Genistein and Procyanidin B2 Reduce Carcinogen-Induced Reactive Oxygen Species and DNA Damage through the Activation of Nrf2/ARE Cell Signaling in Bronchial Epithelial Cells In Vitro.

Cancer is one of the leading causes of death worldwide. Chemotherapy and radiation therapy are currently providing the basis for cancer therapies, although both are associated with significant side effects. Thus, cancer prevention through dietary modifications has been receiving growing interest. The potential of selected flavonoids in reducing carcinogen-induced reactive oxygen species (ROS) and DNA damage through the activation of nuclear factor erythroid 2 p45 (NF-E2)-related factor (Nrf2)/antioxidant response element (ARE) pathway was studied in vitro. Dose-dependent effects of pre-incubated flavonoids on pro-carcinogen 4-[(acetoxymethyl)nitrosamino]-1-(3-pyridyl)-1-butanone (NNKAc)-induced ROS and DNA damage in human bronchial epithelial cells were studied in comparison to non-flavonoids. The most effective flavonoids were assessed for the activation of Nrf2/ARE pathway. Genistein, procyanidin B2 (PCB2), and quercetin significantly suppressed the NNKAc-induced ROS and DNA damage. Quercetin significantly upregulated the phosphorylated protein kinase B/Akt. PCB2 significantly upregulated the activation of Nrf2 and Akt through phosphorylation. Genistein and PCB2 significantly upregulated the phospho-Nrf2 nuclear translocation and catalase activity. In summary, genistein and PCB2 reduced the NNKAc-induced ROS and DNA damage through the activation of Nrf2. Further studies are required to understand the role of dietary flavonoids on the regulation of the Nrf2/ARE pathway in relation to carcinogenesis.

Protective effect of MitoQ on oxidative stress-mediated senescence of canine bone marrow mesenchymal stem cells via activation of the Nrf2/ARE pathway.

The destruction of biological activity such as senescence and apoptosis caused by oxidative stress could play a pivotal role in the poor therapeutic efficiency of bone marrow mesenchymal stem cells (BMSCs) transplantation. Mitoquinone (MitoQ) has a highly effective mitochondrial antioxidant effect, and has been widely used in many oxidative damage models. This study aimed to investigate the protective effect of MitoQ on the oxidative stress-mediated senescence of canine BMSCs and the underlying mechanism. The senescence of BMSCs was determined by senescence-associated ß-galactosidase staining and quantitative real-time PCR. The expression of p-Nrf2 protein was detected by Western blotting. The results demonstrated that, as BMSCs were expanded in vitro, the senescent phenotype appeared. And the senescence of BMSCs may be caused by oxidative stress, manifested by increasing the level of ROS and decreasing the activity of antioxidant enzymes. Treatment of MitoQ down-regulated the mRNA levels of senescence-related and apoptosis-related genes, but up-regulated the mRNA levels of proliferation-related genes. Meanwhile, ROS generation and senescent activity were reduced in MitoQ-treated BMSCs. Further mechanism studies showed that MitoQ obviously promoted Nrf2 phosphorylation, and also facilitated the translocation of Nrf2 into the nucleus. Moreover, treatment of MitoQ increased the mRNA levels of downstream antioxidant genes and enhanced the activities of superoxide dismutase, catalase, and glutathione peroxidase. Thus, our study revealed that MitoQ, via the Nrf2/ARE signaling pathway, exerts an antioxidant effect as well as potentially delays OS-mediated senescence during BMSCs that were expanded in vitro, which may serve as a novel strategy to optimize the clinical application of BMSCs.

The Cell-Permeable Derivative of the Immunoregulatory Metabolite Itaconate, 4-Octyl Itaconate, Is Anti-Fibrotic in Systemic Sclerosis.

