF2-null mice. Lastly, NRF2 and SIRT1 were recruited to MAREs and Antioxidant Response Elements (AREs) in the human ABCC2 promoter. Innovation: Oxidative stress mediated NRF2 activation is well described, yet the influence of basic metabolic processes on NRF2 activation is just emerging. Conclusion: The current data point toward a novel role of nutrient status in regulation of NRF2 activity and the antioxidant response, and indicates that cAMP/PKA and SIRT1 are upstream regulators for fasting-induced activation of the NRF2-ARE pathway. Antioxid. Redox Signal. 20, 15–30. PMID:23725046

Diffusion dynamics of the Keap1–Cullin3 interaction in single live cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Baird, Liam; Dinkova-Kostova, Albena T., E-mail: a.dinkovakostova@dundee.ac.uk; Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, Baltimore, MD

2013-03-29

Highlights: ► We developed a quantitative FRAP-based system to study the Keap1–Cul3 interaction. ► We show that Keap1–EGFP and mCherry–Cul3 interact in single live cells. ► We used inducers which target distinct cysteine sensors of Keap1 and differ 4000-fold in potency. ► Inducers cause Nrf2 stabilization, nuclear translocation, and target gene expression. ► Inducers of four different types do not dissociate the Keap1–EGFP:mCherry–Cul3 complex. -- Abstract: Transcription factor NF-E2 p45-related factor 2 (Nrf2) regulates the expression of a network of genes encoding drug-detoxification, anti-inflammatory, and metabolic enzymes, as well as proteins involved in the regulation of cellular redox homeostasis. Undermore » basal conditions, Kelch-like ECH associated protein 1 (Keap1) targets Nrf2 for ubiquitination and proteasomal degradation via association with Cullin3 (Cul3)-based Rbx1 E3 ubiquitin ligase. Various small molecules (inducers) activate Nrf2 leading to upregulation of cytoprotective gene expression. Inducers chemically modify specific cysteine residues of Keap1 which ultimately loses its ability to target Nrf2 for degradation. Dissociation of the Keap1–Cul3 complex by inducers is one possible mechanism, but evidence in single live cells is lacking. To investigate the diffusion dynamics of the Keap1–Cul3 interaction and the effect of inducers, we developed a quantitative fluorescence recovery after photobleaching (FRAP)-based system using Keap1–EGFP and mCherry–Cul3 fusion proteins. We show that Keap1–EGFP and mCherry–Cul3 interact in single live cells. Exposure for 1 h to small-molecule inducers of 4 different types, the oleanane triterpenoid CDDO, the isothiocyanate sulforaphane, the sulfoxythiocarbamate STCA, and the oxidant hydrogen peroxide which target distinct cysteine sensors within Keap1 with potencies which differ by nearly 4000-fold, does not dissociate the Keap1–Cul3 complex. As inducers cause conformational changes in Keap1, we conclude that changes in conformation rather than dissociation from Cul3 inactivate the repressor function of Keap1 leading to Nrf2 stabilization.« less

Gestational Diabetes Mellitus Impairs Nrf2-Mediated Adaptive Antioxidant Defenses and Redox Signaling in Fetal Endothelial Cells In Utero

PubMed Central

Cheng, Xinghua; Chapple, Sarah J.; Patel, Bijal; Puszyk, William; Sugden, David; Yin, Xiaoke; Mayr, Manuel; Siow, Richard C.M.; Mann, Giovanni E.

2013-01-01

In utero exposure to gestational diabetes mellitus (GDM) is associated with an increased risk of type 2 diabetes and cardiovascular disease in later life, yet the underlying mechanisms remain to be elucidated. We examined the effects of GDM on the proteome, redox status, and nuclear factor erythroid 2–related factor 2 (Nrf2)-mediated antioxidant gene expression in human fetal endothelial cells. Proteomic analysis revealed that proteins involved in redox homeostasis were significantly altered in GDM and associated with increased mitochondrial superoxide generation, protein oxidation, DNA damage, and diminished glutathione (GSH) synthesis. In GDM cells, the lipid peroxidation product 4-hydroxynonenal (HNE) failed to induce nuclear Nrf2 accumulation and mRNA and/or protein expression of Nrf2 and its target genes NAD(P)H:quinone oxidoreductase 1 (NQO1), Bach1, cystine/glutamate transporter, and glutamate cysteine ligase. Although methylation of CpG islands in Nrf2 or NQO1 promoters was unaltered by GDM, decreased DJ-1 and increased phosphorylated glycogen synthase kinase 3β levels may account for impaired Nrf2 signaling. HNE-induced increases in GSH and NQO1 levels were abrogated by Nrf2 small interfering RNA in normal cells, and overexpression of Nrf2 in GDM cells partially restored NQO1 induction. Dysregulation of Nrf2 in fetal endothelium may contribute to the increased risk of type 2 diabetes and cardiovascular disease in offspring. PMID:23974919

Gestational diabetes mellitus impairs Nrf2-mediated adaptive antioxidant defenses and redox signaling in fetal endothelial cells in utero.

PubMed

Cheng, Xinghua; Chapple, Sarah J; Patel, Bijal; Puszyk, William; Sugden, David; Yin, Xiaoke; Mayr, Manuel; Siow, Richard C M; Mann, Giovanni E

2013-12-01

In utero exposure to gestational diabetes mellitus (GDM) is associated with an increased risk of type 2 diabetes and cardiovascular disease in later life, yet the underlying mechanisms remain to be elucidated. We examined the effects of GDM on the proteome, redox status, and nuclear factor erythroid 2-related factor 2 (Nrf2)-mediated antioxidant gene expression in human fetal endothelial cells. Proteomic analysis revealed that proteins involved in redox homeostasis were significantly altered in GDM and associated with increased mitochondrial superoxide generation, protein oxidation, DNA damage, and diminished glutathione (GSH) synthesis. In GDM cells, the lipid peroxidation product 4-hydroxynonenal (HNE) failed to induce nuclear Nrf2 accumulation and mRNA and/or protein expression of Nrf2 and its target genes NAD(P)H:quinone oxidoreductase 1 (NQO1), Bach1, cystine/glutamate transporter, and glutamate cysteine ligase. Although methylation of CpG islands in Nrf2 or NQO1 promoters was unaltered by GDM, decreased DJ-1 and increased phosphorylated glycogen synthase kinase 3β levels may account for impaired Nrf2 signaling. HNE-induced increases in GSH and NQO1 levels were abrogated by Nrf2 small interfering RNA in normal cells, and overexpression of Nrf2 in GDM cells partially restored NQO1 induction. Dysregulation of Nrf2 in fetal endothelium may contribute to the increased risk of type 2 diabetes and cardiovascular disease in offspring.

Deficiency in the nuclear factor E2-related factor 2 renders pancreatic β-cells vulnerable to arsenic-induced cell damage

PubMed Central

Yang, Bei; Fu, Jingqi; Zheng, Hongzhi; Xue, Peng; Yarborough, Kathy; Woods, Courtney G; Hou, Yongyong; Zhang, Qiang; Andersen, Melvin E.; Pi, Jingbo

2012-01-01

Chronic human exposure to inorganic arsenic (iAs), a potent environmental oxidative stressor, is associated with increased prevalence of Type 2 diabetes, where impairment of pancreatic β-cell function is a key pathogenic factor. Nuclear factor E2-related factor 2 (Nrf2) is a central transcription factor regulating cellular adaptive response to oxidative stress. However, persistent activation of Nrf2 in response to chronic oxidative stress, including inorganic arsenite (iAs3+) exposure, blunts glucose-triggered reactive oxygen species (ROS) signaling and impairs glucose-stimulated insulin secretion (GSIS). In the current study, we found that MIN6 pancreatic β-cells with stable knockdown of Nrf2 (Nrf2-KD) by lentiviral shRNA and pancreatic islets isolated from Nrf2-knockout (Nrf2−/−) mice exhibited reduced expression of several antioxidant and detoxification enzymes in response to acute iAs3+ exposure. As a result, Nrf2-KD MIN6 cells and Nrf2−/− islets were more susceptible to iAs3+ and monomethylarsonous acid (MMA3+)-induced cell damage, as measured by decreased cell viability, augmented apoptosis and morphological change. Pretreatment of MIN6 cells with Nrf2 activator tert-butylhydroquinone protected the cells from iAs3+-induced cell damage in an Nrf2-dependent fashion. In contrast, antioxidant N-acetyl cysteine protected Nrf2-KD MIN6 cells against acute cytotoxicity of iAs3+. The present study demonstrates that Nrf2-mediated antioxidant response is critical in the pancreatic β-cell defense mechanism against acute cytotoxicity by arsenic. The findings here, combined with our previous results on the inhibitory effect of antioxidants on ROS signaling and GSIS, suggest that Nrf2 plays paradoxical roles in pancreatic β-cell dysfunction induced by environmental arsenic exposure. PMID:23000044

Molecular Evolution of the Nuclear Factor (Erythroid-Derived 2)-Like 2 Gene Nrf2 in Old World Fruit Bats (Chiroptera: Pteropodidae).

PubMed

Yin, Qiuyuan; Zhu, Lei; Liu, Di; Irwin, David M; Zhang, Shuyi; Pan, Yi-Hsuan

2016-01-01

Mammals developed antioxidant systems to defend against oxidative damage in their daily life. Enzymatic antioxidants and low molecular weight antioxidants (LMWAs) constitute major parts of the antioxidant systems. Nuclear factor (erythroid-derived 2)-like 2 (Nrf2, encoded by the Nrf2 gene) is a central transcriptional regulator, regulating transcription, of many antioxidant enzymes. Frugivorous bats eat large amounts of fruits that contain high levels of LMWAs such as vitamin C, thus, a reliance on LMWAs might greatly reduce the need for antioxidant enzymes in comparison to insectivorous bats. Therefore, it is possible that frugivorous bats have a reduced need for Nrf2 function due to their substantial intake of diet-antioxidants. To test whether the Nrf2 gene has undergone relaxed evolution in fruit-eating bats, we obtained Nrf2 sequences from 16 species of bats, including four Old World fruit bats (Pteropodidae) and one New World fruit bat (Phyllostomidae). Our molecular evolutionary analyses revealed changes in the selection pressure acting on Nrf2 gene and identified seven specific amino acid substitutions that occurred on the ancestral lineage leading to Old World fruit bats. Biochemical experiments were conducted to examine Nrf2 in Old World fruit bats and showed that the amount of catalase, which is regulated by Nrf2, was significantly lower in the brain, heart and liver of Old World fruit bats despite higher levels of Nrf2 protein in Old World fruit bats. Computational predictions suggest that three of these seven amino acid replacements might be deleterious to Nrf2 function. Therefore, the results suggest that Nrf2 gene might have experienced relaxed constraint in Old World fruit bats, however, we cannot rule out the possibility of positive selection. Our study provides the first data on the molecular adaptation of Nrf2 gene in frugivorous bats in compensation to the increased levels of LWMAs from their fruit-diet.

Molecular Evolution of the Nuclear Factor (Erythroid-Derived 2)-Like 2 Gene Nrf2 in Old World Fruit Bats (Chiroptera: Pteropodidae)

PubMed Central

Liu, Di; Irwin, David M.; Zhang, Shuyi; Pan, Yi-Hsuan

2016-01-01

Mammals developed antioxidant systems to defend against oxidative damage in their daily life. Enzymatic antioxidants and low molecular weight antioxidants (LMWAs) constitute major parts of the antioxidant systems. Nuclear factor (erythroid-derived 2)-like 2 (Nrf2, encoded by the Nrf2 gene) is a central transcriptional regulator, regulating transcription, of many antioxidant enzymes. Frugivorous bats eat large amounts of fruits that contain high levels of LMWAs such as vitamin C, thus, a reliance on LMWAs might greatly reduce the need for antioxidant enzymes in comparison to insectivorous bats. Therefore, it is possible that frugivorous bats have a reduced need for Nrf2 function due to their substantial intake of diet-antioxidants. To test whether the Nrf2 gene has undergone relaxed evolution in fruit-eating bats, we obtained Nrf2 sequences from 16 species of bats, including four Old World fruit bats (Pteropodidae) and one New World fruit bat (Phyllostomidae). Our molecular evolutionary analyses revealed changes in the selection pressure acting on Nrf2 gene and identified seven specific amino acid substitutions that occurred on the ancestral lineage leading to Old World fruit bats. Biochemical experiments were conducted to examine Nrf2 in Old World fruit bats and showed that the amount of catalase, which is regulated by Nrf2, was significantly lower in the brain, heart and liver of Old World fruit bats despite higher levels of Nrf2 protein in Old World fruit bats. Computational predictions suggest that three of these seven amino acid replacements might be deleterious to Nrf2 function. Therefore, the results suggest that Nrf2 gene might have experienced relaxed constraint in Old World fruit bats, however, we cannot rule out the possibility of positive selection. Our study provides the first data on the molecular adaptation of Nrf2 gene in frugivorous bats in compensation to the increased levels of LWMAs from their fruit-diet. PMID:26735303

Pyrrolidine dithiocarbamate activates the Nrf2 pathway in astrocytes.

PubMed

Liddell, Jeffrey R; Lehtonen, Sarka; Duncan, Clare; Keksa-Goldsteine, Velta; Levonen, Anna-Liisa; Goldsteins, Gundars; Malm, Tarja; White, Anthony R; Koistinaho, Jari; Kanninen, Katja M

2016-02-26

Endogenous defense against oxidative stress is controlled by nuclear factor erythroid 2-related factor 2 (Nrf2). The normal compensatory mechanisms to combat oxidative stress appear to be insufficient to protect against the prolonged exposure to reactive oxygen species during disease. Counterbalancing the effects of oxidative stress by up-regulation of Nrf2 signaling has been shown to be effective in various disease models where oxidative stress is implicated, including Alzheimer's disease. Stimulation of Nrf2 signaling by small-molecule activators is an appealing strategy to up-regulate the endogenous defense mechanisms of cells. Here, we investigate Nrf2 induction by the metal chelator and known nuclear factor-κB inhibitor pyrrolidine dithiocarbamate (PDTC) in cultured astrocytes and neurons, and mouse brain. Nrf2 induction is further examined in cultures co-treated with PDTC and kinase inhibitors or amyloid-beta, and in Nrf2-deficient cultures. We show that PDTC is a potent inducer of Nrf2 signaling specifically in astrocytes and demonstrate the critical role of Nrf2 in PDTC-mediated protection against oxidative stress. This induction appears to be regulated by both Keap1 and glycogen synthase kinase 3β. Furthermore, the presence of amyloid-beta magnifies PDTC-mediated induction of endogenous protective mechanisms, therefore suggesting that PDTC may be an effective Nrf2 inducer in the context of Alzheimer's disease. Finally, we show that PDTC increases brain copper content and glial expression of heme oxygenase-1, and decreases lipid peroxidation in vivo, promoting a more antioxidative environment. PDTC activates Nrf2 and its antioxidative targets in astrocytes but not neurons. These effects may contribute to the neuroprotection observed for PDTC in models of Alzheimer's disease.

Nrf2: bane or blessing in cancer?

PubMed

Xiang, MingJun; Namani, Akhileshwar; Wu, ShiJun; Wang, XiaoLi

2014-08-01

The Kelch-like ECH-associated protein 1 (Keap1)-nuclear factor-E2-related factor 2 (Nrf2)-antioxidant response element pathway serves a major function in endogenous cytoprotection in normal cells. Nrf2 is a transcription factor that mainly regulates the expression of a wide array of genes that produce the antioxidants and other proteins responsible for the detoxification of xenobiotics and reactive oxygen species. Nrf2 mediates the chemoprevention of cancer in normal cells. Growing body of evidence suggests that Nrf2 is not only involved in the chemoprevention of normal cells but also promotes the growth of cancer cells. However, the mechanism underlying the function of Nrf2 in oncogenesis and tumor protection in cancer cells remains unclear and thus requires further study. This review aims to rationalize the existing functions of Nrf2 in chemoprevention and tumorigenesis, as well as the somatic mutations of Nrf2 and Keap1 in cancer and Nrf2 cross talk with miRNAs. This review also discusses the future challenges in Nrf2 research.

Sargassum fusiforme polysaccharides activate antioxidant defense by promoting Nrf2-dependent cytoprotection and ameliorate stress insult during aging.

PubMed

Chen, Peichao; He, Dan; Zhang, Ya; Yang, Shanshan; Chen, Liujun; Wang, Shengqin; Zou, Huixi; Liao, Zhiyong; Zhang, Xu; Wu, Mingjiang

2016-11-09

Aging is a complex issue, which results in a progressive decline process in cellular protection and physiological functions. Illustrating the causes of aging and pharmaceutical interference with the aging process has been a pivotal issue for thousands of years. Sargassum fusiforme (S. fusiforme), a kind of brown alga, is also named the "longevity vegetable" as it is not only a kind of food, but also used as an herb in traditional Chinese Medicine for maintaining health and treatment of thyroid disease, cardiovascular disease and so on. But how S. fusiforme promotes longevity is vastly equivocal. We got clues from S. fusiforme polysaccharides, which exhibited antioxidant activity, but the underlying mechanisms remained unclear. In this study, we evaluated the antioxidant effect and the possible mechanisms that S. fusiforme polysaccharides have against d-galactose-induced aging and chronic aging. We selected the SFPS as the candidate for antioxidant defense evaluation, which is a type of S. fusiforme polysaccharide with strong free radical scavenging activity and non-toxicity. It revealed that the antioxidant defense of the d-galactose-induced mice was markedly recovered when they were intragastrically administrated with the SFPS. However, oxidative damage may not be the only cause of aging. We further evaluated the function of the SFPS in the chronic aging mice. Intriguingly, we even found an obvious aging phenotype in the middle aged male ICR mice, which showed a significant decline in Nrf2-dependent cytoprotection. When 9-month old male ICR mice were treated with the SFPS for 2 months or even 11 months to their mean survival age, experimental measurements showed that the SFPS significantly promoted the antioxidant defense and mitochondrial integrity during aging. Furthermore, we suggest that the SFPS promotes Nrf2-dependent cytoprotection by upregulating the nuclear Nrf2 translocation, which may be mediated by p21 and JNK dependent pathways. These results suggest that the SFPS may decelerate the aging process by enhancing Nrf2-dependent cytoprotection, especially antioxidant defense.

Effect of cellular ubiquitin levels on the regulation of oxidative stress response and proteasome function via Nrf1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Donghee; Ryu, Kwon-Yul

The polyubiquitin genes Ubb and Ubc are upregulated under oxidative stress induced by arsenite [As(III)]. However, the role of ubiquitin (Ub) under As(III) exposure is not known in detail. In a previous study, we showed that the reduced viability observed in Ubc{sup −/−} mouse embryonic fibroblasts under As(III) exposure was not due to dysregulation of the Nrf2–Keap1 pathway, which prompted us to investigate another NFE2 family protein, nuclear factor erythroid 2-related factor 1 (Nrf1). In this study, we found that Ub deficiency due to Ubc knockdown in N2a cells reduced cell viability and proteasome activity under As(III) exposure. Furthermore, mRNAmore » levels of the proteasome subunit Psma1 were also reduced. In addition, Ub deficiency led to the nuclear accumulation of the p65 isoform of Nrf1 under As(III) exposure. Interestingly, the overexpression of p65-Nrf1 recapitulated the phenotypes of Ub-deficient N2a cells under As(III) exposure. On the other hand, Nrf1 knockdown suppressed the death of Ub-deficient N2a cells upon exposure to As(III). Therefore, the levels of p65-Nrf1 may play an important role in the maintenance of cell viability under oxidative stress induced by As(III). - Highlights: • N2a cells exhibit reduced viability upon exposure to As(III) via Ubc knockdown. • As(III)-induced proteasomal regulation is impaired in Ub-deficient N2a cells. • Ub deficiency leads to the nuclear accumulation of p65-Nrf1 under As(III) exposure. • p65 expression recapitulates As(III)-induced phenotypes of Ub-deficient N2a cells. • Nrf1 knockdown suppressed As(III)-induced death of Ub-deficient N2a cells.« less

Toll-Like Receptor Signaling Induces Nrf2 Pathway Activation through p62-Triggered Keap1 Degradation.

PubMed

Yin, Shasha; Cao, Wangsen

2015-08-01

Toll-like receptors (TLRs) induce inflammation and tissue repair through multiple signaling pathways. The Nrf2 pathway plays a key role in defending against the tissue damage incurred by microbial infection or inflammation-associated diseases. The critical event that mediates TLR-induced Nrf2 activation is still poorly understood. In this study, we found that lipopolysaccharide (LPS) and other Toll-like receptor (TLR) agonists activate Nrf2 signaling and the activation is due to the reduction of Keap1, the key Nrf2 inhibitor. TLR signaling-induced Keap1 reduction promoted Nrf2 translocation from the cytoplasm to the nucleus, where it activated transcription of its target genes. TLR agonists modulated Keap1 at the protein posttranslation level through autophagy. TLR signaling increased the expression of autophagy protein p62 and LC3-II and induced their association with Keap1 in the autophagosome-like structures. We also characterized the interaction between p62 and Keap1 and found that p62 is indispensable for TLR-mediated Keap1 reduction: TLR signaling had no effect on Keap1 if cells lacked p62 or if cells expressed a mutant Keap1 that could not interact with p62. Our study indicates that p62-mediated Keap1 degradation through autophagy represents a critical linkage for TLR signaling regulation of the major defense network, the Nrf2 signaling pathway. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Clinically significant association of elevated expression of nuclear factor E2-related factor 2 expression with higher glucose uptake and progression of upper urinary tract cancer.

PubMed

Nukui, Akinori; Narimatsu, Takahiro; Kambara, Tsunehito; Abe, Hideyuki; Sakamoto, Setsu; Yoshida, Ken-Ichiro; Kamai, Takao

2018-05-02

There is growing evidence that the transcription factor nuclear factor E2-related factor 2 (Nrf2) is the major participant in regulating antioxidants and pathways for detoxifying reactive oxygen species (ROS), as well as having a vital role in tumor proliferation, invasion, and chemoresistance. It was also recently reported that Nrf2 supports cell proliferation by promoting metabolic activity. Thus, Nrf2 is involved in progression of cancer. Upper urinary tract urothelial carcinoma (UTUC) is a biologically aggressive tumor with high rates of recurrence and progression, resulting in a poor prognosis. However, the role of Nrf2 in UTUC is largely unknown. In order to study the role of Nrf2 in UTUC from the metabolic perspective, we retrospectively assessed Nrf2 expression in the surgical specimen and the preoperative maximum standard glucose uptake (SUVmax) on [ 18 F]fluorodeoxy-glucose positron emission tomography ( 18 F-FDG-PET) of 107 patients with UTUC who underwent radical nephroureterectomy. Increased expression of Nrf2 in the primary lesion was correlated with less differentiated histology, local invasion, and lymph node metastasis, and was also an independent indicator of shorter overall survival according to multivariate analysis. Furthermore, increased expression of Nrf2 was associated with higher preoperative SUVmax by the primary tumor on 18 F-FDG-PET, while Nrf2 expression and SUVmax were also significantly correlated in the metastatic lymph nodes. Among the 18 patients with lymph node metastasis at nephroureterectomy who underwent retroperitoneal lymph node dissection and received adjuvant chemotherapy, the patients with higher Nrf2 expression in the primary tumor had worse recurrence-free survival. These results suggest that constitutive activation of Nrf2 might be linked with tumor aerobic glycolysis and progression of UTUC, indicating that Nrf2 signaling in the tumor microenvironment promotes progression of UTUC.

Ischemic Preconditioning Protects Astrocytes against Oxygen Glucose Deprivation Via the Nuclear Erythroid 2-Related Factor 2 Pathway.

PubMed

Narayanan, Srinivasan V; Dave, Kunjan R; Perez-Pinzon, Miguel A

2018-04-01

Induction of ischemic preconditioning (IPC) represents a potential therapy against cerebral ischemia by activation of adaptive pathways and modulation of mitochondria to induce ischemic tolerance to various cells and tissues. Mitochondrial dysfunction has been ascribed to contribute to numerous neurodegenerative conditions and cerebral ischemia. Nuclear erythroid 2-related factor 2 (Nrf2) is a transcription factor that has traditionally been involved in upregulating cellular antioxidant systems to combat oxidative stress in the brain; however, the association of Nrf2 with mitochondria in the brain remains unclear. In the present study, we investigated the effects of Nrf2 on (i) IPC-induced protection of astrocytes; (ii) OXPHOS protein expression; and (iii) mitochondrial supercomplex formation.Oxygen-glucose deprivation (OGD) was used as an in vitro model of cerebral ischemia and IPC in cultured rodent astrocytes derived from WT C57Bl/6J and Nrf2 -/- mice. OXPHOS proteins were probed via western blotting, and supercomplexes were determined by blue native gel electrophoresis.IPC-induced cytoprotection in wild-type, but not Nrf2 -/- mouse astrocyte cultures following a lethal duration of OGD. In addition, our results suggest that Nrf2 localizes to the outer membrane in non-synaptic brain mitochondria, and that a lack of Nrf2 in vivo produces altered supercomplex formation in mitochondria.Our findings support a role of Nrf2 in mediating IPC-induced protection in astrocytes, which can profoundly impact the ischemic tolerance of neurons. In addition, we provide novel evidence for the association of Nrf2 to brain mitochondria and supercomplex formation. These studies offer new targets and pathways of Nrf2, which may be heavily implicated following cerebral ischemia.

Characterization of Nrf2 activation and heme oxygenase-1 expression in NIH3T3 cells exposed to aqueous extracts of cigarette smoke.

PubMed

Knörr-Wittmann, Constanze; Hengstermann, Arnd; Gebel, Stephan; Alam, Jawed; Müller, Thomas

2005-12-01

Cigarette smoke (CS) is a complex chemical mixture estimated to be composed of up to 5000 different chemicals, many of which are prooxidant. Here we show that, at least in vitro, the cellular response designed to combat oxidative stress resulting from CS exposure is primarily controlled by the transcription factor Nrf2, a principal inducer of antioxidant and phase II-related genes. The prominent role of Nrf2 in the cellular response to CS is substantiated by the following observations: In NIH3T3 cells exposed to aqueous extracts of CS (i) Nrf2 is strongly stabilized and becomes detectable in nuclear extracts. (ii) Nuclear localization of Nrf2 coincides with increased DNA binding of a putative Nrf2/MafK heterodimer to its cognate cis-regulatory site, i.e., the antioxidant-responsive element (ARE). (iii) Studies on the regulatory elements of the oxidative stress-inducible gene heme oxygenase-1 (hmox1) using various hmox1 promoter/luciferase reporter constructs revealed that the strong CS-dependent expression of this gene is primarily governed by the distal enhancers 1 ("E1") and 2 ("E2"), which both contain three canonical ARE-like stress-responsive elements (StREs). Notably, depletion of Nrf2 levels caused by RNA interference significantly compromised CS-induced hmox1 promoter activation, based on the distinct Nrf2 sensitivity exhibited by E1 and E2. Finally, (iv) siRNA-dependent knock-down of Nrf2 completely abrogated CS-induced expression of phase II-related genes. Taken together, these results confirm the outstanding role of Nrf2 both in sensing (oxidant) stress and in orchestrating an efficient transcriptional response aimed at resolving the stressing conditions.

Hepatoprotective effect of 2'-O-galloylhyperin against oxidative stress-induced liver damage through induction of Nrf2/ARE-mediated antioxidant pathway.

PubMed

Wang, Peng; Gao, Yi-Meng; Sun, Xing; Guo, Na; Li, Ji; Wang, Wei; Yao, Li-Ping; Fu, Yu-Jie

2017-04-01

2'-O-galloylhyperin (2'-O-GH), an active compound isolated from Pyrola calliantha, possesses remarkable antioxidant activity. The aims of this study were to investigate the hepatoprotective effect of 2'-O-GH against oxidative stress and elucidate the underlying mechanistic signaling pathways in HepG2 cells as well as in an animal model. Results showed that 2'-O-GH significantly inhibited hydrogen peroxide (H 2 O 2 )-induced HepG2 cell death in a dose dependent manner. The mitogen-activated protein kinase activation, ROS production, mitochondrial membrane potential, intracellular calcium level and subsequent apoptotic protein activation in H 2 O 2 -stimulated HepG2 cells were remarkably inhibited by 2'-O-GH. Furthermore, 2'-O-GH stimulation resulted in a fast and dramatic activation of Akt and nuclear translocation of the NF-E2-related factor 2 (Nrf2), along with the increased expression of heme oxygenase-1 (HO-1) and levels of glutathione (GSH). Meanwhile, histopathological evaluation of the liver also revealed that 2'-O-GH effectively ameliorated CCl 4 -induced the hepatic damage by reducing alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities. Therefore, these results suggested the hepatoprotective effect of 2'-O-GH might be correlated with its antioxidant and free radical scavenger effect. Copyright © 2017. Published by Elsevier Ltd.

Deficiency in the nuclear factor E2-related factor 2 renders pancreatic β-cells vulnerable to arsenic-induced cell damage

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Bei; Department of Histology and Embryology, College of Basic Medical Sciences, China Medical University, Shenyang 110001; Fu, Jingqi

2012-11-01

Chronic human exposure to inorganic arsenic (iAs), a potent environmental oxidative stressor, is associated with increased prevalence of type 2 diabetes, where impairment of pancreatic β-cell function is a key pathogenic factor. Nuclear factor E2-related factor 2 (Nrf2) is a central transcription factor regulating cellular adaptive response to oxidative stress. However, persistent activation of Nrf2 in response to chronic oxidative stress, including inorganic arsenite (iAs{sup 3+}) exposure, blunts glucose-triggered reactive oxygen species (ROS) signaling and impairs glucose-stimulated insulin secretion (GSIS). In the current study, we found that MIN6 pancreatic β-cells with stable knockdown of Nrf2 (Nrf2-KD) by lentiviral shRNA andmore » pancreatic islets isolated from Nrf2-knockout (Nrf2−/−) mice exhibited reduced expression of several antioxidant and detoxification enzymes in response to acute iAs{sup 3+} exposure. As a result, Nrf2-KD MIN6 cells and Nrf2−/− islets were more susceptible to iAs{sup 3+} and monomethylarsonous acid (MMA{sup 3+})-induced cell damage, as measured by decreased cell viability, augmented apoptosis and morphological change. Pretreatment of MIN6 cells with Nrf2 activator tert-butylhydroquinone protected the cells from iAs{sup 3+}-induced cell damage in an Nrf2-dependent fashion. In contrast, antioxidant N‐acetyl cysteine protected Nrf2-KD MIN6 cells against acute cytotoxicity of iAs{sup 3+}. The present study demonstrates that Nrf2-mediated antioxidant response is critical in the pancreatic β-cell defense mechanism against acute cytotoxicity by arsenic. The findings here, combined with our previous results on the inhibitory effect of antioxidants on ROS signaling and GSIS, suggest that Nrf2 plays paradoxical roles in pancreatic β-cell dysfunction induced by environmental arsenic exposure. -- Highlights: ► Lack of Nrf2 reduced expression of antioxidant genes induced by iAs{sup 3+} in β-cells. ► Deficiency of Nrf2 in β-cells sensitized to iAs{sup 3+} and MMA{sup 3+}-induced cytotoxicity. ► Nrf2 activation protected β-cells from acute iAs{sup 3+} cytotoxicity.« less

Protective effect of nuclear factor E2-related factor 2 on inflammatory cytokine response to brominated diphenyl ether-47 in the HTR-8/SVneo human first trimester extravillous trophoblast cell line

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Hae-Ryung, E-mail: heaven@umich.edu; Loch-Caruso, Rita

Polybrominated diphenyl ethers (PBDEs) are widely used flame retardants, and BDE-47 is a prevalent PBDE congener detected in human tissues. Exposure to PBDEs has been linked to adverse pregnancy outcomes in humans. Although the underlying mechanisms of adverse birth outcomes are poorly understood, critical roles for oxidative stress and inflammation are implicated. The present study investigated antioxidant responses in a human extravillous trophoblast cell line, HTR-8/SVneo, and examined the role of nuclear factor E2-related factor 2 (Nrf2), an antioxidative transcription factor, in BDE-47-induced inflammatory responses in the cells. Treatment of HTR-8/SVneo cells with 5, 10, 15, and 20 μM BDE-47more » for 24 h increased intracellular glutathione (GSH) levels compared to solvent control. Treatment of HTR-8/SVneo cells with 20 μM BDE-47 for 24 h induced the antioxidant response element (ARE) activity, indicating Nrf2 transactivation by BDE-47 treatment, and resulted in differential expression of redox-sensitive genes compared to solvent control. Pretreatment with tert-butyl hydroquinone (tBHQ) or sulforaphane, known Nrf2 inducers, reduced BDE-47-stimulated IL-6 release with increased ARE reporter activity, reduced nuclear factor kappa B (NF-κB) reporter activity, increased GSH production, and stimulated expression of antioxidant genes compared to non-Nrf2 inducer pretreated groups, suggesting that Nrf2 may play a protective role against BDE-47-mediated inflammatory responses in HTR-8/SVneo cells. These results suggest that Nrf2 activation significantly attenuated BDE-47-induced IL-6 release by augmentation of cellular antioxidative system via upregulation of Nrf2 signaling pathways, and that Nrf2 induction may be a potential therapeutic target to reduce adverse pregnancy outcomes associated with toxicant-induced oxidative stress and inflammation. - Highlights: • BDE-47 stimulated ARE reporter activity and GSH production. • BDE-47 resulted in differential expression of redox-sensitive genes. • Nrf2 inducers upregulated Nrf2-mediated oxidative stress responses. • Nrf2 inducers reduced BDE-47-stimulated IL-6 release and NF-κB activity.« less

Fermented wheat powder induces the antioxidant and detoxifying system in primary rat hepatocytes.

PubMed

La Marca, Margherita; Beffy, Pascale; Pugliese, Annalisa; Longo, Vincenzo

2013-01-01

Many plants exhibit antioxidant properties which may be useful in the prevention of oxidative stress reactions, such as those mediated by the formation of free radical species in different pathological situations. In recent years a number of studies have shown that whole grain products in particular have strong antioxidant activity. Primary cultures of rat hepatocytes were used to investigate whether and how a fermented powder of wheat (Lisosan G) is able to modulate antioxidant and detoxifying enzymes, and whether or not it can activate Nrf2 transcription factor or inhibit NF-kB activation. All of the antioxidant and detoxifying enzymes studied were significantly up-regulated by 0.7 mg/ml Lisosan G treatment. In particular, quinone oxidoreductase and heme oxygenase-1 were induced, although to different degrees, at the transcriptional, protein and/or activity levels by the treatment. As for the Nrf2 transcription factor, a partial translocation of its protein from the cytosol to the nucleus after 1 h of Lisosan G treatment was revealed by immunoblotting. Lisosan G was also observed to decrease H2O2-induced toxicity Taken together, these results show that this powder of wheat is an effective inducer of ARE/Nrf2-regulated antioxidant and detoxifying genes and has the potential to inhibit the translocation of NF-kB into the nucleus.

α-Dihydroxychalcone-glycoside (α-DHC) isolated from the heartwood of Pterocarpus marsupium inhibits LPS induced MAPK activation and up regulates HO-1 expression in murine RAW 264.7 macrophage

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chakraborty, Prarthana; Saraswat, Ghungroo; Kabir, Syed N., E-mail: snkabir@iicb.res.in

2014-05-15

Three phenolic glycosides isolated from the heartwood of Pterocarpus marsupium showed significant free radical and superoxide ion scavenging activity and antioxidant potential that were comparable to, or several folds higher than those of standard antioxidants, trolox and ascorbic acid. The effective concentrations of these compounds were far below their cytotoxic levels. Compound 3, which was characterized to be α-dihydroxychalcone-glycoside (α-DHC), was the most potent one. Subsequent studies demonstrated that α-DHC effectively reduced nitric oxide and cytokine production by the LPS stimulated RAW 264.7 mouse macrophage cell line. The compound effectively attenuated the expression of inflammation-mediating enzymes COX-2 and iNOS atmore » the mRNA as well as protein levels in a concentration dependent manner. It prevented phosphorylation of all the three MAPKs (JNK, ERK, p38) and eventually blocked the activation of downstream elements contributing to inflammation. Phosphorylation of IκB-α and subsequent translocation of NF-κB into the nucleus were restricted, while the expression of stress responsive gene HO-1 was up-regulated. α-DHC targeted Keap-1 by modifying its cysteine thiols, dissociating it from Nrf-2 and facilitating nuclear entry of the latter; and this in turn induced HO-1 expression. Thus α-DHC exerts its anti-inflammatory activity in a dual manner: by down regulating MAPKs and restricting nuclear stabilization of NF-κB at one end, and by disrupting Nrf-2–Keap-1 complex on the other. In conclusion, the anti-inflammatory potential together with its high therapeutic index envisages α-DHC as a prospective candidate molecule for the development of therapeutic strategy against inflammatory disorders. - Highlights: • α-DHC isolated from Pterocarpus marsupium has significant antioxidant potential. • α-DHC inhibits NO, IL-6, IL-1β, TNF-α production in LPS-stimulated RAW 264.7 cells. • α-DHC down-regulates of COX-2, iNOS expression in LPS-stimulated RAW 264.7 cells. • α-DHC down-regulates MAPK phosphorylation and nuclear translocation of NF-κB. • α-DHC induces ARE-regulated gene expression via Keap-1 modification.« less

[CNC proteins in physiology and pathology].

PubMed

Gęgotek, Agnieszka; Skrzydlewska, Elżbieta

2015-07-06

CNC proteins consist of Bach1, Bach2 and 4 homologous transcription factors: Nrf1, Nrf2, Nrf3 and p45NF-E2. Transcription factors belonging to this group of proteins play a crucial role in protection of cells against oxidative stress. Under physiological conditions, they remain in the cytoplasm in the inactive form or are degraded. However, in oxidative stress conditions, they are translocated to the nucleus, and bind to DNA in the ARE sequence. Consequently, there is transcription of genes encoding cytoprotective proteins, such as phase II enzymes, or low molecular weight antioxidant proteins (i.e., thioredoxin, ferritin, metallothionein) responsible for protecting cells from reactive oxygen species (ROS) action. The activity of transcriptional proteins depends directly on the redox state of the cell. ROS as second messenger signals, control inhibitors of cytoplasmic CNC proteins or potentiate the activity of kinases (MAPK, PKC, PI3K, PERK), leading to phosphorylation of transcription factors. This is conducive to translocation of these molecules into the nucleus and to formation of complexes that initiate the gene expression. Disorders of regulation of the activity of transcription factors belonging to the CNC proteins caused by gene mutations, epigenetic modifications or increased activity of p62, p21, or k-Ras, B-Raf and c-Myc oncogenes, induce changes in the level of ARE-dependent gene expression, which can lead even to the development of carcinogenesis. On the other hand, Nrf transcription factors, inducing the expression of antioxidants and enzymes responsible for the detoxification of xenobiotics, can be considered as a potential target of the action of chemopreventive factors in anticancer therapy.

Natural product-derived pharmacological modulators of Nrf2/ARE pathway for chronic diseases.

PubMed

Kumar, Hemant; Kim, In-Su; More, Sandeep Vasant; Kim, Byung-Wook; Choi, Dong-Kug

2014-01-01

Covering: 2000 to 2013. Oxidative stress is the central component of chronic diseases. The nuclear factor erythroid 2-related factor 2/antioxidant response element (Nrf2/ARE) pathway is vital in the up-regulation of cytoprotective genes and enzymes in response to oxidative stress and treatment with certain dietary phytochemicals. Herein, we classify bioactive compounds derived from natural products that are Nrf2/ARE pathway activators and recapitulate the molecular mechanisms for inducing Nrf2 to provide favorable effects in experimental models of chronic diseases. Moreover, pharmacological inhibition of Nrf2 signalling has emerged as promising strategy against multi-drug resistance thereby improving the treatment efficacy. We have also enlisted natural product-derived inhibitors of Nrf2/ARE pathway.

Sulforaphane enhances the activity of the Nrf2-ARE pathway and attenuates inflammation in OxyHb-induced rat vascular smooth muscle cells.

PubMed

Zhao, X-D; Zhou, Y-T; Lu, X-J

2013-09-01

A growing body of evidence indicates that the nuclear factor erythroid 2-related factor 2-antioxidant response element (Nrf2-ARE) pathway plays a protective role in many physiological stress processes such as inflammatory damage, oxidative stress, and the accumulation of toxic metabolites, which are all involved in the cerebral vasospasm following subarachnoid hemorrhage (SAH). We hypothesized that the Nrf2-ARE pathway might have a protective role in cerebral vasospasm following SAH. In our study, we investigate whether the oxyhemoglobin (OxyHb) can induce the activation of the Nrf2-ARE pathway in vascular smooth muscle cells (VSMCs), and evaluate the modulatory effects of sulforaphane (SUL) on OxyHb-induced inflammation in VSMCs. As a result, both the protein level and the mRNA level of the nuclear Nrf2 were significantly increased, while the mRNA levels of two Nrf2-regulated gene products, both heme oxygenase-1 and NAD(P)H: quinone oxidoreductase-1, were also up-regulated in VSMCs induced with OxyHb. A marked increase of inflammatory cytokines such as IL-1β, IL-6 and TNF-α release was observed at 48 h after cells were treated with OxyHb. SUL enhanced the activity of the Nrf2-ARE pathway and suppressed cytokine release. Our results indicate that the Nrf2-ARE pathway was activated in OxyHb-induced VSMCs. SUL suppressed cytokine release via the activation of the Nrf2-ARE pathway in OxyHb-induced VSMCs.

Nuclear Factor (Erythroid-Derived 2)-Like 2 and Thioredoxin-1 in Atherosclerosis and Ischemia/Reperfusion Injury in the Heart

PubMed Central

Jakobs, Philipp; Serbulea, Vlad; Leitinger, Norbert; Eckers, Anna

2017-01-01

Abstract Significance: Redox signaling is one of the key elements involved in cardiovascular diseases. Two important molecules are the transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) and the oxidoreductase thioredoxin-1 (Trx-1). Recent Advances: During the previous years, a lot of studies investigated Nrf2 and Trx-1 as protective proteins in cardiovascular disorders. Moreover, post-translational modifications of those molecules were identified that play an important role in the cardiovascular system. This review will summarize changes in the vasculature in atherosclerosis and ischemia reperfusion injury of the heart and the newest findings achieved with Nrf2 and Trx-1 therein. Interestingly, Nrf2 and Trx-1 can act together as well as independently of each other in protection against atherosclerosis and ischemia and reperfusion injury. Critical Issues: In principle, pharmacological activation of a transcription factor-like Nrf2 can be dangerous, since a transcription regulator has multiple targets and the pleiotropic effects of such activation should not be ignored. Moreover, overactivation of Nrf2 as well as long-term treatment with Trx-1 could be deleterious for the cardiovascular system. Future Directions: Therefore, the length of treatment with Nrf2 activators and/or Trx-1 has first to be studied in more detail in cardiovascular disorders. Moreover, a combination of Nrf2 activators and Trx-1 should be investigated and taken into consideration. Antioxid. Redox Signal. 26, 630–644. PMID:27923281

Naphthazarin protects against glutamate-induced neuronal death via activation of the Nrf2/ARE pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Son, Tae Gen; Kawamoto, Elisa M.; Yu, Qian-Sheng

2013-04-19

Highlights: •Naphthazarin activates the Nrf2/ARE pathway. •Naphthazarin induces Nrf2-driven genes in neurons and astrocytes. •Naphthazarin protects neurons against excitotoxicity. -- Abstract: Nuclear factor E2-related factor 2 (Nrf2)/antioxidant response element (ARE) pathway is an important cellular stress response pathway involved in neuroprotection. We previously screened several natural phytochemicals and identified plumbagin as a novel activator of the Nrf2/ARE pathway that can protect neurons against ischemic injury. Here we extended our studies to natural and synthetic derivatives of plumbagin. We found that 5,8-dimethoxy-1,4-naphthoquinone (naphthazarin) is a potent activator of the Nrf2/ARE pathway, up-regulates the expression of Nrf2-driven genes in primary neuronal andmore » glial cultures, and protects neurons against glutamate-induced excitotoxicity.« less

Estrogen-Dependent Nrf2 Expression Protects Against Reflux-Induced Esophagitis.

PubMed

Torihata, Yudai; Asanuma, Kiyotaka; Iijima, Katsunori; Mikami, Tetsuhiko; Hamada, Shin; Asano, Naoki; Koike, Tomoyuki; Imatani, Akira; Masamune, Atsushi; Shimosegawa, Tooru

2018-02-01

Gastroesophageal reflux disease is more common in males than in females. The enhanced antioxidative capacity of estrogen in females might account for the gender difference. Nuclear factor erythroid 2-related factor 2 (Nrf2) plays a pivotal role in the host defense mechanism against oxidative stress. This study aimed to clarify the role of Nrf2 in reflux-induced esophageal inflammation, focusing on the gender difference and nitric oxide. Gastroesophageal reflux was surgically induced in male and female rats. Nitrite and ascorbic acid were administered for 1 week to provoke nitric oxide in the esophageal lumen. Male rats with gastroesophageal reflux were supplemented with 17β-estradiol or tert-butylhydroquinone, an Nrf2-inducing reagent. Esophageal squamous cell carcinoma KYSE30 cells were treated with 17β-estradiol. Nrf2 expression was examined by Western blotting and quantitative real-time PCR. Antioxidant gene expression profiles were examined by a PCR array. In the presence of nitric oxide, reflux-induced esophageal damage was less evident, whereas esophageal expression of Nrf2 and its target genes such as Nqo1 was more evident in female or male rats supplemented with 17β-estradiol than in male rats. 17β-Estradiol increased nuclear Nrf2 expression in KYSE30 cells. tert-Butylhydroquinone increased tissue Nqo1 mRNA expression, leading to a reduction in reflux-induced esophageal damage. Estrogen-dependent Nrf2 expression might contribute to protection against the development of gastroesophageal reflux disease in females.

Protective effect of nuclear factor E2-related factor 2 on inflammatory cytokine response to brominated diphenyl ether-47 in the HTR-8/SVneo human first trimester extravillous trophoblast cell line.

PubMed

Park, Hae-Ryung; Loch-Caruso, Rita

2014-11-15

Polybrominated diphenyl ethers (PBDEs) are widely used flame retardants, and BDE-47 is a prevalent PBDE congener detected in human tissues. Exposure to PBDEs has been linked to adverse pregnancy outcomes in humans. Although the underlying mechanisms of adverse birth outcomes are poorly understood, critical roles for oxidative stress and inflammation are implicated. The present study investigated antioxidant responses in a human extravillous trophoblast cell line, HTR-8/SVneo, and examined the role of nuclear factor E2-related factor 2 (Nrf2), an antioxidative transcription factor, in BDE-47-induced inflammatory responses in the cells. Treatment of HTR-8/SVneo cells with 5, 10, 15, and 20μM BDE-47 for 24h increased intracellular glutathione (GSH) levels compared to solvent control. Treatment of HTR-8/SVneo cells with 20μM BDE-47 for 24h induced the antioxidant response element (ARE) activity, indicating Nrf2 transactivation by BDE-47 treatment, and resulted in differential expression of redox-sensitive genes compared to solvent control. Pretreatment with tert-butyl hydroquinone (tBHQ) or sulforaphane, known Nrf2 inducers, reduced BDE-47-stimulated IL-6 release with increased ARE reporter activity, reduced nuclear factor kappa B (NF-κB) reporter activity, increased GSH production, and stimulated expression of antioxidant genes compared to non-Nrf2 inducer pretreated groups, suggesting that Nrf2 may play a protective role against BDE-47-mediated inflammatory responses in HTR-8/SVneo cells. These results suggest that Nrf2 activation significantly attenuated BDE-47-induced IL-6 release by augmentation of cellular antioxidative system via upregulation of Nrf2 signaling pathways, and that Nrf2 induction may be a potential therapeutic target to reduce adverse pregnancy outcomes associated with toxicant-induced oxidative stress and inflammation. Copyright © 2014 Elsevier Inc. All rights reserved.

The Crosstalk Between Nrf2 and AMPK Signal Pathways Is Important for the Anti-Inflammatory Effect of Berberine in LPS-Stimulated Macrophages and Endotoxin-Shocked Mice

PubMed Central

Mo, Chunfen; Wang, Ling; Zhang, Jie; Numazawa, Satoshi; Tang, Hong; Tang, Xiaoqiang; Han, XiaoJuan; Li, Junhong; Yang, Ming; Wang, Zhe; Wei, Dandan

2014-01-01

Abstract Aims: The response of AMP-activated protein kinase (AMPK) to oxidative stress has been recently reported but the downstream signals of this response are largely unknown. Meanwhile, the upstream events for the activation of nuclear factor erythroid-2-related factor-2 (Nrf2), a critical transcriptional activator for antioxidative responses, remain unclear. In the present study, we investigated the relationship between AMPK and Nrf2 signal pathways in lipopolysaccharide (LPS)-triggered inflammatory system, in which berberine (BBR), a known AMPK activator, was used for inflammation suppression. Results and Innovation: In inflammatory macrophages, BBR attenuated LPS-induced expression of inflammatory genes (inducible nitric oxide synthase [iNOS], cyclooxygenase-2 [COX2], interleukin [IL]-6), and the generation of nitric oxide and reactive oxygen species, but increased the transcription of Nrf2-targeted antioxidative genes (NADPH quinone oxidoreductase-1 [NQO-1], heme oxygenase-1 [HO-1]), as well as the nuclear localization and phosphorylation of Nrf2 protein. Importantly, we found BBR-induced activation of Nrf2 is AMPK-dependent, as either pharmacologically or genetically inactivating AMPK blocked the activation of Nrf2. Consistent with in vitro experiments, BBR down-regulated the expression of proinflammatory genes but upregulated those of Nrf2-targeted genes in lungs of LPS-injected mice, and these effects were attenuated in Nrf2-deficient mice. Moreover, the effect of BBR on survival time extension and plasma redox regulation in endotoxin-shocked mice was largely weakened when Nrf2-depleted. Conclusions: Our results demonstrate convergence between AMPK and Nrf2 pathways and this intersection is essential for anti-inflammatory effect of BBR in LPS-stimulated macrophages and endotoxin-shocked mice. Uncovering this intersection is significant for understanding the relationship between energy homeostasis and antioxidative responses and may be beneficial for developing new therapeutic strategies against inflammatory diseases. Antioxid. Redox Signal. 20, 574–588. PMID:23875776

Antioxidants for Healthy Skin: The Emerging Role of Aryl Hydrocarbon Receptors and Nuclear Factor-Erythroid 2-Related Factor-2

PubMed Central

Furue, Masutaka; Uchi, Hiroshi; Mitoma, Chikage; Hashimoto-Hachiya, Akiko; Chiba, Takahito; Ito, Takamichi; Nakahara, Takeshi; Tsuji, Gaku

2017-01-01

Skin is the outermost part of the body and is, thus, inevitably exposed to UV rays and environmental pollutants. Oxidative stress by these hazardous factors accelerates skin aging and induces skin inflammation and carcinogenesis. Aryl hydrocarbon receptors (AHRs) are chemical sensors that are abundantly expressed in epidermal keratinocytes and mediate the production of reactive oxygen species. To neutralize or minimize oxidative stress, the keratinocytes also express nuclear factor-erythroid 2-related factor-2 (NRF2), which is a master switch for antioxidant signaling. Notably, there is fine-tuned crosstalk between AHR and NRF2, which mutually increase or decrease their activation states. Many NRF2-mediated antioxidant phytochemicals are capable of up- and downmodulating AHR signaling. The precise mechanisms by which these phytochemicals differentially affect the AHR and NRF2 system remain largely unknown and warrant future investigation. PMID:28273792

Antioxidants for Healthy Skin: The Emerging Role of Aryl Hydrocarbon Receptors and Nuclear Factor-Erythroid 2-Related Factor-2.

PubMed

Furue, Masutaka; Uchi, Hiroshi; Mitoma, Chikage; Hashimoto-Hachiya, Akiko; Chiba, Takahito; Ito, Takamichi; Nakahara, Takeshi; Tsuji, Gaku

2017-03-03

Skin is the outermost part of the body and is, thus, inevitably exposed to UV rays and environmental pollutants. Oxidative stress by these hazardous factors accelerates skin aging and induces skin inflammation and carcinogenesis. Aryl hydrocarbon receptors (AHRs) are chemical sensors that are abundantly expressed in epidermal keratinocytes and mediate the production of reactive oxygen species. To neutralize or minimize oxidative stress, the keratinocytes also express nuclear factor-erythroid 2-related factor-2 (NRF2), which is a master switch for antioxidant signaling. Notably, there is fine-tuned crosstalk between AHR and NRF2, which mutually increase or decrease their activation states. Many NRF2-mediated antioxidant phytochemicals are capable of up- and downmodulating AHR signaling. The precise mechanisms by which these phytochemicals differentially affect the AHR and NRF2 system remain largely unknown and warrant future investigation.

Are Astrocytes the Predominant Cell Type for Activation of Nrf2 in Aging and Neurodegeneration?

PubMed Central

2017-01-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcription factor that regulates hundreds of antioxidant genes, and is activated in response to oxidative stress. Given that many neurodegenerative diseases including Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis, Huntington’s disease and multiple sclerosis are characterised by oxidative stress, Nrf2 is commonly activated in these diseases. Evidence demonstrates that Nrf2 activity is repressed in neurons in vitro, and only cultured astrocytes respond strongly to Nrf2 inducers, leading to the interpretation that Nrf2 signalling is largely restricted to astrocytes. However, Nrf2 activity can be observed in neurons in post-mortem brain tissue and animal models of disease. Thus this interpretation may be false, and a detailed analysis of the cell type expression of Nrf2 in neurodegenerative diseases is required. This review describes the evidence for Nrf2 activation in each cell type in prominent neurodegenerative diseases and normal aging in human brain and animal models of neurodegeneration, the response to pharmacological and genetic modulation of Nrf2, and clinical trials involving Nrf2-modifying drugs. PMID:28820437

Update on NRF Measurements on ^237Np for National Security and Safeguards Applications

NASA Astrophysics Data System (ADS)

Angell, C. T.; Joshi, T.; Yee, R.; Swanberg, E.; Norman, E. B.; Kulp, W. D.; Warren, G.; Hicks, C. L., Jr.; Korbly, S.; Klimenko, A.; Wilson, C.; Bray, T. H.; Copping, R.; Shuh, D. K.

2010-11-01

Nuclear resonance fluorescence (NRF) uses γ rays to excite nuclear levels and measure their properties. This provides a unique isotopic signature, and can be used to identify and assay material. This is particularly important for applications that detect the smuggling of nuclear material or the diversion of fissile material for covert weapon programs, both of which present grave risks to world security. ^237Np presents significant safeguard challenges; it is fissile yet currently has fewer safeguard restrictions potentially making it an attractive material for covert weapon programs. This talk will present the final results of two measurements of NRF on ^237Np using a bremsstrahlung photon source. 15 NRF states have been identified between 1.5 and 2.5 MeV excitation energy.

Distinct Nrf2 Signaling Mechanisms of Fumaric Acid Esters and Their Role in Neuroprotection against 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine-Induced Experimental Parkinson's-Like Disease.

PubMed

Ahuja, Manuj; Ammal Kaidery, Navneet; Yang, Lichuan; Calingasan, Noel; Smirnova, Natalya; Gaisin, Arsen; Gaisina, Irina N; Gazaryan, Irina; Hushpulian, Dmitry M; Kaddour-Djebbar, Ismail; Bollag, Wendy B; Morgan, John C; Ratan, Rajiv R; Starkov, Anatoly A; Beal, M Flint; Thomas, Bobby

2016-06-08

A promising approach to neurotherapeutics involves activating the nuclear-factor-E2-related factor 2 (Nrf2)/antioxidant response element signaling, which regulates expression of antioxidant, anti-inflammatory, and cytoprotective genes. Tecfidera, a putative Nrf2 activator, is an oral formulation of dimethylfumarate (DMF) used to treat multiple sclerosis. We compared the effects of DMF and its bioactive metabolite monomethylfumarate (MMF) on Nrf2 signaling and their ability to block 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced experimental Parkinson's disease (PD). We show that in vitro DMF and MMF activate the Nrf2 pathway via S-alkylation of the Nrf2 inhibitor Keap1 and by causing nuclear exit of the Nrf2 repressor Bach1. Nrf2 activation by DMF but not MMF was associated with depletion of glutathione, decreased cell viability, and inhibition of mitochondrial oxygen consumption and glycolysis rates in a dose-dependent manner, whereas MMF increased these activities in vitro However, both DMF and MMF upregulated mitochondrial biogenesis in vitro in an Nrf2-dependent manner. Despite the in vitro differences, both DMF and MMF exerted similar neuroprotective effects and blocked MPTP neurotoxicity in wild-type but not in Nrf2 null mice. Our data suggest that DMF and MMF exhibit neuroprotective effects against MPTP neurotoxicity because of their distinct Nrf2-mediated antioxidant, anti-inflammatory, and mitochondrial functional/biogenetic effects, but MMF does so without depleting glutathione and inhibiting mitochondrial and glycolytic functions. Given that oxidative damage, neuroinflammation, and mitochondrial dysfunction are all implicated in PD pathogenesis, our results provide preclinical evidence for the development of MMF rather than DMF as a novel PD therapeutic. Almost two centuries since its first description by James Parkinson, Parkinson's disease (PD) remains an incurable disease with limited symptomatic treatment. The current study provides preclinical evidence that a Food and Drug Administration-approved drug, dimethylfumarate (DMF), and its metabolite monomethylfumarate (MMF) can block nigrostriatal dopaminergic neurodegeneration in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mouse model of PD. We elucidated mechanisms by which DMF and its active metabolite MMF activates the redox-sensitive transcription factor nuclear-factor-E2-related factor 2 (Nrf2) to upregulate antioxidant, anti-inflammatory, mitochondrial biosynthetic and cytoprotective genes to render neuroprotection via distinct S-alkylating properties and depletion of glutathione. Our data suggest that targeting Nrf2-mediated gene transcription using MMF rather than DMF is a promising approach to block oxidative stress, neuroinflammation, and mitochondrial dysfunction for therapeutic intervention in PD while minimizing side effects. Copyright © 2016 the authors 0270-6474/16/366333-20$15.00/0.

