Activation of Nrf2/HO-1/NQO1 Signaling Pathway by Shenqi Tangluo Pill Improves Oxidative Stress Injury of Skeletal Muscle of Type 2 Diabetes Mellitus Mice / 中国实验方剂学杂志

ObjectiveTo investigate the effect and mechanism of Shenqi Tangluo pill （SQTLP） on oxidative stress injury of skeletal muscle of type 2 diabetes mellitus （T2DM） mice based on nuclear factor erythroid 2-related factor 2 （Nrf2）/heme oxygenase-1 （HO-1）/NAD（P）H quinone oxidoreductase 1 （NQO1） pathway. MethodA total of 60 7-week-old male db/db mice ［specific pathogen-free （SPF） grade］ were selected and fed for one week for adaption. They were divided into the model control group， SQTLP low-， medium- and high-dose （19， 38， and 76 g·kg-1） groups and metformin group （0.26 g·kg-1） by gavage. Each group consisted of 12 mice. Twelve male db/m mice of the same age were selected as the blank group. The intervention was implemented continuously for 8 weeks. Fasting blood glucose （FBG） was detected. Fasting serum insulin （FINS） levels were detected by enzyme-linked immunosorbent assay （ELISA）， and the homeostasis model assessment-insulin resistance （HOMA-IR） index and the homeostasis model assessment-insulin sensitivity index （HOMA-ISI） were calculated. Oral glucose tolerance test （OGTT） and insulin tolerance test （ITT） were conducted. The activities of superoxide dismutase （SOD） and glutathione peroxidase （GSH-Px） and the contents of malondialdehyde （MDA） and reduced nicotinamide adenine dinucleotide phosphate （NADPH） in skeletal muscle tissues were detected by biochemical kits. Hematoxylin-eosin （HE） staining was used to observe the pathological changes in skeletal muscle tissues. The levels of reactive oxygen species （ROS） and 4-hydroxynonenal （4-HNE） in skeletal muscle tissue were detected by immunofluorescence （IF）. The expression levels of Nrf2， HO-1， NQO1 and glutamate-cysteine ligase catalytic subunit （GCLC） proteins in skeletal muscle tissues were detected by Western blot. ResultCompared with those in the blank group， FBG， FINS and HOMA-IR in the model group were significantly increased （P<0.05）， while HOMA-ISI was decreased （P<0.05）. The results of OGTT and ITT showed that blood glucose was significantly increased at all time points （P<0.05）， and glucose tolerance and insulin tolerance were significantly impaired. SOD and GSH-Px activities in skeletal muscle tissues were significantly decreased （P<0.05）， and MDA and NADPH contents were significantly increased （P<0.05）. In skeletal muscle tissues， the arrangement of muscle fibers was loose， the nucleus was disordered， and inflammatory cells were infiltrated. The expression levels of ROS and 4-HNE in skeletal muscle tissues were significantly increased （P<0.05）. The protein expression levels of Nrf2， HO-1， NQO1 and GCLC in skeletal muscle tissues were significantly decreased （P<0.05）. Compared with those in the model group， FBG， FINS and HOMA-IR in the metformin group were significantly decreased （P<0.05）， while HOMA-ISI was increased （P<0.05）. The results of OGTT and ITT showed that blood glucose in the metformin group was significantly decreased at all time points （P<0.05）. The activities of SOD and GSH-Px in skeletal muscle tissues were significantly increased （P<0.05）， while the contents of MDA and NADPH were significantly decreased （P<0.05）. No obvious abnormality was found in the skeletal muscle tissue of the metformin group. The expressions of ROS and 4-HNE in skeletal muscle tissues were decreased （P<0.05）. The protein expression levels of Nrf2， HO-1， NQO1 and GCLC in skeletal muscle tissues were significantly increased （P<0.05）. Compared with those in the model group， FBG， FINS and HOMA-IR in the SQTLP medium- and high-dose groups were significantly decreased （P<0.05）， while HOMA-ISI was increased （P<0.05）. The results of OGTT and ITT showed that the glucose tolerance and insulin tolerance of mice were improved in each dose group of SQTLP. The GSH-Px activity in the SQTLP low-dose group was significantly increased （P<0.05）， and the NADPH content was decreased （P<0.05）. The activities of SOD and GSH-Px in the SQTLP medium- and high-dose groups were significantly increased （P<0.05）， while the contents of MDA and NADPH were significantly decreased （P<0.05）. The skeletal muscle tissue injury of mice in each dose group of SQTLP was ameliorated to different degrees. In the SQTLP medium- and high-dose groups， the expressions of ROS and 4-HNE were decreased （P<0.05）， and the protein expression levels of Nrf2， HO-1， NQO1 and GCLC were significantly increased （P<0.05）. Compared with those in the SQTLP low-dose group， FBG and HOMA-IR in the SQTLP high-dose group were significantly decreased （P<0.05）， while HOMA-ISI was increased （P<0.05）. The results of OGTT and ITT showed that the SQTLP high-dose group significantly improved the glucose tolerance and insulin tolerance of mice. The activities of SOD and GSH-Px in skeletal muscle tissues were significantly increased （P<0.05）， while the contents of MDA and NADPH were significantly decreased （P<0.05）. No obvious abnormality was found in the skeletal muscle tissue， the expressions of ROS and 4-HNE were decreased （P<0.05）， and the protein expression levels of Nrf2， HO-1， NQO1 and GCLC were significantly increased （P<0.05） in the skeletal muscle tissue of the SQTLP high-dose group. ConclusionSQTLP can significantly improve IR in T2DM mice， and the mechanism is related to SQTLP activating the Nrf2/HO-1/NQO1 signaling pathway， promoting the expression of antioxidant enzymes， and thus improving the oxidative stress injury in the skeletal muscle.

