Nrf2-mediated Antioxidant Enzymes Research Articles

Water extract of lotus leaves has hepatoprotective activity by enhancing Nrf2- and epigenetics-mediated cellular antioxidant capacity in mouse hepatocytes

• The main phenolics in LL-WE were EGCG, rutin, gallic acid, myricetin, quercetin, ellagic acid, etc. • LL-WE reduced the damage of mouse AML-12 hepatocytes caused by APAP-induced oxidative stress. • LL-WE promoted glutathione synthesis and activated the Nrf2 pathway. • LL-WE upregulated Nrf2-mediated antioxidant enzymes and downregulated HDACs and DNMTs. • LL-WE inhibited the in vitro activity of CpG methyltransferase (M.SssI). The liver is an important organ in the human body; however, studies have shown that the excessive intake of acetaminophen (APAP), the primary ingredient in marketing pain relievers, may cause oxidative damage to liver cells. Here we found that lotus leaf water extract (LL-WE) enriched with various phenolic compounds can prevent mice AML-12 hepatocytes from APAP-induced injury, and trigger the Nrf2 pathway in HepG2-C8 cells with ARE-luciferase plasmid. LL-WE effectively induced Nrf2 nuclear translocation and glutathione synthesis, increased gene expression of antioxidant enzymes (HO-1, NQO1, and UGT1A), and reduced protein levels of epigenetic modification enzymes (HDACs and DNMTs) in AML-12 cells. LL-WE also inhibited the activity of M.SssI CpG methyltransferase in vitro . These results suggest that LL-WE can be a potential hepatoprotective agent to reduce oxidative stress-induced liver damage by activating the Nrf2 pathway and downregulating epigenetic modification enzymes.

Sulforaphane induces colorectal cancer cell proliferation through Nrf2 activation in a p53-dependent manner

Sulforaphane is a well-known phytochemical that stimulates nuclear factor erythroid 2-related factor 2 (Nrf2)-mediated antioxidant cellular response. In this study, we found that sulforaphane promoted cell proliferation in HCT116 human colon cancer cells expressing a normal p53 gene in a dose-dependent but biphasic manner. Since p53 has been reported to contribute to cell survival by regulating various metabolic pathways to adapt to mild stress, we further examined cellular responses in both p53-wild-type (WT) and p53-knockout (KO) HCT116 cells exposed to sulforaphane in vitro and in vivo. Results demonstrated that sulforaphane treatment activated Nrf2-mediated antioxidant enzymes in both p53-WT and p53-KO cells, decreased apoptotic protein expression in WT cells but increased in KO cells in a dose-dependent manner, and increased the expression of a mitochondrial biogenesis marker PGC1α in WT cells but decreased in KO cells. Moreover, a low dose of sulforaphane promoted tumor growth, upregulated the Nrf2 signaling pathway, and decreased apoptotic cell death in p53-WT HCT116 xenografts compared to that in p53-KO HCT116 xenografts in BALB/c nude mice. These findings suggest that sulforaphane can influence colon cancer cell proliferation and mitochondrial function through a crosstalk between the Nrf2 signaling pathway and p53 axis.

Bach1 Inhibition Suppresses Osteoclastogenesis via Reduction of the Signaling via Reactive Oxygen Species by Reinforced Antioxidation.