Identifying panaxynol, a natural activator of nuclear factor erythroid-2 related factor 2 (Nrf2) from American ginseng as a suppressor of inflamed macrophage-induced cardiomyocyte hypertrophy

PubMed Central

Qu, Chen; Li, Bin; Lai, Yimu; Li, Hechu; Windust, Anthony; Hofseth, Lorne J.; Nagarkatti, Mitzi; Nagarkatti, Prakash; Wang, Xing Li; Tang, Dongqi; Janicki, Joseph S.; Tian, Xingsong; Cui, Taixing

2015-01-01

Ethnopharmacological relevance American ginseng is capable of ameliorating cardiac dysfunction and activating Nrf2, a master regulator of antioxidant defense, in the heart. This study was designed to isolate compounds from American ginseng and to determine those responsible for the Nrf2-mediated resolution of inflamed macrophage-induced cardiomyocyte hypertrophy. Materials and methods A standardized crude extract of American ginseng was supplied by the National Research Council of Canada, Institute for National Measurement Standards. A bioassay-based fractionization of American ginseng was performed to identify the putative substances which could activate Nrf2-mediated suppression of pro-inflammatory cytokine expression in macrophages and macrophage-mediated pro-hypertrophic growth in cardiomyocytes. Results A hexane fraction of an anti-inflammatory crude extract of American ginseng was found to be most effective in suppressing the inflammatory responses in macrophages. Preparative, reverse-phase HPLC and a comparative analysis by analytical scale LC–UV/MS revealed the hexane fraction contains predominantly C17 polyacetylenes and linolenic acid. Panaxynol, one of the major polyacetylenes, was found to be a potent Nrf2 activator. Panaxynol posttranscriptionally activated Nrf2 by inhibiting Kelch-like ECH-associated protein (Keap) 1-mediated degradation without affecting the binding of Keap1 and Nrf2. Moreover, panaxynol suppressed a selected set of cytokine expression via the activation of Nrf2 while minimally regulating nuclear factor-kappa B (NF-κB)-mediated cytokine expression in macrophages. It also dramatically inhibited the inflamed macrophage-mediated cardiomyocyte death and hypertrophy by activating Nrf2 in macrophages. Conclusions These results demonstrate that American ginseng-derived panaxynol is a specific Nrf2 activator and panaxynol-activated Nrf2 signaling is at least partly responsible for American ginseng-induced health benefit in the heart. PMID:25882312

Diesel Exhaust Particulate Extracts Inhibit Transcription of Nuclear Respiratory Factor-1 and Cell Viability in Human Umbilical Vein Endothelial Cells

PubMed Central

Mattingly, Kathleen A.; Klinge, Carolyn M.

2011-01-01

Endothelial dysfunction precedes cardiovascular disease and is accompanied by mitochondrial dysfunction. Here we tested the hypothesis that diesel exhaust particulate extracts (DEPEs), prepared from a truck run at different speeds and engine loads, would inhibit genomic estrogen receptor activation of nuclear respiratory factor-1 (NRF-1) transcription in human umbilical vein endothelial cells (HUVECs). Additionally, we examined how DEPEs affect NRF-1 regulated TFAM expression and, in turn, Tfam-regulated mtDNA-encoded cytochrome c oxidase subunit I (COI, MTCO1) and NADH dehydrogenase subunit I (NDI) expression as well as cell proliferation and viability. We report that 17β-estradiol (E2), 4-hydroxytamoxifen (4-OHT), and raloxifene increased NRF-1 transcription in HUVECs in an ER-dependent manner. DEPEs inhibited NRF-1 transcription and this suppression was not ablated by concomitant treatment with E2, 4-OHT, or raloxifene, indicating that the effect was not due to inhibition of ER activity. While E2 increased HUVEC proliferation and viability, DEPEs inhibited viability but not proliferation. Resveratrol increased NRF-1 transcription in an ER-dependent manner in HUVECs, and ablated DEPE inhibition of basal NRF-1 expression. Given that NRF-1 is a key nuclear transcription factor regulating genes involved in mitochondrial activity and biogenesis, these data suggest that DEPEs may adversely affect mitochondrial function leading to endothelial dysfunction and resveratrol may block these effects. PMID:22105178

Protective Role of Nuclear Factor E2-Related Factor 2 against Acute Oxidative Stress-Induced Pancreatic β-Cell Damage

PubMed Central

Fu, Jingqi; Zheng, Hongzhi; Wang, Huihui; Yang, Bei; Zhao, Rui; Lu, Chunwei; Liu, Zhiyuan; Hou, Yongyong; Xu, Yuanyuan; Zhang, Qiang; Qu, Weidong; Pi, Jingbo

2015-01-01

Oxidative stress is implicated in the pathogenesis of pancreatic β-cell dysfunction that occurs in both type 1 and type 2 diabetes. Nuclear factor E2-related factor 2 (NRF2) is a master regulator in the cellular adaptive response to oxidative stress. The present study found that MIN6 β-cells with stable knockdown of Nrf2 (Nrf2-KD) and islets isolated from Nrf2-knockout mice expressed substantially reduced levels of antioxidant enzymes in response to a variety of stressors. In scramble MIN6 cells or wild-type islets, acute exposure to oxidative stressors, including hydrogen peroxide (H2O2) and S-nitroso-N-acetylpenicillamine, resulted in cell damage as determined by decrease in cell viability, reduced ATP content, morphology changes of islets, and/or alterations of apoptotic biomarkers in a concentration- and/or time-dependent manner. In contrast, silencing of Nrf2 sensitized MIN6 cells or islets to the damage. In addition, pretreatment of MIN6 β-cells with NRF2 activators, including CDDO-Im, dimethyl fumarate (DMF), and tert-butylhydroquinone (tBHQ), protected the cells from high levels of H2O2-induced cell damage. Given that reactive oxygen species (ROS) are involved in regulating glucose-stimulated insulin secretion (GSIS) and persistent activation of NRF2 blunts glucose-triggered ROS signaling and GSIS, the present study highlights the distinct roles that NRF2 may play in pancreatic β-cell dysfunction that occurs in different stages of diabetes. PMID:25949772

Interplay between VEGF and Nrf2 regulates angiogenesis due to intracranial venous hypertension.

PubMed

Li, Liwen; Pan, Hao; Wang, Handong; Li, Xiang; Bu, Xiaomin; Wang, Qiang; Gao, Yongyue; Wen, Guodao; Zhou, Yali; Cong, Zixiang; Yang, Youqing; Tang, Chao; Liu, Zhengwei

2016-11-21

Venous hypertension(VH) plays an important role in the pathogenesis of cerebral arteriovenous malformations (AVMs) and is closely associated with the HIF-1α/VEGF signaling pathway. Nuclear factor erythroid 2-related factor 2(Nrf2) significantly influences angiogenesis; however, the interplay between Nrf2 and VEGF under VH in brain AVMs remains unclear. Therefore, our study aimed to investigate the interplay between Nrf2 and VEGF due to VH in brain AVMs. Immunohistochemistry indicated that Nrf2 and VEGF were highly expressed in human brain AVM tissues. In vivo, we established a VH model in both wild-type (WT) and siRNA-mediated Nrf2 knockdown rats. VH significantly increased the expression of Nrf2 and VEGF. Loss of Nrf2 markedly inhibited the upregulation of VEGF, as determined by Western blot analysis and qRT-PCR. In vitro, primary brain microvascular endothelial cells (BMECs) were isolated from WT and Nrf2 -/- mice, and a VEGF-Nrf2 positive feed-back loop was observed in BMECs. By trans well assay and angiogenesis assay, Nrf2 knockout significantly inhibited the migration and vascular tube formation of BMECs. These findings suggest that the interplay between Nrf2 and VEGF can contribute to VH-induced angiogenesis in brain AVMs pathogenesis.

Nrf2 protects against oxidative stress induced by SiO2 nanoparticles.

PubMed

Liu, Wei; Hu, Tao; Zhou, Li; Wu, Desheng; Huang, Xinfeng; Ren, Xiaohu; Lv, Yuan; Hong, Wenxu; Huang, Guanqin; Lin, Zequn; Liu, Jianjun

2017-10-01

The aim of our study was to explore the role of nuclear factor erythroid 2 (NF-E2)-related factor 2 (Nrf2) on the exposure of SiO 2 nanoparticles (NPs) and its influence. To understand the mechanism of NP-induced oxidative stress, the involvement of oxidative-stress-responding transcription factors and the Nrf2/antioxidant reactive element (ARE) signaling pathway in the toxicity of SiO 2 NPs' exposure was investigated via in vivo and in vitro models. A549 cells showed a significant cytotoxic effect while A549-shNrf2 cells showed decreased cell viability after nm-SiO 2 exposure. SiO 2 NPs' exposure activated the Nrf2/ARE signaling pathway. Nrf2 -/- exposed mice showed increased reactive oxygen species, 8-hydroxyl deoxyguanosine level and decreased total antioxidant capacity. Nrf2/ARE signaling pathway activation disrupted, leading inhibition of heme oxygenase-1 and upregulation of PKR-like endoplasmic-reticulum-regulated kinase. Our findings suggested that Nrf2 could protect against oxidative stress induced by SiO 2 NPs, and the Nrf2/ARE pathway might be involved in mild-to-moderate SiO 2 NP-induced oxidative stress that was evident from dampened activity of Nrf2.

Neuroprotective efficacy of naringin on 3-nitropropionic acid-induced mitochondrial dysfunction through the modulation of Nrf2 signaling pathway in PC12 cells.

PubMed

Kulasekaran, Gopinath; Ganapasam, Sudhandiran

2015-11-01

Oxidative stress and mitochondrial dysfunction are implicated in neuronal apoptosis associated with Huntington's disease. Naringin is the flavanone present in grapefruit and related citrus species possess diverse pharmacological and therapeutic properties including antioxidant, anti-apoptotic, and neuroprotective properties. The aim of this study was to investigate the protective effect of naringin on 3-nitropropionic acid (3-NP)-induced neurotoxicity in pheochromocytoma cells (PC12) cells and to explore its mechanism of action. Naringin protects PC12 cells from 3-NP neurotoxicity, as evaluated the by cell viability assays. The lactate dehydrogenase release was decreased upon naringin treatment in 3-NP-induced PC12 cells. Naringin treatment enhances the antioxidant defense by increasing the activities of enzymatic antioxidants and the level of reduced glutathione. The increase in levels of reactive oxygen species and lipid peroxidation induced by 3-NP were significantly decreased by naringin. PC12 cells induced with 3-NP showed decrease in the mitochondrial membrane potential and mitochondrial respiratory complex enzymes, succinate dehydrogenase and cytochrome c oxidase activities, and it was significantly altered to near normal upon naringin treatment. Naringin reduced the 3-NP-induced apoptosis through the modulation in expressions of B-cell lymphoma 2 and Bcl-2-associated X protein. Further, naringin enhances the nuclear translocation of Nrf2 and induces the quinone oxidoreductase-1 and Heme oxygenase-1 expressions through the phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathway. Taken together, the above findings suggest that naringin augments cellular antioxidant defense capacity and reduces the 3-NP-induced neurotoxicity in PC12 cells through the PI-3K/Akt-dependent Nrf2 activation in PC12 cells.

The nuclear factor (erythroid-derived 2)-like 2 (NRF2) antioxidant response promotes melanocyte viability and reduces toxicity of the vitiligo-inducing phenol monobenzone.

PubMed

Arowojolu, Omotayo A; Orlow, Seth J; Elbuluk, Nada; Manga, Prashiela

2017-07-01

Vitiligo, characterised by progressive melanocyte death, can be initiated by exposure to vitiligo-inducing phenols (VIPs). VIPs generate oxidative stress in melanocytes and activate the master antioxidant regulator NRF2. While NRF2-regulated antioxidants are reported to protect melanocytes from oxidative stress, the role of NRF2 in the melanocyte response to monobenzone, a clinically relevant VIP, has not been characterised. We hypothesised that activation of NRF2 may protect melanocytes from monobenzone-induced toxicity. We observed that knockdown of NRF2 or NRF2-regulated antioxidants NQO1 and PRDX6 reduced melanocyte viability, but not viability of keratinocytes and fibroblasts, suggesting that melanocytes were preferentially dependent upon NRF2 activity for growth compared to other cutaneous cells. Furthermore, melanocytes activated the NRF2 response following monobenzone exposure and constitutive NRF2 activation reduced monobenzone toxicity, supporting NRF2's role in the melanocyte stress response. In contrast, melanocytes from individuals with vitiligo (vitiligo melanocytes) did not activate the NRF2 response as efficiently. Dimethyl fumarate-mediated NRF2 activation protected normal and vitiligo melanocytes against monobenzone-induced toxicity. Given the contribution of oxidant-antioxidant imbalance in vitiligo, modulation of this pathway may be of therapeutic interest. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Nrf2-peroxiredoxin I axis in polymorphous adenocarcinoma is associated with low matrix metalloproteinase 2 level.

PubMed

Brod, J M; Demasi, Ana Paula Dias; Montalli, V A; Teixeira, L N; Furuse, C; Aguiar, M C; Soares, A B; Sperandio, M; Araujo, V C

2017-12-01

Polymorphous adenocarcinoma (PAC) is a malignant epithelial neoplasm that affects almost exclusively the minor salivary glands, generally described as having a relatively good prognosis. Aberrant nuclear factor erythroid 2 (NF-E2)-related factor (Nrf2) activation in tumor cells has been associated with induction of antioxidant enzymes, such as peroxiredoxin I (Prx I) and increased matrix metalloproteinase (MMP) expression. In this context, the aim of the present study was to evaluate the expression of Nrf2 and correlate it with Prx I and MMP-2 secretion in PAC. Thirty-one cases of PAC from oral biopsies were selected and immunohistochemically analyzed for Nrf2 and Prx I. MMP-2 quantification was performed on primary cell cultures derived from PAC. Oral squamous cell carcinoma (OSCC) cell cultures were used as control. A high immunoexpression of Nrf2 was observed in both the cytoplasm and the nucleus of neoplastic cells from PAC. Nuclear staining for Nrf2 suggested its activation in the majority of the PAC cells, which was confirmed by the high expression of its target gene, Prx I. Quantification of MMP-2 secretion showed lower levels in PAC cell cultures when compared to OSCC cell cultures (p < 0.05). In conclusion, although Nrf2 overexpression has been frequently associated with high-grade malignancies, such relationship is not infallible and, in fact, the opposite may occur in low-grade tumors, such as PAC of minor salivary glands.

Effect of cellular ubiquitin levels on the regulation of oxidative stress response and proteasome function via Nrf1.

PubMed

Lee, Donghee; Ryu, Kwon-Yul

2017-04-01

The polyubiquitin genes Ubb and Ubc are upregulated under oxidative stress induced by arsenite [As(III)]. However, the role of ubiquitin (Ub) under As(III) exposure is not known in detail. In a previous study, we showed that the reduced viability observed in Ubc -/- mouse embryonic fibroblasts under As(III) exposure was not due to dysregulation of the Nrf2-Keap1 pathway, which prompted us to investigate another NFE2 family protein, nuclear factor erythroid 2-related factor 1 (Nrf1). In this study, we found that Ub deficiency due to Ubc knockdown in N2a cells reduced cell viability and proteasome activity under As(III) exposure. Furthermore, mRNA levels of the proteasome subunit Psma1 were also reduced. In addition, Ub deficiency led to the nuclear accumulation of the p65 isoform of Nrf1 under As(III) exposure. Interestingly, the overexpression of p65-Nrf1 recapitulated the phenotypes of Ub-deficient N2a cells under As(III) exposure. On the other hand, Nrf1 knockdown suppressed the death of Ub-deficient N2a cells upon exposure to As(III). Therefore, the levels of p65-Nrf1 may play an important role in the maintenance of cell viability under oxidative stress induced by As(III). Copyright © 2017 Elsevier Inc. All rights reserved.

Nrf2 regulates cellular behaviors and Notch signaling in oral squamous cell carcinoma cells.

PubMed

Fan, Hong; Paiboonrungruan, Chorlada; Zhang, Xinyan; Prigge, Justin R; Schmidt, Edward E; Sun, Zheng; Chen, Xiaoxin

2017-11-04

Oxidative stress is known to play a pivotal role in the development of oral squamous cell carcinoma (OSCC). We have demonstrated that activation of the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway has chemopreventive effects against oxidative stress-associated OSCC. However, Nrf2 have dual roles in cancer development; while it prevents carcinogenesis of normal cells, hyperactive Nrf2 also promotes the survival of cancer cells. This study is aimed to understand the function of Nrf2 in regulating cellular behaviors of OSCC cells, and the potential mechanisms through which Nrf2 facilitates OSCC. We established the Nrf2-overexpressing and Nrf2-knockdown OSCC cell lines, and examined the function of Nrf2 in regulating cell proliferation, migration, invasion, cell cycle and colony formation. Our data showed that Nrf2 overexpression promoted cancer phenotypes in OSCC cells, whereas Nrf2 silencing inhibited these phenotypes. In addition, Nrf2 positively regulated Notch signaling pathway in OSCC cells in vitro. Consistent with this observation, Nrf2 activation in Keap1 -/- mice resulted in not only hyperproliferation of squamous epithelial cells in mouse tongue as evidenced by increased expression of PCNA, but also activation of Notch signaling in these cells as evidenced by increased expression of NICD1 and Hes1. In conclusion, Nrf2 regulates cancer behaviors and Notch signaling in OSCC cells. Copyright © 2017 Elsevier Inc. All rights reserved.

Selenium antagonizes cadmium-induced apoptosis in chicken spleen but not involving Nrf2-regulated antioxidant response.

PubMed

Chen, Menghao; Li, Xiaojing; Fan, Ruifeng; Cao, Changyu; Yao, Haidong; Xu, Shiwen

2017-11-01

The nuclear transcription factor NF-E2-related factor 2 (Nrf2) binds to antioxidant response elements (AREs) and is involved in the regulation of genes participated in defending cells against oxidative damage, which have been confirmed in animal models. Selenium (Se), known as an important element in the regulation of antioxidant activity, can antagonize Cadmium (Cd) toxicity in birds. However, the role of Nrf2 in selenium-cadmium interaction has not been reported in birds. To further explore the mechanism of selenium attenuating spleen toxicity induced by cadmium in chickens, cadmium chloride (CdCl 2 , 150mg/kg) and sodium selenite (Na 2 SeO 3 , 2mg/kg) were co-administrated or individually administered in the diet of chickens for 90 days. The results showed that Cd exposure increased the level of hydrogen peroxide (H 2 O 2 ) and malondialdehyde (MDA) and decreased the antioxidant enzyme activities, including superoxide dismutase (SOD), glutathione peroxidase (Gpx), total antioxidative capacity (T-AOC), catalase (CAT). Cd exposure increased obviously nuclear accumulation of Nrf2, and the expression of Nrf2 downstream heme oxygenase-1 (HO-1) and NAD(P)H: quinine oxidoreductase 1 (NQO1), reduced the expression of Kelch-like ECH-associated protein (keap1), Gpx-1 and thioredoxin reductase-1 (TrxR1). In addition, Cd induced the increase of bak, caspase9, p53, Cyt c mRNA levels, increased bax/bcl-2 ratio, increased caspase3 mRNA and protein levels. Selenium treatment reduced the accumulation of Cd in the spleen, attenuates Cd-induced Nrf2 nuclear accumulation, enhanced antioxidant enzyme activities, ameliorated Cd-induced oxidative stress and apoptosis in the spleen. In summary, our results demonstrate that Se ameliorated spleen toxicity induced by cadmium by modulating the antioxidant system, independently of Nrf2-regulated antioxidant response pathway. Copyright © 2017 Elsevier Inc. All rights reserved.

Sirt1 activation prevents anti-Thy 1.1 mesangial proliferative glomerulonephritis in the rat through the Nrf2/ARE pathway.

PubMed

Huang, Kaipeng; Li, Ruiming; Wei, Wentao

2018-08-05

Mesangial proliferative glomerulonephritis (MsPGN) is characterized by glomerular mesangial cells proliferation and extracellular matrix deposition in mesangial area, which develop into glomerulosclerosis. Both silent information regulator 2-related protein 1 (Sirt1) and nuclear factor erythroid 2-related factor 2/anti-oxidant response element (Nrf2/ARE) pathway had remarkable renoprotective effects. However, whether Sirt1 and Nrf2/ARE pathway can regulate the pathological process of MsPGN remains unknown. Here, we found that Sirt1 activation by SRT1720 decreased mesangial hypercellularity and mesangial matrix areas, reduced renal Col4 and α-SMA expressions, lowered 24 h proteinuria, and eventually reduced FN and TGF-β1 expressions in rats received anti-Thy 1.1 IgG. Further study showed that SRT1720 markedly enhanced the activity of Nrf2/ARE pathway including promoting the nuclear content and ARE-binding ability of Nrf2, elevating the protein levels of HO-1 and SOD1, two target genes of Nrf2, which eventually increased total SOD activity and decreased malondialdehyde level in the kidney tissues of experimental anti-Thy 1.1 MsPGN rats. Taken together, Sirt1 prevented the pathological process of experimental anti-Thy 1.1 MsPGN through promoting the activation of Nrf2/ARE pathway, which warrants further elucidation. Sirt1 might be a potential therapeutic target for treating MsPGN. Copyright © 2018 Elsevier B.V. All rights reserved.

Metallothionein plays a prominent role in the prevention of diabetic nephropathy by sulforaphane via up-regulation of Nrf2

PubMed Central

Wu, Hao; Kong, Lili; Cheng, Yanli; Zhang, Zhiguo; Wang, Yangwei; Lou, Manyu; Tan, Yi; Chen, Xiangmei; Miao, Lining; Cai, Lu

2015-01-01

Sulforaphane (SFN) prevents diabetic nephropathy (DN) in type 1 diabetes via up-regulation of nuclear factor (erythroid-derived 2)-like 2 (Nrf2). However, it has not been addressed whether SFN also prevents DN from type 2 diabetes or which Nrf2 downstream gene(s) play(s) the key role in SFN renal protection. Here we investigated whether Nrf2 is required for SFN protection against type 2 diabetes-induced DN and whether metallothionein (MT) is an Nrf2 downstream antioxidant using Nrf2 knockout (Nrf2-null) mice. In addition, MT knockout mice were used to further verify if MT is indispensable for SFN protection against DN. Diabetes-increased albuminuria, renal fibrosis, and inflammation were significantly prevented by SFN, and Nrf2 and MT expression was increased. However, SFN renal protection was completely lost in Nrf2-null diabetic mice, confirming the pivotal role of Nrf2 in SFN protection from type 2 diabetes-induced DN. Moreover, SFN failed to up-regulate MT in the absence of Nrf2, suggesting that MT is an Nrf2 downstream antioxidant. MT deletion resulted in a partial, but significant attenuation of SFN renal protection from type 2 diabetes, demonstrating a partial requirement for MT for SFN renal protection. Therefore, the present study demonstrates for the first time that as an Nrf2 downstream antioxidant, MT plays an important, though partial, role in mediating SFN renal protection from type 2 diabetes. PMID:26415026

Long-term Autophagy and Nrf2 Signaling in the Hippocampi of Developing Mice after Carbon Ion Exposure

NASA Astrophysics Data System (ADS)

Ye, Fei; Zhao, Ting; Liu, Xiongxiong; Jin, Xiaodong; Liu, Xinguo; Wang, Tieshan; Li, Qiang

2015-12-01

To explore charged particle radiation-induced long-term hippocampus damage, we investigated the expression of autophagy and antioxidant Nrf2 signaling-related proteins in the mouse hippocampus after carbon ion radiation. Heads of immature female Balb/c mice were irradiated with carbon ions of different LETs at various doses. Behavioral tests were performed on the mice after maturation. Acute and chronic expression of LC3-II, p62/SQSTM1, nuclear Nrf2, activated caspase-3 and the Bax/Bcl-2 ratio were measured in the hippocampi. Secondary X-ray insult was adopted to amplify potential damages. Long-term behavioral changes were observed in high-LET carbon ion-irradiated mice. There were no differences in the rates of LC3-II induction and p62/SQSTM1 degradation compared to the control group regardless of whether the mice received the secondary X-ray insult. A high nuclear Nrf2 content and low apoptosis level in hippocampal cells subjected to secondary X-rays were observed for the mice exposed to relatively low-LET carbon ions. Therefore, carbon ion exposure in the immature mouse led to an LET-dependent behavioral change after maturation. Although autophagy was intact, the persistently high nuclear Nrf2 content in the hippocampus might account for the unchanged behavioral pattern in mice exposed to the relatively low-LET carbon ions and the subsequent increased radioresistance of the hippocampus.

Hawthorn (Crataegus oxyacantha L.) bark extract regulates antioxidant response element (ARE)-mediated enzyme expression via Nrf2 pathway activation in normal hepatocyte cell line.

PubMed

Krajka-Kuźniak, Violetta; Paluszczak, Jarosław; Oszmiański, Jan; Baer-Dubowska, Wanda

2014-04-01

Hawthorn (Crataegus oxyacantha L.), a plant used in traditional medicine, is a rich source of procyanidins which have been reported to exhibit antioxidant and anti-carcinogenic activity. In this study, we assessed the effect of hawthorn bark extract (HBE) on Nrf2 pathway activation in THLE-2 and HepG2 cells. Treatment with 1.1 µg/mL, 5.5 µg/mL and 11 µg/mL of HBE resulted in the translocation of Nrf2 from the cytosol to the nucleus in both cell lines; however, the accumulation of phosphorylated Nrf2 was observed only in THLE-2. Accordingly, treatment of cells with HBE was associated with an increase in the mRNA and protein level of such Nrf2-dependent genes as glutathione S-transferases (GSTA, GSTP, GSTM, GSTT), NAD(P)H:quinone oxidoreductase 1 (NQO1) and heme oxygenase-1 (HO-1) (0.2-1.1-fold change, p < 0.05), however, only in normal THLE-2 hepatocytes. The induction of NQO1 correlated with an increased level of p53 (0.21-0.42-fold change, p < 0.05). These effects may be related to induction of phosphorylation of upstream ERK and JNK kinases. Collectively, the results suggest that the Nrf2/ARE pathway may play an important role in the regulation of procyanidin-mediated antioxidant/detoxifying effects in hepatocytes, and this may explain the hepatoprotective and chemopreventive properties of these phytochemicals. Copyright © 2013 John Wiley & Sons, Ltd.

The mitochondria-targeted antioxidant MitoQ ameliorated tubular injury mediated by mitophagy in diabetic kidney disease via Nrf2/PINK1.

PubMed

Xiao, Li; Xu, Xiaoxuan; Zhang, Fan; Wang, Ming; Xu, Yan; Tang, Dan; Wang, Jiahui; Qin, Yan; Liu, Yu; Tang, Chengyuan; He, Liyu; Greka, Anna; Zhou, Zhiguang; Liu, Fuyou; Dong, Zheng; Sun, Lin

2017-04-01

Mitochondria play a crucial role in tubular injury in diabetic kidney disease (DKD). MitoQ is a mitochondria-targeted antioxidant that exerts protective effects in diabetic mice, but the mechanism underlying these effects is not clear. We demonstrated that mitochondrial abnormalities, such as defective mitophagy, mitochondrial reactive oxygen species (ROS) overexpression and mitochondrial fragmentation, occurred in the tubular cells of db/db mice, accompanied by reduced PINK and Parkin expression and increased apoptosis. These changes were partially reversed following an intraperitoneal injection of mitoQ. High glucose (HG) also induces deficient mitophagy, mitochondrial dysfunction and apoptosis in HK-2 cells, changes that were reversed by mitoQ. Moreover, mitoQ restored the expression, activity and translocation of HG-induced NF-E2-related factor 2 (Nrf2) and inhibited the expression of Kelch-like ECH-associated protein (Keap1), as well as the interaction between Nrf2 and Keap1. The reduced PINK and Parkin expression noted in HK-2 cells subjected to HG exposure was partially restored by mitoQ. This effect was abolished by Nrf2 siRNA and augmented by Keap1 siRNA. Transfection with Nrf2 siRNA or PINK siRNA in HK-2 cells exposed to HG conditions partially blocked the effects of mitoQ on mitophagy and tubular damage. These results suggest that mitoQ exerts beneficial effects on tubular injury in DKD via mitophagy and that mitochondrial quality control is mediated by Nrf2/PINK. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Induction of Heme Oxygenase-1 by Sodium 9-Hydroxyltanshinone IIA Sulfonate Derivative Contributes to Inhibit LPS-Mediated Inflammatory Response in Macrophages.

PubMed

Liu, Xin-Hua; Wang, Xi-Ling; Xin, Hong; Wu, Dan; Xin, Xiao-Ming; Miao, Lei; Zhang, Qiu-Yan; Zhou, Yang; Liu, Qian; Zhang, Qian; Zhu, Yi-Zhun

2015-01-01

Sodium 9-acetoxyltanshinone IIA sulfonate (ZY-1A4), a novel compound derived from sodium 9-hydroxyltanshinone IIA sulfonate, was synthesized with potential biological activities. This study aimed to explore the effects of ZY-1A4 on lipopolysaccharide (LPS)-triggered inflammatory response and the underlying mechanisms. Activation of RAW264.7 macrophages was induced by LPS. The effects of ZY-1A4 on inducible nitric oxide synthase (iNOS) expression, nitric oxide (NO) generation, nuclear factor-κB (NF-κB) activation, heme oxygenase-1 (HO-1) expression, and nuclear factor-erythroid 2 related factor 2 (Nrf2) pathway were evaluated to elucidate its underlying mechanisms on inflammatory responses. ZY-1A4 concentration-dependently reduced iNOS expression and NO production, and inhibited c-Jun-N-terminal kinase 1/2 (JNK1/2) phosphorylation and NF-κB activation in LPS-stimulated macrophages. In addition, ZY-1A4 concentration- and time-dependently induced HO-1 expression associated with degradation of Kelch-like ECH-associated protein 1 (Keap1) and nuclear translocation of Nrf2, while the effect of ZY-1A4 was abolished by a phosphoinositide 3-kinase (PI3K) inhibitor LY294002. Intriguingly, pharmacological inactivation of HO-1 with zinc protoporphyrin IX reversed anti-inflammatory effect of ZY- 1A4, but the anti-inflammatory effect of ZY-1A4 was largely mimicked by HO-1 by-products carbon monoxide and bilirubin. Furthermore, the inhibitory effect of ZY-1A4 on LPS-induced iNOS expression and NO release was abolished by HO-1 siRNA or LY294002. Our results demonstrated that ZY-1A4 suppressed LPS-induced iNOS expression and NO generation via modulation of NF-κB activation and HO-1 expression. This new finding might shed light to the prevention and therapy of cardiovascular diseases. © 2015 S. Karger AG, Basel.

Nuclear factor E2-related factor-2 has a differential impact on MCT1 and MCT4 lactate carrier expression in colonic epithelial cells: a condition favoring metabolic symbiosis between colorectal cancer and stromal cells.

PubMed

Diehl, K; Dinges, L-A; Helm, O; Ammar, N; Plundrich, D; Arlt, A; Röcken, C; Sebens, S; Schäfer, H

2018-01-04

Malignant tumors, such as colorectal cancer (CRC), are heterogeneous diseases characterized by distinct metabolic phenotypes. These include Warburg- and reverse Warburg phenotypes depending on differential distribution of the lactate carrier proteins monocarboxylate transporter-4 and -1 (MCT4 and MCT1). Here, we elucidated the role of the antioxidant transcription factor nuclear factor E2-related factor-2 (Nrf2) as the key regulator of cellular adaptation to inflammatory/environmental stress in shaping the metabolism toward a reverse Warburg phenotype in malignant and premalignant colonic epithelial cells. Immunohistochemistry of human CRC tissues revealed reciprocal expression of MCT1 and MCT4 in carcinoma and stroma cells, respectively, accompanied by strong epithelial Nrf2 activation. In colorectal tissue from inflammatory bowel disease patients, MCT1 and Nrf2 were coexpressed as well, relating to CD68+inflammatory infiltrates. Indirect coculture of human NCM460 colonocytes with M1- but not M2 macrophages induces MCT1 as well as G6PD, LDHB and TALDO expression, whereas MCT4 expression was decreased. Nrf2 knockdown or reactive oxygen species (ROS) scavenging blocked these coculture effects in NCM460 cells. Likewise, Nrf2 knockdown inhibited similar effects of tBHQ-mediated Nrf2 activation on NCM460 and HCT15 CRC cells. M1 coculture or Nrf2 activation/overexpression greatly altered the lactate uptake but not glucose uptake and mitochondrial activities in these cells, reflecting the reverse Warburg phenotype. Depending on MCT1-mediated lactate uptake, Nrf2 conferred protection from TRAIL-induced apoptosis in NCM460 and HCT15 cells. Moreover, metabolism-dependent clonal growth of HCT15 cells was induced by Nrf2-dependent activation of MCT1-driven lactate exchange. These findings indicate that Nrf2 has an impact on the metabolism already in premalignant colonic epithelial cells exposed to inflammatory M1 macrophages, an effect accompanied by growth and survival alterations. Favoring the reverse Warburg effect, these Nrf2-dependent alterations add to malignant transformation of the colonic epithelium.

Potential drugs which activate nuclear factor E2-related factor 2 signaling to prevent diabetic cardiovascular complications: A focus on fumaric acid esters.

PubMed

Zhou, Shanshan; Jin, Jingpeng; Bai, Tao; Sachleben, Leroy R; Cai, Lu; Zheng, Yang

2015-08-01

Diabetes and its cardiovascular complications have been a major public health issue. These complications are mainly attributable to a severe imbalance between free radical and reactive oxygen species production and the antioxidant defense systems. Nuclear factor E2-related factor 2 (Nrf2) is a transcription factor that controls the basal and inducible expression of a battery of antioxidant enzyme genes and other cyto-protective phase II detoxifying enzymes. As a result, Nrf2 has gained great attention as a promising drug target for preventing diabetic cardiovascular complications. And while animal studies have shown that several Nrf2 activators manifest a potential to efficiently prevent the diabetic complications, their use in humans has not been approved due to the lack of substantial evidence regarding safety and efficacy of the Nrf2 activation. We provide here a brief review of a few clinically-used drugs that can up-regulate Nrf2 with the potential of extending their usage to diabetic patients for the prevention of cardiovascular complications and conclude with a closer inspection of dimethyl fumarate and its mimic members. Copyright © 2015 Elsevier Inc. All rights reserved.

Nuclear factor-E2-related factor 2 is a major determinant of bile acid homeostasis in the liver and intestine

PubMed Central

Weerachayaphorn, Jittima; Mennone, Albert; Soroka, Carol J.; Harry, Kathy; Hagey, Lee R.; Kensler, Thomas W.

2012-01-01

The transcription factor nuclear factor-E2-related factor 2 (Nrf2) is a key regulator for induction of hepatic detoxification and antioxidant mechanisms, as well as for certain hepatobiliary transporters. To examine the role of Nrf2 in bile acid homeostasis and cholestasis, we assessed the determinants of bile secretion and bile acid synthesis and transport before and after bile duct ligation (BDL) in Nrf2−/− mice. Our findings indicate reduced rates of biliary bile acid and GSH excretion, higher levels of intrahepatic bile acids, and decreased expression of regulators of bile acid synthesis, Cyp7a1 and Cyp8b1, in Nrf2−/− compared with wild-type control mice. The mRNA expression of the bile acid transporters bile salt export pump (Bsep) and organic solute transporter (Ostα) were increased in the face of impaired expression of the multidrug resistance-associated proteins Mrp3 and Mrp4. Deletion of Nrf2 also decreased ileal apical sodium-dependent bile acid transporter (Asbt) expression, leading to reduced bile acid reabsorption and increased loss of bile acid in feces. Finally, when cholestasis is induced by BDL, liver injury was not different from that in wild-type BDL mice. These Nrf2−/− mice also had increased pregnane X receptor (Pxr) and Cyp3a11 mRNA expression in association with enhanced hepatic bile acid hydroxylation. In conclusion, this study finds that Nrf2 plays a major role in the regulation of bile acid homeostasis in the liver and intestine. Deletion of Nrf2 results in a cholestatic phenotype but does not augment liver injury following BDL. PMID:22345550

Enhanced sensitivity of A549 cells to the cytotoxic action of anticancer drugs via suppression of Nrf2 by procyanidins from Cinnamomi Cortex extract

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ohnuma, Tomokazu; Matsumoto, Takashi; Itoi, Ayano

Highlights: {yields} We found a novel inhibitor of Nrf2 known as a chemoresistance factor. {yields} Overexpressed Nrf2 in lung cancer cells was suppressed by Cinnamomi Cortex extract. {yields} Cytotoxic action of anticancer drugs in cells treated with the extract was enhanced. {yields} Procyanidin tetramers and pentamers were active components in suppressing Nrf2. -- Abstract: Nuclear factor-E2-related factor 2 (Nrf2) is an important cytoprotective transcription factor because Nrf2-regulated enzymes play a key role in antioxidant and detoxification processes. Recent studies have reported that lung cancer cells overexpressing Nrf2 exhibit increased resistance to chemotherapy. Suppression of overexpressed Nrf2 is needed for amore » new therapeutic approach against lung cancers. In the present study, we found that Cinnamomi Cortex extract (CCE) has an ability to suppress Nrf2-regulated enzyme activity and Nrf2 expression in human lung cancer A549 cells with high Nrf2 activity. Moreover, we demonstrated that CCE significantly enhances sensitivity of A549 cells to the cytotoxic action of doxorubicin and etoposide as well as increasing the intracellular accumulation of both drugs. These results suggest that CCE might be an effective concomitant agent to reduce anticancer drug resistance derived from Nrf2 overexpression. Bioactivity-guided fractionation revealed that procyanidin tetramers and pentamers contained in CCE were active components in suppressing Nrf2.« less

Analysis of nuclear resonance fluorescence excitation measured with LaBr3(Ce) detectors near 2 MeV

NASA Astrophysics Data System (ADS)

Omer, Mohamed; Negm, Hani; Ohgaki, Hideaki; Daito, Izuru; Hayakawa, Takehito; Bakr, Mahmoud; Zen, Heishun; Hori, Toshitada; Kii, Toshiteru; Masuda, Kai; Hajima, Ryoichi; Shizuma, Toshiyuki; Toyokawa, Hiroyuki; Kikuzawa, Nobuhiro

2013-11-01

The performance of LaBr3(Ce) to measure nuclear resonance fluorescence (NRF) excitations is discussed in terms of limits of detection and in comparison with high-purity germanium (HPGe) detectors near the 2 MeV region where many NRF excitation levels from special nuclear materials are located. The NRF experiment was performed at the High Intensity γ-ray Source (HIγS) facility. The incident γ-rays, of 2.12 MeV energy, hit a B4C target to excite the 11B nuclei to the first excitation level. The statistical-sensitive non-linear peak clipping (SNIP) algorithm was implemented to eliminate the background and enhance the limits of detection for the spectra measured with LaBr3(Ce). Both detection and determination limits were deduced from the experimental data.

Electrophilic nitro-fatty acids prevent astrocyte-mediated toxicity to motor neurons in a cell model of familial amyotrophic lateral sclerosis via nuclear factor erythroid 2-related factor activation.

PubMed

Diaz-Amarilla, Pablo; Miquel, Ernesto; Trostchansky, Andrés; Trias, Emiliano; Ferreira, Ana M; Freeman, Bruce A; Cassina, Patricia; Barbeito, Luis; Vargas, Marcelo R; Rubbo, Homero

2016-06-01

Nitro-fatty acids (NO2-FA) are electrophilic signaling mediators formed in tissues during inflammation, which are able to induce pleiotropic cytoprotective and antioxidant pathways including up regulation of Nuclear factor erythroid 2-related factor 2 (Nrf2) responsive genes. Amyotrophic Lateral Sclerosis (ALS) is a fatal neurodegenerative disease characterized by the loss of motor neurons associated to an inflammatory process that usually aggravates the disease progression. In ALS animal models, the activation of the transcription factor Nrf2 in astrocytes confers protection to neighboring neurons. It is currently unknown whether NO2-FA can exert protective activity in ALS through Nrf2 activation. Herein we demonstrate that nitro-arachidonic acid (NO2-AA) or nitro-oleic acid (NO2-OA) administrated to astrocytes expressing the ALS-linked hSOD1(G93A) induce antioxidant phase II enzyme expression through Nrf2 activation concomitant with increasing intracellular glutathione levels. Furthermore, treatment of hSOD1(G93A)-expressing astrocytes with NO2-FA prevented their toxicity to motor neurons. Transfection of siRNA targeted to Nrf2 mRNA supported the involvement of Nrf2 activation in NO2-FA-mediated protective effects. Our results show for the first time that NO2-FA induce a potent Nrf2-dependent antioxidant response in astrocytes capable of preventing motor neurons death in a culture model of ALS. Copyright © 2016 Elsevier Inc. All rights reserved.

Differential regulation of innate immune cytokine production through pharmacological activation of Nuclear Factor-Erythroid-2-Related Factor 2 (NRF2) in burn patient immune cells and monocytes

PubMed Central

Stepp, Wesley; Sjeklocha, Lucas; Long, Clayton; Riley, Caitlin; Callahan, James; Sanchez, Yolanda; Gough, Peter; Knowlin, Laquanda; van Duin, David; Ortiz-Pujols, Shiara; Jones, Samuel; Maile, Robert; Hong, Zhi; Berger, Scott; Cairns, Bruce

2017-01-01

Burn patients suffer from immunological dysfunction for which there are currently no successful interventions. Similar to previous observations, we find that burn shock patients (≥15% Total Burn Surface Area (TBSA) injury) have elevated levels of the innate immune cytokines Interleukin-6 (IL-6) and Monocyte Chemoattractant Protein-1 (MCP-1)/CC-motif Chemokine Ligand 2(CCL2) early after hospital admission (0–48 Hours Post-hospital Admission (HPA). Functional immune assays with patient Peripheral Blood Mononuclear Cells (PBMCs) revealed that burn shock patients (≥15% TBSA) produced elevated levels of MCP-1/CCL2 after innate immune stimulation ex vivo relative to mild burn patients. Interestingly, treatment of patient PBMCs with the Nuclear Factor-Erythroid-2-Related Factor 2 (NRF2) agonist, CDDO-Me(bardoxolone methyl), reduced MCP-1 production but not IL-6 or Interleukin-10 (IL-10) secretion. In enriched monocytes from healthy donors, CDDO-Me(bardoxolone methyl) also reduced LPS-induced MCP1/CCL2 production but did not alter IL-6 or IL-10 secretion. Similar immunomodulatory effects were observed with Compound 7, which activates the NRF2 pathway through a different and non-covalent Mechanism Of Action (MOA). Hence, our findings with CDDO-Me(bardoxolone methyl) and Compound 7 are likely to reflect a generalizable aspect of NRF2 activation. These observed effects were not specific to LPS-induced immune responses, as NRF2 activation also reduced MCP-1/CCL2 production after stimulation with IL-6. Pharmacological NRF2 activation reduced Mcp-1/Ccl2 transcript accumulation without inhibiting either Il-6 or Il-10 transcript levels. Hence, we describe a novel aspect of NRF2 activation that may contribute to the beneficial effects of NRF2 agonists during disease. Our work also demonstrates that the NRF2 pathway is retained and can be modulated to regulate important immunomodulatory functions in burn patient immune cells. PMID:28886135

Differential regulation of innate immune cytokine production through pharmacological activation of Nuclear Factor-Erythroid-2-Related Factor 2 (NRF2) in burn patient immune cells and monocytes.

PubMed

Eitas, Timothy K; Stepp, Wesley H; Sjeklocha, Lucas; Long, Clayton V; Riley, Caitlin; Callahan, James; Sanchez, Yolanda; Gough, Peter; Knowlin, Laquanda; van Duin, David; Ortiz-Pujols, Shiara; Jones, Samuel W; Maile, Robert; Hong, Zhi; Berger, Scott; Cairns, Bruce A

2017-01-01

Burn patients suffer from immunological dysfunction for which there are currently no successful interventions. Similar to previous observations, we find that burn shock patients (≥15% Total Burn Surface Area (TBSA) injury) have elevated levels of the innate immune cytokines Interleukin-6 (IL-6) and Monocyte Chemoattractant Protein-1 (MCP-1)/CC-motif Chemokine Ligand 2(CCL2) early after hospital admission (0-48 Hours Post-hospital Admission (HPA). Functional immune assays with patient Peripheral Blood Mononuclear Cells (PBMCs) revealed that burn shock patients (≥15% TBSA) produced elevated levels of MCP-1/CCL2 after innate immune stimulation ex vivo relative to mild burn patients. Interestingly, treatment of patient PBMCs with the Nuclear Factor-Erythroid-2-Related Factor 2 (NRF2) agonist, CDDO-Me(bardoxolone methyl), reduced MCP-1 production but not IL-6 or Interleukin-10 (IL-10) secretion. In enriched monocytes from healthy donors, CDDO-Me(bardoxolone methyl) also reduced LPS-induced MCP1/CCL2 production but did not alter IL-6 or IL-10 secretion. Similar immunomodulatory effects were observed with Compound 7, which activates the NRF2 pathway through a different and non-covalent Mechanism Of Action (MOA). Hence, our findings with CDDO-Me(bardoxolone methyl) and Compound 7 are likely to reflect a generalizable aspect of NRF2 activation. These observed effects were not specific to LPS-induced immune responses, as NRF2 activation also reduced MCP-1/CCL2 production after stimulation with IL-6. Pharmacological NRF2 activation reduced Mcp-1/Ccl2 transcript accumulation without inhibiting either Il-6 or Il-10 transcript levels. Hence, we describe a novel aspect of NRF2 activation that may contribute to the beneficial effects of NRF2 agonists during disease. Our work also demonstrates that the NRF2 pathway is retained and can be modulated to regulate important immunomodulatory functions in burn patient immune cells.

Activation of the Nrf2-ARE pathway by siRNA knockdown of Keap1 reduces oxidative stress and provides partial protection from MPTP-mediated neurotoxicity.

PubMed

Williamson, Tracy P; Johnson, Delinda A; Johnson, Jeffrey A

2012-06-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcription factor that binds to the antioxidant response element, a cis-acting regulatory element that increases expression of detoxifying enzymes and antioxidant proteins. Kelch-like ECH associating protein 1 (Keap1) protein is a negative regulator of Nrf2. Previous work has shown that genetic overexpression of Nrf2 is protective in vitro and in vivo. To modulate the Nrf2-ARE system without overexpressing Nrf2, we used short interfering RNA (siRNA) directed against Keap1. Keap1 siRNA administration in primary astrocytes increased the levels of Nrf2-ARE driven genes and protected against oxidative stress. Moreover, Keap1 siRNA resulted in a persistent upregulation of the Nrf2-ARE pathway and protection against oxidative stress in primary astrocytes. Keap1 siRNA injected into the striatum was also modestly protective against MPTP-induced dopaminergic terminal damage. These data indicate that activation of endogenous intracellular levels of Nrf2 is sufficient to protect in models of oxidative stress and Parkinson's disease. Copyright © 2012 Elsevier Inc. All rights reserved.

The effects of chromium picolinate and chromium histidinate administration on NF-κB and Nrf2/HO-1 pathway in the brain of diabetic rats.

PubMed

Sahin, Kazim; Tuzcu, Mehmet; Orhan, Cemal; Gencoglu, Hasan; Ulas, Mustafa; Atalay, Mustafa; Sahin, Nurhan; Hayirli, Armagan; Komorowski, James R

2012-12-01

The objective of this experiment was to investigate the effects of supplemental chromium picolinate (CrPic) and chromium histidinate (CrHis) on nuclear factor-kappa B (NF-κB p65) and nuclear factor (erythroid-derived 2)-like 2 (Nrf2) signaling pathway in diabetic rat brain. Nondiabetic (n = 45) and diabetic (n = 45) male Wistar rats were either not supplemented or supplemented with CrPic or CrHis via drinking water to consume 8 μg elemental chromium (Cr) per day for 12 weeks. Diabetes was induced by streptozotocin injection (40 mg/kg i.p., for 2 weeks) and maintained by high-fat feeding (40 %). Diabetes was associated with increases in cerebral NF-κB and 4-hydroxynonenal (4-HNE) protein adducts and decreased in cerebral nuclear factor of kappa light polypeptide gene enhancer in B cells inhibitor, alpha (IκBα) and Nrf2 levels. Both Cr chelates were effective to decrease levels of NF-κB and 4-HNE protein adducts and to increase levels of IκBα and Nrf2 in the brain of diabetic rats. However, responses of these increases and decreases were more notable when Cr was supplemented as CrHis than as CrPic. In conclusion, Cr may play a protective role in cerebral antioxidant defense system in diabetic subjects via the Nrf2 pathway by reducing inflammation through NF-κB p65 inhibition. Histidinate form of Cr was superior to picolinate form of Cr in reducing NF-κB expression and increasing Nrf2 expression in the brain of diabetic rats.

Green tea polyphenol (−)-epigallocatechin-3-gallate triggered hepatotoxicity in mice: Responses of major antioxidant enzymes and the Nrf2 rescue pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Dongxu; Wang, Yijun; Wan, Xiaochun

(−)-Epigallocatechin-3-gallate (EGCG), a constituent of green tea, has been suggested to have numerous health-promoting effects. On the other hand, high-dose EGCG is able to evoke hepatotoxicity. In the present study, we elucidated the responses of hepatic major antioxidant enzymes and nuclear factor erythroid 2-related factor 2 (Nrf2) rescue pathway to high-dose levels of EGCG in Kunming mice. At a non-lethal toxic dose (75 mg/kg, i.p.), repeated EGCG treatments markedly decreased the levels of superoxide dismutase, catalase, and glutathione peroxidase. As a rescue response, the nuclear distribution of Nrf2 was significantly increased; a battery of Nrf2-target genes, including heme oxygenase 1more » (HO1), NAD(P)H:quinone oxidoreductase 1 (NQO1), glutathione S-transferase (GST), and those involved in glutathione and thioredoxin systems, were all up-regulated. At the maximum tolerated dose (45 mg/kg, i.p.), repeated EGCG treatments did not disturb the major antioxidant defense. Among the above-mentioned genes, only HO1, NQO1, and GST genes were significantly but modestly up-regulated, suggesting a comprehensive and extensive activation of Nrf2-target genes principally occurs at toxic levels of EGCG. At a lethal dose (200 mg/kg, i.p.), a single EGCG treatment dramatically decreased not only the major antioxidant defense but also the Nrf2-target genes, demonstrating that toxic levels of EGCG are able to cause a biphasic response of Nrf2. Overall, the mechanism of EGCG-triggered hepatotoxicity involves suppression of major antioxidant enzymes, and the Nrf2 rescue pathway plays a vital role for counteracting EGCG toxicity. - Highlights: • EGCG at maximum tolerated dose does not disturb hepatic major antioxidant defense. • EGCG at maximum tolerated dose modestly upregulates hepatic Nrf2 target genes. • EGCG at toxic dose suppresses hepatic major antioxidant enzymes. • EGCG at non-lethal toxic dose pronouncedly activates hepatic Nrf2 rescue response. • EGCG at lethal dose substantially suppresses hepatic Nrf2 pathway.« less

Targeting NRF2 for Improved Skin Barrier Function and Photoprotection: Focus on the Achiote-Derived Apocarotenoid Bixin

PubMed Central

Rojo de la Vega, Montserrat; Krajisnik, Andrea; Zhang, Donna D.; Wondrak, Georg T.

2017-01-01

The transcription factor NRF2 (nuclear factor-E2-related factor 2) orchestrates major cellular defense mechanisms including phase-II detoxification, inflammatory signaling, DNA repair, and antioxidant response. Recent studies strongly suggest a protective role of NRF2-mediated gene expression in the suppression of cutaneous photodamage induced by solar UV (ultraviolet) radiation. The apocarotenoid bixin, a Food and Drug Administration (FDA)-approved natural food colorant (referred to as ‘annatto’) originates from the seeds of the achiote tree native to tropical America, consumed by humans since ancient times. Use of achiote preparations for skin protection against environmental insult and for enhanced wound healing has long been documented. We have recently reported that (i) bixin is a potent canonical activator of the NRF2-dependent cytoprotective response in human skin keratinocytes; that (ii) systemic administration of bixin activates NRF2 with protective effects against solar UV-induced skin damage; and that (iii) bixin-induced suppression of photodamage is observable in Nrf2+/+ but not in Nrf2−/− SKH-1 mice confirming the NRF2-dependence of bixin-induced antioxidant and anti-inflammatory effects. In addition, bixin displays molecular activities as sacrificial antioxidant, excited state quencher, PPAR (peroxisome proliferator-activated receptor) α/γ agonist, and TLR (Toll-like receptor) 4/NFκB (nuclear factor kappa-light-chain-enhancer of activated B cells) antagonist, all of which might be relevant to the enhancement of skin barrier function and environmental stress protection. Potential skin photoprotection and photochemoprevention benefits provided by topical application or dietary consumption of this ethno-pharmacologically validated phytochemical originating from the Americas deserves further preclinical and clinical examination. PMID:29258247

Expression of the Nrf2 and Keap1 proteins and their clinical significance in osteosarcoma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Jihong, E-mail: zhangjihong63@163.com; Wang, Xiaojuan, E-mail: yangjian142@163.com; Wu, Wuzhou, E-mail: jiangchunli68@163.com

Objective: To investigate the expression and clinical significance of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) and Kelch-like ECH-associated protein 1 (Keap1) in osteosarcoma tissue. Methods: The data of 102 osteosarcoma patients who underwent surgical treatment at our hospital from June 2000 to March 2009 were collected. The expression levels of the Nrf2 and Keap1 proteins in osteosarcoma tissue and normal peritumour tissues were detected by immunohistochemistry, and the relationship between the expression level and the clinical and pathological features as well as the prognosis was explored. Results: The nuclear expression rate of Nrf2 was 77.5% in osteosarcoma tissue, which wasmore » significantly higher than the rate in normal peritumour bone tissue (9.8%) (P < 0.05). The expression rate of the Keap1 protein in osteosarcoma tissue was 13.7%, which was significantly lower than the rate in normal peritumour tissue (80.4%). In addition, Nrf2/Keap1 expression was unrelated to patient gender and age, tumour site, and histological type and was related to metastasis and patient response to chemotherapy (P < 0.05). The five-year survival rate was significantly lower in patients with positive Nrf2 expression than in those with negative Nrf2 expression (p = 0.023), and it was significantly higher in patients with positive Keap1 expression than in those with negative Keap1 expression (P = 0.018). Conclusion: The expression of Nrf2-Keap1 is abnormal in osteosarcoma tissue and shows significant clinical relevance for determining the prognosis of osteosarcoma.« less

Nrf2 Is an Attractive Therapeutic Target for Retinal Diseases

PubMed Central

2016-01-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a redox-sensitive transcription factor that binds to antioxidant response elements located in the promoter region of genes encoding many antioxidant enzymes and phase II detoxifying enzymes. Activation of Nrf2 functions is one of the critical defensive mechanisms against oxidative stress in many species. The retina is constantly exposed to reactive oxygen species, and oxidative stress is a major contributor to age-related macular diseases. Moreover, the resulting inflammation and neuronal degeneration are also related to other retinal diseases. The well-known Nrf2 activators, bardoxolone methyl and its derivatives, have been the subject of a number of clinical trials, including those aimed at treating chronic kidney disease, pulmonary arterial hypertension, and mitochondrial myopathies. Recent studies suggest that Nrf2 activation protects the retina from retinal diseases. In particular, this is supported by the finding that Nrf2 knockout mice display age-related retinal degeneration. Moreover, the concept has been validated by the efficacy of Nrf2 activators in a number of retinal pathological models. We have also recently succeeded in generating a novel Nrf2 activator, RS9, using a biotransformation technique. This review discusses current links between retinal diseases and Nrf2 and the possibility of treating retinal diseases by activating the Nrf2 signaling pathway. PMID:27818722

Deletion of Nrf2 reduces skeletal mechanical properties and decreases load-driven bone formation.

PubMed

Sun, Yong-Xin; Li, Lei; Corry, Kylie A; Zhang, Pei; Yang, Yang; Himes, Evan; Mihuti, Cristina Layla; Nelson, Cecilia; Dai, Guoli; Li, Jiliang

2015-05-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcription factor expressed in many cell types, including osteoblasts, osteocytes, and osteoclasts. Nrf2 has been considered a master regulator of cytoprotective genes against oxidative and chemical insults. The lack of Nrf2 can induce pathologies in multiple organs. The aim of this study was to investigate the role of Nrf2 in load-driven bone metabolism using Nrf2 knockout (KO) mice. Compared to age-matched littermate wild-type controls, Nrf2 KO mice have significantly lowered femoral bone mineral density (-7%, p<0.05), bone formation rate (-40%, p<0.05), as well as ultimate force (-11%, p<0.01). The ulna loading experiment showed that Nrf2 KO mice were less responsive than littermate controls, as indicated by reduction in relative mineralizing surface (rMS/BS, -69%, p<0.01) and relative bone formation rate (rBFR/BS, -84%, p<0.01). Furthermore, deletion of Nrf2 suppressed the load-driven gene expression of antioxidant enzymes and Wnt5a in cultured primary osteoblasts. Taken together, the results suggest that the loss-of-function mutation of Nrf2 in bone impairs bone metabolism and diminishes load-driven bone formation. Copyright © 2015 Elsevier Inc. All rights reserved.

Fucoxanthin enhances HO-1 and NQO1 expression in murine hepatic BNL CL.2 cells through activation of the Nrf2/ARE system partially by its pro-oxidant activity.

