Quinine oxidoreductase 1 inhibits the secretion of IL-8 in Mycoplasma pneumoniae lipid-associated membrane proteins stimulated the THP-1 monocyte line / 中华微生物学和免疫学杂志

Objective:To investigate the effects and regulation mechanism of lipid-associated membrane proteins (LAMPs) derived from Mycoplasma pneumoniae( Mp) on the expression of quinine oxidoreductase 1 (NQO-1) in human monocyte cell line THP-1 cells, and to know the effect of NQO-1 to interleukin 8 secretion in LAMPs stimulated cells, so as to better understand the regulation mechanism upon Mp infection. Methods:Mp were cultivated and the precipitate was collected to extract LAMPs. The cytotoxicity of LAMPs to THP-1 cells was analyzed by using CCK8 test. THP-1 cells were cultured in vitro with different concentrations of LAMPs for different times, and the expression of NQO-1 protein was detected by Western blot. Nrf2 siRNA was used to investigate the role of Nrf2 in NQO-1 expression in LAMPs induced cells, and NQO-1 inhibitor Diminutol was performed to test whether they blocked interleukin 8 (IL-8) secretion when treated with LAMPs in THP-1 cells. Results:LAMPs extracted from Mp had no cytotoxicity to THP-1 cells. The expression of NQO-1 protein in LAMPs-stimulated THP-1 cells showed a dose-dependent and time-dependent manner. The production of NQO-1 protein reached peaks when treated with 5.0 μg/ml or 7.5 μg/ml of LAMPs for 12 h. Silencing of Nrf2 by siRNA significantly decreased NQO-1 production, and blocking NQO-1 by Dim increased the level of IL-8 in LAMPs-stimulated cells. Conclusions:LAMPs derived from Mp induced the expression of NQO-1 protein in THP-1 cells via Nrf2, and NQO-1 can inhibit IL-8 secretion in LAMPs stimulated monocytes.

Sirtuin 6 mitigated LPS-induced human umbilical vein endothelial cells inflammatory responses through modulating nuclear factor erythroid 2-related factor 2.

BACKGROUND: Nuclear factor erythroid 2-related factor 2 (Nrf2) protects the lung from sepsis-induced injury through activating Nrf2-regulated multiple phase 2 detoxification genes, including NAD(P)H: quinine oxidoreductase-1 (NQO1) and heme oxygenase-1 (HO1). Based on the positive effect of Sirtuin 6 on Nrf2, we aim to explore the potential role of SIRT6 in the mechanism of sepsis-induced acute lung injury (ALI). METHODS: Mouse models of sepsis were constructed by instilling intratracheal of lipopolysaccharide (LPS; 4 ml/kg). After 48-hour treatment, lung tissues were collected to measure the degree of lung injury. The SIRT6, siSIRT6, and siNrf2 plasmids were cotransfected into various concentrations of LPS-treated human umbilical vein endothelial cells (HUVECs; 0, 1, 5, 10, and 50 µg/ml) using Lipofectamine 2000. Tumor necrosis factor-α (TNF-α) and interleukin (IL)-6 levels were determined by enzyme-linked immunosorbent assay. Expression levels of SIRT6, Nrf2, NQO1, and HO1 was measured by quantitative polymerase chain reaction and Western blot analysis. Cell apoptosis was determined by flow cytometry. RESULTS: Lung tissues in the model group already had basic characteristics of ALI. Compared with the control model, TNF-α and IL-6 levels were much higher (P < 0.01), the levels of SIRT6, Nrf2, and Nrf2-modulated detoxification factors were downregulated (P < 0.01). SIRT6 overexpression decreased the apoptosis below to 10% (P < 0.01), significantly increased the Nrf2 expression, effectively inhibited TNF-α and IL-6 releases, and enhanced NQO1 and HO1 levels (P < 0.01). siNrf2 abolished the protective effects of SIRT6 overexpression, including increasing apoptosis and inhibiting anti-inflammatory and antioxidative genes expressions (P < 0.01). CONCLUSIONS: Our study suggested SIRT6 positively regulated Nrf2 expression and activated Nrf2-regulated anti-inflammatory and antioxidative enzymes, which could effectively mitigate LPS-induced HUVECs inflammatory responses. This might reflect the mechanism of ALI induced by sepsis.

N-acetylcysteine amide provides neuroprotection via Nrf2-ARE pathway in a mouse model of traumatic brain injury.

