NADPH Quinine Oxidoreductase Research Articles

Anti-Osteoporotic Effect of Viscozyme-Assisted Polysaccharide Extracts from Portulaca oleracea L. on H2O2-Treated MC3T3-E1 Cells and Zebrafish

This study aims to screen and characterize the protective effect of polysaccharides from Portulaca oleracea L. (POP) against H2O2-stimulated osteoblast apoptosis in vivo and in vitro. The enzymes viscozyme, celluclast, α-amylase, and β-glucanase were used to extract POPs. Among all enzyme-assisted POPs, the first participating fraction of viscozyme extract POP (VPOP1) exhibited the highest antioxidant activity. Hoechst 33342 and acridine orange/ethidium bromide staining and flow cytometry of MC3T3 cells revealed that VPOP1 inhibited apoptosis in a dose-dependent manner. Moreover, VPOP1 increased the expression levels of heme oxygenase-1 (HO-1) and NADPH quinine oxidoreductase 1 (NQO1) and decreased the expression levels of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) and Kelch-like ECH-associated protein 1 (Keap1) in H2O2-induced cells compared with their controls. The results of an in vivo experiment show that VPOP1 significantly reduced reactive oxygen species generation and lipid peroxidation in zebrafish at 72 h post-fertilization and promoted bone growth at 9 days post-fertilization. Furthermore, VPOP1 was identified via 1-phenyl-3-methyl-5-pyrazolone derivatization as an acidic heteropolysaccharide comprising mannose and possessing a molecular weight of approximately 7.6 kDa. Collectively, VPOP1 was selected as a potential anti-osteoporotic functional food because of its protective activity against H2O2-induced damage in vitro and in vivo.

Cancer Chemopreventive Role of Dietary Terpenoids by Modulating Keap1-Nrf2-ARE Signaling System—A Comprehensive Update

ROS, RNS, and carcinogenic metabolites generate excessive oxidative stress, which changes the basal cellular status and leads to epigenetic modification, genomic instability, and initiation of cancer. Epigenetic modification may inhibit tumor-suppressor genes and activate oncogenes, enabling cells to have cancer promoting properties. The nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcription factor that in humans is encoded by the NFE2L2 gene, and is activated in response to cellular stress. It can regulate redox homoeostasis by expressing several cytoprotective enzymes, including NADPH quinine oxidoreductase, heme oxygenase-1, UDP-glucuronosyltransferase, glutathione peroxidase, glutathione-S-transferase, etc. There is accumulating evidence supporting the idea that dietary nutraceuticals derived from commonly used fruits, vegetables, and spices have the ability to produce cancer chemopreventive activity by inducing Nrf2-mediated detoxifying enzymes. In this review, we discuss the importance of these nutraceuticals in cancer chemoprevention and summarize the role of dietary terpenoids in this respect. This approach was taken to accumulate the mechanistic function of these terpenoids to develop a comprehensive understanding of their direct and indirect roles in modulating the Keap1-Nrf2-ARE signaling system.

Effect of Nrf2/HO-1 signaling pathway in intestinal protection by Sishen Pills against ulcerative colitis in mice

The present study aimed to explore the effect of nuclear factor erythroid 2 related factor 2(Nrf2)/heme oxygenase-1(HO-1) signaling pathway in intestinal protection by Sishen Pills against ulcerative colitis(UC). After the UC model was induced by 3% dextran sodium sulfate(DSS), experimental animals were randomly divided into control group, model group, salazosulfapyridine(SASP) group, and low-and high-dose Sishen Pills groups. Drug intervention(ig) was performed for seven consecutive days during modeling. On the 7 th day, the mice were euthanized. The body weight and colon length were recorded, and the histopathological changes of the colon were observed by HE staining. Serum interleukin-6(IL-6), tumor necrosis factor-α(TNF-α), total antioxidant capacity(T-AOC), malondialdehyde(MDA), and reactive oxygen species(ROS) were detected by ELISA. The protein and mRNA expression of Nrf2, HO-1, and NADPH quinine oxidoreductase-1(NQO-1) was determined by Western blot and reverse transcription-polymerase chain reaction(RT-PCR). Compared with the normal group, the model group exhibited reduced body weight, colon length, and T-AOC, increased IL-6, TNF-α, MDA, and ROS, and diminished protein and mRNA expression of Nrf2, HO-1, and NQO-1 in the colon tissues. Compared with the model group, the SASP group and high-dose Sishen Pills group showed elevated body weight, colon length, and T-AOC, lowered IL-6, TNF-α, MDA, and ROS levels, and increased protein and mRNA expression of Nrf2, HO-1, and NQO-1 in the colon tissues. As assessed by HE staining, Sishen Pills could improve the pathological changes of the colon. The findings suggested that Sishen Pills could protect the colon against UC induced by 3% DSS. The specific mechanism of action may be related to the anti-inflammatory and anti-oxidative stress effects by the activation of the Nrf2/HO-1 signaling pathway.