PubMed

Liu, Cheng-Ling; Chiu, Yu-Ting; Hu, Miao-Lin

2011-10-26

To determine whether fucoxanthin, a major carotenoid in brown sea algae, may activate cellular antioxidant enzymes via up-regulation of the Nrf2/antioxidant-response element (ARE) pathway, we incubated mouse hepatic BNL CL.2 cells with fucoxanthin (0.5-20 μM) for 0-24 h. We found that fucoxanthin (≥5 μM) significantly increased cellular reactive oxygen species (ROS) at 6 h of incubation, whereas preincubation with α-d-tocopherol (30 μM) significantly attenuated the increase of ROS, indicating the pro-oxidant nature of fucoxanthin. Fucoxanthin significantly increased the phosphorylation of ERK and p38 and markedly increased nuclear Nrf2 protein accumulation after incubation for 12 h. Moreover, fucoxanthin significantly enhanced binding activities of nuclear Nrf2 with ARE and increased mRNA and protein expression of HO-1 and NQO1 after incubation for 12 h. siRNA inhibition of Nrf2 led to markedly decreased HO-1 and NQO1 protein expression. Thus, fucoxanthin may exert its antioxidant activity, at least partly, through its pro-oxidant actions.

Nuclear resonance fluorescence in U-238 using LaBr detectors for nuclear security

NASA Astrophysics Data System (ADS)

Hayakawa, Takehito; Negm, Hani; Ohgaki, Hideaki; Daito, Izuru; Kii, Toshiteru; Zen, Heishun; Omer, Mohamed; Shizuma, Toshiyuki; Hajima, Ryoichi

2014-09-01

Recently, a nondestructive measurement method of shielded fissional isotopes such as 235U or 239Pu has been proposed for the nuclear security. These isotopes are measured by using nuclear resonance fluorescence (NRF) with monochromatic energy gamma-ray beams generated by laser Compton-scattering (LCS). We have proposed that one measure scattered gamma-rays from NRF with LCS gamma-ray beams using the LaBr3(Ce) detectors. The LaBr3(Ce) crystals include internal radioisotopes of a meta-stable isotope 138La and alpha decay chains from some actinides as 227Ac. There is a broad pump at about 2 MeV. This pump is considered to be an overlap of alpha-rays from decay chains of some actinides but its detailed structure has not been established. Here we have measured NRF spectra of 238U using the LCS gamma-rays with energy of about 2.5 MeV at the HIgS facility of the Duke University. The background has been evaluated using a simulation code GEAT4. The 9 peaks, 8 NRF gamma-rays plus the Compton scattered gamma-ray of the incident beam, are finally assigned in an energy range of about 200 keV at about 2.5 MeV. The 8 integrated NRF cross-sections measured by LaBr3(Ce) have been consistent with results by an HPGe detector. The three levels are newly assigned using the HPGe detector. Two of them are also measured by LaBr3(Ce).

The hypertension drug, verapamil, activates Nrf2 by promoting p62-dependent autophagic Keap1 degradation and prevents acetaminophen-induced cytotoxicity.

PubMed

Lee, Da Hyun; Park, Jeong Su; Lee, Yu Seol; Sung, Su Haeng; Lee, Yong-Ho; Bae, Soo Han

2017-02-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) provides a cellular defense against oxidative stress by inducing the expression of antioxidant and detoxification enzymes. The calcium antagonist, verapamil, is an FDA-approved drug prescribed for the treatment of hypertension. Here, we show that verapamil acts as a potent Nrf2 activator without causing cytotoxicity, through degradation of Kelch-like ECH-associated protein 1 (Keap1), a Nrf2 repressor. Furthermore, verapamilinduced Keap1 degradation is prominently mediated by a p62-dependent autophagic pathway. Correspondingly, verapamil protects cells from acetaminophen-induced oxidative damage through Nrf2 activation. These results demonstrated the underlying mechanisms for the protective role of verapamil against acetaminophen-induced cytotoxicity. [BMB Reports 2017; 50(2): 91-96].

Ulinastatin activates haem oxygenase 1 antioxidant pathway and attenuates allergic inflammation

PubMed Central

Song, Dongmei; Song, Geng; Niu, Yinghao; Song, Wei; Wang, Jiantao; Yu, Lei; Yang, Jianwang; Lv, Xin; Steinberg, Harry; Liu, Shu Fang; Wang, Baoshan

2014-01-01

Background and Purpose Ulinastatin (UTI), a serine protease inhibitor, was recently found to have an anti-inflammatory action. However, the mechanisms mediating this anti-inflammatory effect are not well understood. This study tested the hypothesis that UTI suppresses allergic inflammation by inducing the expression of haem oxygenase 1 (HO1). Experimental Approach Control mice and mice sensitized (on days 1, 9 and 14) and challenged (on days 21 to 27) with ovalbumin (OVA) were treated with UTI. The effects of UTI on basal expression of HO1 and that induced by OVA challenge were examined. The involvement of UTI-induced HO1 expression in anti-inflammatory and antioxidant effects of UTI was also evaluated. Key Results UTI markedly increased basal HO1 protein expression in lungs of control mice in a time- and dose-dependent manner, and augmented HO1 protein expression induced by OVA. The up-regulation of HO1 mediated by UTI in sensitized and OVA-challenged mice was associated with reduced airway inflammation, alleviated tissue injury, reduced oxidant stress and enhanced antioxidant enzyme activities. Inhibition of HO1 activity using HO1 inhibitor, zinc protoporphyrin, attenuated inhibitory effects of UTI on inflammation and oxidant stress, and its stimulant effects on antioxidant enzyme activities. Mechanistic analysis showed that UTI increased nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2), stimulated Nrf2 DNA binding activity and concomitantly up-regulated HO1 mRNA expression. Conclusions and Implications UTI is a potent and naturally occurring inducer of HO1 expression. HO1 up-regulation contributes significantly to the anti-inflammatory and organ-protective effects of UTI, which has important research and therapeutic implications. PMID:24835359

Activation of Nrf2 target enzymes conferring protection against oxidative stress in PC12 cells by ginger principal constituent 6-shogaol.

PubMed

Peng, Shoujiao; Yao, Juan; Liu, Yaping; Duan, Dongzhu; Zhang, Xiaolong; Fang, Jianguo

2015-08-01

Natural compounds containing phenoxyl groups and/or Michael acceptor units appear to possess antioxidant and cytoprotective properties. The ginger principal constituent 6-shogaol (6-S) represents one of such compounds. In this study, we reported that 6-S efficiently scavenges various free radicals in vitro, and displays remarkable cytoprotection against oxidative stress-induced cell damage in the neuron-like rat pheochromocytoma cell line, PC12 cells. Pretreatment of PC12 cells with 6-S significantly upregulates a series of phase II antioxidant molecules, such as glutathione, heme oxygenase 1, NAD(P)H: quinone oxidoreductase 1, thioredoxin reductase 1, and thioredoxin 1. A mechanistic study revealed that 6-S enhanced the translocation of Nrf2 from the cytosol to the nucleus and knockdown of Nrf2 abolished such protection, indicating that this cytoprotection is mediated by the activation of the transcription factor Nrf2. Another ginger constituent 6-gingerol (6-G), having a similar structure of 6-S but lacking the alpha,beta-unsaturated ketone structure (Michael acceptor moiety), failed to shelter PC12 cells from oxidative stress. Our results demonstrate that 6-S is a novel small molecule activator of Nrf2 in PC12 cells, and suggest that 6-S might be a potential candidate for the prevention of oxidative stress-mediated neurodegenerative disorders.

Metallothionein plays a prominent role in the prevention of diabetic nephropathy by sulforaphane via up-regulation of Nrf2.

PubMed

Wu, Hao; Kong, Lili; Cheng, Yanli; Zhang, Zhiguo; Wang, Yangwei; Luo, Manyu; Tan, Yi; Chen, Xiangmei; Miao, Lining; Cai, Lu

2015-12-01

Sulforaphane (SFN) prevents diabetic nephropathy (DN) in type 1 diabetes via up-regulation of nuclear factor (erythroid-derived 2)-like 2 (Nrf2). However, it has not been addressed whether SFN also prevents DN from type 2 diabetes or which Nrf2 downstream gene(s) play(s) the key role in SFN renal protection. Here we investigated whether Nrf2 is required for SFN protection against type 2 diabetes-induced DN and whether metallothionein (MT) is an Nrf2 downstream antioxidant using Nrf2 knockout (Nrf2-null) mice. In addition, MT knockout mice were used to further verify if MT is indispensable for SFN protection against DN. Diabetes-increased albuminuria, renal fibrosis, and inflammation were significantly prevented by SFN, and Nrf2 and MT expression was increased. However, SFN renal protection was completely lost in Nrf2-null diabetic mice, confirming the pivotal role of Nrf2 in SFN protection from type 2 diabetes-induced DN. Moreover, SFN failed to up-regulate MT in the absence of Nrf2, suggesting that MT is an Nrf2 downstream antioxidant. MT deletion resulted in a partial, but significant attenuation of SFN renal protection from type 2 diabetes, demonstrating a partial requirement for MT for SFN renal protection. Therefore, the present study demonstrates for the first time that as an Nrf2 downstream antioxidant, MT plays an important, though partial, role in mediating SFN renal protection from type 2 diabetes. Copyright © 2015 Elsevier Inc. All rights reserved.

Bardoxolone brings Nrf2-based therapies to light.

PubMed

Zhang, Donna D

2013-08-10

The targeted activation of nuclear factor erythroid-derived-2-like 2 (Nrf2) to alleviate symptoms of chronic kidney disease has recently garnered much attention. Unfortunately, the greatest clinical success to date, bardoxolone, failed in phase III clinical trial for unspecified safety reasons. The present letter to the editor discusses the clinical development of bardoxolone and explores potential reasons for the ultimate withdrawal from clinical trials. In particular, was the correct clinical indication pursued and would improved specificity have mitigated the safety concerns? Ultimately, it is concluded that the right clinical indication and heightened specificity will lead to successful Nrf2-based therapies. Therefore, the bardoxolone clinical results do not dampen enthusiasm for Nrf2-based therapies; rather it illuminates the clinical potential of the Nrf2 pathway as a drug target.

Role of Nuclear Factor Erythroid 2-Related Factor 2 in Diabetic Nephropathy

PubMed Central

Min, Xu; Xu, Xiaohong

2017-01-01

Diabetic nephropathy (DN) is manifested as increased urinary protein level, decreased glomerular filtration rate, and final renal dysfunction. DN is the leading cause of end-stage renal disease worldwide and causes a huge societal healthcare burden. Since satisfied treatments are still limited, exploring new strategies for the treatment of this disease is urgently needed. Oxidative stress takes part in the initiation and development of DN. In addition, nuclear factor erythroid 2-related factor 2 (Nrf2) plays a key role in the cellular response to oxidative stress. Thus, activation of Nrf2 seems to be a new choice for the treatment of DN. In current review, we discussed and summarized the therapeutic effects of Nrf2 activation on DN from both basic and clinical studies. PMID:28512642

Phytochemicals and botanical extracts regulate NF-κB and Nrf2/ARE reporter activities in DI TNC1 astrocytes

PubMed Central

Ajit, Deepa; Simonyi, Agnes; Li, Runting; Chen, Zihong; Hannink, Mark; Fritsche, Kevin L.; Mossine, Valeri V.; Smith, Robert E.; Dobbs, Thomas K.; Luo, Rensheng; Folk, William R.; Gu, Zezong; Lubahn, Dennis B.; Weisman, Gary A.; Sun, Grace Y.

2016-01-01

The increase in oxidative stress and inflammatory responses associated with neurodegenerative diseases has drawn considerable attention towards understanding the transcriptional signaling pathways involving NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) and Nrf2 (Nuclear Factor Erythroid 2-like 2). Our recent studies with immortalized murine microglial cells (BV-2) demonstrated effects of botanical polyphenols to inhibit lipopolysaccharide (LPS)-induced nitric oxide (NO) and enhance Nrf2-mediated antioxidant responses (Sun et al., 2015). In this study, an immortalized rat astrocyte (DI TNC1) cell line expressing a luciferase reporter driven by the NF-κB or the Nrf2/Antioxidant Response Element (ARE) promoter was used to assess regulation of these two pathways by phytochemiscals such as quercetin, rutin, cyanidin, cyanidin-3-O-glucoside, as well as botanical extracts from Withania somnifera (Ashwagandha), Sutherlandia frutescens (Sutherlandia) and Euterpe oleracea (Açaí). Quercetin effectively inhibited LPS-induced NF-κB reporter activity and stimulated Nrf2/ARE reporter activity in DI TNC1 astrocytes. Cyanidin and the glycosides showed similar effects but only at much higher concentrations. All three botanical extracts effectively inhibited LPS-induced NF-κB reporter activity. These extracts were capable of enhancing ARE activity by themselves and further enhanced ARE activity in the presence of LPS. Quercetin and botanical extracts induced Nrf2 and HO-1 protein expression. Interestingly, Ashwagandha extract was more active in inducing Nrf2 and HO-1 expression in DI TNC1 astrocytes as compared to Sutherlandia and Açaí extracts. In summary, this study demonstrated NF-kB and Nrf2/ARE promotor activities in DI TNC1 astrocytes, and further showed differences in ability for specific botanical polyphenols and extracts to down-regulate LPS-induced NF-kB and up-regulate the NRF2/ARE activities in these cells. PMID:27166148

Characterization of docosahexaenoic acid (DHA)-induced heme oxygenase-1 (HO-1) expression in human cancer cells: the importance of enhanced BTB and CNC homology 1 (Bach1) degradation.

PubMed

Wang, Shuai; Hannafon, Bethany N; Wolf, Roman F; Zhou, Jundong; Avery, Jori E; Wu, Jinchang; Lind, Stuart E; Ding, Wei-Qun

2014-05-01

The effect of docosahexaenoic acid (DHA) on heme oxygenase-1 (HO-1) expression in cancer cells has never been characterized. This study examines DHA-induced HO-1 expression in human cancer cell model systems. DHA enhanced HO-1 gene expression in a time- and concentration-dependent manner, with maximal induction at 21 h of treatment. This induction of HO-1 expression was confirmed in vivo using a xenograft nude mouse model fed a fish-oil-enriched diet. The increase in HO-1 gene transcription induced by DHA was significantly attenuated by the antioxidant N-acetyl cysteine, suggesting the involvement of oxidative stress. This was supported by direct measurement of lipid peroxide levels after DHA treatment. Using a human HO-1 gene promoter reporter construct, we identified two antioxidant response elements (AREs) that mediate the DHA-induced increase in HO-1 gene transcription. Knockdown of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) expression compromised the DHA-induced increase in HO-1 gene transcription, indicating the importance of the Nrf2 pathway in this event. However, the nuclear protein levels of Nrf2 remained unchanged upon DHA treatment. Further studies demonstrated that DHA reduces nuclear Bach1 protein expression by promoting its degradation and attenuates Bach1 binding to the AREs in the HO-1 gene promoter. In contrast, DHA enhanced Nrf2 binding to the AREs without affecting nuclear Nrf2 expression levels, indicating a new cellular mechanism that mediates DHA's induction of HO-1 gene transcription. To our knowledge, this is the first characterization of DHA-induced HO-1 expression in human malignant cells. Copyright © 2014 Elsevier Inc. All rights reserved.

Activation of the NRF2 pathway and its impact on the prognosis of anaplastic glioma patients

PubMed Central

Kanamori, Masayuki; Higa, Tsuyoshi; Sonoda, Yukihiko; Murakami, Shohei; Dodo, Mina; Kitamura, Hiroshi; Taguchi, Keiko; Shibata, Tatsuhiro; Watanabe, Mika; Suzuki, Hiroyoshi; Shibahara, Ichiyo; Saito, Ryuta; Yamashita, Yoji; Kumabe, Toshihiro; Yamamoto, Masayuki; Motohashi, Hozumi; Tominaga, Teiji

2015-01-01

Background Nuclear factor erythroid 2–related factor 2 (NRF2) plays pivotal roles in cytoprotection. We aimed at clarifying the contribution of the NRF2 pathway to malignant glioma pathology. Methods NRF2 target gene expression and its association with prognosis were examined in 95 anaplastic gliomas with or without isocitrate dehydrogenase (IDH) 1/2 gene mutations and 52 glioblastomas. To explore mechanisms for the altered activity of the NRF2 pathway, we examined somatic mutations and expressions of the NRF2 gene and those encoding NRF2 regulators, Kelch-like ECH-associated protein 1 (KEAP1) and p62/SQSTSM. To clarify the functional interaction between IDH1 mutations and the NRF2 pathway, we introduced a mutant IDH1 to T98 glioblastoma-derived cells and examined the NRF2 activity in these cells. Results NRF2 target genes were elevated in 13.7% and 32.7% of anaplastic gliomas and glioblastomas, respectively. Upregulation of NRF2 target genes correlated with poor prognosis in anaplastic gliomas but not in glioblastomas. Neither somatic mutations of NRF2/KEAP1 nor dysregulated expression of KEAP1/p62 explained the increased expression of NRF2 target genes. In most cases of anaplastic glioma with mutated IDH1/2, NRF2 and its target genes were downregulated. This was reproducible in IDH1 R132H–expressing T98 cells. In minor cases of IDH1/2-mutant anaplastic gliomas with increased expression of NRF2 target genes, the clinical outcomes were significantly poor. Conclusions The NRF2 activity is increased in a significant proportion of malignant gliomas in general but decreased in the majority of IDH1/2-mutant anaplastic gliomas. It is plausible that the NRF2 pathway plays an important role in tumor progression of anaplastic gliomas with IDH1/2 mutations. PMID:25304134

Sulforaphane reduces hepatic glucose production and improves glucose control in patients with type 2 diabetes.

PubMed

Axelsson, Annika S; Tubbs, Emily; Mecham, Brig; Chacko, Shaji; Nenonen, Hannah A; Tang, Yunzhao; Fahey, Jed W; Derry, Jonathan M J; Wollheim, Claes B; Wierup, Nils; Haymond, Morey W; Friend, Stephen H; Mulder, Hindrik; Rosengren, Anders H

2017-06-14

A potentially useful approach for drug discovery is to connect gene expression profiles of disease-affected tissues ("disease signatures") to drug signatures, but it remains to be shown whether it can be used to identify clinically relevant treatment options. We analyzed coexpression networks and genetic data to identify a disease signature for type 2 diabetes in liver tissue. By interrogating a library of 3800 drug signatures, we identified sulforaphane as a compound that may reverse the disease signature. Sulforaphane suppressed glucose production from hepatic cells by nuclear translocation of nuclear factor erythroid 2-related factor 2 (NRF2) and decreased expression of key enzymes in gluconeogenesis. Moreover, sulforaphane reversed the disease signature in the livers from diabetic animals and attenuated exaggerated glucose production and glucose intolerance by a magnitude similar to that of metformin. Finally, sulforaphane, provided as concentrated broccoli sprout extract, reduced fasting blood glucose and glycated hemoglobin (HbA1c) in obese patients with dysregulated type 2 diabetes. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Long-term Autophagy and Nrf2 Signaling in the Hippocampi of Developing Mice after Carbon Ion Exposure

PubMed Central

Ye, Fei; Zhao, Ting; Liu, Xiongxiong; Jin, Xiaodong; Liu, Xinguo; Wang, Tieshan; Li, Qiang

2015-01-01

To explore charged particle radiation-induced long-term hippocampus damage, we investigated the expression of autophagy and antioxidant Nrf2 signaling-related proteins in the mouse hippocampus after carbon ion radiation. Heads of immature female Balb/c mice were irradiated with carbon ions of different LETs at various doses. Behavioral tests were performed on the mice after maturation. Acute and chronic expression of LC3-II, p62/SQSTM1, nuclear Nrf2, activated caspase-3 and the Bax/Bcl-2 ratio were measured in the hippocampi. Secondary X-ray insult was adopted to amplify potential damages. Long-term behavioral changes were observed in high-LET carbon ion-irradiated mice. There were no differences in the rates of LC3-II induction and p62/SQSTM1 degradation compared to the control group regardless of whether the mice received the secondary X-ray insult. A high nuclear Nrf2 content and low apoptosis level in hippocampal cells subjected to secondary X-rays were observed for the mice exposed to relatively low-LET carbon ions. Therefore, carbon ion exposure in the immature mouse led to an LET-dependent behavioral change after maturation. Although autophagy was intact, the persistently high nuclear Nrf2 content in the hippocampus might account for the unchanged behavioral pattern in mice exposed to the relatively low-LET carbon ions and the subsequent increased radioresistance of the hippocampus. PMID:26689155

Comparison of hepatic NRF2 and AHR binding in 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) treated mice demonstrates NRF2-independent PKM2 induction.

PubMed

Nault, Rance; Doskey, Claire M; Fader, Kelly A; Rockwell, Cheryl E; Zacharewski, Timothy R

2018-05-11

2,3,7,8-Tetrachlorodibenzo- p -dioxin (TCDD) induces hepatic oxidative stress following activation of the aryl hydrocarbon receptor (AhR). Our recent studies showed TCDD induced pyruvate kinase muscle isoform 2 ( Pkm2 ) as a novel antioxidant response in normal differentiated hepatocytes. To investigate cooperative regulation between nuclear factor, erythroid derived 2, like 2 ( Nrf2 ) and the AhR in the induction of Pkm2 , hepatic ChIP-seq analyses were integrated with RNA-seq time course data from mice treated with TCDD for 2 - 168h. ChIP-seq analysis 2h after TCDD treatment identified genome-wide NRF2 enrichment. Approximately 842 NRF2 enriched regions were located in the regulatory region of differentially expressed genes (DEGs) while 579 DEGs showed both NRF2 and AhR enrichment. Sequence analysis of regions with overlapping NRF2 and AhR enrichment showed over-representation of either antioxidant or dioxin response elements (ARE and DRE, respectively), although 18 possessed both motifs. NRF2 exhibited negligible enrichment within a closed Pkm chromatin region while the AhR was enriched 29-fold. Furthermore, TCDD induced Pkm2 in primary hepatocytes from wild-type and Nrf2 null mice, indicating NRF2 is not required. Although NRF2 and AhR cooperate to regulate numerous antioxidant gene expression responses, the induction of Pkm2 by TCDD is independent of ROS-mediated NRF2 activation. The American Society for Pharmacology and Experimental Therapeutics.

Nrf2 protects against airway disorders

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cho, Hye-Youn, E-mail: cho2@niehs.nih.go; Kleeberger, Steven R.

Nuclear factor-erythroid 2 related factor 2 (Nrf2) is a ubiquitous master transcription factor that regulates antioxidant response elements (AREs)-mediated expression of antioxidant enzyme and cytoprotective proteins. In the unstressed condition, Kelch-like ECH-associated protein 1 (Keap1) suppresses cellular Nrf2 in cytoplasm and drives its proteasomal degradation. Nrf2 can be activated by diverse stimuli including oxidants, pro-oxidants, antioxidants, and chemopreventive agents. Nrf2 induces cellular rescue pathways against oxidative injury, abnormal inflammatory and immune responses, apoptosis, and carcinogenesis. Application of Nrf2 germ-line mutant mice has identified an extensive range of protective roles for Nrf2 in experimental models of human disorders in the liver,more » gastrointestinal tract, airway, kidney, brain, circulation, and immune or nerve system. In the lung, lack of Nrf2 exacerbated toxicity caused by multiple oxidative insults including supplemental respiratory therapy (e.g., hyperoxia, mechanical ventilation), cigarette smoke, allergen, virus, bacterial endotoxin and other inflammatory agents (e.g., carrageenin), environmental pollution (e.g., particles), and a fibrotic agent bleomycin. Microarray analyses and bioinformatic studies elucidated functional AREs and Nrf2-directed genes that are critical components of signaling mechanisms in pulmonary protection by Nrf2. Association of loss of function with promoter polymorphisms in NRF2 or somatic and epigenetic mutations in KEAP1 and NRF2 has been found in cohorts of patients with acute lung injury/acute respiratory distress syndrome or lung cancer, which further supports the role for NRF2 in these lung diseases. In the current review, we address the role of Nrf2 in airways based on emerging evidence from experimental oxidative disease models and human studies.« less

Decreased histone deacetylase 2 impairs Nrf2 activation by oxidative stress

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mercado, Nicolas; Thimmulappa, Rajesh; Thomas, Catherine M.R.

2011-03-11

Research highlights: {yields} Nrf2 anti-oxidant function is impaired when HDAC activity is inhibited. {yields} HDAC inhibition decreases Nrf2 protein stability. {yields} HDAC2 is involved in reduced Nrf2 stability and both correlate in COPD samples. {yields} HDAC inhibition increases Nrf2 acetylation. -- Abstract: Nuclear factor erythroid 2-related factor 2 (Nrf2) plays a crucial role in cellular defence against oxidative stress by inducing the expression of multiple anti-oxidant genes. However, where high levels of oxidative stress are observed, such as chronic obstructive pulmonary disease (COPD), Nrf2 activity is reduced, although the molecular mechanism for this defect is uncertain. Here, we show thatmore » down-regulation of histone deacetylase (HDAC) 2 causes Nrf2 instability, resulting in reduced anti-oxidant gene expression and increase sensitivity to oxidative stress. Although Nrf2 protein was clearly stabilized after hydrogen peroxide (H{sub 2}O{sub 2}) stimulation in a bronchial epithelial cell line (BEAS2B), Nrf2 stability was decreased and Nrf2 acetylation increased in the presence of an HDAC inhibitor, trichostatin A (TSA). TSA also reduced Nrf2-regulated heme-oxygenase-1 (HO-1) expression in these cells, and this was confirmed in acute cigarette-smoke exposed mice in vivo. HDAC2 knock-down by RNA interference resulted in reduced H{sub 2}O{sub 2}-induced Nrf2 protein stability and activity in BEAS2B cells, whereas HDAC1 knockdown had no effect. Furthermore, monocyte-derived macrophages obtained from healthy volunteers (non-smokers and smokers) and COPD patients showed a significant correlation between HDAC2 expression and Nrf2 expression (r = 0.92, p < 0.0001). Thus, reduced HDAC2 activity in COPD may account for increased Nrf2 acetylation, reduced Nrf2 stability and impaired anti oxidant defences.« less

A novel nuclear factor erythroid 2-related factor 2 (Nrf2) activator RS9 attenuates brain injury after ischemia reperfusion in mice.

PubMed

Yamauchi, Keita; Nakano, Yusuke; Imai, Takahiko; Takagi, Toshinori; Tsuruma, Kazuhiro; Shimazawa, Masamitsu; Iwama, Toru; Hara, Hideaki

2016-10-01

Recanalization of occluded vessels leads to ischemia-reperfusion injury (IRI), with oxidative stress as one of the main causes of injury, despite the fact that recanalization therapy is the most effective treatment for ischemic stroke. The nuclear factor erythroid 2-related factor 2 (Nrf2) is one of the transcription factors which has an essential role in protection against oxidative stress. RS9 is a novel Nrf2 activator obtained from bardoxolone methyl (BARD), an Nrf2 activator that has already been tested in a clinical trial, using a biotransformation technique. RS9 has been reported to lead to higher Nrf2 activation and less cytotoxicity than BARD. In this study, we investigated the effects of RS9 on IRI. Mice were intraperitoneally treated immediately after 2h of transient middle cerebral artery occlusion (MCAO) with a vehicle solution or 0.2mg/kg of RS9. Post-onset treatment of RS9 attenuated the infarct volume and improved neurological deficits 22h after reperfusion. RS9 activated Nrf2 2 and 6h after reperfusion and activated heme oxygenase-1 at 6 and 22h after reperfusion. RS9 also attenuated the phosphorylation of NF-κB p65 2 and 6h after reperfusion. Finally, RS9 improved the survival rate and neurological deficits 7days after MCAO. Our results suggest that the activation of Nrf2 by RS9 has a neuroprotective effect, mediated by attenuating both oxidative stress and neuroinflammation, and that RS9 is an effective therapeutic candidate for the treatment of IRI. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

Acrylamide-induced oxidative stress and inflammatory response are alleviated by N-acetylcysteine in PC12 cells: Involvement of the crosstalk between Nrf2 and NF-κB pathways regulated by MAPKs.

PubMed

Pan, Xiaoqi; Wu, Xu; Yan, Dandan; Peng, Cheng; Rao, Chaolong; Yan, Hong

2018-05-15

Acrylamide (ACR) is a classic neurotoxin in animals and humans. However, the mechanism underlying ACR neurotoxicity remains controversial, and effective prevention and treatment measures against this condition are scarce. This study focused on clarifying the crosstalk between the involved signaling pathways in ACR-induced oxidative stress and inflammatory response and investigating the protective effect of antioxidant N-acetylcysteine (NAC) against ACR in PC12 cells. Results revealed that ACR exposure led to oxidative stress characterized by significant increase in reactive oxygen species (ROS) and malondialdehyde (MDA) levels and glutathione (GSH) consumption. Inflammatory response was observed based on the dose-dependently increased levels of pro-inflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6). NAC attenuated ACR-induced enhancement of MDA and ROS levels and TNF-α generation. In addition, ACR activated nuclear transcription factor E2-related factor 2 (Nrf2) and nuclear factor-κB (NF-κB) signaling pathways. Knockdown of Nrf2 by siRNA significantly blocked the increased NF-κB p65 protein expression in ACR-treated PC12 cells. Down-regulation of NF-κB by specific inhibitor BAY11-7082 similarly reduced ACR-induced increase in Nrf2 protein expression. NAC treatment increased Nrf2 expression and suppressed NF-κB p65 expression to ameliorate oxidative stress and inflammatory response caused by ACR. Further results showed that mitogen-activated protein kinases (MAPKs) pathway was activated prior to the activation of Nrf2 and NF-κB pathways. Inhibition of MAPKs blocked Nrf2 and NF-κB pathways. Collectively, ACR activated Nrf2 and NF-κB pathways which were regulated by MAPKs. A crosstalk between Nrf2 and NF-κB pathways existed in ACR-induced cell damage. NAC protected against oxidative damage and inflammatory response induced by ACR by activating Nrf2 and inhibiting NF-κB pathways in PC12 cells. Copyright © 2018 Elsevier B.V. All rights reserved.

Polydatin promotes Nrf2-ARE anti-oxidative pathway through activating Sirt1 to resist AGEs-induced upregulation of fibronetin and transforming growth factor-β1 in rat glomerular messangial cells.

PubMed

Huang, Kaipeng; Chen, Cheng; Hao, Jie; Huang, Junying; Wang, Shaogui; Liu, Peiqing; Huang, Heqing

2015-01-05

Sirt1 and nuclear factor-E2 related factor 2 (Nrf2)-anti-oxidant response element (ARE) anti-oxidative pathway play important regulatory roles in the pathological progression of diabetic nephropathy (DN) induced by advanced glycation-end products (AGEs). Polydatin (PD), a glucoside of resveratrol, has been shown to possess strong anti-oxidative bioactivity. Our previous study demonstrated that PD markedly resists the progression of diabetic renal fibrosis and thus, inhibits the development of DN. Whereas, whether PD could resist DN through regulating Sirt1 and consequently promoting Nrf2-ARE pathway needs further investigation. Here, we found that concomitant with decreasing RAGE (the specific receptor for AGEs) expression, PD significantly reversed the downregulation of Sirt1 in terms of protein expression and deacetylase activity and attenuated FN and TGF-β1 expression in GMCs exposed to AGEs. Under AGEs-treatment condition, PD could decrease Keap1 expression and promote the nuclear content, ARE-binding ability, and transcriptional activity of Nrf2. In addition, PD increased the protein levels of heme oxygenase 1 (HO-1) and superoxide dismutase 1 (SOD1), two target genes of Nrf2. The activation of Nrf2-ARE pathway by PD eventually led to the quenching of ROS overproduction sharply boosted by AGEs. Depletion of Sirt1 blocked Nrf2-ARE pathway activation and reversed FN and TGF-β1 downregulation induced by PD in GMCs challenged with AGEs. Along with reducing HO-1 and SOD1 expression, silencing of Nrf2 increased FN and TGF-β1 levels. PD treatment elevated Sirt1 and Nrf2 levels in the kidney tissues of diabetic rats, then improved the anti-oxidative capacity and renal dysfunction of diabetic models, and finally reversed the upregulation of FN and TGF-β1. Taken together, the resistance of PD on upregulated FN and TGF-β1 induced by AGEs via oxidative stress in GMCs is closely associated with its activation of Sirt1-Nrf2-ARE pathway. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Nrf2 the rescue: effects of the antioxidative/electrophilic response on the liver.

PubMed

Klaassen, Curtis D; Reisman, Scott A

2010-04-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcription factor that positively regulates the basal and inducible expression of a large battery of cytoprotective genes. These gene products include proteins that catalyze reduction reactions (NAD(P)H:quinone oxidoreductase 1, Nqo1), conjugation reactions (glutathione-S-transferases, Gsts and UDP-glucuronosyltransferases, Ugts), as well as the efflux of potentially toxic xenobiotics and xenobiotic conjugates (multidrug resistance-associated proteins, Mrps). The significance of Nrf2 in the liver has been established, as livers of Nrf2-null mice are more susceptible to various oxidative/electrophilic stress-induced pathologies than wild-type mice. In contrast, both pharmacological and genetic models of hepatic Nrf2 activation are protective against oxidative/electrophilic stress. Furthermore, because certain Nrf2-target genes in the liver could affect the distribution, metabolism, and excretion of xenobiotics, the effects of Nrf2 on the kinetics of drugs and other xenobiotics should also be considered, with a special emphasis on metabolism and excretion. Therefore, this review highlights the research that has contributed to the understanding of the importance of Nrf2 in toxicodynamics and toxicokinetics, especially that which pertains to the liver. 2010 Elsevier Inc. All rights reserved.

α-Dihydroxychalcone-glycoside (α-DHC) isolated from the heartwood of Pterocarpus marsupium inhibits LPS induced MAPK activation and up regulates HO-1 expression in murine RAW 264.7 macrophage.

PubMed

Chakraborty, Prarthana; Saraswat, Ghungroo; Kabir, Syed N

2014-05-15

Three phenolic glycosides isolated from the heartwood of Pterocarpus marsupium showed significant free radical and superoxide ion scavenging activity and antioxidant potential that were comparable to, or several folds higher than those of standard antioxidants, trolox and ascorbic acid. The effective concentrations of these compounds were far below their cytotoxic levels. Compound 3, which was characterized to be α-dihydroxychalcone-glycoside (α-DHC), was the most potent one. Subsequent studies demonstrated that α-DHC effectively reduced nitric oxide and cytokine production by the LPS stimulated RAW 264.7 mouse macrophage cell line. The compound effectively attenuated the expression of inflammation-mediating enzymes COX-2 and iNOS at the mRNA as well as protein levels in a concentration dependent manner. It prevented phosphorylation of all the three MAPKs (JNK, ERK, p38) and eventually blocked the activation of downstream elements contributing to inflammation. Phosphorylation of IκB-α and subsequent translocation of NF-κB into the nucleus were restricted, while the expression of stress responsive gene HO-1 was up-regulated. α-DHC targeted Keap-1 by modifying its cysteine thiols, dissociating it from Nrf-2 and facilitating nuclear entry of the latter; and this in turn induced HO-1 expression. Thus α-DHC exerts its anti-inflammatory activity in a dual manner: by down regulating MAPKs and restricting nuclear stabilization of NF-κB at one end, and by disrupting Nrf-2-Keap-1 complex on the other. In conclusion, the anti-inflammatory potential together with its high therapeutic index envisages α-DHC as a prospective candidate molecule for the development of therapeutic strategy against inflammatory disorders. Copyright © 2014 Elsevier Inc. All rights reserved.

Translational control of Nrf2 within the open reading frame

DOE Office of Scientific and Technical Information (OSTI.GOV)

Perez-Leal, Oscar, E-mail: operez@temple.edu; Barrero, Carlos A.; Merali, Salim, E-mail: smerali@temple.edu

2013-07-19

Highlights: •Identification of a novel Nrf2 translational repression mechanism. •The repressor is within the 3′ portion of the Nrf2 ORF. •The translation of Nrf2 or eGFP is reduced by the regulatory element. •The translational repression can be reversed with synonymous codon substitutions. •The molecular mechanism requires the mRNA sequence, but not the encoded amino acids. -- Abstract: Nuclear Factor Erythroid 2-Related Factor 2 (Nrf2) is a transcription factor that is essential for the regulation of an effective antioxidant and detoxifying response. The regulation of its activity can occur at transcription, translation and post-translational levels. Evidence suggests that under environmental stressmore » conditions, new synthesis of Nrf2 is required – a process that is regulated by translational control and is not fully understood. Here we described the identification of a novel molecular process that under basal conditions strongly represses the translation of Nrf2 within the open reading frame (ORF). This mechanism is dependent on the mRNA sequence within the 3′ portion of the ORF of Nrf2 but not in the encoded amino acid sequence. The Nrf2 translational repression can be reversed with the use of synonymous codon substitutions. This discovery suggests an additional layer of control to explain the reason for the low Nrf2 concentration under quiescent state.« less

Adaptive induction of NF-E2-related factor-2-driven antioxidant genes in endothelial cells in response to hyperglycemia.

PubMed

Ungvari, Zoltan; Bailey-Downs, Lora; Gautam, Tripti; Jimenez, Rosario; Losonczy, Gyorgy; Zhang, Cuihua; Ballabh, Praveen; Recchia, Fabio A; Wilkerson, Donald C; Sonntag, William E; Pearson, Kevin; de Cabo, Rafael; Csiszar, Anna

2011-04-01

Hyperglycemia in diabetes mellitus promotes oxidative stress in endothelial cells, which contributes to development of cardiovascular diseases. Nuclear factor erythroid 2-related factor-2 (Nrf2) is a transcription factor activated by oxidative stress that regulates expression of numerous reactive oxygen species (ROS) detoxifying and antioxidant genes. This study was designed to elucidate the homeostatic role of adaptive induction of Nrf2-driven free radical detoxification mechanisms in endothelial protection under diabetic conditions. Using a Nrf2/antioxidant response element (ARE)-driven luciferase reporter gene assay we found that in a cultured coronary arterial endothelial cell model hyperglycemia (10-30 mmol/l glucose) significantly increases transcriptional activity of Nrf2 and upregulates the expression of the Nrf2 target genes NQO1, GCLC, and HMOX1. These effects of high glucose were significantly attenuated by small interfering RNA (siRNA) downregulation of Nrf2 or overexpression of Keap-1, which inactivates Nrf2. High-glucose-induced upregulation of NQO1, GCLC, and HMOX1 was also prevented by pretreatment with polyethylene glycol (PEG)-catalase or N-acetylcysteine, whereas administration of H(2)O(2) mimicked the effect of high glucose. To test the effects of metabolic stress in vivo, Nrf2(+/+) and Nrf2(-/-) mice were fed a high-fat diet (HFD). HFD elicited significant increases in mRNA expression of Gclc and Hmox1 in aortas of Nrf2(+/+) mice, but not Nrf2(-/-) mice, compared with respective standard diet-fed control mice. Additionally, HFD-induced increases in vascular ROS levels were significantly greater in Nrf2(-/-) than Nrf2(+/+) mice. HFD-induced endothelial dysfunction was more severe in Nrf2(-/-) mice, as shown by the significantly diminished acetylcholine-induced relaxation of aorta of these animals compared with HFD-fed Nrf2(+/+) mice. Our results suggest that adaptive activation of the Nrf2/ARE pathway confers endothelial protection under diabetic conditions.

Nuclear respiratory factor-1 and bioenergetics in tamoxifen-resistant breast cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Radde, Brandie N.; Ivanova, Margarita M.; Mai, Huy Xuan

Acquired tamoxifen (TAM) resistance is a significant clinical problem in treating patients with estrogen receptor α (ERα)+ breast cancer. We reported that ERα increases nuclear respiratory factor-1 (NRF-1), which regulates nuclear-encoded mitochondrial gene transcription, in MCF-7 breast cancer cells and NRF-1 knockdown stimulates apoptosis. Whether NRF-1 and target gene expression is altered in endocrine resistant breast cancer cells is unknown. We measured NRF-1and metabolic features in a cell model of progressive TAM-resistance. NRF-1 and its target mitochondrial transcription factor A (TFAM) were higher in TAM-resistant LCC2 and LCC9 cells than TAM-sensitive MCF-7 cells. Using extracellular flux assays we observed thatmore » LCC1, LCC2, and LCC9 cells showed similar oxygen consumption rate (OCR), but lower mitochondrial reserve capacity which was correlated with lower Succinate Dehydrogenase Complex, Subunit B in LCC1 and LCC2 cells. Complex III activity was lower in LCC9 than MCF-7 cells. LCC1, LCC2, and LCC9 cells had higher basal extracellular acidification (ECAR), indicating higher aerobic glycolysis, relative to MCF-7 cells. Mitochondrial bioenergetic responses to estradiol and 4-hydroxytamoxifen were reduced in the endocrine-resistant cells compared to MCF-7 cells. These results suggest the acquisition of altered metabolic phenotypes in response to long term antiestrogen treatment may increase vulnerability to metabolic stress. - Highlights: • NRF-1 and TFAM expression are higher in endocrine-resistant breast cancer cells. • Oxygen consumption rate is similar in endocrine-sensitive and resistant cells. • Mitochondrial reserve capacity is lower in endocrine-resistant cells. • Endocrine-resistant breast cancer cells have increased glycolysis. • Bioenergetic responses to E2 and tamoxifen are lower in endocrine-resistant cells.« less

AMPK activation represses the human gene promoter of the cardiac isoform of acetyl-CoA carboxylase: Role of nuclear respiratory factor-1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Adam, Tasneem; Opie, Lionel H.; Essop, M. Faadiel, E-mail: mfessop@sun.ac.za

Research highlights: {yields} AMPK inhibits acetyl-CoA carboxylase beta gene promoter activity. {yields} Nuclear respiratory factor-1 inhibits acetyl-CoA carboxylase beta promoter activity. {yields} AMPK regulates acetyl-CoA carboxylase beta at transcriptional level. -- Abstract: The cardiac-enriched isoform of acetyl-CoA carboxylase (ACC{beta}) produces malonyl-CoA, a potent inhibitor of carnitine palmitoyltransferase-1. AMPK inhibits ACC{beta} activity, lowering malonyl-CoA levels and promoting mitochondrial fatty acid {beta}-oxidation. Previously, AMPK increased promoter binding of nuclear respiratory factor-1 (NRF-1), a pivotal transcriptional modulator controlling gene expression of mitochondrial proteins. We therefore hypothesized that NRF-1 inhibits myocardial ACC{beta} promoter activity via AMPK activation. A human ACC{beta} promoter-luciferase construct was transientlymore » transfected into neonatal cardiomyocytes {+-} a NRF-1 expression construct. NRF-1 overexpression decreased ACC{beta} gene promoter activity by 71 {+-} 4.6% (p < 0.001 vs. control). Transfections with 5'-end serial promoter deletions revealed that NRF-1-mediated repression of ACC{beta} was abolished with a pPII{beta}-18/+65-Luc deletion construct. AMPK activation dose-dependently reduced ACC{beta} promoter activity, while NRF-1 addition did not further decrease it. We also investigated NRF-1 inhibition in the presence of upstream stimulatory factor 1 (USF1), a known transactivator of the human ACC{beta} gene promoter. Here NRF-1 blunted USF1-dependent induction of ACC{beta} promoter activity by 58 {+-} 7.5% (p < 0.001 vs. control), reversed with a dominant negative NRF-1 construct. NRF-1 also suppressed endogenous USF1 transcriptional activity by 55 {+-} 6.2% (p < 0.001 vs. control). This study demonstrates that NRF-1 is a novel transcriptional inhibitor of the human ACC{beta} gene promoter in the mammalian heart. Our data extends AMPK regulation of ACC{beta} to the transcriptional level.« less

An overview of the molecular mechanisms and novel roles of Nrf2 in neurodegenerative disorders.

PubMed

Yang, Yang; Jiang, Shuai; Yan, Juanjuan; Li, Yue; Xin, Zhenlong; Lin, Yan; Qu, Yan

2015-02-01

Recently, growing evidence has demonstrated that nuclear factor erythroid 2-related factor 2 (Nrf2) is a pivotal regulator of endogenous defense systems that function via the activation of a set of protective genes, and this is particularly clear in the central nervous system (CNS). Therefore, it is highly useful to summarize the current literature on the molecular mechanisms and role of Nrf2 in the CNS. In this review, we first briefly introduce the molecular features of Nrf2. We then discuss the regulation, cerebral actions, upstream modulators and downstream targets of Nrf2 pathway. Following this background, we expand our discussion to the role of Nrf2 in several major neurodegenerative disorders (NDDs) such as Alzheimer's disease, Parkinson's disease, Huntington's disease, multiple sclerosis and amyotrophic lateral sclerosis. Lastly, we discuss some potential future directions. The information reviewed here may be significant in the design of further experimental research and increase the potential of Nrf2 as a therapeutic target in the future. Copyright © 2014 Elsevier Ltd. All rights reserved.

Glycosylation enables aesculin to activate Nrf2.

PubMed

Kim, Kyun Ha; Park, Hyunsu; Park, Hee Jin; Choi, Kyoung-Hwa; Sadikot, Ruxana T; Cha, Jaeho; Joo, Myungsoo

2016-07-15

Since aesculin, 6,7-dihydroxycoumarin-6-O-β-glucopyranoside, suppresses inflammation, we asked whether its anti-inflammatory activity is associated with the activation of nuclear factor-E2-related factor 2 (Nrf2), a key anti-inflammatory factor. Our results, however, show that aesculin marginally activated Nrf2. Since glycosylation can enhance the function of a compound, we then asked whether adding a glucose makes aesculin activate Nrf2. Our results show that the glycosylated aesculin, 3-O-β-d-glycosyl aesculin, robustly activated Nrf2, inducing the expression of Nrf2-dependent genes, such as heme oxygenase-1, glutamate-cysteine ligase catalytic subunit, and NAD(P)H quinone oxidoreductase 1 in macrophages. Mechanistically, 3-O-β-d-glycosyl aesculin suppressed ubiquitination of Nrf2, retarding degradation of Nrf2. Unlike aesculin, 3-O-β-d-glycosyl aesculin significantly suppressed neutrophilic lung inflammation, a hallmark of acute lung injury (ALI), in mice, which was not recapitulated in Nrf2 knockout mice, suggesting that the anti-inflammatory function of the compound largely acts through Nrf2. In a mouse model of sepsis, a major cause of ALI, 3-O-β-d-glycosyl aesculin significantly enhanced the survival of mice, compared with aesculin. Together, these results show that glycosylation could confer the ability to activate Nrf2 on aesculin, enhancing the anti-inflammatory function of aesculin. These results suggest that glycosylation can be a way to improve or alter the function of aesculin.

Systemic administration of the apocarotenoid bixin protects skin against solar UV-induced damage through activation of NRF2.

PubMed

Tao, Shasha; Park, Sophia L; Rojo de la Vega, Montserrat; Zhang, Donna D; Wondrak, Georg T

2015-12-01

Exposure to solar ultraviolet (UV) radiation is a causative factor in skin photodamage and carcinogenesis, and an urgent need exists for improved molecular photoprotective strategies different from (or synergistic with) photon absorption. Recent studies suggest a photoprotective role of cutaneous gene expression orchestrated by the transcription factor NRF2 (nuclear factor-E2-related factor 2). Here we have explored the molecular mechanism underlying carotenoid-based systemic skin photoprotection in SKH-1 mice and provide genetic evidence that photoprotection achieved by the FDA-approved apocarotenoid and food additive bixin depends on NRF2 activation. Bixin activates NRF2 through the critical Cys-151 sensor residue in KEAP1, orchestrating a broad cytoprotective response in cultured human keratinocytes as revealed by antioxidant gene expression array analysis. Following dose optimization studies for cutaneous NRF2 activation by systemic administration of bixin, feasibility of bixin-based suppression of acute cutaneous photodamage from solar UV exposure was investigated in Nrf2(+/+) versus Nrf2(-/-) SKH-1 mice. Systemic administration of bixin suppressed skin photodamage, attenuating epidermal oxidative DNA damage and inflammatory responses in Nrf2(+/+) but not in Nrf2(-/-) mice, confirming the NRF2-dependence of bixin-based cytoprotection. Taken together, these data demonstrate feasibility of achieving NRF2-dependent cutaneous photoprotection by systemic administration of the apocarotenoid bixin, a natural food additive consumed worldwide. Copyright © 2015 Elsevier Inc. All rights reserved.

Bardoxolone Brings Nrf2-Based Therapies to Light

PubMed Central

2013-01-01

Abstract The targeted activation of nuclear factor erythroid-derived-2-like 2 (Nrf2) to alleviate symptoms of chronic kidney disease has recently garnered much attention. Unfortunately, the greatest clinical success to date, bardoxolone, failed in phase III clinical trial for unspecified safety reasons. The present letter to the editor discusses the clinical development of bardoxolone and explores potential reasons for the ultimate withdrawal from clinical trials. In particular, was the correct clinical indication pursued and would improved specificity have mitigated the safety concerns? Ultimately, it is concluded that the right clinical indication and heightened specificity will lead to successful Nrf2-based therapies. Therefore, the bardoxolone clinical results do not dampen enthusiasm for Nrf2-based therapies; rather it illuminates the clinical potential of the Nrf2 pathway as a drug target. Antioxid. Redox Signal. 19, 517–518. PMID:23227819

Suppression of nuclear factor erythroid‑2‑related factor 2‑mediated antioxidative defense in the lung injury induced by chronic exposure to methamphetamine in rats.

PubMed

Bai, Yang; Wang, Yun; Liu, Ming; Gu, Yu-Han; Jiang, Bin; Wu, Xu; Wang, Huai-Liang

2017-05-01

The imbalance between oxidative stress and antioxidant defense is important in the pathogenesis of lung diseases. Nuclear factor erythroid‑2‑related factor 2 (Nrf2) is a key transcriptional factor that regulates the antioxidant response. The purpose of the present study was to investigate whether Nrf2‑mediated antioxidative defense is involved in methamphetamine (MA)‑induced lung injury in rats. Following establishment of chronic MA toxicity in rats, Doppler ultrasonic detection was used to measure the changes of physiological indexes, followed by hematoxylin and eosin staining, ELISA and western blot analysis. MA was demonstrated to increase the heart rate and peak blood flow velocity of pulmonary arterial valves and to decrease the survival rate of rats, and resulted in lung injury characterized by perivascular exudates, airspace edema, slight hemorrhage and inflammatory cell infiltration. MA significantly inhibited the expression of nuclear Nrf2 protein and its target genes (glutamate‑cysteine ligase catalytic subunit C and heme oxygenase‑1), and dose‑dependently reduced glutathione (GSH) levels and the ratio of GSH/oxidized glutathione, accompanied by increases in reactive oxygen species (ROS) levels in rat lungs. Linear regression analysis revealed that there was a positive correlation between lung ROS level and lung injury indexes. These findings suggested that chronic exposure to MA led to lung injury by suppression of Nrf2‑mediated antioxidative defense, suggesting that Nrf2 may be an important therapeutic target for MA‑induced chronic lung toxicity.

Disruption of the Transcription Factor Nrf2 Promotes Pro-Oxidative Dendritic Cells That Stimulate Th2-Like Immunoresponsiveness upon Activation by Ambient Particulate Matter

PubMed Central

Williams, Marc A.; Rangasamy, Tirumalai; Bauer, Stephen M.; Killedar, Smruti; Karp, Matthew; Kensler, Thomas W.; Yamamoto, Masayuki; Breysse, Patrick; Biswal, Shyam; Georas, Steve N.

2011-01-01

Oxidative stress is important in dendritic cell (DC) activation. Environmental particulate matter (PM) directs pro-oxidant activities that may alter DC function. Nuclear erythroid 2 p45-related factor 2 (Nrf2) is a redox-sensitive transcription factor that regulates expression of antioxidant and detoxification genes. Oxidative stress and defective antioxidant responses may contribute to the exacerbations of asthma. We hypothesized that PM would impart differential responses by Nrf2 wild-type DCs as compared with Nrf2−/− DCs. We found that the deletion of Nrf2 affected important constitutive functions of both bone marrow-derived and highly purified myeloid lung DCs such as the secretion of inflammatory cytokines and their ability to take up exogenous Ag. Stimulation of Nrf2−/− DCs with PM augmented oxidative stress and cytokine production as compared with resting or Nrf2+/+ DCs. This was associated with the enhanced induction of Nrf2-regulated antioxidant genes. In contrast to Nrf2+/+ DCs, coincubation of Nrf2−/− DCs with PM and the antioxidant N-acetyl cysteine attenuated PM-induced up-regulation of CD80 and CD86. Our studies indicate a previously underappreciated role of Nrf2 in innate immunity and suggest that deficiency in Nrf2-dependent pathways may be involved in susceptibility to the adverse health effects of air pollution in part by promoting Th2 cytokine responses in the absence of functional Nrf2. Moreover, our studies have uncovered a hierarchal response to oxidative stress in terms of costimulatory molecule expression and cytokine secretion in DCs and suggest an important role of heightened oxidative stress in proallergic Th2-mediated immune responses orchestrated by DCs. PMID:18802057

The Nrf2-antioxidant response element pathway: a target for regulating energy metabolism

USDA-ARS?s Scientific Manuscript database

The nuclear factor E2-related factor 2 (Nrf2) is a transcription factor that responds to oxidative stress by binding to the antioxidant response element (ARE) in the promoter of genes coding for antioxidant enzymes like NAD(P)H:quinone oxidoreductase 1 (NQO1) and proteins for glutathione synthesis. ...

Nrf2 Deficiency in Dendritic Cells Enhances the Adjuvant Effect of Ambient Ultrafine Particles on Allergic Sensitization

EPA Science Inventory

Airborne particulate matter (PM) is an important risk factor for asthma. Generation of oxidative stress by PM-associated chemicals is a major mechanism of its health effects. Transcription factor nuclear factor (erythroid-derived 2)-like 2 (Nrf2) mediates antioxidant and phase II...

Iron-Inducible Nuclear Translocation of a Myb3 Transcription Factor in the Protozoan Parasite Trichomonas vaginalis

PubMed Central

Hsu, Hong-Ming; Lee, Yu; Indra, Dharmu; Wei, Shu-Yi; Liu, Hsing-Wei; Chang, Lung-Chun; Chen, Chinpan; Ong, Shiou-Jeng

2012-01-01

In Trichomonas vaginalis, a novel nuclear localization signal spanning the folded R2R3 DNA-binding domain of a Myb2 protein was previously identified. To study whether a similar signal is used for nuclear translocation by other Myb proteins, nuclear translocation of Myb3 was examined in this report. When overexpressed, hemagglutinin-tagged Myb3 was localized to nuclei of transfected cells, with a cellular distribution similar to that of endogenous Myb3. Fusion to a bacterial tetracycline repressor, R2R3, of Myb3 that spans amino acids (aa) 48 to 156 was insufficient for nuclear translocation of the fusion protein, unless its C terminus was extended to aa 167. The conserved isoleucine in helix 2 of R2R3, which is important for Myb2's structural integrity in maintaining DNA-binding activity and nuclear translocation, was also vital for the former activity of Myb3, but less crucial for the latter. Sequential nuclear influx and efflux of Myb3, which require further extension of the nuclear localization signal to aa 180, were immediately induced after iron repletion. Sequence elements that regulate nuclear translocation with cytoplasmic retention, nuclear influx, and nuclear efflux were identified within the C-terminal tail. These results suggest that the R2R3 DNA-binding domain also serves as a common module for the nuclear translocation of both Myb2 and Myb3, but there are intrinsic differences between the two nuclear localization signals. PMID:23042127

Nrf2-dependent suppression of azoxymethane/dextran sulfate sodium-induced colon carcinogenesis by the cinnamon-derived dietary factor cinnamaldehyde.

PubMed

Long, Min; Tao, Shasha; Rojo de la Vega, Montserrat; Jiang, Tao; Wen, Qing; Park, Sophia L; Zhang, Donna D; Wondrak, Georg T

2015-05-01

The progressive nature of colorectal cancer and poor prognosis associated with the metastatic phase of the disease create an urgent need for the development of more efficacious strategies targeting colorectal carcinogenesis. Cumulative evidence suggests that the redox-sensitive transcription factor Nrf2 (nuclear factor-E2-related factor 2), a master regulator of the cellular antioxidant defence, represents a promising molecular target for colorectal cancer chemoprevention. Recently, we have identified cinnamon, the ground bark of Cinnamomum aromaticum (cassia cinnamon) and Cinnamomum verum (Ceylon cinnamon), as a rich dietary source of the Nrf2 inducer cinnamaldehyde (CA) eliciting the Nrf2-regulated antioxidant response in human epithelial colon cells, conferring cytoprotection against electrophilic and genotoxic insult. Here, we have explored the molecular mechanism underlying CA-induced Nrf2 activation in colorectal epithelial cells and have examined the chemopreventive potential of CA in a murine colorectal cancer model comparing Nrf2(+/+) with Nrf2(-/-) mice. In HCT116 cells, CA caused a Keap1-C151-dependent increase in Nrf2 protein half-life via blockage of ubiquitination with upregulation of cytoprotective Nrf2 target genes and elevation of cellular glutathione. After optimizing colorectal Nrf2 activation and target gene expression by dietary CA-supplementation regimens, we demonstrated that CA suppresses AOM/DSS-induced inflammatory colon carcinogenesis with modulation of molecular markers of colorectal carcinogenesis. Dietary suppression of colorectal cancer using CA supplementation was achieved in Nrf2(+/+) but not in Nrf2(-/-) mice confirming the Nrf2 dependence of CA-induced chemopreventive effects. Taken together, our data suggest feasibility of colorectal cancer suppression by dietary CA, an FDA-approved food additive derived from the third most consumed spice in the world. ©2015 American Association for Cancer Research.

Role of platelet-released growth factors in detoxification of reactive oxygen species in osteoblasts.

