NADPH Quinone Oxidoreductase Research Articles

Discovery of a dual inhibitor of NQO1 and GSTP1 for treating glioblastoma

BackgroundGlioblastoma (GBM) is a universally lethal tumor with frequently overexpressed or mutated epidermal growth factor receptor (EGFR). NADPH quinone oxidoreductase 1 (NQO1) and glutathione-S-transferase Pi 1 (GSTP1) are commonly upregulated in GBM. NQO1 and GSTP1 decrease the formation of reactive oxygen species (ROS), which mediates the oxidative stress and promotes GBM cell proliferation.MethodsHigh-throughput screen was used for agents selectively active against GBM cells with EGFRvIII mutations. Co-crystal structures were revealed molecular details of target recognition. Pharmacological and gene knockdown/overexpression approaches were used to investigate the oxidative stress in vitro and in vivo.ResultsWe identified a small molecular inhibitor, “MNPC,” that binds to both NQO1 and GSTP1 with high affinity and selectivity. MNPC inhibits NQO1 and GSTP1 enzymes and induces apoptosis in GBM, specifically inhibiting the growth of cell lines and primary GBM bearing the EGFRvIII mutation. Co-crystal structures between MNPC and NQO1, and molecular docking of MNPC with GSTP1 reveal that it binds the active sites and acts as a potent dual inhibitor. Inactivation of both NQO1 and GSTP1 with siRNA or MNPC results in imbalanced redox homeostasis, leading to apoptosis and mitigated cancer proliferation in vitro and in vivo.ConclusionsThus, MNPC, a dual inhibitor for both NQO1 and GSTP1, provides a novel lead compound for treating GBM via the exploitation of specific vulnerabilities created by mutant EGFR.

Identification of New Targets and the Virtual Screening of Lignans against Alzheimer's Disease.

Alzheimer's disease (AD) is characterized by the progressive disturbance in cognition and affects approximately 36 million people, worldwide. However, the drugs used to treat this disease are only moderately effective and do not alter the course of the neurodegenerative process. This is because the pathogenesis of AD is mainly associated with oxidative stress, and current drugs only target two enzymes involved in neurotransmission. Therefore, the present study sought to identify potential multitarget compounds for enzymes that are directly or indirectly involved in the oxidative pathway, with minimal side effects, for AD treatment. A set of 159 lignans were submitted to studies of QSAR and molecular docking. A combined analysis was performed, based on ligand and structure, followed by the prediction of absorption, distribution, metabolism, excretion, and toxicity (ADMET) properties. The results showed that the combined analysis was able to select 139 potentially active and multitarget lignans targeting two or more enzymes, among them are c-Jun N-terminal kinase 3 (JNK-3), protein tyrosine phosphatase 1B (PTP1B), nicotinamide adenine dinucleotide phosphate oxidase 1 (NOX1), NADPH quinone oxidoreductase 1 (NQO1), phosphodiesterase 5 (PDE5), nuclear factor erythroid 2-related factor 2 (Nrf2), cycloxygenase 2 (COX-2), and inducible nitric oxide synthase (iNOS). The authors conclude that compounds (06) austrobailignan 6, (11) anolignan c, (19) 7-epi-virolin, (64) 6-[(2R,3R,4R,5R)-3,4-dimethyl-5-(3,4,5-trimethoxyphenyl)oxolan-2-yl]-4-methoxy-1,3-benzodioxole, (116) ococymosin, and (135) mappiodoinin b have probabilities that confer neuroprotection and antioxidant activity and represent potential alternative AD treatment drugs or prototypes for the development of new drugs with anti-AD properties.

Selenium-independent antioxidant and anti-inflammatory effects of thioredoxin reductase inhibition in alveolar macrophages