Bone destructive diseases such as periodontitis are common worldwide and are caused by excessive osteoclast formation and activation. Receptor activator of nuclear factor-κB ligand (RANKL) is essential factor for osteoclastogenesis. This triggers reactive oxygen species (ROS), which has a key role in intracellular signaling as well exerting cytotoxicity. Cells have protective mechanisms against ROS, such as nuclear factor E2-related factor 2 (Nrf2), which controls the expression of many antioxidant enzyme genes. Conversely, BTB and CNC homology 1 (Bach1), a competitor for Nrf2, transcriptionally represses the expression of anti-oxidant enzymes. Previously, we demonstrated that RANKL induces Bach1 nuclear import and attenuates the expression of Nrf2-mediated antioxidant enzymes, thereby augmenting intracellular ROS signaling and osteoclastogenesis. However, it remains unknown if Bach1 inhibitors attenuate osteoclastogenesis. In this study, we hypothesized that Bach1 inhibition would exert an anti-osteoclastogenic effects via diminishing of intracellular ROS signaling through augmented antioxidation. We used RAW 264.7 cells as osteoclast progenitor cells. Using flow cytometry, we found that Bach1 inhibitors attenuated RANKL-mediated ROS generation, which resulted in the inhibition of osteoclastogenesis. Local injection of a Bach1 inhibitor into the calvaria of male BALB/c mice blocked bone destruction induced by lipopolysaccharide. In conclusion, we demonstrate that Bach1 inhibitor attenuates RANKL-mediated osteoclastogenesis and bone destruction in mice by inducing the expression of Nrf2-regulated antioxidant enzymes that consequently decrease intracellular ROS levels. Bach1 inhibitors have potential in inhibiting bone destructive diseases such as periodontitis, rheumatoid arthritis and osteoporosis.

Remarkable Protective Effects of Nrf2-Mediated Antioxidant Enzymes and Tissue Specificity in Different Skeletal Muscles of Daurian Ground Squirrels Over the Torpor-Arousal Cycle.

Hibernating mammals experience conditions of extreme oxidative stress, such as fasting, muscle disuse, and repeated hypoxic ischemia-reperfusion, during the torpor-arousal cycle. Despite this, they experience little oxidative injury and are thus an interesting model of anti-oxidative damage. Thus, in the current study, we explored the levels and underlying mechanism of oxidative stress and antioxidant capacity in three skeletal muscles [slow-twitch soleus (SOL), fast-twitch extensor digitorum longus (EDL), and mixed gastrocnemius (GAS)] of Daurian ground squirrels (Spermophilus dauricus) during hibernation. Results showed that hydrogen peroxide content in the EDL and GAS decreased significantly during pre-hibernation (PRE) and late torpor (LT) compared to levels in the summer active (SA) group. Furthermore, relative to SA levels, malondialdehyde content decreased significantly during interbout arousal (IBA) and early torpor (ET) in all three skeletal muscles and decreased in the EDL and GAS during LT. Compared with the SA group, glutathione peroxidase 1 (GPx1) and catalase (CAT) protein expression in the SOL and superoxide dismutase 1 (SOD1) and SOD2 expression in the GAS increased significantly during the entire hibernation season. Furthermore, SOD1 in the IBA group and CAT and GPx1 in the ET and LT groups increased significantly in the EDL. The activities of most tested antioxidant enzymes were higher in the IBA group than in the LT group, whereas CAT remained highly active throughout the hibernation season in all three muscles. Nrf2 and p-Nrf2 protein levels were significantly elevated in the SOL and EDL during hibernation, and increased during the PRE, IBA, and ET states in the GAS. Thus, activation of the Nrf2/Keap1 antioxidant pathway resulted in the elimination of excess reactive oxygen species (ROS). Specifically, ROS levels were maintained at physiological levels by the up-regulation of antioxidant enzyme expression in skeletal muscles under oxidative stress during hibernation, thus preventing oxidative injury over the torpor-arousal cycle. Different antioxidant patterns and oxidative stress levels were also observed among the different skeletal muscles of hibernating Daurian ground squirrels.

Notoginsenoside R1 Protects Against Diabetic Cardiomyopathy Through Activating Estrogen Receptor α and Its Downstream Signaling.