PubMed

Tohidnezhad, Mersedeh; Wruck, Christoph-Jan; Slowik, Alexander; Kweider, Nisreen; Beckmann, Rainer; Bayer, Andreas; Houben, Astrid; Brandenburg, Lars-Ove; Varoga, Deike; Sönmez, Tolga-Taha; Stoffel, Marcus; Jahr, Holger; Lippross, Sebastian; Pufe, Thomas

2014-08-01

Oxidative stress can impair fracture healing. To protect against oxidative damage, a system of detoxifying and antioxidative enzymes works to reduce the cellular stress. The transcription of these enzymes is regulated by antioxidant response element (ARE). The nuclear factor (erythroid-derived 2)-like2 (Nrf2) plays a major role in transcriptional activation of ARE-driven genes. Recently it has been shown that vascular endothelial growth factor (VEGF) prevents oxidative damage via activation of the Nrf2 pathway in vitro. Platelet-released growth factor (PRGF) is a mixture of autologous proteins and growth factors, prepared from a determined volume of platelet-rich plasma (PRP). It has already used to enhance fracture healing in vitro. The aim of the present study was to elucidate if platelets can lead to upregulation of VEGF and if platelets can regulate the activity of Nrf2-ARE system in primary human osteoblast (hOB) and in osteoblast-like cell line (SAOS-2). Platelets and PRGF were obtained from healthy human donors. HOB and SAOS-2 osteosarcoma cell line were used. The ARE activity was analysed using a dual luciferase reporter assay system. We used Western blot to detect the nuclear accumulation of Nrf2 and the amount of cytosolic antioxidant Thioredoxin Reductase-1 (TXNRD-1), Heme Oxygenase-1 (HO-1) and NAD(P)H quinine oxidoreductase-1 (NQO1). Gene expression analysis was performed by real-time RT PCR. ELISA was used for the quantification of growth factors. The activity of ARE was increased in the presence of PRGF up to 50%. Western blotting demonstrated enhanced nuclear accumulation of Nrf2. This was followed by an increase in the protein expression of the aforementioned downstream targets of Nrf2. Real-time RT PCR data showed an upregulation in the gene expression of the VEGF after PRGF treatment. This was confirmed by ELISA, where the treatment with PRGF induced the protein level of VEGF in both cells. These results provide a new insight into PRGF's mode of action in osteoblasts. PRGF not only leads to increase the endogenous VEGF, but also it may be involved in preventing oxidative damage through the Nrf2-ARE signalling. Nrf2 activation via PRGF may have great potential as an effective therapeutic drug target in fracture healing. Copyright © 2014 Elsevier Inc. All rights reserved.

The aryl hydrocarbon receptor and estrogen receptor alpha differentially modulate nuclear factor erythroid-2-related factor 2 transactivation in MCF-7 breast cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lo, Raymond; Matthews, Jason, E-mail: jason.matthews@utoronto.ca

2013-07-15

Nuclear factor erythroid-2-related factor 2 (NRF2; NFE2L2) plays an important role in mediating cellular protection against reactive oxygen species. NRF2 signaling is positively modulated by the aryl hydrocarbon receptor (AHR) but inhibited by estrogen receptor alpha (ERα). In this study we investigated the crosstalk among NRF2, AHR and ERα in MCF-7 breast cancer cells treated with the NRF2 activator sulforaphane (SFN), a dual AHR and ERα activator, 3,3′-diindolylmethane (DIM), 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or 17β-estradiol (E2). SFN-dependent increases in NADPH-dependent oxidoreductase 1 (NQO1) and heme oxygenase I (HMOX1) mRNA levels were significantly reduced after co-treatment with E2. E2-dependent repression of NQO1 andmore » HMOX1 was associated with increased ERα but reduced p300 recruitment and reduced histone H3 acetylation at both genes. In contrast, DIM + SFN or TCDD + SFN induced NQO1 and HMOX1 mRNA expression to levels higher than SFN alone, which was prevented by RNAi-mediated knockdown of AHR. DIM + SFN but not TCDD + SFN also induced recruitment of ERα to NQO1 and HMOX1. However, the presence of AHR at NQO1 and HMOX1 restored p300 recruitment and histone H3 acetylation, thereby reversing the ERα-dependent repression of NRF2. Taken together, our study provides further evidence of functional interplay among NRF2, AHR and ERα signaling pathways through altered p300 recruitment to NRF2-regulated target genes. - Highlights: • We examined crosstalk among ERα, AHR, and NRF2 in MCF-7 breast cancer cells. • AHR enhanced the mRNA expression levels of two NRF2 target genes – HMOX1 and NQO1. • ERα repressed HMOX1 and NQO1 expression via decreased histone acetylation. • AHR prevented ERα-dependent repression of HMOX1 and NQO1.« less

The membrane-topogenic vectorial behaviour of Nrf1 controls its post-translational modification and transactivation activity.

PubMed

Zhang, Yiguo; Hayes, John D

2013-01-01

The integral membrane-bound Nrf1 transcription factor fulfils important functions in maintaining cellular homeostasis and organ integrity, but how it is controlled vectorially is unknown. Herein, creative use of Gal4-based reporter assays with protease protection assays (GRAPPA), and double fluorescence protease protection (dFPP), reveals that the membrane-topogenic vectorial behaviour of Nrf1 dictates its post-translational modification and transactivation activity. Nrf1 is integrated within endoplasmic reticulum (ER) membranes through its NHB1-associated TM1 in cooperation with other semihydrophobic amphipathic regions. The transactivation domains (TADs) of Nrf1, including its Asn/Ser/Thr-rich (NST) glycodomain, are transiently translocated into the ER lumen, where it is glycosylated in the presence of glucose to become a 120-kDa isoform. Thereafter, the NST-adjoining TADs are partially repartitioned out of membranes into the cyto/nucleoplasmic side, where Nrf1 is subject to deglycosylation and/or proteolysis to generate 95-kDa and 85-kDa isoforms. Therefore, the vectorial process of Nrf1 controls its target gene expression.

Increased Energy Expenditure, Ucp1 Expression, and Resistance to Diet-induced Obesity in Mice Lacking Nuclear Factor-Erythroid-2-related Transcription Factor-2 (Nrf2).

PubMed

Schneider, Kevin; Valdez, Joshua; Nguyen, Janice; Vawter, Marquis; Galke, Brandi; Kurtz, Theodore W; Chan, Jefferson Y

2016-04-01

The NRF2 (also known as NFE2L2) transcription factor is a critical regulator of genes involved in defense against oxidative stress. Previous studies suggest thatNrf2plays a role in adipogenesisin vitro, and deletion of theNrf2gene protects against diet-induced obesity in mice. Here, we demonstrate that resistance to diet-induced obesity inNrf2(-/-)mice is associated with a 20-30% increase in energy expenditure. Analysis of bioenergetics revealed thatNrf2(-/-)white adipose tissues exhibit greater oxygen consumption. White adipose tissue showed a >2-fold increase inUcp1gene expression. Oxygen consumption is also increased nearly 2.5-fold inNrf2-deficient fibroblasts. Oxidative stress induced by glucose oxidase resulted in increasedUcp1expression. Conversely, antioxidant chemicals (such asN-acetylcysteine and Mn(III)tetrakis(4-benzoic acid)porphyrin chloride) and SB203580 (a known suppressor ofUcp1expression) decreasedUcp1and oxygen consumption inNrf2-deficient fibroblasts. These findings suggest that increasing oxidative stress by limitingNrf2function in white adipocytes may be a novel means to modulate energy balance as a treatment of obesity and related clinical disorders. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Morris Water Maze Training in Mice Elevates Hippocampal Levels of Transcription Factors Nuclear Factor (Erythroid-derived 2)-like 2 and Nuclear Factor Kappa B p65

PubMed Central

Snow, Wanda M.; Pahlavan, Payam S.; Djordjevic, Jelena; McAllister, Danielle; Platt, Eric E.; Alashmali, Shoug; Bernstein, Michael J.; Suh, Miyoung; Albensi, Benedict C.

2015-01-01

Research has identified several transcription factors that regulate activity-dependent plasticity and memory, with cAMP-response element binding protein (CREB) being the most well-studied. In neurons, CREB activation is influenced by the transcription factor nuclear factor kappa B (NF-κB), considered central to immunity but more recently implicated in memory. The transcription factor early growth response-2 (Egr-2), an NF-κB gene target, is also associated with learning and memory. Nuclear factor (erythroid-derived 2)-like 2 (Nrf2), an antioxidant transcription factor linked to NF-κB in pathological conditions, has not been studied in normal memory. Given that numerous transcription factors implicated in activity-dependent plasticity demonstrate connections to NF-κB, this study simultaneously evaluated protein levels of NF-κB, CREB, Egr-2, Nrf2, and actin in hippocampi from young (1 month-old) weanling CD1 mice after training in the Morris water maze, a hippocampal-dependent spatial memory task. After a 6-day acquisition period, time to locate the hidden platform decreased in the Morris water maze. Mice spent more time in the target vs. non-target quadrants of the maze, suggestive of recall of the platform location. Western blot data revealed a decrease in NF-κB p50 protein after training relative to controls, whereas NF-κB p65, Nrf2 and actin increased. Nrf2 levels were correlated with platform crosses in nearly all tested animals. These data demonstrate that training in a spatial memory task results in alterations in and associations with particular transcription factors in the hippocampus, including upregulation of NF-κB p65 and Nrf2. Training-induced increases in actin protein levels caution against its use as a loading control in immunoblot studies examining activity-dependent plasticity, learning, and memory. PMID:26635523

Silencing Nrf2 impairs glioma cell proliferation via AMPK-activated mTOR inhibition

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jia, Yue; Wang, Handong, E-mail: njhdwang@hotmail.com; Wang, Qiang

Gliomas are the leading cause of death among adults with primary brain malignancies. Treatment for malignant gliomas remains limited, and targeted therapies have been incompletely explored. Nuclear factor erythroid 2-related factor 2 (Nrf2), a key transcription regulator for antioxidant and detoxification enzymes, is abundantly expressed in cancer cells. In this study, the role and mechanism of Nrf2 in cancer cell proliferation was investigated in multiple glioma cell lines. We first evaluated the expression patterns of Nrf2 in four glioma cell lines and found all four cell lines expressed Nrf2, but the highest level was observed in U251 cells. We further evaluatedmore » the biological functions of Nrf2 in U251 glioma cell proliferation by specific inhibition of Nrf2 using short hairpin RNA (shRNA). We found that Nrf2 depletion inhibited glioma cell proliferation. Nrf2 depletion also decreased colony formation in U251 cells stably expressing Nrf2 shRNA compared to scrambled control shRNA. Moreover, suppression of Nrf2 expression could lead to ATP depletion (with concomitant rise in AMP/ATP ratio) and consequently to AMPK-activated mTOR inhibition. Finally, activation of adenosine monophosphate–activated protein kinase (AMPK) by treated with phenformin, an AMPK agonist, can mimic the inhibitory effect of Nrf2 knockdown in U251 cells. In conclusion, our findings will shed light to the role and mechanism of Nrf2 in regulating glioma proliferation via ATP-depletion-induced AMPK activation and consequent mTOR inhibition, a novel insight into our understanding the role and mechanism of Nrf2 in glioma pathoetiology. To our knowledge, this is also the first report to provide a rationale for the implication of cross-linking between Nrf2 and mTOR signaling.« less

Partial contribution of the Keap1–Nrf2 system to cadmium-mediated metallothionein expression in vascular endothelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shinkai, Yasuhiro; Kimura, Tomoki; Itagaki, Ayaka

Cadmium is an environmental electrophile that modifies protein reactive thiols such as Kelch-like ECH-associated protein 1 (Keap1), a negative regulator of nuclear factor-erythroid 2-related factor 2 (Nrf2). In the present study, we investigated a role of the Keap1–Nrf2 system in cellular response to cadmium in vascular endothelial cells. Exposure of bovine aortic endothelial cells to cadmium resulted in modification of Keap1 and Nrf2 activation, thereby up-regulating not only its typical downstream proteins but also metallothionein-1/2. Experiments with siRNA-mediated knockdown of Nrf2 or Keap1 supported participation of the Keap1–Nrf2 system in the modulation of metallothionein-1/2 expression. Furthermore, chromatin immunoprecipitation assay showedmore » that Nrf2 was recruited to the antioxidant response element of the promoter region of the bovine metallothionein-2 gene in the presence of cadmium. These results suggest that the transcription factor Nrf2 plays, at least in part, a role in the changes in metallothionein expression mediated by exposure to cadmium. - Highlights: • Role of the Keap1–Nrf2 system in cellular response to cadmium was examined. • We used bovine aortic endothelial cells as a model of the vascular endothelium. • Exposure of cells to cadmium resulted in modification of Keap1 and Nrf2 activation. • Keap1–Nrf2 system participated in the modulation of metallothionein-1/2 expression. • Nrf2 was recruited to the antioxidant response element of MT2 promoter region.« less

Genetic silencing of Nrf2 enhances X-ROS in dysferlin-deficient muscle

PubMed Central

Kombairaju, Ponvijay; Kerr, Jaclyn P.; Roche, Joseph A.; Pratt, Stephen J. P.; Lovering, Richard M.; Sussan, Thomas E.; Kim, Jung-Hyun; Shi, Guoli; Biswal, Shyam; Ward, Christopher W.

2014-01-01

Oxidative stress is a critical disease modifier in the muscular dystrophies. Recently, we discovered a pathway by which mechanical stretch activates NADPH Oxidase 2 (Nox2) dependent ROS generation (X-ROS). Our work in dystrophic skeletal muscle revealed that X-ROS is excessive in dystrophin-deficient (mdx) skeletal muscle and contributes to muscle injury susceptibility, a hallmark of the dystrophic process. We also observed widespread alterations in the expression of genes associated with the X-ROS pathway and redox homeostasis in muscles from both Duchenne muscular dystrophy patients and mdx mice. As nuclear factor erythroid 2-related factor 2 (Nrf2) plays an essential role in the transcriptional regulation of genes involved in redox homeostasis, we hypothesized that Nrf2 deficiency may contribute to enhanced X-ROS signaling by reducing redox buffering. To directly test the effect of diminished Nrf2 activity, Nrf2 was genetically silenced in the A/J model of dysferlinopathy—a model with a mild histopathologic and functional phenotype. Nrf2-deficient A/J mice exhibited significant muscle-specific functional deficits, histopathologic abnormalities, and dramatically enhanced X-ROS compared to control A/J and WT mice, both with functional Nrf2. Having identified that reduced Nrf2 activity is a negative disease modifier, we propose that strategies targeting Nrf2 activation may address the generalized reduction in redox homeostasis to halt or slow dystrophic progression. PMID:24600403

Structure activity relationship of phenolic diterpenes from Salvia officinalis as activators of the nuclear factor E2-related factor 2 pathway

PubMed Central

Fischedick, Justin T; Standiford, Miranda; Johnson, Delinda A.; Johnson, Jeffrey A.

2013-01-01

Nuclear factor E2-related factor 2 (Nrf2) is a transcription factor known to activate cytoprotective genes which may be useful in the treatment of neurodegenerative disease. In order to better understand the structure activity relationship of phenolic diterpenes from Salvia officinalis L., we isolated carnosic acid, carnosol, epirosmanol, rosmanol, 12-methoxy-carnosic acid, sageone, and carnosaldehyde using polyamide column, centrifugal partition chromatography, and semi-preparative high performance liquid chromatography. Isolated compounds were screened in-vitro for their ability to active the Nrf2 and general cellular toxicity using mouse primary cortical cultures. All compounds except 12-methoxy-carnosic acid were able to activate the antioxidant response element. Furthermore both carnosol and carnoasldehyde were able to induce Nrf2-dependent gene expression as well as protect mouse primary cortical neuronal cultures from H2O2 induced cell death. PMID:23507152

Resveratrol protects primary rat hepatocytes against oxidative stress damage: activation of the Nrf2 transcription factor and augmented activities of antioxidant enzymes.

PubMed

Rubiolo, Juan Andrés; Mithieux, Gilles; Vega, Félix Victor

2008-09-04

Oxidative stress is recognized as an important factor in the development of liver pathologies. The reactive oxygen species endogenously generated or as a consequence of xenobiotic metabolism are eliminated by enzymatic and nonenzymatic cellular systems. Besides endogen defences, the antioxidant consumption in the diet has an important role in the protection against the development of diseases product of oxidative damage. Resveratrol is a naturally occurring compound which is part of the human diet. This molecule has been shown to have many biological properties, including antioxidant activity. We decided to test if resveratrol could protect primary hepatocytes in culture from oxidative stress damage and if so, to determine if this compound affects the cellular detoxifying systems and their regulation through the Nrf2 transcription factor that regulates the expression of antioxidant and phase II detoxifying enzymes. Cell death by necrosis was detected by measuring the activity of lactate dehydrogenase liberated to the medium. The activities of antioxidant and phase II enzymes were measured using previously described methods. Activation of the Nrf2 transcription factor was studied by confocal microscopy and the Nrf2 and its coding mRNA levels were determined by western blot and quantitative PCR respectively. Resveratrol pre-treatment effectively protected hepatocytes in culture exposed to oxidative stress, increasing the activities of catalase, superoxide dismutase, glutathione peroxidase, NADPH quinone oxidoreductase and glutathione-S-transferase. Resveratrol increases the level of Nrf2 and induces its translocation to the nucleus. Also, it increases the concentration of the coding mRNA for Nrf2. In this work we show that resveratrol could be a useful drug for the protection of liver cells from oxidative stress induced damage.

NAD(P)H:quinone oxidoreductase 1 activity reduces hypertrophy in 3T3-L1 adipocytes

USDA-ARS?s Scientific Manuscript database

The nuclear factor E2-related factor 2 (Nrf2)/Kelch-like ECH-associated protein 1 (Keap1) pathway responds to oxidative stress via control of the expression of several antioxidant genes. Recent efforts demonstrate that Nrf2 modulates development of adiposity and adipogenesis. However little is kno...

The regulatory role of Nrf2 in antioxidants phase2 enzymes and IL-17A expression in patients with ulcerative colitis.

PubMed

Sabzevary-Ghahfarokhi, Milad; Shohan, Mojtaba; Shirzad, Hedayatollah; Rahimian, Ghorbanali; Soltani, Amin; Ghatreh-Samani, Mahdi; Deris, Fatemeh; Bagheri, Nader; Shafigh, Mohammedhadi; Tahmasbi, Kamran

2018-06-22

Reactive oxygen species (ROS) is one of the pathogenic factors responsible for intestinal injury in Ulcerative colitis (UC). Nuclear factor erythroid-2 related factor 2 (Nrf2) plays a critical role against ROS factors to conserve epithelial integrity. This study aimed to localize Nrf2 and IL-17A protein in the inflamed mucosa of patients with ulcerative colitis. The gene expression of Nrf2 was also correlated with GST-A4 and PRDX1. A total of 20 patients and 20 healthy controls with definite UC based on the clinical criteria were enrolled for this study. The expression pattern of Nrf2 and IL-17A protein was compared in inflamed and non-inflamed colonic biopsies by immunohistochemical staining. Nrf2, GST-A4 and PRDX1 gene expression were determined by real-time polymerase chain reaction (RT-PCR). In inflamed colonic biopsies, an increased level of Nrf2 protein factor was detected in epithelial cells. Conversely, IL-17A protein was presented more in mononuclear cells in mucosa and lamina propria regions. A significant increase of Nrf2, GST-A4 gene expression was observed in both mild and severe patients with ulcerative colitis. GST-A4 gene expression indicated a high exponential rate in logistic regression. Oxidative stress in inflamed colonic tissue can induce Nrf2 gene expression. The performance of Nrf2 transcription factor may lead to the induction of GST-A4 and PRDX1. IL-17A is less detected in intestinal inflammation, presenting Nrf2 factor. The present findings suggest that Nrf2 function in the gut plays a role in arresting both inflammatory response and oxidative damages of UC. Copyright © 2018 Elsevier GmbH. All rights reserved.

Absence of Nrf2 or Its Selective Overexpression in Neurons and Muscle Does Not Affect Survival in ALS-Linked Mutant hSOD1 Mouse Models

PubMed Central

Vargas, Marcelo R.; Burton, Neal C.; Gan, Li; Johnson, Delinda A.; Schäfer, Matthias; Werner, Sabine; Johnson, Jeffrey A.

2013-01-01

The nuclear factor erythroid 2-related factor 2 (Nrf2) governs the expression of antioxidant and phase II detoxifying enzymes. Nrf2 activation can prevent or reduce cellular damage associated with several types of injury in many different tissues and organs. Dominant mutations in Cu/Zn-superoxide dismutase (SOD1) cause familial forms of amyotrophic lateral sclerosis (ALS), a fatal disorder characterized by the progressive loss of motor neurons and subsequent muscular atrophy. We have previously shown that Nrf2 activation in astrocytes delays neurodegeneration in ALS mouse models. To further investigate the role of Nrf2 in ALS we determined the effect of absence of Nrf2 or its restricted overexpression in neurons or type II skeletal muscle fibers on symptoms onset and survival in mutant hSOD1 expressing mice. We did not observe any detrimental effect associated with the lack of Nrf2 in two different mutant hSOD1 animal models of ALS. However, restricted Nrf2 overexpression in neurons or type II skeletal muscle fibers delayed disease onset but failed to extend survival in hSOD1G93A mice. These results highlight the concept that not only the pharmacological target but also the cell type targeted may be relevant when considering a Nrf2-mediated therapeutic approach for ALS. PMID:23418589

Absence of Nrf2 or its selective overexpression in neurons and muscle does not affect survival in ALS-linked mutant hSOD1 mouse models.

PubMed

Vargas, Marcelo R; Burton, Neal C; Kutzke, Jennifer; Gan, Li; Johnson, Delinda A; Schäfer, Matthias; Werner, Sabine; Johnson, Jeffrey A

2013-01-01

The nuclear factor erythroid 2-related factor 2 (Nrf2) governs the expression of antioxidant and phase II detoxifying enzymes. Nrf2 activation can prevent or reduce cellular damage associated with several types of injury in many different tissues and organs. Dominant mutations in Cu/Zn-superoxide dismutase (SOD1) cause familial forms of amyotrophic lateral sclerosis (ALS), a fatal disorder characterized by the progressive loss of motor neurons and subsequent muscular atrophy. We have previously shown that Nrf2 activation in astrocytes delays neurodegeneration in ALS mouse models. To further investigate the role of Nrf2 in ALS we determined the effect of absence of Nrf2 or its restricted overexpression in neurons or type II skeletal muscle fibers on symptoms onset and survival in mutant hSOD1 expressing mice. We did not observe any detrimental effect associated with the lack of Nrf2 in two different mutant hSOD1 animal models of ALS. However, restricted Nrf2 overexpression in neurons or type II skeletal muscle fibers delayed disease onset but failed to extend survival in hSOD1(G93A) mice. These results highlight the concept that not only the pharmacological target but also the cell type targeted may be relevant when considering a Nrf2-mediated therapeutic approach for ALS.

Adipose Deficiency of Nrf2 in ob/ob Mice Results in Severe Metabolic Syndrome

PubMed Central

Xue, Peng; Hou, Yongyong; Chen, Yanyan; Yang, Bei; Fu, Jingqi; Zheng, Hongzhi; Yarborough, Kathy; Woods, Courtney G.; Liu, Dianxin; Yamamoto, Masayuki; Zhang, Qiang; Andersen, Melvin E.; Pi, Jingbo

2013-01-01

Nuclear factor E2–related factor 2 (Nrf2) is a transcription factor that functions as a master regulator of the cellular adaptive response to oxidative stress. Our previous studies showed that Nrf2 plays a critical role in adipogenesis by regulating expression of CCAAT/enhancer-binding protein β and peroxisome proliferator–activated receptor γ. To determine the role of Nrf2 in the development of obesity and associated metabolic disorders, the incidence of metabolic syndrome was assessed in whole-body or adipocyte-specific Nrf2-knockout mice on a leptin-deficient ob/ob background, a model with an extremely positive energy balance. On the ob/ob background, ablation of Nrf2, globally or specifically in adipocytes, led to reduced white adipose tissue (WAT) mass, but resulted in an even more severe metabolic syndrome with aggravated insulin resistance, hyperglycemia, and hypertriglyceridemia. Compared with wild-type mice, WAT of ob/ob mice expressed substantially higher levels of many genes related to antioxidant response, inflammation, adipogenesis, lipogenesis, glucose uptake, and lipid transport. Absence of Nrf2 in WAT resulted in reduced expression of most of these factors at mRNA or protein levels. Our findings support a novel role for Nrf2 in regulating adipose development and function, by which Nrf2 controls the capacity of WAT expansion and insulin sensitivity and maintains glucose and lipid homeostasis. PMID:23238296

An Essential Role of NRF2 in Diabetic Wound Healing

PubMed Central

Long, Min; Rojo de la Vega, Montserrat; Wen, Qing; Bharara, Manish; Jiang, Tao; Zhang, Rui; Zhou, Shiwen; Wong, Pak K.

2016-01-01

The high mortality and disability of diabetic nonhealing skin ulcers create an urgent need for the development of more efficacious strategies targeting diabetic wound healing. In the current study, using human clinical specimens, we show that perilesional skin tissues from patients with diabetes are under more severe oxidative stress and display higher activation of the nuclear factor-E2–related factor 2 (NRF2)–mediated antioxidant response than perilesional skin tissues from normoglycemic patients. In a streptozotocin-induced diabetes mouse model, Nrf2−/− mice have delayed wound closure rates compared with Nrf2+/+ mice, which is, at least partially, due to greater oxidative DNA damage, low transforming growth factor-β1 (TGF-β1) and high matrix metalloproteinase 9 (MMP9) expression, and increased apoptosis. More importantly, pharmacological activation of the NRF2 pathway significantly improves diabetic wound healing. In vitro experiments in human immortalized keratinocyte cells confirm that NRF2 contributes to wound healing by alleviating oxidative stress, increasing proliferation and migration, decreasing apoptosis, and increasing the expression of TGF-β1 and lowering MMP9 under high-glucose conditions. This study indicates an essential role for NRF2 in diabetic wound healing and the therapeutic benefits of activating NRF2 in this disease, laying the foundation for future clinical trials using NRF2 activators in treating diabetic skin ulcers. PMID:26718502

Lack of association between nuclear factor erythroid-derived 2-like 2 promoter gene polymorphisms and oxidative stress biomarkers in amyotrophic lateral sclerosis patients.

PubMed

LoGerfo, Annalisa; Chico, Lucia; Borgia, Loredana; Petrozzi, Lucia; Rocchi, Anna; D'Amelio, Antonia; Carlesi, Cecilia; Caldarazzo Ienco, Elena; Mancuso, Michelangelo; Siciliano, Gabriele

2014-01-01

Oxidative stress involvement has been strongly hypothesized among the possible pathogenic mechanisms of motor neuron degeneration in amyotrophic lateral sclerosis (ALS). The intracellular redox balance is finely modulated by numerous complex mechanisms critical for cellular functions, among which the nuclear factor erythroid-derived 2-like 2 (NFE2L2/Nrf2) pathways. We genotyped, in a cohort of ALS patients (n = 145) and healthy controls (n = 168), three SNPs in Nrf2 gene promoter: -653 A/G, -651 G/A, and -617 C/A and evaluated, in a subset (n = 73) of patients, advanced oxidation protein products (AOPP), iron-reducing ability of plasma (FRAP), and plasma thiols (-SH) as oxidative damage peripheral biomarkers. Nrf2 polymorphisms were not different among patients and controls. Increased levels of AOPP (P < 0.05) and decreased levels of FRAP (P < 0.001) have been observed in ALS patients compared with controls, but no difference in -SH values was found. Furthermore, no association was found between biochemical markers of redox balance and Nrf2 polymorphisms. These data confirm an altered redox balance in ALS and indicate that, while being abnormally modified compared to controls, the oxidative stress biomarkers assessed in this study are independent from the -653 A/G, -651 G/A, and -617 C/A Nrf2 SNPs in ALS patients.

Identification of an Unfavorable Immune Signature in Advanced Lung Tumors from Nrf2-Deficient Mice.

PubMed

Zhang, Di; Rennhack, Jonathan; Andrechek, Eran R; Rockwell, Cheryl E; Liby, Karen T

2018-04-16

Activation of the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) pathway in normal cells inhibits carcinogenesis, whereas constitutive activation of Nrf2 in cancer cells promotes tumor growth and chemoresistance. However, the effects of Nrf2 activation in immune cells during lung carcinogenesis are poorly defined and could either promote or inhibit cancer growth. Our studies were designed to evaluate tumor burden and identify immune cell populations in the lungs of Nrf2 knockout (KO) versus wild-type (WT) mice challenged with vinyl carbamate. Nrf2 KO mice developed lung tumors earlier than the WT mice and exhibited more and larger tumors over time, even at late stages. T cell populations were lower in the lungs of Nrf2 KO mice, whereas tumor-promoting macrophages and myeloid-derived suppressor cells were elevated in the lungs and spleen, respectively, of Nrf2 KO mice relative to WT mice. Moreover, 34 immune response genes were significantly upregulated in tumors from Nrf2 KO mice, especially a series of cytokines (Cxcl1, Csf1, Ccl9, Cxcl12, etc.) and major histocompatibility complex antigens that promote tumor growth. Our studies discovered a novel immune signature, characterized by the infiltration of tumor-promoting immune cells, elevated cytokines, and increased expression of immune response genes in the lungs and tumors of Nrf2 KO mice. A complementary profile was also found in lung cancer patients, supporting the clinical significance of our findings. Overall, our results confirmed a protective role for Nrf2 in late-stage carcinogenesis and, unexpectedly, suggest that activation of Nrf2 in immune cells may be advantageous for preventing or treating lung cancer. Antioxid. Redox Signal. 00, 000-000.

Nuclear EGFRvIII resists hypoxic microenvironment induced apoptosis via recruiting ERK1/2 nuclear translocation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xie, Hui; Yang, Jinfeng; Xing, Wenjing

2016-02-05

Glioblastoma (GBM) is the most aggressive type of primary brain tumor. Its interaction with the tumor microenvironment promotes tumor progression. Furthermore, GBM bearing expression of EGFRvIII displays more adaptation to tumor microenvironment related stress. But the mechanisms were poorly understood. Here, we presented evidence that in the human U87MG glioblastoma tumor model, EGFRvIII overexpression led aberrant kinase activation and nuclear translocation of EGFRvIII/ERK1/2 under hypoxia, which induced growth advantage by resisting apoptosis. Additionally, EGFRvIII defective in nuclear entry impaired this capacity in hypoxia adaptation, and partially interrupted ERK1/2 nuclear translocation. Pharmacology or genetic interference ERK1/2 decreased hypoxia resistance triggered bymore » EGFRvIII expression, but not EGFRvIII nuclear translocation. In summary, this study identified a novel role for EGFRvIII in hypoxia tolerance, supporting an important link between hypoxia and subcellular localization alterations of the receptor. - Highlights: • Nuclear translocation of EGFRvIII contributes to GBM cell apoptotic resistance by hypoxia. • Nuclear ERK1/2 facilitates EGFRvIII in hypoxia resistance. • EGFRvIII nuclear translocation is not dependent on ERK1/2.« less

Internal Pudental Artery Dysfunction in Diabetes Mellitus Is Mediated by NOX1-Derived ROS-, Nrf2-, and Rho Kinase-Dependent Mechanisms.

PubMed

Alves-Lopes, Rhéure; Neves, Karla B; Montezano, Augusto C; Harvey, Adam; Carneiro, Fernando S; Touyz, Rhian M; Tostes, Rita C

2016-10-01

Oxidative stress plays an important role in diabetes mellitus (DM)-associated vascular injury. DM is an important risk factor for erectile dysfunction. Functional and structural changes in internal pudendal arteries (IPA) can lead to erectile dysfunction. We hypothesized that downregulation of nuclear factor E2-related factor 2 (Nrf2), consequent to increased nicotinamide adenine dinucleotide phosphate oxidase 1 (NOX1)-derived reactive oxygen species (ROS), impairs IPA function in DM. IPA and vascular smooth muscle cells from C57BL/6 (control) and NOX1 knockout mice were used. DM was induced by streptozotocin in C57BL/6 mice. Functional properties of IPA were assessed using a myograph, protein expression and peroxiredoxin oxidation by Western blot, RNA expression by polymerase chain reaction, carbonylation by oxyblot assay, ROS generation by lucigenin, nitrotyrosine, and amplex red, and Rho kinase activity and nuclear accumulation of Nrf2 by ELISA. IPA from diabetic mice displayed increased contractions to phenylephrine (control 138.5±9.5 versus DM 191.8±15.5). ROS scavenger, Nrf2 activator, NOX1 and Rho kinase inhibitors normalized vascular function. High glucose increased ROS generation in IPA vascular smooth muscle cell. This effect was abrogated by Nrf2 activation and not observed in NOX1 knockout vascular smooth muscle cell. High glucose also increased levels of nitrotyrosine, protein oxidation/carbonylation, and Rho kinase activity, but reduced Nrf2 activity and expression of Nrf2-regulated genes (catalase [25.6±0.05%], heme oxygenase-1 [21±0.1%], and quinone oxidoreductase 1 [22±0.1%]) and hydrogen peroxide levels. These effects were not observed in vascular smooth muscle cell from NOX1 knockout mice. In these cells, high glucose increased hydrogen peroxide levels. In conclusion, Rho kinase activation, via NOX1-derived ROS and downregulation of Nrf2 system, impairs IPA function in DM. These data suggest that Nrf2 is vasoprotective in DM-associated erectile dysfunction. © 2016 American Heart Association, Inc.

Nrf2-Dependent and -Independent Effects of tert-Butylhydroquinone, CDDO-Im, and H2O2 in Human Jurkat T Cells as Determined by CRISPR/Cas9 Gene Editing.

PubMed

Zagorski, Joseph W; Maser, Tyler P; Liby, Karen T; Rockwell, Cheryl E

2017-05-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a stress-activated transcription factor activated by stimuli such as electrophilic compounds and other reactive xenobiotics. Previously, we have shown that the commonly used food additive and Nrf2 activator tert -butylhydroquinone (tBHQ) suppresses interleukin-2 (IL-2) production, CD25 expression, and NF κ B activity in human Jurkat T cells. The purpose of the current studies was to determine whether these effects were dependent upon Nrf2 by developing a human Nrf2-null T cell model using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 technology. The current studies show that suppression of CD25 expression by tBHQ is partially dependent on Nrf2, whereas inhibition of IL-2 secretion is largely Nrf2-independent. Interestingly, tBHQ inhibited NF κ B activation in an Nrf2-independent manner. This was an unexpected finding since Nrf2 inhibits NF κ B activation in other models. These results led us to investigate another more potent Nrf2 activator, the synthetic triterpenoid 1[2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oyl]imidazole (CDDO-Im). Treatment of wild-type and Nrf2-null Jurkat T cells with CDDO-Im resulted in an Nrf2-dependent suppression of IL-2. Furthermore, susceptibility to reactive oxygen species was significantly enhanced in the Nrf2-null clones as determined by decreased mitochondrial membrane potential and cell viability. Importantly, this study is the first to describe the generation of a human Nrf2-null model, which is likely to have multiple applications in immunology and cancer biology. Collectively, this study demonstrates a role for Nrf2 in the effects of CDDO-Im on CD25 and IL-2 expression, whereas the effect of tBHQ on these parameters is complex and likely involves modulation of multiple stress-activated transcription factors, including NF κ B and Nrf2. Copyright © 2017 by The American Society for Pharmacology and Experimental Therapeutics.

Impact of Nrf2 on UVB-induced skin inflammation/photoprotection and photoprotective effect of sulforaphane.

PubMed

Saw, Constance L; Huang, Mou-Tuan; Liu, Yue; Khor, Tin Oo; Conney, Allan H; Kong, Ah-Ng

2011-06-01

Ultraviolet (UV) of sunlight is a complete carcinogen that can burn skin, enhance inflammation, and drive skin carcinogenesis. Previously, we have shown that sulforaphane (SFN) inhibited chemically induced skin carcinogenesis via nuclear factor (erythroid-derived 2)-like 2 (Nrf2) and others have shown that broccoli sprout extracts containing high SFN protected against UV-induced skin carcinogenesis in SKH-1 hairless mice. A recent study showed that there was no difference between Nrf2 knockout (Nrf2 KO) and Nrf2 wild-type (WT) BALB/C mice after exposing to high dose of UVB. Since Nrf2 plays critical roles in the anti-oxidative stress/anti-inflammatory responses, it is relevant to assess the role of Nrf2 for photoprotection against UV. In this context, the role of Nrf2 in UVB-induced skin inflammation in Nrf2 WT and Nrf2 KO C57BL/6 mice was studied. A single dose of UVB (300 mJ/cm(2)) resulted in skin inflammation in both WT and Nrf2 KO (-/-) mice (KO mice) at 8 h and 8 d following UVB irradiation. In the WT mice inflammation returned to the basal level to a greater extent when compared to the KO mice. SFN treatment of Nrf2 WT but not Nrf2 KO mice restored the number of sunburn cells back to their basal level by 8 d after UVB irradiation. Additionally, UVB-induced short-term inflammatory biomarkers (interleukin-1β and interleukin-6) were increased in the KO mice and UVB-induced apoptotic cells in the KO mice were significantly higher as compared to that in the WT. Taken together, our results show that functional Nrf2 confers a protective effect against UVB-induced inflammation, sunburn reaction, and SFN-mediated photoprotective effects in the skin. Copyright © 2010 Wiley-Liss, Inc.

Nuclear factor erythroid-derived 2-related factor 2 activates glutathione S-transferase expression in the midgut of Spodoptera litura (Lepidoptera: Noctuidae) in response to phytochemicals and insecticides.

PubMed

Chen, S; Lu, M; Zhang, N; Zou, X; Mo, M; Zheng, S

2018-05-10

Detoxication enzymes play an important role in insect resistance to xenobiotics such as insecticides and phytochemicals. We studied the pathway for activating the expression of glutathione S-transferases (GSTs) in response to selected xenobiotics. An assay of the promoter activity of GST epsilon 1 (Slgste1) of Spodoptera litura led to the discovery of a cis-regulating element. An antioxidant response element was activated in response to indole-3-carbinol (I3C) and chlorpyrifos (CPF) and was able to bind with the xenobiotic sensor protein nuclear factor erythroid-derived 2-related factor 2 (SlNrf2). SlNrf2 and Slgste1 were responsive to reactive oxygen species induced by I3C and CPF in a S. litura cell line, as well as in S. litura midguts. SlNrf2 RNA interference (RNAi) reduced the message RNA levels of Slgste1 and the peroxidase activity of GSTs in response to I3C, xanthotoxin, CPF and deltamethrin. SlNrf2 RNAi and inhibitor treatment of GST activity decreased the viability of I3C-treated cells. These results indicate that SlNrf2 activates the expression of GSTs in response to oxidative stresses caused by exposure to xenobiotics. © 2018 The Royal Entomological Society.

Alteration of Nrf2 and Glutamate Cysteine Ligase expression contribute to lesions growth and fibrogenesis in ectopic endometriosis.

PubMed

Marcellin, L; Santulli, P; Chouzenoux, S; Cerles, O; Nicco, C; Dousset, B; Pallardy, M; Kerdine-Römer, S; Just, P A; Chapron, C; Batteux, F

2017-09-01

The redox-sensitive nuclear factor erythroid-derived 2-like 2 (NRF2) controls endogenous antioxidant enzymes' transcription and protects against oxidative damage which is triggered by inflammation and known to favor progression of endometriosis. Glutamate Cysteine Ligase (GCL), a target gene of NRF2, is the first enzyme in the synthesis cascade of glutathione, an important endogenous antioxidant. Sixty-one patients, with thorough surgical examination of the abdominopelvic cavity, were recruited for the study: 31 with histologically-proven endometriosis and 30 disease-free women taken as controls. Expressions of NRF2 and GCL were investigated by quantitative RT-PCR and immunohistochemistry in eutopic and ectopic endometria from endometriosis-affected women and in endometrium of disease-free women. Ex vivo stromal and epithelial cells were extracted and purified from endometrial and endometriotic biopsies to explore expression of NRF2 and GCL in both stromal and epithelial compartments by western blot. Finally, in order to strengthen the role of NRF2 in endometriosis pathogenesis, we evaluated the drop of NRF2 expression in a mouse model of endometriosis using NRF2 knockout (NRF2 -/- ) mice. The mRNA levels of NRF2 and GCL were significantly lower in ectopic endometria of endometriosis-affected women compared to eutopic endometria of disease-free women. The immunohistochemical analysis confirmed the decreased expression of both NRF2 and GCL in ectopic endometriotic tissues compared to eutopic endometria of endometriosis-affected and disease-free women. Immunoblotting revealed a significant decreased of NRF2 and GCL expression in epithelial and stroma cells from ectopic lesions of endometriosis-affected women compared to eutopic endometria from controls. Using a murine model of endometriosis, NRF2 -/- implants were more fibrotic compared to wild-type with an increased weight and volume. These findings indicate that expression of the transcription factor NRF2 and its effector GCL are both profoundly deregulated in endometriotic lesions towards increased growth and fibrogenetic processes. Copyright © 2017 Elsevier Inc. All rights reserved.

Metallothionein Is Downstream of Nrf2 and Partially Mediates Sulforaphane Prevention of Diabetic Cardiomyopathy.

PubMed

Gu, Junlian; Cheng, Yanli; Wu, Hao; Kong, Lili; Wang, Shudong; Xu, Zheng; Zhang, Zhiguo; Tan, Yi; Keller, Bradley B; Zhou, Honglan; Wang, Yuehui; Xu, Zhonggao; Cai, Lu

2017-02-01

We have reported that sulforaphane (SFN) prevented diabetic cardiomyopathy in both type 1 and type 2 diabetes (T2DM) animal models via the upregulation of nuclear transcription factor erythroid 2-related factor 2 (Nrf2) and metallothionein (MT). In this study, we tested whether SFN protects the heart from T2DM directly through Nrf2, MT, or both. Using Nrf2-knockout (KO), MT-KO, and wild-type (WT) mice, T2DM was induced by feeding a high-fat diet for 3 months followed by a small dose of streptozotocin. Age-matched controls were given a normal diet. Both T2DM and control mice were then treated with or without SFN for 4 months by continually feeding a high-fat or normal diet. SFN prevented diabetes-induced cardiac dysfunction as well as diabetes-associated cardiac oxidative damage, inflammation, fibrosis, and hypertrophy, with increases in Nrf2 and MT expressions in the WT mice. Both Nrf2-KO and MT-KO diabetic mice exhibited greater cardiac damage than WT diabetic mice. SFN did not provide cardiac protection in Nrf2-KO mice, but partially or completely protected the heart from diabetes in MT-KO mice. SFN did not induce MT expression in Nrf2-KO mice, but stimulated Nrf2 function in MT-KO mice. These results suggest that Nrf2 plays the indispensable role for SFN cardiac protection from T2DM with significant induction of MT and other antioxidants. MT expression induced by SFN is Nrf2 dependent, but is not indispensable for SFN-induced cardiac protection from T2DM. © 2017 by the American Diabetes Association.

Dihydro-CDDO-trifluoroethyl amide suppresses inflammatory responses in macrophages via activation of Nrf2

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Bin; Department of Cell Biology and Anatomy, University of South Carolina School of Medicine, Columbia, SC 29208; Abdalrahman, Akram

2014-02-21

Highlights: • Dh404 suppresses the expression of a selected set of pro-inflammatory cytokines in inflamed macrophages via activating Nrf2. • Dh404 activates Nrf2 while keeping Keap1 function intact in macrophages. • Dh404 minimally regulates NF-κB pathway in macrophages. - Abstract: Nuclear factor erythroid 2-related factor (Nrf2) is the major regulator of cellular defenses against various pathological stresses in a variety of organ systems, thus Nrf2 has evolved to be an attractive drug target for the treatment and/or prevention of human disease. Several synthetic oleanolic triterpenoids including dihydro-CDDO-trifluoroethyl amide (dh404) appear to be potent activators of Nrf2 and exhibit chemopreventive promisesmore » in multiple disease models. While the pharmacological efficacy of Nrf2 activators may be dependent on the nature of Nrf2 activation in specific cell types of target organs, the precise role of Nrf2 in mediating biological effects of Nrf2 activating compounds in various cell types remains to be further explored. Herein we report a unique and Nrf2-dependent anti-inflammatory profile of dh404 in inflamed macrophages. In lipopolysaccharide (LPS)-inflamed RAW264.7 macrophages, dh404 dramatically suppressed the expression of pro-inflammatory cytokines including inducible nitric oxide synthase (iNOS), monocyte chemotactic protein-1 (MCP-1), and macrophage inflammatory protein-1 beta (MIP-1β), while minimally regulating the expression of interleulin-6 (IL-6), IL-1β, and tumor necrosis factor alpha (TNFα). Dh404 potently activated Nrf2 signaling; however, it did not affect LPS-induced NF-κB activity. Dh404 did not interrupt the interaction of Nrf2 with its endogenous inhibitor Kelch-like ECH associating protein 1 (Keap1) in macrophages. Moreover, knockout of Nrf2 blocked the dh404-induced anti-inflammatory responses in LPS-inflamed macrophages. These results demonstrated that dh404 suppresses pro-inflammatory responses in macrophages via an activation of Nrf2 independently of Keap1 and NF-κB, suggesting a unique therapeutic potential of dh404 for specific targeting a Nrf2-mediated resolution of inflammation.« less

Acetaminophen Enhances Cisplatin- and Paclitaxel-mediated Cytotoxicity to SKOV3 Human Ovarian Carcinoma

PubMed Central

Wu, Y. Jeffrey; Neuwelt, Alexander J.; Muldoon, Leslie L.; Neuwelt, Edward A.

2013-01-01

Background Ovarian cancer is commonly treated with cisplatin/paclitaxel but many tumors become resistant. Acetaminophen reduced glutathione and enhanced chemotherapy efficacy in treating hepatic cancer. The objective of this study was to examine if acetaminophen enhances the cytotoxicity of cisplatin/paclitaxel in ovarian cancer. Materials and Methods SKOV3 human ovarian carcinoma cells in vitro and a subcutaneous tumor nude rat model were used and treated with cisplatin/paclitaxel with or without acetaminophen. Results In vitro, acetaminophen enhanced apoptosis induced by cisplatin and paclitaxel with similar effects on glutathione, reactive oxygen species and mitochondrial membrane potential but different effects on nuclear factor erythroid 2-related factor 2 (NRF2) translocation. In vivo, acetaminophen was uniformly distributed in tissue and significantly reduced hepatic glutathione. Acetaminophen enhanced cisplatin chemotherapeutic effect by reducing tumor recurrence Conclusion Our results suggest that acetaminophen as a chemoenhancing adjuvant could improve the efficacy of cisplatin and paclitaxel in treating patients with ovarian carcinoma and other tumor types. PMID:23749887

Structural and Functional Characterization of Nrf2 Degradation by the Glycogen Synthase Kinase 3/β-TrCP Axis

PubMed Central

Rada, Patricia; Rojo, Ana I.; Evrard-Todeschi, Nathalie; Innamorato, Nadia G.; Cotte, Axelle; Jaworski, Tomasz; Tobón-Velasco, Julio C.; Devijver, Herman; García-Mayoral, María Flor; Van Leuven, Fred; Hayes, John D.

2012-01-01

The transcription factor NF-E2-related factor 2 (Nrf2) is a master regulator of a genetic program, termed the phase 2 response, that controls redox homeostasis and participates in multiple aspects of physiology and pathology. Nrf2 protein stability is regulated by two E3 ubiquitin ligase adaptors, Keap1 and β-TrCP, the latter of which was only recently reported. Here, two-dimensional (2D) gel electrophoresis and site-directed mutagenesis allowed us to identify two serines of Nrf2 that are phosphorylated by glycogen synthase kinase 3β (GSK-3β) in the sequence DSGISL. Nuclear magnetic resonance studies defined key residues of this phosphosequence involved in docking to the WD40 propeller of β-TrCP, through electrostatic and hydrophobic interactions. We also identified three arginine residues of β-TrCP that participate in Nrf2 docking. Intraperitoneal injection of the GSK-3 inhibitor SB216763 led to increased Nrf2 and heme oxygenase-1 levels in liver and hippocampus. Moreover, mice with hippocampal absence of GSK-3β exhibited increased levels of Nrf2 and phase 2 gene products, reduced glutathione, and decreased levels of carbonylated proteins and malondialdehyde. This study establishes the structural parameters of the interaction of Nrf2 with the GSK-3/β-TrCP axis and its functional relevance in the regulation of Nrf2 by the signaling pathways that impinge on GSK-3. PMID:22751928

Modulation of Nrf2 by Olive Oil and Wine Polyphenols and Neuroprotection

PubMed Central

Martínez-Huélamo, Miriam; Boronat, Anna; de la Torre, Rafael

2017-01-01

Strong adherence to a Mediterranean diet is associated with improved cognitive function and a lower prevalence of mild cognitive impairment. Olive oil and red wine are rich sources of polyphenols which are responsible in part for the beneficial effects on cognitive functioning. Polyphenols induce endogenous antioxidant defense mechanisms by modulating transcription factors such as the nuclear factor (erythroid-derived 2)-like 2 (Nrf2). This review discusses the scientific data supporting the modulating effect of olive oil and red wine polyphenols on Nrf2 expression, and the potential health benefits associated with cognitive functioning. PMID:28954417

Sulforaphane and Other Nutrigenomic Nrf2 Activators: Can the Clinician's Expectation Be Matched by the Reality?

PubMed

Houghton, Christine A; Fassett, Robert G; Coombes, Jeff S

2016-01-01

The recognition that food-derived nonnutrient molecules can modulate gene expression to influence intracellular molecular mechanisms has seen the emergence of the fields of nutrigenomics and nutrigenetics. The aim of this review is to describe the properties of nutrigenomic activators of transcription factor Nrf2 (nuclear factor erythroid 2-related factor 2), comparing the potential for sulforaphane and other phytochemicals to demonstrate clinical efficacy as complementary medicines. Broccoli-derived sulforaphane emerges as a phytochemical with this capability, with oral doses capable of favourably modifying genes associated with chemoprevention. Compared with widely used phytochemical-based supplements like curcumin, silymarin, and resveratrol, sulforaphane more potently activates Nrf2 to induce the expression of a battery of cytoprotective genes. By virtue of its lipophilic nature and low molecular weight, sulforaphane displays significantly higher bioavailability than the polyphenol-based dietary supplements that also activate Nrf2. Nrf2 activation induces cytoprotective genes such as those playing key roles in cellular defense mechanisms including redox status and detoxification. Both its high bioavailability and significant Nrf2 inducer capacity contribute to the therapeutic potential of sulforaphane-yielding supplements.

The nuclear factor-erythroid 2-related factor/heme oxygenase-1 axis is critical for the inflammatory features of type 2 diabetes-associated osteoarthritis.

PubMed

Vaamonde-Garcia, Carlos; Courties, Alice; Pigenet, Audrey; Laiguillon, Marie-Charlotte; Sautet, Alain; Houard, Xavier; Kerdine-Römer, Saadia; Meijide, Rosa; Berenbaum, Francis; Sellam, Jérémie

2017-09-01

Epidemiological findings support the hypothesis that type 2 diabetes mellitus (T2DM) is a risk factor for osteoarthritis (OA). Moreover, OA cartilage from patients with T2DM exhibits a greater response to inflammatory stress, but the molecular mechanism is unclear. To investigate whether the antioxidant defense system participates in this response, we examined here the expression of nuclear factor-erythroid 2-related factor (Nrf-2), a master antioxidant transcription factor, and of heme oxygenase-1 (HO-1), one of its main target genes, in OA cartilage from T2DM and non-T2DM patients as well as in murine chondrocytes exposed to high glucose (HG). Ex vivo experiments indicated that Nrf-2 and HO-1 expression is reduced in T2DM versus non-T2DM OA cartilage (0.57-fold Nrf-2 and 0.34-fold HO-1), and prostaglandin E 2 (PGE 2 ) release was increased in samples with low HO-1 expression. HG-exposed, IL-1β-stimulated chondrocytes had lower Nrf-2 levels in vitro , particularly in the nuclear fraction, than chondrocytes exposed to normal glucose (NG). Accordingly, HO-1 levels were also decreased (0.49-fold) in these cells. The HO-1 inducer cobalt protoporphyrin IX more efficiently attenuated PGE 2 and IL-6 release in HG+IL-1β-treated cells than in NG+IL-1β-treated cells. Greater reductions in HO-1 expression and increase in PGE 2 /IL-6 production were observed in HG+IL-1β-stimulated chondrocytes from Nrf-2 -/- mice than in chondrocytes from wild-type mice. We conclude that the Nrf-2/HO-1 axis is a critical pathway in the hyperglucidic-mediated dysregulation of chondrocytes. Impairments in this antioxidant system may explain the greater inflammatory responsiveness of OA cartilage from T2DM patients and may inform treatments of such patients. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Quercetin attenuates toosendanin-induced hepatotoxicity through inducing the Nrf2/GCL/GSH antioxidant signaling pathway.

PubMed

Jin, Yao; Huang, Zhen-Lin; Li, Li; Yang, Yang; Wang, Chang-Hong; Wang, Zheng-Tao; Ji, Li-Li

2018-06-19

Toosendanin (TSN) is the main active compound in Toosendan Fructus and Meliae Cortex, two commonly used traditional Chinese medicines. TSN has been reported to induce hepatotoxicity, but its mechanism remains unclear. In this study, we demonstrated the critical role of nuclear factor erythroid 2-related factor 2 (Nrf2) in protecting against TSN-induced hepatotoxicity in mice and human normal liver L-02 cells. In mice, administration of TSN (10 mg/kg)-induced acute liver injury evidenced by increased serum alanine/aspartate aminotransferase (ALT/AST) and alkaline phosphatase (ALP) activities, and total bilirubin (TBiL) content as well as the histological changes. Furthermore, TSN markedly increased liver reactive oxygen species (ROS) and malondialdehyde (MDA) levels, and decreased liver glutathione (GSH) content and Nrf2 expression. In L-02 cells, TSN (2 μM) time-dependently reduced glutamate-cysteine ligase (GCL) activity and cellular expression of the catalytic/modify subunit of GCL (GCLC/GCLM). Moreover, TSN reduced cellular GSH content and the increased ROS formation, and time-dependently decreased Nrf2 expression and increased the expression of the Nrf2 inhibitor protein kelch-like ECH-associated protein-1 (Keap1). Pre-administration of quercetin (40, 80 mg/kg) effectively inhibited TSN-induced liver oxidative injury and reversed the decreased expression of Nrf2 and GCLC/GCLM in vivo and in vitro. In addition, the quercetin-provided protection against TSN-induced hepatotoxicity was diminished in Nrf2 knock-out mice. In conclusion, TSN decreases cellular GSH content by reducing Nrf2-mediated GCLC/GCLM expression via decreasing Nrf2 expression. Quercetin attenuates TSN-induced hepatotoxicity by inducing the Nrf2/GCL/GSH antioxidant signaling pathway. This study implies that inducing Nrf2 activation may be an effective strategy to prevent TSN-induced hepatotoxicity.

Sulforaphane Ameliorates Okadaic Acid-Induced Memory Impairment in Rats by Activating the Nrf2/HO-1 Antioxidant Pathway.

PubMed

Dwivedi, Subhash; Rajasekar, N; Hanif, Kashif; Nath, Chandishwar; Shukla, Rakesh

2016-10-01

Okadaic acid (OKA) causes memory impairment and attenuates nuclear factor erythroid 2-related factor 2 (Nrf2) along with oxidative stress and neuroinflammation in rats. Sulforaphane (dietary isothiocyanate compound), an activator of Nrf2 signaling, exhibits neuroprotective effects. However, the protective effect of sulforaphane in OKA-induced neurotoxicity remains uninvestigated. Therefore, in the present study, the role of sulforaphane in OKA-induced memory impairment in rats was explored. A significant increased Nrf2 expression in the hippocampus and cerebral cortex was observed in trained (Morris water maze) rats, and a significant decreased Nrf2 expression in memory-impaired (OKA, 200 ng icv) rats indicated its involvement in memory function. Sulforaphane administration (5 and 10 mg/kg, ip, days 1 and 2) ameliorates OKA-induced memory impairment in rats. The treatment also restored Nrf2 and its downstream antioxidant protein expression (GCLC, HO-1) and attenuated oxidative stress (ROS, nitrite, GSH), neuroinflammation (NF-κB, TNF-α, IL-10), and neuronal apoptosis in the cerebral cortex and hippocampus of OKA-treated rats. Further, to determine whether modulation of Nrf2 signaling is responsible for the protective effect of sulforaphane, in vitro, Nrf2 siRNA and its downstream HO-1 inhibition studies were carried out in a rat astrocytoma cell line (C6). The protective effects of sulforaphane were abolished with Nrf2 siRNA and HO-1 inhibition in astrocytes. The results suggest that Nrf2-dependent activation of cellular antioxidant machinery results in sulforaphane-mediated protection against OKA-induced memory impairment in rats. Graphical Abstract ᅟ.

Aluminium induced oxidative stress results in decreased mitochondrial biogenesis via modulation of PGC-1α expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sharma, Deep Raj; Sunkaria, Aditya; Wani, Willayat Yousuf

The present investigation was carried out to elucidate a possible molecular mechanism related to the effects of aluminium-induced oxidative stress on various mitochondrial respiratory complex subunits with special emphasis on the role of Peroxisome proliferator activated receptor gamma co-activator 1α (PGC-1α) and its downstream targets i.e. Nuclear respiratory factor-1(NRF-1), Nuclear respiratory factor-2(NRF-2) and Mitochondrial transcription factor A (Tfam) in mitochondrial biogenesis. Aluminium lactate (10 mg/kg b.wt./day) was administered intragastrically to rats for 12 weeks. After 12 weeks of exposure, we found an increase in ROS levels, mitochondrial DNA oxidation and decrease in citrate synthase activity in the Hippocampus (HC) andmore » Corpus striatum (CS) regions of rat brain. On the other hand, there was a decrease in the mRNA levels of the mitochondrial encoded subunits–NADH dehydrogenase (ND) subunits i.e. ND1, ND2, ND3, Cytochrome b (Cytb), Cytochrome oxidase (COX) subunits i.e. COX1, COX3, ATP synthase (ATPase) subunit 6 along with reduced expression of nuclear encoded subunits COX4, COX5A, COX5B of Electron transport chain (ETC). Besides, a decrease in mitochondrial DNA copy number and mitochondrial content in both regions of rat brain was observed. The PGC-1α was down-regulated in aluminium treated rats along with NRF-1, NRF-2 and Tfam, which act downstream from PGC-1α in aluminium treated rats. Electron microscopy results revealed a significant increase in the mitochondrial swelling, loss of cristae, chromatin condensation and decreases in mitochondrial number in case of aluminium treated rats as compared to control. So, PGC-1α seems to be a potent target for aluminium neurotoxicity, which makes it an almost ideal target to control or limit the damage that has been associated with the defective mitochondrial function seen in neurodegenerative diseases. - Highlights: • Aluminium decreases the mRNA levels of mitochondrial and nuclear encoded subunits. • It decreases the mtDNA copy number and mitochondrial content in rat brain. • It down-regulates the mRNA and protein levels of PGC-1α, NRF-1, NRF-2 and Tfam. • It also disturbs the mitochondrial or nuclear architecture of neurons. • Finally it also decreases mitochondrial number in HC and CS regions of rat brain.« less

Novel Nrf2 activators from microbial transformation products inhibit blood-retinal barrier permeability in rabbits.