Mangiferin Promotes Bregs Level, Activates Nrf2 Antioxidant Signaling, and Inhibits Proinflammatory Cytokine Expression in Murine Splenic Mononuclear Cells In Vitro.

Recent studies indicated that regulatory B cells (Bregs) and nuclear factor erythroid 2-related factor 2 (Nrf2) antioxidant signaling pathway play important roles in the pathogenesis of chronic graft-versus-host disease (cGVHD). Mangiferin (MA), a polyphenol compound, has been reported to activate Nrf2/antioxidant-responsive element (ARE) signaling pathway. This study was aimed to investigate the effects of MA on Bregs and Nrf2 antioxidant signaling in murine splenic mononuclear cells (MNCs) in vitro. Our results revealed that MA could increase the Bregs level in murine splenic MNCs. Moreover, MA up-regulated the expression of Bregs-associated immunosuppressive factor interleukin-10 (IL-10) by activating the Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) and extracellular signal-regulated kinase (ERK) signaling in murine splenic MNCs. Meanwhile, MA inhibited the proinflammatory cytokines IL-2 and interferon-γ (INF-γ) at both mRNA and protein levels. MA also enhanced the transcription and protein expression of Nrf2 and NADPH quinine oxidoreductase 1 (NQO1), whereas decreased that of Kelch-like ECH-associated protein 1 (Keap1) in murine splenic MNCs. Moreover, MA promoted the proliferation and inhibited the apoptosis of murine splenic MNCs. These results suggested that MA exerts immunosuppressive effects by upregulating the Bregs level, activating the Nrf2 antioxidant pathway, and inhibiting the expression of pro-immunoinflammatory factors. MA, as a natural immunomodulatory and anti-inflammatory agent, may have a potential role in the prophylaxis and treatment of cGVHD.

Rutin-loaded liquid crystalline nanoparticles attenuate oxidative stress in bronchial epithelial cells: a PCR validation.

Aim: In the present study, the inhibitory potential of rutin-loaded liquid crystalline nanoparticles (LCNs) on oxidative stress was determined in human bronchial epithelial cells (BEAS-2B) by analysing the expression levels of different antioxidant (NADPH quinine oxidoreductase-1(NQO1); γ-glutamyl cysteine synthetase catalytic subunit (GCLC)) and pro-oxidant (NADPH oxidase (Nox)-4; Nox2B) genes. Results: Our findings revealed that the rutin-loaded LCNs inhibited the genes, namelyNox2B and Nox4, which caused oxidative stress. In addition, these nanoparticles demonstrated an upregulation in the expression of the antioxidant genesGclc and Nqo-1 in a dose-dependent manner. Conclusion: The study indicates the promising potential of rutin-loaded LCNs as an effective treatment strategy in patients with high oxidant loadsin various respiratory diseases.