Discovery of proqodine A derivatives with antitumor activity targeting NAD(P)H: quinone oxidoreductase 1 and nicotinamide phosphoribosyltransferase / 中国天然药物

NAD(P)H: quinone oxidoreductase 1 (NQO1) is a flavin protease highly expressed in various cancer cells. NQO1 catalyzes a futile redox cycle in substrates, leading to substantial reactive oxygen species (ROS) production. This ROS generation results in extensive DNA damage and elevated poly (ADP-ribose) polymerase 1 (PARP1)-mediated consumption of nicotinamide adenine dinucleotide (NAD+), ultimately causing cell death. Nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in the NAD+ salvage synthesis pathway, emerges as a critical target in cancer therapy. The concurrent inhibition of NQO1 and NAMPT triggers hyperactivation of PARP1 and intensive NAD+ depletion. In this study, we designed, synthesized, and assessed a novel series of proqodine A derivatives targeting both NQO1 and NAMPT. Among these, compound T8 demonstrated potent antitumor properties. Specifically, T8 selectively inhibited the proliferation of MCF-7 cells and induced apoptosis through mechanisms dependent on both NQO1 and NAMPT. This discovery offers a promising new molecular entity for advancing anticancer research.

Effect and Mechanism of Banxia Xiexintang on Rats with Chronic Atrophic Gastritis Based on Keap1/Nrf2/ARE Signaling Pathway / 中国实验方剂学杂志

Objective:To explore the mechanism of Banxia Xiexintang （BXXX） in preventing and treating chronic atrophic gastritis （CAG） through Kelch-like ECH-associated protein 1 （Keap1）/nuclear factor erythroid 2-related factor 2 （Nrf2）/antioxidant response element （ARE） signaling pathway. Method:SD rats were divided into a normal group （<italic>n</italic>=12） and an experimental group for CAG model induction. The model rats were then randomly divided into a model group， a vatacoenayme （VG） group （60 mg·kg^-1）， and high- （280 mg·kg^-1）， medium- （140 mg·kg^-1）， and low-dose （70 mg·kg^-1） BXXX groups. The doses in the BXXX groups were equivalent to 28， 14， and 7 g·kg^-1 crude drugs. The rats in the normal group and the model group received distilled water at an equal volume， and those in the VG group and the BXXX groups were treated correspondingly by gavage. After 12 weeks of treatment， hematoxylin-eosin （HE） staining was carried out to observe pathological changes in the gastric mucosa of CAG rats. Western blot and real-time fluorescence-based quantitative PCR was used to detect the protein and mRNA expression levels of Nrf2， glutathione S-transferase （GST）， and NAD （P）H：quinone oxidoreductase 1 （NQO1） in the gastric mucosa of CAG rats. Result:Compared with the normal group， the model group showed increased protein and mRNA expression levels of Nrf2， NQO1， and GST in the gastric mucosa of the rats （<italic>P</italic><0.05）， atrophic gastric mucosa， and even intestinal metaplasia. The protein and mRNA expression levels of Nrf2， NQO1， and GST in the VG group and the high- and medium-dose BXXX groups were lower than those in the model group （<italic>P</italic><0.05）， and gastric mucosa atrophy and intestinal metaplasia were significantly improved， especially in the high-dose BXXX group. However， the effect in the low-dose BXXX group was not significant. Conclusion:BXXX can blunt the transcriptional activity of Nrf2， shut down Nrf2 signaling pathway， and reduce the expression levels of NQO1 and GST to achieve normal oxidation-anti-oxidation balance， which may be one of its action mechanisms in the treatment of CAG.