PubMed

Nakagami, Yasuhiro; Masuda, Kayoko; Hatano, Emiko; Inoue, Tatsuya; Matsuyama, Takuya; Iizuka, Mayumi; Ono, Yasunori; Ohnuki, Takashi; Murakami, Yoko; Iwasaki, Masaru; Yoshida, Kazuhiro; Kasuya, Yuji; Komoriya, Satoshi

2015-03-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a redox-sensitive transcription factor that binds to antioxidant response elements located in the promoter region of genes encoding many antioxidant enzymes and phase II detoxifying enzymes. Activation of the Nrf2 pathway seems protective for many organs, and although a well-known Nrf2 activator, bardoxolone methyl, was evaluated clinically for treating chronic kidney disease, it was found to induce adverse events. Many bardoxolone methyl derivatives, mostly derived by chemical modifications, have already been studied. However, we adopted a biotransformation technique to obtain a novel Nrf2 activator. The potent novel Nrf2 activator, RS9, was obtained from microbial transformation products. Its Nrf2 activity was evaluated by determining NADPH:quinone oxidoreductase-1 induction activity in Hepa1c1c7 cells. We also investigated the effects of RS9 on oxygen-induced retinopathy in rats and glycated albumin-induced blood-retinal barrier permeability in rabbits because many ocular diseases are associated with oxidative stress and inflammation. Bardoxolone methyl doubled the specific activity of Nrf2 in Hepa1c1c7 cells at a much higher concentration than RS9. Moreover, the induction of Nrf2-targeted genes was observed at a one-tenth lower concentration of RS9. Interestingly, the cytotoxicity of RS9 was substantially reduced compared with bardoxolone methyl. Oral and intravitreal administration of RS9 ameliorated the pathological scores and leakage in the models of retinopathy in rats and ocular inflammation in rabbits respectively. Nrf2 activators are applicable for treating ocular diseases and novel Nrf2 activators have potential as a unique method for prevention and treatment of retinovascular disease. © 2014 The British Pharmacological Society.

Novel Nrf2 activators from microbial transformation products inhibit blood–retinal barrier permeability in rabbits

PubMed Central

Nakagami, Yasuhiro; Masuda, Kayoko; Hatano, Emiko; Inoue, Tatsuya; Matsuyama, Takuya; Iizuka, Mayumi; Ono, Yasunori; Ohnuki, Takashi; Murakami, Yoko; Iwasaki, Masaru; Yoshida, Kazuhiro; Kasuya, Yuji; Komoriya, Satoshi

2015-01-01

Background and Purpose Nuclear factor erythroid 2-related factor 2 (Nrf2) is a redox-sensitive transcription factor that binds to antioxidant response elements located in the promoter region of genes encoding many antioxidant enzymes and phase II detoxifying enzymes. Activation of the Nrf2 pathway seems protective for many organs, and although a well-known Nrf2 activator, bardoxolone methyl, was evaluated clinically for treating chronic kidney disease, it was found to induce adverse events. Many bardoxolone methyl derivatives, mostly derived by chemical modifications, have already been studied. However, we adopted a biotransformation technique to obtain a novel Nrf2 activator. Experimental Approach The potent novel Nrf2 activator, RS9, was obtained from microbial transformation products. Its Nrf2 activity was evaluated by determining NADPH:quinone oxidoreductase-1 induction activity in Hepa1c1c7 cells. We also investigated the effects of RS9 on oxygen-induced retinopathy in rats and glycated albumin-induced blood–retinal barrier permeability in rabbits because many ocular diseases are associated with oxidative stress and inflammation. Key Results Bardoxolone methyl doubled the specific activity of Nrf2 in Hepa1c1c7 cells at a much higher concentration than RS9. Moreover, the induction of Nrf2-targeted genes was observed at a one-tenth lower concentration of RS9. Interestingly, the cytotoxicity of RS9 was substantially reduced compared with bardoxolone methyl. Oral and intravitreal administration of RS9 ameliorated the pathological scores and leakage in the models of retinopathy in rats and ocular inflammation in rabbits respectively. Conclusion and Implications Nrf2 activators are applicable for treating ocular diseases and novel Nrf2 activators have potential as a unique method for prevention and treatment of retinovascular disease. PMID:25363737

Nrf2 activation prevents cadmium-induced acute liver injury

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu, Kai C.; Liu, Jie J.; Klaassen, Curtis D., E-mail: cklaasse@kumc.edu

2012-08-15

Oxidative stress plays an important role in cadmium-induced liver injury. Nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcription factor that up-regulates cytoprotective genes in response to oxidative stress. To investigate the role of Nrf2 in cadmium-induced hepatotoxicity, Nrf2-null mice, wild-type mice, kelch-like ECH-associated protein 1-knockdown (Keap1-KD) mice with enhanced Nrf2, and Keap1-hepatocyte knockout (Keap1-HKO) mice with maximum Nrf2 activation were treated with cadmium chloride (3.5 mg Cd/kg, i.p.). Blood and liver samples were collected 8 h thereafter. Cadmium increased serum alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) activities, and caused extensive hepatic hemorrhage and necrosis in the Nrf2-nullmore » mice. In contrast, Nrf2-enhanced mice had lower serum ALT and LDH activities and less morphological alternations in the livers than wild-type mice. H{sub 2}DCFDA (2′,7′-dichlorodihydrofluoresein diacetate) staining of primary hepatocytes isolated from the four genotypes of mice indicated that oxidative stress was higher in Nrf2-null cells, and lower in Nrf2-enhanced cells than in wild-type cells. To further investigate the mechanism of the protective effect of Nrf2, mRNA of metallothionein (MT) and other cytoprotective genes were determined. Cadmium markedly induced MT-1 and MT-2 in livers of all four genotypes of mice. In contrast, genes involved in glutathione synthesis and reducing reactive oxygen species, including glutamate-cysteine ligase (Gclc), glutathione peroxidase-2 (Gpx2), and sulfiredoxin-1 (Srxn-1) were only induced in Nrf2-enhanced mice, but not in Nrf2-null mice. In conclusion, the present study shows that Nrf2 activation prevents cadmium-induced oxidative stress and liver injury through induction of genes involved in antioxidant defense rather than genes that scavenge Cd. -- Highlights: ► Cadmium caused extensive hepatic hemorrhage and necrosis in Nrf2-null mice. ► Keap1-KD and Keap1-HKO mice were resistant to cadmium-induced liver injury. ► Cadmium increased ROS in hepatocytes isolated from Nrf2-null and wild-type mice. ► Mt-1 and Mt‐2 were induced over 200-fold in both Nrf2-null and Nrf2-enhanced mice. ► Gclc, Gpx2, and Srxn-1 were induced in Nrf2-enhanced mice, not in Nrf2-null mice.« less

Nrf2 inhibits oxaliplatin-induced peripheral neuropathy via protection of mitochondrial function.

PubMed

Yang, Yang; Luo, Lan; Cai, Xueting; Fang, Yuan; Wang, Jiaqi; Chen, Gang; Yang, Jie; Zhou, Qian; Sun, Xiaoyan; Cheng, Xiaolan; Yan, Huaijiang; Lu, Wuguang; Hu, Chunping; Cao, Peng

2018-05-20

Oxaliplatin-induced peripheral neuropathy (OIPN) is a severe, dose-limiting toxicity associated with cancer chemotherapy. The efficacy of antioxidant administration in OIPN is debatable, as the promising preliminary results obtained with a number of antioxidants have not been confirmed in larger clinical trials. Besides its antioxidant activity, the transcription factor, nuclear factor-erythroid 2 (NF-E2) p45-related factor 2 (Nrf2) plays a crucial role in the maintenance of mitochondrial homeostasis, and mitochondrial dysfunction is a key contributor to OIPN. Here, we have investigated the protective properties of Nrf2 in OIPN. Nrf2 -/- mice displayed severe mechanical allodynia and cold sensitivity and thus experienced increased peripheral nervous system injury compared to Nrf2 +/+ mice. Furthermore, Nrf2 knockout aggravated oxaliplatin-induced reactive oxygen species production, decreased the mitochondrial membrane potential, led to abnormal intracellular calcium levels, and induced cytochrome c-related apoptosis and overexpression of the TRP protein family. Sulforaphane-induced activation of the Nrf2 signaling pathway alleviated morphological alterations, mitochondrial dysfunction in dorsal root ganglion neurons, and nociceptive sensations in mice. Our findings reveal that Nrf2 may play a critical role in ameliorating OIPN, through protection of mitochondrial function by alleviating oxidative stress and inhibiting TRP protein family expression. This suggests that pharmacological or therapeutic activation of Nrf2 may be used to prevent or slow down the progression of OIPN. Copyright © 2018 Elsevier Inc. All rights reserved.

Oxidative stress and dysfunctional NRF2 underlie pachyonychia congenita phenotypes

PubMed Central

Kerns, Michelle L.; Hakim, Jill M.C.; Lu, Rosemary G.; Guo, Yajuan; Berroth, Andreas; Kaspar, Roger L.

2016-01-01

Palmoplantar keratoderma (PPK) are debilitating lesions that arise in individuals with pachyonychia congenita (PC) and feature upregulation of danger-associated molecular patterns and skin barrier regulators. The defining features of PC-associated PPK are reproduced in mice null for keratin 16 (Krt16), which is commonly mutated in PC patients. Here, we have shown that PPK onset is preceded by oxidative stress in footpad skin of Krt16–/– mice and correlates with an inability of keratinocytes to sustain nuclear factor erythroid–derived 2 related factor 2–dependent (NRF2-dependent) synthesis of the cellular antioxidant glutathione (GSH). Additionally, examination of plantar skin biopsies from individuals with PC confirmed the presence of high levels of hypophosphorylated NRF2 in lesional tissue. In Krt16–/– mice, genetic ablation of Nrf2 worsened spontaneous skin lesions and accelerated PPK development in footpad skin. Hypoactivity of NRF2 in Krt16–/– footpad skin correlated with decreased levels or activity of upstream NRF2 activators, including PKCδ, receptor for activated C kinase 1 (RACK1), and p21. Topical application of the NRF2 activator sulforaphane to the footpad of Krt16–/– mice prevented the development of PPK and normalized redox balance via regeneration of GSH from existing cellular pools. Together, these findings point to oxidative stress and dysfunctional NRF2 as contributors to PPK pathogenesis, identify K16 as a regulator of NRF2 activation, and suggest that pharmacological activation of NRF2 should be further explored for PC treatment. PMID:27183391

Transcription Factor NRF2 as a Therapeutic Target for Chronic Diseases: A Systems Medicine Approach.

PubMed

Cuadrado, Antonio; Manda, Gina; Hassan, Ahmed; Alcaraz, María José; Barbas, Coral; Daiber, Andreas; Ghezzi, Pietro; León, Rafael; López, Manuela G; Oliva, Baldo; Pajares, Marta; Rojo, Ana I; Robledinos-Antón, Natalia; Valverde, Angela M; Guney, Emre; Schmidt, Harald H H W

2018-04-01

Systems medicine has a mechanism-based rather than a symptom- or organ-based approach to disease and identifies therapeutic targets in a nonhypothesis-driven manner. In this work, we apply this to transcription factor nuclear factor (erythroid-derived 2)-like 2 (NRF2) by cross-validating its position in a protein-protein interaction network (the NRF2 interactome) functionally linked to cytoprotection in low-grade stress, chronic inflammation, metabolic alterations, and reactive oxygen species formation. Multiscale network analysis of these molecular profiles suggests alterations of NRF2 expression and activity as a common mechanism in a subnetwork of diseases (the NRF2 diseasome). This network joins apparently heterogeneous phenotypes such as autoimmune, respiratory, digestive, cardiovascular, metabolic, and neurodegenerative diseases, along with cancer. Importantly, this approach matches and confirms in silico several applications for NRF2-modulating drugs validated in vivo at different phases of clinical development. Pharmacologically, their profile is as diverse as electrophilic dimethyl fumarate, synthetic triterpenoids like bardoxolone methyl and sulforaphane, protein-protein or DNA-protein interaction inhibitors, and even registered drugs such as metformin and statins, which activate NRF2 and may be repurposed for indications within the NRF2 cluster of disease phenotypes. Thus, NRF2 represents one of the first targets fully embraced by classic and systems medicine approaches to facilitate both drug development and drug repurposing by focusing on a set of disease phenotypes that appear to be mechanistically linked. The resulting NRF2 drugome may therefore rapidly advance several surprising clinical options for this subset of chronic diseases. Copyright © 2018 by The Author(s).

Celecoxib activates PI-3K/Akt and mitochondrial redox signaling to enhance heme oxygenase-1-mediated anti-inflammatory activity in vascular endothelium.

PubMed

Hamdulay, Shahir S; Wang, Bufei; Birdsey, Graeme M; Ali, Faisal; Dumont, Odile; Evans, Paul C; Haskard, Dorian O; Wheeler-Jones, Caroline P; Mason, Justin C

2010-04-15

Although nonsteroidal anti-inflammatory drugs (NSAIDs) provide important control of pain and inflammation, they have been overshadowed by concerns regarding atherothrombotic complications. However, celecoxib seems to have a relatively good cardiovascular profile and may improve endothelial function in coronary heart disease. This led us to the hypothesis that celecoxib induces the vasculoprotective enzyme heme oxygenase-1 (HO-1). In human umbilical vein and aortic endothelial cells, 24-48 h treatment with celecoxib induced HO-1 mRNA and protein expression and increased HO-1 enzyme activity. This effect was not seen with rofecoxib or indomethacin. Supplementation of culture medium with iloprost or prostaglandin E(2) failed to reverse celecoxib-mediated HO-1 induction, indicating a cyclooxygenase-independent mechanism. Rather, this action of celecoxib involved generation of mitochondria-derived reactive oxygen species, Akt phosphorylation, and nuclear translocation of the transcription factor Nrf2, with N-acetylcysteine, PI-3K antagonist LY290042, and dominant-negative Akt abrogating the effects. Furthermore, celecoxib-induced HO-1 was inhibited by dominant-negative Nrf2. The functional significance of HO-1 induction was revealed by celecoxib-mediated inhibition of VCAM-1 expression, a response reversed by the HO-1 antagonist zinc protoporphyrin. HO-1 induction provides a molecular mechanism for clinical observations indicating relative freedom from atherothrombotic complications in patients taking celecoxib compared to other NSAIDs with comparable anti-inflammatory activity. Copyright 2010 Elsevier Inc. All rights reserved.

Continuous activation of Nrf2 and its target antioxidant enzymes leads to arsenite-induced malignant transformation of human bronchial epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Xu; Wang, Dapeng; Department of Toxicology, School of Public Health, Guizhou Medical University, Guiyang, Guizhou

Long-term exposure to arsenite leads to human lung cancer, but the underlying mechanisms of carcinogenesis remain obscure. The transcription factor of nuclear factor-erythroid-2 p45-related factor (Nrf2)-mediated antioxidant response represents a critical cellular defense mechanism and protection against various diseases. Paradoxically, emerging data suggest that the constitutive activation of Nrf2 is associated with cancer development, progression and chemotherapy resistance. However, the role of Nrf2 in the occurrence of cancer induced by long-term arsenite exposure remains to be fully understood. By establishing transformed human bronchial epithelial (HBE) cells via chronic low-dose arsenite treatment, we showed that, in acquiring this malignant phenotype, continuousmore » low level of ROS and sustained enhancement of Nrf2 and its target antioxidant enzyme levels were observed in the later-stage of arsenite-induced cell transformation. The downregulation of Keap1 level may be responsible for the over-activation of Nrf2 and its target enzymes. To validate these observations, Nrf2 was knocked down in arsenite-transformed HBE cells by SiRNA transfection, and the levels of Nrf2 and its target antioxidant enzymes, ROS, cell proliferation, migration, and colony formation were determined following these treatments. Results showed that blocked Nrf2 expression significantly reduced Nrf2 and its target antioxidant enzyme levels, restored ROS levels, and eventually suppressed cell proliferation, migration, and colony formation of the transformed cells. In summary, the results of the study strongly suggested that the continuous activation of Nrf2 and its target antioxidant enzymes led to the over-depletion of intracellular ROS levels, which contributed to arsenite-induced HBE cell transformation. - Highlights: • Low level, long term arsenite exposure induces malignant transformation in vitro. • Long term arsenite exposure reduces ROS and MDA levels. • Long term arsenite exposure enhances Nrf2-mediated antioxidant levels. • Knockdown of Nrf2 reduces malignant degree of arsenite-transformed cells.« less

NRF1 Is an ER Membrane Sensor that Is Central to Cholesterol Homeostasis.

PubMed

Widenmaier, Scott B; Snyder, Nicole A; Nguyen, Truc B; Arduini, Alessandro; Lee, Grace Y; Arruda, Ana Paula; Saksi, Jani; Bartelt, Alexander; Hotamisligil, Gökhan S

2017-11-16

Cholesterol is a critical nutrient requiring tight constraint in the endoplasmic reticulum (ER) due to its uniquely challenging biophysical properties. While the mechanisms by which the ER defends against cholesterol insufficiency are well described, it remains unclear how the ER senses and effectively defends against cholesterol excess. Here, we identify the ER-bound transcription factor nuclear factor erythroid 2 related factor-1, Nrf1/Nfe2L1, as a critical mediator of this process. We show that Nrf1 directly binds to and specifically senses cholesterol in the ER through a defined domain and that cholesterol regulates Nrf1 turnover, processing, localization, and activity. In Nrf1 deficiency, in vivo cholesterol challenges induce massive hepatic cholesterol accumulation and damage, which is rescued by replacing Nrf1 exogenously. This Nrf1-mediated mechanism involves the suppression of CD36-driven inflammatory signaling and derepression of liver X receptor activity. These findings reveal Nrf1 as a guardian of cholesterol homeostasis and a core component of adaptive responses to excess cellular cholesterol. Copyright © 2017. Published by Elsevier Inc.

Insulin is a potential antioxidant for diabetes-associated cognitive decline via regulating Nrf2 dependent antioxidant enzymes.

PubMed

Song, Ying; Ding, Wei; Bei, Yun; Xiao, Yan; Tong, Hai-Da; Wang, Li-Bo; Ai, Li-Yao

2018-05-21

To investigate the neuroprotective effects of insulin on diabetic encephalopathy and its mechanism. The diabetic model was established by injection of streptozotocin. Behavior examinations were conducted by the Morris water maze. Histopathological alterations were detected by HE staining. ROS, CAT levels and SOD activity were measured using a microplate reader. In vitro, the viability of wild type and knock-down PC12 cells was detected by MTT assay, the morphology of cells was monitored under a microscope. The subcellular distribution of Nrf2 was observed by western blotting and immunohistochemistry. Evident oxidative stress injury was observed in diabetic rats and H 2 O 2 -induced PC12 cells. Insulin not only protect diabetic rat from oxidative stress injury but also significantly inhibited H 2 O 2 -induced apoptosis and intracellular ROS in cells. In addition, the level of malondialdehyde was reduced, and the activities of superoxide dismutase, catalase and glutathione peroxidase were augmented in both diabetic rats and PC12 cells. Interestingly, insulin promoted the translocation of Nrf2 into the nucleus and activation of downstream antioxidant protein expression. Further, the Nrf2 knockdown cells suffered more serious H 2 O 2 -induced damage than the wild PC12 cells. Moreover, insulin had no significant protective effect on knockdown cells with H 2 O 2 -damage. Collectively, our results suggested that insulin significantly inhibited neuronal damage through the Nrf2 signaling pathway, which regulates endogenous oxidant-antioxidant balance, therefore, insulin may be a potential protective agent for the treatment of oxidative stress-induced diabetic encephalopathy. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Anti-inflammatory and antioxidant activity of polyphenolic extracts from Lactuca sativa (var. Maravilla de Verano) under different farming methods.

PubMed

Adesso, Simona; Pepe, Giacomo; Sommella, Eduardo; Manfra, Michele; Scopa, Antonio; Sofo, Adriano; Tenore, Gian Carlo; Russo, Mariateresa; Di Gaudio, Francesca; Autore, Giuseppina; Campiglia, Pietro; Marzocco, Stefania

2016-09-01

Besides their nutritional value, vegetables are a source of health-promoting compounds, such as polyphenols, and their content can be influenced by the particular farming method. In this study polyphenolic extracts from Lactuca sativa (var. Maravilla de verano) plants cultivated with different farming methods were chemically characterised and tested in vitro and ex vivo inflammation models. The tested extacts (250-2.5 µg mL(-1) ) were able to reduce both the inflammatory and oxidative stress in LPS-stimulated J774A.1 murine monocyte macrophage cells, by lowering the release of nitric oxide (NO) and reactive oxygen species (ROS) and promoting nuclear translocation of nuclear factor (erythroid-derived 2)-like 2; (Nrf2) and nuclear factor-κB (NF-κB). In this regard, quantitative profiles revealed different amounts of polyphenols, in particular quercetin levels were higher in plants under mineral fertilised treatment. Those extract showed an enhanced anti-inflammatory and antioxidant activity. Our data showed the anti-inflammatory and antioxidant potential of Maravilla de Verano polyphenolic extracts. The effect of farming methods on polyphenolic levels was highlighted. The higher reduction of inflammatory mediators release in extracts from plants cultivated under mineral fertilisation treatment was correlated to the higher amount of quercetin. These results can be useful for both nutraceutical or agronomic purposes. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Nrf2-mediated antioxidant response by ethanolic extract of Sida cordifolia provides protection against alcohol-induced oxidative stress in liver by upregulation of glutathione metabolism.

PubMed

Rejitha, S; Prathibha, P; Indira, M

2015-03-01

Objective The study aimed to evaluate the antioxidant property of ethanolic extract of Sida cordifolia (SAE) on alcohol-induced oxidative stress and to elucidate its mechanism of action. Methods Male albino rats of the Sprague-Dawley strain were grouped into four: (1) control, (2) alcohol (4 g/kg body weight), (3) SAE (50 mg/100 g body weight), and (4) alcohol (4 g/kg body weight) + SAE (50 mg/100 g body weight). Alcohol and SAE were given orally each day by gastric intubation. The duration of treatment was 90 days. Results The activities of toxicity markers in liver and serum increased significantly in alcohol-treated rats and to a lesser extent in the group administered SAE + alcohol. The activity of alcohol dehydrogenase and the reactive oxygen species level were increased significantly in alcohol-treated rats but attenuated in the SAE co-administered group. Oxidative stress was increased in alcohol-treated rats as evidenced by the lowered activities of antioxidant enzymes, decreased level of reduced glutathione (GSH), increased lipid peroxidation products, and decreased expression of γ-glutamyl cysteine synthase in liver. The co-administration of SAE with alcohol almost reversed these changes. The activity of glutathione-S-transferase and translocation of Nrf2 from cytosol to nucleus in the liver was increased in both the alcohol and alcohol + SAE groups, but the maximum changes were observed in the latter group. Discussion The SAE most likely elicits its antioxidant potential by reducing oxidative stress, enhancing the translocation of Nrf2 to nucleus and thereby regulating glutathione metabolism, leading to enhanced GSH content.

Sulforaphane induces Nrf2 target genes and attenuates inflammatory gene expression in microglia from brain of young adult and aged mice.

PubMed

Townsend, Brigitte E; Johnson, Rodney W

2016-01-01

Increased neuroinflammation and oxidative stress resulting from heightened microglial activation are associated with age-related cognitive impairment. The objectives of this study were to examine the effects of the bioactive sulforaphane (SFN) on the nuclear factor E2-related factor 2 (Nrf2) pathway in BV2 microglia and primary microglia, and to evaluate proinflammatory cytokine expression in lipopolysaccharide (LPS)-stimulated primary microglia from adult and aged mice. BV2 microglia and primary microglia isolated from young adult and aged mice were treated with SFN and LPS. Changes in Nrf2 activity, expression of Nrf2 target genes, and levels of proinflammatory markers were assessed by quantitative PCR and immunoassay. SFN increased Nrf2 DNA-binding activity and upregulated Nrf2 target genes in BV2 microglia, while reducing LPS-induced interleukin (IL-)1β, IL-6, and inducible nitric oxide synthase (iNOS). In primary microglia from adult and aged mice, SFN increased expression of Nrf2 target genes and attenuated IL-1β, IL-6, and iNOS induced by LPS. These data indicate that SFN is a potential beneficial supplement that may be useful for reducing microglial mediated neuroinflammation and oxidative stress associated with aging. Copyright © 2015 Elsevier Inc. All rights reserved.

MDM2 controls NRF2 antioxidant activity in prevention of diabetic kidney disease.

PubMed

Guo, Weiying; Tian, Dan; Jia, Ye; Huang, Wenlin; Jiang, Mengnan; Wang, Junnan; Sun, Weixia; Wu, Hao

2018-04-26

Oxidative stress and P53 contribute to the pathogenesis of diabetic kidney disease (DKD). Nuclear factor erythroid 2-related factor 2 (NRF2) is a master regulator of cellular antioxidant defense system, is negatively regulated by P53 and prevents DKD. Recent findings revealed an important role of mouse double minute 2 (MDM2) in protection against DKD. However, the mechanism remained unclear. We hypothesized that MDM2 enhances NRF2 antioxidant signaling in DKD given that MDM2 is a key negative regulator of P53. The MDM2 inhibitor nutlin3a elevated renal P53, inhibited NRF2 signaling and induced oxidative stress, inflammation, fibrosis, DKD-like renal pathology and albuminuria in the wild-type (WT) non-diabetic mice. These effects exhibited more prominently in nutlin3a-treated WT diabetic mice. Interestingly, nutlin3a failed to induce greater renal injuries in the Nrf2 knockout (KO) mice under both the diabetic and non-diabetic conditions, indicating that NRF2 predominantly mediates MDM2's action. On the contrary, P53 inhibition by pifithrin-α activated renal NRF2 signaling and the expression of Mdm2, and attenuated DKD in the WT diabetic mice, but not in the Nrf2 KO diabetic mice. In high glucose-treated mouse mesangial cells, P53 gene silencing completely abolished nutlin3a's inhibitory effect on NRF2 signaling. The present study demonstrates for the first time that MDM2 controls renal NRF2 antioxidant activity in DKD via inhibition of P53, providing MDM2 activation and P53 inhibition as novel strategies in the management of DKD. Copyright © 2018 Elsevier B.V. All rights reserved.

Nrf2/Keap1 system regulates vascular smooth muscle cell apoptosis for vascular homeostasis: role in neointimal formation after vascular injury

PubMed Central

Ashino, Takashi; Yamamoto, Masayuki; Numazawa, Satoshi

2016-01-01

Abnormal increases in vascular smooth muscle cells (VSMCs) in the intimal region after a vascular injury is a key event in developing neointimal hyperplasia. To maintain vascular function, proliferation and apoptosis of VSMCs is tightly controlled during vascular remodeling. NF-E2-related factor 2 (Nrf2)/Kelch-like ECH-associated protein 1 (Keap1) system, a key component of the oxidative stress response that acts in maintaining homeostasis, plays an important role in neointimal hyperplasia after a vascular injury; however, the role of Nrf2/Keap1 in VSMC apoptosis has not been clarified. Here we report that 14 days after arterial injury in mice, TUNEL-positive VSMCs are detected in both the neointimal and medial layers. These layers contain cells expressing high levels of Nrf2 but low Keap1 expression. In VSMCs, Keap1 depletion induces features of apoptosis, such as positive TUNEL staining and annexin V binding. These changes are associated with an increased expression of nuclear Nrf2. Simultaneous Nrf2 depletion inhibits Keap1 depletion-induced apoptosis. At 14 days after the vascular injury, Nrf2-deficient mice demonstrated fewer TUNEL-positive cells and increased neointimal formation in the neointimal and medial areas. The results suggest that the Nrf2/Keap1 system regulates VSMC apoptosis during neointimal formation, thereby inhibiting neointimal hyperplasia after a vascular injury. PMID:27198574

CNC-bZIP protein Nrf1-dependent regulation of glucose-stimulated insulin secretion.

PubMed

Zheng, Hongzhi; Fu, Jingqi; Xue, Peng; Zhao, Rui; Dong, Jian; Liu, Dianxin; Yamamoto, Masayuki; Tong, Qingchun; Teng, Weiping; Qu, Weidong; Zhang, Qiang; Andersen, Melvin E; Pi, Jingbo

2015-04-01

The inability of pancreatic β-cells to secrete sufficient insulin in response to glucose stimulation is a major contributing factor to the development of type 2 diabetes (T2D). We investigated both the in vitro and in vivo effects of deficiency of nuclear factor-erythroid 2-related factor 1 (Nrf1) in β-cells on β-cell function and glucose homeostasis. Silencing of Nrf1 in β-cells leads to a pre-T2D phenotype with disrupted glucose metabolism and impaired insulin secretion. Specifically, MIN6 β-cells with stable knockdown of Nrf1 (Nrf1-KD) and isolated islets from β-cell-specific Nrf1-knockout [Nrf1(b)-KO] mice displayed impaired glucose responsiveness, including elevated basal insulin release and decreased glucose-stimulated insulin secretion (GSIS). Nrf1(b)-KO mice exhibited severe fasting hyperinsulinemia, reduced GSIS, and glucose intolerance. Silencing of Nrf1 in MIN6 cells resulted in oxidative stress and altered glucose metabolism, with increases in both glucose uptake and aerobic glycolysis, which is associated with the elevated basal insulin release and reduced glucose responsiveness. The elevated glycolysis and reduced glucose responsiveness due to Nrf1 silencing likely result from altered expression of glucose metabolic enzymes, with induction of high-affinity hexokinase 1 and suppression of low-affinity glucokinase. Our study demonstrated a novel role of Nrf1 in regulating glucose metabolism and insulin secretion in β-cells and characterized Nrf1 as a key transcription factor that regulates the coupling of glycolysis and mitochondrial metabolism and GSIS. Nrf1 plays critical roles in regulating glucose metabolism, mitochondrial function, and insulin secretion, suggesting that Nrf1 may be a novel target to improve the function of insulin-secreting β-cells.

Evaluation of the Protective Effects of Sarains on H2O2-Induced Mitochondrial Dysfunction and Oxidative Stress in SH-SY5Y Neuroblastoma Cells.

PubMed

Alvariño, Rebeca; Alonso, Eva; Tribalat, Marie-Aude; Gegunde, Sandra; Thomas, Olivier P; Botana, Luis M

2017-10-01

Sarains are diamide alkaloids isolated from the Mediterranean sponge Haliclona (Rhizoniera) sarai that have previously shown antibacterial, insecticidal and anti-fouling activities. In this study, we examined for the first time the neuroprotective effects of sarains 1, 2 and A against oxidative stress in a human neuronal model. SH-SY5Y cells were co-incubated with sarains at concentrations ranging from 0.01 to 10 μM, and the well-known oxidant hydrogen peroxide at 150 μM for 6 h and the protective effects of the compounds were evaluated. Among the sarains tested, sarain A was the most promising compound, improving mitochondrial function and decreasing reactive oxygen species levels in human neuroblastoma cells treated with the compound at 0.01, 0.1 and 1 μM. This compound was also able to increase the activity of the antioxidant enzymes superoxide dismutases by inducing the translocation of the nuclear factor E2-related factor 2 (Nrf2) to the nucleus at the lower concentrations tested (0.01 and 0.1 μM). Moreover, sarain A at 0.1 and 1 μM blocked the mitochondrial permeability transition pore (mPTP) opening through cyclophilin D inhibition. These results suggest that the protective effects produced by the treatment with sarain A are related with its ability to block the mPTP and to enhance the Nrf2 pathway, indicating that sarain A may be a candidate compound for further studies in neurodegenerative diseases.

Lack of Association between Nuclear Factor Erythroid-Derived 2-Like 2 Promoter Gene Polymorphisms and Oxidative Stress Biomarkers in Amyotrophic Lateral Sclerosis Patients

PubMed Central

Chico, Lucia; Borgia, Loredana; Rocchi, Anna; D'Amelio, Antonia; Carlesi, Cecilia; Mancuso, Michelangelo; Siciliano, Gabriele

2014-01-01

Oxidative stress involvement has been strongly hypothesized among the possible pathogenic mechanisms of motor neuron degeneration in amyotrophic lateral sclerosis (ALS). The intracellular redox balance is finely modulated by numerous complex mechanisms critical for cellular functions, among which the nuclear factor erythroid-derived 2-like 2 (NFE2L2/Nrf2) pathways. We genotyped, in a cohort of ALS patients (n = 145) and healthy controls (n = 168), three SNPs in Nrf2 gene promoter: −653 A/G, −651 G/A, and −617 C/A and evaluated, in a subset (n = 73) of patients, advanced oxidation protein products (AOPP), iron-reducing ability of plasma (FRAP), and plasma thiols (-SH) as oxidative damage peripheral biomarkers. Nrf2 polymorphisms were not different among patients and controls. Increased levels of AOPP (P < 0.05) and decreased levels of FRAP (P < 0.001) have been observed in ALS patients compared with controls, but no difference in -SH values was found. Furthermore, no association was found between biochemical markers of redox balance and Nrf2 polymorphisms. These data confirm an altered redox balance in ALS and indicate that, while being abnormally modified compared to controls, the oxidative stress biomarkers assessed in this study are independent from the −653 A/G, −651 G/A, and −617 C/A Nrf2 SNPs in ALS patients. PMID:24672634

Introducing the "TCDD-inducible AhR-Nrf2 gene battery".

PubMed

Yeager, Ronnie L; Reisman, Scott A; Aleksunes, Lauren M; Klaassen, Curtis D

2009-10-01

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) induces genes via the transcription factor aryl hydrocarbon receptor (AhR), including Cyp1a1, NAD(P)H:quinone oxidoreductase 1 (Nqo1), UDP-glucuronosyltransferase 1a6 (Ugt1a6), and glutathione S-transferase a1 (Gsta1). These genes are referred to as the "AhR gene battery." However, Nqo1 is also considered a prototypical target gene of the transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2). In mice, TCDD induction of Nrf2 and Nrf2 target, Nqo1, is dependent on AhR, and thus TCDD induction of drug-processing genes may be routed through an AhR-Nrf2 sequence. There has been speculation that Nrf2 may be involved in the TCDD induction of drug-processing genes; however, the data are not definitive. Therefore, to address whether TCDD induction of Nqo1, Ugts, and Gsts is dependent on Nrf2, we conducted the definitive experiment by administering TCDD (50 mug/kg, ip) to Nrf2-null and wild-type (WT) mice and collecting livers 24 h later to quantify the mRNA of drug-processing genes. TCDD induction of Cyp1a1 and Ugt1a1 was similar in WT and Nrf2-null mice, whereas TCDD induction of Ugt1a5 and 1a9 was blunted in Nrf2-null mice. TCDD induced Nqo1, Ugt1a6, 2b34, 2b35, 2b36, UDP-glucuronic acid-synthesizing gene UDP-glucose dehydrogenase, and Gsta1, m1, m2, m3, m6, p2, t2, and microsomal Gst1 in WT mice but not in Nrf2-null mice. Therefore, the present study demonstrates the novel finding that Nrf2 is required for TCDD induction of classical AhR battery genes Nqo1, Ugt1a6, and Gsta1, as well as most Ugt and Gst isoforms in livers of mice.

Nrf2 reduces levels of phosphorylated tau protein by inducing autophagy adaptor protein NDP52

NASA Astrophysics Data System (ADS)

Jo, Chulman; Gundemir, Soner; Pritchard, Susanne; Jin, Youngnam N.; Rahman, Irfan; Johnson, Gail V. W.

2014-03-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a pivotal transcription factor in the defence against oxidative stress. Here we provide evidence that activation of the Nrf2 pathway reduces the levels of phosphorylated tau by induction of an autophagy adaptor protein NDP52 (also known as CALCOCO2) in neurons. The expression of NDP52, which we show has three antioxidant response elements (AREs) in its promoter region, is strongly induced by Nrf2, and its overexpression facilitates clearance of phosphorylated tau in the presence of an autophagy stimulator. In Nrf2-knockout mice, phosphorylated and sarkosyl-insoluble tau accumulates in the brains concurrent with decreased levels of NDP52. Moreover, NDP52 associates with phosphorylated tau from brain cortical samples of Alzheimer disease cases, and the amount of phosphorylated tau in sarkosyl-insoluble fractions is inversely proportional to that of NDP52. These results suggest that NDP52 plays a key role in autophagy-mediated degradation of phosphorylated tau in vivo.

Carnosic Acid Induces Anti-Inflammatory Effects in Paraquat-Treated SH-SY5Y Cells Through a Mechanism Involving a Crosstalk Between the Nrf2/HO-1 Axis and NF-κB.

PubMed

de Oliveira, Marcos Roberto; de Souza, Izabel Cristina Custódio; Fürstenau, Cristina Ribas

2018-01-01

Carnosic acid (CA) is a phenolic diterpene obtained from Rosmarinus officinalis L. and has demonstrated cytoprotective properties in several experimental models. CA exerts antioxidant effects by upregulating the transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2), which controls the expression of antioxidant and phase II detoxification enzymes. Heme oxygenase-1 (HO-1) expression is modulated by Nrf2 and has been demonstrated as part of the mechanism underlying the CA-induced cytoprotection. Nonetheless, it remains to be studied whether and how HO-1 would mediate CA-elicited anti-inflammatory effects. Therefore, we have investigated here whether and how CA would prevent paraquat (PQ)-induced inflammation-related alterations in human neuroblastoma SH-SY5Y cells. SH-SY5Y cells were pretreated for 12 h with CA at 1 μM before exposure to PQ for further 24 h. CA suppressed the PQ-induced alterations on the levels of interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and cyclooxygenase-2 (COX-2) through a mechanism involving the activation of the Nrf2/HO-1 axis. Furthermore, we observed a crosstalk between the Nrf2/HO-1 signaling pathway and the activation of the nuclear factor-κB (NF-κB) transcription factor, since administration of ZnPP IX (specific inhibitor of HO-1) or Nrf2 knockdown using small interfering RNA (siRNA) abolished the anti-inflammatory effects induced by CA. Moreover, administration of SN50 (specific inhibitor of NF-κB) inhibited the PQ-induced inflammation-related effects in SH-SY5Y cells. Therefore, CA exerted anti-inflammatory effects in SH-SY5Y cells through an Nrf2/HO-1 axis-dependent manner associated with downregulation of NF-κB.

Monomeric cocoa catechins enhance β-cell function by increasing mitochondrial respiration.

PubMed

Rowley, Thomas J; Bitner, Benjamin F; Ray, Jason D; Lathen, Daniel R; Smithson, Andrew T; Dallon, Blake W; Plowman, Chase J; Bikman, Benjamin T; Hansen, Jason M; Dorenkott, Melanie R; Goodrich, Katheryn M; Ye, Liyun; O'Keefe, Sean F; Neilson, Andrew P; Tessem, Jeffery S

2017-11-01

A hallmark of type 2 diabetes (T2D) is β-cell dysfunction and the eventual loss of functional β-cell mass. Therefore, mechanisms that improve or preserve β-cell function could be used to improve the quality of life of individuals with T2D. Studies have shown that monomeric, oligomeric and polymeric cocoa flavanols have different effects on obesity, insulin resistance and glucose tolerance. We hypothesized that these cocoa flavanols may have beneficial effects on β-cell function. INS-1 832/13-derived β-cells and primary rat islets cultured with a monomeric catechin-rich cocoa flavanol fraction demonstrated enhanced glucose-stimulated insulin secretion, while cells cultured with total cocoa extract and with oligomeric or polymeric procyanidin-rich fraction demonstrated no improvement. The increased glucose-stimulated insulin secretion in the presence of the monomeric catechin-rich fraction corresponded with enhanced mitochondrial respiration, suggesting improvements in β-cell fuel utilization. Mitochondrial complex III, IV and V components are up-regulated after culture with the monomer-rich fraction, corresponding with increased cellular ATP production. The monomer-rich fraction improved cellular redox state and increased glutathione concentration, which corresponds with nuclear factor, erythroid 2 like 2 (Nrf2) nuclear localization and expression of Nrf2 target genes including nuclear respiratory factor 1 (Nrf1) and GA binding protein transcription factor alpha subunit (GABPA), essential genes for increasing mitochondrial function. We propose a model by which monomeric cocoa catechins improve the cellular redox state, resulting in Nrf2 nuclear migration and up-regulation of genes critical for mitochondrial respiration, glucose-stimulated insulin secretion and ultimately improved β-cell function. These results suggest a mechanism by which monomeric cocoa catechins exert their effects as an effective complementary strategy to benefit T2D patients. Copyright © 2017 Elsevier Inc. All rights reserved.

[Synergism inhibition of curcumin combined with cisplatin on T24 bladder carcinoma cells and its related mechanism].

PubMed

Zhang, Shao-nan; Yong, Qun; Wu, Xin-li; Liu, Xiao-ping

2014-11-01

To investigate the synergism inhibition of curcumin combined with cisplatin on T24 bladder carcinoma cells and the down-regulating effect of curcumin on the Keapl-Nrf2 pathway, a well recognized anti-drug pathway in almost drugged tumor cells. T24 cells were cultured and treated with increasing concentrations of curcumin(5 ,10 and 20 µmol/mL) combined with cisplatin(30 µg/mL) for 24 hours. The inhibitory effects on T24 cells were tested with MTI colorimetric assay. Nuclear Nrf2 and Keapl , cytoplasmic Keapl and two typical phase II enzymes (GSTP1 and NQOl) were checked with Western blotting. The proliferation of T24 cells was significantly inhibited by different concentrations of curcumin combined with cisplatin. After the treatment with different concentrations of curcumin, Nuclear Nrf2 was decreased but Keapl was increased, and GSTP1 and NQO1 were decreased. Synergism inhibition of curcumin combined with cisplatin on T24 bladder carcinoma cells is observed in this research. The Keapl-Nrf2 pathway in T24 cells is down-regulated by curcumin. The expression of typical phase I enzymes (GSTP1 and NQO1) mediated by Nrf2 are decreased by curcumin. The sensitivity of tumor cells to chemotherapeutic drugs is then enhanced. These may be the mechanism of synergism effect of curcumin combined with cisplatin.

Nrf2 mediates redox adaptation in NOX4-overexpressed non-small cell lung cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu, Qipeng; Yao, Bei; Li, Ning

The redox adaptation mechanisms in cancer cells are very complex and remain largely unclear. Our previous studies have confirmed that NADPH oxidase 4 (NOX4) is abundantly expressed in non-small cell lung cancer (NSCLC) and confers apoptosis resistance on NSCLC cells. However, the comprehensive mechanisms for NOX4-mediated oxidative resistance of cancer cells remain still undentified. The present study found that NOX4-derived H{sub 2}O{sub 2} enhanced the nuclear factor erythroid 2-related factor 2 (Nrf2) stability via disruption of redox-dependent proteasomal degradation and stimulated its activity through activation of PI3K signaling. Specifically, the results showed that ectopic NOX4 expression did not induce apoptosismore » of A549 cells; however, inhibition of Nrf2 resulted in obvious apoptotic death of NOX4-overexpressed A549 cells, accompanied by a significant increase in H{sub 2}O{sub 2} level and decrease in GSH content. Besides, inhibition of Nrf2 could suppress cell growth and efficiently reverse the enhancement effect of NOX4 on cell growth. The in vivo data confirmed that inhibition of Nrf2 could interfere apoptosis resistance in NOX4-overexpressed A549 tumors and led to cell growth inhibition. In conclusion, these results reveal that Nrf2 is critically involved in redox adaptation regulation in NOX4-overexpressed NSCLC cells. Therefore, NOX4 and Nrf2 may be promising combination targets against malignant progression of NSCLC. - Highlights: • NOX4-derived H{sub 2}O{sub 2} upregulates Nrf2 expression and activity in NSCLC. • Nrf2 confers apoptosis resistance in NOX4-overexpressed NSCLC cells. • Inhibition of Nrf2 reverses the enhancement effect of NOX4 on cell growth.« less

Bardoxolone methyl (BARD) ameliorates aristolochic acid (AA)-induced acute kidney injury through Nrf2 pathway.

PubMed

Wu, Juan; Liu, Xinhui; Fan, Jinjin; Chen, Wenfang; Wang, Juan; Zeng, Youjia; Feng, Xiaorang; Yu, Xueqing; Yang, Xiao

2014-04-06

Bardoxolone methyl (BARD) is an antioxidant modulator that acts through induction of the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway. This study aimed to investigate the role of BARD in protecting kidneys from aristolochic acid (AA)-induced acute kidney injury (AKI). Male C57BL/6 mice received intraperitoneal (i.p.) injections of aristolochic acid I (AAI) (5mg/kg/day) for 5 days to produce acute AA nephropathy (AAN) model. BARD (10mg/kg/day, i.p.) was applied for 7 consecutive days, starting 2 days prior to AAI administration. The mice in the AA group showed AKI as evidenced by worsening kidney function evaluated by blood urea nitrogen (BUN) and serum creatinine (SCr) levels, and severe tubulointerstitial injury marked by massive tubule necrosis in kidney tissues. BARD significantly reduced BUN and SCr levels which were elevated by AAI. Additionally, AAI-induced histopathological renal damage was ameliorated by BARD. Furthermore, the expression of Nrf2 was reduced, and its repressor Kelch-like ECH-associated protein 1 (Keap1) was increased significantly, whereas heme oxygenase-1 (HO-1) was upregulated and NAD(P)H quinone oxidoreductase-1 (NQO1) was barely increased in the cytoplasm of tubules in kidneys after treatment with AAI. BARD significantly upregulated renal Nrf2, NQO1 and HO-1 expression and downregulated Keap1 expression compared with those in the AA group. Moreover, it was found that Nrf2 was expressed both in the cytoplasm and nuclear of glomeruli and tubules, whereas NQO1 and HO-1 were localized in the cytoplasm of tubules only. In conclusion, AA-induced acute renal injury was associated with impaired Nrf2 activation and expression of its downstream target genes in renal tissues. BARD prevented renal damage induced by AAI, and this renoprotective effect may be exerted by activating the Nrf2 signaling pathway and increasing expression of the downstream target genes. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Scopoletin protects against methylglyoxal-induced hyperglycemia and insulin resistance mediated by suppression of advanced glycation endproducts (AGEs) generation and anti-glycation.

PubMed

Chang, Wen-Chang; Wu, Shinn-Chih; Xu, Kun-Di; Liao, Bo-Chieh; Wu, Jia-Feng; Cheng, An-Sheng

2015-02-09

Recently, several types of foods and drinks, including coffee, cream, and cake, have been found to result in high methylglyoxal (MG) levels in the plasma, thus causing both nutritional and health concerns. MG can be metabolized by phase-II enzymes in liver through the positive regulation of nuclear factor-erythroid 2-related factor 2 (Nrf2). In this study, we investigated the ability of scopoletin (SP) to protect against MG-induced hyperglycemia and insulin resistance. Recently, SP was shown to be a peroxisome proliferator-activated receptor-γ activator to elevate insulin sensitivity. We investigated the effects of oral administration of SP on the metabolic, biochemical, and molecular abnormalities characteristic of type 2 diabetes in MG-treated Wistar rats to understand the potential mechanism of scopoletin for diabetes protection. Our results suggested that SP activated Nrf2 by Ser40 phosphorylation, resulting in the metabolism of MG into d-lactic acid and the inhibition of AGEs generation, which reduced the accumulation of AGEs in the livers of MG-induced rats. In this manner, SP improved the results of the oral glucose tolerance test and dyslipidemia. Moreover, SP also increased the plasma translocation of glucose transporter-2 and promoted Akt phosphorylation caused by insulin treatment in MG-treated FL83B hepatocytes. In contrast, SP effectively suppressed protein tyrosine phosphatase 1B (PTP1B) expression, thereby alleviating insulin resistance. These findings suggest that SP acts as an anti-glycation and anti-diabetic agent, and thus has therapeutic potential for the prevention of diabetes.

JSH-23 prevents depressive-like behaviors in mice subjected to chronic mild stress: Effects on inflammation and antioxidant defense in the hippocampus.

PubMed

Wang, Qi; Dong, Xiaomei; Li, Nannan; Wang, Yan; Guan, Xiaofeng; Lin, Yiwei; Kang, Jiguang; Zhang, Xia; Zhang, Yuchen; Li, Xiaobai; Xu, Tianchao

2018-06-01

Nuclear factor-kappa B (NF-κB), which is reported to play an important role in the pathogenesis of depression, also has a central role in the genesis and progression of inflammation. Here, we have targeted the nuclear translocation of NF-κB using 4-methyl-N1-(3-phenyl-propyl)-benzene-1,2-diamine (JSH-23) to elucidate its role in depression. We investigated the antidepressant-like effects of JSH-23 in the chronic mild stress (CMS) mouse model, which is a valid, reasonably reliable, and useful model of depression. The antidepressant-like effects of JSH-23 were evaluated using the sucrose preference test (SPT) and the forced swimming test (FST). We also assessed inflammatory markers [interleukin (IL)-6 and tumor necrosis factor-α (TNF-α)] and components of antioxidant defense [superoxide dismutase (SOD) and nuclear factor erythroid-2-related factor 2 (Nrf 2)] in the hippocampus. Fluoxetine, a classical antidepressant, was used in this study as a positive control. Administration of JSH-23 significantly prevented the decreased sucrose preference in the SPT and prevented the increased immobility time in the FST caused by CMS, but had no effect on locomotor activity. Expression of NF-κB p65 protein in the hippocampus was decreased, and elevated levels of IL-6 and TNF-α were reduced, after JSH-23 administration. In addition to its anti-inflammatory effect, JSH-23 treatment increased the expression of SOD and Nrf 2 in the hippocampus, suggesting that it strengthens antioxidant defense. The current study demonstrated that inhibiting the NF-κB signaling cascade using JSH-23 prevented depressive-like behaviors by decreasing inflammation and improving antioxidant defense in the hippocampus. We concluded that NF-κB activation plays an important role in the pathophysiology of depression and that targeting NF-κB signaling may provide a novel and effective therapy for depression. Additional preclinical studies and clinical trials are, however, needed to further elucidate the effects of this therapeutic strategy. Copyright © 2018 Elsevier Inc. All rights reserved.

Nrf2 deficiency potentiates methamphetamine-induced dopaminergic axonal damage and gliosis in the striatum.

PubMed

Granado, Noelia; Lastres-Becker, Isabel; Ares-Santos, Sara; Oliva, Idaira; Martin, Eduardo; Cuadrado, Antonio; Moratalla, Rosario

2011-12-01

Oxidative stress that correlates with damage to nigrostriatal dopaminergic neurons and reactive gliosis in the basal ganglia is a hallmark of methamphetamine (METH) toxicity. In this study, we analyzed the protective role of the transcription factor Nrf2 (nuclear factor-erythroid 2-related factor 2), a master regulator of redox homeostasis, in METH-induced neurotoxicity. We found that Nrf2 deficiency exacerbated METH-induced damage to dopamine neurons, shown by an increase in loss of tyrosine hydroxylase (TH)- and dopamine transporter (DAT)-containing fibers in striatum. Consistent with these effects, Nrf2 deficiency potentiated glial activation, indicated by increased striatal expression of markers for microglia (Mac-1 and Iba-1) and astroglia (GFAP) one day after METH administration. At the same time, Nrf2 inactivation dramatically potentiated the increase in TNFα mRNA and IL-15 protein expression in GFAP+ cells in the striatum. In sharp contrast to the potentiation of striatal damage, Nrf2 deficiency did not affect METH-induced dopaminergic neuron death or expression of glial markers or proinflammatory molecules in the substantia nigra. This study uncovers a new role for Nrf2 in protection against METH-induced inflammatory and oxidative stress and striatal degeneration. Copyright © 2011 Wiley‐Liss, Inc.

4-Methoxychalcone Enhances Cisplatin-Induced Oxidative Stress and Cytotoxicity by Inhibiting the Nrf2/ARE-Mediated Defense Mechanism in A549 Lung Cancer Cells

PubMed Central

Lim, Juhee; Lee, Sung Ho; Cho, Sera; Lee, Ik-Soo; Kang, Bok Yun; Choi, Hyun Jin

2013-01-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a key transcriptional regulator for the protection of cells against oxidative and xenobiotic stresses. Recent studies have demonstrated that high constitutive expression of Nrf2 is observed in many types of cancer cells showing resistance to anti-cancer drugs, suggesting that the suppression of overexpressed Nrf2 could be an attractive therapeutic strategy to overcome cancer drug resistance. In the present study, we aimed to find small molecule compounds that enhance the sensitivity of tumor cells to cisplatin induced cytotoxicity by suppressing Nrf2-mediated defense mechanism. A549 lung cancer cells were shown to be more resistant to the anti-cancer drug cisplatin than HEK293 cells, with higher Nrf2 signaling activity; constitutively high amounts of Nrf2-downstream target proteins were observed in A549 cells. Among the three chalcone derivatives 4-methoxy-chalcone (4-MC), hesperidin methylchalcone, and neohesperidin dihydrochalcone, 4-MC was found to suppress transcriptional activity of Nrf2 in A549 cells but to activate it in HEK293 cells. 4-MC was also shown to down-regulate expression of Nrf2 and the downstream phase II detoxifying enzyme NQO1 in A549 cells. The PI3K/Akt pathway was found to be involved in the 4-MC-induced inhibition of Nrf2/ARE activity in A549 cells. This inhibition of Nrf2 signaling results in the accelerated generation of reactive oxygen species and exacerbation of cytotoxicity in cisplatin-treated A549 cells. Taken together, these results suggest that the small molecule compound 4-MC could be used to enhance the sensitivity of tumor cells to the therapeutic effect of cisplatin through the regulation of Nrf2/ARE signaling. PMID:24046186

4-methoxychalcone enhances cisplatin-induced oxidative stress and cytotoxicity by inhibiting the Nrf2/ARE-mediated defense mechanism in A549 lung cancer cells.

PubMed

Lim, Juhee; Lee, Sung Ho; Cho, Sera; Lee, Ik-Soo; Kang, Bok Yun; Choi, Hyun Jin

2013-10-01

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a key transcriptional regulator for the protection of cells against oxidative and xenobiotic stresses. Recent studies have demonstrated that high constitutive expression of Nrf2 is observed in many types of cancer cells showing resistance to anti-cancer drugs, suggesting that the suppression of overexpressed Nrf2 could be an attractive therapeutic strategy to overcome cancer drug resistance. In the present study, we aimed to find small molecule compounds that enhance the sensitivity of tumor cells to cisplatin induced cytotoxicity by suppressing Nrf2-mediated defense mechanism. A549 lung cancer cells were shown to be more resistant to the anti-cancer drug cisplatin than HEK293 cells, with higher Nrf2 signaling activity; constitutively high amounts of Nrf2-downstream target proteins were observed in A549 cells. Among the three chalcone derivatives 4-methoxy-chalcone (4-MC), hesperidin methylchalcone, and neohesperidin dihydrochalcone, 4-MC was found to suppress transcriptional activity of Nrf2 in A549 cells but to activate it in HEK293 cells. 4-MC was also shown to down-regulate expression of Nrf2 and the downstream phase II detoxifying enzyme NQO1 in A549 cells. The PI3K/Akt pathway was found to be involved in the 4-MC-induced inhibition of Nrf2/ARE activity in A549 cells. This inhibition of Nrf2 signaling results in the accelerated generation of reactive oxygen species and exacerbation of cytotoxicity in cisplatin-treated A549 cells. Taken together, these results suggest that the small molecule compound 4-MC could be used to enhance the sensitivity of tumor cells to the therapeutic effect of cisplatin through the regulation of Nrf2/ARE signaling.

The effect of Mastin® on expression of Nrf2 in the rat heart with subtotally nephrectomy chronic Kidney disease model

NASA Astrophysics Data System (ADS)

Nathania, J.; Soetikno, V.

2017-08-01

Chronic kidney disease (CKD) is increasingly prevalent in Indonesia and worldwide. One of the major causes of morbidity and mortality in CKD is the complication of cardiovascular disease. Mastin® is a supplement that is locally produced in Indonesia and is made from extract of mangosteen pericarp, which is reported to have antioxidative, anti-inflammatory, and antitumor properties. The present study aimed to investigate whether Mastin® could improve antioxidant responses in the rat heart during CKD by measuring the expression of nuclear factor erythroid-2-related factor (Nrf)2, a master regulator of antioxidant response elements. RNA was extracted from the heart tissue of three groups of rats: a normal group, a nephrectomy group, and a nephrectomy with Mastin® group. Two-step real-time RT-PCR was then conducted to calculate the relative expression of the Nrf2 gene. Nrf2 expression was markedly decreased in the nephrectomy group vs the normal group, but slightly increas ed in the nephrectomy with Mastin® group vs the nephrectomy group. CKD resulted in impaired activation of the Nrf2 pathway in the rat heart. Although the administration of Mastin® slightly increased Nrf2 expression, it was not enough to confer cardioprotective effects through the Nrf2 pathway.

