scholarly journals Curcumin Protects Human Keratinocytes against Inorganic Arsenite-Induced Acute Cytotoxicity through an NRF2-Dependent Mechanism

2013 ◽  
Vol 2013 ◽  
pp. 1-11 ◽  
Author(s):  
Rui Zhao ◽  
Bei Yang ◽  
Linlin Wang ◽  
Peng Xue ◽  
Baocheng Deng ◽  
...  
Keyword(s):  
Inorganic Arsenic ◽  
Human Keratinocytes ◽  
Antioxidant Response ◽  
Nuclear Accumulation ◽  
Related Factor ◽  
High Concentration ◽  
Low Concentrations ◽  
Acute Cytotoxicity ◽  
Nrf2 Protein ◽  
Dependent Mechanism

Human exposure to inorganic arsenic leads to various dermal disorders, including hyperkeratosis and skin cancer. Curcumin is demonstrated to induce remarkable antioxidant activity in a variety of cells and tissues. The present study aimed at identifying curcumin as a potent activator of nuclear factor erythroid 2-related factor 2 (NRF2) and demonstrating its protective effect against inorganic arsenite- (iAs3+-) induced cytotoxicity in human keratinocytes. We found that curcumin led to nuclear accumulation of NRF2 protein and increased the expression of antioxidant response element- (ARE-) regulated genes in HaCaT keratinocytes in concentration- and time-dependent manners. High concentration of curcumin (20 μM) also increased protein expression of long isoforms of NRF1. Treatment with low concentrations of curcumin (2.5 or 5 μM) effectively increased the viability and survival of HaCaT cells against iAs3+-induced cytotoxicity as assessed by the MTT assay and flow cytometry and also attenuated iAs3+-induced expression of cleaved caspase-3 and cleaved PARP protein. Selective knockdown ofNRF2orKEAP1by lentiviral shRNAs significantly diminished the cytoprotection conferred by curcumin, suggesting that the protection against iAs3+-induced cytotoxicity is dependent on the activation of NRF2. Our results provided a proof of the concept of using curcumin to activate the NRF2 pathway to alleviate arsenic-induced dermal damage.

scholarly journals Activation of the Nrf2 Pathway by Inorganic Arsenic in Human Hepatocytes and the Role of Transcriptional Repressor Bach1

2013 ◽  
Vol 2013 ◽  
pp. 1-10 ◽  
Author(s):  
Dan Liu ◽  
Xiaoxu Duan ◽  
Dandan Dong ◽  
Caijun Bai ◽  
Xin Li ◽  
...  
Keyword(s):  
Biological Effects ◽  
Transcriptional Repressor ◽  
Inorganic Arsenic ◽  
Cell Types ◽  
Antioxidant Response ◽  
Human Hepatocytes ◽  
Heme Oxygenase 1 ◽  
Related Factor ◽  
Nrf2 Pathway ◽  
Nrf2 Protein

Previous studies have proved that the environmental toxicant, inorganic arsenic, activates nuclear factor erythroid 2-related factor 2 (Nrf2) pathway in many different cell types. This study tried to explore the hepatic Nrf2 pathway upon arsenic treatment comprehensively, since liver is one of the major target organs of arsenical toxicity. Our results showed that inorganic arsenic significantly induced Nrf2 protein and mRNA expression in Chang human hepatocytes. We also observed a dose-dependent increase of antioxidant response element- (ARE-) luciferase activity. Both the mRNA and protein levels of NAD(P)H:quinone oxidoreductase 1 (NQO1) and heme oxygenase-1 (HO-1) were all upregulated dramatically. On the other hand, entry and accumulation of Nrf2 protein in the nucleus, while exportting the transcriptional repressor BTB and CNC homology 1 (Bach1) from nucleus to cytoplasm, were also confirmed by western blot and immunofluorescence assay. Our results therefore confirmed the arsenic-induced Nrf2 pathway activation in hepatocytes and also suggested that the translocation of Bach1 was associated with the regulation of Nrf2 pathway by arsenic. Hepatic Nrf2 pathway plays indispensable roles for cellular defenses against arsenic hepatotoxicity, and the interplay of Bach1 and Nrf2 may be helpful to understand the self-defensive responses and the diverse biological effects of arsenicals.

