Igalan induces detoxifying enzymes mediated by the Nrf2 pathway in HepG2 cells

Igalan is one of the sesquiterpene lactones found in Inula helenium L., which is used as the traditional medicine to treat inflammatory diseases. However, the pharmacological effects of igalan have not been characterized. In this study, we isolated igalan from I. helenium L. and evaluated the effects of igalan on signaling pathways and expression of target genes in HepG2 cells. Igalan activated the nuclear factor erythroid 2‐related factor 2 (Nrf2) pathway by increasing the inactive form of GSK3β, the phosphorylated form of AKT, and the nuclear accumulation of Nrf2. Thus, target genes of Nrf2 such as HO‐1 and NQO1 increased in HepG2 cells. Moreover, igalan inhibited the tumor necrosis factor‐α (TNF‐α)‐induced nuclear factor‐κB activation and suppressed the expression of its target genes, including TNF‐α, interleukin (IL)‐6, and IL‐8 in HepG2 cells. Our results indicate the potential of igalan as an activator of cellular defense mechanisms and a detoxifying agent.     

Nrf2 attenuates inflammatory response in COPD/emphysema: Crosstalk with Wnt3a/β‐catenin and AMPK pathways

Chronic obstructive pulmonary disease (COPD) is characterized by persistent airflow limitation and abnormal inflammatory response. Wnt/β‐catenin and AMP‐activated protein kinase (AMPK) have been shown to modulate lung inflammatory responses and injury. However, it remains elusive whether Wnt/β‐catenin and AMPK modulate nuclear factor erythroid‐2 related factor‐2 (Nrf2)‐mediated protective responses during the development of emphysema. Here we showed that treatment with a Wnt pathway activator (LiCl) reduced elastase‐induced airspace enlargement and cigarette smoke extract (CSE)‐induced lung inflammatory responses in WT mice, which was associated with increased activation of Nrf2 pathway. Interestingly, these effects of LiCl were not observed in Nrf2^?/? mice exposed to elastase. In normal human bronchial epithelial (NHBE) cells, Wnt3a overexpression up‐regulated, whereas Wnt3a knockdown further down‐regulated the levels of Nrf2 and its target proteins heme oxygenase‐1 (HO‐1) and NAD(P)H: quinone oxidoreductase 1 (NQO1) by CSE treatment. In contrast, Nrf2 deficiency did not have any effects on Wnt/β‐catenin pathway in mouse lungs and NHBE cells. Both elastase and CSE exposures reduced AMPK phosphorylation. A specific AMPK activator metformin increased Wnt3a, β‐catenin, Nrf2 phosphorylation and activation but reduced the levels of IL‐6 and IL‐8 in NHBE cells and mouse lungs exposed to CSE. Furthermore, Nrf2 deficiency abolished the protection of metformin against CSE‐induced increase in IL‐6 and IL‐8 in NHBE cells. In conclusion, Nrf2 mediates the protective effects of both Wnt3a/β‐catenin and AMPK on lung inflammatory responses during the development of COPD/emphysema. These findings provide potential therapeutic targets for the intervention of COPD/emphysema.     

Proteasome dysfunction in Drosophila signals to an Nrf2‐dependent regulatory circuit aiming to restore proteostasis and prevent premature aging

The ubiquitin–proteasome system is central to the regulation of cellular proteostasis. Nevertheless, the impact of in vivo proteasome dysfunction on the proteostasis networks and the aging processes remains poorly understood. We found that RNAi‐mediated knockdown of 20S proteasome subunits in Drosophila melanogaster resulted in larval lethality. We therefore studied the molecular effects of proteasome dysfunction in adult flies by developing a model of dose‐dependent pharmacological proteasome inhibition. Impaired proteasome function promoted several ‘old‐age’ phenotypes and markedly reduced flies' lifespan. In young somatic tissues and in gonads of all ages, loss of proteasome activity induced higher expression levels and assembly rates of proteasome subunits. Proteasome dysfunction was signaled to the proteostasis network by reactive oxygen species that originated from malfunctioning mitochondria and triggered an Nrf2‐dependent upregulation of the proteasome subunits. RNAi‐mediated Nrf2 knockdown reduced proteasome activities, flies' resistance to stress, as well as longevity. Conversely, inducible activation of Nrf2 in transgenic flies upregulated basal proteasome expression and activity independently of age and conferred resistance to proteotoxic stress. Interestingly, prolonged Nrf2 overexpression reduced longevity, indicating that excessive activation of the proteostasis pathways can be detrimental. Our in vivo studies add new knowledge on the proteotoxic stress‐related regulation of the proteostasis networks in higher metazoans. Proteasome dysfunction triggers the activation of an Nrf2‐dependent tissue‐ and age‐specific regulatory circuit aiming to adjust the cellular proteasome activity according to temporal and/or spatial proteolytic demands. Prolonged deregulation of this proteostasis circuit accelerates aging.     

