Chloroform inhalation exposure conditions necessary to initiate liver toxicity in female B6C3F1 mice

Abstract

Chloroform is a nongenotoxic-cytotoxic carcinogen in rodent liver and kidney, including the female B6C3F1 mouse liver. Because tumors are secondary to events associated with cytolethality and regenerative cell proliferation, these end points are valid surrogates for tumor formation in cancer risk assessments. The purpose of the experiments presented here was to more clearly define the combinations of atmospheric concentration and duration of exposure necessary to induce cytolethality and regenerative cell proliferation in the sensitive female B6C3F1 mouse liver. Female B6C3F1 mice were exposed to chloroform by inhalation for 7 consecutive days using atmospheres of 10, 30, or 90 ppm and selected exposure times of 2, 6, 12, or 18 h/day. Bromodeoxyuridine (BrdU) was given the last 3.5 days via an implanted osmotic pump to label cells in S-phase. Labeled hepatocytes were visualized immunohistochemically, and the labeling index (LI) was determined as the percentage of cells in S-phase. LI was a more sensitive indicator of cellular damage than histopathological examination and is the more conservative end point for use in risk assessments. Significant concentration and exposure time related increases in LI were observed at 30 and 90 ppm but not at any 10-ppm exposure. These data defined an empirical relationship for the combinations of airborne exposure concentration and duration needed to induce cytolethality. These results suggest that concentrations of about 10 ppm or below will not induce hepatotoxicity in these mice regardless of exposure duration. Thus, the rate of production of toxic metabolites and the subsequent rate of cellular damage produced by a continual exposure of approximately 10 ppm chloroform are less than the maximum rates at which hepatocytes can detoxify those metabolites and repair any induced cellular damage. A physiologically based pharmacokinetic (PBPK) dosimetry model was used to compare anticipated responses in mice and humans and predicted that chloroform concentrations of approximately an order of magnitude greater than 10 ppm would be required to induce human liver toxicity. Thus, no safety factor to account for species to species extrapolation should be required in formulating a chloroform inhalation cancer risk assessment based on the dose × time inhalation data presented here.