Akiko H. Hashimoto

Enhanced Spontaneous and Benzo(a)pyrene-Induced Mutations in the Lung of Nrf2-Deficient gpt Delta Mice

The lung is an organ that is sensitive to mutations induced by chemicals in ambient air, and transgenic mice harboring guanine phosphoribosyltransferase (gpt) gene as a target gene are a well-established model system for assessing genotoxicity in vivo. Transcription factor Nrf2 mediates inducible and constitutive expression of cytoprotective enzymes against xenobiotics and mutagens. To address whether Nrf2 is also involved in DNA protection, we generated nrf2+/-::gpt and nrf2-/-::gpt mice. The spontaneous mutation frequency of the gpt gene in the lung was approximately three times higher in nrf2-null (nrf2-/-) mice than nrf2 heterozygous (nrf2+/-) and wild-type (nrf2+/+) mice, whereas in the liver, the mutation frequency was higher in nrf2-/- and nrf2+/- mice than in nrf2+/+ wild-type mice. By contrast, no difference in mutation frequency was observed in testis among the three genotypes. A single intratracheal instillation of benzo(a)pyrene (BaP) increased the lung mutation frequency 3.1- and 6.1-fold in nrf2+/- and nrf2-/- mice, respectively, compared with BaP-untreated nrf2+/- mice, showing that nrf2-/- mice are more susceptible to genotoxic carcinogens. Surprisingly, mutation profiles of the gpt gene in BaP-treated nrf2+/- mice was substantially different from that in BaP-untreated nrf2-/- mice. In nrf2-/- mice, spontaneous and BaP-induced mutation hotspots were observed at nucleotides 64 and 140 of gpt, respectively. These results thus show that Nrf2 aids in the prevention of mutations in vivo and suggest that Nrf2 protects genomic DNA against certain types of mutations.

Degradation pathways of trichloroethylene and 1,1,1-trichloroethane by Mycobacterium sp. TA27

We analyzed the kinetics and metabolic pathways of trichloroethylene and 1,1,1-trichloroethane degradation by the ethane-utilizing Mycobacterium sp. TA27. The apparent V max and K m of trichloroethylene were 9.8 nmol min−1 mg of cells−1 and 61.9 μM, respectively. The apparent V max and K m of 1,1,1-trichloroethane were 0.11 nmol min−1 mg of cells−1 and 3.1 μM, respectively. 2,2,2-trichloroethanol, trichloroacetic acid, chloral, and dichloroacetic acid were detected as metabolites of trichloroethylene. 2,2,2-trichloroethanol, trichloroacetic acid, and dichloroacetic acid were also detected as metabolites of 1,1,1-trichloroethane. The amounts of 2,2,2-trichloroethanol, trichloroacetic acid, chloral, and dichloroacetic acid derived from the degradation of 3.60 μmol trichloroethylene were 0.16 μmol (4.4%), 0.11 μmol (3.1%), 0.02 μmol (0.6%), and 0.02 μmol (0.6%), respectively. The amounts of 2,2,2-trichloroethanol, trichloroacetic acid and dichloroacetic acid derived from the degradation of 1.73 μmol 1,1,1-trichloroethane were 1.48 μmol (85.5%), 0.22 μmol (12.7%), and 0.02 μmol (1.2%), respectively. More than 90% of theoretical total chloride was released in trichloroethylene degradation. Chloral and 2,2,2-trichloroethanol were transformed into each other, and were finally converted to trichloroacetic acid, and dichloroacetic acid. Trichloroacetic acid and dichloroacetic acid were not degraded by strain TA27.

Degradation of trichloroethylene and related compounds by Mycobacterium spp. isolated from soil

Mycobacterium spp. strains TA5 and TA27 (ethane-utilizing bacteria), which can degrade trichloroethylene (TCE) and 1,1,1-trichloroethane (1,1,1-TCA), were isolated from soil. Both bacteria could cometabolically degrade dichloromethane, chloroform, 1,1-dichloroethane, 1,2-dichloroethane, 1,1,1-TCA, 1,1,2-TCA, 1,1,1,2-tetrachloroethane, 1,1,2,2-tetrachloroethane, 1,1-dichloroethylene, cis-1,2-dichloroethylene, trans-1,2-dichloroethylene, and TCE with ethane as a carbon source. They could not degrade carbon tetrachloride, freon 113, or tetrachloroethylene. The TCE degradation characteristics of strain TA27 were determined. Under a head-space gas containing 3% ethane, strain TA27 degraded more than 95% of TCE at an initial concentration of 1 mg l–1 within 3 days. We observed good growth and TCE degradation between 25 and 35 °C. At an initial TCE concentration of 30 mg l–1, it degraded 30% of TCE within 7 days. Although growth was inhibited for more than 50 mg l–1 TCE at 3% ethane concentration, good growth and 50% degradation of TCE were observed at 12% ethane concentration within 14 days. High ethane concentration may mitigate the toxicity of TCE.