scholarly journals Overexpression of NRF1-742 or NRF1-772 Reduces Arsenic-Induced Cytotoxicity and Apoptosis in Human HaCaT Keratinocytes

2020 ◽  
Vol 21 (6) ◽  
pp. 2014
Author(s):  
Shuai Wang ◽  
Hao Cheng ◽  
Linlin Wang ◽  
Rui Zhao ◽  
Dawei Guan
Keyword(s):  
Leucine Zipper ◽  
Nuclear Translocation ◽  
Inorganic Arsenic ◽  
Human Keratinocytes ◽  
Antioxidant Response ◽  
Bzip Transcription Factor ◽  
Nuclear Factor ◽  
Hacat Keratinocytes ◽  
Protein Levels ◽  
Antioxidant Protein

Increasing evidence indicates that human exposure to inorganic arsenic causes cutaneous diseases and skin cancers. Nuclear factor erythroid 2-like 1 (NRF1) belongs to the cap “n” collar (CNC) basic-region leucine zipper (bZIP) transcription factor family and regulates antioxidant response element (ARE) genes. The human NRF1 gene is transcribed into multiple isoforms, which contain 584, 616, 742, 761, or 772 amino acids. We previously demonstrated that the long isoforms of NRF1 (i.e., NRF1-742, NRF1-761 and NRF1-772) are involved in the protection of human keratinocytes from acute arsenic cytotoxicity by enhancing the cellular antioxidant response. The aim of the current study was to investigate the roles of NRF1-742 and NRF1-772 in the arsenic-induced antioxidant response and cytotoxicity. We found that overexpression of NRF1-742 or NRF1-772 in human HaCaT keratinocytes decreased susceptibility to arsenic-induced apoptosis and cytotoxicity. In addition, we characterized the different protein bands observed for NRF1-742 and NRF1-772 by western blotting. The posttranslational modifications and nuclear translocation of these isoforms differed and were partially affected by arsenic exposure. Antioxidant protein levels were increased in the NRF1-742 and NRF1-772-overexpressing cell lines. The upregulation of antioxidant protein levels was partly due to the translation of nuclear factor erythroid 2-like 2 (NRF2) and its increased nuclear transport. Overall, overexpression of NRF1-742 and NRF1-772 protected HaCaT cells from arsenic-induced cytotoxicity, mainly through translational modifications and the promotion of antioxidant gene expression.

scholarly journals Gain-of-function genetic screen of the kinome reveals BRSK2 as an inhibitor of the NRF2 transcription factor

2020 ◽  
Vol 133 (14) ◽  
pp. jcs241356 ◽  
Author(s):  
Tigist Y. Tamir ◽  
Brittany M. Bowman ◽  
Megan J. Agajanian ◽  
Dennis Goldfarb ◽  
Travis P. Schrank ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Protein Kinase ◽  
Human Cancer ◽  
Antioxidant Response ◽  
Genetic Screen ◽  
Related Factor ◽  
Dependent Manner ◽  
Gain Of Function ◽  
Protein Levels ◽  
Nrf2 Protein

ABSTRACTNuclear factor erythroid 2-related factor 2 (NFE2L2, also known as NRF2) is a transcription factor and master regulator of cellular antioxidant response. Aberrantly high NRF2-dependent transcription is recurrent in human cancer, but conversely NRF2 activity diminishes with age and in neurodegenerative and metabolic disorders. Although NRF2-activating drugs are clinically beneficial, NRF2 inhibitors do not yet exist. Here, we describe use of a gain-of-function genetic screen of the kinome to identify new druggable regulators of NRF2 signaling. We found that the under-studied protein kinase brain-specific kinase 2 (BRSK2) and the related BRSK1 kinases suppress NRF2-dependent transcription and NRF2 protein levels in an activity-dependent manner. Integrated phosphoproteomics and RNAseq studies revealed that BRSK2 drives 5′-AMP-activated protein kinase α2 (AMPK) signaling and suppresses the mTOR pathway. As a result, BRSK2 kinase activation suppresses ribosome-RNA complexes, global protein synthesis and NRF2 protein levels. Collectively, our data illuminate the BRSK2 and BRSK1 kinases, in part by functionally connecting them to NRF2 signaling and mTOR. This signaling axis might prove useful for therapeutically targeting NRF2 in human disease.This article has an associated First Person interview with the first author of the paper.