Diabetic cardiomyopathy (DCM) leads to heart failure and death in diabetic patients, no effective treatment is available. Notoginsenoside R1 (NGR1) is a novel saponin that is derived from Panax notoginseng and our previous studies have showed cardioprotective and neuroprotective effects of NGR1. However, its role in protecting against DCM remains unexplored. Herein, we examine potential effects of NGR1 on cardiac function of diabetic db/db mice and H9c2 cardiomyocytes treated by advanced glycation end products (AGEs). In vitro experiments revealed that pretreatment with NGR1 significantly decreased AGEs-induced mitochondria injury, limited an increase in ROS, and reduced apoptosis in H9c2 cells. NGR1 eliminated ROS by promoting estrogen receptor α expression, which subsequently activated Akt and Nrf2-mediated anti-oxidant enzymes. In vivo investigation demonstrated that NGR1 significantly reduced serum lipid levels, insulin resistance, the expression of enzymes related to cardiomyopathy, and the expression of apoptotic proteins. Finally, NGR1 improved cardiac dysfunction and attenuated histological abnormalities, as evidenced by elevating ejection fraction and fractional shortening, and reducing cardiac fibrosis. Mechanistically, NGR1 promoted ERα expression, which led to the activation of Akt-Nrf2 signaling and the inhibition of the TGFβ pathway. Collectively, these results strongly indicate that NGR1 exerts cardioprotective effects against DCM through its inhibition of oxidative stress and apoptosis, and eventually suppresses cardiac fibrosis and hypertrophy, which suggests that NGR1 is a potential therapeutic medicine for the treatment of DCM.

Protective effects of chebulic acid from Terminalia chebula Retz. against t-BHP-induced oxidative stress by modulations of Nrf2 and its related enzymes in HepG2 cells.

Although chebulic acid isolated from Terminalia chebular has diverse biological effects, its effects on the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and the expression of downstream genes have not been elucidated. The purpose of this research is to investigate the hepatoprotective mechanism of chebulic acid against oxidative stress produced by tert-butyl hydroperoxide (t-BHP) in liver cells. The treatment with chebulic acid attenuated cell death in t-BHP-induced HepG2 liver cells and increased intracellular glutathione content, upregulated the activity of heme oxygenase-1, and also increased the translocation of Nrf2 into the nucleus and Nrf2 target gene expression in a dose-dependent manner. The exposure of chebulic acid activated the phosphorylation of mitogen-activated protein kinases. The overall result is that chebulic acid has cytoprotective effect on t-BHP-induced hepatotoxicity in HepG2 cells through Nrf2-mediated antioxidant enzymes.

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

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.

Different dietary protein levels affect flesh quality, fatty acids and alter gene expression of Nrf2-mediated antioxidant enzymes in the muscle of grass carp (Ctenopharyngodon idella)

This study investigated the effects of dietary protein levels on the flesh quality, fatty acids, antioxidant capacity and antioxidant-related signalling molecule gene expression in the muscle of grass carp (Ctenopharyngodon idella). A total of 540 grass carp (264.11 ± 0.76 g) were fed six diets containing graded levels of protein (143.1, 176.7, 217.2, 257.5, 292.2 and 322.8 g digestible protein kg−1 diet) for 8 weeks. The results indicated that optimal dietary protein levels: (1) improved physical (or chemical) and flavor quality of grass carp fillets by increasing shear force, pH, hydroxyproline, protein, lipid, free amino acids (aspartic acid, threonine, serine, glutamic acid, glycine, alanine, valine, isoleucine, lysine, histidine, arginine and proline) and nucleotide (5′-inosinic acid) contents, and decreasing cooking loss, cathepsin B and L activities and lactate content. (2) enhanced the potential health benefits to humans of grass carp fillets by decreasing the saturated fatty acid (C14:0, C15:0, C16:0, C18:0 and C20:0) concentrations, and increasing unsaturated fatty acids [monounsaturated fatty acids (C14:1 and C16:1) and polyunsaturated fatty acids (C18:2c+t, C18:3n-3, C20:4 and C22:6)] concentrations. (3) enhanced antioxidant capacity of grass carp fillets by decreasing reactive oxygen species, malondialdehyde and protein carbonyl contents, and increasing the activities and gene expression of copper/zinc superoxide dismutase, manganese superoxide dismutase, glutathione peroxidase, glutathione-S-transferase and glutathione reductase and the content of glutathione. (4) regulated antioxidant-related signalling molecules gene expression by up-regulating the mRNA levels of NF-E2-related factor 2, casein kinase 2α and 2β, and decreasing Kelch-like-ECH-associated protein 1a and 1b in the muscle of grass carp. In conclusion, optimal dietary protein level improved the physical and flavor characteristics, fatty acid profile, antioxidant capacity and regulated the gene expression of antioxidant-related signalling molecules in fish fillets.

