Extent and Timeliness of Tissue Repair Determines the Dose-Related Hepatotoxicity of Chloroform

Abstract

As a part of mixture toxicity studies, the objective of the present investigation was to validate the hypothesis that the rate and extent of liver tissue repair response to a given dose determines the end result of toxicity (death or recovery), regardless of the mechanisms by which injury is inflicted, using a well-known environmental pollutant, chloroform (CHCl(3)). In future, the data will be used to compare with the results of mixtures containing CHCl(3) to aid in characterizing the safety of chemical mixtures and to construct a physiologically based pharmacokinetic (PBPK) model for dose, route, and species extrapolation. Hepatotoxicity and tissue repair were measured in male Sprague-Dawley rats (S-D) receiving a 10-fold dose range of CHCl(3) (74, 185, 370, and 740 mg/kg, IP) during a time course of 0 to 96 hours. Liver injury, as assessed by plasma alanine aminotransferase (ALT) and sorbitol dehydrogenase (SDH) elevation, increased with dose over the 10-fold dose range. Because CHCl(3) is also known to cause kidney damage, blood urea nitrogen (BUN) and creatinine were measured to evaluate the kidney injury. With doses up to 370 mg/kg, liver injury increased in a dose-related fashion, which peaked at 24 hours and returned to normal after 48 hours, whereas at highest dose (740 mg/kg), the injury was progressive resulting in 90% mortality. Blood and liver CHCl(3) levels were quantified using gas chromatography (GC) over a time course of 30 to 360 minutes. The dose-related increase in the blood and liver CHCl(3) levels were consistent with dose-dependent liver injury. Tissue regeneration response, as measured by [(3)H]-thymidine incorporation into hepatocellular nuclear DNA peaked at 36 hours in rats treated with the lower two doses of CHCl(3) (74 and 185 mg/kg). Further increase in CHCl(3) dose to 370 mg/kg resulted in an earlier increase in [(3)H]-thymidine incorporation at 24 hours, which peaked at 36 hours. However, at the highest dose of CHCl(3) (740 mg/kg), tissue repair was delayed and attenuated, allowing for unrestrained progression of liver injury. The kidney injury markers after CHCl(3) administration were not different from controls. These results support the concept that in addition to the magnitude of tissue repair response, the time at which this response occurs is critical in restraining the progression of injury. Measuring tissue repair and injury as simultaneous biological responses to toxic agents might increase the usefulness of dose-response paradigms in predictive toxicology and risk assessment. Although the dosimetry of the present study was well beyond the environmental exposure levels of CHCl(3), a PBPK model will be developed in future based upon these data to evaluate the effects at environmental levels.

Application of Key Events Analysis to Chemical Carcinogens and Noncarcinogens

“…Similarly, the rate of systemic removal can influence systemic levels. There is evidence that the elimination of chloroform at higher doses is saturable, so that systemic, and hence target tissue, exposure increases disproportionately at high doses (Anand et al, 2003; 2006). This is particularly notable after a single exposure.…”

“…It is very unlikely that accumulation in fat per se is saturable, even at doses used experimentally. Repeated exposure results in increased elimination of chloroform, at least in mice, by mechanisms that have yet to be determined (Anand et al, 2006). …”

“…Cmax for chloroform also showed nonlinear increases with dose, 7-fold in blood, 8-fold in liver, and 9-fold in kidney. Following repeated exposure for 30 days, the target organ AUC increased only 3–4-fold over this dose range, while Cmax increased less than 2-fold (Anand et al, 2006). As a consequence of these changes, while the target organ Cmax showed very little variation with repeated exposure at a dose of 150 mg/kg bw, at high doses of 300 mg/kg the target organ Cmax varied 8-fold in liver and 18-fold in kidney between one and 30 days exposure.…”

“…However, repeated exposure at low doses appears to engage physiological response mechanisms that lead to increased elimination, reduction of AUC and Cmax, and decreased target tissue dose—i.e., changes that would be expected to affect the magnitude of the biological response, and there is evidence that this is so with tolerance to the target organ toxicity of chloroform developing on repeated exposures (Anand et al, 2006). The effects may be of such a magnitude as to reduce target organ exposure to below that level necessary to provoke any biological response at all.…”

<b>3-methyadenine attenuates chloroform-induced hepatotoxicity via autophagy </b><b>activation </b>

“…Exposure of human to chloroform leads to severe hepatotoxicity (13,21). Chloroform has been classified as one of the hazardous substances in the list of top 20 priority substances by Agency for Toxic Substances and Disease Registry (ATSDR 1995) (2). However, even nowadays, chloroform-induced acute liver injuries are also frequently reported by clinicians (4,13,15).…”