Systemic sclerosis (SSc) is an autoimmune connective tissue disease that leads to skin fibrosis. Altered metabolism has recently been described in autoimmune diseases and SSc. Itaconate is a product of the Krebs cycle intermediate cis-aconitate and is an immunomodulator. This work examines the role of the cell-permeable derivative of itaconate, 4-octyl itaconate (4-OI), in SSc. SSc and healthy dermal fibroblasts were exposed to 4-OI. The levels of collagen Nrf2-target genes and pro-inflammatory cytokines interleukin 6 (IL-6) and monocyte chemotactic protein 1 (MCP-1) were determined. Levels of reactive oxygen species (ROS) as well as the gene expression of collagen and Cellular Communication Network Factor 2 (CCN2) were measured after transforming growth factor beta 1 (TGF-ß1) stimulation in the presence or absence of 4-OI. Wild-type or Nrf2-knockout (Nrf2-KO) mouse embryonic fibroblasts (MEFs) were also treated with 4-OI to determine the role of Nrf2 in 4-OI-mediated effects. 4-OI reduced the levels of collagen in SSc dermal fibroblasts. Incubation with 4-OI led to activation of Nrf2 and its target genes heme oxygenase 1 (HO-1) and NAD(P)H quinone oxidoreductase 1 (NQO1). 4-OI activated antioxidant response element (ARE)-dependent gene expression, reduced inflammatory cytokine release and reduced TGF-ß1-induced collagen and ROS production in dermal fibroblasts. The effects of 4-OI are dependent on Nrf2. The cell-permeable derivative of itaconate 4-OI is anti-fibrotic through upregulation of Nrf2 and could be a potential therapeutic option in an intractable disease.

Discovery and characterization of novel peptide inhibitors of the NRF2/MAFG/DNA ternary complex for the treatment of cancer.

Pathway activating mutations of the transcription factor NRF2 and its negative regulator KEAP1 are strongly correlative with poor clinical outcome with pemetrexed/carbo(cis)platin/pembrolizumab (PCP) chemo-immunotherapy in lung cancer. Despite the strong genetic support and therapeutic potential for a NRF2 transcriptional inhibitor, currently there are no known direct inhibitors of the NRF2 protein or its complexes with MAF and/or DNA. Herein we describe the design of a novel and high-confidence homology model to guide a medicinal chemistry effort that resulted in the discovery of a series of peptides that demonstrate high affinity, selective binding to the Antioxidant Response Element (ARE) DNA and thereby displace NRF2-MAFG from its promoter, which is an inhibitory mechanism that to our knowledge has not been previously described. In addition to their activity in electrophoretic mobility shift (EMSA) and TR-FRET-based assays, we show significant dose-dependent ternary complex disruption of NRF2-MAFG binding to DNA by SPR, as well as cellular target engagement by thermal destabilization of HiBiT-tagged NRF2 in the NCI-H1944 NSCLC cell line upon digitonin permeabilization, and SAR studies leading to improved cellular stability. We report the characterization and unique profile of lead peptide 18, which we believe to be a useful in vitro tool to probe NRF2 biology in cancer cell lines and models, while also serving as an excellent starting point for additional in vivo optimization toward inhibition of NRF2-driven transcription to address a significant unmet medical need in non-small cell lung cancer (NSCLC).

Neuroprotective Effects of Rhynchophylline Against Aß1-42-Induced Oxidative Stress, Neurodegeneration, and Memory Impairment Via Nrf2-ARE Activation.

Extensive studies have shown that oxidative stress is a crucial pathogenic factor in Alzheimer's disease (AD). Nuclear factor E2-related factor 2 (Nrf2) is a master cytoprotective regulator against oxidative stress, and thus represents an attractive therapeutic target in AD. The goal of our study is to investigate the contribution of Nrf2 in Rhynchophylline (Rhy)-induced neuroprotection in AD. The data showed that intraperitoneal administration of Rhy (10 or 20 mg/kg) could ameliorate Aß1-42-induced cognitive impairment, evidenced by performance improvement in memory tests. The result of Antioxidant response element (ARE)-luciferase activity assay indicated that Rhy treatment improved ARE promoter activity. The results of reactive oxygen species (ROS), malondialdehyde (MDA) and glutathione (GSH) assessment in the frontal cortex and hippocampus showed that Rhy treatment could attenuate Aß1-42-induced oxidative stress to some extent, evidenced by reversion of these cytokines compared to Aß1-42 + Veh group. Rhy treatment also restored expression of Nrf2 and its downstream protein heme oxygenase-1 (HO-1), NAD(P)H/quinone oxidoreductase 1 (NOQ1), and recombinant glutamate cysteine ligase, modifier subunit (GCLM) in the frontal cortex and hippocampus of Aß1-42-treated mice. In addition, to investigate whether activation of Nrf2-mediated pathway is responsible for the neuroprotection of Rhy, Nrf2 siRNA was used in human neuroblastoma cells (SH-SY5Y). Interestingly, the results showed that the protective effects of Rhy, including anti-oxidative, anti-apoptosis and elevation of Nrf2 and its downstream proteins, were abolished in Nrf2 siRNA-transfected cells. These findings indicate that Rhynchophylline is protective against Aß1-42-induced neurotoxicity via Nrf2-ARE activation, and suggest that Rhy may serve as a potential candidate and promising Nrf2 activator for management of AD.