Chebulic Acid against t‐BHP‐induced effect via Modulations of Nrf2 and Its Related Enzymes in HepG2 cells

Chebulic acid, phenolic compound isolated from the ripe fruits of Terminalia chebular Retz. shows diverse biological effects. The purpose of this study is to investigate protective effects of chebulic acid against tert‐butyl hydroperoxide (t‐BHP) induced oxidative stress and its mechanism related to Nrf2 signal pathway in HepG2 cells. Chebulic acid attenuated cell death and production of reactive oxygen species in t‐BHP‐induced HepG2 cells and increased intracellular glutathione (GSH) content and the ratio of GSH to glutathione disulfide (GSSG). Chebulic acid also upregulated the activity of heme oxygenase‐1 (HO‐1). The treatment with chebulic acid increased the translocation of Nrf2 into the nucleus and the expression of Nrf2 target genes such as γ‐glutamate cysteine ligase and HO‐1 in a dose‐dependent manner. In addition, the exposure of chebulic acid enhanced the phosphorylation of mitogen‐activated protein kinases (MAPK). The overall result is that chebulic acid has cytoprotective effect on t‐BHP‐induced hepatotoxicity in HepG2 cells through Nrf2‐mediated antioxidant enzymes and MAPK.Support or Funding InformationThis work was supported by a Grant from the Institute of Life Science and Natural Resources, Korea University.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Protective effects of total glucosides of Yupingfeng against CCl4 induced hepatic fibrosis in mice by modulating Nrf2-mediated antioxidant enzymes and NOX4

Protective effects of total glucosides of Yupingfeng against CCl4 induced hepatic fibrosis in mice by modulating Nrf2-mediated antioxidant enzymes and NOX4

Evaluation of the Liver Cancer Prevention of Anthocyanin Extracts from Mulberry (<i>Morus alba</i> L.) Variety PR-01

This study aims to evaluate the preventive effects of anthocyanins extracts (MAEs) from mulberry variety PR-01 against N-nitrosodiethylamine (NDEA)-induced hepatocarcinogenesis in rats. It was found that 150 mg·kg-1 MAEs treatment significantly reduced the NDEA-induced hepatic nodules incidence and hepatocellular carcinoma incidence by 58.30% and 41.70% compared to the model group. Meanwhile, MAEs significantly restored the elevated the liver function enzymes, inhibited the tumor necrosis factor alpha and interleukin-6 levels, elevated the serum interleukin-10 and interferon-γ and increased hepatic glutathione-S-transferase and UDP-glucuronosyltransferase 2B1 enzyme activity. Moreover, 150 mg·kg-1 MAEs supplement enhanced glutathione content and the activities of superoxide dismutase, catalase, glutathione peroxidase activities but reduced the malondialdehyde and thiobarbituric acid-reactive substances content by 37.90% and 44.52%. Furthermore, MAEs pretreatment maintained nuclear factor erythroid 2-related factor 2 (Nrf2), Kelch-like ECH-associated protein 1, heme oxygenase-1, and NAD(P)H: quinine oxidoreductase1 stimulation and inhibited the expression of TNF-α, nuclear factor-kappaB (NF-κB), and cyclooxygenase-2 (COX-2), indicating that MAEs exhibit effectively prevention effects against liver cancer via decreased lipid peroxidation, induced Nrf2-mediated antioxidant enzymes and attenuating the inflammatory mediators COX-2 through NF-κB pathway. Thus, MAEs of mulberry variety PR-01 may be used as a good functional dietary supplement against liver cancer.