scholarly journals AMPK Facilitates Nuclear Accumulation of Nrf2 by Phosphorylating at Serine 550

2016 ◽  
Vol 36 (14) ◽  
pp. 1931-1942 ◽  
Author(s):  
Min Sung Joo ◽  
Won Dong Kim ◽  
Ki Young Lee ◽  
Ji Hyun Kim ◽  
Ja Hyun Koo ◽  
...  
Keyword(s):  
Cell Survival ◽  
Nuclear Export ◽  
Glycogen Synthase ◽  
Nuclear Export Signal ◽  
Direct Interaction ◽  
Antioxidant Response ◽  
Nuclear Accumulation ◽  
Related Factor ◽  
Ampk Activation

Nrf2 (nuclear factor erythroid 2-related factor 2) is an antioxidant transcription factor. AMP-activated protein kinase (AMPK) functions as a central regulator of cell survival in response to stressful stimuli. Nrf2 should be coordinated with the cell survival pathway controlled by AMPK, but so far the mechanistic connections remain undefined. This study investigated the role of AMPK in Nrf2 trafficking and its activity regulation. A subnetwork integrating neighbor molecules suggested direct interaction between AMPK and Nrf2. In cells, AMPK activation caused nuclear accumulation of Nrf2. In thein vitrokinase and peptide competition assays, AMPK phosphorylated Nrf2 at the Ser558 residue (Ser550 in mouse) located in the canonical nuclear export signal. Nrf2 with an S550A mutation failed to be accumulated in the nucleus after AMPK activation. Leptomycin B, a nuclear export inhibitor, did not enhance nuclear accumulation of wild-type Nrf2 (WT-Nrf2) activated by AMPK or a phospho-Ser550-mimetic Nrf2 mutant, corroborating the finding that AMPK facilitated nuclear accumulation of Nrf2, probably by inhibiting nuclear export. Activated glycogen synthase kinase 3β (GSK3β) diminished the basal nuclear level of Myc-S550A-Nrf2. Taking the data collectively, AMPK phosphorylates Nrf2 at the Ser550 residue, which, in conjunction with AMPK-mediated GSK3β inhibition, promotes nuclear accumulation of Nrf2 for antioxidant response element (ARE)-driven gene transactivation.

scholarly journals Investigating polystyrene nanoplastics-induced reproductive toxicity in vitro: Focus on Nrf2-PKM2-autophagy signaling pathway

2021 ◽  
Author(s):  
Siwen Li ◽  
Yu Ma ◽  
Shuzi Ye ◽  
Ying Su ◽  
Die Hu ◽  
...  
Keyword(s):  
Reproductive Toxicity ◽  
Environmental Pollutant ◽  
Nuclear Accumulation ◽  
Seminiferous Tubules ◽  
Related Factor ◽  
Mitochondrial Apoptosis ◽  
Inverted Microscope ◽  
Toxicity In Vitro ◽  
Nrf2 Protein

