Previous studies in animals and humans have shown that selenium compounds can prevent cancer development. In this work we studied the tumor preventive effect of selenium supplementation, administrated as selenite, in the initiation, promotion and progression phases in a synchronized rat model for chemically induced hepatocarcinogenesis, the resistant hepatocyte model. Selenite in supra-nutritional but subtoxic doses (1 and 5 p.p.m.) was administrated to the animals through the drinking water. Such supplementation during the initiation phase did not have a tumor preventive effect. However, selenite treatment during the promotion phase decreased the volume fraction of pre-neoplastic liver nodules from 38% in control animals to 25 (1 p.p.m.) and 14% (5 p.p.m.) in the selenite-supplemented groups. In addition the cell proliferation within the nodules decreased from 42% in the control to 22 (1 p.p.m.) and 17% (5 p.p.m.). Immunohistochemical staining for the selenoenzyme thioredoxin reductase 1 revealed an increased expression of the enzyme in liver nodules compared with the surrounding tissue. The activity was reduced to 50% in liver homogenates from selenium-treated animals but the activity of the selenoenzyme glutathione peroxidase was essentially unaltered. Selenite treatment (5 p.p.m.) during the progression phase resulted in a significantly lower volume fraction of liver tumors (14 compared with 26%) along with a decrease in cell proliferation within the tumors (34 compared with 63%). Taken together our data indicate that the carcinogenetic process may be prevented by selenium supplementation both during the promotion and the progression phase.

Previous studies have indicated that isolated preneoplastic rat hepatocytes in vitro fail to induce nuclear p53 protein and fail to block replication in response to genotoxic compounds. This suggests that defects in the protection of genomic integrity are part of their premalignant character. In the present study, we have investigated if similar defects occur in vivo. Preneoplastic glutathione-S-transferase (GST) 7-7-positive foci were induced in male Wistar rats by diethylnitrosamine (DEN) initiation and promotion with 2-acetylaminofluorene (2-AAF)/partial hepatectomy (PH). The response to genotoxic damage was studied by X-irradiation. p53 protein was moderately expressed in nuclei in surrounding hepatocytes. This nuclear p53 staining had decreased 2 weeks after 2-AAF treatment. In foci, the protein was detected in the cytoplasm whereas the nuclei were negative. Levels of p21 waf1/cip1 protein were high in nuclei and cytoplasm of surrounding hepatocytes, whereas the expression in foci was low. A low level of Mdm2 in nuclei was observed in surrounding liver, while both Mdm2 and Bcl-2 protein were strongly expressed in the cytoplasm in foci. X-ray exposure further induced nuclear expression of p53, p21 waf1/cip1 , and Mdm2 in surrounding hepatocytes, but focal nuclei were still negative. DNA replication was strongly reduced by X-irradiation in surrounding hepatocytes, but only partially reduced in the foci. These results indicate that the p53 pathway of response to genomic stress is impaired in preneoplastic Over the last decades the process of experimental hepatocarcinogenesis in rats has been intensively studied (reviewed in Schulte-Hermann 1 and Pitot 2 ). Early in the process, preneoplastic cells emerge, which are able to proliferate during exposure to cytotoxic/genotoxic tumor promoters 3 while phenotypically normal cells are blocked in their growth. 4 Changes in expression of several proteins have been related to the premalignant nature of the preneoplastic cells, such as increased expression of glutathione-S-transferase (GST) 7-7, p-glycoprotein, and ␥-glutamyltranspeptidase, and decreased expression of a sulfotransferase and alterations in cytochrome P450s and c-myc expression. [5][6][7][8][9][10][11][12] The molecular mechanisms of their escape from growth inhibition as normally seen in hepatocytes after exposure to genotoxic compounds are, however, not clear.The tumor suppressor p53 plays an important role in maintaining the integrity of the genome. 13 Wild-type p53 protein blocks the cell cycle in G1 after DNA damage and may stimulate DNA repair. [13][14][15][16] The p53 protein is mutated in over 50% of the human tumors indicating that loss of p53 is an important event in tumor development. 13,14 We have shown earlier that compounds with hepatic tumor promoting activity are able to induce p53 in rat liver in vivo, 17,18 and that cells from preneoplastic foci in vitro and in vivo express less nuclear p53 compared with surrounding hepatocytes during treatment with tumor promoters 19 (Ohlson et al., subm...