Astaxanthin protects mesenchymal stem cells from oxidative stress by direct scavenging of free radicals and modulation of cell signaling

Recent evidence has shown that mesenchymal stem cells (MSCs) play vital roles in cell therapy of ischemia/hypoxia damaged tissues. However, after the transplantation, they might undergo apoptosis due to oxidative stress. Thus, some strategies have been developed to support stem cells in harsh conditions, including pre-treatment of the cells with antioxidants. Of various antioxidants, in this study, astaxanthin (ATX) was used to protect adipose-derived MSCs against oxidative stress. The MSCs were exposed to different doses of hydrogen peroxide, and then the expression of key genes involved in the redox signaling pathway was studied, including nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), and NADPH quinine oxidoreductase 1 (NQO1). The balance of intracellular reactive oxygen species was detected with the H2DCFDA molecular probe. Additionally, for the detection of apoptosis and protective effect of ATX, the DAPI/Phallacidin and annexin V cell staining were performed. The results of cellular studies revealed that ATX reduced the H2O2-induced cell apoptosis and oxidative stress. Furthermore, after the induction of oxidative stress, the cells’ native antioxidants (HO-1 and NQO1) were overexpressed but they were modulated with ATX treatments (p < 0.023). Based on our findings, ATX could increase the expression of Nrf2 as a key transcription factor of antioxidant enzymes (p < 0.05). These findings support the notion that ATX can act as an effective antioxidant in the pre-treatment of MSCs before cell therapy. Thus, to enhance the viability of stem cells during the transplantation in harsh conditions, the concurrent use of ATX in cell therapy modalities is proposed.

Radiosensitization by Kinase Inhibition Revealed by Phosphoproteomic Analysis of Pancreatic Cancer Cells

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive cancers and known for its extensive genetic heterogeneity, high therapeutic resistance, and strong variation in intrinsic radiosensitivity. To understand the molecular mechanisms underlying radioresistance, we screened the phenotypic response of 38 PDAC cell lines to ionizing radiation. Subsequent phosphoproteomic analysis of two representative sensitive and resistant lines led to the reproducible identification of 7,800 proteins and 13,000 phosphorylation sites (p-sites). Approximately 700 p-sites on 400 proteins showed abundance changes after radiation in all cell lines regardless of their phenotypic sensitivity. Apart from recapitulating known radiation response phosphorylation markers such as on proteins involved in DNA damage repair, the analysis uncovered many novel members of a radiation-responsive signaling network that was apparent only at the level of protein phosphorylation. These regulated p-sites were enriched in potential ATM substrates and in vitro kinase assays corroborated 10 of these. Comparing the proteomes and phosphoproteomes of radiosensitive and -resistant cells pointed to additional tractable radioresistance mechanisms involving apoptotic proteins. For instance, elevated NADPH quinine oxidoreductase 1 (NQO1) expression in radioresistant cells may aid in clearing harmful reactive oxygen species. Resistant cells also showed elevated phosphorylation levels of proteins involved in cytoskeleton organization including actin dynamics and focal adhesion kinase (FAK) activity and one resistant cell line showed a strong migration phenotype. Pharmacological inhibition of the kinases FAK by Defactinib and of CHEK1 by Rabusertib showed a statistically significant sensitization to radiation in radioresistant PDAC cells. Together, the presented data map a comprehensive molecular network of radiation-induced signaling, improves the understanding of radioresistance and provides avenues for developing radiotherapeutic strategies.

Tannic acid attenuates hepatic oxidative stress, apoptosis and inflammation by activating the Keap1‑Nrf2/ARE signaling pathway in arsenic trioxide‑toxicated rats

The present study was performed to investigate the protective effects of tannic acid (TA) on liver injury induced by arsenic trioxide (ATO) and to elucidate the mechanism involved as related to the Kelch‑like ECH‑associated protein 1 (Keap1)‑nuclear factor erythroid 2‑related factor2 (Nrf2)/antioxidant response element (ARE) signaling pathway. Adult rats were intraperitoneally injected with TA, while ATO was administered 1h later. On the 11th day, the rats were euthanized to determine any liver histological changes, liver function, and the activities of antioxidant, antiapoptosis and proinflammatory cytokines in the liver. Furthermore, the protein expression levels of nuclear Nrf2, total Nrf2, Keap1, Heme oxygenase‑1 (HO‑1), NADPH quinine oxidoreductase‑1 (NQO1), and γ‑glutamylcysteine synthetase (γ‑GCS) were determined using western blot analysis. The results showed that TA treatment ameliorated ATO‑induced liver histological changes and decreased the ATO‑induced increased alanine aminotransferase (ALT) and aspartate transaminase (AST) serum levels. Activities of the antioxidant enzymes significantly were increased, while the levels of malondialdehyde (MDA) and reactive oxygen species (ROS) were attenuated following TA treatment. In addition, TA treatment inhibited ATO‑induced liver apoptosis and inflammatory responses, increased Bcl‑2 protein expression level and reduced the levels of Bax, caspase‑3, interleukin (IL)‑1β, IL‑6 and tumor necrosis factor (TNF)‑α. Furthermore, TA treatment increased the protein expression levels of Nrf2 and Keap1, HO‑1, NQO1 and γ‑GCS. The results demonstrated that TA has a protective effect on ATO‑treated hepatic toxicity and that its underlying mechanism could be due to TA activation of the Keap1‑Nrf2/ARE signaling pathway, to reduce oxidative stress, apoptosis and inflammation in ATO‑intoxicated rats.