Curcumin protects BEAS­2B cells from PM2.5­induced oxidative stress and inflammation by activating NRF2/antioxidant response element pathways.

Fine particulate matter (PM2.5) with an average aerodynamic diameter of <2.5 µm can cause severe lung injury. Oxidative stress and inflammation are considered the main outcomes of PM2.5 exposure. Curcumin is a well­known antioxidant; however, its effect on PM2.5­induced oxidative injury in airway epithelial cells remains unclear. In the present study, it was demonstrated that pre­treatment with curcumin significantly reduced the PM2.5­induced apoptosis of BEAS­2B human bronchial epithelial cells by decreasing the level of intercellular reactive oxygen species. Western blot analysis revealed that curcumin increased the expression of nuclear factor erythroid 2­related factor 2 (NRF2) and regulated the transcription of downstream genes, particularly those encoding antioxidant enzymes. Moreover, curcumin reduced the PM2.5­induced expression and production of inflammatory factors, and induced the expression of the anti­inflammatory factors, interleukin (IL)­5 and IL­13. Taken together, the present study demonstrates that curcumin protects BEAS­2B cells against PM2.5­induced oxidative damage and inflammation, and prevents cell apoptosis by increasing the activation of NRF2­related pathways. It is thus suggested that curcumin may be a potential compound for use in the prevention of PM2.5­induced tissue injury.

(E)-N-(2-(3, 5-Dimethoxystyryl) phenyl) furan-2-carboxamide (BK3C231) induces cytoprotection in CCD18-Co human colon fibroblast cells through Nrf2/ARE pathway activation.

Cytoprotection involving the nuclear factor erythroid 2-related factor 2 (Nrf2)/antioxidant response element (ARE) signaling pathway is an important preventive strategy for normal cells against carcinogenesis. In our previous study, the chemopreventive potential of (E)-N-(2-(3, 5-Dimethoxystyryl) phenyl) furan-2-carboxamide (BK3C231) has been elucidated through its cytoprotective effects against DNA and mitochondrial damages in the human colon fibroblast CCD-18Co cell model. Therefore this study aimed to investigate the molecular mechanisms underlying BK3C231-induced cytoprotection and the involvement of the Nrf2/ARE pathway. The cells were pretreated with BK3C231 before exposure to carcinogen 4-nitroquinoline N-oxide (4NQO). BK3C231 increased the protein expression and activity of cytoprotective enzymes namely NAD(P)H:quinone oxidoreductase 1 (NQO1), glutathione S-transferase (GST) and heme oxygenase-1 (HO-1), as well as restoring the expression of glutamate-cysteine ligase catalytic subunit (GCLC) back to the basal level. Furthermore, dissociation of Nrf2 from its inhibitory protein, Keap1, and ARE promoter activity were upregulated in cells pretreated with BK3C231. Taken together, our findings suggest that BK3C231 exerts cytoprotection by activating the Nrf2 signaling pathway which leads to ARE-mediated upregulation of cytoprotective proteins. This study provides new mechanistic insights into BK3C231 chemopreventive activities and highlights the importance of stilbene derivatives upon development as a potential chemopreventive agent.

The protective role of luteolin against the methotrexate-induced hepato-renal toxicity via its antioxidative, anti-inflammatory, and anti-apoptotic effects in rats.

