Extent and timeliness of tissue repair determines the dose-related hepatotoxicity of chloroform

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 (CHCl3). In future, the data will be used to compare with the results of mixtures containing CHCl3 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 CHCl3 (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 CHCl3 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 CHCl3 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 CHCl3 levels were consistent with dose-dependent liver injury. Tissue regeneration response, as measured by [H-3]-thymidine incorporation into hepatocellular nuclear DNA peaked at 36 hours in rats treated with the lower two doses of CHCl3 (74 and 185 mg/kg). Further increase in CHCl3 dose to 370 mg/kg resulted in an earlier increase in [H-3]-thymidine incorporation at 24 hours, which peaked at 36 hours. However, at the highest dose of CHCl3 (740 mg/kg), tissue repair was delayed and attenuated, allowing for unrestrained progression of liver injury. The kidney injury markers after CHCl3 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 CHCl3, a PBPK model will be developed in future based upon these data to evaluate the effects at environmental levels.