Thresholds for the effects of 2-acetylaminofluorene in rat liver

To explore for practical thresholds for DNA-reactive carcinogens in rat liver carcinogenicity, we have conducted a series of exposure-response studies using 2 well-studied hepatocarcinogens, 2-acetylaminofluorene (AAF) and diethylnitrosamine (DEN). Findings with AAF, including as yet unpublished experiments, are reviewed here and related to DEN observations. In these studies, we have administered exact intragastric doses during an initiation segment (IS) of 12-16 weeks followed in some experiments by phenobarbital (PB) as a liver tumor promoter for 24 weeks to enhance manifestation of initiation. The cumulative doses (CD) of AAF at the end of ISs ranged from 0.094 to 282.2 mg/kg. Our findings for AAF in the IS can be summarized as follows: (1) the earliest parameter to be affected with administration of low doses was the appearance of DNA adducts (around 4 weeks), followed at higher doses by cell proliferation; (2) formation of DNA adducts was nonlinear, with a no-observed effect level (NOEL) at a CD of 0.094 mg/kg and a plateau at higher doses (94.1 mg/kg); (3) cytotoxicity (necrosis) showed a NOEL at a CD of 28.2 mg/kg; (4) compensatory hepatocellular proliferation showed a NOEL at a CD of 28.2 mg/kg and was supralinear at a high CD (282.2 mg/kg); (5) formation of preneoplastic hepatocellular altered foci (HAF) showed a NOEL at a CD of 28.2 mg/kg, and was supralinear at a high CD (282.2 mg/kg); (6) a NOEL (CD 28.2 mg/kg) was found for tumor development and the exposure-response was supralinear. We interpret these findings to reflect practical thresholds for hepatocellular initiating effects of AAF and exaggerated responses at high-exposures doses, as also found for DEN. Thus, mechanisms of carcinogenesis can differ between low and high doses.     

Nasal cytotoxic and carcinogenic activities of systemically distributed organic chemicals

Toxicity and carcinogenicity in the mucosa of the nasal passages in rodents has been produced by a variety of organic chemicals which are systemically distributed. In this review, 14 such chemicals or classes were identified that produced rodent nasal cytotoxicity, but not carcinogenicity, and 11 were identified that produced nasal carcinogenicity. Most chemicals that affect the nasal mucosa were either concentrated in that tissue or readily activated there, or both. All chemicals with effects in the nasal mucosa that were DNA-reactive, were also carcinogenic, if adequately tested. None of the rodent nasal cytotoxins has been identified as a human systemic nasal toxin. This may reflect the lesser biotransformation activity of human nasal mucosa compared to rodent and the much lower levels of human exposures. None of the rodent carcinogens lacking DNA reactivity has been identified as a nasal carcinogen or other cancer hazard to humans. Some DNA-reactive rodent carcinogens that affect the nasal mucosa, as well as other tissues, have been associated with cancer at various sites in humans, but not the nasal cavity. Thus, findings in only the rodent nasal mucosa do not necessarily predict either a toxic or carcinogenic hazard to that tissue in humans.     

Quantitative comparison between in vivo DNA adduct formation from exposure to selected DNA-reactive carcinogens, natural background levels of DNA adduct formation and tumour incidence in rodent bioassays

This study aimed at quantitatively comparing the occurrence/formation of DNA adducts with the carcinogenicity induced by a selection of DNA-reactive genotoxic carcinogens. Contrary to previous efforts, we used a very uniform set of data, limited to in vivo rat liver studies in order to investigate whether a correlation can be obtained, using a benchmark dose (BMD) approach. Dose-response data on both carcinogenicity and in vivo DNA adduct formation were available for six compounds, i.e. 2-acetylaminofluorene, aflatoxin B1, methyleugenol, safrole, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline and tamoxifen. BMD(10) values for liver carcinogenicity were calculated using the US Environmental Protection Agency BMD software. DNA adduct levels at this dose were extrapolated assuming linearity of the DNA adduct dose response. In addition, the levels of DNA adducts at the BMD(10) were compared to available data on endogenous background DNA damage in the target organ. Although for an individual carcinogen the tumour response increases when adduct levels increase, our results demonstrate that when comparing different carcinogens, no quantitative correlation exists between the level of DNA adduct formation and carcinogenicity. These data confirm that the quantity of DNA adducts formed by a DNA-reactive compound is not a carcinogenicity predictor but that other factors such as type of adduct and mutagenic potential may be equally relevant. Moreover, comparison to background DNA damage supports the notion that the mere occurrence of DNA adducts above or below the level of endogenous DNA damage is neither correlated to development of cancer. These data strongly emphasise the need to apply the mode of action framework to understand the contribution of other biological effect markers playing a role in carcinogenicity.     