Methotrexate (MTX) is frequently used drug in treatment of cancer and autoimmune diseases. Unfortunately, MTX has many side effects including the hepato-renal toxicity. In this study, we hypothesized that Luteolin (Lut) exhibits protective effect against the MTX-induced hepato-renal toxicity. In order to investigate our hypothesis, the experiment was designed to examine the effect of exposure of male rats to MTX (20 mg/kg, i.p., at day 9) alone or together with Lut (50 mg/kg, oral for 14 days) compared to the control rats (received saline). The findings demonstrated that MTX treatment induced significant increases in the liver and kidney functions markers in serum samples including Aspartate transaminase (AST), Alanine transaminase (ALT), creatinine, urea and uric acid. MTX also mediated an oxidative stress expressed by elevated malondialdehyde (MDA) level and decreased level of reduced glutathione (GSH), antioxidant enzyme activities, and downregulation of the Nrf2 gene expression as an antioxidant trigger. Moreover, the inflammatory markers (NF-κB, TNF-α, and IL-1ß) were significantly elevated upon MTX treatment. In addition, MTX showed an apoptotic response mediated by elevating the pro-apoptotic (Bax) and lowering the anti-apoptotic (Bcl-2) proteins. All of these changes were confirmed by the observed alterations in the histopathological examination of the hepatic and renal tissues. Lut exposure significantly reversed all the MTX-induced changes in the measured parameters suggesting its potential protective role against the MTX-induced toxicity. Finally, our findings concluded the antioxidative, anti-inflammatory and anti-apoptotic effects of Lut as a mechanism of its protective role against the MTX-induced hepato-renal toxicity in rats.

Astragaloside IV, a saponin from Astragalus membranaceus var. mongholicus, induces expressions of heme recycle proteins via signaling of Nrf2/ARE in cultured macrophages.

ETHNOPHARMACOLOGICAL RELEVANCE: In traditional Chinese medicine (TCM) theory, "Qi" is classified as energetic essence supporting the life activities in human. "Blood" is categorized as nourishing essence and circulating in the body. "Blood" and "Qi" have an intimate relationship. Astragali Radix (AR; root of Astragalus membranaceus (Fisch.) Bge. Var. mongholicus (Bge.) Hsiao) has a broad spectrum of application for "Qi-Blood" enrichment. Astragaloside IV, a major saponin in AR, has therapeutic functions in erythropoietic, cardiovascular and immune systems. However, the efficacy of astragaloside IV in erythrophagocytosis has not been elucidated. AIM OF THE STUDY: The possible functions of astragaloside IV in heme iron recycling during erythrophagocytosis in cultured macrophage were elucidated. METHODS: The translational and transcriptional expressions of heme recycling enzymes were determined after incubating of astragaloside IV for 24 h in cultured macrophage. RESULTS: In astragaloside IV-treated macrophage, the expressions, both RNA and protein levels, of regulators of heme recycling, e.g. heme oxygenase-1 (HO-1), ferroportin (FPN), biliverdin reductase A and B (BVRA, BVRB), were markedly induced in dose-dependent manners. In parallel, the transcriptional activity of antioxidant response element, cloned within an expression vector as pARE-Luc and transfected in cultured macrophages, was markedly induced after a challenge with astragaloside IV in a dose-dependent manner. Moreover, the translocation of Nrf2, a transcriptional factor in regulating expression of heme recycling protein, was induced by astragaloside IV, leading to an enrichment at nucleus fraction. CONCLUSION: Astragaloside IV shed lights in enhancing the expression of heme recycle proteins via Nrf2/ARE signaling pathway.

Puerarin induces apoptosis in prostate cancer cells via inactivation of the Keap1/Nrf2/ARE signaling pathway.

In this study, we examined the antitumor effects of Puerarin (PEU) on androgen-independent (DU145 and PC-3) and androgen-dependent (LNCaP) prostate cancer cells, and explored its potential mechanisms. Supplement with PEU (2.5 µM, 5 µM, and 10 µM) exhibited a marked inhibitory effect against the growth of DU145 and PC-3 cells, especially beyond 24 h, whereas there is only slight growth inhibitory effect on LNCaP cells at the high concentration of 10 µM at 72 h. This loss of cell viability in DU145 and PC-3 cells by PEU was mediated by the induction of apoptosis via up-regulation of Bax and cleaved-caspase-3, but downregulation of Bcl-2. Moreover, more intracellular ROS and LDH production were observed in DU145 and PC-3 cells upon PEU treatment. Meanwhile, the amount of pro-inflammatory cytokines (IL-1ß and IL-6) was increased, but the content of anti-inflammatory cytokines IL-10 was attenuated. Additionally, PEU pretreatment resulted in an increase of Keap1 protein expression, and a decline of Nrf2, HO-1 and NQO1 protein expression in DU145 and PC3 cells. The present findings indicated that PEU exerted its antitumor activities toward androgen-independent prostate cancer cells via inactivation of Keap1/NrF2/ARE signaling pathway.

