Risk assessment of chemical carcinogens and thresholds.

The controversial arguments about the existence of "thresholds" for carcinogens are discussed and some conclusions are drawn: (1) The meaning of "threshold" has changed considerably during the last decades. Initially, the discussion focused on the genotoxic properties of chemicals. In dose-response studies the endpoint was tumor incidence. Later, DNA adducts represented the biologically active target dose and whether saturation of metabolic activation could lead to non-linear relationships was tested as a hypothesis. (2) In a next step, the implications of the initiation-promotion model were studied. Carcinogens with tumor-initiating properties showed linear dose-response relationships at low doses without a definable threshold, whereas those with tumor-promoting properties showed non-linear characteristics compatible with the existence of a threshold. However, the results are difficult to transfer to the human situation, and many critical endpoints are subject to other risk factors so that a meaningful value cannot be given. (3) Eventually, it turned out that most carcinogens exhibit genotoxic as well as non-genotoxic properties, and toxicity may be equally important as genotoxicity. (4) In view of the discussion for more than 60 years about the existence of thresholds for carcinogens, it is suggested that the threshold approach not be used to establish acceptable risk limits. (5) Instead of calculating an acceptable risk from cancer risk data, the recommended method is to assess the incremental contribution of exposure to the background of avoidable and unavoidable exposures by using biomonitoring data from human individuals. Such data could help in risk management, in order to reach acceptable limits of exposures on the basis of the "as low as reasonably achievable" or "ALARA" principle.

Early resistance to cell death and to onset of the mitochondrial permeability transition during hepatocarcinogenesis with 2-acetylaminofluorene.

A hallmark of tumorigenesis is resistance to apoptosis. To explore whether resistance to cell death precedes tumor formation, we have studied the short-term effects of the hepatocarcinogen 2-acetylaminofluorene (AAF) on liver mitochondria, on hepatocytes, and on the response to bacterial endotoxin lipopolysaccharide (LPS) in albino Wistar rats. We show that after as early as two weeks of AAF feeding liver mitochondria developed an increased resistance to opening of the permeability transition pore (PTP), an inner membrane channel that is involved in various forms of cell death. Consistent with a mitochondrial adaptive response in vivo, (i) AAF feeding increased the expression of BCL-2 in mitochondria, and (ii) hepatocytes isolated from AAF-fed rats became resistant to PTP-dependent depolarization, cytochrome c release, and cell death, which were instead observed in hepatocytes from rats fed a control diet. AAF-fed rats were fully protected from the hepatotoxic effects of the injection of 20-30 microg of LPS plus 700 mg of d-galactosamine (d-GalN) x kg-1 of body weight, a treatment that in control rats readily caused a large increase of terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling-positive cells in liver cryosections and release of alanine and aspartate aminotransferase into the bloodstream. Treatment with LPS and d-GalN triggered cleavage of BID, a BCL-2 family member, in the livers of both control- and AAF-fed animals, whereas caspase 3 was cleaved only in control-fed animals, indicating that the mitochondrial proapoptotic pathway had been selectively suppressed during AAF feeding. Phenotypic reversion was observed after stopping the carcinogenic diet. These results underscore a key role of mitochondria in apoptosis and demonstrate that regulation of the mitochondrial PTP is altered early during AAF carcinogenesis, which matches, and possibly causes, the increased resistance of hepatocytes to death stimuli in vivo. Both events precede tumor formation, suggesting that suppression of apoptosis may contribute to the selection of a resistant phenotype, eventually increasing the probability of cell progression to the transformed state.