The toxicology of aflatoxins as a basis for public health decisions

Aflatoxins have been extensively studied with respect to their mechanisms of toxicity. An understanding of metabolism, DNA adduct induction, mutagenicity and carcinogenicity has been paralleled by the development of biomarkers of aflatoxin exposure and biological effects (e.g. mutations) applied to human populations. The improvements in exposure assessment and their application in prospective epidemiological studies and the demonstration of a specific mutation in the TP53 gene in hepatocellular carcinomas from areas of high aflatoxin exposure have contributed significantly to the classification of aflatoxins as human carcinogens. In addition to establishing the carcinogenicity of aflatoxins in humans, understanding molecular mechanisms of action has provided the scientific rationale for prevention strategies, including primary and chemoprevention approaches. Overall, integrated, multidisciplinary research on aflatoxins has provided the platform on which to base decisions regarding acceptable exposures and priorities for interventions to reduce human risk in a public health context.     

Furan-induced dose–response relationships for liver cytotoxicity,cell proliferation,and tumorigenicity (furan-induced liver tumorigenicity)

Rodent studies of furan are associated with liver cell necrosis, release of liver-associated enzymes, increased hepatocyte proliferation, and hepatocarcinogenesis. For carcinogens whose proposed mode of action is cytolethality, it is hypothesized that the dose–response curve for tumor development would parallel the dose–response curve for cell death with compensatory proliferation in the target organ. To prospectively test this hypothesis, female B6C3F₁ mice were exposed to furan at carcinogenic doses and lower for 3 weeks or 2 years. At 3 weeks and in the 2-year study, there were dose-dependent and significant increases in hepatic cytotoxicity at 1.0, 2.0, 4.0, and 8.0 mg furan/kg. For cell proliferation as measured by 5-bromo-2′-deoxyuridine (BrdU) labeling index (LI), there was a statistically significant trend with increasing dose levels of furan and increased LI at 8.0 mg/kg. There was an increased incidence of foci of altered hepatocytes, hepatocellular adenomas, and adenomas or carcinomas at 4.0 and 8.0 mg/kg and carcinomas at 8.0 mg/kg. The multiplicity of microscopic tumors was increased and latency was decreased in mice exposed to 8.0 mg/kg. Prevalence of hepatic nodules at necropsy was increased in mice exposed to 4.0 and 8.0 mg/kg. Data demonstrate an association among furan-induced hepatic cytotoxicity, compensatory cell replication, and liver tumor formation in mice; at high doses ?4.0 mg/kg, furan induced hepatotoxicity, compensatory cell replication and tumorigenesis in a dose-related manner, while furan did not produce tumors at cytotoxic doses of 1.0 and 2.0 mg/kg.     

Nongenotoxic (epigenetic) carcinogens: pesticides as an example. A critical review