Metformin-ROS-Nrf2 connection in the host defense mechanism against oxidative stress,apoptosis, cancers,and ageing

Reactive oxygen species (ROS) acts as a second messenger to trigger biological responses in low concentrations, while it is implicated to be toxic to biomolecules in high concentrations. Mild inhibition of respiratory chain Complex I by metformin at physiologically relevant concentrations stimulates production of low-level mitochondrial ROS. The ROS seems to induce anti-oxidative stress response via activation of nuclear factor erythroid 2-related factor 2 (Nrf2) and glutathione peroxidase (GPx), which results in not only elimination of ROS but also activation of cellular responses including resistance to apoptosis, metabolic changes, cell proliferation, senescence prevention, lifespan extension, and immune T cell activation against cancers, regardless of its effect controlling blood glucose level and T2DM. Although metformin's effect against T2DM, cancers, and ageing, are believed mostly attributed to the activation of AMP-activated protein kinase (AMPK), the cellular responses involving metformin-ROS-Nrf2 axis might be another natural asset to improve healthspan and lifespan.     

Increasing autophagy and blocking Nrf2 suppress laminopathy‐induced age‐dependent cardiac dysfunction and shortened lifespan

Mutations in the human LMNA gene cause a collection of diseases known as laminopathies. These include myocardial diseases that exhibit age‐dependent penetrance of dysrhythmias and heart failure. The LMNA gene encodes A‐type lamins, intermediate filaments that support nuclear structure and organize the genome. Mechanisms by which mutant lamins cause age‐dependent heart defects are not well understood. To address this issue, we modeled human disease‐causing mutations in the Drosophila melanogaster Lamin C gene and expressed mutant Lamin C exclusively in the heart. This resulted in progressive cardiac dysfunction, loss of adipose tissue homeostasis, and a shortened adult lifespan. Within cardiac cells, mutant Lamin C aggregated in the cytoplasm, the CncC(Nrf2)/Keap1 redox sensing pathway was activated, mitochondria exhibited abnormal morphology, and the autophagy cargo receptor Ref2(P)/p62 was upregulated. Genetic analyses demonstrated that simultaneous over‐expression of the autophagy kinase Atg1 gene and an RNAi against CncC eliminated the cytoplasmic protein aggregates, restored cardiac function, and lengthened lifespan. These data suggest that simultaneously increasing rates of autophagy and blocking the Nrf2/Keap1 pathway are a potential therapeutic strategy for cardiac laminopathies.     

Inhibition of Nrf2 promotes the antitumor effect of Pinelliae rhizome in papillary thyroid cancer

We previously reported that Xiaotan Sanjie (XTSJ) decoction can prevent the progression of gastric cancer in vitro and in vivo. Pinelliae rhizome (PR), one component of XTSJ decoction, has an inhibitory effect on the growth and proliferation of tumor cells. The present study investigated the underlying mechanisms of action of PR. Using the human papillary thyroid cancer cell lines, TPC-1 and BCPAP, we found that XTSJ decoction and PR alone decreased cell viability to a similar extent in both cell lines, whereas treatment with XTJS decoction without PR [PR (−)] had a lesser effect. PR treatment inhibited the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) in a dose-dependent manner. To investigate the role of Nrf2 in the PR-mediated effects of XTSJ, knockdown of Nrf2 in the tumor cell lines using Nrf2 siRNA (siNrf2) was performed and transfected cells were treated with PR. Silencing of Nrf2 amplified the effects on autophagy, cell viability, apoptosis, and colony formation. Similar results were obtained following treatment with the autophagy inhibitor 3-methyladenine (3-MA). Furthermore, treatment with PR, siNrf2, and/or 3-MA inhibited the MAPK pathway, and analysis of the MAPK pathway components confirmed the role of this pathway in the PR-mediated cellular effects. In mice implanted with siNrf2-transfected cells, the effects of PR were amplified. Taken together, these findings indicate that PR is critical for the inhibitory effects of XTSJ decoction on tumor cell viability and that downregulation of Nrf2 promotes the antitumor effects of PR on papillary thyroid cancer cells.     