Potential neuroprotection of wheat alkylresorcinols in hippocampal neurons via Nrf2/ARE pathway.

5-n-Alkylresorcinols (ARs) are abundant in wheat bran and potentially antioxidative, although the neuroprotective mechanism is not fully understood. The neuroprotective effect of wheat bran ARs on H2O2-induced neuronal cells and the relationship between neuroprotection and the nuclear factor erythroid 2-related factor 2 (Nrf2)/antioxidant-response element (ARE) pathway were investigated in this study. Seven homologs were identified in the purified ARs by high-performance liquid chromatography-tandem mass spectrometry. Pretreatment with 80 µg mL-1 ARs alleviated 23% HT22 cell death and the up-regulation of intracellular reactive oxygen species level and malondialdehyde under H2O2 stimulation. The neuroprotection effect was proved by the increase in the Nrf2 nuclear location and up-regulation of mRNA and protein expressions of heme oxygenase-1, NAD(P)H quinone dehydrogenase 1, glutamate-cysteine ligase catalytic subunit, and glutamate-cysteine ligase modifier subunit l. Wheat bran ARs displayed a neuroprotective function, possibly by promoting the endogenous antioxidant defense system. ARs may be regarded as a functional food ingredient for preventing neurodegenerative diseases in the future.

5-Caffeoylquinic acid ameliorates oxidative stress-mediated cell death via Nrf2 activation in hepatocytes.

CONTEXT: 5-Caffeoylquinic acid (5-CQA) is one of the most abundant compounds found in natural foods including coffee. OBJECTIVE: We investigated whether 5-CQA had a cytoprotective effect through the NF-E2-related factor 2 (Nrf2)-antioxidant response element (ARE) signalling pathway. MATERIALS AND METHODS: Nrf2 activation in response to 5-CQA treatment at the concentration of 10-100 µM is evaluated by Western blotting of Nrf2 and ARE reporter gene assay as well as its target gene expression in HepG2 cells. Intracellular reactive oxygen species (ROS) and glutathione (GSH) levels were measured in the tert-butyl hydroperoxide-induced hepatocytes to examined cytoprotective effect of 5-CQA (10-100 µM). The specific role of 5-CQA on Nrf2 activation was examined using Nrf2 knockout cells or Nrf2 specific inhibitor, ML-385. RESULTS: Nuclear translocation of Nrf2 is increased by 5-CQA in HepG2 cells which peaked at 6 h. Consequently, 5-CQA significantly increases the ARE reporter gene activity and downstream antioxidant proteins, including glutamate cysteine ligase (GCL), hemeoxygenase-1 (HO-1), NAD(P)H quinone oxidoreductase 1, and Sestrin2. Nrf2 deficiency or inhibition completely antagonized ability of 5-CQA to induce HO-1 and GCL expression. Cells pre-treated with 5-CQA were rescued from tert-butyl hydroperoxide-induced ROS production and GSH depletion. Nrf2 activation by 5-CQA was due to increased phosphorylation of MAPKs, AMPK and PKCδ. DISCUSSION AND CONCLUSIONS: Taken together, our results demonstrate that as a novel Nrf2 activator, 5-CQA, may be a promising candidate against oxidative stress-mediated liver injury. Additional efforts are needed to assess 5-CQA, as a potential therapeutic in liver diseases in vivo and in humans.

S-Allylcysteine Protects Against Excitotoxic Damage in Rat Cortical Slices Via Reduction of Oxidative Damage, Activation of Nrf2/ARE Binding, and BDNF Preservation.

