Cooperation of NAD(P)H:quinone oxidoreductase 1 and UDP-glucuronosyltransferases reduces menadione cytotoxicity in HEK293 cells
Section snippets
Materials and methods
Chemicals. DCIP, menadione, NADPH and UDPGA were purchased from Sigma–Aldrich (St. Louis, MO). TransIT®-LT1 was purchased from Mirus Bio LLC., (Madison, WI). pIRESpuro3, pTRE2hyg and 293 Tet-Off™ cell line (HEK293) were purchased from Clontech Laboratories, Inc. (Mountain View, CA). 2′,7′-Dichlorodihydrofluorescein diacetate (DCF-DA) was purchased from Molecular Probes, Inc. (Eugene, OR). [3H]menadione (185 GBq/mmol) was purchased from Moravek Biochemicals, Inc. (Brea, CA). Menadiol,
Establishment of stably transfected HEK293 cell lines
In this study, we established three HEK293 cell lines, namely, the parent HEK293 stably expressing NQO1 (HEK293/NQO1) and HEK293/NQO1 cell lines with DOX-regulated expression of UGT1A6 (HEK293/NQO1/UGT1A6) and of UGT1A10 (HEK293/NQO1/UGT1A10). In each cell line, NQO1 activity toward DCIP and menadione was significantly higher than in the HEK293 wild-type cell line. NQO1 activity did not differ significantly among the established cell lines (Table 1). In addition, we assessed the effect of DOX
Discussion
We previously suggested that menadiol, which is formed from the NQO1-mediated two-electron reduction of menadione, is not a detoxification pathway because menadiol undergoes the reformation of menadione with concomitant formation of ROS. Consequently, glucuronide conjugates remain as reduced species and therefore their formation results in detoxification of menadione [15]. In the present study, to evaluate the role of NQO1 and UGTs in menadione detoxification pathways, we established HEK293
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2021, ToxicologyCitation Excerpt :NQO1 is a cytosolic enzyme that catalyzes the two-electron reduction reaction of various quinones and quinoneimines to corresponding hydroxyquinones and hydroxyanilines, respectively (Vredenburg et al., 2014; Nishiyama et al., 2010; Zhang et al., 2018). Nishiyama et al. (2010) have reported that cytotoxicity of menadione, which possesses quinone structure, is attenuated by overexpression of NQO1 in HEK293 cells. Vredenburg et al. (2014) have reported, by an in vitro study using recombinant NQO1, that MFA-quinoneimine is reduced by NQO1.
Oxidative stress and neurodegeneration: The possible contribution of quinone reductase 2
2018, Free Radical Biology and MedicineCitation Excerpt :Previous experiments that were carried out using S29434, a specific QR2 inhibitor, in native CHO cells and in cells overexpressing QR2 demonstrated that QR2 increased ROS following exposure to ortho-quinones but decreased ROS following exposure to para-quinones [22]. Our hypothesis is that the differences in ROS production with ortho-quinones versus para-quinones could arise due to variations in the conjugation process, as proposed previously for QR1 [25]. Accordingly, we studied the ROS production consecutive to the metabolism of ortho- and para-quinones in human myelogenous leukemia (K562) cells that naturally express high levels of QR2 but only trace amounts of QR1.
In cellulo monitoring of quinone reductase activity and reactive oxygen species production during the redox cycling of 1,2 and 1,4 quinones
2015, Free Radical Biology and MedicineCitation Excerpt :One-electron reduction of quinones or their derivatives by enzymes such as cytochrome P450 reductase or other flavoproteins generates unstable semiquinones, which undergo redox cycling in the presence of molecular oxygen leading to the formation of highly reactive oxygen species. Quinone reductases catalyze the two-electron reduction of quinones to form hydroquinone that can be removed from the cell by conjugation of the hydroxyl with glucuronide or sulfate [36], thus avoiding its autoxidation and the formation of free radicals and highly reactive oxygen species. In the absence of overexpressed QR, as was the case for CHO naive cells, the quinone reduction was fully supported by one-electron reductases, which results in an overproduction of radical species.
Airborne quinones induce cytotoxicity and DNA damage in human lung epithelial A549 cells: The role of reactive oxygen species
2014, ChemosphereCitation Excerpt :PQ had the lowest LC50, indicating it had the greatest acute cytotoxic effects on A549 cells, followed by 1,2-NQ, MNQ, MAQ and ACQ. The decreased viability caused by quinones has been investigated in many cell types (Klaus et al., 2010; Nishiyama et al., 2010). The previously reported LC50 values of PQ and ACQ in A549 cells (Das et al., 2012) are identical to the data of the present study.
The chemical defensome of five model teleost fish
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