The following groups of pesticides are considered in this review by supposed mechanisms of their carcinogenicity: hepatocarcinogenic pesticides, pesticides - peroxisome proliferators, pesticides as endocrine disruptors, goitrogenic pesticides, pesticides producing sustained cell proliferation and some others. With very rare exceptions, pesticides do not react with DNA directly and the mechanisms of their carcinogenicity are, in general, similar to those of other nongenotoxic (epigenetic) carcinogens, namely: promotion of spontaneous initiation, cytotoxicity with sustained cell proliferation, oxidative stress, formation of activated receptors and some others. Genotoxicity of pesticides varies from its complete absence (propiconazol as an example) to a very pronounced one (captafol) with remaining compounds in between. These two compounds demonstrate full correlation between genotoxicity and carcinogenicity (or their absence). Many pesticides give positive results in some tests for genotoxicity but these results are frequently controversial, not readily reproducible, or obtained only at toxic dose levels. The weak genotoxicity of the majority of pesticides is easily explainable by their rather severe testing before their introduction into practical use. The above mechanisms are threshold-based and therefore pesticides are regulated through NOEL/safety factor. There exist examples of lack of correlation between genotoxicity and carcinogenicity: some pesticides are genotoxic (although not strongly) but noncarcinogenic, others are considered as nongenotoxic but are strongly carcinogenic (chlorothalonil, acetochlor). The general scheme of the promoters' effect is presented in which an important role is attributed to the cytochrome P-450 induction (some pesticides are the cytochrome P-450 inducers), formation of reactive oxygen species and peroxitome proliferation. Teratogenesis Carcinog. Mutagen. 20:229-240, 2000.     

Non-reversibility of vinyl chloride carcinogenesis in rodents

A review of the data obtained from various studies on carcinogenicity of vinyl chloride (VC) in rodents, particularly on the effect of dose, age, duration of exposure and potential reversibility of lesions, revealed that vinyl chloride-induced carcinogenicity in rodents was dose and time related; no recovery occurred in mice even after only 1 month of VC exposures or in rats after 6-month exposures. In addition, younger animals (2 months old) were more susceptible to VC-induced carcinogenicity than animals held for 6 or 12 months prior to exposure. Initial 6 or 12 month exposures were adequate to detect the carcinogenic potential of VC. The above information was used as a basis for discussion on design of carcinogenicity studies. Possibility of determining the carcinogenic potential of a compound in a shorter period than the traditional 2 year studies in rodents was discussed in consideration with appropriate doses, species, age and exposure duration. Although this approach may be applicable to a strong carcinogen, it was not considered practicable in case of weak or unknown carcinogens.     

Prediction of rodent carcinogenesis: an evaluation of prechronic liver lesions as forecasters of liver tumors in NTP carcinogenicity studies

The National Toxicology Program (NTP) developed the chronic 2-year bioassay as a mechanism for predicting the carcinogenic potential of chemicals in humans. The cost and duration of these studies has limited their use to small numbers of selected chemicals. Many different short-term methods aimed at increasing predictive accuracy and the number of chemicals evaluated have been developed in attempts to successfully correlate their results with evidence of carcinogenicity (or lack of carcinogenicity) are assessed. Using NTP studies, the effectiveness of correlating prechronic liver lesions with liver cancer encompassing multiple studies using mice (83 compounds) and rats (87 compounds). These lesions include hepatocellular necrosis, hepatocellular hypertrophy, hepatocellular cytomegaly, bile duct hyperplasia, and hepatocellular degeneration, along with increased liver weight. Our results indicate that pooling 3 of these prechronic data points (hepatocellular necrosis, hepatocellular hypertrophy, and hepatocellular cytomegaly) can be very predictive of carcinogenicity in the 2-year study (p < 0.05). The inclusion of increased liver weight as an endpoint in the pool of data points increases the number of rodent liver carcinogens that are successfully predicted (p < 0.05), but also results in the prediction of increased numbers of noncarcinogenic chemicals as carcinogens. The use of multiple prechronic study endpoints provides supplementary information that enhances the predictivity of identifying chemicals with carcinogenic potential.     

The mutagenic and carcinogenic activity of chemical compounds

The article presents data on the association between the mutagenicity and carcinogenicity of chemical compounds. Genotoxic carcinogens, which universally act via interaction with DNA, are positive in tests for mutagenicity. The mechanisms of the carcinogenicity of non-genotoxic carcinogens include promotion, cytotoxicity and oxidative stress. As mutagenicity tests do not allow determination the carcinogenicity of chemical substances, long-term experiments on rodents should be considerred the only reliable method of carcinogenicity detection.     

