Dual effects of N-acetyl-l-cysteine dependent on NQO1 activity: Suppressive or promotive of 9,10-phenanthrenequinone-induced toxicity
Toyooka, T. ; Shinmen, T. ; Aarts, J.M.M.J.G. ; Ibuki, Y. - \ 2012
Toxicology and Applied Pharmacology 264 (2012)3. - ISSN 0041-008X - p. 404 - 412.
double-strand breaks - yeast saccharomyces-cerevisiae - diesel exhaust particles - dt-diaphorase - nad(p)h-quinone oxidoreductase - oxidative stress - dna-damage - cell-death - quinone oxidoreductases - histone h2ax
A typical antioxidant, N-acetyl-L-cysteine (NAC) generally protects cells from oxidative damage induced by reactive oxygen species (ROS). 9,10-Phenanthrenequinone (9,10-PQ), a major quinone in diesel exhaust particles, produces ROS in redox cycling following two-electron reduction by NAD(P)H:quinone oxidoreductase 1 (NQO1), which has been considered as a cause of its cyto- and genotoxicity. In this study, we show that NAC unexpectedly augments the toxicity of 9,10-PQ in cells with low NQO1 activity. In four human skin cell lines, the expression and the activity of NQO1 were lower than in human adenocarcinoma cell lines, A549 and MCF7. In the skin cells, the cytotoxicity of 9,10-PQ was significantly enhanced by addition of NAC. The formation of DNA double strand breaks accompanying phosphorylation of histone H2AX, was also remarkably augmented. On the other hand, the cyto- and genotoxicity were suppressed by addition of NAC in the adenocarcinoma cells. Two contrasting experiments: overexpression of NQO1 in CHO-K1 cells which originally expressed low NQO1 levels, and knock-down of NQO1 in the adenocarcinoma cell line A549 by transfection of RNAi, also showed that NAC suppressed 9,10-PQ-induced toxicity in cell lines expressing high NQO1 activity and enhanced it in cell lines with low NQO1 activity. The results suggested that dual effects of NAC on the cyto- and genotoxicity of 9,10-PQ were dependent on tissue-specific NQO1 activity.
NQO1 and NFE2L2 polymorphisms, fruit and vegetable intake and smoking, and the risk of colorectal adenomas in an endoscopy-based population.
Tijhuis, M.J. ; Visker, M.H.P.W. ; Aarts, J.M.M.J.G. ; Laan, A. van der; Boer, S.Y. van; Kok, F.J. ; Kampman, E. - \ 2008
International Journal of Cancer 122 (2008)8. - ISSN 0020-7136 - p. 1842 - 1848.
food frequency questionnaire - quinone oxidoreductase 1 - nad(p)h-quinone oxidoreductase - dt-diaphorase - relative validity - meat consumption - cancer - alcohol - gene - association
Both environment and genetics contribute to the pathogenesis and prevention of colorectal neoplasia. NAD(P)H:quinone oxidoreductase (NQO1) is a detoxification enzyme that is polymorphic and inducible. We investigated interactions between lifestyle factors and polymorphisms in NQO1 and its key regulatory transcription factor NFE2L2 in colorectal adenoma risk. The NQO1 c.609C>T and g.-718A>G and NFE2L2 g.-650C>A, g.-684G>A and g.-686A>G polymorphisms were determined among 740 Dutch adenoma cases and 698 endoscopy-based controls. Dietary intake was assessed by food frequency questionnaire, other lifestyle information by questionnaire. The NQO1 609CT genotype was associated with a higher adenoma risk (OR 1.27, 95% CI 1.00-1.62) compared with the 609CC genotype, whereas the 609TT genotype was not (OR 1.03, 95% CI 0.56-1.88). The higher risk with the NQO1 609CT-genotype was seen among smokers (OR 1.96, 95% CI 1.40-2.76), but not among nonsmokers (OR 0.91, 95% CI 0.62-1.35; interaction p = 0.030). Fruit and vegetable consumption did not protect smokers from adenomas and did not interact with the NQO1 609C>T polymorphism or the NFE2L2 polymorphisms. A higher adenoma risk seen with high fruit and vegetable consumption among NQO1 -718GG genotypes was absent among -718GA genotypes (interaction p = 0.071). Gene-gene interactions were observed between the NQO1 609C>T and NFE2L2 -686A>G polymorphisms (interaction p = 0.056) and between the NQO1 -718 G>A and NFE2L2 -650C>A polymorphisms (interaction p = 0.013). In conclusion: the NQO1 609CT genotype is associated with increased adenoma risk among smokers, which is not diminished by high fruit and vegetable consumption. The observed gene-gene interactions may point to a role for NFE2L2 polymorphisms in NQO1-related adenoma formation
In vivo relevance of two critical levels for NAD(P)H:quinone oxidoreductase (NQO1)-mediated cellular protection against electrophile toxicity found in vitro
Haan, L.H.J. de; Pot, G.K. ; Aarts, J.M.M.J.G. ; Rietjens, I.M.C.M. ; Alink, G.M. - \ 2006
Toxicology in Vitro 20 (2006)5. - ISSN 0887-2333 - p. 594 - 600.
