Comparison of three rat liver foci bioassays-incidence of preneoplastic foci initiated by diethylnitrosamine

Abstract

Three rat liver foci bioassays have been compared with respect to their sensitivity by the histochemical demonstration of preneoplastic foci, and by the biochemical determination of alterations in enzyme activities of serum indicating hepatotoxicity. We studied the initiation/promotion schedules according to Oesterle and Deml (A), and according to Pereira (B, Broad Spectrum Protocol), and the initiation/selection protocol according to Tatematsu et al. (C), with diethylnitrosamine (DEN), given as a single initiating dose of 10 and 30 mg/kg body wt respectively. With all schedules Sprague-Dawley rats, either females, 3 weeks old (A), or males, 6 weeks old (B, C) were used. For promotion polychlorinated biphenyls (A) or phenobarbital (B) were administered. Selection was performed with 2-acetylaminofluorene (C). The rats in schemes (B) and (C) underwent partial hepatectomy one day prior to initiation. The number and total area of foci deficient in adenosine-5'-triphosphatase (ATPase) and positive in gamma-glutamyltranspeptidase (GGTase) was evaluated. In the complete schedule with 30 mg of DEN in system (A) foci incidence exceeded that of the other systems by about 7-fold (ATPase) and 2-fold (GGTase) respectively. The lower dose of DEN and all control experiments resulted in a respective lower foci yield. With scheme (C), but not with schemes (A) and (B), e.g. serum fructose-1.6-bisphosphatase and alkaline phosphatase were increased, suggesting liver cell damage. Thus tested with DEN, scheme (A) is most sensitive and causes a low impairment of animals' welfare.

Significance of Hepatic Preneoplasia in Risk Identification and Early Detection of Neoplasia *

“…With respect to peroxisome proliferators the application of the protocol proposed by Oesterle et al (26) had the same false-negative result (16) as the medium-term carcinogenesis bioassay used by Ito et al (18). The observation that some peroxisome proliferators including dehydroepiandrosterone (5), and particularly T 3 (23), inhibit hepatocarcinogenesis under certain experimental conditions implies, however, that a reduction in the area fraction of FAH by such compounds (19,22) actually indicates a preventive potential, the possible application of which to patients remains to be clarified under well-controlled clinical conditions.…”

“…Many research groups have used these lesions in mechanistic studies on hepatocarcinogenesis, and some groups have developed mediumterm carcinogenesis bioassays based on the predictive value of FAH (3,18,26,27,34,36,37). It is evident, however, that there is a confusing variety of experimental approaches (Figure 1), which make a complete consensus on the interpretation of the results difficult (5,7).…”

Rapid Induction of Preneoplastic Liver Foci in Embryonal Turkey Liver by Diethylnitrosamine

“…Various types of FAH have been described and are regarded as preneoplastic lesions (3). A number of experiments indicated a predominant sequential development from glycogenotic (clear and acidophilic) cell foci through mixed and basophilic cell foci to benign and malignant neoplasms (1,8,27,28,(42)(43)(44). FAH may develop to hepatocellular tumors without any further treatment and, therefore, are a favorite subject in investigations on hepatocarcinogenesis and for carcinogenesis bioassay (5,17,18,32).…”

Prevalidation of a Rat Liver Foci Bioassay (RLFB) Based on Results from 1600 Rats: A Study Report

“…in juvenile rats as test animals, has been developed in one of the participating laboratories and employed in several studies prior to the present prevalidation study (16,41,42). During the initial phases of the RLFB prevalidation project, Standard Operating Procedures (SOPs) were developed, which describe in detail all essential aspects of the animal study, including the morphometric evaluation of FAH and the statistical analysis of the data.…”

“…FAH have been accepted as early indicators of carcinogenic response by a number of international and national working groups (22,37,57,58,63). Several medium-term carcinogenesis assays using FAH as an endpoint have been proposed (32,37,42,62,66), and have been suggested to become part of a general testing strategy taking its place between in vitro mutagenicity tests and the chronic rodent bioassay (CRB) (19,66). The limitation of these in vivo test systems to the liver is an obvious disadvantage for the identification of cancer risk factors in general, but the liver is the prevailing target organ of chemical carcinogens in rodents (1), being affected by ∼30% of all chemicals positively tested in long-term carcinogenesis bioassays in the National Toxicology Program (29).…”