Alternative strategies for carcinogenicity assessment: an efficient and simplified approach based on in vitro mutagenicity and cell transformation assays

The need for tools able to predict chemical carcinogens in less time and at a lower cost in terms of animal lives and money is still a research priority, even after several decades of effort in that direction. Now, new regulatory requirements (e.g. the Registration, Evaluation, Authorisation and Restriction of Chemical substances recently implemented in Europe) have even increased the pressure to develop new tools in this field. Drawbacks of the present testing strategies have come to light again recently especially in view of new requirements in worldwide regulations. Among these are (i) the lack of assays able to identify non-genotoxic carcinogens, (ii) the exaggerated rate of misleading (false) positive results of the in vitro mammalian cell-based short-term mutagenicity tests and (iii) the extremely low sensitivity of in vivo short-term mutagenicity tests. Within this perspective, we analyse the contribution of cell transformation assays (CTAs), and we show that they are a valid complement to tools able to detect DNA-reactive carcinogens. We also show that a tiered strategy, with inexpensive and fast tests in Tier 1 (e.g. the Ames test or structural alerts) and the Syrian hamster embryo CTA in Tier 2, is able to identify up to 90% of carcinogens.     

Inter-laboratory comparison of turkey in ovo carcinogenicity assessment (IOCA) of hepatocarcinogens

In three independent laboratories carcinogens (diethylnitrosamine, DEN, 4-(N-methyl-N-nitrosamino)-1-(3-pyridyl)-1-butanone, NNK) and non-carcinogens (N-nitrosoproline, nicotine) were evaluated in turkey eggs for in ovo carcinogenicity assessment (IOCA). Compounds were injected into aseptic fertilized eggs. After incubation for 24 days, foci of altered hepatocytes (FAH), some with a pseudoglandular structure and/or signs of compression of the surrounding tissue were observed in the fetal liver. All laboratories were able to distinguish unequivocally the hepatocarcinogen-exposed groups from those exposed to non-carcinogens or the vehicle controls, based on the pre-specified evaluation parameters: tumor-like lesions, pseudoglandular areas and FAH. In addition to focal changes, only the carcinogens induced hepatocellular karyomegaly. Lower doses of the carcinogens, which did not induce FAH, were sufficient to induce hepatocellular karyomegaly. After exposure to 4 mg DEN, gall bladder agenesis was observed in all fetuses. The IOCA may be a valuable tool for early investigative studies on carcinogenicity and since it does not use rodents may complement chronic rat or mouse bioassays. Test substances that are positive in both rodents and fertilized turkey eggs are most probably trans-species carcinogens with particular significance for humans. The good concordance observed among the three laboratories demonstrates that the IOCA is a reliable and robust method.     

DNA adducts: endogenous and induced

Human exposure to DNA damaging agents can arise from exogenous sources or endogenous processes that occur normally or in pathological states. DNA isolated from human tissues, obtained from the very young to the old, contains detectable amounts of a number of different types of DNA adducts that reflect exposure to both known carcinogens and as yet unidentified genotoxic agents. The levels of DNA damage observed in human studies as a result of exogenous exposures (noniatrogenic) is of the order of 1 adduct per 10(7)-10(9) normal DNA bases, whereas that arising from endogenous exposures may potentially be several orders of magnitude higher. Large interindividual variations in DNA adduct levels have been reported, and these are probably the result of host and environmental factors, although variation in analytical and sampling procedures may also play a role. It is important to recognize that the presence of DNA adducts in a tissue does not necessarily indicate a specific tumorigenic risk for that tissue, as other factors downstream of DNA adduct formation (including DNA repair and cell proliferation) play an important role in determining overall risk.     

