Lack of genotoxic activity of di(2-ethylhexyl)phthalate (DEHP) in rat and human hepatocytes

Abstract

Di(2-ethylhexyl)phthalate (DEHP) is a widely used plasticizer which has been reported to induce a statistically significant increase in the incidence of hepatocellular carcinomas in female Fischer-344 rats (8/50) when administered in the diet at 12 000 p.p.m. for two years. Numerous studies with cells in culture have failed to show any genotoxic activity associated with DEHP. Because DEHP induces multiple changes in the liver, such as peroxisomal proliferation, it was possible that these alterations could result in genotoxic effects in the treated whole animal that would not be seen in cells in culture. Accordingly, the ability of DEHP to induce DNA damage or repair was examined in rat hepatocytes in vivo and in vitro and in human hepatocytes in vitro. Unscheduled DNA synthesis was measured by incorporation of [3H]thymidine into primary hepatocyte cultures immediately isolated from treated animals or hepatocyte cultures incubated directly with DEHP. DNA damage was measured by alkaline elution of cellular DNA from the same cultures. In vivo-in vitro treatment regimens were: (i) female rats, 12 000 p.p.m. DEHP in the diet for 30 days; (ii) female rats, 12 000 p.p.m. in the diet for 30 days, followed by 500 mg/kg DEHP by gavage 2 h before sacrifice; (iii) male rats, 500 mg/kg DEHP by gavage 2, 12, 24, or 48 h before sacrifice; and (iv) male rats, 150 mg/kg/day by gavage for 14 days. In vitro conditions were 0.1, 1.0 and 10.0 mM DEHP in the cultures for 18 h. Primary cultures of human hepatocytes were prepared from freshly discarded surgical material and exposed to the same concentration of DEHP. Concentrations up to 0.5 mM mono(2-ethylhexyl)phthalate, a principal metabolite of DEHP, were also examined in the human hepatocyte assay. No chemically induced DNA damage or repair was observed in vivo or in vitro in rat or human hepatocytes under any of the conditions employed. However, an increase in the percentage of cells in S-phase in the animals given DEHP was observed. These data indicate that DEHP does not exhibit direct genotoxic activity in the animals even with a treatment regimen which eventually produced tumors in a long term bioassay, and that both rat and human hepatocytes are similar in their lack of a genotoxic response to DEHP exposure in culture.

Mitogenic activity of quinoline to the rat, mouse, and guinea pig liver: Empirical correlations with hepatic carcinogenicity

“…The overall insensitivity of guinea pigs to chemical mitogenesis and hepatocarcinogenesis may be related to the recent claim that they are not in fact rodents [Graur et al, 19911. Although the stimulation of rodent hepatocytes into DNA synthesis has been causally linked with nongenotoxic carcinogenesis for a wide range of compounds [ Butterworth et al, 1984;Goldsworthy et al, 1991;Marsman et al, 1988;Mirsalis, 1987;Mirsalis et al, 19851, the mechanistic basis "Data on the higher dose levels of quinoline in the rat are abstracted from Ashby et al (1989).…”

DEHP: Genotoxicity and potential carcinogenic mechanisms—A review

“…Its solubility in water is low and is even lower in salt water. A number of authors have given estimates or assumptions of 3900 mg/ml or higher as the concentration in which precipitation occurs [28,91,129,141], while others report precipitation at a lower range of 390-2000 mg/ml [25,149,156,161]. These studies have usually been conducted with the addition of DMSO to aid in solubility.…”

Measurement of ploidy and cell proliferation in the rodent liver.

“…(31). The compounds in the latter group are non-DNA reactive (32)(33)(34)(35)(36)(37)(38)(39)(40), and they elicit liver growth and cancer that is rodent specific (31). The liver growth phenomena that are induced are hepatocyte hypertrophy (smooth endoplasmic reticulum proliferation and usually peroxisome proliferation) and hyperplasia (41)(42)(43)(44)(45)(46).…”