Aldosterone Activates Transcription Factor Nrf2 in Kidney Cells Both In Vitro and In Vivo

PubMed Central

Oteiza, Patricia I.; Link, Samuel; Hey, Valentin; Stopper, Helga; Schupp, Nicole

2014-01-01

Abstract Aims: An increased kidney cancer risk was found in hypertensive patients, who frequently exhibit hyperaldosteronism, known to contribute to kidney injury, with oxidative stress playing an important role. The capacity of kidney cells to up-regulate transcription factor nuclear factor-erythroid-2-related factor 2 (Nrf2), a key regulator of the cellular antioxidative defense, as a prevention of aldosterone-induced oxidative damage was investigated both in vitro and in vivo. Results: Aldosterone activated Nrf2 and increased the expression of enzymes involved in glutathione (GSH) synthesis and detoxification. This activation depended on the mineralocorticoid receptor (MR) and oxidative stress. In vitro, Nrf2 activation, GSH amounts, and target gene levels decreased after 24 h, while oxidant levels remained high. Nrf2 activation could not protect cells against oxidative DNA damage, as aldosterone-induced double-strand breaks and 7,8-dihydro-8-oxo-guanine (8-oxodG) lesions steadily rose. The Nrf2 activator sulforaphane enhanced the Nrf2 response both in vitro and in vivo, thereby preventing aldosterone-induced DNA damage. In vivo, Nrf2 activation further had beneficial effects on the aldosterone-caused blood pressure increase and loss of kidney function. Innovation: This is the first study showing the activation of Nrf2 by aldosterone. Moreover, the results identify sulforaphane as a substance that is capable of preventing aldosterone-induced damage both in vivo and in vitro. Conclusion: Aldosterone-induced Nrf2 adaptive response cannot neutralize oxidative actions of chronically increased aldosterone, which, therefore could be causally involved in the increased cancer incidence of hypertensive individuals. Enhancing the cellular antioxidative defense with sulforaphane might exhibit beneficial effects. Antioxid. Redox Signal. 21, 2126–2142. PMID:24512358

Britanin Ameliorates Cerebral Ischemia-Reperfusion Injury by Inducing the Nrf2 Protective Pathway.

PubMed

Wu, Guozhen; Zhu, Lili; Yuan, Xing; Chen, Hao; Xiong, Rui; Zhang, Shoude; Cheng, Hao; Shen, Yunheng; An, Huazhang; Li, Tiejun; Li, Honglin; Zhang, Weidong

2017-10-10

Oxidative stress is considered the major cause of tissue injury after cerebral ischemia. The nuclear factor erythroid 2-related factor 2 (Nrf2) pathway is one of the most important defensive mechanisms against oxidative stresses and has been confirmed as a target for stroke treatment. Thus, we desired to find new Nrf2 activators and test their neuronal protective activity both in vivo and in vitro. The herb-derived compound, Britanin, is a potent inducer of the Nrf2 system. Britanin can induce the expression of protective enzymes and reverse oxygen-glucose deprivation, followed by reperfusion (OGD-R)-induced neuronal injury in primary cortical neurons in vitro. Furthermore, the administration of Britanin significantly ameliorated middle cerebral artery occlusion-reperfusion (MCAO-R) insult in vivo. We report here the crystal structure of the complex of Britanin and the BTB domain of Keap1. Britanin selectively binds to a conserved cysteine residue, cysteine 151, of Keap1 and inhibits Keap1-mediated ubiquitination of Nrf2, leading to induction of the Nrf2 pathway. Britanin is a potent inducer of Nrf2. The complex crystal structure of Britanin and the BTB domain of Keap1 help clarify the mechanism of Nrf2 induction. Britanin was proven to protect primary cortical neurons against OGD-R-induced injury in an Nrf2-dependant way. Additionally, Britanin had excellent cerebroprotective effect in an MCAO-R model. Our results demonstrate that the natural product Britanin with potent Nrf2-activating and neural protective activities both in vitro and in vivo could be developed into a cerebroprotective therapeutic agent. Antioxid. Redox Signal. 27, 754-768.

High Nrf2 expression in alveolar type I pneumocytes is associated with low recurrences in primary spontaneous pneumothorax.

PubMed

Chen, Yu-Wen; Chiu, Wen-Chin; Chou, Shah-Hwa; Su, Yu-Han; Huang, Ying-Fong; Lee, Yen-Lung; Yuan, Shyng-Shiou F; Lee, Yi-Chen

2017-10-01

Recurrent primary spontaneous pneumothorax (PSP) is a troublesome problem and a major concern for the patients. This study examined whether nuclear factor erythroid 2-related factor 2 (Nrf2) expression in alveolar type I pneumocytes was associated with the clinical manifestations of PSP patients including disease recurrence. Eighty-eight PSP patients who were managed with needlescopic video-assisted thoracoscopic surgery (NVATS) were included in this study. Immunohistochemistry (IHC) was assessed to determine Nrf2 expression in resected lung tissues and the results were correlated with clinicopathological characteristics by the chi-square or the Fisher's exact test. The prognostic value of Nrf2 for overall recurrence was evaluated by univariate and multivariable Cox regression model. The expression of Nrf2 was observed in type I pneumocytes of lung tissues from PSP patients by IHC. We found that low Nrf2 expression in PSP patients, especially in young (age ≤ 20, p = 0.033) and body mass index (BMI) ≥18 kg/m 2 (p = 0.019) groups, was significantly correlated with PSP recurrence. In the univariate and multivariate analyses, high Nrf2 expression was a significant protective factor for overall recurrence in PSP patients (univariate: p = 0.026; multivariate: p = 0.004). The expression level of Nrf2 in alveolar type I pneumocytes was a potential factor involved in PSP recurrence. Our findings suggest that elevated Nrf2 expression in PSP patients may be a promising way for reducing PSP recurrence. Copyright © 2017. Published by Elsevier Taiwan.

Propolis reversed cigarette smoke-induced emphysema through macrophage alternative activation independent of Nrf2.

PubMed

Barroso, Marina Valente; Cattani-Cavalieri, Isabella; de Brito-Gitirana, Lycia; Fautrel, Alain; Lagente, Vincent; Schmidt, Martina; Porto, Luís Cristóvão; Romana-Souza, Bruna; Valença, Samuel Santos; Lanzetti, Manuella

2017-10-15

Chronic obstructive pulmonary disease (COPD) is an incurable and progressive disease. Emphysema is the principal manifestation of COPD, and the main cause of this condition is cigarette smoke (CS). Natural products have shown antioxidant and anti-inflammatory properties that can prevent acute lung inflammation and emphysema, but there are few reports in the literature regarding therapeutic approaches to emphysema. We hypothesized that supplementation with natural extracts would repair lung damage in emphysema caused by CS exposure. Mice were exposed to 60days of CS and then treated or not with three different natural extracts (mate tea, grape and propolis) orally for additional 60days. Histological analysis revealed significant improvements in lung histoarchitecture, with recovery of alveolar spaces in all groups treated with natural extracts. Propolis was also able to recovery alveolar septa and elastic fibers. Propolis also increased MMP-2 and decreased MMP-12 expression, favoring the process of tissue repair. Additionally, propolis recruited leukocytes, including macrophages, without ROS release. These findings led us to investigate the profile of these macrophages, and we showed that propolis could promote macrophage alternative activation, thus increasing the number of arginase-positive cells and IL-10 levels and favoring an anti-inflammatory microenvironment. We further investigated the participation of Nrf2 in lung repair, but no Nrf2 translocation to the nucleus was observed in lung cells. Proteins and enzymes related to Nrf2 were not altered, other than NQO1, which seemed to be activated by propolis in a Nrf2-independent manner. Finally, propolis downregulated IGF1 expression. In conclusion, propolis promoted lung repair in a mouse emphysema model via macrophage polarization from M1 to M2 in parallel to the downregulation of IGF1 expression in a Nrf2-independent manner. Copyright © 2017 Elsevier Ltd. All rights reserved.

Brain ischemic preconditioning protects against ischemic injury and preserves the blood-brain barrier via oxidative signaling and Nrf2 activation.

PubMed

Yang, Tuo; Sun, Yang; Mao, Leilei; Zhang, Meijuan; Li, Qianqian; Zhang, Lili; Shi, Yejie; Leak, Rehana K; Chen, Jun; Zhang, Feng

2018-05-06

Brain ischemic preconditioning (IPC) with mild ischemic episodes is well known to protect the brain against subsequent ischemic challenges. However, the underlying mechanisms are poorly understood. Here we demonstrate the critical role of the master redox transcription factor, nuclear factor (erythroid-derived 2)-like 2 (Nrf2), in IPC-mediated neuroprotection and blood-brain barrier (BBB) preservation. We report that IPC causes generation of endogenous lipid electrophiles, including 4-hydroxy-2-nonenal (4-HNE), which release Nrf2 from inhibition by Keap1 (via Keap1-C288) and inhibition by glycogen synthase kinase 3β (via GSK3β-C199). Nrf2 then induces expression of its target genes, including a new target, cadherin 5, a key component of adherens junctions of the BBB. These effects culminate in mitigation of BBB leakage and of neurological deficits after stroke. Collectively, these studies are the first to demonstrate that IPC protects the BBB against ischemic injury by generation of endogenous electrophiles and activation of the Nrf2 pathway through inhibition of Keap1- and GSK3β-dependent Nrf2 degradation. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Protective function of pyridoxamine on retinal photoreceptor cells via activation of the p‑Erk1/2/Nrf2/Trx/ASK1 signalling pathway in diabetic mice.

PubMed

Ren, Xiang; Sun, Hong; Zhang, Chenghong; Li, Chen; Wang, Jinlei; Shen, Jie; Yu, Dong; Kong, Li

2016-07-01

The present study aimed to investigate the mechanisms that mediate the protective effects of pyridoxamine (PM) on light‑damaged retinal photoreceptor cells in diabetic mice. A high‑fat diet and streptozotocin were used to induce a mouse model of type II diabetes. During the experiment, mice were divided the mice into three types of group, as follows: Control groups (negative control and light‑damaged groups); experimental groups (diabetic and diabetic light‑damaged groups); and treatment groups (25, 50 and 100 mg/kg PM‑treated groups). Using hematoxylin‑eosin staining, the number of nuclear layer cells were counted. Western blotting and immunohistochemistry were performed to measure the levels of thioredoxin (Trx), phospho‑extracellular signal‑regulated kinase 1/2 (p‑Erk1/2), nuclear factor erythroid 2‑related factor 2 (Nrf2) and apoptosis signal‑regulating kinase 1 (ASK1). The photoreceptor cell count in the outer nuclear layer of the light‑damaged, diabetic control and diabetic light‑damaged groups were significantly reduced compared with the negative control group (P<0.001). The cell counts in the PM‑treated groups were significantly increased compared with the diabetic group (P<0.001). Compared with the negative control group, the light‑damaged, diabetic and diabetic light‑damaged groups exhibited significantly decreased Trx, p‑Erk1/2 and Nrf2 expression levels (P<0.001), and significantly increased ASK1 expression levels (P<0.001). However, in the PM‑treated groups, Trx, p‑Erk1/2 and Nrf2 expression levels were significantly increased (P<0.001), and ASK1 expression was significantly decreased (P<0.001). The results of the present study demonstrate that PM protects retinal photoreceptor cells against light damage in diabetic mice, and that its mechanism may be associated with the upregulation of Trx, p‑Erk1/2 and Nrf2 expression, and the downregulation of ASK1 expression.

An Essential Role of NRF2 in Diabetic Wound Healing.

PubMed

Long, Min; Rojo de la Vega, Montserrat; Wen, Qing; Bharara, Manish; Jiang, Tao; Zhang, Rui; Zhou, Shiwen; Wong, Pak K; Wondrak, Georg T; Zheng, Hongting; Zhang, Donna D

2016-03-01

The high mortality and disability of diabetic nonhealing skin ulcers create an urgent need for the development of more efficacious strategies targeting diabetic wound healing. In the current study, using human clinical specimens, we show that perilesional skin tissues from patients with diabetes are under more severe oxidative stress and display higher activation of the nuclear factor-E2-related factor 2 (NRF2)-mediated antioxidant response than perilesional skin tissues from normoglycemic patients. In a streptozotocin-induced diabetes mouse model, Nrf2(-/-) mice have delayed wound closure rates compared with Nrf2(+/+) mice, which is, at least partially, due to greater oxidative DNA damage, low transforming growth factor-β1 (TGF-β1) and high matrix metalloproteinase 9 (MMP9) expression, and increased apoptosis. More importantly, pharmacological activation of the NRF2 pathway significantly improves diabetic wound healing. In vitro experiments in human immortalized keratinocyte cells confirm that NRF2 contributes to wound healing by alleviating oxidative stress, increasing proliferation and migration, decreasing apoptosis, and increasing the expression of TGF-β1 and lowering MMP9 under high-glucose conditions. This study indicates an essential role for NRF2 in diabetic wound healing and the therapeutic benefits of activating NRF2 in this disease, laying the foundation for future clinical trials using NRF2 activators in treating diabetic skin ulcers. © 2016 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

A Geant4-based Simulation to Evaluate the Feasibility of Using Nuclear Resonance Fluorescence (NRF) in Determining Atomic Compositions of Body Tissue in Cancer Diagnostics and Irradiation

NASA Astrophysics Data System (ADS)

Gilbo, Yekaterina; Wijesooriya, Krishni; Liyanage, Nilanga

2017-01-01

Customarily applied in homeland security for identifying concealed explosives and chemical weapons, NRF (Nuclear Resonance Fluorescence) may have high potential in determining atomic compositions of body tissue. High energy photons incident on a target excite the target nuclei causing characteristic re-emission of resonance photons. As the nuclei of each isotope have well-defined excitation energies, NRF uniquely indicates the isotopic content of the target. NRF radiation corresponding to nuclear isotopes present in the human body is emitted during radiotherapy based on Bremsstrahlung photons generated in a linear electron accelerator. We have developed a Geant4 simulation in order to help assess NRF capabilities in detecting, mapping, and characterizing tumors. We have imported a digital phantom into the simulation using anatomical data linked to known chemical compositions of various tissues. Work is ongoing to implement the University of Virginia's cancer center treatment setup and patient geometry, and to collect and analyze the simulation's physics quantities to evaluate the potential of NRF for medical imaging applications. Preliminary results will be presented.

Sulforaphane and Other Nutrigenomic Nrf2 Activators: Can the Clinician's Expectation Be Matched by the Reality?

PubMed Central

Houghton, Christine A.; Fassett, Robert G.; Coombes, Jeff S.

2016-01-01

The recognition that food-derived nonnutrient molecules can modulate gene expression to influence intracellular molecular mechanisms has seen the emergence of the fields of nutrigenomics and nutrigenetics. The aim of this review is to describe the properties of nutrigenomic activators of transcription factor Nrf2 (nuclear factor erythroid 2-related factor 2), comparing the potential for sulforaphane and other phytochemicals to demonstrate clinical efficacy as complementary medicines. Broccoli-derived sulforaphane emerges as a phytochemical with this capability, with oral doses capable of favourably modifying genes associated with chemoprevention. Compared with widely used phytochemical-based supplements like curcumin, silymarin, and resveratrol, sulforaphane more potently activates Nrf2 to induce the expression of a battery of cytoprotective genes. By virtue of its lipophilic nature and low molecular weight, sulforaphane displays significantly higher bioavailability than the polyphenol-based dietary supplements that also activate Nrf2. Nrf2 activation induces cytoprotective genes such as those playing key roles in cellular defense mechanisms including redox status and detoxification. Both its high bioavailability and significant Nrf2 inducer capacity contribute to the therapeutic potential of sulforaphane-yielding supplements. PMID:26881038

The Role of the Nrf2/ARE Antioxidant System in Preventing Cardiovascular Diseases

PubMed Central

Smith, Robert E.; Tran, Kevin; Smith, Cynthia C.; McDonald, Miranda; Shejwalkar, Pushkar; Hara, Kenji

2016-01-01

It is widely believed that consuming foods and beverages that have high concentrations of antioxidants can prevent cardiovascular diseases and many types of cancer. As a result, many articles have been published that give the total antioxidant capacities of foods in vitro. However, many antioxidants behave quite differently in vivo. Some of them, such as resveratrol (in red wine) and epigallocatechin gallate or EGCG (in green tea) can activate the nuclear erythroid-2 like factor-2 (Nrf2) transcription factor. It is a master regulator of endogenous cellular defense mechanisms. Nrf2 controls the expression of many antioxidant and detoxification genes, by binding to antioxidant response elements (AREs) that are commonly found in the promoter region of antioxidant (and other) genes, and that control expression of those genes. The mechanisms by which Nrf2 relieves oxidative stress and limits cardiac injury as well as the progression to heart failure are described. Also, the ability of statins to induce Nrf2 in the heart, brain, lung, and liver is mentioned. However, there is a negative side of Nrf2. When over-activated, it can cause (not prevent) cardiovascular diseases and multi-drug resistance cancer. PMID:28933413

Regressive Effect of Myricetin on Hepatic Steatosis in Mice Fed a High-Fat Diet

PubMed Central

Xia, Shu-Fang; Le, Guo-Wei; Wang, Peng; Qiu, Yu-Yu; Jiang, Yu-Yu; Tang, Xue

2016-01-01

Myricetin is an effective antioxidant in the treatment of obesity and obesity-related metabolic disorders. The objective of this study was to explore the regressive effect of myricetin on pre-existing hepatic steatosis induced by high-fat diet (HFD). C57BL/6 mice were fed either a standard diet or a HFD for 12 weeks and then half of the mice were treated with myricetin (0.12% in the diet, w/w) while on their respective diets for further 12 weeks. Myricetin treatment significantly alleviated HFD-induced steatosis, decreased hepatic lipid accumulation and thiobarbituric acid reactive substance (TBARS) levels, and increased antioxidative enzyme activities, including catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx) activities. Microarray analysis of hepatic gene expression profiles showed that myricetin significantly altered the expression profiles of 177 genes which were involved in 12 biological pathways, including the peroxisome proliferator activated receptor (PPAR) signaling pathway and peroxisome. Further research indicated that myricetin elevated hepatic nuclear Nrf2 translocation, increased the protein expression of heme oxygenase-1 (HO-1) and NAD(P)H quinone dehydrogenase 1 (NQO1), reduced the protein expression of PPARγ, and normalized the expressions of genes that were involved in peroxisome and the PPAR signaling pathway. Our data indicated that myricetin might represent an effective therapeutic agent to treat HFD-induced hepatic steatosis via activating the Nrf2 pathway and the PPAR signaling pathway. PMID:27973423

Taxifolin Activates the Nrf2 Anti-Oxidative Stress Pathway in Mouse Skin Epidermal JB6 P+ Cells through Epigenetic Modifications

PubMed Central

Kuang, Haixue; Tang, Zhenqiu; Zhang, Chengyue; Wang, Zhibin; Li, Wenji; Yang, Chunjuan; Wang, Qiuhong; Yang, Bingyou; Kong, Ah-Ng

2017-01-01

Nuclear factor erythroid-2 related factor 2 (Nrf2) is a vital transcription factor that regulates the anti-oxidative defense system. Previous reports suggested that the expression of the Nrf2 gene can be regulated by epigenetic modifications. The potential epigenetic effect of taxifolin (TAX), a potent cancer chemopreventive agent, in skin cancer chemoprotection is unknown. In this study, we investigated how Nrf2 is epigenetically regulated by TAX in JB6 P+ cells. TAX was found to inhibit the 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced colony formation of JB6 P+ cells. TAX induced antioxidant response element (ARE)-luciferase activity in HepG2-C8 cells and up-regulated mRNA and protein levels of Nrf2 and its downstream genes heme oxygenase-1 (HO-1) and NAD(P)H quinone oxidoreductase 1 (NQO1), in JB6 P+ cells. Furthermore, bisulfite genomic sequencing revealed that TAX treatment reduces the methylation level of the first 15 CpGs sites in the Nrf2 promoter. Western blotting showed that TAX inhibits the expression levels of DNA methyltransferase (DNMT) and histone deacetylase (HDAC) proteins. In summary, our results revealed that TAX can induce expression of Nrf2 and its downstream target genes in JB6 P+ cells by CpG demethylation. These finding suggest that TAX may exhibit a skin cancer preventive effect by activating Nrf2 via an epigenetic pathway. PMID:28714938

The involvement of Nrf2 in the protective effects of (-)-Epigallocatechin-3-gallate (EGCG) on NaAsO2-induced hepatotoxicity.

PubMed

Han, Xiao-Dong; Zhang, Yan-Yan; Wang, Ke-Lei; Huang, Yong-Pan; Yang, Zhong-Bao; Liu, Zhi

2017-09-12

Arsenic exposure produces hepatotoxicity. The common mechanism determining its toxicity is the generation of oxidative stress. Oxidative stress induced by arsenic leads to the activation of nuclear factor erythroid 2-related factor 2 (Nrf2) pathway. (-)-Epigallocatechin-3-gallate (EGCG) possesses a potent antioxidant capacity and exhibits extensive pharmacological activities. This study aims to evaluate effects of EGCG on arsenic-induced hepatotoxicity and activation of Nrf2 pathway. Plasma activities of alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, and lactate dehydrogenase were measured; Histological analyses were conducted to observe morphological changes; Biochemical indexes such as oxidative stress (Catalase (CAT), malonyldialdehyde (MDA), superoxide dismutase (SOD), glutathione (GSH), reactive oxygen species (ROS)), Nrf2 signaling related genes ( Nrf2, Nqo1, and Ho-1 ) were assessed. The results showed that EGCG inhibited arsenic-induced hepatic pathological damage, liver ROS level and MDA level. Arsenic decreases the antioxidant enzymes SOD, GPX, and CAT activity and the decrease was inhibited by treatment of EGCG. Furthermore, EGCG attenuated the retention of arsenic in liver tissues and improved the expressions of Nrf2 signaling related genes ( Nrf2, Nqo1, and Ho-1 ). These findings provide evidences that EGCG may be useful for reducing hepatotoxicity associated with oxidative stress by the activation of Nrf2 signaling pathway. Our findings suggest a possible mechanism of antioxidant EGCG in preventing hepatotoxicity, which implicate that EGCG may be a potential treatment for arsenicosis therapy.

The Keap1/Nrf2 pathway in health and disease: from the bench to the clinic.

PubMed

O'Connell, Maria A; Hayes, John D

2015-08-01

The transcription factor nuclear factor-erythroid 2 p45-related factor 2 (Nrf2, with gene called NFE2L2) is a master regulator of the antioxidant response. In the last decade, interest has intensified in this research area as its importance in several physiological and pathological processes has become widely recognized; these include redox signalling and redox homoeostasis, drug metabolism and disposition, intermediary metabolism, cellular adaptation to stress, chemoprevention and chemoresistance, toxicity, inflammation, neurodegeneration, lipogenesis and aging. Regulation of Nrf2 is complex and although much attention has focussed on its repression by Kelch-like ECH-associated protein-1 (Keap1), recently it has become increasingly apparent that it is also controlled by cross-talk with other signalling pathways including the glycogen synthase kinase-3 (GSK-3)-β-transducin repeat-containing protein (β-TrCP) axis, ERAD (endoplasmic reticulum-associated degradation)-associated E3 ubiquitin-protein ligase (Hrd1, also called synoviolin), nuclear factor-kappa B (NF-κB), Notch and AMP kinase. Due to its beneficial role in several diseases, Nrf2 has become a major therapeutic target, with novel natural, synthetic and targeted small molecules currently under investigation to modulate the pathway and in clinical trials. © 2015 Authors; published by Portland Press Limited.

Experimental demonstration of an isotope-sensitive warhead verification technique using nuclear resonance fluorescence.

PubMed

Vavrek, Jayson R; Henderson, Brian S; Danagoulian, Areg

2018-04-24

Future nuclear arms reduction efforts will require technologies to verify that warheads slated for dismantlement are authentic without revealing any sensitive weapons design information to international inspectors. Despite several decades of research, no technology has met these requirements simultaneously. Recent work by Kemp et al. [Kemp RS, Danagoulian A, Macdonald RR, Vavrek JR (2016) Proc Natl Acad Sci USA 113:8618-8623] has produced a novel physical cryptographic verification protocol that approaches this treaty verification problem by exploiting the isotope-specific nature of nuclear resonance fluorescence (NRF) measurements to verify the authenticity of a warhead. To protect sensitive information, the NRF signal from the warhead is convolved with that of an encryption foil that contains key warhead isotopes in amounts unknown to the inspector. The convolved spectrum from a candidate warhead is statistically compared against that from an authenticated template warhead to determine whether the candidate itself is authentic. Here we report on recent proof-of-concept warhead verification experiments conducted at the Massachusetts Institute of Technology. Using high-purity germanium (HPGe) detectors, we measured NRF spectra from the interrogation of proxy "genuine" and "hoax" objects by a 2.52 MeV endpoint bremsstrahlung beam. The observed differences in NRF intensities near 2.2 MeV indicate that the physical cryptographic protocol can distinguish between proxy genuine and hoax objects with high confidence in realistic measurement times.

Ebselen attenuates cisplatin-induced ROS generation through Nrf2 activation in auditory cells.

PubMed

Kim, Se-Jin; Park, Channy; Han, A Lum; Youn, Myung-Ja; Lee, Jeong-Han; Kim, Yunha; Kim, Eun-Sook; Kim, Hyung-Jin; Kim, Jin-Kyung; Lee, Ho-Kyun; Chung, Sang-Young; So, Hongseob; Park, Raekil

2009-05-01

Ebselen, an organoselenium compound that acts as a glutathione peroxidase mimetic, has been demonstrated to possess antioxidant and anti-inflammatory activities. However, the molecular mechanism underlying this effect is not fully understood in auditory cells. The purpose of the present study is to investigate the protective effect of ebselen against cisplatin-induced toxicity in HEI-OC1 auditory cells, organotypic cultures of cochlear explants from two-day postnatal rats (P(2)) and adult Balb/C mice. Pretreatment with ebselen ameliorated apoptotic death induced by cisplatin in HEI-OC1 cells and organotypic cultures of Corti's organ. Ebselen pretreatment also significantly suppressed cisplatin-induced increases in intracellular reactive oxygen species (ROS), intracellular reactive nitrogen species (RNS) and lipid peroxidation levels. Ebselen dose-dependently increased the expression level of an antioxidant response element (ARE)-luciferase reporter in HEI-OC1 cells through the translocation of Nrf2 into the nucleus. Furthermore, we found that pretreatment with ebselen significantly restored Nrf2 function, whereas it ameliorated the cytotoxicity of cisplatin in cells transfectants with either a pcDNA3.1 (control) or a DN-Nrf2 (dominant-negative) plasmid. We also observed that Nrf2 activation by ebselen increased the expression of phase II antioxidant genes, including heme oxygenase (HO-1), NAD(P)H:quinine oxidoreductase, and gamma-glutamylcysteine synthetase (gamma-GCS). Treatment with ebselen resulted in an increased expression of HO-1 and intranuclear Nrf2 in hair cells of organotypic cultured cochlea. After intraperitoneal injection with cisplatin, auditory brainstem responses (ABRs) threshold was measured on 8th day in Balb/C mice. ABR threshold shift was marked occurred in mice injected with cisplatin (16 mg/kg, n=5; Click and 8-kHz stimuli, p<0.05; 4, 16 and 32 kHz, p<0.01), whereas that of animal group which was treated with cisplatin and ebselen was not significantly changed. These results suggest that ebselen activates the Nrf2-ARE signaling pathway, which ultimately prevents free radical stresses from cisplatin and further contributes to protect auditory sensory hair cells from free radicals produced by cisplatin.

Maresin 1 Ameliorates Lung Ischemia/Reperfusion Injury by Suppressing Oxidative Stress via Activation of the Nrf-2-Mediated HO-1 Signaling Pathway

PubMed Central

Wu, You; Zhao, Feng

2017-01-01

Lung ischemia/reperfusion (I/R) injury occurs in various clinical conditions and heavily damaged lung function. Oxidative stress reaction and antioxidant enzymes play a pivotal role in the etiopathogenesis of lung I/R injury. In the current study, we investigated the impact of Maresin 1 on lung I/R injury and explored the possible mechanism involved in this process. MaR 1 ameliorated I/R-induced lung injury score, wet/dry weight ratio, myeloperoxidase, tumor necrosis factor, bronchoalveolar lavage fluid (BALF) leukocyte count, BALF neutrophil ratio, and pulmonary permeability index levels in lung tissue. MaR 1 significantly reduced ROS, methane dicarboxylic aldehyde, and 15-F2t-isoprostane generation and restored antioxidative enzyme (superoxide dismutase, glutathione peroxidase, and catalase) activities. Administration of MaR 1 improved the expression of nuclear Nrf-2 and cytosolic HO-1 in I/R-treated lung tissue. Furthermore, we also found that the protective effects of MaR 1 on lung tissue injury and oxidative stress were reversed by HO-1 activity inhibitor, Znpp-IX. Nrf-2 transcription factor inhibitor, brusatol, significantly decreased MaR 1-induced nuclear Nrf-2 and cytosolic HO-1 expression. In conclusion, these results indicate that MaR 1 protects against lung I/R injury through suppressing oxidative stress. The mechanism is partially explained by activation of the Nrf-2-mediated HO-1 signaling pathway. PMID:28751936

Negative transcriptional regulation of mitochondrial transcription factor A (TFAM) by nuclear TFAM

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Eun Jin; Kang, Young Cheol; Park, Wook-Ha

2014-07-18

Highlights: • TFAM localizes in nuclei and mitochondria of neuronal cells. • Nuclear TFAM does not bind the Tfam promoter. • Nuclear TFAM reduced the Tfam promoter activity via suppressing NRF-1 activity. • A novel self-negative feedback regulation of Tfam gene expression is explored. • FAM may play different roles depending on its subcellular localizations. - Abstract: The nuclear DNA-encoded mitochondrial transcription factor A (TFAM) is synthesized in cytoplasm and transported into mitochondria. TFAM enhances both transcription and replication of mitochondrial DNA. It is unclear, however, whether TFAM plays a role in regulating nuclear gene expression. Here, we demonstrated thatmore » TFAM was localized to the nucleus and mitochondria by immunostaining, subcellular fractionation, and TFAM-green fluorescent protein hybrid protein studies. In HT22 hippocampal neuronal cells, human TFAM (hTFAM) overexpression suppressed human Tfam promoter-mediated luciferase activity in a dose-dependent manner. The mitochondria targeting sequence-deficient hTFAM also repressed Tfam promoter activity to the same degree as hTFAM. It indicated that nuclear hTFAM suppressed Tfam expression without modulating mitochondrial activity. The repression required for nuclear respiratory factor-1 (NRF-1), but hTFAM did not bind to the NRF-1 binding site of its promoter. TFAM was co-immunoprecipitated with NRF-1. Taken together, we suggest that nuclear TFAM down-regulate its own gene expression as a NRF-1 repressor, showing that TFAM may play different roles depending on its subcellular localizations.« less

Role of Nrf2/HO-1 system in development, oxidative stress response and diseases: an evolutionarily conserved mechanism.

PubMed

Loboda, Agnieszka; Damulewicz, Milena; Pyza, Elzbieta; Jozkowicz, Alicja; Dulak, Jozef

2016-09-01

The multifunctional regulator nuclear factor erythroid 2-related factor (Nrf2) is considered not only as a cytoprotective factor regulating the expression of genes coding for anti-oxidant, anti-inflammatory and detoxifying proteins, but it is also a powerful modulator of species longevity. The vertebrate Nrf2 belongs to Cap 'n' Collar (Cnc) bZIP family of transcription factors and shares a high homology with SKN-1 from Caenorhabditis elegans or CncC found in Drosophila melanogaster. The major characteristics of Nrf2 are to some extent mimicked by Nrf2-dependent genes and their proteins including heme oxygenase-1 (HO-1), which besides removing toxic heme, produces biliverdin, iron ions and carbon monoxide. HO-1 and their products exert beneficial effects through the protection against oxidative injury, regulation of apoptosis, modulation of inflammation as well as contribution to angiogenesis. On the other hand, the disturbances in the proper HO-1 level are associated with the pathogenesis of some age-dependent disorders, including neurodegeneration, cancer or macular degeneration. This review summarizes our knowledge about Nrf2 and HO-1 across different phyla suggesting their conservative role as stress-protective and anti-aging factors.

Targeting Nrf2 Signaling Improves Bacterial Clearance by Alveolar Macrophages in Patients with COPD and in a Mouse Model

PubMed Central

Harvey, Christopher J.; Thimmulappa, Rajesh K.; Sethi, Sanjay; Kong, Xiaoni; Yarmus, Lonny; Brown, Robert H.; David, Feller-Kopman; Wise, Robert; Biswal, Shyam

2016-01-01

Patients with chronic obstructive pulmonary disease (COPD) have innate immune dysfunction in the lung largely due to defective macrophage phagocytosis. This deficiency results in periodic bacterial infections that cause acute exacerbations of COPD, a major source of morbidity and mortality. Recent studies indicate that a decrease in Nrf2 (nuclear erythroid–related factor 2) signaling in patients with COPD may hamper their ability to defend against oxidative stress, although the role of Nrf2 in COPD exacerbations has not been determined. Here, we test whether activation of Nrf2 by the phytochemical sulforaphane restores phagocytosis of clinical isolates of nontypeable Haemophilus influenza (NTHI) and Pseudomonas aeruginosa (PA) by alveolar macrophages from patients with COPD. Sulforaphane treatment restored bacteria recognition and phagocytosis in alveolar macrophages from COPD patients. Furthermore, sulforaphane treatment enhanced pulmonary bacterial clearance by alveolar macrophages and reduced inflammation in wild-typemice but not in Nrf2-deficientmice exposed to cigarette smoke for 6 months. Gene expression and promoter analysis revealed that Nrf2 increased phagocytic ability of macrophages by direct transcriptional up-regulation of the scavenger receptor MARCO. Disruption of Nrf2 or MARCO abrogated sulforaphane-mediated bacterial phagocytosis by COPD alveolar macrophages. Our findings demonstrate the importance of Nrf2 and its downstream target MARCO in improving antibacterial defenses and provide a rationale for targeting this pathway, via pharmacological agents such as sulforaphane, to prevent exacerbations of COPD caused by bacterial infection. PMID:21490276

Interplay between the chalcone cardamonin and selenium in the biosynthesis of Nrf2-regulated antioxidant enzymes in intestinal Caco-2 cells.

PubMed

De Spirt, Silke; Eckers, Anna; Wehrend, Carina; Micoogullari, Mustafa; Sies, Helmut; Stahl, Wilhelm; Steinbrenner, Holger

2016-02-01

Selenoenzymes and nuclear factor erythroid 2-related factor 2 (Nrf2)-regulated phase II enzymes comprise key components of the cellular redox and antioxidant systems, which show multiple interrelations. Deficiency of the micronutrient selenium (Se) and impaired biosynthesis of selenoproteins have been reported to result in induction of Nrf2 target genes. Conversely, transcription of the selenoenzymes glutathione peroxidase 2 (GPx2) and thioredoxin reductase 1 (TrxR1) is up-regulated upon Nrf2 activation. Here, we have studied the interplay between Se and the secondary plant metabolite cardamonin, an Nrf2-activating chalcone, in the regulation of Nrf2-controlled antioxidant enzymes. Se-deficient and Se-repleted (sodium selenite-supplemented) human intestinal Caco-2 cells were exposed to cardamonin. Uptake of cardamonin by the Caco-2 cells was independent of their Se status. Cardamonin strongly induced gene expression of GPx2 and TrxR1. However, cardamonin treatment did not result in elevated GPx or TrxR activity and protein levels, possibly relating to a concomitant down-regulation of O-phosphoseryl-tRNA(Sec) kinase (PSTK), an enzyme involved in translation of selenoprotein mRNAs. On the other hand, induction of the Nrf2-regulated enzyme heme oxygenase 1 (HO-1) by cardamonin was diminished in Se-replete compared to Se-deficient cells. Our findings suggest that cardamonin interferes with the biosynthesis of Nrf2-regulated selenoenzymes, in contrast to the Nrf2-activating isothiocyanate compound sulforaphane, which has been shown earlier to synergize with Se-mediated cytoprotection. Conversely, the cellular Se status apparently affects the cardamonin-mediated induction of non-selenoprotein antioxidant enzymes such as HO-1. Copyright © 2015 Elsevier Inc. All rights reserved.

Ameliorating mitochondrial dysfunction restores carbon ion-induced cognitive deficits via co-activation of NRF2 and PINK1 signaling pathway.

PubMed

Liu, Yang; Yan, Jiawei; Sun, Cao; Li, Guo; Li, Sirui; Zhang, Luwei; Di, Cuixia; Gan, Lu; Wang, Yupei; Zhou, Rong; Si, Jing; Zhang, Hong

2018-07-01

Carbon ion therapy is a promising modality in radiotherapy to treat tumors, however, a potential risk of induction of late normal tissue damage should still be investigated and protected. The aim of the present study was to explore the long-term cognitive deficits provoked by a high-linear energy transfer (high-LET) carbon ions in mice by targeting to hippocampus which plays a crucial role in memory and learning. Our data showed that, one month after 4 Gy carbon ion exposure, carbon ion irradiation conspicuously resulted in the impaired cognitive performance, neurodegeneration and neuronal cell death, as well as the reduced mitochondrial integrity, the disrupted activities of tricarboxylic acid cycle flux and electron transport chain, and the depressed antioxidant defense system, consequently leading to a decline of ATP production and persistent oxidative damage in the hippocampus region. Mechanistically, we demonstrated the disruptions of mitochondrial homeostasis and redox balance typically characterized by the disordered mitochondrial dynamics, mitophagy and glutathione redox couple, which is closely associated with the inhibitions of PINK1 and NRF2 signaling pathway as the key regulators of molecular responses in the context of neurotoxicity and neurodegenerative disorders. Most importantly, we found that administration with melatonin as a mitochondria-targeted antioxidant promoted the PINK1 accumulation on the mitochondrial membrane, and augmented the NRF2 accumulation and translocation. Moreover, melatonin pronouncedly enhanced the molecular interplay between NRF2 and PINK1. Furthermore, in the mouse hippocampal neuronal cells, overexpression of NRF2/PINK1 strikingly protected the hippocampal neurons from carbon ion-elicited toxic insults. Thus, these data suggest that alleviation of the sustained mitochondrial dysfunction and oxidative stress through co-modulation of NRF2 and PINK1 may be in charge of restoration of the cognitive impairments in a mouse model of high-LET carbon ion irradiation. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Attenuation of Aβ{sub 25–35}-induced parallel autophagic and apoptotic cell death by gypenoside XVII through the estrogen receptor-dependent activation of Nrf2/ARE pathways

DOE Office of Scientific and Technical Information (OSTI.GOV)

Meng, Xiangbao; Wang, Min; Sun, Guibo, E-mail: sunguibo@126.com

Amyloid-beta (Aβ) has a pivotal function in the pathogenesis of Alzheimer's disease. To investigate Aβ neurotoxicity, we used an in vitro model that involves Aβ{sub 25–35}-induced cell death in the nerve growth factor-induced differentiation of PC12 cells. Aβ{sub 25–35} (20 μM) treatment for 24 h caused apoptotic cell death, as evidenced by significant cell viability reduction, LDH release, phosphatidylserine externalization, mitochondrial membrane potential disruption, cytochrome c release, caspase-3 activation, PARP cleavage, and DNA fragmentation in PC12 cells. Aβ{sub 25–35} treatment led to autophagic cell death, as evidenced by augmented GFP-LC3 puncta, conversion of LC3-I to LC3-II, and increased LC3-II/LC3-I ratio.more » Aβ{sub 25–35} treatment induced oxidative stress, as evidenced by intracellular ROS accumulation and increased production of mitochondrial superoxide, malondialdehyde, protein carbonyl, and 8-OHdG. Phytoestrogens have been proved to be protective against Aβ-induced neurotoxicity and regarded as relatively safe targets for AD drug development. Gypenoside XVII (GP-17) is a novel phytoestrogen isolated from Gynostemma pentaphyllum or Panax notoginseng. Pretreatment with GP-17 (10 μM) for 12 h increased estrogen response element reporter activity, activated PI3K/Akt pathways, inhibited GSK-3β, induced Nrf2 nuclear translocation, augmented antioxidant responsive element enhancer activity, upregulated heme oxygenase 1 (HO-1) expression and activity, and provided protective effects against Aβ{sub 25–35}-induced neurotoxicity, including oxidative stress, apoptosis, and autophagic cell death. In conclusion, GP-17 conferred protection against Aβ{sub 25–35}-induced neurotoxicity through estrogen receptor-dependent activation of PI3K/Akt pathways, inactivation of GSK-3β and activation of Nrf2/ARE/HO-1 pathways. This finding might provide novel insights into understanding the mechanism for neuroprotective effects of phytoestrogens or gypenosides. - Highlights: • GP-17 showed protection against Aβ{sub 25–35}-induced neurotoxicity. • The neuroprotective effects of GP-17 are dependent on estrogen receptors. • GP-17 activates Nrf2/ARE/HO-1 pathways. • GP-17 activates PI3K/Akt/GSK-3β pathways.« less

Knockdown of Nrf2 Inhibits the Angiogenesis of Rat Cardiac Micro-vascular Endothelial Cells under Hypoxic Conditions

PubMed Central

Kuang, Lihong; Feng, Jian; He, Guoxiang; Jing, Tao

2013-01-01

Angiogenesis plays an important role in myocardial repair after myocardial infarction (MI). Cardiac micro-vascular endothelial cells (CMECs) are important participants in myocardial angiogenesis processes. Recent studies have revealed that Nuclear factor-erythroid 2-related factor 2 (Nrf2), a master transcription factor of endogenous anti-oxidative defense systems, exerts cardio-protection in the cardiovascular system. However, the role of Nrf2 in the process of myocardial angiogenesis and corresponding mechanisms are not fully understood. Thus, the present study investigated the role of Nrf2 in the angiogenesis of rat CMECs to hypoxia. Trans-well assay, three-dimensional Matrigel assay were used to determine cell migration and vascular tube formation. Real-time RT-PCR, ELISA and Western blot were measured mRNA and protein expression. Here, we report that the mRNA and protein expression of Nrf2 and heme oxygenase-1(HO-1) were temporarily upregulated under hypoxic condition. Furthermore, knock down of Nrf2 significantly suppressed the migration and vascular tube formation of rat CMECs to hypoxia, Nrf2 knockdown also significantly decreased HO-1 and vascular endothelial growth factor (VEGF) expression at 48 h after transfection under hypoxic condition. Finally, transfection of CMECs with the Nrf2 over-expressing lentiviral vector upregulated HO-1 expression with a concomitant increase in cell migration and vascular tube formation induced by hypoxia, and this effect was greatly attenuated in the presence of ZnPP (a HO-1 inhibitor). Taken together, these results suggest that Nrf2 may mediate the angiogenesis of CMECs under hypoxic condition, and HO-1 is involved in regulating the angiogenesis of CMECs through Nrf2. Therefore, Nrf2 is a potent regulator of hypoxia-condition mediated angiogenesis in CMECs, which may provide a therapeutic strategy for myocardial repair after MI. PMID:23904790

ROS enhance angiogenic properties via regulation of NRF2 in tumor endothelial cells

PubMed Central

Towfik, Alam Mohammad; Akiyama, Kosuke; Ohga, Noritaka; Shindoh, Masanobu; Hida, Yasuhiro; Minowa, Kazuyuki; Fujisawa, Toshiaki; Hida, Kyoko

2017-01-01

Reactive oxygen species (ROS) are unstable molecules that activate oxidative stress. Because of the insufficient blood flow in tumors, the tumor microenvironment is often exposed to hypoxic condition and nutrient deprivation, which induces ROS accumulation. We isolated tumor endothelial cells (TECs) and found that they have various abnormalities, although the underlying mechanisms are not fully understood. Here we showed that ROS were accumulated in tumor blood vessels and ROS enhanced TEC migration with upregulation of several angiogenesis related gene expressions. It was also demonstrated that these genes were upregulated by regulation of Nuclear factor erythroid 2-related factor 2 (NRF2). Among these genes, we focused on Biglycan, a small leucine-rich proteoglycan. Inhibition of Toll-like receptors 2 and 4, known BIGLYCAN (BGN) receptors, cancelled the TEC motility stimulated by ROS. ROS inhibited NRF2 expression in TECs but not in NECs, and NRF2 inhibited phosphorylation of SMAD2/3, which activates transcription of BGN. These results indicated that ROS-induced BGN caused the pro-angiogenic phenotype in TECs via NRF2 dysregulation. PMID:28525375

The effect of S100A6 on nuclear translocation of CacyBP/SIP in colon cancer cells

PubMed Central

Yang, Bo; Li, Qianqian; Liu, Aiqin; Zhao, Yingying; Qiu, Changqing; Ge, Jun

2018-01-01

Background Calcyclin Binding Protein/(Siah-1 interacting protein) (CacyBP/SIP) acts as an oncogene in colorectal cancer. The nuclear accumulation of CacyBP/SIP has been linked to the proliferation of cancer cells. It has been reported that intracellular Ca2+ induces the nuclear translocation of CacyBP/SIP. However, the molecular mechanism of CacyBP/SIP nuclear translocation has yet to be elucidated. The purpose of this study was to test whether the Ca2+-dependent binding partner S100 protein is involved in CacyBP/SIP nuclear translocation in colon cancer SW480 cells. Methods The subcellular localization of endogenous CacyBP/SIP was observed following the stimulation of ionomycin or BAPTA/AM by immunofluorescence staining in SW480 cells. S100A6 small interfering RNAs (siRNA) were transfected into SW480 cells. Immunoprecipitation assays detected whether S100 protein is relevant to the nuclear translocation of CacyBP/SIP in response to changes in [Ca2+]i. Results We observed that endogenous CacyBP/SIP is translocated from the cytosol to the nucleus following the elevation of [Ca2+]i by ionomycin in SW480 cells. Co-immunoprecipitation experiments showed that the interaction between S100A6 and CacyBP/SIP was increased simultaneously with elevated Ca2+. Knockdown of S100A6 abolished the Ca2+ effect on the subcellular translocation of CacyBP/SIP. Conclusion Thus, we demonstrated that S100A6 is required for the Ca2+-dependent nuclear translocation of CacyBP/SIP in colon cancer SW480 cells. PMID:29534068

3H-1,2-dithiole-3-thione protects retinal pigment epithelium cells against Ultra-violet radiation via activation of Akt-mTORC1-dependent Nrf2-HO-1 signaling.

PubMed

Li, Ke-Ran; Yang, Su-Qing; Gong, Yi-Qing; Yang, Hong; Li, Xiu-Miao; Zhao, Yu-Xia; Yao, Jin; Jiang, Qin; Cao, Cong

2016-05-06

Excessive UV radiation and reactive oxygen species (ROS) cause retinal pigment epithelium (RPE) cell injuries. Nrf2 regulates transcriptional activation of many anti-oxidant genes. Here, we tested the potential role of 3H-1,2-dithiole-3-thione (D3T) against UV or ROS damages in cultured RPE cells (both primary cells and ARPE-19 line). We showed that D3T significantly inhibited UV-/H2O2-induced RPE cell death and apoptosis. UV-stimulated ROS production was dramatically inhibited by D3T pretreatment. D3T induced Nrf2 phosphorylation in cultured RPE cells, causing Nrf2 disassociation with KEAP1 and its subsequent nuclear accumulation. This led to expression of antioxidant response elements (ARE)-dependent gene heme oxygenase-1 (HO-1). Nrf2-HO-1 activation was required for D3T-mediated cytoprotective effect. Nrf2 shRNA knockdown or S40T dominant negative mutation as well as the HO-1 inhibitor Zinc protoporphyrin (ZnPP) largely inhibited D3T's RPE cytoprotective effects against UV radiation. Yet, exogenous overexpression Nrf2 enhanced D3T's activity in RPE cells. Further studies showed that D3T activated Akt/mTORC1 in cultured RPE cells. Akt-mTORC1 inhibitors, or Akt1 knockdown by shRNA, not only inhibited D3T-induced Nrf2-HO-1 activation, but also abolished the RPE cytoprotective effects. In vivo, D3T intravitreal injection protected from light-induced retinal dysfunctions in mice. Thus, D3T protects RPE cells from UV-induced damages via activation of Akt-mTORC1-Nrf2-HO-1 signaling axis.

Schisandrin B elicits a glutathione antioxidant response and protects against apoptosis via the redox-sensitive ERK/Nrf2 pathway in H9c2 cells.

PubMed

Chiu, Po Yee; Chen, Na; Leong, Po Kuan; Leung, Hoi Yan; Ko, Kam Ming

2011-04-01

This study investigated the signal transduction pathway involved in the cytoprotective action of (-)schisandrin B [(-)Sch B, a stereoisomer of Sch B]. Using H9c2 cells, the authors examined the effects of (-)Sch B on MAPK and Nrf2 activation, as well as the subsequent eliciting of glutathione response and protection against apoptosis. Pharmacological tools, such as cytochrome P-450 (CYP) inhibitor, antioxidant, MAPK inhibitor, and Nrf2 RNAi, were used to delineate the signaling pathway. (-)Sch B caused a time-dependent activation of MAPK in H9c2 cells, with the degree of ERK activation being much larger than that of p38 or JNK. The MAPK activation was followed by an increase in the level of nuclear Nrf2, an indirect measure of Nrf2 activation, and the eliciting of a glutathione antioxidant response. The activation of MAPK and Nrf2 seemed to involve oxidants generated from a CYP-catalyzed reaction with (-)Sch B. Both ERK inhibition by U0126 and Nrf2 suppression by Nrf2 RNAi transfection largely abolished the cytoprotection against hypoxia/reoxygenation-induced apoptosis in (-)Sch B-pretreated cells. (-)Sch B pretreatment potentiated the reoxygenation-induced ERK activation, whereas both p38 and JNK activations were suppressed. Under the condition of ERK inhibition, Sch B treatment did not protect against ischemia/reperfusion injury in an ex vivo rat heart model. The results indicate that (-)Sch B triggers a redox-sensitive ERK/Nrf2 signaling, which then elicits a cellular glutathione antioxidant response and protects against hypoxia/reoxygenation-induced apoptosis in H9c2 cells. The ERK-mediated signaling is also likely involved in the cardioprotection afforded by Sch B in vivo.

Isothiocyanates Reduce Mercury Accumulation via an Nrf2-Dependent Mechanism during Exposure of Mice to Methylmercury

PubMed Central

Toyama, Takashi; Shinkai, Yasuhiro; Yasutake, Akira; Uchida, Koji; Yamamoto, Masayuki

2011-01-01

Background: Methylmercury (MeHg) exhibits neurotoxicity through accumulation in the brain. The transcription factor Nrf2 (nuclear factor erythroid 2-related factor 2) plays an important role in reducing the cellular accumulation of MeHg. Objectives: We investigated the protective effect of isothiocyanates, which are known to activate Nrf2, on the accumulation of mercury after exposure to MeHg in vitro and in vivo. Methods: We used primary mouse hepatocytes in in vitro experiments and mice as an in vivo model. We used Western blotting, luciferase assays, atomic absorption spectrometry assays, and MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assays, and we identified toxicity in mice based on hind-limb flaccidity and mortality. Results: The isothiocyanates 6-methylsulfinylhexyl isothiocyanate (6-HITC) and sulforaphane (SFN) activated Nrf2 and up-regulated downstream proteins associated with MeHg excretion, such as glutamate-cysteine ligase, glutathione S-transferase, and multidrug resistance–associated protein, in primary mouse hepatocytes. Under these conditions, intracellular glutathione levels increased in wild-type but not Nrf2-deficient primary mouse hepatocytes. Pretreatment with 6-HITC and SFN before MeHg exposure suppressed cellular accumulation of mercury and cytotoxicity in wild-type but not Nrf2-deficient primary mouse hepatocytes. In comparison, in vivo administration of MeHg to Nrf2-deficient mice resulted in increased sensitivity to mercury concomitant with an increase in mercury accumulation in the brain and liver. Injection of SFN before administration of MeHg resulted in a decrease in mercury accumulation in the brain and liver of wild-type, but not Nrf2-deficient, mice. Conclusions: Through activation of Nrf2, 6-HITC and SFN can suppress mercury accumulation and intoxication caused by MeHg intake. PMID:21382770

The antioxidant and anti-inflammatory activities of tocopherols are independent of Nrf2 in mice.

PubMed

Li, Guangxun; Lee, Mao-Jung; Liu, Anna Ba; Yang, Zhihong; Lin, Yong; Shih, Weichung Joe; Yang, Chung S

2012-04-01

The present study investigated the antioxidant and anti-inflammatory actions of tocopherols in mice and determined whether the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is involved in these activities. A mixture of tocopherols (γ-TmT) that is rich in γ-tocopherol was used. Nrf2 knockout (Nrf2 -/-) and wild-type mice were maintained on 0.03, 0.1, or 0.3% γ-TmT-enriched diet starting 2 weeks before the administration of dextran sulfate sodium (DSS) in drinking water (for 1 week, to induce colonic inflammation), until the termination of the experiment at 3 days after the DSS treatment. Dietary γ-TmT dose dependently lowered the levels of 8-oxo-deoxyguanosine, nitrotyrosine, inflammation index, and leukocyte infiltration in colon tissues, as well as 8-isoprostane and prostaglandin E2 in the serum, in both Nrf2 (-/-) and wild-type mice. No significant difference on the inhibitory actions of γ-TmT between the Nrf2 (-/-) and the wild-type mice was observed. The γ-TmT treatment significantly increased the serum levels of γ- and δ-tocopherols. Interestingly, the serum levels of tocopherol metabolites, specifically the γ- and δ-forms of carboxymethylbutyl hydroxychroman and carboxyethyl hydroxychroman, in Nrf2 (-/-) mice were significantly higher than those in wild-type mice. These findings suggest that the antioxidant and anti-inflammatory activities of γ-TmT in the colon are mostly due to the direct action of tocopherols in trapping reactive oxygen and nitrogen species, independent of the antioxidant enzymes and anti-inflammatory proteins that are regulated by Nrf2; however, Nrf2 knockout appears to affect the serum levels of tocopherol metabolites. Copyright © 2011. Published by Elsevier Inc.

Marine Natural Product Honaucin A Attenuates Inflammation by Activating the Nrf2-ARE Pathway.

PubMed

Mascuch, Samantha J; Boudreau, Paul D; Carland, Tristan M; Pierce, N Tessa; Olson, Joshua; Hensler, Mary E; Choi, Hyukjae; Campanale, Joseph; Hamdoun, Amro; Nizet, Victor; Gerwick, William H; Gaasterland, Teresa; Gerwick, Lena

2018-03-23

The cyanobacterial marine natural product honaucin A inhibits mammalian innate inflammation in vitro and in vivo. To decipher its mechanism of action, RNA sequencing was used to evaluate differences in gene expression of cultured macrophages following honaucin A treatment. This analysis led to the hypothesis that honaucin A exerts its anti-inflammatory activity through activation of the cytoprotective nuclear erythroid 2-related factor 2 (Nrf2)-antioxidant response element/electrophile response element (ARE/EpRE) signaling pathway. Activation of this pathway by honaucin A in cultured human MCF7 cells was confirmed using an Nrf2 luciferase reporter assay. In vitro alkylation experiments with the natural product and N-acetyl-l-cysteine suggest that honaucin A activates this pathway through covalent interaction with the sulfhydryl residues of the cytosolic repressor protein Keap1. Honaucin A presents a potential therapeutic lead for diseases with an inflammatory component modulated by Nrf2-ARE.

Comparison of Adaptive Neuroprotective Mechanisms of Sulforaphane and its Interconversion Product Erucin in in Vitro and in Vivo Models of Parkinson's Disease.

PubMed

Morroni, Fabiana; Sita, Giulia; Djemil, Alice; D'Amico, Massimo; Pruccoli, Letizia; Cantelli-Forti, Giorgio; Hrelia, Patrizia; Tarozzi, Andrea

2018-01-31

Several studies suggest that an increase of glutathione (GSH) through activation of the transcriptional nuclear factor (erythroid-derived 2)-like 2 (Nrf2) in the dopaminergic neurons may be a promising neuroprotective strategy in Parkinson's disease (PD). Among Nrf2 activators, isothiocyanate sulforaphane (SFN), derived from precursor glucosinolate present in Brassica vegetables, has gained attention as a potential neuroprotective compound. Bioavailability studies also suggest the contribution of SFN metabolites, including erucin (ERN), to the neuroprotective effects of SFN. Therefore, we compared the in vitro neuroprotective effects of SFN and ERN at the same dose level (5 μM) and oxidative treatment with 6-hydroxydopamine (6-OHDA) in SH-SY5Y cells. The pretreatment of SH-SY5Y cells with SFN recorded a higher (p < 0.05) active nuclear Nrf2 protein (12.0 ± 0.4 vs 8.0 ± 0.2 fold increase), mRNA Nrf2 (2.0 ± 0.3 vs 1.4 ± 0.1 fold increase), total GSH (384.0 ± 9.0 vs 256.0 ± 8.0 μM) levels, and resistance to neuronal apoptosis elicited by 6-OHDA compared to ERN. By contrast, the simultaneous treatment of SH-SY5Y cells with either SFN or ERN and 6-OHDA recorded similar neuroprotective effects with both the isothiocyanates (Nrf2 protein 2.2 ± 0.2 vs 2.1 ± 0.1 and mRNA Nrf2 2.1 ± 0.3 vs 1.9 ± 0.2 fold increase; total GSH 384.0 ± 4.8 vs 352.0 ± 6.4 μM). Finally, in vitro finding was confirmed in a 6-OHDA-PD mouse model. The metabolic oxidation of ERN to SFN could account for their similar neuroprotective effects in vivo, raising the possibility of using vegetables containing a precursor of ERN for systemic antioxidant benefits in a similar manner to SFN.

The redox-sensing gene Nrf2 affects intestinal homeostasis, insecticide resistance, and Zika virus susceptibility in the mosquito Aedes aegypti.

