Coronary risk factors, including age, hypertension, diabetes mellitus, hyperlipidemia, and smoking, are associated with enhanced oxidative stress, which is implicated as a potential mechanism for atherogenesis and atherosclerotic cardiovascular diseases. Male sex is one of the well-known cardiovascular risk factors. We tested the hypothesis that oxidative stress is greater in men than in women. Plasma thiobarbituric acid-reactive substances (TBARS) and urinary 8-isoprostaglandin F2alpha (8-iso-PGF2alpha) were measured in 52 young men and 51 age-matched women. The subjects were healthy, were not smokers, and were not taking any medications or vitamins. Age, blood pressure, plasma cholesterol, and glucose did not differ between the groups. Baseline TBARS (2.32 +/- 0.11 [men] versus 1.87 +/- 0.09 [women] nmol/mL, P<0.01) and 8-iso-PGF2alpha (292 +/- 56 [men] versus 164 +/- 25 [women] pg/mg creatinine, P<0.05) were higher in men than in women. Supplementation of antioxidant vitamins for 4 weeks in men produced a significant reduction in TBARS and 8-iso-PGF2alpha by 34% (P<0.01) and 48% (P<0.05), respectively. Plasma superoxide dismutase, catalase, and vitamin E levels were comparable between the groups. Enhanced oxidative stress in men may provide a biochemical link between male sex and atherosclerotic diseases related to oxidative stress.

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The repeated-dose liver micronucleus (RDLMN) assay using young adult rats has the potential to detect hepatocarcinogens. We conducted a collaborative study to assess the performance of this assay and to evaluate the possibility of integrating it into general toxicological studies. Twenty-four testing laboratories belonging to the Mammalian Mutagenicity Study Group, a subgroup of the Japanese Environmental Mutagen Society, participated in this trial. Twenty-two model chemicals, including some hepatocarcinogens, were tested in 14- and/or 28-day RDLMN assays. As a result, 14 out of the 16 hepatocarcinogens were positive, including 9 genotoxic hepatocarcinogens, which were reported negative in the bone marrow/peripheral blood micronucleus (MN) assay by a single treatment. These outcomes show the high sensitivity of the RDLMN assay to hepatocarcinogens. Regarding the specificity, 4 out of the 6 non-liver targeted genotoxic carcinogens gave negative responses. This shows the high organ specificity of the RDLMN assay. In addition to the RDLMN assay, we simultaneously conducted gastrointestinal tract MN assays using 6 of the above carcinogens as an optional trial of the collaborative study. The MN assay using the glandular stomach, which is the first contact site of the test chemical when administered by oral gavage, was able to detect chromosomal aberrations with 3 test chemicals including a stomach-targeted carcinogen. The treatment regime was the 14- and/or 28-day repeated-dose, and the regime is sufficiently promising to incorporate these methods into repeated-dose toxicological studies. The outcomes of our collaborative study indicated that the new techniques to detect chromosomal aberrations in vivo in several tissues worked successfully.