Effects of diclofenac on the expression of Nrf2 and its downstream target genes in mosquito fish (Gambusia affinis)

Diclofenac (DCF) is one of widely used non-steroidal anti-inflammatory drugs. Recently, this drug has been universally detected in aquatic environment. However, its potential adverse effects and oxidative stress toxic mechanisms on fish remain unclear. In the present study, we first cloned the crucial partial sequences of some key oxidative stress related genes, which include NF-E2-related factor 2 (Nrf2), NAD(P)H: quinoneoxidoreductase (NQO1), glutamate-cysteine ligase catalytic subunit (GCLC), Cu–Zn superoxide dismutase (SOD2), catalase (CAT), alpha-glutathione S-transferase (GSTA), and UDP-glucuronosyltransferases (UGT) in mosquito fish (Gambusia affinis). We also deduced amino acids of Nrf2 and then constructed the phylogenetic trees of Nrf2, NQO1 and GCLC, respectively. Results showed that a high identity percentage was founded between G. affinis and other bony fish species, such as Xiphophorus maculates and Poecilia reticulate. The transcriptional expression of these genes and partly related enzymes activities were then investigated under the included environmental relevant concentration DCF exposure (0μmolL−1, 1.572×10−3μmolL−1, 1.572×10−2μmolL−1, 0.1572μmolL−1 and 1.572μmolL−1) for 24h and 168h. The expression of Nrf2 was inhibited at 24h but induced at 168h, exhibiting a significant time and/or dose-effect relationship under DCF exposure. Similar observation was found in its downstream target genes. However, Nrf2-mediated antioxidant enzymes activities displayed differently under the same concentration of DCF exposure for the same time. Under DCF exposure for 168h, the genes exhibited dramatic induction trend, but there were no significant changes in enzyme activities and MDA content. Overall, mRNA responses were more sensitive than enzyme changes in mosquito fish under DCF exposure.

Fraxin Prevents Chemically Induced Hepatotoxicity by Reducing Oxidative Stress.

Fraxin isolated from Acer tegmentosum is reported to exert potent anti-oxidative stress action. However, pharmacological activities of fraxin remain to be elucidated. This study investigated the potential hepatoprotective effects of fraxin and the underlying signaling mechanism involved. Treatment with fraxin significantly lowered the serum levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in a CCl4-induced hepatotoxicity rat model. In the fraxin-treated group, glutathione (GSH) significantly increased, while the malondialdehyde (MDA) in the liver significantly decreased. Fraxin also showed radical-scavenging activity. Furthermore, it significantly reduced the t-BHP-induced cytotoxicity and production of reactive oxygen species (ROS) in Hep G2. Fraxin protected Hep G2 cells through Nrf2 pathway-dependent HO-1 expression. The results of this study indicate that fraxin shows potent hepatoprotective effects in vitro and in vivo, presumably through direct antioxidant activity and the Nrf2-mediated antioxidant enzyme system.

DJ-1 regulating PI3K-Nrf2 signaling plays a significant role in bibenzyl compound 20C-mediated neuroprotection against rotenone-induced oxidative insult