AimsInterleukin-1β (IL-1β) contributes to the development of bronchopulmonary dysplasia (BPD). Thioredoxin reductase-1 (Txnrd1) inhibition activates nuclear factor erythroid 2-related factor 2 (Nrf2)-dependent responses. Txnrd1 activity is selenium (Se) dependent and Se deficiency is common in prematurity. Auranofin (AFN), a Txnrd1 inhibitor, decreases IL-1β levels and increases Nrf2 activation in lipopolysaccharide (LPS) treated alveolar macrophages. In lung epithelia, AFN-induced Nrf2 activation is Se dependent. We tested the hypothesis that the effects of Txnrd1 inhibition in alveolar macrophages are Se dependent. Main methodsTo establish Se sufficient (Se+) and deficient (Se−) conditions, alveolar (MH-S) macrophages were cultured in 2.5% fetal bovine serum (FBS) ± 25 nM Na2SeO3. Se− (2.5% FBS) and Se+ (2.5% FBS + 25 nM Na2SeO3) cells were cultured in the presence or absence of 0.05 μg/mL LPS and/or 0.5 μM AFN. Nrf2 activation was determined by measuring NADPH quinone oxidoreductase-1 (Nqo1) and glutathione levels. IL-1β mRNA (Il1b) and protein levels were measured using qRT-PCR and ELISA. Data were analyzed by ANOVA followed by Tukey's post-hoc. Key findingsWe detected an independent effect of AFN, but not LPS, on Nqo1 expression and GSH levels in Se+ and Se− cells. LPS significantly increased Il1b and IL-1β levels in both groups. AFN-mediated attenuation of this effect was not impacted by Se status. SignificanceThe beneficial effects of Txnrd1 inhibition in alveolar macrophages are Se-independent and therefore unlikely to be diminished by clinical Se deficiency.

NADPH-quinone oxidoreductase-1 mediates Benzo-[a]-pyrene-1,6-quinone-induced cytotoxicity and reactive oxygen species production in human EA.hy926 endothelial cells

Numerous studies conducted in the past have reported deaths in the human population due to cardiovascular diseases (CVD) on exposure to air particulate matter (APM). BP-1,6-quinone (BP-1,6-Q) is one of the significant components of APM. However, the mechanism(s) by which it can exert its toxicity in endothelial cells is not yet completely understood. NAD(P)H: quinone oxidoreductase-1 (NQO1) is expressed highly in myocardium and vasculature tissues of the heart and plays a vital role in maintaining vascular homeostasis. This study, demonstrated that BP-1,6-Q diminishes NQO1 enzyme activity in a dose-dependent manner in human EA.hy926 endothelial cells. The decrease in the NQO1 enzyme causes potentiation in BP-1,6-Q-mediated toxicity in EA.hy926 endothelial cells. The enhancement of NQO1 in endothelial cells showed cytoprotection against BP-1,6-Q-induced cellular toxicity, lipid, and protein damage suggesting an essential role of NQO1 in cytoprotection against BP-1,6-Q toxicity. Using various biochemical assays and genetic approaches, results from this study further demonstrated that NQO1 also plays a crucial role in BP-1,6-Q-induced production of reactive oxygen species (ROS). These findings will contribute to elucidating BP-1,6-Q mediated toxicity and its role in the development of atherosclerosis.

Enhancement of piperlongumine chemosensitivity by silencing heme oxygenase-1 expression in cholangiocarcinoma cell lines.

Piperlongumine (PL) produces reactive oxygen species (ROS) and induces G2/M-phase arrest in cholangiocarcinoma (CCA) cells via the JNK/ERK pathway. A differential response to PL was observed among all CCA cell lines However, the underlying mechanisms have remained to be fully elucidated. The aim of the present study was to investigate the molecular mechanisms of PL-induced heme oxygenase-1 (HO-1) expression in CCA cell lines. The anti-proliferative action of PL in the CCA cell lines KKU-100 and KKU-213A was analyzed using sulforhodamine B assays. Reverse transcription-quantitative PCR and western blot analyses were used to examine mRNA and protein expression. HO-1 inhibition was achieved using the chemical inhibitor zinc protophoryn or specific small interfering RNA to HO-1. Intracellular ROS was detected using a 2,7-dichlorodihydrofluorescein diacetate fluorescence assay. High expression of phase-II detoxification enzymes, including NADPH quinone oxidoreductase-1, heme oxygenase-1, superoxide dismutases and aldo-keto reductase 1 subunits C-1 and 3, were detected in the KKU-100 cell line. Of the CCA cell lines tested, KKU-100 was the least sensitive to PL. Dose-dependent upregulation of HO-1 expression via PI3K/Akt activation was detected in PL-treated CCA cells. Inhibition of HO-1 eliminated the antioxidant defense mechanisms, leading to increased anti-cancer activity of PL in the CCA cell lines via an increase in intracellular ROS levels and apoptotic protein expression. These observations indicated that HO-1 inhibition had a chemosensitizing effect on CCA to PL.

Oltipraz Prevents High Glucose-Induced Oxidative Stress and Apoptosis in RSC96 Cells through the Nrf2/NQO1 Signalling Pathway.