This article is available online at http://dmd.aspetjournals.org ABSTRACT:A metabolite formed by incubation of human liver microsomes, etoposide, and UDP-glucuronic acid was identified as etoposide glucuronide by liquid chromatography-tandem mass spectrometry analysis. According to the derivatization with trimethylsilylimidazole (Tri-Sil-Z), it was confirmed that the glucuronic acid is linked to an alcoholic hydroxyl group of etoposide and not to a phenolic group. Among nine recombinant human UGT isoforms (UGT1A1, UGT1A3, UGT1A4, UGT1A6, UGT1A8, UGT1A9. UGT1A10, UGT2B7, and UGT2B15), only UGT1A1 exhibited the catalytic activity of Etoposide [4Ј-demethylepipodophyllotoxin-9-(4,6-O-ethylidene)-␤-D-glucopyranoside] is one of the clinically important antitumor agents derived from 4Ј-demethylepipodophyllotoxin, which is an extract from the plants Podophyllum peltatum and Podophyllum emodi (Clark and Slevin, 1987;Stähelin and von Wartburg, 1991). It is widely used in the treatment of testicular cancer, small cell lung cancer, and certain lymphomas (O'Dwyer et al., 1985). Etoposide causes tumor cell killing through DNA strand breakage resulting from the interaction of etoposide with the enzyme topoisomerase II and DNA (Ross et al., 1984).In humans, the disposition of etoposide is described as a biphasic process with a distribution half-life of about 1.5 h and terminal elimination half-life ranging from 4 to 11 h (PDR, 2000). Clearance of etoposide occurs by direct renal excretion and metabolism. Roughly 35% of the administered drug is excreted into urine as a parent drug (Hande et al., 1984;Sinkule et al., 1984), but less than 3% is excreted into bile (Joel et al., 1996). Several metabolites were identified in human plasma and urine such as hydroxy acid derivatives, cis-(picro) lactone, 3Ј-demethyletoposide, and etoposide glucuronide (Clark and Slevin, 1987;Stewart, 1994). It has been reported that 58 and 19% of the administered drug is excreted as hydroxy acid into urine and bile, respectively (Clark and Slevin, 1987); less than 5 and 1% of the administered drug is found as cis-(picro) lactone in plasma and urine, respectively (Holthuis et al., 1986). It has been reported that 3Ј-demethylation of etoposide, a minor metabolite, is catalyzed by CYP3A4 (Relling et al., 1994). Etoposide glucuronide accounts for the disposition of 15 to 35% of administered etoposide (D'Incalci et al., 1986;Hande et al., 1988).Glucuronidation is catalyzed by UDP-glucuronosyltransferase (UGT 1 ) enzymes (Miners and Mackenzie, 1991). It is well known that there are many isoforms of mammalian UGT enzymes (Tukey and Strassburg, 2000). UGT1 and UGT2 have been shown to catalyze the glucuronidation of xenobiotics. The UGT1 and UGT2 genes seem to be structurally different in that UGT1 proteins result from alternate splicing of different first exons with five shared exons encoded by the UGT1 gene complex, whereas UGT2 proteins seem to be encoded by unique genes. In the human genome, at least 13 different first exons have been identified for the UGT1...

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ABSTRACT:A method for the direct determination of imipramine N-glucuronidation in human liver microsomes by high-performance liquid chromatography with UV detection was developed. Imipramine was incubated with human liver microsomes and UDP-glucuronic acid. The Eadie-Hofstee plots of imipramine N-glucuronidation in human liver microsomes were biphasic. For the high-affinity component, the

BackgroundResynthesis of triglycerides in enterocytes of the small intestine plays a critical role in the absorption of dietary fat. Acyl-CoA:monoacylglycerol acyltransferase-2 (MGAT2) is highly expressed in the small intestine and catalyzes the synthesis of diacylglycerol from monoacylglycerol and acyl-CoA. To determine the physiological importance of MGAT2 in metabolic disorders and lipid metabolism in the small intestine, we constructed and analyzed Mgat2-deficient mice.ResultsIn oral fat tolerance test (OFTT), Mgat2-deficient mice absorbed less fat into the circulation. When maintained on a high-fat diet (HFD), Mgat2-deficient mice were protected from HFD-induced obesity and insulin resistance. Heterozygote (Mgat2+/−) mice had an intermediate phenotype between Mgat2+/+ and Mgat2−/− and were partially protected from metabolic disorders. Despite of a decrease in fat absorption in the Mgat2-deficient mice, lipid levels in the feces and small intestine were comparable among the genotypes. Oxygen consumption was increased in the Mgat2-deficient mice when maintained on an HFD. A prominent upregulation of the genes involved in fatty acid oxidation was observed in the duodenum but not in the liver of the Mgat2-deficient mice.ConclusionThese results suggest that MGAT2 has a pivotal role in lipid metabolism in the small intestine, and the inhibition of MGAT2 activity may be a promising strategy for the treatment of obesity-related metabolic disorders.