Caffeic acid and protocatechuic acid modulate Nrf2 and inhibit Ehrlich ascites carcinomas in mice

Objective: To assess the nuclear factor-erythroid 2-related factor-2 (Nrf2) modulatory effect of caffeic acid and protocatechuic acid and determine the anti-tumor activity of these phenolic compounds against Ehrlich ascites carcinoma growth in mice. Methods: Antioxidant activity of protocatechuic acid and caffeic acid was assessed using ferric reducing antioxidant power (FRAP) and 2,2-diphenyl-1-picrylhydrazyl (DPPH). Nrf2 activation potential of phenolic compounds was tested by quantitative realtime polymerase chain reaction, and luciferase complementation reporter assays. In vivo efficacy was tested using the Ehrlich ascites carcinoma model. Results: FRAP and DPPH radical scavenging assays showed that caffeic acid and protocatechuic acid were more potent compared with cinnamic acid and benzoic acid. Luciferase complementation reporter assays identified caffeic acid and protocatechuic acid as the activators of Nrf2. Both caffeic acid and protocatechuic acid upregulated the expression of Nrf2 target genes heme oxygenase-1 (HO-1), glutamate-cysteine ligase catalytic subunit (GCLC), and glutamate-cysteine ligase modifier subunit (GCLM) and the activity of NAD(P)H:quinone oxidoreductase 1 (NQO1) when tested on HCT-116 cells using a cell-based assay system at 9 h. In addition, intraperitoneal administration of caffeic acid and protocatechuic acid to Ehrlich ascites carcinoma bearing mice suppressed tumor growth and angiogenesis. Conclusions: Caffeic acid and protocatechuic acid can modulate Nrf2 and inhibit Ehrlich ascites carcinoma cells.

Effect of Nrf2/HO-1 signaling pathway in intestinal protection by Sishen Pills against ulcerative colitis in mice / 中国中药杂志

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.

Naringenin increases Nrf2/phase II detoxifying enzymes to reduce vascular inflammation / 中国药理学通报

Aim To investigate the role of naringenin in nuclear factor erythroid 2-related factor 2 (Nrf2)/ phase II detoxifying enzyme activities and evaluate its effects on vascular inflammation. Methods Western blot, immunofluorescence and reverse transcription- qPCR were used to detect the protein expression. The activities of phase II detoxifying enzymes were measured by commercial kits. Immunoprecipitation technology was used to detect the interaction between Nrf2 and kelch-like ECH-associated protein 1 ( Keap-1). Results Naringenin promoted the dislocation of Nr£2 from Keap-1 and increased Nrf2 nuclear accumulation in RAW264. 7 macrophages. Naringenin up-regulated expressions of phase II detoxifying enzymes such as NAD(P)H quinone oxidoreductase ( NQO-1), gluta thione S-transferase (GST) and glutamate-cysteine lig- ase (GCL). It also reduced the levels of cytokines in macrophages. Moreover, the Nrf2 inhibitor ML385 reduced phase II detoxifying enzyme expressions and increased cytokine levels. In addition, we found naringenin increased the expressions and activities of liver phase II detoxifying enzymes ( NQO-1, GST and GCL) and reduced aortic inflammation in atherosclerotic model mice. The effects were dependent on Nr£2 activity. Conclusions Naringenin activates Nrf2 and promotes phase II detoxifying enzyme activities, which leads to the inhibition of vascular inflammation.