Neuroprotective approaches comprising different mechanisms to counteract the noxious effects of excitotoxicity and oxidative stress need validation and detailed characterization. Although S-allylcysteine (SAC) is a natural compound exhibiting a broad spectrum of protective effects characterized by antioxidant, anti-inflammatory, and neuromodulatory actions, the mechanisms underlying its protective role on neuronal cell damage triggered by early excitotoxic insults remain elusive. In this study, we evaluated if the preconditioning or the post-treatment of isolated rat cortical slices with SAC (100 µM) can ameliorate the toxic effects induced by the excitotoxic metabolite quinolinic acid (QUIN, 100 µM), and whether this protective response involves the early display of specific antioxidant and neuroprotective signals. For this purpose, cell viability/mitochondrial reductive capacity, lipid peroxidation, levels of reduced and oxidized glutathione (GSH and GSSG, respectively), the rate of cell damage, the NF-E2-related factor 2/antioxidant response element (Nrf2/ARE) binding activity, heme oxygenase 1 (HO-1) regulation, extracellular signal-regulated kinase (ERK1/2) phosphorylation, and the levels of tumor necrosis factor-alpha (TNF-α) and the neurotrophin brain-derived neurotrophic factor (BDNF) were all estimated in tissue slices exposed to SAC and/or QUIN. The incubation of slices with QUIN augmented all toxic endpoints, whereas the addition of SAC prevented and/or recovered all toxic effects of QUIN, exhibiting better results when administered 60 min before the toxin and demonstrating protective and antioxidant properties. The early stimulation of Nrf2/ARE binding activity, the upregulation of HO-1, the ERK1/2 phosphorylation and the preservation of BDNF tissue levels by SAC demonstrate that this molecule displays a wide range of early protective signals by triggering orchestrated antioxidant responses and neuroprotective strategies. The relevance of the characterization of these mechanisms lies in the confirmation that the protective potential exerted by SAC begins at the early stages of excitotoxicity and neurodegeneration and supports the design of integral prophylactic/therapeutic strategies to reduce the deleterious effects observed in neurodegenerative disorders with inherent excitotoxic events.

Bioassay-Guided Separation of Centipeda minima Using Comprehensive Linear Gradient Centrifugal Partition Chromatography.

A comprehensive linear gradient solvent system for centrifugal partition chromatography (CPC) was developed for the bioassay-guided isolation of natural compounds. The gradient solvent system consisted of three different ternary biphasic solvents types: n-hexane-acetonitrile-water (10:2:8, v/v), ethyl acetate-acetonitrile-water (10:2:8, v/v), and water-saturated n-butanol-acetonitrile-water (10:2:8, v/v). The lower phase of the n-hexane-acetonitrile-water (10:2:8, v/v) was used as the stationary phase, while its upper phase, as well as ethyl acetate-acetonitrile-water (10:2:8), and water-saturated n-butanol-acetonitrile-water (10:2:8, v/v) were pumped to generate a linear gradient elution, increasing the mobile phase polarity. We used the gradient CPC to identify antioxidant response elements (AREs), inducing compounds from Centipeda minima, using an ARE-luciferase assay in HepG2 cells, which led to the purification of the active molecules 3-methoxyquercetin and brevilin A. The developed CPC solvent systems allow the separation and isolation of compounds with a wide polarity range, allowing active molecule identification in the complex crude extract of natural products.

Vinpocetine reduces cisplatin-induced acute kidney injury through inhibition of NF-κB pathway and activation of Nrf2/ARE pathway in rats.

Acute kidney injury is a complex clinical disease that is associated with a high incidence of morbidity and mortality. Drug-induced acute kidney injury occurs in approximately 19-33% of hospitalized patients. Cisplatin, one of the most commonly used and effective chemotherapeutic drugs not only exerts anti-tumor effects but also causes renal toxicity damage, affecting its clinical application. Vinpocetine is an anti-inflammatory and antioxidant drug that predominately acts in the nervous system. In this study, we investigated the effects and mechanisms of vinpocetine in an animal model of cisplatin-induced acute renal injury. Rats were randomly divided into three experimental groups. During a 10-day trial, rats in the control group were administered a physiological saline solution; rats in the model group received a 5 mg/kg intraperitoneal injection of cisplatin; and rats in the cisplatin + vinpocetine group received a 5 mg/kg intraperitoneal injection of cisplatin as well as a 5 mg/kg dose of vinpocetine via gavage. We observed that following cisplatin administration, the rats exhibited an increase in blood urea and creatinine levels as well as an increase in their inflammation and oxidative stress levels. In renal tissue, cisplatin caused the morphological changes typical of acute tubular injury. Vinpocetine reduced the cisplatin-induced acute renal function damage and tubular injury. In both in vivo and in vitro experiments, we found that vinpocetine can confer protection of rat renal cells by inhibiting the NF-κB signaling pathway and activating the Nrf2/ARE signaling pathway. Therefore, vinpocetine is a promising therapeutic drug for the treatment of cisplatin-induced acute kidney injury.