Inverse relationship between epidermal growth factor induced proliferation and expression of high affinity surface epidermal growth factor receptors in rat hepatocytes

Rat hepatocytes express large numbers of high and low affinity surface membrane receptors (EGFR) for epidermal growth factor (EGF) but the roles of EGF and EGFRs in hepatocyte proliferation in vivo are unclear. F344 rat hepatocytes in primary culture proliferated maximally in response to continuous serum-free culture with 3.3 nM (20 ng/ml) EGF, as quantified by cumulative [3H]thymidine labeling index. However, serum concentrations of EGF in rats with normal livers or induced hepatocyte proliferation due to partial hepatectomy, carbon tetrachloride-induced necrosis, or hepatic neoplasia were consistently below 0.1 nM. The 3- or 6-hour pulse exposures to EGF (1.7 nM) between 0 to 16 hours had minimal effect on labeling index at 48 hours, but these pulse exposures at 24 or 32 hours were equivalent to continuous exposure. At 24 and 32 hours, the total specific surface binding of [125I]EGF to hepatocytes cultured free of EGF decreased to 43 and 24% of the initial values, respectively. Scatchard analysis of EGF binding indicated that hepatocytes lost all high affinity EGFRs (Kd of 0.08 nM) by 24 hours. Low affinity [125I]EGF binding at 0 hour (Kd 0.8 nM) was further reduced at 24 hours (Kd = 3.9 nM) and corresponded more closely to mitogenic concentrations of EGF in culture. These studies demonstrate that proliferative responsiveness of hepatocytes to EGF increases during culture by a process that involves prior loss of constitutive high affinity EGFRs. These results suggest that constitutive high affinity EGFRs do not elicit the proliferative response to EGF.     

Evidence of a threshold-effect for 2-amino-3,8-dimethylimidazo-[4,5-f]quinoxaline liver carcinogenicity in F344/DuCrj rats

To estimate potential human risk of exposure to a food-derived, genotoxic hepatocarcinogen, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), a 2-year carcinogenicity test was conducted using male F344 rats administered MeIQx-containing diet at doses of 0 (control), 0.001, 1, and 100 ppm. The lowest dose 0.001 ppm was established as equivalent to the daily intake of this carcinogen in humans (0.2 to 2.6 microg/man/day). Significant decreases of survival rate and body weight gain were observed in rats treated with 100 ppm MeIQx. Histopathological examination revealed significant induction of hepatocellular carcinomas, adenomas, and development of glutathione S-transferase placental form-positive foci with MeIQx at 100 ppm. Moreover, the incidences of Zymbal's glands carcinoma, mammary fibroadenoma, and subcutaneous fibroma were found significantly increased in a 100 ppm MeIQx group. However, no significant induction of altered preneoplastic hepatocellular foci was observed in 0.001 and 1 ppm groups as compared to the controls. 8-Hydroxy-2'-deoxyguanosine levels in the rat liver DNA of the 100 ppm-treated group were not elevated, but MeIQx-DNA adduct formation increased as compared with the 1 ppm case, albeit without significance. No significant induction of any other neoplastic lesions related to the carcinogen administration was found in MeIQx-administered groups except for 100 ppm. These results imply that 1 ppm may be a no-effect level for MeIQx carcinogenesis.     

Cytomimetic engineering of hepatocyte morphogenesis and function by substrate-based presentation of acellular E-cadherin