Nur77 attenuates inflammatory responses and oxidative stress by inhibiting phosphorylated IκB-α in Parkinson's disease cell model.

Neuroinflammation and oxidative stress play key roles in the pathological development of Parkinson’s disease (PD). Nerve growth factor-induced gene B (Nur77) is closely related to dopamine neurotransmission, and its pathogenesis is unclear. This study aims to investigate the role and mechanism of Nur77 in a cell model of Parkinson’s disease. Silencing Nur77 with siRNA can aggravate intracellular LDH release, increase the expression of pro-inflammatory genes (such as tumor necrosis factor α, nuclear factor κB (p65), monocyte chemotactic protein 1, interleukin-6), and decrease cell survival, decrease expression of nuclear factor E2-related factor(Nrf2), heme oxygenase 1, NADPH quinineoxidoreductase-1. Cytosporone B (Nur77 agonist) has the opposite effect to Nur77 silencing. PDTC (NF-κB inhibitor / antioxidant) can also inhibit pro-inflammatory genes to a similar degree as Cytosporone B. Phosphorylated IκB-α can be inhibited by Cytosporone B, while silencing Nur77 can increase the protein expression level of phosphorylated IκB-α. After silencing IκB-α, both Cytosporone B and siNur77 did not affect pro-inflammatory genes and antioxidant stress. These findings reveal the first evidence that Nur77 exerts anti-inflammatory and antioxidant stress effects by inhibiting IκB-α phosphorylation expression in a Parkinson cell model. Nur77 may be a potential therapeutic target for Parkinson’s disease.

Luteolin Exerts Neuroprotection via Modulation of the p62/Keap1/Nrf2 Pathway in Intracerebral Hemorrhage

Upregulation of neuronal oxidative stress is involved in the progression of secondary brain injury (SBI) following intracerebral hemorrhage (ICH). In this study, we investigated the potential effects and underlying mechanisms of luteolin on ICH-induced SBI. Autologous blood and oxyhemoglobin (OxyHb) were used to establish in vivo and in vitro models of ICH, respectively. Luteolin treatment effectively alleviated brain edema and ameliorated neurobehavioral dysfunction and memory loss in vivo. Also, in vivo, we found that luteolin promoted the activation of the sequestosome 1 (p62)/kelch‐like enoyl-coenzyme A hydratase (ECH)‐associated protein 1 (Keap1)/nuclear factor erythroid 2-related factor 2 (Nrf2) pathway by enhancing autophagy and increasing the translocation of Nrf2 to the nucleus. Meanwhile, luteolin inhibited the ubiquitination of Nrf2 and increased the expression levels of downstream antioxidant proteins, such as heme oxygenase-1 (HO-1) and reduced nicotinamide adenine dinucleotide phosphate (NADPH): quinine oxidoreductase 1 (NQO1). This effect of luteolin was also confirmed in vitro, which was reversed by the autophagy inhibitor, chloroquine (CQ). Additionally, we found that luteolin inhibited the production of neuronal mitochondrial superoxides (MitoSOX) and alleviated neuronal mitochondrial injury in vitro, as indicated via tetrachloro-tetraethylbenzimidazol carbocyanine-iodide (JC-1) staining and MitoSOX staining. Taken together, our findings demonstrate that luteolin enhances autophagy and anti-oxidative processes in both in vivo and in vitro models of ICH, and that activation of the p62-Keap1-Nrf2 pathway, is involved in such luteolin-induced neuroprotection. Hence, luteolin may represent a promising candidate for the treatment of ICH-induced SBI.