Anti-Oxidation Mechanism of Andrographolide on HaCaT Cells Via Nrf2/ARE Pathway / 中国药学杂志

OBJECTIVE: To investigate the anti-oxidant mechanism of andrographolide on HaCaT cells via Nrf2/ARE signal pathway. METHODS: The effect of andrographolide on the viability of HaCaT cells and the effect of H2O2-induced cell viability were measured by CCK-8. HaCaT cells were pretreated with andrographolide of different concentration for 24 h. The protein and mRNA expression levels of Nrf2, HO-1, AKR1C1 and NQO1 in HaCaT cells were detected by Western blot and RT-qPCR, respectively. The expression of Nrf2 protein in the nucleus was analyzed by nuclear cytoplasmic separation and immunofluorescence. RESULTS: Andrographolide had no significant effect on cell viability and dose-dependently decreased H2O2-induced cell death, the difference was statistically significant. Andrographolide significantly enhanced the expression of protein and mRNA of antioxidant enzymes Nrf2, HO-1, AKR1C1, NQO1, increased the distribution of Nrf2 in the nucleus, and up-regulated the expression of ARE. Besides, andrographolide upregulated the phosphorylation level of the upstream protein kinase AMPKα (p-AMPKα). CONCLUSION: Andrographolide could decrease H2O2-induced cell death, and its mechanism may be through the activation of Nrf2/ARE signaling pathway, thereby regulating the expression levels of HO-1, AKR1C1, and NQO1.

Effect of Duzhong Butiansu Capsule on improving spermatogenesis obstacle in mice induced by cyclophosphamide / 中草药

Objective: To investigate the reproductive protective effect of Duzhong Butiansu Capsule (DBC) by using cyclophosphamide induced spermatogenic disorder model, and explore its mechanism. Methods: The model of spermatogenic disorder was established by intraperitoneal injection of cyclophosphamide (60 mg/kg) for 5 d. Drug intervention at high and low doses (1.388 g/kg and 0.694 g/kg) was given for 4 weeks from the 8th day. The body weight and organ index of each group were measured. The pathological structure of testis was detected by HE staining. ELISA method was used to detect the levels of T, FSH, LH, MDA, SOD, GSH-Px. Western blotting and immunohistochemistry were used to analyze the expression of Nrf2/ARE signaling pathway related factors Nrf2, HO-1, NQO1, HDAC2, and p-PKC in testicular tissue. Results: Compared with the model group, DBC significantly reduced the weight of mice, increased the index of testis, epididymis and kidney, improved the pathological morphology of testis, increased the number of spermatozoa, increased the motility of sperm, decreased the rate of abnormal sperm, increased the level of T and decreasd the level of LH and FSH, increased the content of MDA, decreased the content of SOD and GSH-Px, increased the expression of Nrf2, HO-1, NQO1, HDAC2, and p-PKC protein, and increased the area of positive expression of Nrf2, HO-1 protein (P < 0.05 or P < 0.01). Conclusion: DBC can obviously improve the spermatogenic disorder induced by cyclophosphamide, and the mechanism may be related to the regulation of Nrf2/ARE signal pathway associated with oxidative stress.

Expression and significance of NAD (P) H: quinone oxidoreductase 1 and heme oxygen-ase-1 proteins in T-cell lymphoma / 中国肿瘤临床

Objective: To investigate the expression of NAD (P) H: quinone oxidoreductase 1 (NQO1) and heme oxygenase-1 (HO-1) in T-cell lymphoma (TCL), and investigate the correlation between these two indicators and other clinicopathological parameters in TCL. Methods: Clinical data of 61 patients with TCL who were initially diagnosed in Gansu Provincial Hospital were analyzed retrospectively. Immunohistochemical examination was performed to detect NQO1 and HO-1 expression levels in 61 TCL tissues (TCL group) and 20 lymph node reactive hyperplasia tissues (control group). Results: Positive expression rates of NQO1 and HO-1 were significantly higher in TCL tissues than in lymph node reactive hyperplasia tissues (P<0.05). NQO1 expression was closely related with Ann-Arbor clinical stage and B symptoms (P<0.05); HO-1 expression was correlated with clinical stage, bone marrow invasion, and B symptoms (P<0.05). NQO1 and HO-1 expression levels were not related to age, sex, lactate dehydrogenase level, and pathological type (P>0.05); there was a correlation between NQO1 and HO-1 expression (r=0.264; P=0.040). Conclusions: NQO1 and HO-1 are highly expressed in TCL and may interact and contribute to the occurrence and development of TCL.