BACKGROUND: Increasing evidence demonstrate N-acetylcysteine amide (NACA) provides neuroprotection and attenuated oxidative stress in rats following traumatic brain injury (TBI). The nuclear factor erythroid 2-related factor 2 (Nrf2)-antioxidant response element (ARE) signal pathway is activated after TBI and provides a protective effect against TBI. However, the function and mechanism of NACA in mice after TBI remain unknown. This study was to evaluate the neuroprotection of NACA and the potential action of the Nrf2-ARE pathway in a weight-drop mouse model of TBI. MATERIALS AND METHODS: Four groups of animals were randomly divided into sham, TBI, TBI+vehicle, and TBI+NACA (100 mg/kg, administered intraperitoneally). The protein levels of Nrf2, heme oxygenase-1 (HO-1), NAD(P)H: quinine oxidoreductase-1 (NQO1), cleaved caspase-3 and the mRNA levels of HO-1 and NQO1 were detected. The neurobehavior, neuronal degeneration, apoptosis and oxidative stress were also assessed. RESULTS: Treatment with NACA significantly improved neurologic status at days 1 and 3 following TBI. Moreover, NACA promoted Nrf2 activation a day after TBI. The protein and mRNA levels of HO-1 and NQO1 were upregulated by NACA. Meanwhile, NACA treatment significantly reduced the level of malondialdehyde (MDA) and enhanced the activity of superoxide dismutase (SOD) and glutathione peroxidase (GPx), which indicated NACA attenuated oxidative stress following TBI. NACA prominently reduced the protein level of cleaved caspase-3 and TUNEL-positive cells, indicating its antiapoptotic effect. Additionally, Fluoro-Jade C staining showed NACA alleviated neuronal degeneration a day after TBI. CONCLUSIONS: Our study reveals that NACA potentially provides neuroprotection via the activation of the Nrf2-ARE signaling pathway after TBI in mice.

Neuroprotective effect of polysaccharide separated from Perilla frutescens Britton var. acuta Kudo against H2O2-induced oxidative stress in HT22 hippocampus cells.

This study was carried out to evaluate the neuroprotective activity of polysaccharide extracts isolated from Perilla frutescens (PEPF) in H2O2-treated HT22 hippocampus cells. The PEPF treatment was found to increase the anti-oxidant activities of HT22 hippocampus cells. PEPF treatment resulted in a significant protection of HT22 hippocampus cells against H2O2-induced neurotoxicity, this protection ultimately occurred through an inhibition of ROS-mediated intracellular Ca2+ levels leading to MAPKs and NF-κB, as well as the accumulation of PI3K/AKT and Nrf2-mediated HO-1/NQO1 pathways. Furthermore, PEPF not only decreased the expression of Bax, cytochrome c, and cleaved caspases-3, -8, and -9, but also increased the expression of PARP and Bcl-2 in the H2O2-treated HT22 hippocampus cells, which overall contributed to the neuroprotective action. PEPF retains its mitochondrial membrane potential and reduces the elevated levels of sub-G1 phase and apoptotic morphological features induced by H2O2. It also reduces the malondialdehyde levels and enhances the intracellular SOD activity.

Cytoprotective dihydronaphthalenones from the wood of Catalpa ovata.

Three previously undescribed dihydronaphthalenones, 7-hydroxycatalponol, (4S)-3,4-dihydro-4-hydroxy-2-[(2R)-2,3-dihydroxy-3-methylbutylidene]naphthalen-1(2H)-one, and (6S)-5,6-dihydro-6-hydroxy-2,2-dimethyl-2H-benzo[h]chromen-4(3H)-one and one phthalide, (±)-3-(5-hydroxy-5-methyl-2-oxohex-3-en-1-yl)isobenzofuran-1(3H)-one, were isolated from the wood of Catalpa ovata G. Don (Bignoniaceae), together with six known compounds. The structures of the previously undescribed compounds were elucidated by interpretation of 1D and 2D NMR data. The absolute configurations of the dihydronaphthalenones were deduced by analysis of the ECD data and application of Mosher ester methodology. All isolates were investigated for their cytoprotective effects against hydrogen peroxide (H2O2)-induced oxidative damage in HepG2 cells. Moreover, the mRNA expression levels of antioxidant enzymes such as heme oxygenase-1 (HO-1) and NAD(P)H:quinine oxidoreductase 1 (NQO1) in HepG2 cells were examined by RT-PCR analysis. As a result, catalponol and epi-catalponol showed antioxidant activities via directly scavenging of intracellular ROS and inducing the antioxidant enzymes in vitro.