Diabetic peripheral neuropathy (DPN) is a common complication of diabetes mellitus (DM). Schwann cell (SC) apoptosis contributes to the occurrence and development of DPN. Effective drugs to prevent SC apoptosis are required to relieve and reverse peripheral nerve injury caused by DM. Oltipraz [4-methyl-5-(2-pyrazinyl)-1,2-dithiole-3-thione], an agonist of nuclear factor erythroid derived-2-related factor 2 (Nrf2), exerts strong effect against oxidative stress in animal models or clinical patients in certain diseases, including heart failure, acute kidney injury, and liver injury. The aim of the present study was to determine the effectiveness of oltipraz in preventing SC apoptosis induced by high glucose levels. RSC96 cells pretreated with oltipraz were cultured in high-glucose medium (50 mM glucose) for 24 h, and cells cultured in medium containing 5 mM glucose were used as the control. Flow cytometry was used to evaluate the degree of apoptosis. A Cell Counting Kit-8 assay was used to assess cell viability. The mitochondrial membrane potential was assessed using JC-1 staining, and reactive oxygen species (ROS) generation was measured using 20,70-dichlorodihydrofluorescein diacetate staining. In addition, the levels of malondialdehyde (MDA) and superoxide dismutase (SOD) levels were also evaluated using the corresponding kits. Flow cytometry was subsequently used to detect apoptosis, and western blotting was used to measure the expression levels of nuclear factor erythroid derived-2-related factor 2 and NADPH quinone oxidoreductase 1. The results showed that high glucose concentration increased oxidative stress and apoptosis in RSC96 cells. Oltipraz improved cell viability and reduced apoptosis of RSC96 cells in the high glucose environment. Additionally, oltipraz exhibited a significant antioxidative effect, as shown by the decrease in MDA levels, increased SOD levels, and reduced ROS generation in RSC96 cells. The results of the present study suggest that oltipraz exhibits potential as an effective drug for treatment with DPN.

Treatment with slow-releasing hydrogen sulfide donors inhibits the nociceptive and depressive-like behaviours accompanying chronic neuropathic pain: Endogenous antioxidant system activation.

Therapies to treat chronic neuropathic pain and its associated comorbidities are limited. Recent studies demonstrated that the administration of slow-releasing hydrogen sulfide (H2S) donors inhibited chemotherapy-induced neuropathic pain. However, the antidepressant or anxiolytic effects of these compounds and their mechanisms of action during chronic neuropathic pain have not been evaluated. To determine whether the administration of two slow-releasing H2S donors, allyl isothiocyanate (A-ITC) and phenyl isothiocyanate (P-ITC), inhibits the nociceptive and emotional disorders associated with chronic neuropathic pain. In C57BL/6 male mice with neuropathic pain caused by the chronic constriction of the sciatic nerve, we assessed the effects of intraperitoneal administration of A-ITC and P-ITC in (a) the mechanical allodynia, thermal hyperalgesia and thermal allodynia induced by nerve ligation; (b) the anxiety- and depressive-like behaviours linked with neuropathic pain; (c) glial activation and mitogen-activated protein kinases phosphorylation, and (d) expression of the antioxidant enzymes, heme oxygenase 1 (HO-1), NADPH quinone oxidoreductase1, and glutathione S-transferase mu-1 (GSTM1), and alpha-1 (GSTA1), in hippocampus and prefrontal cortex (PFC). Both treatments inhibited the allodynia and hyperalgesia, depressive-like behaviours, astroglial activation, and the extracellular signal-regulated kinase 1/2 phosphorylation but were unable to abolish the anxiety-like behaviours accompanying neuropathic pain. A-ITC and P-ITC also augmented the expression of HO-1, GSTM1, and GSTA1 in the hippocampus and/or PFC. The administration of slow-releasing H2S donors might be a promising treatment for the management of chronic neuropathic pain and some associated comorbidities via inhibiting the inflammatory and plasticity changes, and activating the endogenous antioxidant responses.