dt-diaphorase - menadione toxicity - quinone toxicity - human colon - cells - nqo1 - reductase - enzymes - lines - sensitivity
NAD(P)H:quinone oxidoreductase (NQO1)-mediated detoxification of quinones is suggested to be involved in cancer prevention. In the present study, using transfected CHO cells, it was demonstrated that the relation between NQO1 activity and the resulting protection against the cytotoxicity of menadione shows a steep dose¿response curve revealing a `lower protection threshold¿ of 0.5 ¿mol DCPIP/min/mg protein and an `upper protection threshold¿ at 1 ¿mol DCPIP/min/mg protein. In an additional in vivo experiment it was investigated how both in vitro critical activity levels of NQO1, relate to NQO1 activities in mice and man, either without or upon induction of the enzyme by butylated hydroxyanisol (BHA) or indole-3-carbinol (I3C). Data from an experiment with CD1 mice revealed that base-line NQO1 levels in liver, kidney, small intestine, colon and lung are generally below the observed `lower protection threshold¿ in vitro, this also holds for most human tissue S-9 samples. To achieve NQO1 levels above this `lower protection threshold¿ will require 5¿20 fold NQO1 induction. Discussion focuses on the relevance of the in vitro NQO1 activity thresholds for the in vivo situation. We conclude that increased protection against menadione toxicity can probably not be achieved by NQO1 induction but should be achieved by other mechanisms. Whether this conclusion also holds for other electrophiles and the in vivo situation awaits further definition of their NQO1 protection thresholds
Human NAD(P)H:Quinone oxidoreductase inhibition by flavonoids in living cells
Lee, Y.Y. ; Westphal, A.H. ; Haan, L.H.J. de; Aarts, J.M.M.J.G. ; Rietjens, I.M.C.M. - \ 2005
Free Radical Biology and Medicine 39 (2005)2. - ISSN 0891-5849 - p. 257 - 265.
hamster ovary cells - dt-diaphorase - quinone oxidoreductase - acceptor oxidoreductase - antitumor quinones - human plasma - cancer risk - quercetin - reductase - rat
Procedures for assessing enzyme inhibition in living cells are an important tool in the study of the relevance of enzyme-catalyzed reactions and interactions in the human body. This paper presents the effects of flavonoids on NAD(P)H:quinone oxidoreductase 1 (NQO1) activity, by a newly developed method to measure NQO1 inhibition in intact cells. The principle of this method is based on the resorufin reductase activity of NQO1. The change in fluorescence in time was used to determine NQO1 activity in intact Chinese hamster ovary (CHO) cells genetically engineered to overexpress human NQO1. Applying this method to determine the inhibitory effects of reported in vitro NQO1 inhibitors (dicoumarol, 7,8-dihydroxyflavone, chrysin) showed that for all inhibitors tested, the IC50 in intact cells was at least 3 orders of magnitude higher than the IC50 in cell lysates. This result demonstrates that in vitro studies with purified NQO1 or with extracts from disrupted tissues are of limited value for obtaining insight into the situation in living cells. Possible factors underlying this discrepancy are being discussed. For the first time, we determined NQO1 inhibition by flavonoids in cells without disruption of the cells or addition of cofactors, enabling the assessment of enzymatic activity and the interaction of modulators of enzymatic activity in an intracellular situation.
The role of quinone reductase (NQO1) and quinone chemistry in quercetin cytotoxicity
Gliszczynska-Swiglo, A. ; Woude, H. van der; Haan, L.H.J. de; Tyrakowska, B. ; Aarts, J.M.M.J.G. ; Rietjens, I.M.C.M. - \ 2003
Toxicology in Vitro 17 (2003). - ISSN 0887-2333 - p. 423 - 431.
dt-diaphorase - dietary flavonoids - mutagenic activity - brussels-sprouts - tyrosine kinase - cell-growth - o-quinones - in-vitro - methide - cancer
The effects of quercetin on viability and proliferation of Chinese Hamster Ovary (CHO) cells and CHO cells overexpressing human quinone reductase (CHO+NQO1) were studied to investigate the involvement of the pro-oxidant quinone chemistry of quercetin. The toxicity of menadione was significantly reduced in CHO+NQO1 cells compared to wild-type CHO cells, validating the NQO1-overexpression in the CHO+NQO1 transfectant. Quercetin inhibited the proliferation of wild-type CHO and CHO+NQO1 cells to a similar extent without affecting cell viability, indicating that NQO1 enrichment of CHO cells did not provide increased protection. On the other hand, inhibition of NQO1 in both types of cells by dicoumarol significantly potentiated the inhibitory effect of quercetin on cell proliferation, revealing the role of NQO1 in cellular protection against quercetin. Altogether, these results can be explained by the hypothesis that both wild-type CHO and CHO+NQO1 cells contain sufficient NQO1 activity for optimal protection against the pro-oxidant effect of quercetin on cell proliferation. The results also point at a cellular NQO1 threshold for optimal protection against quercetin. This NQO1 threshold seems to be in the range of NQO1 activities already present in various tissues. (C) 2003 Elsevier Ltd. All rights reserved.