Geniposide alleviates non‐alcohol fatty liver disease via regulating Nrf2/AMPK/mTOR signalling pathways

Non‐alcohol fatty liver disease (NAFLD) is a common disease which causes serious liver damage. Geniposide (GEN), a kind of iridoid glycoside extracted from Gardenia jasminoides fruit, has many biological effects, such as resistance to cell damage and anti‐neurodegenerative disorder. Lipid accumulation was obvious in tyloxapol‐induced liver and oil acid (OA) with palmitic acid (PA)‐induced HepG2 cells compared with the control groups while GEN improved the increasing conditions. GEN significantly lessened the total cholesterol (TC), the triglyceride (TG), low‐density lipoprotein (LDL), very low‐density lipoprotein (VLDL), myeloperoxidase (MPO), reactive oxygen species (ROS) and increased high‐density lipoprotein (HDL), superoxide dismutase (SOD) to response the oxidative stress via activating nuclear factor erythroid‐2–related factor 2 (Nrf2), haeme oxygenase (HO)‐1 and peroxisome proliferator‐activated receptor (PPAR)α which may influence the phosphorylation of adenosine 5’‐monophosphate–activated protein kinase (AMPK) signalling pathway in mice and cells. Additionally, GEN evidently decreased the contents of sterol regulatory element‐binding proteins (SREBP)‐1c, phosphorylation (P)‐mechanistic target of rapamycin complex (mTORC), P‐S6K, P‐S6 and high mobility group protein (HMGB) 1 via inhibiting the expression of phosphoinositide 3‐kinase (PI3K), and these were totally abrogated in Nrf2^?/? mice. Our study firstly proved the protective effect of GEN on lipid accumulation via enhancing the ability of antioxidative stress and anti‐inflammation which were mostly depend on up‐regulating the protein expression of Nrf2/HO‐1 and AMPK signalling pathways, thereby suppressed the phosphorylation of mTORC and its related protein.     

Preventive effects of ZPDC glycoprotein (24 kDa) on hepatotoxicity induced by mercury chloride in vitro and in vivo

Mercury is a potent environmental contaminant that exerts toxic effect on various vital organs in the human body. Recently, we isolated glycoprotein from Zanthoxylum piperitum DC (ZPDC), which has antioxidant and anticancer effects. In the present study, we determined the preventive effects of ZPDC glycoprotein on hepatic damage induced by mercury chloride (HgCl₂). We evaluated the activities of lactate dehydrogenase (LDH), alanine aminotransferase (ALT), antioxidant enzymes [superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx)], extracellular signal‐regulated kinase (ERK)1/2, p38 mitogen‐activated protein kinase (MAPK), cyclo‐oxygenase (COX‐2), inducible nitric oxide synthetase (iNOS), and activator protein (AP‐1) and the quantitative expressions of nuclear factor E2‐related factor (Nrf2), heme oxygenase (HO‐1), metallothionein (MT) and reduced glutathione (GSH) in mercury‐chloride‐exposed (50 μM and 10 mg/kg body weight) primary cultured hepatocytes and ICR mice, using biochemical assays, radioactivity and immunoblot analysis. The results demonstrated that ZPDC glycoprotein decreased the levels of LDH, ALT, HO‐1 and MT, whereas it increased the activities of hepatic antioxidant enzymes (SOD, CAT and GPx) and reduced GSH in mercury‐chloride‐exposed primary cultured hepatocytes. Also, it suppressed arachidonic acid release and expression of ERK, p38 MAPK, COX‐2, iNOS, AP‐1 and Nrf‐2 in primary cultured hepatocytes and ICR mice exposed to mercury chloride. Collectively, ZPDC glycoprotein may have potential applications to prevent hepatotoxicity induced by mercury chloride. Copyright © 2014 John Wiley & Sons, Ltd.