NQO1 dependent non-canonical necroptosis mediated by ROS and RIP1/RIP3 in parallel in glioma cancer cells / 中国药理学与毒理学杂志

OBJECTIVE Glioblastomas(GBM)are the most malignant brain tumors in humans and have a very poor prognosis. New therapeutics are urgently needed. Here, we reported 2-methoxy-6-acetyl-7-methyljuglone (MAM)-induced cell death in U87 and U251 glioma cancer cells. METHODS Cells were cultured and treated with MAM, the cell viability was determined by MTT assay and LDH assay. Intracellular reactive oxygen species (ROS) generation was observed by DCF fluorescence. The protein expression was determined by Western blotting. RESULTS MAM induced glioma cancer cell death without caspase activation. The cell death induced by MAM was attenuated by the pharmacological or genetic blockage of necroptosis signaling,including RIP1 inhibitor necrostatin-1s (Nec-1s)and siRNA-mediated gene silencing of RIP1 and RIP3,but was unaffected by caspase inhibitor z-vad-fmk or necrosis inhibitor 2-(1H-Indol-3-yl)-3-pentylamino-maleimide (IM54). MAM treated U87 and U251 glioma cancer cells induced RIP1/RIP3 complex formation, ROS level increased, ATP concentration decreased and loss of plasma membrane integrity, further confirmed this process was necroptosis.The essential role of ROS was confirmed by the protective effect of ROS scavenger NAC. Interestingly, MAM induced necroptosis both triggered by RIP1/RIP3 complex and ROS generation. Moreover, MAM induced necroptosis through cytosolic calcium (Ca2 +) accumulation and sustained c-Jun N-terminal kinase (JNK) activation. Both calcium chelator BAPTA-AM and JNK inhibitor SP600125 could attenuate cell death. Further, we found there exists a feedback loop between RIP1 and JNK activation.Finally,MAM induced necroptosis was inhibited by dicoumarol(a NQO1 inhibitor). Dicoumarol exposed glioma cancer cells were resistant to RIP1/RIP3 complex formation and ROS generation. MAM induced necroptosis was independent of MLKL. CONCLUSION MAM induced non-canonical necroptosis through the NQO1-dependent ROS and RIP1/RIP3 pathway.This study also provided new insights into the molecular regulation of necroptosis in human glioma cancer cells and a promising approach for GBM treatment.

Expressions of NQO1 and iNOS in different molecular subtypes of breast cancer and their clinical signifi-cance / 实用医学杂志

Objective To investigate the difference of iNOS and NQO1 in breast cancer with different mo-lecular subtypes and the correlation between clinical parameters,and to explore the clinical treatment of breast can-cer. Methods The data on 100 patients with breast cancer who had undergone modified radical mastectomy was retrospectively reviewed.Expressions of iNOS and NQO1 were detected by immunohistochemistry.Numeration data was processed using χ2test or Fisher′s exact test.Spearman correlation was applied to analyze different clinical mo-lecular pathological features.Results NQO1 expression was correlated with TNM staging and lymph node metasta-sis(χ2=6.603,4.938,P<0.05),and the expression rate of NQO1 in overexpressing of HER-2 was higher than that in type luminal A(χ2=8.341,P < 0.008). Expression of iNOS was related to expression of Ki-67,stage of TNM and lymph node metastasis(χ2=5.243,8.157,and 5.929,P<0.05),the expression rate of iNOS in lumi-nal A was significantly lower than that in type luminal B(χ2=7.990,P<0.008).Conclusions The expression of NQO1 and iNOS may be involved the process of oxidative stress associated with breast cancer,and it plays an im-portant role in the development of breast cancer. Different molecular level of oxidative stress between the types of breast cancer may be different,whose molecular mechanism needs to be further studied.

Tocotrienol-rich Palm Oil Extract Induces NAD(P)H:quinone Oxidoreductase 1 (NQO1) Expression in Mice Liver.