Although cadherin-mediated intercellular contacts can be integral to the maintenance of functionally competent hepatocytes in vitro, the ability to engineer hepatocellular differentiated function via acellular E-cadherin has yet to be thoroughly explored. To investigate the potential of substrate-presented, acellular E-cadherin to modulate hepatocellular self-assembly and functional fate, rat hepatocytes were cultured at sparse densities on surfaces designed to display recombinant E-cadherin/Fc chimeras. On these substrates, hepatocytes were observed to recognize microdisplayed E-cadherin/Fc and responded by modulating the spatial distribution of the intracellular cadherin-complexing protein beta-catenin. Substrate-presented E-cadherin/Fc was also found to markedly alter patterns of hepatocyte morphogenesis, as cellular spreading and two-dimensional reorganization were significantly inhibited under these conditions, leading to multicellular aggregates that were considerably more three-dimensional in nature. Increasing cadherin exposure was also associated with elevated levels of albumin and urea secretion, two markers of hepatocyte differentiation, over control cultures. This suggested that cell-substrate cadherin engagement established more functionally competent hepatocellular phenotypes, coinciding with the notion that E-cadherin is a differentiation-inducing ligand for these cells. The morphogenetic and function-promoting effects of substrate-bound E-cadherin/Fc were further enhanced under conditions in which protein A was utilized as an anchoring molecule to present cadherin molecules, suggesting that ligand mobility may play an important role in the effective establishment of cell-to-substrate cadherin interactions. Interestingly, the percent increase in function detected for conditions of high cadherin exposure versus control cultures was found to be substantially higher at extremely low cell densities. This observation indicated that hepatocytes respond to substrate-presented E-cadherin even in the absence of native intercellular interactions and associated juxtacrine signaling. The incorporation of acellular E-cadherin on biomaterial substrates may thus potentially present a means to prevent hepatocellular dedifferentiation by maintaining liver-specific function in otherwise severely functionally repressive culture conditions.     

Forestomach carcinogens: pathology and relevance to man

Squamous cell carcinomas of the forestomach have been observed in many carcinogenicity studies in rodents, especially after oral or gavage exposure. The histopathological diagnosis of forestomach lesions and the relevance of the data for human risk estimation can be controversial. The pathological classification may be troublesome because of the low-grade malignancy and the pseudoepitheliomatous hyperplasia that may develop after ulceration and inflammation. For human risk estimation it is important to understand the mechanism of action; this is illustrated by examples using butylated hydroxyanisole, methyl bromide, and epichlorohydrin. Another feature that complicates risk estimation is the absence of a homologue for the forestomach in man. The potential risk from non-genotoxic forestomach carcinogens in man involves exposure of the mouth, pharynx, and esophagus at dose levels that exert irritating action. It is assumed that exposure to non-genotoxic chemicals at concentrations far below those having irritating potential is not hazardous to humans.     

New structural concepts for predicting carcinogenicity in rodents: an artificial intelligence approach

The Computer Automated Structure Evaluation (CASE) method for studying structure-activity relationships has been applied to a data base of rodent carcinogens. It has been demonstrated that CASE is able to identify determinants embedded in the molecular structure which, with a high probability, predict rodent carcinogenicity. CASE has also identified determinants associated with the activity of non-genotoxic carcinogens, thereby suggesting that there is a structural commonality in the activity of these molecules. The present study reveals that there are "universal" as well as species-specific structural determinants of carcinogenicity. CASE was able to predict the carcinogenicity in rodents of certain endogenous pesticides in edible plants.     

Systematical analysis of impacts of heat stress on the proliferation, apoptosis and metabolism of mouse hepatocyte

Heat stress will stimulate cells of living organisms to generate heat shock proteins (Hsps). In the mouse liver, impacts of heat stress on hepatocyte proliferation, apoptosis and metabolism have not been studied systematically at different temperatures. In this research, the test mice were heated to 40, 42, 44 and 46°C, respectively, for 20?min and recovered at room temperature for 8?h in normal feeding conditions; the control animals were kept at room temperature without heat stress. The expression levels of Hsp70, Pcna, Bax, Bcl2, cytochrome P450 1A2 (CYP1A2), CYP2E1 and analog of CYP3A4 (not reported in mouse before), the parameters reflecting stress strength, cell proliferation, apoptosis and metabolism, were detected by western blotting, immunohistochemistry and semi-quantitative RT-PCR in test and control mice. Haematoxylin-eosin (H&E) staining and TUNEL analysis were further used to study the impacts of heat stress at different temperatures on hepatocellular necrosis and apoptosis. Serum AST and ALT levels, the markers of liver injury, were measured after heat stress at different temperatures. The data show that Hsp70 expression was significantly increased when temperature increased (P??0.05). At higher temperatures (44 or 46°C), expression of Pcna was decreased while hepatocyte necrosis and apoptosis were induced (P?相似文献   

Overexpression of transforming growth factor-alpha causes liver enlargement and increased hepatocyte proliferation in transgenic mice.