TBHQ improved neurological recovery after traumatic brain injury by inhibiting the overactivation of astrocytes

Traumatic brain injury (TBI) is a major leading cause of death and long-term disability. Although astrocytes play a key role in neuroprotection after TBI in the early stage, the overactivation of astrocytes can lead to long-term functional deficits, and the underlying pathophysiological mechanisms remain unclear. In addition, it is unknown whether the nuclear factor erythroid 2-related factor2/haem oxygenase-1 (Nrf-2/HO-1) pathway could elicit a neuroprotective effect by decreasing astrocyte overactivation after TBI. We aimed to study the effects of tert-butylhydroquinone (TBHQ) in reducing astrocyte overactivation after TBI and explored the underlying mechanisms. We first established a controlled cortical impact (CCI) model in rats and performed Haematoxylin and eosin (H&E) staining to observe brain tissue damage. The cognitive function of rats was assessed by modified neurological severity scoring (mNSS) and Morris water maze (MWM) test. Astrocyte and microglia activation was detected by immunofluorescence staining. Oxidative stress conditions were investigated using Western blotting. An enzyme-linked immunosorbent assay (ELISA) was designed to assess the level of the proinflammatory factor tumour necrosis factor-alpha (TNF-α). Dihydroethidium (DHE) staining was used to detect reactive oxygen species (ROS). Apoptosis was assessed by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. The results showed that the administration of TBHQ ameliorated motor function and cognitive deficits and decreased the lesion volume. In addition, TBHQ significantly decreased astrocyte overactivation, diminished the pro-inflammatory phenotype M1 and inflammatory cytokines production after TBI, increased Nrf-2 nuclear accumulation, and enhanced the levels of the Nrf-2 downstream antioxidative genes HO-1 and NADPH-quinone oxidoreductase-1 (NQO-1). Furthermore, TBHQ treatment alleviated apoptosis and neuronal death in the cerebral cortex. Overall, our data indicated that the upregulation of Nrf-2 expression could enhance neuroprotection and decrease astrocyte overactivation and might represent a new theoretical basis for treating TBI.

Nrf2 transcription factor inhibition aggravates oxidative stress induced D1R dysfunction and hypertension in mice

The renal dopamine 1 receptor (D1R) dysfunction is strongly associated with oxidative stress and high blood pressure (BP). Nuclear factor E2‐related factor 2 (Nrf2), a redox‐sensitive transcriptional factor is a key regulator in antioxidant response element (ARE)‐mediated induction of phase II antioxidant enzymes. The present study was conducted to investigate the role of Nrf2/ARE signaling pathway in renal D1R function and BP in mice. C57BL/6J mice were treated with a pro‐oxidant buthionine sulfoximine (BSO) (10 mmol/L in drinking water) and ML385, a Nrf2 inhibitor that binds to Nrf2 and inhibits its downstream target gene expression (10 mg/kg body weight/day, intraperitoneally through osmotic pumps) for 8 weeks. BSO administration to wild type mice resulted in marked increased in oxidative stress and mean arterial pressure (MAP) which was accompanied by decreased renal D1R expression and glomerular filtration rate (GFR). Also, SKF38393 (D1R agonist) infusion failed to promote sodium excretion indicating D1R dysfunction in BSO treated animals. ML385 treatment per se had no significant change in MAP, D1R, heme oxygenase‐1 (HO‐1) and NADPH quinone oxidoreductase1 (NQO‐1) expression, GFR and sodium excretion. However, co‐treatment of ML385 and BSO exhibited robust increase in MAP, aggravated oxidative damage and significantly decreased GFR. ML385 treatment to BSO administered animals further impaired the D1R function that resulted in decrease in sodium excretion. Moreover, ML385 and BSO co‐treatment showed decreased D1R and phase II antioxidant enzyme (HO‐1 and NQO‐1) expression in comparison to BSO alone animals. In conclusion, the results suggest Nrf2 inhibition exacerbated oxidative stress induced D1R dysfunction and hypertension possibly by attenuating the phase II antioxidant defense response.Support or Funding InformationThis work is funded by National Institutes of Health (National Heart, Lung, and Blood Institute) Grant HL‐139808.

Modulation of Nqo1 activity intercepts anoikis resistance and reduces metastatic potential of hepatocellular carcinoma.