Oxidative stress is thought to be involved in the development of Parkinson’s disease (PD). We previously reported that 20C, a bibenzyl compound isolated from Gastrodia elata, possesses antioxidative properties, but its in-depth molecular mechanisms against rotenone-induced neurotoxicity remains unknown. Recent studies indicate that without intact DJ-1, nuclear factor erythroid 2-related factor (Nrf2) protein becomes unstable, and the activity of Nrf2-mediated downstream antioxidant enzymes are thereby suppressed. In this study, we showed that 20C clearly protected PC12 and SH-SY5Y cells against rotenone-induced oxidative injury. Furthermore, 20C markedly up-regulated the levels of DJ-1, which in turn activated phosphoinositide-3-kinase (PI3K)/Akt signaling and inhibited glycogen synthase kinase 3β (GSK3β) activation, eventually promoted the nuclear translocation of Nrf2 and induced the expression of hemeoxygenase-1 (HO-1). The antioxidant effects of 20C could be partially blocked by ShRNA-mediated knockdown of DJ-1 and inhibition of the PI3K/Akt pathways with Akt1/2 kinase inhibitor, respectively. Conclusively, our findings confirm that DJ-1 is necessary for 20C-mediated protection against rotenone-induced oxidative damage, at least in part, by activating PI3K/Akt signaling, and subsequently enhancing the nuclear accumulation of Nrf2. The findings from our investigation suggest that 20C should be developed as a novel candidate for alleviating the consequences of PD in the future.

Saponins from the roots of Platycodon grandiflorum ameliorate high fat diet-induced non-alcoholic steatohepatitis.

Platycodon grandiflorum has been healthy effects due to its various nutritious compounds and is considered as a functional food. Platycodon grandiflorum root-derived saponins (CKS) have been reported to show a variety of effects including anti-inflammatory and anti-oxidative activity. Although CKS have been studied on various bioactivities, the inhibitory effect of CKS on non-alcoholic steatohepatitis (NASH) is not examined. In this study, the inhibitory effects on HFD-induced NASH by CKS were determined. CKS suppressed HFD-induced hepatic lipid peroxidation level, collagen deposition, pro-fibrogenic and pro-inflammatory cytokines expression. CKS treatment suppressed HFD-induced COX-2 expression via inhibition of NF-κB p65 nuclear translocation and IκBα degradation. CKS treatment restored HFD-reduced Nrf2-mediated antioxidant enzymes expression. Furthermore, CKS treatment reinstated HFD-reduced peroxisomal proliferator-activated receptor alpha (PPARα)-regulated acyl-coA oxidase and carnitine-palmitoyl-coA transferase-1 expression. These findings suggest that CKS reduces HFD-induced NASH by up-regulation of Nrf2-mediated anti-oxidant enzymes and PPARα-regulated fatty acid oxidation.

RANKL induces Bach1 nuclear import and attenuates Nrf2-mediated antioxidant enzymes, thereby augmenting intracellular reactive oxygen species signaling and osteoclastogenesis in mice.

Reactive oxygen species (ROS) play a role in intracellular signaling during osteoclastogenesis. We previously reported that transcriptional factor nuclear factor E2-related factor 2 (Nrf2) was exported from the nucleus to the cytoplasm by receptor activator of nuclear factor-κB ligand (RANKL), and that Nrf2 negatively regulated osteoclastogenesis via antioxidant enzyme up-regulation. Knockout mice of BTB and CNC homology 1 (Bach1)-the competitor for Nrf2 in transcriptional regulation-was known to attenuate RANKL-mediated osteoclastogenesis, although the mechanism remains unclear. Therefore, we hypothesized that RANKL could be involved in the nuclear translocation of Bach1, which would attenuate Nrf2-mediated antioxidant enzymes, thereby augmenting intracellular ROS signaling in osteoclasts. RANKL induced Bach1 nuclear import and Nrf2 nuclear export. Induction of Bach1 nuclear export increased Nrf2 nuclear import, augmented antioxidant enzyme expression, and, thus, diminished RANKL-mediated osteoclastogenesis via attenuated intracellular ROS signaling. Finally, an in vivo mouse bone destruction model clearly demonstrated that induction of Bach1 nuclear export inhibited bone destruction. In this study, we report that RANKL favors osteoclastogenesis via attenuation of Nrf2-mediated antioxidant enzyme expression by competing with Bach1 nuclear accumulation. Of importance, induction of Bach1 nuclear export activates Nrf2-dependent antioxidant enzyme expression, thereby attenuating osteoclastogenesis. Bach1 nuclear export might be a therapeutic target for such bone destructive diseases as rheumatoid arthritis, osteoporosis, and periodontitis.-Kanzaki, H., Shinohara, F., Itohiya, K., Yamaguchi, Y., Katsumata, Y., Matsuzawa, M., Fukaya, S., Miyamoto, Y., Wada, S., Nakamura, Y. RANKL induces Bach1 nuclear import and attenuates Nrf2-mediated antioxidant enzymes, thereby augmenting intracellular reactive oxygen species signaling and osteoclastogenesis in mice.