Transforming growth factor-alpha (TGF-alpha) expression is associated with hepatocyte DNA replication both in vivo and in culture. Our previous work using TGF-alpha transgenic mice showed that constitutive overexpression of this growth factor in the liver causes hepatic tumors in 75 to 80% of the animals at 12 to 15 months of age. To understand the cellular events by which TGF-alpha overexpression leads to abnormal liver growth, we examined hepatocyte proliferative activity in young and old TGF-alpha transgenic mice and hepatocyte ploidy in normal, dysplastic, and neoplastic livers of these animals. At 4 weeks of age, transgenic mice had higher liver weights and liver weight/body weight ratios than non-transgenic mice of the same age and hepatocyte proliferative activity, measured by 3H-thymidine incorporation after 3- and 7-day infusion, proliferating cell nuclear antigen staining, and mitotic index determination, was 2 to 3 times higher than in controls. In both transgenic and non-transgenic mice hepatocyte proliferation declined with age but the decrease was much more pronounced in control animals, so that at 8 months of age, hepatocyte replication was 8 to 10 times higher in transgenic animals. Surprisingly, however, transgenic and non-transgenic mice at this age had similar liver weight/body weight ratios. Labeling studies done in 3-month-old animals revealed that hepatocyte turnover was much faster in transgenic than in control animals, suggesting that a homeostatic compensatory mechanism involving cell death tended to restore normal liver weight/body weight ratios in older transgenic mice. Ploidy analyses showed that at 4 weeks of age transgenic mice had a higher proportion of diploid and tetraploid hepatocytes and that the hepatocellular tumors which developed in TGF-alpha transgenic mice at 13 months of age contained a higher fraction of diploid hepatocytes than that present in adjacent tissue or in dysplastic livers. The results demonstrate that constitutive overexpression of TGF-alpha causes increased hepatocyte proliferation and liver enlargement in young animals and is associated with a delay in the establishment of hepatic polyploidy. These findings as well as the response of transgenic mice to partial hepatectomy show that constitutive overexpression of TGF-alpha initially caused increased but regulated hepatocyte proliferation which in older animals was compensated in part by a faster cell turnover. At 8 to 10 months of age, proliferative activity may become constitutive in some TGF-alpha expressing hepatocytes.(ABSTRACT TRUNCATED AT 400 WORDS)     

The slow induction of resistant hepatocytes during initiation of hepatocarcinogenesis by the nongenotoxic carcinogen clofibrate

This study was designed to explore whether a well-known nongenotoxic liver carcinogen, clofibrate, would induce rare resistant hepatocytes similar to those seen during initiation of hepatocarcinogenesis with many genotoxic carcinogens. Male young adult F344 rats were exposed to a control diet containing 0.5% (w/w) clofibrate for 3, 6, or 10 months. After 1 month on a diet free of clofibrate, the animals were assayed for resistant hepatocytes by a standardized selection procedure using 2-acetylaminofluorene as the inhibitor and partial hepatectomy as a strong stimulus for cell proliferation. No resistant hepatocytes were found in the animals exposed to clofibrate for 3 months or in any of a series of control animals. However, animals on the clofibrate for 6 and 10 months contained resistant hepatocytes that were clonally expanded to produce hepatocyte nodules. These nodules were indistinguishable on gross and microscopic examination from hepatocyte nodules seen in animals in which nodules are induced with one of many different genotoxic carcinogens. Also, like those nodules, the nodules seen in the animals exposed to clofibrate stained positively for glutathione S-transferase 1-1 and gamma-glutamyl transpeptidase and negatively for ATPase. The evidence from this study indicates that the nongenotoxic carcinogen, clofibrate, induces early cellular changes in the liver that are very similar to those induced by many different genotoxic carcinogens. These changes are manifest as a resistance phenotype in a few scattered hepatocytes that now can be clonally expanded selectively to form hepatocyte nodules. However, the resistant hepatocytes are induced by clofibrate much more slowly. Whether this basic similarity pertains to the later steps in the hepatocarcinogenic process remains to be studied.