The processing of intracellular reactive oxygen species (ROS) by nuclear factor erythroid‐derived 2‐like 2 (Nrf2) and NADPH quinone oxidoreductase 1 (Nqo1) is important for tumor metastasis. However, the clinical and biological significance of Nrf2/Nqo1 expression in hepatocellular carcinoma (HCC) remains unclear. We aimed to clarify the clinical importance of Nrf2/Nqo1 expression in HCC and evaluate the association of Nrf2/Nqo1 expression with HCC metastasis. We also evaluated the impact of Nqo1 modulation on HCC metastatic potential. We used spheroids derived from HCC cell lines. In anchorage‐independent culture, HCC cells showed increased ROS, leading to the upregulation of Nrf2/Nqo1. Futile stimulation of Nqo1 by β‐lapachone induces excessive oxidative stress and dramatically increased anoikis sensitivity, finally diminishing the spheroid formation ability, which was far stronger than depletion of Nqo1. We analyzed 117 cases of primary HCC who underwent curative resection. Overexpression of Nrf2/Nqo1 in primary HCC was associated with tumor size, high α‐fetoprotein, and des‐γ‐carboxy‐prothrombin levels. Overexpression of Nrf2/Nqo1 was also associated with multiple intrahepatic recurrences (P = .0073) and was an independent risk factor for poor prognosis (P = .0031). NADPH quinone oxidoreductase 1 plays an important role in anchorage‐independent survival, which is essential for survival for circulation and distant metastasis of HCC cells. These results suggest that targeting Nqo1 activity could be a potential strategy for HCC adjuvant therapy.

Inonotus obliquus polysaccharide ameliorates impaired reproductive function caused by Toxoplasma gondii infection in male mice via regulating Nrf2-PI3K/AKT pathway

This study was carried out to investigate the effects of Inonotus obliquus polysaccharide (IOP) on impaired reproductive function and its mechanisms in Toxoplasma gondii (T. gondii)-infected male mice. Results showed that IOP significantly improved the spermatogenic capacity and ameliorated pathological damage of testis, increased serum testosterone (T), luteinizing hormone (LH) and follicular-stimulating hormone (FSH) levels in T. gondii-infected male mice. IOP effectively up-regulated testicular steroidogenic acute regulatory protein (StAR), P450scc and 17β-HSD expressions. IOP also significantly decreased the levels of malondialdehyde (MDA) and nitric oxide (NO), but increased the activities of antioxidant enzyme superoxide dismutase (SOD) and glutathione (GSH). Furthermore, IOP up-regulated the expressions of nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1) and NADPH quinoneoxidoreductase-1 (NQO-1), and suppressed the apoptosis of testicular cells by decreasing Bcl-2 associated x protein (Bax) and cleaved caspase-3 expressions. IOP further enhanced testicular phosphatidylinositol 3-kinase (PI3K), phospho-protein kinase B (p-AKT) and phospho-mammalian target of rapamycin (p-mTOR) expression levels. It demonstrates the beneficial effects of IOP on impaired reproductive function in T. gondii-infected male mice due to its anti-oxidative stress and anti-apoptosis via regulating Nrf2-PI3K/AKT signaling pathway.

Properties of melanin from Lachnum YM156 and its hepatoprotective effect in Cd-exposure mice

Natural melanin was extracted from Lachnum YM156 (LIM). LIM had better thermostability and light resistance, and its solubility was relatively high under alkaline conditions. Simultaneously, we examined its hepatoprotective effect in Cd-exposure mice. Cd-exposure resulted in decreasing weight growth rate, raised liver index, elevated serum levels of alanine aminotransferase (ALT) and aspartate aminotransaminase (AST), and increased Cd accumulation in livers. Hepatic oxidative stress was evidenced by increased malondialdehyde (MDA) contents, decreased glutathione (GSH) contents, and decreased activities of dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT) after CdCl2 administration. Additionally, Cd-exposure also increased the liver level of tumor necrosis factor (TNF)-α and interleukin(IL)-1β. Furthermore, the result of quantitative real-time polymerase chain reaction(qRT-PCR) showed that Cd-exposure reduced the mRNA expression level of NF-E2-related factor 2 (Nrf2), heme oxygenase (HO-1), NADPH quinone oxidoreductase 1 (NQO-1), and elevated the mRNA expression level of factor-kappaB (NF-κB) p65, inducible nitric oxide synthase (iNOS) in livers. However, all these changes were dose-dependently reversed by LIM. Overall, the present findings demonstrated that LIM have the hepatoprotective effect through antioxidant and anti-inflammatory responses in Cd-exposure mice.