A diet rich in tocotrienols has been shown to be beneficial for health. However, its detailed mechanism of action is still not fully understood. NAD(P)H:quinone oxidoreductase 1 (NQO1) is important in cellular defence due to its ability to detoxify reactive quinones and quinoneimines to their less toxic hydroquinones forms. The objective of this study is to investigate the effects of different doses of palm oil-derived tocotrienol rich fraction (palm TRF) supplementation on NQO1 gene and protein expression in mice livers. Western blot and qPCR assays were used to detect NQO1 expression levels. It was found that palm TRF significantly induced NQO1 expression at all doses given. In conclusion, palm TRF treatment increased NQO1 gene and protein expression in mice liver dose dependently, with the highest expression seen in mice treated with 1000 mg/kg palm TRF, followed by 500 and 200 mg/kg respectively.

Increased Cellular NAD⁺ Level through NQO1 Enzymatic Action Has Protective Effects on Bleomycin-Induced Lung Fibrosis in Mice / 결핵및호흡기질환

BACKGROUND: Idiopathic pulmonary fibrosis is a common interstitial lung disease; it is a chronic, progressive, and fatal lung disease of unknown etiology. Over the last two decades, knowledge about the underlying mechanisms of pulmonary fibrosis has improved markedly and facilitated the identification of potential targets for novel therapies. However, despite the large number of antifibrotic drugs being described in experimental pre-clinical studies, the translation of these findings into clinical practices has not been accomplished yet. NADH:quinone oxidoreductase 1 (NQO1) is a homodimeric enzyme that catalyzes the oxidation of NADH to NAD+ by various quinones and thereby elevates the intracellular NAD⁺ levels. In this study, we examined the effect of increase in cellular NAD⁺ levels on bleomycin-induced lung fibrosis in mice. METHODS: C57BL/6 mice were treated with intratracheal instillation of bleomycin. The mice were orally administered with β-lapachone from 3 days before exposure to bleomycin to 1-3 weeks after exposure to bleomycin. Bronchoalveolar lavage fluid (BALF) was collected for analyzing the infiltration of immune cells. In vitro, A549 cells were treated with transforming growth factor β1 (TGF-β1) and β-lapachone to analyze the extracellular matrix (ECM) and epithelial-mesenchymal transition (EMT). RESULTS: β-Lapachone strongly attenuated bleomycin-induced lung inflammation and fibrosis, characterized by histological staining, infiltrated immune cells in BALF, inflammatory cytokines, fibrotic score, and TGF-β1, α-smooth muscle actin accumulation. In addition, β-lapachone showed a protective role in TGF-β1–induced ECM expression and EMT in A549 cells. CONCLUSION: Our results suggest that β-lapachone can protect against bleomycin-induced lung inflammation and fibrosis in mice and TGF-β1–induced EMT in vitro, by elevating the NAD+/NADH ratio through NQO1 activation.

NAMPT inhibition synergizes with NQO1-targeting agents in inducing apoptotic cell death in non-small cell lung cancer cells / 中国天然药物

Nicotinamide phosphoribosyltransferase (NAMPT) catalyzes the first rate-limiting step in converting nicotinamide to NAD(+), essential for a number of enzymes and regulatory proteins involved in a variety of cellular processes, including deacetylation enzyme SIRT1 which modulates several tumor suppressors such as p53 and FOXO. Herein we report that NQO1 substrates Tanshione IIA (TSA) and β-lapachone (β-lap) induced a rapid depletion of NAD(+) pool but adaptively a significant upregulation of NAMPT. NAMPT inhibition by FK866 at a nontoxic dose significantly enhanced NQO1-targeting agent-induced apoptotic cell death. Compared with TSA or β-lap treatment alone, co-treatment with FK866 induced a more dramatic depletion of NAD(+), repression of SIRT1 activity, and thereby the increased accumulation of acetylated FOXO1 and the activation of apoptotic pathway. In conclusion, the results from the present study support that NAMPT inhibition can synergize with NQO1 activation to induce apoptotic cell death, thereby providing a new rationale for the development of combinative therapeutic drugs in combating non-small lung cancer.

