Mutagenicity testing of dichloromethane in short-term mammalian test systems

Jongen, W.M.F.; Lohman, P.H.M.; Kottenhagen, M.J.; Alink, G.M.; Berends, Frits J.; Koeman, J.H.

doi:10.1016/0027-5107(81)90035-x

Cited by 40 publications

(5 citation statements)

References 16 publications

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“…Both types of damage were induced in DNA from CHO cells incubated with formaldehyde, and with MC metabolized exogenously by mouse liver S9. There was no DNA damage in CHO cells incubated widi MC alone, suggesting a lack of metabolic activation and providing an explanation for the absence of MC-induced mutations in diese cells (5,6). In contrast 1,2-DBE, which is metabolized to a DNA-binding GSH conjugate (38), is mutagenic (54) and causes DNA ss breaks (Table I) Addition of semicarbazide to the incubation medium inhibited formaldehyde-induced DNA ss breaks and DNA-protein crosslinks, but inhibited only the DNA-protein cross-links induced by MC.…”

Section: Discussionmentioning

confidence: 90%

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Relationship between hepatic DNA damage and methylene chloride-induced hepatocarcinogenicity in B6C3F1 mice

et al. 1994

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Methylene chloride (MC) induced DNA damage in freshly isolated hepatocytes from mice and rats, which was detectable as single-strand (ss) breaks by alkaline elution. The lowest in vitro concentration of MC needed to induce DNA damage in mouse hepatocytes (0.4 mM) was much lower than for rat hepatocytes (30 mM), and is close to the calculated steady-state concentration of MC in the mouse liver (1.6 mM) at a carcinogenic dose (4000 p.p.m. by inhalation). DNA ss breaks were also detectable in hepatocyte DNA from mice which had inhaled 4000 p.p.m. MC for 6 h, but not in hepatocyte DNA from rats similarly exposed. In studies with hepatocytes cultured overnight in the presence of buthionine sulfoximine to deplete glutathione (GSH), subsequent exposure to MC resulted in less DNA damage in the GSH-depleted cells. This shows that conjugation of MC with GSH is important in its activation of DNA-damaging species in the liver. The GSH pathway of MC metabolism produces two potential DNA-damaging species, formaldehyde and S-chloromethylglutathione (GSCH2Cl). Formaldehyde is known to cause DNA ss breaks in cells. However, the lowest concentration of formaldehyde required to induce a significant amount of DNA ss breaks in mouse hepatocytes (0.25 mM) is unlikely to be formed following in vitro or in vivo metabolism of MC at concentrations that induce similar amounts of DNA damage. That formaldehyde does not play a role in this DNA damage has been confirmed in experiments with CHO cells exposed to MC and an exogenous activation system from mouse liver (S9 fraction). Formaldehyde was responsible for the DNA- protein cross-linking effect of MC, but did not cause the DNA damage leading to ss breaks. These DNA ss breaks are likely to be caused by GSCH2Cl. The results suggest a genotoxic mechanism for MC carcinogenicity in the mouse liver, and support the proposal that the observed species differences in liver carcinogenicity result from differences in the amount of MC metabolism via the GSH pathway in the target organ.

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Section: Discussionmentioning

confidence: 90%

“…The reason for this carcinogenic effect is unknown. In in vitro assays MC is positive as a mutagen in bacteria (2,3) and yeast (4), but is not a mutagen in mammalian cells (5,6). The possible lack of metabolic activation of MC by the cells used in the latter assays has not been addressed.…”

Section: Introductionmentioning

confidence: 99%

Relationship between hepatic DNA damage and methylene chloride-induced hepatocarcinogenicity in B6C3F1 mice

et al. 1994

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“…The mutagenicity of this material to Salmonella is well established, but this is not reflected in mammalian cells. For example, several negative responses have been reported for DCM in mammalian cell gene mutation assays and in assays for the induction of SCE and UDS [Thilagar and Kumaroo, 1983;Jongen et al, 1981;Thilagar et al, 1984a,b;Andrae and Wolff, 19831. This leaves the positive in vitro clastogenic effects as apparently the most compelling mammalian cell mutagenicity data [Thilagar et al, 1984al.…”