A novel Xanthomonas citri subsp. citri NADPH quinone reductase involved in salt stress response and virulence

BackgroundXanthomonas citri subsp. citri (Xcc), the causal agent of citrus canker is maintained as an epiphyte on citrus leaves until entering the plant tissue. During epiphytic survival, bacteria may encounter low water availability that challenges the infection process. Proteomics analyses of Xcc under saline stress, mimicking the conditions found during epiphytic survival, showed increased abundance of a putative NAD(P)H dehydrogenase encoded by XAC2229. MethodsExpression levels of XAC2229 and a Xcc mutant in XAC2229 were analyzed in salt and oxidative stress and during plant-pathogen interaction. An Escherichia coli expressing XAC2229 was obtained, and the role of this protein in oxidative stress resistance and in reactive oxygen species production was studied. Finally, Xac2229 protein was purified, spectrophotometric and cofactor analyses were done and enzymatic activities determined. ResultsXAC2229 was expressed under salt stress and during plant-pathogen interaction. ΔXAC2229 mutant showed less number of cankers and impaired epiphytic survival than the wild type strain. ΔXAC2229 survived less in the presence of H2O2 and produced more reactive oxygen species and thiobarbituric acid-reactive substances than the wild type strain. Similar results were observed for E. coli expressing XAC2229. Xac2229 is a FAD containing flavoprotein, displays diaphorase activity with an optimum at pH 6.0 and has quinone reductase activity using NADPH as an electron donor. ConclusionsA FAD containing flavoprotein from Xcc is a new NADPH quinone reductase required for bacterial virulence, particularly in Xcc epiphytic survival on citrus leaves. General significanceA novel protein involved in the worldwide disease citrus canker was characterized.

Jujube (Ziziphus jujuba Mill.) Protects Hepatocytes against Alcohol-Induced Damage through Nrf2 Activation.

This study aimed at evaluating the cytoprotective activity of jujube water extract (JWE) against alcohol-induced oxidative stress via the activation of the Nrf2 pathway in HepG2 cells. JWE had various phenolic compounds, and the vanillic acid content was the highest in the extract. To determine the cytoprotective effect of JWE against alcohol-induced damage, hepatocytes were treated with JWE and 3% ethanol. JWE (100 μg/mL) markedly increased cell viability by approximately 100% in a dose-dependent manner. Moreover, JWE attenuated the production of malondialdehyde, reactive oxygen species, aspartate, and alanine aminotransferase and the depletion of glutathione. Moreover, JWE enhanced the expression of antioxidant defense enzymes including heme oxygenase-1, NADPH quinone oxidoreductase 1, and γ-glutamate-cysteine ligase catalytic against alcohol-induced oxidative damage in hepatocytes via the activation of Nrf2. Taken together, JWE possesses the protective effect against alcohol-induced oxidative injury in hepatocytes through the upregulation of the Nrf2 signaling pathway. Therefore, jujube fruit might have the potential to improve alcohol-related liver problems.

The compound, diallyl disulfide, enriched in garlic, prevents the progression of doxorubicin-induced nephropathy

Abstract The main medicinal property of garlic is mostly attributed to its organosulfur compounds, of which the oil-soluble diallyl disulfide (DADS) is the principal component of distilled garlic oil. Nuclear factor erythroid 2-related factor 2 (Nrf2) is an important cellular regulator in response to oxidative stress. This study assessed the possible protective effect of DADS on doxorubicin (Dox)-induced nephrotoxicity and its potential regulation of the Nrf2 pathway. Treatment with DADS (200 μM) induced heme oxygenase (HO)-1 and NADPH quinone oxidoreductase 1 (NQO-1) expression in a human proximal tubular cell line. The induction of HO-1 expression was suppressed in Nrf2-silenced cells. In an animal study, pretreatment with DADS relieved Dox-induced albuminuria, increased catalase activity, and reduced the urinary 8-hydroxy-2'-deoxyguanosine level. In addition, DADS ameliorated the severity of glomerulosclerosis and suppressed expressions of fibrotic and inflammatory gene expressions. Our data indicate that DADS, a major component of garlic, showed protective effects of preventing the progression of Dox-induced nephropathy through enhancing Nrf2-mediated antioxidant activity. DADS, a normal constituent of the human diet, merits investigation as a potential antioxidant daily food supplement against free radical-mediated chronic diseases.