Hepatoprotective efficacy of gallic acid during Nitrosodiethylamine-induced liver inflammation in Wistar rats

Gallic acid (GA), a popular phenolic acid is found in gallnuts, grapes, pomegranates, tea and oak bark. It possesses anti-cancer, anti-bacterial, anti-depressant, anti-asthmatic and anti-obesity effects. N′-Nitrosodiethylamine (NDEA) is a well-known hepatotoxin, carcinogen and mutagen. In this study, we have examined the hepatoprotective effect of gallic acid against liver inflammation induced by NDEA in Wistar rats. Hepatic damage in the animals was induced by 10mlkg−1b.wt of 1% NDEA (i.p.) solution in normal saline once in a week. Another group received GA supplement (i.p.) in 100mgkg−1b.wtwk−1. Animals belonging to control group were administered equal amounts of saline or GA. LPO, SOD and membrane-bound ATPase (Ca2+- and Mg2+-ATPase) activities were determined in liver homogenate of control and treated rats. Alterations in liver architecture were assessed by H&E and Masson’s trichrome stainings of 5μm thick liver sections. Immunohistochemistry (IHC) was performed to localize the inflammatory marker, Cyclooxygenase-2 (COX-2). Our results demonstrate a significant increase in malondialdehyde, and decrease in SOD and ATPases (Ca2+/Mg2+) in NDEA-treated rats. Histopathology data showed inflammation, activated HSCs, deposition of collagen, periportal as well as bridging fibrosis in NDEA-treated liver specimens. Immunohistochemistry of NDEA-treated liver sections exhibited COX-2 positive cells. Gallic acid supplement revert the hepatic functioning in rats injured with NDEA probably by inducing Nrf2-mediated antioxidant enzymes and attenuating the inflammatory mediators COX-2 through NF-κB inhibition pathway. Therefore, gallic acid supplement may be a useful promising bioagent in combating liver injury.

Neuroprotective and Cognition-Enhancing Effects of Compound K Isolated from Red Ginseng.

The present study was aimed at elucidating the effect of compound K derived from red ginseng on memory function in mouse model and glutamate-induced cytotoxicity in mouse hippocampal HT22 cells. Compound K induced antioxidant enzymes in nuclear factor (erythroid-derived 2)-like 2 (Nrf2)-mediated manner, and effectively attenuated cytotoxicity and mitochondrial damage induced by glutamate in HT22 cells. However, the cytoprotective effect by compound K was abolished by heme oxygenase-1 inhibitor, tin protophorphyrin IX, suggesting that neuroprotective effect of compound K was caused by its Nrf2-mediated induction of antioxidant enzymes. Further, memory deficit induced by scopolamine was restored by compound K, which did not inhibit acetylcholine esterase, in C57BL/6 mice but not in Nrf2 knockout mice as assessed by passive avoidance test, Y-maze and water maze tests, suggesting that scopolamine-induced memory impairment was overcome by the induction of Nrf2-mediated antioxidant enzymes by the compound K. Overall, our data indicate that compound K could be useful in prevention and treatment of reactive oxygen species-induced neurological disorders such as Alzheimer's disease.