Coumarins as Modulators of the Keap1/Nrf2/ARE Signaling Pathway.

The Keap1/Nrf2/ARE system is a central defensive mechanism against oxidative stress which plays a key role in the pathogenesis and progression of many diseases. Nrf2 is a redox-sensitive transcription factor controlling a variety of downstream antioxidant and cytodefensive genes. Nrf2 has a powerful anti-inflammatory activity mediated via modulating NF-κB. Therefore, pharmacological activation of Nrf2 is a promising therapeutic strategy for the treatment/prevention of several diseases that are underlined by both oxidative stress and inflammation. Coumarins are natural products with promising pharmacological activities, including antioxidant, anticancer, antimicrobial, and anti-inflammatory efficacies. Coumarins are found in many plants, fungi, and bacteria and have been widely used as complementary and alternative medicines. Some coumarins have shown an ability to activate Nrf2 signaling in different cells and animal models. The present review compiles the research findings of seventeen coumarin derivatives of plant origin (imperatorin, visnagin, urolithin B, urolithin A, scopoletin, esculin, esculetin, umbelliferone, fraxetin, fraxin, daphnetin, anomalin, wedelolactone, glycycoumarin, osthole, hydrangenol, and isoimperatorin) as antioxidant and anti-inflammatory agents, emphasizing the role of Nrf2 activation in their pharmacological activities. Additionally, molecular docking simulations were utilized to investigate the potential binding mode of these coumarins with Keap1 as a strategy to disrupt Keap1/Nrf2 protein-protein interaction and activate Nrf2 signaling.

Luteolin prevents liver from tunicamycin-induced endoplasmic reticulum stress via nuclear factor erythroid 2-related factor 2-dependent sestrin 2 induction.

Endoplasmic reticulum (ER) stress designates a cellular response to the accumulation of misfolded proteins, which is related to disease progression in the liver. Luteolin (3',4',5,7-tetrahydroxyflavone) is a phytochemical found frequently in medicinal herbs. Although luteolin has been reported to possess the therapeutic potential to prevent diverse stage of liver diseases, its role in hepatic ER stress has not been established. Thus, the present study aimed to determine the role of luteolin in tunicamycin (Tm)-induced ER stress, and to identify the relevant mechanisms involved in its hepatoprotective effects. In hepatocyte-derived cells and primary hepatocytes, luteolin significantly decreased Tm- or thapsigargin-mediated C/EBP homologous protein (CHOP) expression. In addition, luteolin reduced the activation of three canonical signaling pathways related to the unfolded protein response, and decreased mRNA levels of glucose-regulated protein 78, ER DNA J domain-containing protein 4, and asparagine synthetase. Luteolin also significantly upregulated sestrin 2 (SESN2), and luteolin-mediated CHOP inhibition was blocked in SESN2 (+/-) cells. Moreover, luteolin resulted in phosphorylation of nuclear factor erythroid 2-related factor 2 (Nrf2), as well as increased nuclear Nrf2 expression. Deletion of the antioxidant response element in the human SESN2 promoter inhibited increased luciferase activation by luteolin, suggesting that Nrf2 is a critical transcription factor for luteolin-dependent SESN2 expression. In a Tm-mediated liver injury model, luteolin decreased serum alanine aminotransferase and aspartate aminotransferase activities, prevented degenerative changes and apoptosis of hepatocytes, and inhibited CHOP and glucose-regulated protein 78 expression in hepatic tissues. Therefore, luteolin may be an effective phytochemical to manage ER stress-related liver injury.

Perilipin 5 Reduces Oxidative Damage Associated With Lipotoxicity by Activating the PI3K/ERK-Mediated Nrf2-ARE Signaling Pathway in INS-1 Pancreatic ß-Cells.