The HO-1 Signal Prevents HMGB1-Mediated Activation of NLRP3 Inflammasomes in Lipopolysaccharide-Induced Acute Lung Injury In Vitro

BackgroundCurrent treatments of lipopolysaccharide (LPS)-induced acute lung injury (ALI) are unsatisfactory due to the insufficient understanding of the pathogenesis of LPS-induced ALI. The NLRP3 inflammasome is an essential part of the innate protection system and is involved in LPS-induced ALI; however, comprehensive understanding of molecular pathogenesis of the disease is lacking. Our study explored the effect of heme oxygenase-1 (HO-1) on NLRP3 inflammasomes in vitro. MethodsAlveolar macrophages (NR8383) were preincubated with high-mobility group box-1 (HMGB1) or HO-1 CRISPR plasmids before LPS stimulation. Then, we detected the effect of HO-1 on NLRP3 inflammasomes. ResultsOur study demonstrates that the activation of HO-1 represses the level of NLRP3 inflammasomes and the subsequent increases of the level of IL-1β. Moreover, NLRP3 inflammasome activation was sensitive to the HMGB1 activity, and HO-1 was able to reduce the amount of HMGB1 released. Furthermore, downregulation of NLRP3 inflammasomes was related to NADPH quinone oxidoreductase 1 (an HO-1–related gene). ConclusionsOur study clarifies the constrained coordination of the HO-1 signal in the HMGB1-mediated activation of NLRP3 inflammasomes in NR8383 alveolar macrophages after LPS stimulation.

NADPH: Quinone oxidoreductase 1 (NQO1) mediated anti-cancer effects of plumbagin in endocrine resistant MCF7 breast cancer cells

BackgroundPLB is a natural naphthoquinone compound isolated from the roots of Plumbago indica plant. Our previous study reported the inhibitory effect of Plumbagin (PLB) on human endocrine resistant breast cancer cell growth and cell invasion. Hypothesis/PurposeSince PLB is a naphthoquinone compound, it can be reduced by the cytosolic NADPH: quinone oxidoreductase 1 (NQO1) enzyme. NQO1 expression is upregulated in various types of aggressive cancer including breast cancer. This study investigated the impact of NQO1 on anti-cancer effects of PLB in endocrine-resistant breast cancer cells. Study DesignThis study was an in vitro study using ER-positive cell line (MCF7) and endocrine-resistant cell lines (MCF7/LCC2 and MCF7/LCC9 cells). MethodsThe roles of NQO1 in anti-cancer activity of PLB were investigated by using NQO1 knockdown cells, NQO1 inhibitor and NQO1 overexpressed cells. To study the impact of NQO1 on the effects of PLB on cell viability, apoptosis, invasion and generation of ROS, the following assays were used: MTT assays, annexin V-PE/7-ADD staining flow cytometry, matrigel invasion assays and DCFHDA assays. To study the mechanism of how NQO1 mediated PLB effects in tamoxifen response and apoptosis, we assessed the levels of mRNA expression by using qRT-PCR. Results1. In this study, NQO1 was upregulated in endocrine-resistant cells.2. PLB did not change the expression of NQO1 but it was able to increase NQO1 activity.3. The inhibitory effects of PLB on cell proliferation, cell invasion and expression of tamoxifen resistant gene were attenuated in NQO1 knockdown cells or in the presence of NQO1 inhibitor.4. The effects of PLB to induce apoptosis and generate ROS were also decreased when NQO1 activity was inhibited or when the NQO1 expression was reduced.5. The anti-cancer effects of PLB increased when NQO1 was upregulated. ConclusionThe effects of PLB in endocrine-resistant breast cancer cells is dependent on NQO1’s activity.

Gene Polymorphism of Xenobiotic Biotransformation Enzymes in Patients with Classical Ph-Negative Myeloproliferative Neoplasms.

The correlation of gene polymorphisms rs4025935 (large deletion), rs1695 (313A>G), rs71748309 (large deletion), and rs1800566 (609C>T) of GSTM1, GSTT1, and NQO1 genes encoding glutathione-S-transferases (GST) M1, P1, and T1 and NADPH-quinone oxidoreductase with the risk of development of classical Ph-negative myeloproliferative neoplasms (polycythemia vera, essential thrombocythemia, and primary myelofibrosis) was studied in the Caucasian ethnicity population of Vyatka region of the Russian Federation. It was found that NQO1*609T allele, NQO1*609T genotypes, and homozygous carriage of a deletion (null) allele of GSTT1 gene are associated with the risk of development of myeloproliferative neoplasms (OR=1.29, 95%CI=1.02-1.64, p=0.04; OR=1.39, 95%CI=1.04-1.85, p=0.03; and OR=1.48, 95%CI=1.03-2.12, p=0.03, respectively). However, no influence of GSTM1 and GSTP1 gene polymorphisms on the risk of development of myeloproliferative disorders was registered.

Coptisine ameliorates renal injury in diabetic rats through the activation of Nrf2 signaling pathway.