Polysaccharides from Enteromorpha prolifera protect against carbon tetrachloride-induced acute liver injury in mice via activation of Nrf2/HO-1 signaling, and suppression of oxidative stress, inflammation and apoptosis.

To investigate whether polysaccharides from Enteromorpha prolifera (EPP) could protect against acute hepatic injury induced by CCl4, ICR mice were pretreated with EPP (150, 300, and 450 mg kg-1) and silymarin (100 mg kg-1) for 28 days before CCl4 induction. Pretreatment with EPP attenuated CCl4-induced elevated serum transaminase activities and histopathological alterations in the liver. In addition, EPP prevented CCl4-induced reduction of protein levels of phosphorylated nuclear factor E2-related factor 2 (p-Nrf2)/Nrf2, heme oxygenase-1 (HO-1), and mRNA levels of NADPH quinineoxidoreductase-1 (NQO-1), which, in turn, reduced hepatic oxidative stress injury. Furthermore, the hepatic protein levels of inflammatory mediators and the phosphorylation of nuclear factor-kappaB p65 (NF-κB p65) and I kappaB alpha (IκBα), and the mRNA levels of Toll-like receptor 2 (TLR2), TLR4, and prolyl-isomerase-1 (Pin-1) in the inflammatory signaling pathway were recovered in the EPP pretreated groups. Moreover, EPP prevented the hepatocellular apoptotic changes with inhibition of B-cell lymphoma 2 (Bcl-2), and the induction of Bcl-2-associated X (Bax) and Cleaved caspase-3 caused by CCl4. Taken together, these results indicated that EPP protected against hepatic injury induced by CCl4-derived reactive intermediates through the activation of Nrf2/HO-1 signaling, and suppression of oxidative stress, inflammation and apoptosis.

Theaflavins alleviate sevoflurane-induced neurocytotoxicity via Nrf2 signaling pathway

Aim: Sevoflurane could induce apoptosis of rat hippocampal neurons, while theaflavins (TFs) have antioxidant and anti-inflammatory properties. This study aims to explore whether TFs could alleviate sevoflurane-induced neuronal cell injury.Materials and methods: Cells were treated by concentration gradient of sevoflurane and TFs. Cell viability, level of reactive oxygen species (ROS) and apoptosis rate were determined by cell counting kit-8 (CCK-8) and flow cytometry, respectively. Quantitative PCR (qPCR) and western blot were performed to determine mRNA and protein expressions.Results: TFs promoted viability of cells under the treatment of sevoflurane, while it suppressed apoptosis and down-regulated ROS level in a concentration-dependent manner. TFs could also down-regulate expression levels of caspase-3 and caspase-9 and cytosol and intranuclear nuclear factor E2-related factor 2 (Nrf2) in rat hippocampal nerve cells, while it up-regulated those of heme oxygenase 1 (HO-1), NADPH quinine oxidoreductase 1 (NQO1), glutamate cysteine ligase (GCL) and peroxiredoxin 1 (Prx1).Conclusions: Our study suggests that TFs exert protective effects on sevoflurane-induced neurocytotoxicity and therefore could be used as a potential drug for treatment of neuronal injury.

The antioxidant system mediated by Nrf2 in C2C12 cells responding to H2O2 stimulus under different oxygen concentration