Celastrol enhances Nrf2 mediated antioxidant enzymes and exhibits anti-fibrotic effect through regulation of collagen production against bleomycin-induced pulmonary fibrosis

Pulmonary fibrosis (PF) is characterized by excessive accumulation of extracellular matrix components in the alveolar region which distorts the normal lung architecture and impairs the respiratory function. The aim of this study is to evaluate the anti-fibrotic effect of celastrol, a quinine-methide tri-terpenoid mainly found in Thunder God Vine root extracts against bleomycin (BLM)-induced PF through the enhancement of antioxidant defense system. A single intratracheal instillation of BLM (3 U/kg.bw) was administered in rats to induce PF. Celastrol (5 mg/kg) was given intraperitoneally, twice a week for a period of 28 days. BLM-induced rats exhibits declined activities of enzymatic and non-enzymatic antioxidants which were restored upon treatment with celastrol. BLM-induced rats show increased total and differential cell counts as compared to control and celastrol treated rats. Histopathological analysis shows increased inflammation and alveolar damage; while assay of hydroxyproline and Masson's trichrome staining shows an increased collagen deposition in BLM-challenged rats that were decreased upon celastrol treatment. Celastrol also reduces inflammation in BLM-induced rats as evidenced by decrease in the expressions of mast cells, Tumor necrosis factor-alpha (TNF- α) and matrix metalloproteinases (MMPs) 2 and 9. Further, Western blot analysis shows that celastrol is a potent inducer of NF-E2-related factor 2 (Nrf2) and it restores the activities of Phase II enzymes such as hemoxygenase-1 (HO-1), glutathione-S-transferase (GSTs) and NADP(H): quinine oxidoreductase (NQO1) which were declined upon BLM administration. The results of this study show evidence on the protective effect of celastrol against BLM-induced PF through its antioxidant and anti-fibrotic effects.

Dietary leucine improves flesh quality and alters mRNA expressions of Nrf2-mediated antioxidant enzymes in the muscle of grass carp (Ctenopharyngodon idella)

The objective of the study was to assess the influence of dietary leucine on flesh quality and the mRNA level of Nrf2-mediated antioxidant enzymes in the muscle of grass carp (Ctenopharyngodon idella). A total of 540 grass carp (295.85±2.07g) were fed six diets with graded levels of leucine (7.1, 8.9, 11.0, 13.3, 15.2 and 17.1gkg−1 diet) for eight weeks. The result indicated that muscle protein, lipid content, muscle EAA (essential amino acid) levels, shear force, and hydroxyproline concentration were significantly improved by 11.0–17.1, 8.9–17.1, 8.9–17.1, 11.0–13.3, 8.9–13.3gleucinekg−1 diet supplementations, respectively. On the other hand, cooking loss, activities of cathepsin L, malondialdehyde and protein carbonyl contents exhibited an opposite tendency. Moreover, the results indicated that compared with leucine unsupplemented group (7.1gleucinekg−1 diet), 8.9–17.1, 11.0–17.1, 11.0–15.2 and 8.9–15.2gkg−1 leucine supplementations significantly enhanced the activities of copper/zinc superoxide dismutase (SOD1), catalase (CAT) and glutathione peroxidase (GPx), and glutathione content, respectively. Furthermore, the changes in the relative mRNA expressions of SOD1, CAT, GPx, nuclear factor erythroid 2-related factor 2 (Nrf2), target of rapamycin (TOR), and ribosomal protein S6 kinase beta-1 (S6K1) induced by leucine were similar to those noted in their enzyme activities. Conversely, the mRNA levels of Kelch-like ECH-associated protein 1 (Keap1) were significantly down-regulated by leucine supplementation. In conclusion, optimum leucine improved flesh quality, partly due to the enhancement of antioxidant capacity through regulating antioxidant-related signaling molecule. Based on muscle MDA, the optimum dietary leucine requirements of young grass carp (296–690g) for flesh quality were estimated to be 12.8gkg−1 diet, corresponding to 39.3gkg−1 of dietary protein.