PubMed

Bottino-Rojas, Vanessa; Talyuli, Octavio A C; Carrara, Luana; Martins, Ademir J; James, Anthony A; Oliveira, Pedro L; Paiva-Silva, Gabriela O

2018-06-08

Production and degradation of reactive oxygen species (ROS) are extensively regulated to ensure proper cellular responses to various environmental stimuli and stresses. Moreover, physiologically generated ROS function as secondary messengers that can influence tissue homeostasis. The cap'n'collar transcription factor known as nuclear factor erythroid-derived factor 2 (Nrf2) coordinates an evolutionarily conserved transcriptional activation pathway that mediates antioxidant and detoxification responses in many animal species, including insects and mammals. Here, we show that Nrf2-mediated signaling affects embryo survival, midgut homeostasis, and redox biology in Aedes aegypti , a mosquito species vector of dengue, Zika, and other disease-causing viruses. We observed that AeNrf2 silencing increases ROS levels and stimulates intestinal stem cell proliferation. Because ROS production is a major aspect of innate immunity in mosquito gut, we found that a decrease in Nrf2 signaling results in reduced microbiota growth and Zika virus infection. Moreover, we provide evidence that AeNrf2 signaling also controls transcriptional adaptation of A. aegypti to insecticide challenge. Therefore, we conclude that Nrf2-mediated response regulates assorted gene clusters in A. aegypti that determine cellular and midgut redox balance, affecting overall xenobiotic resistance and vectorial adaptation of the mosquito. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Nuclear Resonance Fluorescence for Materials Assay

DOE Office of Scientific and Technical Information (OSTI.GOV)

Quiter, Brian; Ludewigt, Bernhard; Mozin, Vladimir

This paper discusses the use of nuclear resonance fluorescence (NRF) techniques for the isotopic and quantitative assaying of radioactive material. Potential applications include age-dating of an unknown radioactive source, pre- and post-detonation nuclear forensics, and safeguards for nuclear fuel cycles Examples of age-dating a strong radioactive source and assaying a spent fuel pin are discussed. The modeling work has ben performed with the Monte Carlo radiation transport computer code MCNPX, and the capability to simulate NRF has bee added to the code. Discussed are the limitations in MCNPX's photon transport physics for accurately describing photon scattering processes that are importantmore » contributions to the background and impact the applicability of the NRF assay technique.« less

Nrf2 activation ameliorates cytotoxic effects of arsenic trioxide in acute promyelocytic leukemia cells through increased glutathione levels and arsenic efflux from cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nishimoto, Shoichi; Suzuki, Toshihiro; Koike, Shin

Carnosic acid (CA), a phenolic diterpene isolated from Rosmarinus officinalis, has been shown to activate nuclear transcription factor E2-related factor 2 (Nrf2), which plays a central role in cytoprotective responses to oxidative and electrophilic stress. Recently, the Nrf2-Kelch ECH associating protein 1 (Keap1) pathway has been associated with cancer drug resistance attributable to modulation of the expression and activation of antioxidant and detoxification enzymes. However, the exact mechanisms by which Nrf2 activation results in chemoresistance are insufficiently understood to date. This study investigated the mechanisms by which the cytotoxic effects of arsenic trioxide (ATO), an anticancer drug, were decreased inmore » acute promyelocytic leukemia cells treated with CA, a typical activator of Nrf2 used to stimulate the Nrf2/Keap1 system. Our findings suggest that arsenic is non-enzymatically incorporated into NB4 cells and forms complexes that are dependent on intracellular glutathione (GSH) concentrations. In addition, the arsenic complexes are recognized as substrates by multidrug resistance proteins and subsequently excreted from the cells. Therefore, Nrf2-associated activation of the GSH biosynthetic pathway, followed by increased levels of intracellular GSH, are key mechanisms underlying accelerated arsenic efflux and attenuation of the cytotoxic effects of ATO. - Highlights: • Nrf2 activation attenuates the effect of arsenic trioxide to acute promyelocytic leukemia cells. • The sensitivity of arsenic trioxide to NB4 cells was dependent on efflux rate of arsenic. • Activation of the GSH biosynthesis is essential in Nrf2-regulated responses for arsenic efflux.« less

EGFR mediates astragaloside IV-induced Nrf2 activation to protect cortical neurons against in vitro ischemia/reperfusion damages

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gu, Da-min; Lu, Pei-Hua, E-mail: lphty1_1@163.com; Zhang, Ke

In this study, we tested the potential role of astragaloside IV (AS-IV) against oxygen and glucose deprivation/re-oxygenation (OGD/R)-induced damages in murine cortical neurons, and studied the associated signaling mechanisms. AS-IV exerted significant neuroprotective effects against OGD/R by reducing reactive oxygen species (ROS) accumulation, thereby attenuating oxidative stress and neuronal cell death. We found that AS-IV treatment in cortical neurons resulted in NF-E2-related factor 2 (Nrf2) signaling activation, evidenced by Nrf2 Ser-40 phosphorylation, and its nuclear localization, as well as transcription of antioxidant-responsive element (ARE)-regulated genes: heme oxygenase-1 (HO-1), NAD(P)H:quinone oxidoreductase 1 (NQO-1) and sulphiredoxin 1 (SRXN-1). Knockdown of Nrf2 throughmore » lentiviral shRNAs prevented AS-IV-induced ARE genes transcription, and abolished its anti-oxidant and neuroprotective activities. Further, we discovered that AS-IV stimulated heparin-binding-epidermal growth factor (HB-EGF) release to trans-activate epidermal growth factor receptor (EGFR) in cortical neurons. Blockage or silencing EGFR prevented Nrf2 activation by AS-IV, thus inhibiting AS-IV-mediated anti-oxidant and neuroprotective activities against OGD/R. In summary, AS-IV protects cortical neurons against OGD/R damages through activating of EGFR-Nrf2 signaling. - Highlights: • Pre-treatment of astragaloside IV (AS-IV) protects murine cortical neurons from OGD/R. • AS-IV activates Nrf2-ARE signaling in murine cortical neurons. • Nrf2 is required for AS-IV-mediated anti-oxidant and neuroprotective activities. • AS-IV stimulates HB-EGF release to trans-activate EGFR in murine cortical neurons. • EGFR mediates AS-IV-induced Nrf2 activation and neuroprotection against OGD/R.« less

Nrf2 Knockdown Disrupts the Protective Effect of Curcumin on Alcohol-Induced Hepatocyte Necroptosis.

PubMed

Lu, Chunfeng; Xu, Wenxuan; Zhang, Feng; Shao, Jiangjuan; Zheng, Shizhong

2016-12-05

It has emerged that hepatocyte necroptosis plays a critical role in chronic alcoholic liver disease (ALD). Our previous study has identified that the beneficial therapeutic effect of curcumin on alcohol-caused liver injury might be attributed to activation of nuclear factor (erythroid-derived 2)-like 2 (Nrf2), whereas the role of curcumin in regulating necroptosis and the underlying mechanism remain to be determined. We first found that chronic alcohol consumption triggered obvious hepatocyte necroptosis, leading to increased expression of receptor-interacting protein 1, receptor-interacting protein 3, high-mobility group box 1, and phosphorylated mixed lineage kinase domain-like in murine livers. Curcumin dose-dependently ameliorated hepatocyte necroptosis and alleviated alcohol-caused decrease in hepatic Nrf2 expression in alcoholic mice. Then Nrf2 shRNA lentivirus was introduced to generate Nrf2-knockdown mice. Our results indicated that Nrf2 knockdown aggravated the effects of alcohol on liver injury and necroptosis and even abrogated the inhibitory effect of curcumin on necroptosis. Further, activated Nrf2 by curcumin inhibited p53 expression in both livers and cultured hepatocytes under alcohol stimulation. The next in vitro experiments, similar to in vivo ones, revealed that although Nrf2 knockdown abolished the suppression of curcumin on necroptosis of hepatocytes exposed to ethanol, p53 siRNA could clearly rescued the relative effect of curcumin. In summary, for the first time, we concluded that curcumin attenuated alcohol-induced hepatocyte necroptosis in a Nrf2/p53-dependent mechanism. These findings make curcumin an excellent candidate for ALD treatment and advance the understanding of ALD mechanisms associated with hepatocyte necroptosis.

Synthesis, evaluation, and metabolism of novel [6]-shogaol derivatives as potent Nrf2 activators.

PubMed

Zhu, Yingdong; Wang, Pei; Zhao, Yantao; Yang, Chun; Clark, Anderson; Leung, TinChung; Chen, Xiaoxin; Sang, Shengmin

2016-06-01

Oxidative stress is a central component of many chronic diseases. The Kelch-like ECH-associated protein 1 (Keap1)-nuclear factor erythroid 2 p45-related factor 2 (Nrf2) system is a major regulatory pathway of cytoprotective genes against oxidative and electrophilic stress. Activation of the Nrf2 pathway plays crucial roles in the chemopreventive effects of various inducers. In this study, we developed a novel class of potent Nrf2 activators derived from ginger compound, [6]-shogaol (6S), using the Tg[glutathione S-transferase pi 1 (gstp1):green fluorescent protein (GFP)] transgenic zebrafish model. Investigation of structure-activity relationships of 6S derivatives indicates that the combination of an α,β-unsaturated carbonyl entity and a catechol moiety in one compound enhances the Tg(gstp1:GFP) fluorescence signal in zebrafish embryos. Chemical reaction and in vivo metabolism studies of the four most potent 6S derivatives showed that both α,β-unsaturated carbonyl entity and catechol moiety act as major active groups for conjugation with the sulfhydryl groups of the cysteine residues. In addition, we further demonstrated that 6S derivatives increased the expression of Nrf2 downstream target, heme oxygenase-1, in both a dose- and time-dependent manner. These results suggest that α,β-unsaturated carbonyl entity and catechol moiety of 6S derivatives may react with the cysteine residues of Keap1, disrupting the Keap1-Nrf2 complex, thereby liberating and activating Nrf2. Our findings of natural product-derived Nrf2 activators lead to design options of potent Nrf2 activators for further optimization. Copyright © 2016 Elsevier Inc. All rights reserved.

Chronic Endurance Exercise Impairs Cardiac Structure and Function in Middle-Aged Mice with Impaired Nrf2 Signaling

PubMed Central

Shanmugam, Gobinath; Narasimhan, Madhusudhanan; Conley, Robbie L.; Sairam, Thiagarajan; Kumar, Ashutosh; Mason, Ronald P.; Sankaran, Ramalingam; Hoidal, John R.; Rajasekaran, Namakkal S.

2017-01-01

Nuclear factor erythroid 2 related factor 2 (Nrf2) signaling maintains the redox homeostasis and its activation is shown to suppress cardiac maladaptation. Earlier we reported that acute endurance exercise (2 days) evoked antioxidant cytoprotection in young WT animals but not in aged WT animals. However, the effect of repeated endurance exercise during biologic aging (WT) characterized by an inherent deterioration in Nrf2 signaling and pathological aging (pronounced oxidative susceptibility—Nrf2 absence) in the myocardium remains elusive. Thus, the purpose of our study was to determine the effect of chronic endurance exercise-induced cardiac adaptation in aged mice with and without Nrf2. Age-matched WT and Nrf2-null mice (Nrf2−/−) (>22 months) were subjected to 6 weeks chronic endurance exercise (25 meter/min, 12% grade). The myocardial redox status was assessed by expression of antioxidant defense genes and proteins along with immunochemical detection of DMPO-radical adduct, GSH-NEM, and total ubiquitination. Cardiac functions were assessed by echocardiography and electrocardiogram. At sedentary state, loss of Nrf2 resulted in significant downregulation of antioxidant gene expression (Nqo1, Ho1, Gclm, Cat, and Gst-α) with decreased GSH-NEM immuno-fluorescence signals. While Nrf2−/− mice subjected to CEE showed an either similar or more pronounced reduction in the transcript levels of Gclc, Nqo1, Gsr, and Gst-α in relation to WT littermates. In addition, the hearts of Nrf2−/− on CEE showed a substantial reduction in specific antioxidant proteins, G6PD and CAT along with decreased GSH, a pronounced increase in DMPO-adduct and the total ubiquitination levels. Further, CEE resulted in a significant upregulation of hypertrophy genes (Anf, Bnf, and β-Mhc) (p < 0.05) in the Nrf2−/− hearts in relation to WT mice. Moreover, the aged Nrf2−/− mice exhibited a higher degree of cardiac remodeling in association with a significant decrease in fractional shortening, pronounced ST segment, and J wave elevation upon CEE compared to age-matched WT littermates. In conclusion, our findings indicate that while the aged WT and Nrf2 knockout animals both exhibit hypertrophy after CEE, the older Nrf2 knockouts showed ventricular remodeling coupled with profound cardiac functional abnormalities and diastolic dysfunction. PMID:28515695

Lithium Promotes Longevity through GSK3/NRF2-Dependent Hormesis

PubMed Central

Castillo-Quan, Jorge Iván; Li, Li; Kinghorn, Kerri J.; Ivanov, Dobril K.; Tain, Luke S.; Slack, Cathy; Kerr, Fiona; Nespital, Tobias; Thornton, Janet; Hardy, John; Bjedov, Ivana; Partridge, Linda

2016-01-01

Summary The quest to extend healthspan via pharmacological means is becoming increasingly urgent, both from a health and economic perspective. Here we show that lithium, a drug approved for human use, promotes longevity and healthspan. We demonstrate that lithium extends lifespan in female and male Drosophila, when administered throughout adulthood or only later in life. The life-extending mechanism involves the inhibition of glycogen synthase kinase-3 (GSK-3) and activation of the transcription factor nuclear factor erythroid 2-related factor (NRF-2). Combining genetic loss of the NRF-2 repressor Kelch-like ECH-associated protein 1 (Keap1) with lithium treatment revealed that high levels of NRF-2 activation conferred stress resistance, while low levels additionally promoted longevity. The discovery of GSK-3 as a therapeutic target for aging will likely lead to more effective treatments that can modulate mammalian aging and further improve health in later life. PMID:27068460

Suppression of NRF2–ARE activity sensitizes chemotherapeutic agent-induced cytotoxicity in human acute monocytic leukemia cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Peng, Hui; Institute of Disease Control and Prevention, Academy of Military Medical Sciences, Beijing; Wang, Huihui

2016-02-01

Nuclear factor erythroid 2-related factor 2 (NRF2), a master regulator of the antioxidant response element (ARE)-dependent transcription, plays a pivotal role in chemical detoxification in normal and tumor cells. Consistent with previous findings that NRF2–ARE contributes to chemotherapeutic resistance of cancer cells, we found that stable knockdown of NRF2 by lentiviral shRNA in human acute monocytic leukemia (AML) THP-1 cells enhanced the cytotoxicity of several chemotherapeutic agents, including arsenic trioxide (As{sub 2}O{sub 3}), etoposide and doxorubicin. Using an ARE-luciferase reporter expressed in several human and mouse cells, we identified a set of compounds, including isonicotinic acid amides, isoniazid and ethionamide,more » that inhibited NRF2–ARE activity. Treatment of THP-1 cells with ethionamide, for instance, significantly reduced mRNA expression of multiple ARE-driven genes under either basal or As{sub 2}O{sub 3}-challenged conditions. As determined by cell viability and cell cycle, suppression of NRF2–ARE by ethionamide also significantly enhanced susceptibility of THP-1 and U937 cells to As{sub 2}O{sub 3}-induced cytotoxicity. In THP-1 cells, the sensitizing effect of ethionamide on As{sub 2}O{sub 3}-induced cytotoxicity was highly dependent on NRF2. To our knowledge, the present study is the first to demonstrate that ethionamide suppresses NRF2–ARE signaling and disrupts the transcriptional network of the antioxidant response in AML cells, leading to sensitization to chemotherapeutic agents. - Highlights: • Identification of novel inhibitors of ARE-dependent transcription • Suppression of NRF2–ARE sensitizes THP-1 cells to chemotherapy. • Ethionamide suppresses ARE-dependent transcriptional activity. • Ethionamide and isoniazid increase the cytotoxicity of As{sub 2}O{sub 3} in AML cells. • Sensitization of THP-1 cells to As{sub 2}O{sub 3} toxicity by ethionamide is NRF2-dependent.« less

Carvedilol, a third-generation β-blocker prevents oxidative stress-induced neuronal death and activates Nrf2/ARE pathway in HT22 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ouyang, Ying; Chen, Ziwei; Tan, Min

Highlights: •Carvedilol significantly prevented oxidative stress-induced cell death. •Carvedilol significantly decreased the production of ROS. •Carvedilol activated Nrf2/ARE pathway. •Carvedilol increased the protein levels of HO-1 and NQO-1. -- Abstract: Carvedilol, a nonselective β-adrenoreceptor blocker with pleiotropic activities has been shown to exert neuroprotective effect due to its antioxidant property. However, the neuroprotective mechanism of carvedilol is still not fully uncovered. Nuclear factor E2-related factor 2 (Nrf2)/antioxidant response element (ARE) pathway is an important cellular stress response pathway involved in neuroprotection. Here we investigated the effect of carvedilol on oxidative stress-induced cell death (glutamate 2 mM and H{sub 2}O{sub 2}more » 600 μM) and the activity of Nrf2/ARE pathway in HT22 hippocampal cells. Carvedilol significantly increased cell viability and decreased ROS in HT22 cells exposed to glutamate or H{sub 2}O{sub 2}. Furthermore, carvedilol activated the Nrf2/ARE pathway in a concentration-dependent manner, and increased the protein levels of heme oxygenase-1(HO-1) and NAD(P)H quinone oxidoreductase-1(NQO-1), two downstream factors of the Nrf2/ARE pathway. Collectively, our results indicate that carvedilol protects neuronal cell against glutamate- and H{sub 2}O{sub 2}-induced neurotoxicity possibly through activating the Nrf2/ARE signaling pathway.« less

Methylglyoxal, the foe and friend of glyoxalase and Trx/TrxR systems in HT22 nerve cells.

PubMed

Dafre, A L; Goldberg, J; Wang, T; Spiegel, D A; Maher, P

2015-12-01

Methylglyoxal (MGO) is a major glycating agent that reacts with basic residues of proteins and promotes the formation of advanced glycation end products (AGEs) which are believed to play key roles in a number of pathologies, such as diabetes, Alzheimer's disease, and inflammation. Here, we examined the effects of MGO on immortalized mouse hippocampal HT22 nerve cells. The endpoints analyzed were MGO and thiol status, the glyoxalase system, comprising glyoxalase 1 and 2 (GLO1/2), and the cytosolic and mitochondrial Trx/TrxR systems, as well as nuclear Nrf2 and its target genes. We found that nuclear Nrf2 is induced by MGO treatment in HT22 cells, as corroborated by induction of the Nrf2-controlled target genes and proteins glutamate cysteine ligase and heme oxygenase 1. Nrf2 knockdown prevented MGO-dependent induction of glutamate cysteine ligase and heme oxygenase 1. The cystine/glutamate antiporter, system xc(-), which is also controlled by Nrf2, was also induced. The increased cystine import (system xc(-)) activity and GCL expression promoted GSH synthesis, leading to increased levels of GSH. The data indicate that MGO can act as both a foe and a friend of the glyoxalase and the Trx/TrxR systems. At low concentrations of MGO (0.3mM), GLO2 is strongly induced, but at high MGO (0.75 mM) concentrations, GLO1 is inhibited and GLO2 is downregulated. The cytosolic Trx/TrxR system is impaired by MGO, where Trx is downregulated yet TrxR is induced, but strong MGO-dependent glycation may explain the loss in TrxR activity. We propose that Nrf2 can be the unifying element to explain the observed upregulation of GSH, GCL, HO1, TrxR1, Trx2, TrxR2, and system xc(-) system activity. Copyright © 2015. Published by Elsevier Inc.

Methylglyoxal, the foe and friend of glyoxalase and Trx/TrxR systems in HT22 nerve cells

PubMed Central

Dafre, A.L.; Goldberg, J.; Wang, T.; Spiegel, D.A.; Maher, P.

2017-01-01

Methylglyoxal (MGO) is a major glycating agent that reacts with basic residues of proteins and promotes the formation of advanced glycation end products (AGEs) which are believed to play key roles in a number of pathologies, such as diabetes, Alzheimer’s disease, and inflammation. Here, we examined the effects of MGO on immortalized mouse hippocampal HT22 nerve cells. The endpoints analyzed were MGO and thiol status, the glyoxalase system, comprising glyoxalase 1 and 2 (GLO1/2), and the cytosolic and mitochondrial Trx/TrxR systems, as well as nuclear Nrf2 and its target genes. We found that nuclear Nrf2 is induced by MGO treatment in HT22 cells, as corroborated by induction of the Nrf2-controlled target genes and proteins glutamate cysteine ligase and heme oxygenase 1. Nrf2 knockdown prevented MGO-dependent induction of glutamate cysteine ligase and heme oxygenase 1. The cystine/glutamate antiporter, system xc−, which is also controlled by Nrf2, was also induced. The increased cystine import (system xc−) activity and GCL expression promoted GSH synthesis, leading to increased levels of GSH. The data indicate that MGO can act as both a foe and a friend of the glyoxalase and the Trx/TrxR systems. At low concentrations of MGO (0.3 mM), GLO2 is strongly induced, but at high MGO (0.75 mM) concentrations, GLO1 is inhibited and GLO2 is downregulated. The cytosolic Trx/TrxR system is impaired by MGO, where Trx is downregulated yet TrxR is induced, but strong MGO-dependent glycation may explain the loss in TrxR activity. We propose that Nrf2 can be the unifying element to explain the observed upregulation of GSH, GCL, HO1, TrxR1, Trx2, TrxR2, and system xc− system activity. PMID:26165190

Synthetic Lignan Secoisolariciresinol Diglucoside (LGM2605) Reduces Asbestos-Induced Cytotoxicity in an Nrf2-Dependent and -Independent Manner

PubMed Central

Pietrofesa, Ralph A.; Chatterjee, Shampa; Park, Kyewon; Arguiri, Evguenia; Albelda, Steven M.; Christofidou-Solomidou, Melpo

2018-01-01

Asbestos exposure triggers inflammatory processes associated with oxidative stress and tissue damage linked to malignancy. LGM2605 is the synthetic lignan secoisolariciresinol diglucoside (SDG) with free radical scavenging, antioxidant, and anti-inflammatory properties in diverse inflammatory cell and mouse models, including exposure to asbestos fibers. Nuclear factor-E2 related factor 2 (Nrf2) activation and boosting of endogenous tissue defenses were associated with the protective action of LGM2605 from asbestos-induced cellular damage. To elucidate the role of Nrf2 induction by LGM2605 in protection from asbestos-induced cellular damage, we evaluated LGM2605 in asbestos-exposed macrophages from wild-type (WT) and Nrf2 disrupted (Nrf2−/−) mice. Cells were pretreated with LGM2605 (50 µM and 100 µM) and exposed to asbestos fibers (20 µg/cm2) and evaluated 8 h and 24 h later for inflammasome activation, secreted cytokine levels (interleukin-1β (IL-1β), interleukin-18 (IL-18), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNFα)), cytotoxicity and cell death, nitrosative stress, and Nrf2-regulated enzyme levels. Asbestos exposure induced robust oxidative and nitrosative stress, cell death and cytotoxicity, which were equally mitigated by LGM2605. Inflammasome activation was significantly attenuated in Nrf2−/− macrophages compared to WT, and the protective action of LGM2605 was seen only in WT cells. In conclusion, in a cell model of asbestos-induced toxicity, LGM2605 acts via protective mechanisms that may not involve Nrf2 activation. PMID:29498660

Minocycline attenuates sevoflurane-induced cell injury via activation of Nrf2

PubMed Central

Tian, Yue; Wu, Xiuying; Guo, Shanbin; Ma, Ling; Huang, Wei; Zhao, Xiaochun

2017-01-01

Minocycline has been demonstrated to exert neuroprotective effects in various experimental models. In the present study, we investigated the mechanisms underlying the protective effects of minocycline on cell injury induced by the inhalation of the anesthetic, sevoflurane. In our in vivo experiments using rats, minocycline attenuated sevoflurane-induced neuronal degeneration and apoptosis in the rat hippocampus, and this effect was associated with the minocycline-mediated suppression of oxidative stress in the hippocampus. In in vitro experiments, minocycline inhibited sevoflurane-induced apoptosis and the production of reactive oxygen species (ROS) in H4 human neuroglioma cells. In addition, minocycline suppressed the sevoflurane-induced upregulation of interleukin (IL)-6 and the activation of the nuclear factor-κB (NF-κB) signaling pathway in H4 cells. Furthermore, we found that nuclear factor E2-related factor 2 (Nrf2), an activator of the stress response, was upregulated and activated upon sevoflurane treatment both in the rat hippocampus and in H4 cells. In addition, minocycline further augmented the upregulation and activation of Nrf2 when used in conjunction with sevoflurane. Moreover, the knockdown of Nrf2 in H4 cells by small interfering RNA (siRNA) diminished the cytoprotective effect of minocycline, and attenuated the inhibitory effect of minocycline on ROS production, IL-6 upregulation and the activation of the NF-κB signaling pathway. On the whole, our findings indicate that minocycline may exert protective effects against sevoflurane-induced cell injury via the Nrf2-modulated antioxidant response and the inhibition of the activation of the NF-κB signaling pathway. PMID:28260081

Role of Nrf2 antioxidant defense in mitigating cadmium-induced oxidative stress in the olfactory system of zebrafish

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Lu; Gallagher, Evan P., E-mail: evang3@uw.edu

2013-01-15

Exposure to trace metals can disrupt olfactory function in fish leading to a loss of behaviors critical to survival. Cadmium (Cd) is an olfactory toxicant that elicits cellular oxidative stress as a mechanism of toxicity while also inducing protective cellular antioxidant genes via activation of the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) pathway. However, the molecular mechanisms of Cd-induced olfactory injury have not been characterized. In the present study, we investigated the role of the Nrf2-mediated antioxidant defense pathway in protecting against Cd-induced olfactory injury in zebrafish. A dose-dependent induction of Nrf2-regulated antioxidant genes associated with cellular responses to oxidativemore » stress was observed in the olfactory system of adult zebrafish following 24 h Cd exposure. Zebrafish larvae exposed to Cd for 3 h showed increased glutathione S-transferase pi (gst pi), glutamate–cysteine ligase catalytic subunit (gclc), heme oxygenase 1 (hmox1) and peroxiredoxin 1 (prdx1) mRNA levels indicative of Nrf2 activation, and which were blocked by morpholino-mediated Nrf2 knockdown. The inhibition of antioxidant gene induction in Cd-exposed Nrf2 morphants was associated with disruption of olfactory driven behaviors, increased cell death and loss of olfactory sensory neurons (OSNs). Nrf2 morphants also exhibited a downregulation of OSN-specific genes after Cd exposure. Pre-incubation of embryos with sulforaphane (SFN) partially protected against Cd-induced olfactory tissue damage. Collectively, our results indicate that oxidative stress is an important mechanism of Cd-mediated injury in the zebrafish olfactory system. Moreover, the Nrf2 pathway plays a protective role against cellular oxidative damage and is important in maintaining zebrafish olfactory function. -- Highlights: ► Oxidative stress is an important mechanism of Cd-mediated olfactory injury. ► Cd induces antioxidant gene expression in the zebrafish olfactory system. ► The olfactory antioxidant response is blocked by Nrf2 knockdown. ► Disruption of olfactory neurobehaviors is associated with Nrf2 knockdown. ► Nrf2 morphants show increased cell death and olfactory sensory neuron loss.« less

Impaired redox signaling and antioxidant gene expression in endothelial cells in diabetes: a role for mitochondria and the nuclear factor-E2-related factor 2-Kelch-like ECH-associated protein 1 defense pathway.

PubMed

Cheng, Xinghua; Siow, Richard C M; Mann, Giovanni E

2011-02-01

Type 2 diabetes is an age-related disease associated with vascular pathologies, including severe blindness, renal failure, atherosclerosis, and stroke. Reactive oxygen species (ROS), especially mitochondrial ROS, play a key role in regulating the cellular redox status, and an overproduction of ROS may in part underlie the pathogenesis of diabetes and other age-related diseases. Cells have evolved endogenous defense mechanisms against sustained oxidative stress such as the redox-sensitive transcription factor nuclear factor E2-related factor 2 (Nrf2), which regulates antioxidant response element (ARE/electrophile response element)-mediated expression of detoxifying and antioxidant enzymes and the cystine/glutamate transporter involved in glutathione biosynthesis. We hypothesize that diminished Nrf2/ARE activity contributes to increased oxidative stress and mitochondrial dysfunction in the vasculature leading to endothelial dysfunction, insulin resistance, and abnormal angiogenesis observed in diabetes. Sustained hyperglycemia further exacerbates redox dysregulation, thereby providing a positive feedback loop for severe diabetic complications. This review focuses on the role that Nrf2/ARE-linked gene expression plays in regulating endothelial redox homeostasis in health and type 2 diabetes, highlighting recent evidence that Nrf2 may provide a therapeutic target for countering oxidative stress associated with vascular disease and aging.

Anti-fibrotic effects of L-2-oxothiazolidine-4-carboxylic acid via modulation of nuclear factor erythroid 2-related factor 2 in rats.

PubMed

Kim, In Hee; Kim, Dae-Ghon; Hao, Peipei; Wang, Yunpeng; Kim, Seong Hun; Kim, Sang Wook; Lee, Seung Ok; Lee, Soo Teik

2012-06-01

L-2-Oxothiazolidine-4-carboxylic acid (OTC) is a cysteine prodrug that maintains glutathione in tissues. The present study was designed to investigate anti-fibrotic and anti-oxidative effects of OTC via modulation of nuclear factor erythroid 2-related factor 2 (Nrf2) in an in vivo thioacetamide (TAA)-induced hepatic fibrosis model. Treatment with OTC (80 or 160 mg/kg) improved serum liver function parameters and significantly ameliorated liver fibrosis. The OTC treatment groups exhibited significantly lower expression of α-smooth muscle actin, transforming growth factor-β 1, and collagen α 1 mRNA than that in the TAA model group. Furthermore, the OTC treatment groups showed a significant decrease in hepatic malondialdehyde level compared to that in the TAA model group. Nrf2 and heme oxygenase-1 expression increased significantly in the OTC treatment groups compared with that in the TAA model group. Taken together, these results suggest that OTC restores the anti- oxidative system by upregulating Nrf2; thus, ameliorating liver injury and a fibrotic reaction.

Neuroprotection against 6-OHDA toxicity in PC12 cells and mice through the Nrf2 pathway by a sesquiterpenoid from Tussilago farfara.

PubMed

Lee, Joohee; Song, Kwangho; Huh, Eugene; Oh, Myung Sook; Kim, Yeong Shik

2018-06-01

Oxidative stress plays a key role in neurodegenerative diseases such as Alzheimer's and Parkinson's diseases. Therefore, the nuclear factor-E2-related factor 2 (Nrf2), a key regulator of the antioxidative response, is considered to be important as a therapeutic target for neurodegenerative diseases. We investigated the underlying mechanism of Nrf2-mediated neuroprotective effects against oxidative stress in the PC12 cell line by 7β-(3-ethyl-cis-crotonoyloxy)-1α-(2-methylbutyryloxy)-3,14-dehydro-Z-notonipetranone (ECN), one of the sesquiterpenoids in Farfarae Flos. Pretreatment of PC12 cells with ECN had a protective effect against hydrogen peroxide (H 2 O 2 )- or 6-hydroxydopamine (6-OHDA)-induced cytotoxicity. ECN upregulated the ARE-luciferase activity and induced the mRNA expression of Nrf2 and antioxidant enzyme heme oxygenase-1 (HO-1). Knockdown of Nrf2 by small, interfering RNA (siRNA) abrogated the upregulation of HO-1, indicating that ECN had induced HO-1 via the Nrf2 pathway. Pretreatment with the thiol reducing agents, N-acetylcysteine (NAC) or dithiothreitol (DTT), attenuated Nrf2 activation and HO-1 expression. However, the non-thiol reducing antioxidant, Trolox, failed to inhibit HO-1 induction by ECN. These results suggest that ECN may directly interact with Kelch-like ECH-associated protein 1 (Keap1) and modify critical cysteine thiols present in the proteins responsible for Nrf2-mediated upregulation of HO-1. In a 6-OHDA-induced mouse model of PD, administration of ECN ameliorated motor impairments and dopaminergic neuronal damage. Taken together, ECN exerts neuroprotective effects by activating the Nrf2/HO-1 signaling pathway in both PC12 cells and mice. Thus, ECN, as an Nrf2 activator, could be an attractive therapeutic candidate for the neuroprotection or treatment of neurodegenerative diseases. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Synthetic Lignan Secoisolariciresinol Diglucoside (LGM2605) Reduces Asbestos-Induced Cytotoxicity in an Nrf2-Dependent and -Independent Manner.

PubMed

Pietrofesa, Ralph A; Chatterjee, Shampa; Park, Kyewon; Arguiri, Evguenia; Albelda, Steven M; Christofidou-Solomidou, Melpo

2018-03-02

Asbestos exposure triggers inflammatory processes associated with oxidative stress and tissue damage linked to malignancy. LGM2605 is the synthetic lignan secoisolariciresinol diglucoside (SDG) with free radical scavenging, antioxidant, and anti-inflammatory properties in diverse inflammatory cell and mouse models, including exposure to asbestos fibers. Nuclear factor-E2 related factor 2 (Nrf2) activation and boosting of endogenous tissue defenses were associated with the protective action of LGM2605 from asbestos-induced cellular damage. To elucidate the role of Nrf2 induction by LGM2605 in protection from asbestos-induced cellular damage, we evaluated LGM2605 in asbestos-exposed macrophages from wild-type (WT) and Nrf2 disrupted (Nrf2 - / - ) mice. Cells were pretreated with LGM2605 (50 µM and 100 µM) and exposed to asbestos fibers (20 µg/cm²) and evaluated 8 h and 24 h later for inflammasome activation, secreted cytokine levels (interleukin-1β (IL-1β), interleukin-18 (IL-18), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNFα)), cytotoxicity and cell death, nitrosative stress, and Nrf2-regulated enzyme levels. Asbestos exposure induced robust oxidative and nitrosative stress, cell death and cytotoxicity, which were equally mitigated by LGM2605. Inflammasome activation was significantly attenuated in Nrf2 -/- macrophages compared to WT, and the protective action of LGM2605 was seen only in WT cells. In conclusion, in a cell model of asbestos-induced toxicity, LGM2605 acts via protective mechanisms that may not involve Nrf2 activation.

Cul3-mediated Nrf2 ubiquitination and antioxidant response element (ARE) activation are dependent on the partial molar volume at position 151 of Keap1.

PubMed

Eggler, Aimee L; Small, Evan; Hannink, Mark; Mesecar, Andrew D

2009-07-29

Nrf2 (nuclear factor erythroid 2-related factor 2) is a transcription factor that activates transcription of a battery of cytoprotective genes by binding to the ARE (antioxidant response element). Nrf2 is repressed by the cysteine-rich Keap1 (kelch-like ECH-associated protein 1) protein, which targets Nrf2 for ubiquitination and subsequent degradation by a Cul3 (cullin 3)-mediated ubiquitination complex. We find that modification of Cys(151) of human Keap1, by mutation to a tryptophan, relieves the repression by Keap1 and allows activation of the ARE by Nrf2. The Keap1 C151W substitution has a decreased affinity for Cul3, and can no longer serve to target Nrf2 for ubiquitination, though it retains its affinity for Nrf2. A series of 12 mutant Keap1 proteins, each containing a different residue at position 151, was constructed to explore the chemistry required for this effect. The series reveals that the extent to which Keap1 loses the ability to target Nrf2 for degradation, and hence the ability to repress ARE activation, correlates well with the partial molar volume of the residue. Other physico-chemical properties do not appear to contribute significantly to the effect. Based on this finding, a structural model is proposed whereby large residues at position 151 cause steric clashes that lead to alteration of the Keap1-Cul3 interaction. This model has significant implications for how electrophiles which modify Cys(151), disrupt the repressive function of Keap1.

Sulforaphane Prevents Angiotensin II-Induced Testicular Cell Death via Activation of NRF2.

PubMed

Wang, Yonggang; Wu, Hao; Xin, Ying; Bai, Yang; Kong, Lili; Tan, Yi; Liu, Feng; Cai, Lu

2017-01-01

Although angiotensin II (Ang II) was reported to facilitate sperm motility and intratesticular sperm transport, recent findings shed light on the efficacy of Ang II in stimulating inflammatory events in testicular peritubular cells, effect of which may play a role in male infertility. It is still unknown whether Ang II can induce testicular apoptotic cell death, which may be a more direct action of Ang II in male infertility. Therefore, the present study aims to determine whether Ang II can induce testicular apoptotic cell death and whether this action can be prevented by sulforaphane (SFN) via activating nuclear factor (erythroid-derived 2)-like 2 (NRF2), the governor of antioxidant-redox signalling. Eight-week-old male C57BL/6J wild type (WT) and Nrf2 gene knockout mice were treated with Ang II, in the presence or absence of SFN. In WT mice, SFN activated testicular NRF2 expression and function, along with a marked attenuation in Ang II-induced testicular oxidative stress, inflammation, endoplasmic reticulum stress, and apoptotic cell death. Deletion of the Nrf2 gene led to a complete abolishment of these efficacies of SFN. The present study indicated that Ang II may result in testicular apoptotic cell death, which can be prevented by SFN via the activation of NRF2.

Nuclear Import of β-Dystroglycan Is Facilitated by Ezrin-Mediated Cytoskeleton Reorganization

PubMed Central

Vásquez-Limeta, Alejandra; Wagstaff, Kylie M.; Ortega, Arturo; Crouch, Dorothy H.; Jans, David A.; Cisneros, Bulmaro

2014-01-01

The β-dystroglycan (β-DG) protein has the ability to target to multiple sites in eukaryotic cells, being a member of diverse protein assemblies including the transmembranal dystrophin-associated complex, and a nuclear envelope-localised complex that contains emerin and lamins A/C and B1. We noted that the importin α2/β1-recognised nuclear localization signal (NLS) of β-DG is also a binding site for the cytoskeletal-interacting protein ezrin, and set out to determine whether ezrin binding might modulate β-DG nuclear translocation for the first time. Unexpectedly, we found that ezrin enhances rather than inhibits β-DG nuclear translocation in C2C12 myoblasts. Both overexpression of a phosphomimetic activated ezrin variant (Ez-T567D) and activation of endogenous ezrin through stimulation of the Rho pathway resulted in both formation of actin-rich surface protrusions and significantly increased nuclear translocation of β-DG as shown by quantitative microscopy and subcellular fractionation/Western analysis. In contrast, overexpression of a nonphosphorylatable inactive ezrin variant (Ez-T567A) or inhibition of Rho signaling, decreased nuclear translocation of β-DG concomitant with a lack of cell surface protrusions. Further, a role for the actin cytoskeleton in ezrin enhancement of β-DG nuclear translocation was implicated by the observation that an ezrin variant lacking its actin-binding domain failed to enhance nuclear translocation of β-DG, while disruption of the actin cytoskeleton led to a reduction in β-DG nuclear localization. Finally, we show that ezrin-mediated cytoskeletal reorganization enhances nuclear translocation of the cytoplasmic but not the transmembranal fraction of β-DG. This is the first study showing that cytoskeleton reorganization can modulate nuclear translocation of β-DG, with the implication that β-DG can respond to cytoskeleton-driven changes in cell morphology by translocating from the cytoplasm to the nucleus to orchestrate nuclear processes in response to the functional requirements of the cell. PMID:24599031

The Peptide Near the C Terminus Regulates Receptor CAR Nuclear Translocation Induced by Xenochemicals in Mouse Liver

PubMed Central

Zelko, Igor; Sueyoshi, Tatsuya; Kawamoto, Takeshi; Moore, Rick; Negishi, Masahiko

2001-01-01

In response to phenobarbital (PB) and other PB-type inducers, the nuclear receptor CAR translocates to the mouse liver nucleus (T. Kawamoto et al., Mol. Cell. Biol. 19:6318–6322, 1999). To define the translocation mechanism, fluorescent protein-tagged human CAR (hCAR) was expressed in the mouse livers using the in situ DNA injection and gene delivery systems. As in the wild-type hCAR, the truncated receptor lacking the C-terminal 10 residues (i.e., AF2 domain) translocated to the nucleus, indicating that the PB-inducible translocation is AF2 independent. Deletion of the 30 C-terminal residues abolished the receptor translocation, and subsequent site-directed mutagenesis delineated the PB-inducible translocation activity of the receptor to the peptide L313GLL316AEL319. Ala mutations of Leu313, Leu316, or Leu319 abrogated the translocation of CAR in the livers, while those of Leu312 or Leu315 did not affect the nuclear translocation. The leucine-rich peptide dictates the nuclear translocation of hCAR in response to various PB-type inducers and appears to be conserved in the mouse and rat receptors. PMID:11283262

Effects of Nrf2 knockdown on the properties of irradiated cell conditioned medium from A549 human lung cancer cells.

PubMed

Yoshino, Hironori; Murakami, Kanna; Nawamaki, Mikoto; Kashiwakura, Ikuo

2018-05-01

The nuclear factor erythroid 2-related factor 2 (Nrf2) plays an important role in cellular defense against oxidative stress. Recent studies have demonstrated that Nrf2 is a useful target for cancer treatment, including radiation therapy. Ionizing radiation affects, not only the irradiated cells, but also the non-irradiated neighboring cells, and this effect is known as radiation-induced bystander effect. Upon exposure to radiation, the irradiated cells transmit signals to the non-irradiated cells via gap junctions or soluble factors. These signals in turn cause biological effects, such as a decrease in the clonogenic potential and cell death, in the non-irradiated neighboring cells. Nrf2 inhibition enhances cellular radiosensitivity. However, whether this modification of radiosensitivity by Nrf2 inhibition affects the radiation-induced bystander effects is unknown. In this study, we prepared an Nrf2 knockdown human lung cancer cell A549 and investigated whether the effects of irradiated cell conditioned medium (ICCM) on cell growth and cell death induction of non-irradiated cells vary depending on the Nrf2 knockdown. We found that Nrf2 knockdown resulted in a decrease in the cell growth and an increase in the radiosensitivity of A549 cells. When non-irradiated A549 cells were transfected with control siRNA and treated with ICCM, no significant difference was observed in the cell growth and proportion of Annexin V + dead cells between ICCM from non-irradiated cells and that from 2 or 8 Gy-irradiated cells. Similarly, no significant difference was observed in the cell growth and cell death induction upon treatment with ICCM in the Nrf2 knockdown A549 cells. Taken together, these results suggest that Nrf2 knockdown decreases cell growth and enhances the radiosensitivity of A549 cells; however, it does not alter the effect of ICCM on cell growth.

Experimental demonstration of an isotope-sensitive warhead verification technique using nuclear resonance fluorescence

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vavrek, Jayson R.; Henderson, Brian S.; Danagoulian, Areg

Future nuclear arms reduction efforts will require technologies to verify that warheads slated for dismantlement are authentic without revealing any sensitive weapons design information to international inspectors. Despite several decades of research, no technology has met these requirements simultaneously. Recent work by Kemp et al. [Kemp RS, Danagoulian A, Macdonald RR, Vavrek JR (2016) Proc Natl Acad Sci USA 113:8618–8623] has produced a novel physical cryptographic verification protocol that approaches this treaty verification problem by exploiting the isotope-specific nature of nuclear resonance fluorescence (NRF) measurements to verify the authenticity of a warhead. To protect sensitive information, the NRF signal frommore » the warhead is convolved with that of an encryption foil that contains key warhead isotopes in amounts unknown to the inspector. The convolved spectrum from a candidate warhead is statistically compared against that from an authenticated template warhead to determine whether the candidate itself is authentic. Here in this paper we report on recent proof-of-concept warhead verification experiments conducted at the Massachusetts Institute of Technology. Using high-purity germanium (HPGe) detectors, we measured NRF spectra from the interrogation of proxy “genuine” and “hoax” objects by a 2.52 MeV endpoint bremsstrahlung beam. The observed differences in NRF intensities near 2.2 MeV indicate that the physical cryptographic protocol can distinguish between proxy genuine and hoax objects with high confidence in realistic measurement times.« less

Experimental demonstration of an isotope-sensitive warhead verification technique using nuclear resonance fluorescence

DOE PAGES

Vavrek, Jayson R.; Henderson, Brian S.; Danagoulian, Areg

2018-04-10

Future nuclear arms reduction efforts will require technologies to verify that warheads slated for dismantlement are authentic without revealing any sensitive weapons design information to international inspectors. Despite several decades of research, no technology has met these requirements simultaneously. Recent work by Kemp et al. [Kemp RS, Danagoulian A, Macdonald RR, Vavrek JR (2016) Proc Natl Acad Sci USA 113:8618–8623] has produced a novel physical cryptographic verification protocol that approaches this treaty verification problem by exploiting the isotope-specific nature of nuclear resonance fluorescence (NRF) measurements to verify the authenticity of a warhead. To protect sensitive information, the NRF signal frommore » the warhead is convolved with that of an encryption foil that contains key warhead isotopes in amounts unknown to the inspector. The convolved spectrum from a candidate warhead is statistically compared against that from an authenticated template warhead to determine whether the candidate itself is authentic. Here in this paper we report on recent proof-of-concept warhead verification experiments conducted at the Massachusetts Institute of Technology. Using high-purity germanium (HPGe) detectors, we measured NRF spectra from the interrogation of proxy “genuine” and “hoax” objects by a 2.52 MeV endpoint bremsstrahlung beam. The observed differences in NRF intensities near 2.2 MeV indicate that the physical cryptographic protocol can distinguish between proxy genuine and hoax objects with high confidence in realistic measurement times.« less

Treatment of Parkinson's disease in rats by Nrf2 transfection using MRI-guided focused ultrasound delivery of nanomicrobubbles

DOE Office of Scientific and Technical Information (OSTI.GOV)

Long, Ling; Cai, Xiaodong; Guo, Ruomi

Parkinson's disease (PD) is a very common neurological disorder. However, effective therapy is lacking. Although the blood-brain-barrier (BBB) protects the brain, it prevents the delivery of about 90% of drugs and nucleotides into the brain, thereby hindering the development of gene therapy for PD. Magnetic resonance imaging (MRI)-guided focused ultrasound delivery of microbubbles enhances the delivery of gene therapy vectors across the BBB and improves transfection efficiency. In the present study, we delivered nuclear factor E2-related factor 2 (Nrf2, NFE2L2) contained in nanomicrobubbles into the substantia nigra of PD rats by MRI-guided focused ultrasound, and we examined the effect ofmore » Nrf2 over-expression in this animal model of PD. The rat model of PD was established by injecting 6-OHDA in the right substantia nigra stereotactically. Plasmids (pDC315 or pDC315/Nrf2) were loaded onto nanomicrobubbles, and then injected through the tail vein with the assistance of MRI-guided focused ultrasound. MRI-guided focused ultrasound delivery of nanomicrobubbles increased gene transfection efficiency. Furthermore, Nrf2 gene transfection reduced reactive oxygen species levels, thereby protecting neurons in the target region. - Highlights: • MRI-guided focused ultrasound enhances gene transfection into the brain of rats. • Increased Nrf2 expression protects neurons in the rat model of PD. • Nrf2 protects neurons in PD by inhibiting ROS production.« less

Low-Concentration Tributyltin Decreases GluR2 Expression via Nuclear Respiratory Factor-1 Inhibition

PubMed Central

Ishida, Keishi; Aoki, Kaori; Takishita, Tomoko; Miyara, Masatsugu; Sakamoto, Shuichiro; Sanoh, Seigo; Kimura, Tomoki; Kanda, Yasunari; Ohta, Shigeru; Kotake, Yaichiro

2017-01-01

Tributyltin (TBT), which has been widely used as an antifouling agent in paints, is a common environmental pollutant. Although the toxicity of high-dose TBT has been extensively reported, the effects of low concentrations of TBT are relatively less well studied. We have previously reported that low-concentration TBT decreases α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-type glutamate receptor subunit 2 (GluR2) expression in cortical neurons and enhances neuronal vulnerability to glutamate. However, the mechanism of this TBT-induced GluR2 decrease remains unknown. Therefore, we examined the effects of TBT on the activity of transcription factors that control GluR2 expression. Exposure of primary cortical neurons to 20 nM TBT for 3 h to 9 days resulted in a decrease in GluR2 mRNA expression. Moreover, TBT inhibited the DNA binding activity of nuclear respiratory factor-1 (NRF-1), a transcription factor that positively regulates the GluR2. This result indicates that TBT inhibits the activity of NRF-1 and subsequently decreases GluR2 expression. In addition, 20 nM TBT decreased the expression of genes such as cytochrome c, cytochrome c oxidase (COX) 4, and COX 6c, which are downstream of NRF-1. Our results suggest that NRF-1 inhibition is an important molecular action of the neurotoxicity induced by low-concentration TBT. PMID:28800112

Low-Concentration Tributyltin Decreases GluR2 Expression via Nuclear Respiratory Factor-1 Inhibition.

PubMed

Ishida, Keishi; Aoki, Kaori; Takishita, Tomoko; Miyara, Masatsugu; Sakamoto, Shuichiro; Sanoh, Seigo; Kimura, Tomoki; Kanda, Yasunari; Ohta, Shigeru; Kotake, Yaichiro

2017-08-11

Tributyltin (TBT), which has been widely used as an antifouling agent in paints, is a common environmental pollutant. Although the toxicity of high-dose TBT has been extensively reported, the effects of low concentrations of TBT are relatively less well studied. We have previously reported that low-concentration TBT decreases α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-type glutamate receptor subunit 2 ( GluR2 ) expression in cortical neurons and enhances neuronal vulnerability to glutamate. However, the mechanism of this TBT-induced GluR2 decrease remains unknown. Therefore, we examined the effects of TBT on the activity of transcription factors that control GluR2 expression. Exposure of primary cortical neurons to 20 nM TBT for 3 h to 9 days resulted in a decrease in GluR2 mRNA expression. Moreover, TBT inhibited the DNA binding activity of nuclear respiratory factor-1 (NRF-1), a transcription factor that positively regulates the GluR2 . This result indicates that TBT inhibits the activity of NRF-1 and subsequently decreases GluR2 expression. In addition, 20 nM TBT decreased the expression of genes such as cytochrome c, cytochrome c oxidase (COX) 4, and COX 6c, which are downstream of NRF-1. Our results suggest that NRF-1 inhibition is an important molecular action of the neurotoxicity induced by low-concentration TBT.

Postprandial antioxidant gene expression is modified by Mediterranean diet supplemented with coenzyme Q(10) in elderly men and women.

PubMed

Yubero-Serrano, Elena M; Gonzalez-Guardia, Lorena; Rangel-Zuñiga, Oriol; Delgado-Casado, Nieves; Delgado-Lista, Javier; Perez-Martinez, Pablo; Garcia-Rios, Antonio; Caballero, Javier; Marin, Carmen; Gutierrez-Mariscal, Francisco M; Tinahones, Francisco J; Villalba, Jose M; Tunez, Isaac; Perez-Jimenez, Francisco; Lopez-Miranda, Jose

2013-02-01

Postprandial oxidative stress is characterized by an increased susceptibility of the organism towards oxidative damage after consumption of a meal rich in lipids and/or carbohydrates. We have investigated whether the quality of dietary fat alters postprandial gene expression and protein levels involved in oxidative stress and whether the supplementation with coenzyme Q(10) (CoQ) improves this situation in an elderly population. Twenty participants were randomized to receive three isocaloric diets each for 4 weeks: Mediterranean diet supplemented with CoQ (Med + CoQ diet), Mediterranean diet (Med diet), saturated fatty acid-rich diet (SFA diet). After 12-h fast, volunteers consumed a breakfast with a fat composition similar to that consumed in each of the diets. Nrf2, p22(phox) and p47(phox), superoxide dismutase 1 and 2 (SOD1 and SOD2), glutathione peroxidase 1 (GPx1), thiorredoxin reductase (TrxR) gene expression and Kelch-like ECH associating protein 1 (Keap-1) and citoplasmic and nuclear Nrf2 protein levels were determined. Med and Med + CoQ diets induced lower Nrf2, p22(phox), p47(phox), SOD1, SOD2 and TrxR gene expression and higher cytoplasmic Nrf2 and Keap-1 protein levels compared to the SFA diet. Moreover, Med + CoQ diet produced lower postprandial Nrf2 gene expression and lower nuclear Nrf2 protein levels compared to the other diets and lower GPx1 gene expression than the SFA diet. Our results support the antioxidant effect of a Med diet and that exogenous CoQ supplementation has a protective effects against free radical overgeneration through the lowering of postprandial oxidative stress modifying the postprandial antioxidant protein levels and reducing the postprandial expression of antioxidant genes in peripheral blood mononuclear cells.

Induction of activation of the antioxidant response element and stabilization of Nrf2 by 3-(3-pyridylmethylidene)-2-indolinone (PMID) confers protection against oxidative stress-induced cell death

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yao, Jia-Wei; Beijing Institute of Radiation Medicine, Beijing 100850; Liu, Jing

2012-03-01

The antioxidant response elements (ARE) are a cis-acting enhancer sequence located in regulatory regions of antioxidant and detoxifying genes. Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is a member of the Cap ‘n’ Collar family of transcription factors that binds to the ARE and regulates the transcription of specific ARE-containing genes. Under oxidative stress, Nrf2/ARE induction is fundamental to defense against reactive oxygen species (ROS) and serves as a key factor in the protection against toxic xenobiotics. 3-(3-Pyridylmethylidene)-2-Indolinone (PMID) is a derivative of 2-indolinone compounds which act as protein kinase inhibitors and show anti-tumor activity. However, the role of PMID inmore » the oxidative stress remains unknown. In the present study, we showed that PMID induced the activation of ARE-mediated transcription, increased the DNA-binding activity of Nrf2 and then up-regulated the expression of antioxidant genes such as HO-1, SOD, and NQO1. The level of Nrf2 protein was increased in cells treated with PMID by a post-transcriptional mechanism. Under CHX treatment, the stability of Nrf2 protein was enhanced by PMID with decreased turnover rate. We showed that PMID reduced the ubiquitination of Nrf2 and disrupted the Cullin3 (Cul3)-Keap1 interaction. Furthermore, cells treated with PMID showed resistance to cytotoxicity by H{sub 2}O{sub 2} and pro-oxidant 6-OHDA. PMID also up-regulated the antioxidant level in BALB/c mice. Taken together, the compound PMID induces the ARE-mediated gene expression through stabilization of Nrf2 protein and activation of Nrf2/ARE pathway and protects against oxidative stress-mediated cell death. -- Highlights: ► PMID up-regulates ARE-mediated antioxidant gene expression in vitro and in vivo. ► PMID enhances the stabilization of Nrf2 protein, decreasing Nrf2 turnover rate. ► PMID disrupted the Cullin3 (Cul3)-Keap1 interaction. ► PMID protects against cell death induced by H{sub 2}O{sub 2} and pro-oxidant 6-OHDA.« less

Ursolic acid sensitizes cisplatin-resistant HepG2/DDP cells to cisplatin via inhibiting Nrf2/ARE pathway

PubMed Central

Wu, Shouhai; Zhang, Tianpeng; Du, Jingsheng

2016-01-01

Background Combinations of adjuvant sensitizers with anticancer drugs is a promising new strategy to reverse chemoresistance. Ursolic acid (UA) is one of the natural pentacyclic triterpene compounds known to have many pharmacological characteristics such as anti-inflammatory and anticancer properties. This study investigates whether UA can sensitize hepatocellular carcinoma cells to cisplatin. Materials and methods Cells were transfected with nuclear factor erythroid-2-related factor 2 (Nrf2) small interfering RNA and Nrf2 complementary DNA by using Lipofectin 2000. The cytotoxicity of cells was investigated by Cell Counting Kit 8 assay. Cell apoptosis, cell cycle, reactive oxygen species, and mitochondrial membrane potential were detected by flow cytometry fluorescence-activated cell sorting. The protein level of Nrf2, NAD(P)H quinone oxidoreductase 1 (NQO1), glutathione S-transferase (GST), and heme oxygenase-1 (HO-1) was detected by Western blot analysis. Results The results showed that the reverse index was 2.9- and 9.69-fold by UA of 1.125 μg/mL and 2.25 μg/mL, respectively, for cisplatin to HepG2/DDP cells. UA–cisplatin combination induced cell apoptosis and reactive oxygen species, blocked the cell cycle in G0/G1 phase, and reduced the mitochondrial membrane potential. Mechanistically, UA–cisplatin dramatically decreased the expression of Nrf2 and its downstream genes. The sensibilization of UA–cisplatin combination was diminished in Nrf2 small interfering RNA-transfected HepG2/DDP cells, as well as in Nrf2 complementary DNA-transfected HepG2/DDP cells. Conclusion The results confirmed the sensibilization of UA on HepG2/DDP cells to cisplatin, which was possibly mediated via the Nrf2/antioxidant response element pathway. PMID:27822011

Anti-inflammatory effects of 4′-demethylnobiletin, a major metabolite of nobiletin

PubMed Central

Rakariyatham, Kanyasiri; Zheng, Jinkai; Guo, Shanshan; Tang, Zhonghai; Zhou, Shuangde; Xiao, Hang

2015-01-01

Nobiletin, a citrus flavonoid has been associated with various beneficial biological activities. 4′-Demethylnobiletin (4DN) is a major metabolite of nobiletin and its tissue level was found to be much higher than that of nobiletin after oral administration of nobiletin in mice. Anti-inflammatory effects of 4DN were studied in lipopolysaccharide (LPS)-treated RAW 264.7 macrophages. The results showed 4DN not only dose-dependently inhibited LPS-induced nitric oxide production, but also significantly reduced expression of pro-inflammatory mediators, namely PGE2, IL-1β and IL-6. 4DN potently suppressed the expression of iNOS and COX-2 at both protein and mRNA levels. 4DN also inhibited nuclear translocation of NF-κB and AP-1. Furthermore, we demonstrated that 4DN activated transcription factor Nrf2 and its dependent genes including HO-1 and NQO1 whose expression may contribute to anti-inflammatory effects. The results demonstrated anti-inflammatory effects of 4DN and provided a scientific basis for using nobiletin as a nutraceutical to inhibit inflammation–driven diseases. PMID:26770275

Nuclear Resonance Fluorescence for Materials Assay

DOE Office of Scientific and Technical Information (OSTI.GOV)

Quiter, Brian J.; Ludewigt, Bernhard; Mozin, Vladimir

This paper discusses the use of nuclear resonance fluorescence (NRF) techniques for the isotopic and quantitative assaying of radioactive material. Potential applications include age-dating of an unknown radioactive source, pre- and post-detonation nuclear forensics, and safeguards for nuclear fuel cycles Examples of age-dating a strong radioactive source and assaying a spent fuel pin are discussed. The modeling work has ben performed with the Monte Carlo radiation transport computer code MCNPX, and the capability to simulate NRF has bee added to the code. Discussed are the limitations in MCNPX?s photon transport physics for accurately describing photon scattering processes that are importantmore » contributions to the background and impact the applicability of the NRF assay technique.« less

Sulforaphane protects rodent retinas against ischemia-reperfusion injury through the activation of the Nrf2/HO-1 antioxidant pathway.