The present study has been designed and carried out to evaluate the potential of coptisine on diabetic nephropathy. Diabetes was induced in SD rats through one single intraperitoneal injection of streptozotocin (65mg/kg) method, and then diabetic rats were orally administered with 25mg/kg/day coptisine or 50mg/kg/day coptisine for 8weeks. Severe impairment of renal function in rats with diabetes was observed as indicated by increased urine protein excretion, kidney hypertrophy index, serum creatinine level, and blood urea nitrogen level. Oxidative stress damage was observed as indicated by increased levels of reactive oxygen species, malondialdehyde, and decreased levels of glutathione, superoxide dismutase, and catalase. However, these alterations in kidneys of rats with diabetes were alleviated by administration of coptisine. Furthermore, the expression levels of nuclear factor-erythroid 2-related factor 2 (Nrf2) and its targeted antioxidative genes heme oxygenase 1 and NADPH quinone oxidoreductase 1 in the diabetic kidneys were significantly increased after coptisine treatment. These results suggested that coptisine ameliorated oxidative renal injury in diabetic rats, and the possible mechanisms for the renoprotective effects of coptisine may be related to activation of the Nrf2 signaling pathway.

Dendrobium Officinale Polysaccharides Protect against MNNG-Induced PLGC in Rats via Activating the NRF2 and Antioxidant Enzymes HO-1 and NQO-1.

Dendrobium officinale polysaccharides (DOP) are the main effective ingredient in Dendrobium officinale. Nuclear factor erythroid 2-related factor 2 (NRF2) signaling is regarded as an important way to mitigate the effects of reactive oxygen species (ROS) damage and inhibit gastric cancer progress. This study introduces a previously unknown effect of DOP on precancerous lesions of gastric cancer (PLGC). The mechanism discussed herein is based on the NRF2 signal pathway as well as its downstream antioxidant enzymes heme oxygenase-1 (HO-1) and NADPH quinone oxidoreductase-1 (NQO-1). DOP was prepared by the alcohol deposition method, and its molecular weight was determined using High-Performance Gel-Permeation Chromatography (HPGPC). Sixty male rats were randomly divided into five groups: normal control group (NC), PLGC model group (PLGC), model treated with low dose (2.4 g/kg) of DOP (L-DOP), model treated with middle dose (4.8 g/kg) of DOP (M-DOP), and model treated with high dose (9.6 g/kg) of DOP (H-DOP). DOP was orally administered to rats for 15 consecutive days prior to the start of a seven-month course of 1-methyl-3-nitro-1-nitrosoguanidine (MNNG) exposure. Histological evaluation was observed by hematoxylin and eosin (HE) and alcian blue/periodic acid-Schiff (AB-PAS) staining. Alanine aminotransferase (ALT), aspartate transaminase (AST), serum creatinine (Scr), serum uric acid (UA), blood urea nitrogen (BUN), and HE staining were detected for liver and kidney function. The level of 8-hydroxy-deoxyguanosine (8-OHdG) in serum was detected by kits. The NRF2 protein expression was detected by immunohistochemistry, and western blotting was utilized to compare differential protein expression levels among cytoplasmic and nuclear cell fractions. Expression levels of antioxidant enzymes heme oxygenase 1 (HO-1), Glutamate-Cysteine Ligase Catalytic Subunit (GCLC), Glutamate-Cysteine Ligase Modifier Subunit (GCLM), and NAD(P)H: quinone oxidoreductase-1 (NQO-1) were analyzed by reverse transcriptase polymerase chain reaction (RT-PCR); furthermore, the protein expression of NRF2, HO-1, and NQO-1 was detected by western blotting. The results showed that the average content of DOP is 83%, and its molecular weight is mainly contained within 3500 and 1000000. The H-DOP experimental group exhibited noticeable weight gain after seven months, reduced intestinal metaplasia, and made the atypical hyperplasia to be kept in moderate or mild degree. Data also showed DOP to be capable of decreasing levels of ALT, UA, and BUN, all of which had been elevated following the appearance of MNNG-induced PLGCs. DOP was also seen to reduce the expression of 8-OHdG and promote the expression of NRF2 in the gastric mucosa. Furthermore, RT-PCR and western blotting results showed that DOP upregulated the gene and protein expression of HO-1 and NQO-1. These findings show that DOP prevents MNNG-induced PLGC along with subsequent liver and kidney damage. The protective effects of DOP are associated with the reduction of 8-OHdG levels as well as the activation of the NRF2 pathway and its related antioxidant enzymes, HO-1 and NQO-1.