Oxidative stress induced by free fatty acid overload in pancreatic ß-cells is a potential contributory factor to dysfunction of insulin secretion and apoptotic cell death. Perilipin 5 (Plin5) has been reported to ameliorate oxidative stress-mediated damage in non-insulin-secreting tissues. We tested the hypothesis that Plin5 plays a similar role in pancreatic ß-cells, which are extremely sensitive to oxidative stress. Here, our in vitro data showed that Plin5-mediated alleviation of palmitate-triggered apoptosis involves the mitochondrial pathway. And the protective role of Plin5 on ß-cells was partially dependent on its modulation in oxidative stress. Upregulation of Plin5 in INS-1 cells decreased reactive oxygen species production, enhanced cellular glutathione levels, and induced expression of antioxidant enzymes glutamate-cysteine ligase catalytic subunit and heme oxygenase-1. However, knocking out of Plin5 abolished all of these beneficial effects. Furthermore, by using the O2- scavenger MnTMPyP, we verified that altering Plin5 expression impacted lipotoxic cell death partially via modulating oxidative stress. Mechanistic experiments revealed that Plin5 induced Nrf2-ARE system, a master regulator in the cellular adaptive response to oxidative stress, by activating PI3K/Akt and ERK signal pathways, contributing to the increase of antioxidant defense and consequently improving ß-cell function and survival in the presence of lipotoxic oxidative stress. Overall, our findings indicate that Plin5 abrogates oxidative damage in INS-1 ß-cells during lipotoxic stress partially through the enhancement of antioxidant defense involving the PI3K/Akt and ERK mediated Nrf2-ARE system.

Polysaccharide from Mulberry Fruit (Morus alba L.) Protects against Palmitic-Acid-Induced Hepatocyte Lipotoxicity by Activating the Nrf2/ARE Signaling Pathway.

This study was aimed to investigate the protective effects of three different mulberry fruit polysaccharide fractions (MFP-I, MFP-II, and MFP-III) against palmitic acid (PA)-induced hepatocyte lipotoxicity and characterize the functional polysaccharide fraction using gel permeation chromatography, high-performance liquid chromatography, Fourier transform infrared spectroscopy, and nuclear magnetic resonance analyses. MFP-I, MFP-II, and MFP-III were isolated from mulberry fruit by stepwise precipitation with 30, 60, and 90% ethanol, respectively. MFP-II at 0.1 and 0.2 mg/mL dramatically attenuated PA-induced hepatic lipotoxicity, while MFP-I and MFP-III showed weak protection. It was demonstrated that MFP-II not only increased nuclear factor erythroid-2-related factor 2 (Nrf2) phosphorylation and its nuclear translocation, thereby activating the Nrf2/ARE signaling pathway, but also enhanced heme oxygenase 1, NAD(P)H:quinone oxidoreductase 1, and γ-glutamate cysteine ligase gene expressions and promoted catalase and glutathione peroxidase activities, which protected hepatocytes against PA-induced oxidative stress and lipotoxicity. Further investigation indicated that the molecular weight of MFP-II was 115.0 kDa, and MFP-II mainly consisted of galactose (30.5%), arabinose (26.2%), and rhamnose (23.1%). Overall, our research might provide in-depth insight into mulberry fruit polysaccharide in ameliorating lipid metabolic disorders.

Mitochondrial biology in airway pathogenesis and the role of NRF2.

A constant improvement in understanding of mitochondrial biology has provided new insights into mitochondrial dysfunction in human disease pathogenesis. Impaired mitochondrial dynamics caused by various stressors are characterized by structural abnormalities and leakage, compromised turnover, and reactive oxygen species overproduction in mitochondria as well as increased mitochondrial DNA mutation frequency, which leads to modified energy production and mitochondria-derived cell signaling. The mitochondrial dysfunction in airway epithelial, smooth muscle, and endothelial cells has been implicated in diseases including chronic obstructive lung diseases and acute lung injury. Increasing evidence indicates that the NRF2-antioxidant response element (ARE) pathway not only enhances redox defense but also facilitates mitochondrial homeostasis and bioenergetics. Identification of functional or potential AREs further supports the role for Nrf2 in mitochondrial dysfunction-associated airway disorders. While clinical reports indicate mixed efficacy, NRF2 agonists acting on respiratory mitochondrial dynamics are potentially beneficial. In lung cancer, growth advantage provided by sustained NRF2 activation is suggested to be through increased cellular antioxidant defense as well as mitochondria reinforcement and metabolic reprogramming to the preferred pathways to meet the increased energy demands of uncontrolled cell proliferation. Further studies are warranted to better understand NRF2 regulation of mitochondrial functions as therapeutic targets in airway disorders.