To apply hypoxia of different oxygen concentration on C2C12 cells to study the changes of Nrf2 antioxidant system under H2O2. The perfect simulative effect time and concentration of H2O2 were chosen. Cell vitality was tested after C2C12 cells cultured in 0.1 mmol/L, 0.25 mmol/L, 0.5 mmol/L, 0.75 mmol/L, 1 mmol/L and 2 mmol/L H2O2 for 1 or 2 h respectively. The C2C12 cells were divided into different oxygen concentration group: 21%O2, 12%O2, 8%O2, 5%O2 respectively. And then cells were treated with H2O2 for 1 h, and collected for determination. Immunofluorescence of Nrf2 and the protein expression of Nrf2 were detected. The expressions of antioxidant enzymes superoxide dismutase 1 (SOD1), superoxide dismutase 2 (SOD2), catalase(CAT), NADPH quinine oxidoreductase-1 (NQO-1), glutathione peroxidase-1 (GPX-1), Heme oxygenase-1 (HO-1) mRNA and cellular ROS levels were tested by high quality fluorescence assay. 0.5 mmol/L H2O2 for 1 h was selected as the conditions of H2O2stimulation. Compared with 21% O2 group, the expressions of Nrf2 mRNA and protein, antioxidant enzymes SOD1, SOD2, CAT, HO-1, NQO-1, GPX-1 mRNA were increased significantly (P＜0.05 or P＜0.01), and ROS level was lower (P＜0.01) in 12%O2 group cells; only the expression of GPX-1 mRNA was increased (P＜0.05) in 8%O2 group; the expressions of Nrf2 mRNA and protein expression, antioxidant enzymes SOD1, SOD2, NQO-1, GPX-1 mRNA were decreased significantly(P＜0.05 or P＜0.01), and ROS level was higher (P＜0.01) in 5%O2 group. Hypoxia can affect the Nrf2 antioxidant system, and the different oxygen concentrations have different impact. In addition, 12% O2 for 12 h could promote the Nrf2 antioxidant system, and 5% extremely low oxygen may inhibit it.

Spatholobus suberectus Ameliorates Diabetes-Induced Renal Damage by Suppressing Advanced Glycation End Products in db/db Mice.

Spatholobus suberectus (SS) is a medicinal herb commonly used in Asia to treat anemia, menoxenia and rheumatism. However, its effect of diabetes-induced renal damage and mechanisms of action against advanced glycation end-products (AGEs) are unclear. In this study, we evaluated the effects of SS on diabetes-induced renal damage and explored the possible underlying mechanisms using db/db type 2 diabetes mice. db/db mice were administered SS extract (50 mg/kg) orally for 6 weeks. SS-treated group did not change body weight, blood glucose and glycated hemoglobin (HbA1c) levels. However, SS treatment reversed diabetes-induced dyslipidemia and urinary albumin/creatinine ratio in db/db mice. Moreover, SS administration showed significantly increased protein expression of nuclear factor erythroid 2-related factor 2 (Nrf2), which is a transcription factor for antioxidant enzyme. SS significantly upregulated glyoxalase 1 (Glo1) and NADPH quinine oxidoreductase 1 (NQO1) expression but reduced CML accumulation and downregulated receptor for AGEs (RAGE). Furthermore, SS showed significant decrease of periodic acid–Schiff (PAS)-positive staining and AGEs accumulation in histological and immunohistochemical analyses of kidney tissues. Taken together, we concluded that SS ameliorated the renal damage by inhibiting diabetes-induced glucotoxicity, dyslipidemia and oxidative stress, through the Nrf2/antioxidant responsive element (ARE) stress-response system.

FTY720 inhibits the Nrf2/ARE pathway in human glioblastoma cell lines and sensitizes glioblastoma cells to temozolomide.

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a redox-sensitive transcription factor regulating the expression of various cytoprotective genes. Constitutive Nrf2 activation in many cancers enhances cell survival and resistance to anti-cancer drugs. Our previous studies have shown that FTY720 induced autophagy-related apoptosis and necroptosis and inhibited invasion and migration in human glioblastoma cells. However, whether FTY720 regulated Nrf2 in glioblastoma cells remained unclear. Cell viability assay, western blot, migration and invasion assay, short hairpin RNA (shRNA) were used. We found that FTY720 suppressed the protein and mRNA levels of Nrf2 in human U251MG and U87MG glioblastoma cells lines. In addition, the protein and mRNA levels of heme oxygenase-1 (HO-1) and NADPH:quinine oxidoreductase-1 (NQO-1), two representative target factors of Nrf2, also decreased upon FTY720 treatment. Knockdown of Nrf2 further promoted the anti-cancer effects of FTY720, while activation of Nrf2 exist the opposite effects. In addition, FTY720 significantly sensitized glioblastoma cells to temozolomide (TMZ). However, activation of Nrf2 essentially abolished the induced sensitivity by FTY720. Our results indicated the potential application of FTY720 in treatment of glioblastoma and demonstrated that inhibition of Nrf2 can enhance the sensitivity of cancer cells to chemotherapeutic drugs.