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.

Dieckol enhances the expression of antioxidant and detoxifying enzymes by the activation of Nrf2-MAPK signalling pathway in HepG2 cells.

Dieckol was previously reported to exhibit antioxidant and anticancer activities in vitro studies. In this study, we characterised the mechanism underlying the dieckol-mediated expression of antioxidant and detoxifying enzymes. Dieckol suppressed the production of intracellular reactive oxygen species in the presence or absence of H2O2 and increased glutathione level in HepG2 cells. Dieckol enhanced the activities of antioxidant enzymes, and the expression of detoxifying enzymes including heme oxygenase-1 (HO-1), NAD(P)H:quinine oxidoreductase 1 (NQO1), and glutathione S-transferase (GST) in HepG2 cells. Enhanced expression of antioxidant and detoxifying enzymes by dieckol was presumed to be the activation of the nuclear factor erythroid-derived 2-like 2 (Nrf2) demonstrated by its nuclear translocation and transcriptional activity via activation of mitogen-activated protein kinases in HepG2 cells. Furthermore, we demonstrated dieckol induced the expression of HO-1 in mouse liver. These results demonstrate that the dieckol-mediated cytoprotection in HepG2 cells is mediated through a ROS-independent up-regulation of antioxidant and detoxifying enzymes via Nrf2 activation as well as its intrinsic antioxidant activity, suggesting that dieckol may be used as a natural cytoprotective agent.

Correlation of Nrf2, NQO1, MRP1, cmyc and p53 in colorectal cancer and their relationships to clinicopathologic features and survival.

Due to emergence of resistant tumor populations, prognosis for metastatic colorectal cancer (CRC) patients remains poor and five-year survival rate is still very low. To guide clinicians in selecting treatment option for CRC patients, reliable markers predictive of poor clinical outcome are desirable. This study analyzed the correlation of NF-E2-related factor 2 (Nrf2), NAD(P)H quinine oxidoreductase 1 (NQO1), multidrug resistant protein 1 (MRP1), cmyc and p53 in CRC and their relationships to Duke's stage and clinical prognosis. 76 specimens of CRC tissues were immunohistochemically investigated using Nrf2, NQO1, MRP1, cmyc and p53 antibodies. IHC stain showed that Nrf2, NQO1, MRP1, cmyc and p53 were highly expressed in CRC tissues compared with adjacent non-tumor tissues. Significant positive correlations were found between the expression of Nrf2 and that of NQO1, MRP1, cmyc and p53. Moreover, there was significant correlation between the high level of Nrf2, NQO1, MRP1, p53 expression and Duke's stage, as well as poor clinical prognosis. We confirmed that Nrf2, NQO1, MRP1, and p53 expression exhibits considerable heterogeneity according to CRC clinical stage and prognosis. Nrf2 is the most promising biomarker in identifying a poor prognostic group of CRC.

Curcumin attenuates arsenic-induced hepatic injuries and oxidative stress in experimental mice through activation of Nrf2 pathway, promotion of arsenic methylation and urinary excretion.

Oxidative stress is one of the major mechanisms implicated in inorganic arsenic poisoning. Curcumin is a natural phenolic compound with impressive antioxidant properties. What's more, curcumin is recently proved to exert its chemopreventive effects partly through the activation of nuclear factor (erythroid-2 related) factor 2 (Nrf2) and its antioxidant and phase II detoxifying enzymes. In vivo, we investigated the protective effects of curcumin against arsenic-induced hepatotoxicity and oxidative injuries. Our results showed that arsenic-induced elevation of serum alanine amino transferase (ALT) and aspartate aminotransferase (AST) activities, augmentation of hepatic malonaldehyde (MDA), as well as the reduction of blood and hepatic glutathione (GSH) levels, were all consistently relieved by curcumin. We also observed the involvement of curcumin in promoting arsenic methylation and urinary elimination in vivo. Furthermore, both the hepatic Nrf2 protein and two typically recognized Nrf2 downstream genes, NADP(H) quinine oxidoreductase 1 (NQO1) and heme oxygenase-1 (HO-1), were consistently up-regulated in curcumin-treated mice. Our study confirmed the antagonistic roles of curcumin to counteract inorganic arsenic-induced hepatic toxicity in vivo, and suggested that the potent Nrf2 activation capability might be valuable for the protective effects of curcumin against arsenic intoxication. This provides a potential useful chemopreventive dietary component for human populations.