Abstract As an issue of widespread concern, microplastics pollution has emerged as a harmful environmental pollutant. Nanoplastics (NaPs) has reported to accumulate in the testes and cause degeneration in the seminiferous tubules. However, the current research involving NaPs-induced reproductive toxicity remains poorly understood. The current work aimed to investigate the mechanisms of NaPs-induced reproductive injury in vitro. At first, we found that 80 nm fluorescent NaPs could enter into GC-2spd(ts) cells by fluorescent inverted microscope. Our results also demonstrated that suppression of reactive oxygen species (ROS) inhibited NaPs-triggered mitochondrial apoptosis and autophagy in GC-2spd(ts) cells. We also found that NaPs treatment did not change the interaction between nuclear factor erythroid-derived 2-related factor (Nrf2) and Kelch-like ECH associated protein 1 (Keap1), while inhibiting nuclear accumulation of Nrf2 protein. Further in vitro experiments showed that NaPs-induced reproductive toxicity associated with reducing dimerize pyruvate kinase M2 (PKM2), which are ascribed to the loss of Nrf2. Meanwhile, improving nuclear accumulation of Nrf2 might interact with PKM2 to rescue mitochondrial apoptosis caused by NaPs. Together, this study highlight that disturbing Nrf2-PKM2 signaling is essential process of NaPs-induced reproductive toxicity and provide valuable insights into the mechanism of microplastics-induced reproductive toxicity.

scholarly journals DCAF1-targeting microRNA-3175 activates Nrf2 signaling and inhibits dexamethasone-induced oxidative injury in human osteoblasts

2021 ◽  
Vol 12 (11) ◽  
Author(s):  
Jing Chen ◽  
Jin-qian Liang ◽  
Yun-Fang Zhen ◽  
Lei Chang ◽  
Zhen-tao Zhou ◽  
...  
Keyword(s):  
Oxidative Injury ◽  
Antioxidant Response ◽  
Protein Stabilization ◽  
Related Factor ◽  
Activity Increase ◽  
Human Osteoblasts ◽  
Argonaute 2 ◽  
Ubiquitin E3 Ligase ◽  
Rna Immunoprecipitation ◽  
Nrf2 Protein

AbstractActivation of nuclear-factor-E2-related factor 2 (Nrf2) signaling can protect human osteoblasts from dexamethasone-induced oxidative injury. DDB1 and CUL4 associated factor 1 (DCAF1) is a novel ubiquitin E3 ligase for Nrf2 protein degradation. We identified a novel DCAF1-targeting miRNA, miR-3175. RNA pull-down, Argonaute 2 RNA-immunoprecipitation, and RNA fluorescent in situ hybridization results confirmed a direct binding between miR-3175 and DCAF1 mRNA in primary human osteoblasts. DCAF1 3′-untranslated region luciferase activity and its expression were significantly decreased after miR-3175 overexpression but were augmented with miR-3175 inhibition in human osteoblasts and hFOB1.19 osteoblastic cells. miR-3175 overexpression activated Nrf2 signaling, causing Nrf2 protein stabilization, antioxidant response (ARE) activity increase, and transcription activation of Nrf2-dependent genes in human osteoblasts and hFOB1.19 cells. Furthermore, dexamethasone-induced oxidative injury and apoptosis were largely attenuated by miR-3175 overexpression in human osteoblasts and hFOB1.19 cells. Importantly, shRNA-induced silencing or CRISPR/Cas9-mediated Nrf2 knockout abolished miR-3175 overexpression-induced osteoblast cytoprotection against dexamethasone. Conversely, DFAC1 knockout, by the CRISPR/Cas9 method, activated the Nrf2 cascade and inhibited dexamethasone-induced cytotoxicity in hFOB1.19 cells. Importantly, miR-3175 expression was decreased in necrotic femoral head tissues of dexamethasone-taking patients, where DCAF1 mRNA was upregulated. Together, silencing DCAF1 by miR-3175 activated Nrf2 signaling to inhibit dexamethasone-induced oxidative injury and apoptosis in human osteoblasts.

scholarly journals Liver macrophages inhibit the endogenous antioxidant response in obesity-associated insulin resistance

2020 ◽  
Vol 12 (532) ◽  
pp. eaaw9709 ◽  
Author(s):  
Valerio Azzimato ◽  
Jennifer Jager ◽  
Ping Chen ◽  
Cecilia Morgantini ◽  
Laura Levi ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Insulin Resistance ◽  
Liver Disease ◽  
Lipid Accumulation ◽  
Therapeutic Potential ◽  
Antioxidant Response ◽  
Related Factor ◽  
Liver Macrophages ◽  
Endogenous Antioxidant ◽  
Nrf2 Protein