PubMed

Pan, Hong; He, Meihua; Liu, Ruixing; Brecha, Nicholas C; Yu, Albert Cheung Hoi; Pu, Mingliang

2014-01-01

Retinal ischemia-reperfusion (I/R) injury induces oxidative stress, leukocyte infiltration, and neuronal cell death. Sulforaphane (SF), which can be obtained in cruciferous vegetables such as broccoli, exerts protective effects in response to oxidative stress in various tissues. These effects can be initiated through nuclear factor E2-related factor 2 (Nrf2)-mediated induction of heme oxygenase-1 (HO-1). This investigation was designed to elucidate the neural protective mechanisms of SF in the retinal I/R rat model. Animals were intraperitoneally (i.p.) injected with SF (12.5 mg/kg) or vehicle (corn oil) once a day for 7 consecutive days. Then, retinal I/R was made by elevating the intraocular pressure (IOP) to 130 mmHg for 1 h. To determine if HO-1 was involved in the Nrf2 antioxidant pathway, rats were subjected to protoporphyrin IX zinc (II) (ZnPP, 30 mg/kg, i.p.) treatments at 24 h before retinal ischemia. The neuroprotective effects of SF were assessed by determining the morphology of the retina, counting the infiltrating inflammatory cells and the surviving retinal ganglion cells (RGCs) and amacrine cells, and measuring apoptosis in the retinal layers. The expression of Nrf2 and HO-1 was studied by immunofluorescence analysis and western blotting. I/R induced a marked increase of ROS generation, caused pronounced inflammation, increased the apoptosis of RGCs and amacrine cells and caused the thinning of the inner retinal layer (IRL), and these effects were diminished or abolished by SF pretreatment. Meanwhile, SF pretreatment significantly elevated the nuclear accumulation of Nrf2 and the level of HO-1 expression in the I/R retinas; however, ZnPP reversed the protective effects of SF on I/R retinas. Together, we offer direct evidence that SF had protective effects on I/R retinas, which could be attributed, at least in part, to the activation of the Nrf2/HO-1 antioxidant pathway.

Sulforaphane Protects Rodent Retinas against Ischemia-Reperfusion Injury through the Activation of the Nrf2/HO-1 Antioxidant Pathway

PubMed Central

Liu, Ruixing; Brecha, Nicholas C.; Yu, Albert Cheung Hoi; Pu, Mingliang

2014-01-01

Retinal ischemia-reperfusion (I/R) injury induces oxidative stress, leukocyte infiltration, and neuronal cell death. Sulforaphane (SF), which can be obtained in cruciferous vegetables such as broccoli, exerts protective effects in response to oxidative stress in various tissues. These effects can be initiated through nuclear factor E2-related factor 2 (Nrf2)-mediated induction of heme oxygenase-1 (HO-1). This investigation was designed to elucidate the neural protective mechanisms of SF in the retinal I/R rat model. Animals were intraperitoneally (i.p.) injected with SF (12.5 mg/kg) or vehicle (corn oil) once a day for 7 consecutive days. Then, retinal I/R was made by elevating the intraocular pressure (IOP) to 130 mmHg for 1 h. To determine if HO-1 was involved in the Nrf2 antioxidant pathway, rats were subjected to protoporphyrin IX zinc (II) (ZnPP, 30 mg/kg, i.p.) treatments at 24 h before retinal ischemia. The neuroprotective effects of SF were assessed by determining the morphology of the retina, counting the infiltrating inflammatory cells and the surviving retinal ganglion cells (RGCs) and amacrine cells, and measuring apoptosis in the retinal layers. The expression of Nrf2 and HO-1 was studied by immunofluorescence analysis and western blotting. I/R induced a marked increase of ROS generation, caused pronounced inflammation, increased the apoptosis of RGCs and amacrine cells and caused the thinning of the inner retinal layer (IRL), and these effects were diminished or abolished by SF pretreatment. Meanwhile, SF pretreatment significantly elevated the nuclear accumulation of Nrf2 and the level of HO-1 expression in the I/R retinas; however, ZnPP reversed the protective effects of SF on I/R retinas. Together, we offer direct evidence that SF had protective effects on I/R retinas, which could be attributed, at least in part, to the activation of the Nrf2/HO-1 antioxidant pathway. PMID:25470382

Cross Talk in HEK293 Cells Between Nrf2, HIF, and NF-κB Activities upon Challenges with Redox Therapeutics Characterized with Single-Cell Resolution.

PubMed

Johansson, Katarina; Cebula, Marcus; Rengby, Olle; Dreij, Kristian; Carlström, Karl E; Sigmundsson, Kristmundur; Piehl, Fredrik; Arnér, Elias S J

2017-02-20

Many transcription factors with importance in health and disease are redox regulated. However, how their activities may be intertwined in responses to redox-perturbing stimuli is poorly understood. To enable in-depth characterization of this aspect, we here developed a methodology for simultaneous determination of nuclear factor E2-related factor 2 (Nrf2), hypoxia-inducible factor (HIF), and nuclear factor kappa-light-chain-enhancer of activated B cell (NF-κB) activation at single-cell resolution, using a new tool named pTRAF (plasmid for transcription factor reporter activation based upon fluorescence). The pTRAF allowed determination of Nrf2, HIF, and NF-κB activities in a high-resolution and high-throughput manner, and we here assessed how redox therapeutics affected the activities of these transcription factors in human embryonic kidney cells (HEK293). Cross talk was detected between the three signaling pathways upon some types of redox therapeutics, also by using inducers typically considered specific for Nrf2, such as sulforaphane or auranofin, hypoxia for HIF activation, or tumor necrosis factor alpha (TNFα) for NF-κB stimulation. Doxorubicin, at low nontoxic doses, potentiated TNFα-induced activation of NF-κB and HIF, without effects in stand-alone treatment. Stochastic activation patterns in cell cultures were also considerable upon challenges with several redox stimuli. A novel strategy was here used to study simultaneous activation of Nrf2, HIF, and NF-κB in single cells. The method can also be adapted for studies of other transcription factors. The pTRAF provides new opportunities for in-depth studies of transcription factor activities. In this study, we found that upon challenges of cells with several redox-perturbing conditions, Nrf2, HIF, and NF-κB are uniquely responsive to separate stimuli, but can also display marked cross talk to each other within single cells. Antioxid. Redox Signal. 26, 229-246.

γ-Tocotrienol prevents 5-FU-induced reactive oxygen species production in human oral keratinocytes through the stabilization of 5-FU-induced activation of Nrf2.

PubMed

Takano, Hideyuki; Momota, Yukihiro; Kani, Kouichi; Aota, Keiko; Yamamura, Yoshiko; Yamanoi, Tomoko; Azuma, Masayuki

2015-04-01

Chemotherapy-induced oral mucositis is a common adverse event in patients with oral squamous cell carcinoma, and is initiated through a variety of mechanisms, including the generation of reactive oxygen species (ROS). In this study, we examined the preventive effect of γ-tocotrienol on the 5-FU-induced ROS production in human oral keratinocytes (RT7). We treated RT7 cells with 5-FU and γ-tocotrienol at concentrations of 10 µg/ml and 10 nM, respectively. When cells were treated with 5-FU alone, significant growth inhibition was observed as compared to untreated cells. This inhibition was, in part, due to the ROS gene-rated by 5-FU treatment, because N-acetyl cysteine (NAC), a ROS scavenger, significantly ameliorated the growth of RT7 cells. γ-tocotrienol showed no cytotoxic effect on the growth of RT7 cells. Simultaneous treatment of cells with these agents resulted in the significant recovery of cell growth, owing to the suppression of ROS generation by γ-tocotrienol. Whereas 5-FU stimulated the expression of NF-E2-related factor 2 (Nrf2) protein in the nucleus up to 12 h after treatment of RT7 cells, γ-tocotrienol had no obvious effect on the expression of nuclear Nrf2 protein. Of note, the combined treatment with both agents stabilized the 5-FU-induced nuclear Nrf2 protein expression until 24 h after treatment. In addition, expression of Nrf2-dependent antioxidant genes, such as heme oxygenase-1 (HO-1) and quinone oxidoreductase-1 (NQO-1), was significantly augmented by treatment of cells with both agents. These findings suggest that γ-tocotrienol could prevent 5-FU-induced ROS generation by stabilizing Nrf2 activation, thereby leading to ROS detoxification and cell survival in human oral keratinocytes.

γ-tocotrienol prevents 5-FU-induced reactive oxygen species production in human oral keratinocytes through the stabilization of 5-FU-induced activation of Nrf2

PubMed Central

TAKANO, HIDEYUKI; MOMOTA, YUKIHIRO; KANI, KOUICHI; AOTA, KEIKO; YAMAMURA, YOSHIKO; YAMANOI, TOMOKO; AZUMA, MASAYUKI

2015-01-01

Chemotherapy-induced oral mucositis is a common adverse event in patients with oral squamous cell carcinoma, and is initiated through a variety of mechanisms, including the generation of reactive oxygen species (ROS). In this study, we examined the preventive effect of γ-tocotrienol on the 5-FU-induced ROS production in human oral keratinocytes (RT7). We treated RT7 cells with 5-FU and γ-tocotrienol at concentrations of 10 μg/ml and 10 nM, respectively. When cells were treated with 5-FU alone, significant growth inhibition was observed as compared to untreated cells. This inhibition was, in part, due to the ROS generated by 5-FU treatment, because N-acetyl cysteine (NAC), a ROS scavenger, significantly ameliorated the growth of RT7 cells. γ-tocotrienol showed no cytotoxic effect on the growth of RT7 cells. Simultaneous treatment of cells with these agents resulted in the significant recovery of cell growth, owing to the suppression of ROS generation by γ-tocotrienol. Whereas 5-FU stimulated the expression of NF-E2-related factor 2 (Nrf2) protein in the nucleus up to 12 h after treatment of RT7 cells, γ-tocotrienol had no obvious effect on the expression of nuclear Nrf2 protein. Of note, the combined treatment with both agents stabilized the 5-FU-induced nuclear Nrf2 protein expression until 24 h after treatment. In addition, expression of Nrf2-dependent antioxidant genes, such as heme oxygenase-1 (HO-1) and NAD(P)H:quinone oxidoreductase-1 (NQO-1), was significantly augmented by treatment of cells with both agents. These findings suggest that γ-tocotrienol could prevent 5-FU-induced ROS generation by stabilizing Nrf2 activation, thereby leading to ROS detoxification and cell survival in human oral keratinocytes. PMID:25625649

NRF2-regulated metabolic gene signature as a prognostic biomarker in non-small cell lung cancer

PubMed Central

Namani, Akhileshwar; Cui, Qin Qin; Wu, Yihe; Wang, Hongyan; Wang, Xiu Jun; Tang, Xiuwen

2017-01-01

Mutations in Kelch-like ECH-associated protein 1 (KEAP1) cause the aberrant activation of nuclear factor erythroid-derived 2-like 2 (NRF2), which leads to oncogenesis and drug resistance in lung cancer cells. Our study was designed to identify the genes involved in lung cancer progression targeted by NRF2. A series of microarray experiments in normal and cancer cells, as well as in animal models, have revealed regulatory genes downstream of NRF2 that are involved in wide variety of pathways. Specifically, we carried out individual and combinatorial microarray analysis of KEAP1 overexpression and NRF2 siRNA-knockdown in a KEAP1 mutant-A549 non-small cell lung cancer (NSCLC) cell line. As a result, we identified a list of genes which were mainly involved in metabolic functions in NSCLC by using functional annotation analysis. In addition, we carried out in silico analysis to characterize the antioxidant responsive element sequences in the promoter regions of known and putative NRF2-regulated metabolic genes. We further identified an NRF2-regulated metabolic gene signature (NRMGS) by correlating the microarray data with lung adenocarcinoma RNA-Seq gene expression data from The Cancer Genome Atlas followed by qRT-PCR validation, and finally showed that higher expression of the signature conferred a poor prognosis in 8 independent NSCLC cohorts. Our findings provide novel prognostic biomarkers for NSCLC. PMID:29050246

Adipose-specific ablation of Nrf2 transiently delayed high-fat diet-induced obesity by altering glucose, lipid and energy metabolism of male mice.

PubMed

Zhang, Le; Dasuri, Kalavathi; Fernandez-Kim, Sun-Ok; Bruce-Keller, Annadora J; Keller, Jeffrey N

2016-01-01

Nuclear factor E2-related factor 2 (NRF2) is a well-known master controller of the cellular adaptive antioxidant and detoxification response. Recent studies demonstrated altered glucose, lipid and energy metabolism in mice with a global Nrf2 knockout. In the present study, we aim to determine the effects of an adipose-specific ablation of Nrf2 (ASAN) on diet-induced obesity (DIO) in male mice. The 6-week-old adipose-specific Nrf2 knockout (NK) and its Nrf2 control (NC) mice were fed with either control diet (CD) or high-fat diet (HFD) for 14 weeks. NK mice exhibited transiently delayed body weight (BW) growth from week 5 to week 11 of HFD feeding, higher daily physical activity levels and preferential use of fat over carbohydrates as a source of energy at week 8 of the CD-feeding period. After 14 weeks of feeding, NK mice showed comparable results with NC mice with respect to the overall BW and body fat content, but exhibited reduced blood glucose, reduced number but increased size of adipocytes, accompanied with elevated expression of many genes and proteins in the visceral fat related to glucose, lipid and energy metabolism (e.g. Fgf21 , Pgc1a ). These results indicated that NRF2 is an important mediator for glucose, lipid and energy metabolism in adipose tissue, and ASAN could have beneficial effect for prevention of DIO during the early development of mice.

NOS1 mediates AP1 nuclear translocation and inflammatory response.

PubMed

Srivastava, Mansi; Baig, Mirza S

2018-06-01

A hallmark of the AP1 functioning is its nuclear translocation, which induces proinflammatory cytokine expression and hence the inflammatory response. After endotoxin shock AP1 transcription factor, which comprises Jun, ATF2, and Fos family of proteins, translocates into the nucleus and induces proinflammatory cytokine expression. In the current study, we found, NOS1 inhibition prevents nuclear translocation of the AP1 transcription factor subunits. Pharmacological inhibition of NOS1 impedes translocation of subunits into the nucleus, suppressing the transcription of inflammatory genes causing a diminished inflammatory response. In conclusion, the study shows the novel mechanism of NOS1- mediated AP1 nuclear translocation, which needs to be further explored. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

The Nrf1 CNC-bZIP protein is regulated by the proteasome and activated by hypoxia.

PubMed

Chepelev, Nikolai L; Bennitz, Joshua D; Huang, Ting; McBride, Skye; Willmore, William G

2011-01-01

Nrf1 (nuclear factor-erythroid 2 p45 subunit-related factor 1) is a transcription factor mediating cellular responses to xenobiotic and pro-oxidant stress. Nrf1 regulates the transcription of many stress-related genes through the electrophile response elements (EpREs) located in their promoter regions. Despite its potential importance in human health, the mechanisms controlling Nrf1 have not been addressed fully. We found that proteasomal inhibitors MG-132 and clasto-lactacystin-β-lactone stabilized the protein expression of full-length Nrf1 in both COS7 and WFF2002 cells. Concomitantly, proteasomal inhibition decreased the expression of a smaller, N-terminal Nrf1 fragment, with an approximate molecular weight of 23 kDa. The EpRE-luciferase reporter assays revealed that proteasomal inhibition markedly inhibited the Nrf1 transactivational activity. These results support earlier hypotheses that the 26 S proteasome processes Nrf1 into its active form by removing its inhibitory N-terminal domain anchoring Nrf1 to the endoplasmic reticulum. Immunoprecipitation demonstrated that Nrf1 is ubiquitinated and that proteasomal inhibition increased the degree of Nrf1 ubiquitination. Furthermore, Nrf1 protein had a half-life of approximately 5 hours in COS7 cells. In contrast, hypoxia (1% O(2)) significantly increased the luciferase reporter activity of exogenous Nrf1 protein, while decreasing the protein expression of p65, a shorter form of Nrf1, known to act as a repressor of EpRE-controlled gene expression. Finally, the protein phosphatase inhibitor okadaic acid activated Nrf1 reporter activity, while the latter was repressed by the PKC inhibitor staurosporine. Collectively, our data suggests that Nrf1 is controlled by several post-translational mechanisms, including ubiquitination, proteolytic processing and proteasomal-mediated degradation as well as by its phosphorylation status. © 2011 Chepelev et al.

Protective role of Nrf2 against mechanical-stretch-induced apoptosis in mouse fibroblasts: a potential therapeutic target of mechanical-trauma-induced stress urinary incontinence.

PubMed

Li, Qiannan; Li, Bingshu; Liu, Cheng; Wang, Linlin; Tang, Jianming; Hong, Li

2018-01-10

We investigated the protective effect and underlying molecular mechanism of nuclear factor-E2-related factor 2 (Nrf2) against mechanical-stretch-induced apoptosis in mouse fibroblasts. Normal cells, Nrf2 silencing cells, and Nrf2 overexpressing cells were respectively divided into two groups-nonintervention and cyclic mechanical strain (CMS)-subjected to CMS of 5333 μ (1.0 Hz for 4 h), six groups in total (control, CMS, shNfe212, shNfe212 + CMS, LV-shNfe212, and LV-shNfe212 + CMS). After treatment, cell apoptosis; cell-cycle distribution; expressions of Nrf2, Bax, Bcl-2, Cyt-C, caspase-3, caspase-9, cleaved-caspase-3, and cleaved-caspase-9; mitochondrial membrane potential (ΔΨm); reactive oxygen species (ROS); and malondialdehyde (MDA) levels were measured. Thirty virgin female C57BL/6 mice were divided into two groups: control (without intervention) and vaginal distension (VD) groups, which underwent VD for 1 h with an 8-mm dilator (0.3 ml saline). Leak-point pressure (LPP) was tested on day 7 after VD; Nrf2 expression, apoptosis, and MDA levels were then measured in urethra and anterior vaginal wall. Mechanical stretch decreased Nrf2 messenger RNA (mRNA) and protein expressions. Overexpression of Nrf2 alleviated mechanical-stretch-induced cell apoptosis; S-phase arrest of cell cycle; up-regulation of Bax, cytochrome C (Cyt-C), ROS, MDA, ratio of cleaved-caspase-3/caspase-3 and cleaved-caspase-9/caspase-9; and exacerbated the decrease of Bcl2 and ΔΨm in L929 cells. On the contrary, silencing of Nrf2 showed opposite effects. Besides, VD reduced LPP levels and Nrf2 expression and increased cell apoptosis and MDA generation in the urethra and anterior vaginal wall. Nrf2 exhibits a protective role against mechanical-stretch -induced apoptosis on mouse fibroblasts, which might indicate a potential therapeutic target of mechanical-trauma-induced stress urinary incontinence (SUI).

Inorganic Arsenic Induces NRF2-Regulated Antioxidant Defenses in Both Cerebral Cortex and Hippocampus in Vivo.

PubMed

Zhang, Yang; Duan, Xiaoxu; Li, Jinlong; Zhao, Shuo; Li, Wei; Zhao, Lu; Li, Wei; Nie, Huifang; Sun, Guifang; Li, Bing

2016-08-01

Inorganic arsenic is reported to induce the reactive oxygen species-mediated oxidative stress, which is supposed to be one of the main mechanisms of arsenic-related neurological diseases. Nuclear factor erythroid 2-related factor 2 (NRF2), a master regulator of antioxidant defense systems, up-regulates the expression of target genes to fight against oxidative damages caused by harmful substances, including metals. In the present study, mice were used as a model to investigate the oxidative stress levels and the expressions of NRF2-regulated antioxidant substances in both cerebral cortex and hippocampus with 5, 10 and 20 mg/kg NaAsO2 exposure intra-gastrically. Our results showed that acute NaAsO2 treatment resulted in decreased total anti-oxidative capacity (T-AOC) and increased maleic dialdehyde production in the nervous system. We also detected rapidly elevation of NRF2 protein levels by enhancement of Nrf2 transcription, especially at 20 mg/kg NaAsO2 exposure group. In the meantime, mRNA and protein levels of Nrf2 encoding antioxidant enzymes heme oxygenase-1 (HO-1), NAD(P)H: quinine oxidoreductase 1 (NQO1) and glutathione S-transferase (GST) were consistently elevated time- and dose-dependently both in the cerebral cortex and hippocampus. Taken together, the presence study demonstrated the activation of NRF2 pathway, an early antioxidant defensive response, in both cerebral cortex and hippocampus upon inorganic arsenic (iAs) exposure in vivo. A better knowledge on the roles of NRF2 pathway in maintaining cellular redox homeostasis would be helpful for the strategies on improvement of neurotoxicity related to this metalloid.

S-Glutathionylation of Keap1: a new role for glutathione S-transferase pi in neuronal protection.

PubMed

Carvalho, Andreia Neves; Marques, Carla; Guedes, Rita C; Castro-Caldas, Margarida; Rodrigues, Elsa; van Horssen, Jack; Gama, Maria João

2016-05-01

Oxidative stress is a key pathological feature of Parkinson's disease (PD). Glutathione S-transferase pi (GSTP) is a neuroprotective antioxidant enzyme regulated at the transcriptional level by the antioxidant master regulator nuclear factor-erythroid 2-related factor 2 (Nrf2). Here, we show for the first time that upon MPTP-induced oxidative stress, GSTP potentiates S-glutathionylation of Kelch-like ECH-associated protein 1 (Keap1), an endogenous repressor of Nrf2, in vivo. S-glutathionylation of Keap1 leads to Nrf2 activation and subsequently increases expression of GSTP. This positive feedback regulatory loop represents a novel mechanism by which GSTP elicits antioxidant protection in the brain. © 2016 Federation of European Biochemical Societies.

Paeonol extracted from Paeonia suffruticosa Andr. ameliorated UVB-induced skin photoaging via DLD/Nrf2/ARE and MAPK/AP-1 pathway.

PubMed

Sun, ZhengWang; Du, Juan; Hwang, Eunson; Yi, Tae-Hoo

2018-05-10

Paeonia suffruticosa Andr. (PS) has been used in traditional Chinese medicine for a long time. However, there are no studies that investigate the preventive effects of PS on ultraviolet B (UVB)-induced photoaging. In this study, paeonol (PA) was detected the main compound in PS root. In vitro, PS and PA significantly inhibited UVB-induced phosphorylation of mitogen-activated protein kinase and activator protein 1 in keratinocytes, which consequently led to degradation of procollagen type I. On the other hand, PS and PA increased NAD(P)H:quinone oxidoreductase 1 and heme oxygenase-1 expression, confirmed by greater nuclear accumulation of nuclear factor E2-releated factor 2 (Nrf2). Furthermore, this study proved that the endogenous antioxidant system Nrf2/antioxidant response element was regulated by dihydrolipoamide dehydrogenase, a tricarboxylic acid (TCA) cycle-associated protein whose level was decreased after UVB exposure. PS and PA promoted the production of dihydrolipoamide dehydrogenase, as well as the activation of Nrf2 and antioxidant response element, resulting in preventing procollagen type I ruined caused by UVB. In vivo, topical application of PS and PA attenuated UVB-induced matrix metalloproteinase-1 production and promoted procollagen type I in hairless mice. These results suggested PA a promising botanical in protecting skin from UVB-induced photoaging. Copyright © 2018 John Wiley & Sons, Ltd.

Impact of epigallocatechin‑3‑gallate on expression of nuclear factor erythroid 2‑related factor 2 and γ‑glutamyl cysteine synthetase genes in oxidative stress‑induced mouse renal tubular epithelial cells.

PubMed

Du, Xuanyi; Yu, Jinfeng; Sun, Xiaohan; Qu, Shaochuan; Zhang, Haitao; Hu, Mengying; Yang, Shufen; Zhou, Ping

2018-06-01

The aim of the present study was to investigate the antioxidant response mechanism of epigallocatechin‑3‑gallate (EGCG) in H2O2‑induced mouse renal tubular epithelial cells (MRTECs). The cultured MRTECs were divided into normal, H2O2 (control) and EGCG treatment groups. The MTT assay was used to assess cell viability, and reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR), immunocytochemical and western blot analyses were performed to detect the expression of nuclear factor erythroid 2‑related factor 2 (Nrf2) and γ‑glutamyl cysteine synthetase (γ‑GCS). EGCG was able to mitigate H2O2‑mediated cell damage. The RT‑qPCR results demonstrated that EGCG was able to upregulate the gene expression of Nrf2 and γ‑GCS in MRTECs in a dose‑dependent manner. The immunocytochemistry and western blot analyses demonstrated that EGCG was able to increase the protein expression of Nrf2 and γ‑GCS in MRTECs in a dose‑dependent manner. Oxidative stress may lead to a decrease in the viability of MRTECs, while EGCG was able to promote the expression of Nrf2 and γ‑GCS in MRTECs, thereby improving the antioxidant capacity of the cells and promoting the repair of oxidative stress injury.

Sesquiterpene Lactone Composition and Cellular Nrf2 Induction of Taraxacum officinale Leaves and Roots and Taraxinic Acid β-d-Glucopyranosyl Ester.

PubMed

Esatbeyoglu, Tuba; Obermair, Betina; Dorn, Tabea; Siems, Karsten; Rimbach, Gerald; Birringer, Marc

2017-01-01

Taraxacum officinale, the common dandelion, is a plant of the Asteraceae family, which is used as a food and medical herb. Various secondary plant metabolites such as sesquiterpene lactones, triterpenoids, flavonoids, phenolic acids, coumarins, and steroids have been described to be present in T. officinale. Dandelion may exhibit various health benefits, including antioxidant, anti-inflammatory, and anticarcinogenic properties. We analyzed the leaves and roots of the common dandelion (T. officinale) using high-performance liquid chromatography/mass spectrometry to determine its sesquiterpene lactone composition. The main compound of the leaf extract taraxinic acid β-d-glucopyranosyl ester (1), a sesquiterpene lactone, was isolated and the structure elucidation was conducted by nuclear magnetic resonance spectrometry. The leaf extract and its main compound 1 activated the transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) in human hepatocytes more significantly than the root extract. Furthermore, the leaf extract induced the Nrf2 target gene heme oxygenase 1. Overall, present data suggest that compound 1 may be one of the active principles of T. officinale.

Antioxidant Opuntia ficus-indica Extract Activates AHR-NRF2 Signaling and Upregulates Filaggrin and Loricrin Expression in Human Keratinocytes.

PubMed

Nakahara, Takeshi; Mitoma, Chikage; Hashimoto-Hachiya, Akiko; Takahara, Masakazu; Tsuji, Gaku; Uchi, Hiroshi; Yan, Xianghong; Hachisuka, Junichi; Chiba, Takahito; Esaki, Hitokazu; Kido-Nakahara, Makiko; Furue, Masutaka

2015-10-01

Opuntia ficus-indica (OFI) is a cactus species widely used as an anti-inflammatory, antilipidemic, and hypoglycemic agent. It has been shown that OFI extract (OFIE) inhibits oxidative stress in animal models of diabetes and hepatic disease; however, its antioxidant mechanism remains largely unknown. In this study, we demonstrated that OFIE exhibited potent antioxidant activity through the activation of nuclear factor erythroid 2-related factor 2 (NRF2) and the downstream antioxidant enzyme quinone oxidoreductase 1 (NQO1), which inhibited the generation of reactive oxygen species in keratinocytes challenged with tumor necrosis factor α or benzo[α]pyrene. The antioxidant capacity of OFIE was canceled in NRF2 knockdown keratinocytes. OFIE exerted this NRF2-NQO1 upregulation through activation of the aryl hydrocarbon receptor (AHR). Moreover, the ligation of AHR by OFIE upregulated the expression of epidermal barrier proteins: filaggrin and loricrin. OFIE also prevented TH2 cytokine-mediated downregulation of filaggrin and loricrin expression in an AHR-dependent manner because it was canceled in AHR knockdown keratinocytes. Antioxidant OFIE is a potent activator of AHR-NRF2-NQO1 signaling and may be beneficial in treating barrier-disrupted skin disorders.

Expression of Nrf2 Promotes Schwann Cell-Mediated Sciatic Nerve Recovery in Diabetic Peripheral Neuropathy.

PubMed

Tang, Wei; Chen, Xiangfang; Liu, Haoqi; Lv, Qian; Zou, Junjie; Shi, Yongquan; Liu, Zhimin

2018-04-26

High glucose-induced oxidative stress and inflammatory responses play an important role in painful diabetic neuropathy by activating the TLR4/NFκB signal pathway. Schwann cells (SCs) are integral to peripheral nerve biology, contributing to saltatory conduction along axons, nerve and axon development, and axonal regeneration. SCs provide a microenvironment favoring vascular regeneration but their low survival ratio in hyperglycemic conditions suppress the function to promote nerve growth. Nuclear factor erythroid 2-related factor 2 (Nrf2) promotes remyelination after peripheral nerve injury. The aim of this study was to identify the role of Nrf2 in SC-mediated functional recovery after sciatic nerve injury. We compared plasma inflammatory factors in diabetic patients (DN) with/without diabetic peripheral neuropathy (DPN) and assessed whether Nrf2 expression in SCs could repair peripheral nerve injury in a rat model. Nrf2, TLR4/NFκB signal pathway and apoptosis relative protein expression were detected by western blot. Apoptosis and angiogenesis were determined by immunofluorescence and tubule formation assay, respectively. Regenerated nerves were determined by transmission electron microscope. Higher levels of inflammatory factors and VEGF expression were found in DPN patients. Cellular experiments indicate that Nrf2 expression inhibits hyperglycemia-induced apoptosis and promotes angiogenesis by regulating the TLR4/NFκB signal pathway. Animal experiments show that nerve conduction velocity, myelin sheath thickness, and sciatic vasa nervorum are restored with transplantation of SCs overexpressing Nrf2. Taken together, the high survival ratio of SCs in a DPN rat model indicates that overexpression of Nrf2 restores nerve injury. © 2018 The Author(s). Published by S. Karger AG, Basel.

Luteolin inhibits the Nrf2 signaling pathway and tumor growth in vivo

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chian, Song; Thapa, Ruby; Chi, Zhexu

Highlights: • Luteolin inhibits the Nrf2 pathway in mouse liver and in xenografted tumors. • Luteolin markedly inhibits the growth of xenograft tumors. • Luteolin enhances the anti-cancer effect of cisplatin in mice in vivo. • Luteolin could serve as an adjuvant in the chemotherapy of NSCLC. - Abstract: Nuclear factor erythroid 2-related factor 2 (Nrf2) is over-expressed in many types of tumor, promotes tumor growth, and confers resistance to anticancer therapy. Hence, Nrf2 is regarded as a novel therapeutic target in cancer. Previously, we reported that luteolin is a strong inhibitor of Nrf2 in vitro. Here, we showed thatmore » luteolin reduced the constitutive expression of NAD(P)H quinone oxidoreductase 1 in mouse liver in a time- and dose-dependent manner. Further, luteolin inhibited the expression of antioxidant enzymes and glutathione transferases, decreasing the reduced glutathione in the liver of wild-type mice under both constitutive and butylated hydroxyanisole-induced conditions. In contrast, such distinct responses were not detected in Nrf2{sup −/−} mice. In addition, oral administration of luteolin, either alone or combined with intraperitoneal injection of the cytotoxic drug cisplatin, greatly inhibited the growth of xenograft tumors from non-small-cell lung cancer (NSCLC) cell line A549 cells grown subcutaneously in athymic nude mice. Cell proliferation, the expression of Nrf2, and antioxidant enzymes were all reduced in tumor xenograft tissues. Furthermore, luteolin enhanced the anti-cancer effect of cisplatin. Together, our findings demonstrated that luteolin inhibits the Nrf2 pathway in vivo and can serve as an adjuvant in the chemotherapy of NSCLC.« less

Concerted action of p62 and Nrf2 protects cells from palmitic acid-induced lipotoxicity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Park, Jeong Su; Yonsei Biomedical Research Institute, Yonsei University College of Medicine, 50 Yonsei-ro, Seodaemun-gu, Seoul 120-752; Kang, Dong Hoon

Nonalcoholic fatty liver disease (NAFLD), frequently associated with obesity and diabetes mellitus, is caused by the accumulation of excess fatty acids within liver cells. Palmitic acid (PA), a common saturated fatty acid found in mammals, induces the generation of reactive oxygen species (ROS) and elicits apoptotic cell death, known as lipotoxicity. However, protective mechanisms against PA-induced lipotoxicity have not been elucidated. In this study, we aimed to clarify the role of p62, an adapter protein in the autophagic process, as well as the nuclear factor erythroid 2-related factor 2 (Nrf2)-Kelch-like ECH-associated protein 1 (Keap1) pathway, in protecting cells from PA-inducedmore » lipotoxicity. The Nrf2-Keap1 pathway is essential for the protection of cells from oxidative stress. p62 enhances its binding to Keap1 and leads to Nrf2 activation. Here, we show that PA potentiates Keap1 degradation and thereby activates the transcription of Nrf2 target genes partially through autophagy. Furthermore, this PA-mediated Keap1 degradation depends on p62. Correspondingly, a lack of p62 attenuates the PA-mediated Nrf2 activation and increases the susceptibility of cells to oxidative stress. These results indicate that p62 plays an important role in protecting cells against lipotoxicity through Keap1 degradation-mediated Nrf2 activation. - Highlights: • PA induces Keap1 downregulation and activates Nrf2 target gene transcription. • PA-induced Keap1 degradation is partly mediated by the autophagic pathway. • PA-induced Keap1 degradation depends on p62. • Ablation of p62 exacerbates PA-mediated apoptotic cell death.« less

Activation of Nrf2 Reduces UVA-Mediated MMP-1 Upregulation via MAPK/AP-1 Signaling Cascades: The Photoprotective Effects of Sulforaphane and Hispidulin

PubMed Central

Chaiprasongsuk, Anyamanee; Lohakul, Jinaphat; Soontrapa, Kitipong; Sampattavanich, Somponnat; Akarasereenont, Pravit

2017-01-01

UVA irradiation plays a role in premature aging of the skin through triggering oxidative stress-associated stimulation of matrix metalloproteinase-1 (MMP-1) responsible for collagen degradation, a hallmark of photoaged skin. Compounds that can activate nuclear factor E2-related factor 2 (Nrf2), a transcription factor regulating antioxidant gene expression, should therefore serve as effective antiphotoaging agents. We investigated whether genetic silencing of Nrf2 could relieve UVA-mediated MMP-1 upregulation via activation of mitogen-activated protein kinase (MAPK)/activator protein 1 (AP-1) signaling using human keratinocyte cell line (HaCaT). Antiphotoaging effects of hispidulin (HPD) and sulforaphane (SFN) were assessed on their abilities to activate Nrf2 in controlling MMP-1 and collagen expressions in association with phosphorylation of MAPKs (extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38), c-Jun, and c-Fos, using the skin of BALB/c mice subjected to repetitive UVA irradiation. Our findings suggested that depletion of Nrf2 promoted both mRNA expression and activity of MMP-1 in the UVA-irradiated HaCaT cells. Treatment of Nrf2 knocked-down HaCaT cells with MAPK inhibitors significantly suppressed UVA-induced MMP-1 and AP-1 activities. Moreover, pretreatment of the mouse skin with HPD and SFN, which could activate Nrf2, provided protective effects against UVA-mediated MMP-1 induction and collagen depletion in correlation with the decreased levels of phosphorylated MAPKs, c-Jun, and c-Fos in the mouse skin. In conclusion, Nrf2 could influence UVA-mediated MMP-1 upregulation through the MAPK/AP-1 signaling cascades. HPD and SFN may therefore represent promising antiphotoaging candidates. PMID:28011874

Comparing the effects of mitochondrial targeted and localized antioxidants with cellular antioxidants in human skin cells exposed to UVA and hydrogen peroxide.

PubMed

Oyewole, Anne O; Wilmot, Marie-Claire; Fowler, Mark; Birch-Machin, Mark A

2014-01-01

Skin cancer and aging are linked to increased cellular reactive oxygen species (ROS), particularly following exposure to ultraviolet A (UVA) in sunlight. As mitochondria are the main source of cellular ROS, this study compared the protective effects of mitochondria-targeted and -localized antioxidants (MitoQ and tiron, respectively) with cellular antioxidants against oxidative stress-induced [UVA and hydrogen peroxide (H2O2)] mitochondrial DNA (mtDNA) damage in human dermal fibroblasts. With the use of a long quantitative PCR assay, tiron (EC50 10 mM) was found to confer complete (100%) protection (P<0.001) against both UVA- and H2O2-induced mtDNA damage, whereas MitoQ (EC50 750 nM) provided less protection (17 and 32%, respectively; P<0.05). This particular protective effect of tiron was greater than a range of cellular antioxidants investigated. The nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway provides cellular protection against oxidative stress. An ELISA assay for the Nrf2 target gene heme oxygenase-1 (HO-1) and studies using Nrf2 small interfering RNA both indicated that tiron's mode of action was Nrf2 independent. The comet assay showed that tiron's protective effect against H2O2-induced nuclear DNA damage was greater than the cellular antioxidants and MitoQ (P<0.001). This study provides a platform to investigate molecules with similar structure to tiron as potent and clinically relevant antioxidants.

Suppression of antioxidant Nrf-2 and downstream pathway in H9c2 cells by advanced glycation end products (AGEs) via ERK phosphorylation.

PubMed

Ko, Shun-Yao; Chang, Shu-Shing; Lin, I-Hsuan; Chen, Hong-I

2015-11-01

Diabetic cardiomyopathy is related to oxidative stress and correlated with the presence of advanced glycation end products (AGEs). In a clinical setting, AGEs can be detected in patients presenting diabetic cardiomyopathy; however, the underlying mechanism has yet to be elucidated. In our previous study, AGEs increase cell hypertrophy via ERK phosphorylation in a process closely related to ROS production. Thus, we propose that AGEs regulate the antioxidant gene nuclear factor-erythroid 2-related factor (Nrf-2). In H9c2 cells treated with AGEs, the expression of Nrf-2 was reduced; however, ERK phosphorylation was shown to increase. Treatment with H2O2 was also shown to increase Nrf-2 and ERK phosphorylation. In cells pretreatment with ROS scavenger NAC, the effects of H2O2 were reduced; however, the effects of the AGEs remained largely unchanged. Conversely, when cells were pretreated with PD98059 (ERK inhibitor), the expression of Nrf-2 was recovered following treatment with AGEs. Our results suggest that AGEs inhibit Nrf-2 via the ERK pathway; however, this influence is partly associated with ROS. Our finding further indicated that AGEs possess both ROS-dependent and ROS-independent pathways, resulting in a reduction in Nrf-2. This report reveals an important mechanism underlying the regulation of diabetic cardiomyopathy progression by AGEs. Copyright © 2015 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

Butein induction of HO-1 by p38 MAPK/Nrf2 pathway in adipocytes attenuates high-fat diet induced adipose hypertrophy in mice.

PubMed

Wang, Zheng; Ka, Sun-O; Lee, Youngyi; Park, Byung-Hyun; Bae, Eun Ju

2017-03-15

Adipose tissue inflammation and oxidative stress are key components in the development of obesity and insulin resistance. Heme oxygenase (HO)-1 in adipocytes protects against obesity and adipose dysfunction. In this study, we report the identification of butein, a flavonoid chalcone, as a novel inducer of HO-1 expression in adipocytes in vitro and in vivo. Butein upregulated HO-1 mRNA and protein expression in 3T3-L1 adipocytes, accompanied by Kelch-Like ECH-Associated Protein (Keap) 1 degradation and increase in the nuclear level of nuclear factor erythroid 2-related factor 2 (Nrf2). Butein modulation of Keap1 and Nrf2 as well as HO-1 upregulation was reversed by pretreatment with p38 MAPK inhibitor SB203580, indicating the involvement of p38 MAPK in butein activation of Nrf2 in adipocytes. In addition, HO-1 activation by butein led to the inhibitions of reactive oxygen species and adipocyte differentiation, as evidenced by the fact that butein repression of reactive oxygen species and adipogenesis was reversed by pretreatment with HO-1 inhibitor SnPP. Induction of HO-1 expression by butein was also demonstrated in the adipose tissue of C57BL/6 mice fed a high-fat diet administered along with butein for three weeks, and correlated with the inhibitions of adiposity and adipose tissue inflammation, which were reversed by co-administration of SnPP. Altogether, our results demonstrate that butein activates the p38 MAPK/Nrf2/HO-1 pathway to act as a potent inhibitor of adipose hypertrophy and inflammation in a diet-induced obesity model and thus has potential for suppressing obesity-linked metabolic syndrome. Copyright © 2017 Elsevier B.V. All rights reserved.

The effect of natural antioxidants in cyclophosphamide-induced hepatotoxicity: Role of Nrf2/HO-1 pathway.

PubMed

Sherif, Iman O

2018-05-22

Hepatotoxicity induced by cyclophosphamide (Cyclo) is a major concern in clinical practice. This study was designed to investigate the possible cytoprotective effect of natural antioxidants as oleuropein and quercetin against Cyclo induced hepatotoxicity via the nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) signaling pathway. Male Wistar rats were randomly divided into six groups and treated for 10 days as follow: Group I (Normal control) received saline, group II (Oleu control): received orally oleuropein 30 mg/kg/day, group III (Quer control): administered orally quercetin 50 mg/kg/day, group IV (Cyclo): received saline and injected with single intraperitoneal (i.p) dose of Cyclo 200 mg/kg at day 5, group V (Oleu ttt): treated with oleuropein plus Cyclo i.p. injection at day 5, and group VI (Quer ttt): treated with quercetin plus Cyclo i.p. injection at day 5. Injection of Cyclo showed marked increase in serum transaminases and alkaline phosphatase, hepatic malondialdehyde (MDA) and tumor necrosis factor-alpha (TNF-⍺) levels along with significant reduction in hepatic reduced glutathione (GSH), superoxide dismutase (SOD), and catalase levels in addition to downregulation of hepatic Nrf2 and HO-1 expressions and reduction in hepatic nuclear Nrf2 binding activity when compared with normal group. Histopathological examination of Cyclo treated rats revealed hepatic damage. Both oleuropein and quercetin exhibited an improvement in the biochemical and histopathological findings. In conclusion, the natural antioxidants oleuropein and quercetin counteract the Cyclo induced hepatotoxicity through activation of Nrf2/HO-1 signaling pathway with subsequent suppression of oxidative stress and inflammation. Copyright © 2018 Elsevier B.V. All rights reserved.

Minocycline attenuates sevoflurane-induced cell injury via activation of Nrf2.

PubMed

Tian, Yue; Wu, Xiuying; Guo, Shanbin; Ma, Ling; Huang, Wei; Zhao, Xiaochun

2017-04-01

Minocycline has been demonstrated to exert neuroprotective effects in various experimental models. In the present study, we investigated the mechanisms underlying the protective effects of minocycline on cell injury induced by the inhalation of the anesthetic, sevoflurane. In our in vivo experiments using rats, minocycline attenuated sevoflurane-induced neuronal degeneration and apoptosis in the rat hippocampus, and this effect was associated with the minocycline-mediated suppression of oxidative stress in the hippocampus. In in vitro experiments, minocycline inhibited sevoflurane-induced apoptosis and the production of reactive oxygen species (ROS) in H4 human neuroglioma cells. In addition, minocycline suppressed the sevoflurane-induced upregulation of interleukin (IL)-6 and the activation of the nuclear factor-κB (NF-κB) signaling pathway in H4 cells. Furthermore, we found that nuclear factor E2-related factor 2 (Nrf2), an activator of the stress response, was upregulated and activated upon sevoflurane treatment both in the rat hippocampus and in H4 cells. In addition, minocycline further augmented the upregulation and activation of Nrf2 when used in conjunction with sevoflurane. Moreover, the knockdown of Nrf2 in H4 cells by small interfering RNA (siRNA) diminished the cytoprotective effect of minocycline, and attenuated the inhibitory effect of minocycline on ROS production, IL-6 upregulation and the activation of the NF-κB signaling pathway. On the whole, our findings indicate that minocycline may exert protective effects against sevoflurane-induced cell injury via the Nrf2-modulated antioxidant response and the inhibition of the activation of the NF-κB signaling pathway.

RAGE-Specific Inhibitor FPS-ZM1 Attenuates AGEs-Induced Neuroinflammation and Oxidative Stress in Rat Primary Microglia.

PubMed

Shen, Chao; Ma, Yingjuan; Zeng, Ziling; Yin, Qingqing; Hong, Yan; Hou, Xunyao; Liu, Xueping

2017-10-01

Advanced glycation end products (AGEs) enhance microglial activation and intensify the inflammatory response and oxidative stress in the brain. This process may occur due to direct cytotoxicity or interacting with AGEs receptors (RAGE), which are expressed on the surface of microglia. FPS-ZM1 is a high-affinity but nontoxic RAGE-specific inhibitor that has been recently shown to attenuate the Aβ-induced inflammatory response by blocking the ligation of Aβ to RAGE. In this study, we further investigated the effect of FPS-ZM1 on the AGEs/RAGE interaction and downstream elevation of neuroinflammation and oxidative stress in primary microglia cells. The results suggested that FPS-ZM1 significantly suppressed AGEs-induced RAGE overexpression, RAGE-dependent microglial activation, nuclear translocation of nuclear factor kappaB p65 (NF-κB p65), and the expression of downstream inflammatory mediators such as tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), cyclooxygenase 2 (COX-2)/prostaglandin E2 (PGE2) and inducible nitric oxide synthase (iNOS)/nitric oxide (NO). Furthermore, FPS-ZM1 attenuated AGEs-stimulated NADPH oxidase (NOX) activation and reactive oxygen species (ROS) expression. Finally, FPS-ZM1 elevated the levels of transcription factors nuclear-factor (erythroid-derived 2)-like 2 (Nrf2) and heme oxygenase-1 (HO-1), as well as decreased antioxidant capacity and increased production of oxidative species. Our results suggest that FPS-ZM1 may be neuroprotective through attenuating microglial activation, oxidative stress and inflammation by blocking RAGE.

Cyclo(His-Pro) promotes cytoprotection by activating Nrf2-mediated up-regulation of antioxidant defence

PubMed Central

Minelli, Alba; Conte, Carmela; Grottelli, Silvia; Bellezza, Maria; Cacciatore, Ivana; Bolaños, Juan P

2009-01-01

Hystidyl-proline [cyclo(His-Pro)] is an endogenous cyclic dipeptide produced by the cleavage of thyrotropin releasing hormone. Previous studies have shown that cyclo(His-Pro) protects against oxidative stress, although the underlying mechanism has remained elusive. Here, we addressed this issue and found that cyclo(His-Pro) triggered nuclear accumulation of NF-E2-related factor-2 (Nrf2), a transcription factor that up-regulates antioxidant-/electrophile-responsive element (ARE-EpRE)-related genes, in PC12 cells. Cyclo(His-Pro) attenuated reactive oxygen species production, and prevented glutathione depletion caused by glutamate, rotenone, paraquat and β-amyloid treatment. Moreover, real-time PCR analyses revealed that cyclo(His-Pro) induced the expression of a number of ARE-related genes and protected cells against hydrogen peroxide-mediated apoptotic death. Furthermore, these effects were abolished by RNA interference-mediated Nrf2 knockdown. Finally, pharmacological inhibition of p-38 MAPK partially prevented both cyclo(His-Pro)-mediated Nrf2 activation and cellular protection. These results suggest that the signalling mechanism responsible for the cytoprotective actions of cyclo(His-Pro) would involve p-38 MAPK activation leading to Nrf2-mediated up-regulation of antioxidant cellular defence. PMID:18373731

Recent Updates on Acetaminophen Hepatotoxicity: The Role of Nrf2 in Hepatoprotection

PubMed Central

Gum, Sang Il

2013-01-01

Acetaminophen (APAP) known as paracetamol is the main ingredient in Tylenol, which has analgesic and anti-pyretic properties. Inappropriate use of APAP causes major morbidity and mortality secondary to hepatic failure. Overdose of APAP depletes the hepatic glutathione (GSH) rapidly, and the metabolic intermediate leads to hepatocellular death. This article reviews the mechanisms of hepatotoxicity and provides an overview of current research studies. Pharmacokinetics including metabolism (activation and detoxification), subsequent transport (efflux)-facilitating excretion, and some other aspects related to toxicity are discussed. Nuclear factor erythroid 2-related factor 2 (Nrf2)-regulated gene battery plays a critical role in the multiple steps associated with the mitigation of APAP toxicity. The role of Nrf2 as a protective target is described, and potential natural products inhibiting APAP toxicity are outlined. This review provides an update on the mechanism of APAP toxicity and highlights the beneficial role of Nrf2 and specific natural products in hepatoprotection. PMID:24386516

Subcellular localization and cytoplasmic complex status of endogenous Keap1.

PubMed

Watai, Yoriko; Kobayashi, Akira; Nagase, Hiroko; Mizukami, Mio; McEvoy, Justina; Singer, Jeffrey D; Itoh, Ken; Yamamoto, Masayuki

2007-10-01

Keap1 acts as a sensor for oxidative/electrophilic stress, an adaptor for Cullin-3-based ubiquitin ligase, and a regulator of Nrf2 activity through the interaction with Nrf2 Neh2 domain. However, the mechanism(s) of Nrf2 migration into the nucleus in response to stress remains largely unknown due to the lack of a reliable antibody for the detection of endogenous Keap1 molecule. Here, we report the generation of a new monoclonal antibody for the detection of endogenous Keap1 molecules. Immunocytochemical analysis of mouse embryonic fibroblasts with the antibody revealed that under normal, unstressed condition, Keap1 is localized primarily in the cytoplasm with minimal amount in the nucleus and endoplasmic reticulum. This subcellular localization profile of Keap1 appears unchanged after treatment of cells with diethyl maleate, an electrophile, and/or Leptomycin B, a nuclear export inhibitor. Subcellular fractionation analysis of mouse liver cells showed similar results. No substantial change in the subcellular distribution profile could be observed in cells isolated from butylated hydroxyanisole-treated mice. Analyses of sucrose density gradient centrifugation of mouse liver cells indicated that Keap1 appears to form multiprotein complexes in the cytoplasm. These results demonstrate that endogenous Keap1 remains mostly in the cytoplasm, and electrophiles promote nuclear accumulation of Nrf2 without altering the subcellular localization of Keap1.

NRF2 Mediates Neuroblastoma Proliferation and Resistance to Retinoic Acid Cytotoxicity in a Model of In Vitro Neuronal Differentiation.

PubMed

de Miranda Ramos, Vitor; Zanotto-Filho, Alfeu; de Bittencourt Pasquali, Matheus Augusto; Klafke, Karina; Gasparotto, Juciano; Dunkley, Peter; Gelain, Daniel Pens; Moreira, José Cláudio Fonseca

2016-11-01

Retinoic acid (RA) morphogenetic properties have been used in different kinds of therapies, from neurodegenerative disorders to some types of cancer such as promyelocytic leukemia and neuroblastoma. However, most of the pathways responsible for RA effects remain unknown. To investigate such pathways, we used a RA-induced differentiation model in the human neuroblastoma cells, SH-SY5Y. Our data showed that n-acetyl-cysteine (NAC) reduced cells' proliferation rate and increased cells' sensitivity to RA toxicity. Simultaneously, NAC pre-incubation attenuated nuclear factor erythroid 2-like factor 2 (NRF2) activation by RA. None of these effects were obtained with Trolox ® as antioxidant, suggesting a cysteine signalization by RA. NRF2 knockdown increased cell sensibility to RA after 96 h of treatment and diminished neuroblastoma proliferation rate. Conversely, NRF2 overexpression limited RA anti-proliferative effects and increased cell proliferation. In addition, a rapid and non-genomic activation of the ERK 1/2 and PI3K/AKT pathways revealed to be equally required to promote NRF2 activation and necessary for RA-induced differentiation. Together, we provide data correlating NRF2 activity with neuroblastoma proliferation and resistance to RA treatments; thus, this pathway could be a potential target to optimize neuroblastoma chemotherapeutic response as well as in vitro neuronal differentiation protocols.

Prediction of Binding Energy of Keap1 Interaction Motifs in the Nrf2 Antioxidant Pathway and Design of Potential High-Affinity Peptides.

PubMed

Karttunen, Mikko; Choy, Wing-Yiu; Cino, Elio A

2018-06-07

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcription factor and principal regulator of the antioxidant pathway. The Kelch domain of Kelch-like ECH-associated protein 1 (Keap1) binds to motifs in the N-terminal region of Nrf2, promoting its degradation. There is interest in developing ligands that can compete with Nrf2 for binding to Kelch, thereby activating its transcriptional activities and increasing antioxidant levels. Using experimental Δ G bind values of Kelch-binding motifs determined previously, a revised hydrophobicity-based model was developed for estimating Δ G bind from amino acid sequence and applied to rank potential uncharacterized Kelch-binding motifs identified from interaction databases and BLAST searches. Model predictions and molecular dynamics (MD) simulations suggested that full-length MAD2A binds Kelch more favorably than a high-affinity 20-mer Nrf2 E78P peptide, but that the motif in isolation is not a particularly strong binder. Endeavoring to develop shorter peptides for activating Nrf2, new designs were created based on the E78P peptide, some of which showed considerable propensity to form binding-competent structures in MD, and were predicted to interact with Kelch more favorably than the E78P peptide. The peptides could be promising new ligands for enhancing the oxidative stress response.

PAK1 translocates into nucleus in response to prolactin but not to estrogen

DOE Office of Scientific and Technical Information (OSTI.GOV)

Oladimeji, Peter, E-mail: Peter.Oladimeji@rockets.utoledo.edu; Diakonova, Maria, E-mail: mdiakon@utnet.utoledo.edu

2016-04-22

Tyrosyl phosphorylation of the p21-activated serine–threonine kinase 1 (PAK1) has an essential role in regulating PAK1 functions in breast cancer cells. We previously demonstrated that PAK1 serves as a common node for estrogen (E2)- and prolactin (PRL)-dependent pathways. We hypothesize herein that intracellular localization of PAK1 is affected by PRL and E2 treatments differently. We demonstrate by immunocytochemical analysis that PAK1 nuclear translocation is ligand-dependent: only PRL but not E2 stimulated PAK1 nuclear translocation. Tyrosyl phosphorylation of PAK1 is essential for this nuclear translocation because phospho-tyrosyl-deficient PAK1 Y3F mutant is retained in the cytoplasm in response to PRL. We confirmedmore » these data by Western blot analysis of subcellular fractions. In 30 min of PRL treatment, only 48% of pTyr-PAK1 is retained in the cytoplasm of PAK1 WT clone while 52% re-distributes into the nucleus and pTyr-PAK1 shuttles back to the cytoplasm by 60 min of PRL treatment. In contrast, PAK1 Y3F is retained in the cytoplasm. E2 treatment causes nuclear translocation of neither PAK1 WT nor PAK1 Y3F. Finally, we show by an in vitro kinase assay that PRL but not E2 stimulates PAK1 kinase activity in the nuclear fraction. Thus, PAK1 nuclear translocation is ligand-dependent: PRL activates PAK1 and induces translocation of activated pTyr-PAK1 into nucleus while E2 activates pTyr-PAK1 only in the cytoplasm. - Highlights: • Prolactin but not estrogen causes translocation of PAK1 into nucleus. • Tyrosyl phosphorylation of PAK1 is required for nuclear localization. • Prolactin but not estrogen stimulates PAK1 kinase activity in nucleus.« less

The trypanocidal benznidazole promotes adaptive response to oxidative injury: Involvement of the nuclear factor-erythroid 2-related factor-2 (Nrf2) and multidrug resistance associated protein 2 (MRP2)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rigalli, Juan Pablo

Oxidative stress is a frequent cause underlying drug-induced hepatotoxicity. Benznidazole (BZL) is the only trypanocidal agent available for treatment of Chagas disease in endemic areas. Its use is associated with side effects, including increases in biomarkers of hepatotoxicity. However, BZL potential to cause oxidative stress has been poorly investigated. Here, we evaluated the effect of a pharmacologically relevant BZL concentration (200 μM) at different time points on redox status and the counteracting mechanisms in the human hepatic cell line HepG2. BZL increased reactive oxygen species (ROS) after 1 and 3 h of exposure, returning to normality at 24 h. Additionally,more » BZL increased glutathione peroxidase activity at 12 h and the oxidized glutathione/total glutathione (GSSG/GSSG + GSH) ratio that reached a peak at 24 h. Thus, an enhanced detoxification of peroxide and GSSG formation could account for ROS normalization. GSSG/GSSG + GSH returned to control values at 48 h. Expression of the multidrug resistance-associated protein 2 (MRP2) and GSSG efflux via MRP2 were induced by BZL at 24 and 48 h, explaining normalization of GSSG/GSSG + GSH. BZL activated the nuclear erythroid 2-related factor 2 (Nrf2), already shown to modulate MRP2 expression in response to oxidative stress. Nrf2 participation was confirmed using Nrf2-knockout mice in which MRP2 mRNA expression was not affected by BZL. In summary, we demonstrated a ROS increase by BZL in HepG2 cells and a glutathione peroxidase- and MRP2 driven counteracting mechanism, being Nrf2 a key modulator of this response. Our results could explain hepatic alterations associated with BZL therapy. - Highlights: • BZL triggers a redox imbalance in the human hepatic cell line HepG2. • Concomitantly BZL triggers compensatory mechanisms to alleviate the redox injury. • Response mechanisms comprise an enhanced glutathione peroxidase and MRP2 activity. • Transcription factor Nrf2 plays a key role orchestrating compensatory mechanisms.« less