Omega-3 polyunsaturated fatty acids ameliorates testicular ischemia-reperfusion injury through the induction of Nrf2 and inhibition of NF-κB in rats

ObjectiveThis study was designed to investigate the protective effect of Omega-3 polyunsaturated fatty acids (n-3 PUFAs) on testicular ischemia-reperfusion (I/R) injury in rats. MethodsA total of 24 rats were randomly divided into the following three groups: Group A (Control group, n=8), Group B (I/R group, n=8), Group C (I/R group treated with n-3 PUFAs, n=8). Histological examination was used to assess changes in testicular structure. Tissue oxidative stress biomarkers, malondialdehyde (MDA) as well as Superoxide, and antioxidant indexes, including total antioxidant capacity (T-AOC); catalase (CAT); glutathione (GSH); glutathione/glutathione disulfide ratio (GSH/GSSG); superoxide dismutase (SOD) were determined. In addition, nuclear transcription factor erythroid 2-related factor 2 (Nrf2), Nrf2-dependent antioxidant enzymes, such as heme oxygenase-1 (HO-1) and NADPH quinine oxidoreductase-1 (NQO-1), and nuclear factor kappa B (NF-κB) were detected. ResultsCompared to I/R group, n-3 PUFAs could obviously increase the mean seminiferous tubular diameter in the histological examination. After n-3 PUFAs treatment, the level of tissue MDA and Superoxide were significantly decreased, while tissue T-AOC, CAT, GSH, GSH/GSSG and SOD levels were significantly increased, compared to I/R group (P<0.05). Besides, the expression levels of Nrf2, HO-1, and NQO-1 were significantly higher and the NF-κB expression level was significantly lower in the n-3 PUFAs treated group than that in I/R group (P<0.05). ConclusionsThese results provided evidences that n-3 PUFAs ameliorated testes damage caused by testicular I/R injury through its antioxidative capacity and anti-inflammatory effects, involving the activation of Nrf2 and the inhibition of NF-κB.

Betulin exhibits anti-inflammatory activity in LPS-stimulated macrophages and endotoxin-shocked mice through an AMPK/AKT/Nrf2-dependent mechanism

Continued oxidative stress can lead to chronic inflammation, which in turn could mediate most chronic diseases including cancer. Nuclear factor erythroid 2-related factor (Nrf2), a critical transcriptional activator for antioxidative responses, has envolved to be an attractive drug target for the treatment or prevention of human diseases. In the present study, we investigated the effects and mechanisms of betulin on Nrf2 activation and its involvement in the lipopolysaccharide (LPS)-triggered inflammatory system. In macrophages, betulin activated the Nrf2 signaling pathway and increased Nrf2-targeted antioxidant and detoxifying enzymes, including NADPH, quinine oxidoreductase 1 (NQO1), heme oxygenase-1 (HO-1), γ-glutamyl cysteine synthetase catalytic subunit (GCLC) and modifier subunit (GCLM) in a dose and time dependent manner. Importantly, we found betulin-induced activation of Nrf2 is AMPK/AKT/GSK3β dependent, as pharmacologically inactivating AMPK blocked the activating effect of betulin on AKT, GSK3β and Nrf2. Furthermore, betulin attenuated LPS-induced inflammatory mediators (iNOS and COX-2) and MAPK inflammatory signaling pathway. The effect of betulin on HO-1 and NQO1 upregulation, iNOS and COX-2 the downregulation, and survival time extension was largely weakened when Nrf2 was depleted in vitro and in vivo. Our results demonstrate that the AMPK/AKT/Nrf2 pathways are essential for the anti-inflammatory effects of betulin in LPS-stimulated macrophages and endotoxin-shocked mice.

Preventive and Protective Roles of Dietary Nrf2 Activators Against Central Nervous System Diseases.