Effect of ulinastatin on oxidative stress in the lung tissue of acute H2 S-intoxicated rats / 中华急诊医学杂志

Objective To observe the dynamic changes of heme oxygenase 1,NAD(P)H:quinine oxidoreductase 1 and Nuclear factor-E2-related factor 2 in the lung tissue of acute H2S-intoxicated rats and intervention effects of ulinastratin(UTI).Methods A total of 96 SD rats of clean grade were divided randomly(random number)into four groups:normal control group(NS group,n =8),UTI control group(UTI group,n =8),H2S-intoxicated model group(H2S group,n =40,rats were exposed to H2S(200 × 10-6)for 1 h to establish the H2S-intoxicated model)and UTI treatment group(H2S +UTI group,n =40,rats were intraperitoneal injected with the dose of UTI 105 U/kg).H2S group and H2S + UTI group were sacrificed 2,6,12,24 and 48 h after modeling.The activity and mRNA expression of HO-1 and NQO-1 in the lung tissue were measured by ELISA and RT-PCR methods,and the expression of Nrf2 mRNA and protein in the lung tissue was detected by RT-PCR and Western Blot methods.Pathological changes of lung tissue were observed by lightmicroscope and the lung injury score was used to evaluate inhalation injury.Results The pulmonary HO-1 activity and mRNA expression in rats of H2S group at 2,6,12 h(P ＜ 0.01)after intoxication were markedly increased than that in NS group:In comparison with H2S group,the pulmonary HO-1 activity and mRNA expression increased at 6,12,24,48 h(P ＜0.01).The pulmonary NQO-1 activity and mRNA expression in rats of H2S group at 2,6,12,24 h(P＜ 0.01)after intoxication were markedly increased than that in NS group; In comparison with H2S group,the pulmonary NQO-1 activity and mRNA expression increased at 6,12,24,48 h(P ＜ 0.01).The pulmonary Nrf2 mRNA and protein expression in rats of H2S group at 2,6,12 h(P ＜0.01 or P ＜0.05)after modeling were markedly increased than that in NS group and reached peak 2 hour after modeling; In comparison with H2S group,the pulmonary Nrf2 mRNA and protein expression increased at 6,12,24,48 h(P ＜0.01).At 24 h after modeling,the degree of lung damage were also decreased in H2S group compared with H2S + UTI group in the lightmicroscope.Histopathological examination showed that the degree of lung injury in H2S + UTI group was less severe than that in H2S group especially in the 12,24 and 48 h (P ＜0.01).Conclusions HO-1,NQO-1 and Nrf2 are involved in the pathogenesis of acute lung injury induced by H2S-intoxicated in rats.UTI may improve the imbalance in redox and activate HO-1,NQO-1 and Nrf2 can reduce lung injury and protect the lung injury induced by H2S in rats.

NAD(P)H:quinone oxidoreductase 1 Pro187Ser polymorphism and the risk of lung cancer: A meta-analysis.

BACKGROUND: NAD(P)H:quinine oxidoreductase 1 (NQO1) is a two-electron reductase that catalyzes quinine to hydroquinone. Variant genotypes of NQO1 Pro187Ser may be related to low enzyme activity and thus are suggested as affecting the risk of lung cancer. Our purpose was to study the NQO1 Pro187Ser polymorphism and the risk of lung cancer. METHODS: We conducted a PubMed search and a China National Knowledge Infrastructure search using "NAD(P)H quinine reductase 1" ,"lung cancer", and " polymorphism" for articles published from January 1997 to March 2010. For each study, we calculated the crude odds ratios (OR) and 95% confidence intervals (CI) for lung cancer. Summary estimates for crude as well as adjusted OR were calculated with the statistical program stata. RESULTS: The summary OR for carrying one variant allele and the homozygous variant genotype was 1.04 (95%CI, 0.97-1.13). In the Asian population, the summary OR for carrying one variant genotype was 1.09 (95%CI, 0.95-1.25). CONCLUSIONS: Through our meta-analysis, we found that the Pro/Ser and Ser/Ser genotypes involving NQO1 were non-significantly associated with lung cancer. In the Asian population, the combined genotype was marginally associated with increased risk of lung cancer. When stratified on gender and smoking, the meta-analysis showed that there was no statistically significant difference between men and women, smokers and non-smokers, respectively.