Transferrin binding was found to be around 60-fold higher in hepatocyte nodules compared to normal liver, with no apparent differences in binding affinity or molecular weight of binding proteins, indicating that the increase in [125I]transferrin binding was the result of an increased number of binding sites with similar properties as in normal liver. The relative 'induction' of transferrin receptors was most marked in the total membrane fraction followed by membrane subfractions comprising endoplasmic reticulum, the Golgi complex and endocytic vesicles. Despite the increased number of transferrin receptors, the in vivo endocytosis of transferrin, measured as uptake of [125I]ferrotransferrin, was quantitatively similar in nodular cells compared to normal liver. The number of transferrin receptors in regenerating liver, 48 h after partial hepatectomy, was increased 20-fold over normal levels, but binding affinity, receptor structure and kinetics of transferrin uptake were normal. A slower than normal rate of 59Fe accumulation in hepatocyte nodules may suggest an alteration in the dissociation of iron from ferrotransferrin, thereby suggesting one mechanism relevant to the iron storage deficiency in nodular cells.

2-Nitrofluorene (NF), a model substance for nitrated polycyclic aromatic hydrocarbons, is present in exhaust from diesel- and petrol-driven vehicles, in exhaust from kerosene heaters, and in urban air and river sediments. Therefore the possible consequences of human exposure are important to elucidate. In the present study the initiating and promoting activity of NF was studied in a liver model for chemical carcinogenesis in the rat. Furthermore, the in vivo metabolism and formation of genotoxic metabolites of NF were investigated. NF was found to be a moderate initiator and a weak promoter when compared to diethylnitrosamine (DEN) and 2-acetylaminofluorene (AAF) respectively, both of which are potent carcinogens. Animals given NF excreted more urinary mutagenicity when compared to animals given AAF or DEN.

2-Nitrofluorene (NF) is a model compound for nitroarenes which has been identified in diesel exhaust and in urban air. The current study was carried out to observe the carcinogenicity of different doses of NF to rats and DNA adduct formation in different organs at an early stage of NF administration. One group of rats was fed basal diet as a control, whereas the other three groups of rats were fed basal diet supplemented with different amounts of NF (0.24, 0.95 and 2.37 mmol NF/kg diet, referred to as low, medium and high dose, respectively). The rats were exposed to NF continuously for 11 months, after which all groups of rats were fed basal diet without NF for another 13 months. In the high dose group hepatocellular carcinomas were found in all rats (20/20), forestomach squamous carcinomas in 11 and cortical kidney carcinomas in 10 rats. Fifteen out of 19 rats fed the medium dose of NF had hepatocellular carcinomas, 16 had forestomach squamous carcinomas and 15 had cortical kidney carcinomas. The major tumors of the rats fed the low dose of NF were forestomach squamous carcinomas (10/18). DNA adducts formed in tumor target organs after 1, 2, 6 and 10 days NF administration were dose- and time-dependent. Ten days after the start of NF administration DNA adduct levels were found to be 54, 11 and 6 DNA adducts/10(8) normal nucleotides in forestomach, liver and kidney respectively. In the non-tumor target organs levels in the range 1.7-4.8 DNA adducts/10(8) normal nucleotides were found. DNA adduct formation in this study showed a good correlation with the localization of tumors, although there is a need for additional factors for tumor formation. The results indicate that DNA adduct formation is an important factor for tumor formation and suggest that DNA adducts could be used as biomarkers for genotoxic risk.