Central nervous system diseases are major health issues and are often associated with disability or death. Most central nervous system disorders are characterized by high levels of oxidative stress. Nuclear factor erythroid 2 related factor (Nrf2) is known for its ability to regulate the expression of a series of enzymes with antioxidative, prosurvival, and detoxification effects. Under basal conditions, Nrf2 forms a complex with Kelch-like ECH associated protein 1, leading to Nrf2 inactivation via ubiquitination and degradation. However, following exposure of Keap1 to oxidative stress, Nrf2 is released from Keap1, activated, and translocated into the nucleus. Upon nuclear entry, Nrf2 binds to antioxidant response elements (ARE), thereby inducing the expression of genes such as glutathione s-transferase, heme oxygenase 1, and NADPH quinine oxidoreductase 1. Many dietary phytochemicals have been reported to activate the protective Nrf2/ARE pathway. Here, we review the preventive and protective effects of dietary Nrf2 activators against CNS diseases, including stroke, traumatic brain injury, Alzheimer's disease, and Parkinson's disease.

Role of Nrf2 signaling pathway in the radiation tolerance of patients with head and neck squamous cell carcinoma: an in vivo and in vitro study.

We aimed to investigate the relationship between the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway and the radiation tolerance of patients with head and neck squamous cell carcinoma (HNSCC). From January 2015 to January 2016, 117 patients with HNSCC were enrolled in our study and assigned into the sensitive and tolerance groups based on curative effect. Immunohistochemistry (IHC) was conducted to measure protein expressions of Nrf2, heme oxygenase-1 (HO1), NADPH quinine oxidoreductase 1 (NQO1) and glutathione S-transferase (GST). Human squamous cell carcinoma cell line, HSC-4, was induced by radiation to construct the HSC-4-radiation resistance (RR) cell line. HSC-4 and HSC-4-RR were also assigned into the blank, negative control (NC) and Nrf2 siRNA groups. Cell Counting Kit-8 (CCK-8), quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting were employed to detect cell viability, mRNA expression and protein expression, respectively, of Nrf2, HO1, NQO1 and GST. A total of 40 nude mice were equally assigned into the untreated, Nrf2 siRNA, radiation therapy (RT) and RT + Nrf2 siRNA groups. Compared with the sensitive group, patients in the tolerance group had upregulated Nrf2, HO1, NQO1 and GST expressions. HSC-4-RR cell line had improved cell viability and higher protein and mRNA expressions of Nrf2, HO1, NQO1 and GST compared with HSC-4 cell line. Compared with the HSC-4-NC and HSC-4-blank groups, the HSC-4-Nrf2 siRNA group had downregulated cell viability. Compared with the HSC-4-RR-NC and HSC-4-RR-blank groups, the HSC-4-RR-Nrf2 siRNA group had lower cell viability. However, the HSC-4-RR-Nrf2 siRNA group had elevated cell viability than the HSC-4-Nrf2 siRNA group. Tumor volume and tumor weight in the RT and RT + Nrf2 siRNA groups decreased evidently. The RT + Nrf2 siRNA group exhibited decreased tumor volume and tumor weight in comparison with the RT group. Our data demonstrated that downregulation of HO1, NQO1 and GST via inhibiting Nrf2 signaling pathway reduces the radiation tolerance of patients with HNSCC.

Targeting the NF-E2-Related Factor 2 Pathway: a Novel Strategy for Traumatic Brain Injury.

As an essential component of cellular defense against a variety of endogenous and exogenous stresses, nuclear factor erythroid 2-related factor 2 (Nrf2) has received increased attention in the past decades. Multiple studies indicate that Nrf2 acts not only as an important protective factor in injury models but also as a downstream target of therapeutic agents. Activation of Nrf2 has increasingly been linked to many human diseases, especially in central nervous system (CNS) injury such as traumatic brain injury (TBI). Several researches have deciphered that activation of Nrf2 exerts antioxidative stress, antiapoptosis, and antiinflammation influence in TBI via different molecules and pathways including heme oxygenase-1 (HO-1), NADPH:quinine oxidoreductase-1 (NQO-1), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), and nicotinamide adenine dinucleotide phosphate oxidase 2 (NOX2). Hence, Nrf2 shows great promise as a molecular target in TBI. In the present article, we provide an updated review of the current state of our knowledge about relationship between Nrf2 and TBI, highlighting the specific roles of Nrf2 in TBI.