Obesity and insulin resistance are risk factors for nonalcoholic fatty liver disease (NAFLD), the most common chronic liver disease worldwide. Because no approved medication nor an accurate and noninvasive diagnosis is currently available for NAFLD, there is a clear need to better understand the link between obesity and NAFLD. Lipid accumulation during obesity is known to be associated with oxidative stress and inflammatory activation of liver macrophages (LMs). However, we show that although LMs do not become proinflammatory during obesity, they display signs of oxidative stress. In livers of both humans and mice, antioxidant nuclear factor erythroid 2–related factor 2 (NRF2) was down-regulated with obesity and insulin resistance, yielding an impaired response to lipid accumulation. At the molecular level, a microRNA-targeting NRF2 protein, miR-144, was elevated in the livers of obese insulin-resistant humans and mice, and specific silencing of miR-144 in murine and human LMs was sufficient to restore NRF2 protein expression and the antioxidant response. These results highlight the pathological role of LMs and their therapeutic potential to restore the impaired endogenous antioxidant response in obesity-associated NAFLD.

scholarly journals Role of Nrf2 in the regulation of the Mrp2 (ABCC2) gene

2006 ◽  
Vol 395 (3) ◽  
pp. 599-609 ◽  
Author(s):  
Valeska Vollrath ◽  
Ana M. Wielandt ◽  
Mirentxu Iruretagoyena ◽  
Jose Chianale
Keyword(s):  
Gene Expression ◽  
Transcriptional Activation ◽  
Protein Complexes ◽  
Antioxidant Response ◽  
Proximal Region ◽  
Common Mechanism ◽  
Related Factor ◽  
Key Enzyme ◽  
Chloramphenicol Acetyltransferase Gene ◽  
Nrf2 Protein

The Nrf2 (nuclear factor-erythroid 2 p45-related factor 2) transcription factor regulates gene expression of the GCLC (glutamate–cysteine ligase catalytic subunit), which is a key enzyme in glutathione synthesis, and GSTs (glutathione S-transferases) via the ARE (antioxidant-response element). The Mrp2 (multidrug-resistance protein 2) pump mediates the excretion of GSH and GSSG excretion as well as endo- and xeno-biotics that are conjugated with GSH, glucuronate or sulphate. Considering that Mrp2 acts synergistically with these enzymes, we hypothesized that the regulation of Mrp2 gene expression is also dependent on Nrf2. Using BHA (butylated hydroxyanisole), which is a classical activator of the ARE–Nrf2 pathway, we observed an increase in the transcriptional activity of Mrp2, GCLC and Gsta1/Gsta2 genes in the mouse liver. A similar pattern of co-induction of Mrp2 and GCLC genes was also observed in mouse (Hepa 1-6) and human (HepG2) hepatoma cells treated with BHA, β-NF (β-naphthoflavone), 2,4,5-T (trichlorophenoxyacetic acid) or 2AAF (2-acetylaminofluorene), suggesting that these genes share common mechanism(s) of transcriptional activation in response to exposure to xenobiotics. To define the mechanism of Mrp2 gene induction, the 5′-flanking region of the mouse Mrp2 gene (2.0 kb) was isolated, and two ARE-like sequences were found: ARE-2 (−1391 to −1381) and ARE-1 (−95 to −85). Deletion analyses demonstrated that the proximal region (−185 to +99) contains the elements for the basal expression and xenobiotic-mediated induction of the Mrp2 gene. Gel-shift and supershift assays indicated that Nrf2–protein complexes bind ARE sequences of the Mrp2 promoter, preferentially to the ARE-1 sequence. Overexpression of Nrf2 increased ARE-1-mediated CAT (chloramphenicol acetyltransferase) gene activity, while overexpression of mutant Nrf2 protein repressed the activity. Thus Nrf2 appears to regulate Mrp2 gene expression via an ARE element located at the proximal region of its promoter in response to exposure to xenobiotics.