NQO1 overexpression is associated with poor prognosis in the uterine cervix carcinoma / 实用肿瘤学杂志

Objective To investigate the clinicopathological significance of upregulated NQO 1 protein expression in uterine cervix carcinoma ( UCC) .Methods Immunohistochemical staining was performed on paraf-fin-embedded UCC specimens from 123 patients.Disease-free survival(DFS)and overall survival(OS)rates for all cervical UCC patients were calculated using the Kaplan -Meier method ,and univariate or multivariate analyses were performed using the Cox proportional hazards regression model .Results The NQO1 protein showed a main-ly cytoplasmic staining pattern in cervical cancer cells ,and the strongly positive rate of NQO 1 was significantly higher in UCC.High-level NQO1 was closely associated with poor differentiation ,late-stage,lymph node metas-tasis and high-risk for HPV infection.Additionally,high-level NQO1 was associated with lower DFS and OS rates .Furthermore ,Cox analysis revealed that NQO 1 expression emerged as a significant independent hazard factor for DFS rate in patients with UCC .Conclusion NQO1 overexpression might be an independent biomarker for prognostic evaluation of UCCs .

Interactive Effect of Smoking and NQO1 Haplotypes on Lung Cancer Risk

The role of genetic polymorphisms of NAD(P)H:quinone oxidoreductase 1 (NQO1), which is known to be related to carcinogen metabolism and oxidative status, was evaluated for lung cancer development. The genotypes of two NQO1 polymorphisms, namely, IVS1-27C>G and Ex6+40C>T, were determined in 616 lung cancer cases and 616 lung cancer-free controls and haplotypes composed of the two polymorphisms were estimated. In the evaluation of the effect of the NQO1 genotypes or diplotypes, we did not find any significant association with lung cancer risk after adjusting for body mass index and smoking status. However, when we evaluated the effect of the NQO1 diplotypes for lung cancer risk in combination with smoking, smokers without the C-T/C-T diplotype showed a significantly increased risk of lung cancer compared with nonsmokers without the C-T/C-T diplotype (adjusted OR, 2.2; 95% CI, 1.67-3.02), and smokers with the C-T/C-T diplotype showed the highest OR of lung cancer (adjusted OR, 2.7; 95% CI, 1.78-4.21). Moreover, a trend test showed an additive interaction between smoking and the NQO1 C-T/C-T diplotype (P(trend) < 0.01). The additive effect of smoking and the NQO1 C-T/C-T diplotype was more apparent in squamous cell carcinoma, although this effect was statistically significant in all lung cancer cell types (all cell types, P(trend) < 0.05). This result suggests that haplotypes of the NQO1 gene play an important role in the development of lung cancer by interaction with smoking.

Relationship and significance between C609 T polymorphism of NQO1 gene and breast cancer molecular subtype / 临床与实验病理学杂志

Purpose To investigate the distribution of polymorphisms of quinone oxidoreductase 1 (NQO1) C609T gene in breast cancer patients, and to analyze the relationship with breast cancer molecular subtype. Methods Genotyping of C609T rs1800566 lo-cus of NQO1 gene in peripheral blood of 248 cases of female breast cancer were detected using high-throughput TaqMan MGB real-time fluorescence quantitative PCR technology, while the detection of ER, PR, HER-2 and Ki-67 in cancerous tissues were used with immu-nohistochemical staining and FISH gene amplification. Results Among 248 cases of breast cancer patients, CC genotype accounted for 27. 42% (68/248), CT genotype accounted for 49. 60% (123/248), TT genotype accounted for 22. 98% (57/248), which con-sistent with Hardy-Weinberg equilibrium law genetic (P>0. 05). 5 cases of HER-2 (++) who did not undergo FISH testing were re-moved, all the rest were done with FISH detection. Luminal A type accounted for 15. 2% (37/243), Luminal B type accounted for 51. 4% (125/243), HER-2 overexpression type accounted for 19. 8% (48/243), basal cell type accounted for 13. 6% (33/243). Compared with patients carrying the CC genotype, ER and PR positive rates in breast cancer patients carrying CT and TT genotype was significantly higher (P0. 05). There was no statistically difference on distribution of C609T polymorphism of NQO1 gene among different molecular sub-types of breast cancer (P>0. 05). Conclusions Here is no relationship between C609T polymorphism of NQO1 gene and breast cancer molecular subtype, miss rate of NQO1 ( CT+TT) in basal cell carcinoma is lower, and its gene polymorphism may provide the reasonable explanation to the heterogeneity of breast cancer molecular subtype.