Section: Discussionmentioning

confidence: 99%

Lack of uds activity in the livers of mice and rats exposed to dichloromethane

Trueman¹,

Ashby²

1987

Environ and Mol Mutagen

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Dichloromethane (DCM) has been evaluated for its ability to initiate unscheduled DNA synthesis (UDS) in the livers of male mice and rats in vivo. Two types of experiment were conducted. In the first, Alpk:AP rats were exposed by oral gavage to 100, 500, or 1,000 mg/kg DCM and hepatocytes assessed for UDS via autoradiography 4 and 12 hours later. In the second, Fischer F344 rats or B6C3F1 mice were exposed by inhalation to either 2,000 or 4,000 ppm of DCM for either 2 or 6 hours, and hepatocytes assessed for UDS immediately after exposure. The dose levels and strains of rodent employed in the latter protocol correspond to those employed in a recent cancer bioassay of DCM conducted by the U.S. National Toxicology Program. DCM failed to induce UDS in any of the experiments. These data are discussed within the context of other evidence indicating DCM to be nongenotoxic in vivo, despite its reported carcinogenicity in the mouse.

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“…It produces tumors at other sites by inhalation and oral administration (IARC, 1987). DCM was mutagenic in bacteria only when tested as a vapour (IARC, 1986), but was not mutagenic (Jongen et al, 1981), or produced an equivocal response, in mammalian cells in culture (Myhr et al, 1990). Conflicting results have been reported regarding its clastogenicity in vitro (Jongen et al, 1981;Thilagar and Kumaroo, 1983;Anderson et al, 1990;Ishidate, 1988).…”

Section: Introductionmentioning

confidence: 99%

The role of glutathione in the bacterial mutagenicity of vapour phase dichloromethane

Dillon

Edwards

Combes

et al. 1992

Environ and Mol Mutagen

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Dichloromethane (DCM) vapour by inhalation is carcinogenic to rodents and is an in vivo rodent cell clastogen and a bacterial mutagen. It has been suggested that the bacterial mutagenicity of DCM is mediated by glutathione (GSH) conjugation. The involvement of endogenous and exogenous GSH in the conversion of DCM to a bacterial mutagen has been studied in a vapour phase protocol using wild-type and GSH-deficient (NG54; gsh) Salmonella typhimurium TA100 strains in the presence and absence of various rat liver fractions. The effect of the duration of exposure was also investigated in these Salmonella strains and in E. coli WP2 uvrA pKM101. Dose- and time-related increases in revertants occurred with all metabolic activation systems used (without exogenous metabolic activation; with Aroclor-induced rat liver S9, microsomes, or cytosol fractions), with minor quantitative differences among the 3 strains. Mutagenicity was marginally highest in the presence of cytosol at the highest DCM concentrations. Strain NG54 gsh, which contains approximately 25% of the TA100 level of GSH/microgram protein, was slightly less responsive to DCM-induced mutagenicity than TA100. Addition of 0.33 mumoles/plate of GSH had little effect on the mutagenic responses of TA100 or NG54 in the presence or absence of S9. In these 2 strains, exogenous S9 produced small increases in mutagenicity at the highest concentrations of DCM (2 and 4% v/v). These results suggest that if an interaction between DCM and GSH is required for the activation of DCM to a bacterial mutagen, it occurs at low levels of endogenous GSH and is not significantly affected by GSH supplementation.

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Mutagenicity testing of dichloromethane in short-term mammalian test systems

Cited by 40 publications

References 16 publications

Relationship between hepatic DNA damage and methylene chloride-induced hepatocarcinogenicity in B6C3F1 mice

Relationship between hepatic DNA damage and methylene chloride-induced hepatocarcinogenicity in B6C3F1 mice

Lack of uds activity in the livers of mice and rats exposed to dichloromethane

The role of glutathione in the bacterial mutagenicity of vapour phase dichloromethane

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