scholarly journals Delivery of Cinnamic Aldehyde Antioxidant Response Activating nanoParticles (ARAPas) for Vascular Applications

Antioxidants ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 709
Author(s):  
Ana E. Cartaya ◽  
Halle Lutz ◽  
Sophie Maiocchi ◽  
Morgan Nalesnik ◽  
Edward M. Bahnson
Keyword(s):  
Ex Vivo ◽  
Antioxidant Response ◽  
Related Factor ◽  
Cinnamic Aldehyde ◽  
Vsmc Proliferation ◽  
Nrf2 Activation ◽  
Selective Delivery ◽  
And Migration ◽  
Inner Vessel ◽  
Macrophage Uptake

Selective delivery of nuclear factor erythroid 2-related factor 2 (Nrf2) activators to the injured vasculature at the time of vascular surgical intervention has the potential to attenuate oxidative stress and decrease vascular smooth muscle cell (VSMC) hyperproliferation and migration towards the inner vessel wall. To this end, we developed a nanoformulation of cinnamic aldehyde (CA), termed Antioxidant Response Activating nanoParticles (ARAPas), that can be readily loaded into macrophages ex vivo. The CA-ARAPas-macrophage system was used to study the effects of CA on VSMC in culture. CA was encapsulated into a pluronic micelle that was readily loaded into both murine and human macrophages. CA-ARAPas inhibits VSMC proliferation and migration, and activates Nrf2. Macrophage-mediated transfer of CA-ARAPas to VSMC is evident after 12 h, and Nrf2 activation is apparent after 24 h. This is the first report, to the best of our knowledge, of CA encapsulation in pluronic micelles for macrophage-mediated delivery studies. The results of this study highlight the feasibility of CA encapsulation and subsequent macrophage uptake for delivery of cargo into other pertinent cells, such as VSMC.

scholarly journals Ethosomes and Transethosomes for Mangiferin Transdermal Delivery

Antioxidants ◽  
2021 ◽  
Vol 10 (5) ◽  
pp. 768
Author(s):  
Maddalena Sguizzato ◽  
Francesca Ferrara ◽  
Supandeep Singh Hallan ◽  
Anna Baldisserotto ◽  
Markus Drechsler ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Radical Scavenging ◽  
Human Keratinocytes ◽  
Antioxidant Response ◽  
Correlation Spectroscopy ◽  
Inflammatory Status ◽  
Transmission Electron ◽  
Anti Inflammatory

Mangiferin is a natural glucosyl xanthone with antioxidant and anti-inflammatory activity, making it suitable for protection against cutaneous diseases. In this study ethosomes and transethosomes were designed as topical delivery systems for mangiferin. A preformulation study was conducted using different surfactants in association with phosphatidylcholine. Vesicle dimensional distribution was monitored by photon correlation spectroscopy, while antioxidant capacity and cytotoxicity were respectively assessed by free radical scavenging analysis and MTT on HaCaT keratinocytes. Selected nanosystems were further investigated by cryogenic transmission electron microscopy, while mangiferin entrapment capacity was evaluated by ultracentrifugation and HPLC. The diffusion kinetics of mangiferin from ethosomes and transethosomes evaluated by Franz cell was faster in the case of transethosomes. The suitability of mangiferin-containing nanovesicles in the treatment of skin disorders related to pollutants was investigated, evaluating, in vitro, the antioxidant and anti-inflammatory effect of ethosomes and transethosomes on human keratinocytes exposed to cigarette smoke as an oxidative and inflammatory challenger. The ability to induce an antioxidant response (HO-1) and anti-inflammatory status (IL-6 and NF-kB) was determined by RT-PCR and immunofluorescence. The data demonstrated the effectiveness of mangiferin loaded in nanosystems to protect cells from damage. Finally, to gain insight into the keratinocytes’ uptake of ethosome and transethosome, transmission electron microscopy analyses were conducted, showing that both nanosystems were able to pass intact within the cells.

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