Inhibition of thymoquinone on tumor growth of ovarian cancer and its mechanism / 中草药

Objective: To study the effect and mechanism of thymoquinone in the growth inhibition of ovarian cancer. Methods: After ovarian cancer cells SKOV3 were treated with thymoquinone, cell growth was observed by the CCK-8 assay and apoptosis was evaluated by ELISA. ROS in SKOV3 cells was detected by fluorescent probe. The expression of Keap1 protein, nuclear Nrf2 protein, P-Akt protein, Akt protein, NQO1 protein, and GCLC protein in SKOV3 cells was analyzed by Western blotting assay. NQO1 and GCLC mRNA were analyzed by RT-PCR. SKOV3 cells were injected into nude mice to establish engrafted tumor model. Immunohistochemistry was used to detect the positive expression of NQO1, GCLC, and Nrf2 in the ovarian tumors. Results: Compared with the control group, thymoquinone significantly inhibited the proliferation of SKOV3 cells and increased the apoptosis (P < 0.05, 0.01). Thymoquinone could lessen the expression of nuclear Nrf2 protein and P-Akt protein (P < 0.05), but significantly increase the expression of Keap1 protein (P < 0.001). Protein and mRNA expression of NQO1 and GCLC in SKOV3 cells treaded by thymoquinone, were lower than those in the control group (P < 0.05, 0.01). The positive expression of NQO1, GCLC, and Nrf2 was also decreased in ovarian tumors after treatment of thymoquinone (P < 0.05, 0.01). Conclusion: Thymoquinone could inhibit the proliferation of ovarian cancer cells by inhibiting the expression of nuclear Nrf2, increasing the generation of ROS in ovarian cancer cells to promote apoptosis.

A Review of NAD(P)H:Quinone Oxidoreductase 1 (NQO1); A Multifunctional Antioxidant Enzyme.

NAD(P)H:quinone-oxidoreductase-1 (NQO1) is a cytosolic enzyme that catalyses two- or four-electron reduction of many endogenous and environmental quinones using flavin adenine dinucleotide (FAD) as a cofactor. It is a cytosolic enzyme exists as a homodimer and is biochemically distinguished by its prominent ability to use either NADH or NADPH as reducing cofactors and by its suppression by the anticoagulant dicumarol. This enzyme generally considered as a detoxification enzyme due to of its ability to diminish reactive quinones and quinoneimines to its less reactive and less toxic hydroquinones forms. NQO1 is a substantially inducible enzyme that is controlled by the Nrf2-Keap1/ARE pathway. Evidence for the significance of the antioxidant functions of NQO1 in suppression of oxidative stress is provided by manifestations that induction of NQO1 levels or their reduction are associated with reduced and raised susceptibilities to oxidative stress, respectively. The gene coding for NQO1 has two common polymorphisms at nucleotide position 609(C-T) and 465 (C-T) of the human cDNA. C465T causes reduction in enzyme activity, whereas the C609T results in complete loss of enzymatic activity due to protein instability.In this review, we discuss the protective functions of NQO1 and present its possible transcriptional pathways regulating its induction by Nrf2-Keap1/ARE pathway.

CYP17, CYP19, and NQO1 Genetic Polymorphisms and Breast Cancer Susceptibility in Young Women in Brazil.

Aims: Breast cancer is the most common cancer in women worldwide, being exposure to estrogens the acknowledged main risk factor. CYP17, CYP19 and NQO1 are enzymes involved in the estrogen metabolism, so their polymorphisms may be involved in breast carcinogenesis. The aim of this study was to determine the magnitude of the association between CYP17 MspA1, CYP19 Arg264Cys, and NQO1 C609T polymorphisms and breast cancer in young women. Methods: This is a hospital-based case-control study carried out in Rio de Janeiro. Cases were 270 women with age range 18-35 years and a histopathological diagnosis of breast cancer between 1999-2009. Controls were 270 women without cancer at the same age range. Results: An association between CYP17 MspA1 or CYP19 Arg264Cys polymorphisms and breast cancer were not observed (OR = 1.02, 95% CI 0.72-1.44 for CYP17 genotypes TC/CC and OR = 0.85, 95% CI 0.48-1.49 for CYP19 genotypes CT/TT). However, a statistically significant increased risk estimate was identified in women who had at least one NQO1 polymorphic allele (T), OR= 1.96, 95% CI 1.13-3.40 following adjustment for selected confounders. Conclusion: This study suggests that the NQO1 609T polymorphism may be a risk factor for breast carcinogenesis in women less than 36 years in Brazil.