Promotion by dietary phenobarbital of hepatocarcinogenesis by 2-methyl-N,N-dimethyl-4-aminoazobenzene in the rat.
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The hepatocarcinogenicity of 2-methyl-N,N-dimethyl-4-aminoazobenzene, previously shown to be noncarcinogenic in adult rats in the absence of further treatment, was observed by following a 1- to 6-week period of feeding this dye to weanling rats with the dietary administration of 0.05% phenobarbital for up to 70 weeks. Many large hepatocellular carcinomas developed in the phenobarbital-treated animals by 72 weeks, whereas a very small number of tiny neoplastic nodules, including one carcinoma, were seen in the rats not given this drug. This study suggests that the use of promoting agents, following the short-term administration of weak carcinogens for the liver, can be useful in demonstrating the initiating activity of such compounds. This system may be useful in the identification of such agents in the environment. (+info)
A carcinogen-resistant inbred strain DRH/Sea has been developed from the Crj:Donryu strain. The rats had a very low incidence of liver tumors when they were fed diets containing a hepatocarcinogen such as 3'-methyl-4-dimethylamino-azobenzene (3'-Me-DAB). Despite using 3'-Me-DAB during the stage of selection, the DRH/Sea rats developed normally, reproduced and did not have any spontaneous tumor in the lung, liver or uterus at over 1 year of age. Although their growth curves were similar to the Crj:Donryu rats, the progression of polyploidization in the liver was significantly delayed when compared with Crj:Donryu rats. Mitogenic changes that occurred in the liver caused by either 3'-Me-DAB or lead nitrate were less significant in the DRH rats than in Crj:Donryu rats. Furthermore, the growth rate of cultured fibroblasts derived from the DRH rats was slower than that of Crj:Donryu rats. These results, together with our previous results, suggest that slow growth potential is present under certain conditions in DRH rats. These findings may explain partly the meaning of the different susceptibility to hepatocarcinogens. (+info)
Immunohistochemical localization of transforming growth factor alpha in chemically induced rat hepatocellular carcinomas with reference to differentiation and proliferation.
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Hepatocellular carcinomas (HCCs) were induced in male Fischer 344 rats with dietary 3'-methyl-4-(dimethylamino)-azobenzene treatment and were classified into solid, glandular (well- or poorly differentiated), and trabecular types. Investigation of cell proliferation kinetics and immunohistochemical localization of transforming growth factor alpha (TGF-alpha) demonstrated all solid (n = 24) and poorly differentiated glandular type (n = 6) HCCs to have TGF-alpha-positive nuclei. Nuclear staining of TGF-alpha was also observed in 13 of 28 (46%) trabecular-type HCCs, whereas 12 (43%) exhibited cytoplasmic staining, and 3 (11%) were negative. As for well-differentiated glandular HCCs, 7 of 20 (35%) were positively stained in their nucleus, another 7 (35%) demonstrated antibody binding in the cytoplasm, and 6 (30%) were negative. The order for growth rate evaluated by bromodeoxyuridine (BrdU) labeling was solid (38.22%), poorly differentiated glandular (26.82%), trabecular (7.98%), and well-differentiated glandular (2.57%) types. For trabecular HCCs with nuclear, cytoplasmic, or negative TGF reactions, values were 13.39% (n = 13), 3.61% (n = 12), and 2.01% (n = 3), respectively. Likewise, BrdU-labeling indices for the counterpart groups of well-differentiated glandular type HCCs were 4.53, 1.91, and 1.29%, respectively. The results indicate that TGF-alpha expression might be linked to histopathological differentiation and cell proliferation in rat HCCs. (+info)
Resistance of DRH strain rats to chemical carcinogenesis of liver: genetic analysis of later progression stage.
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The inbred DRH rats are highly resistant to the induction of hepatocellular carcinoma (HCC) by feeding of 3'-methyl-4-dimethylaminoazobenzene (3'-Me-DAB). Previously, we found that two quantitative trait loci (QTLs), Drh1 and Drh2, significantly reduced the number, size and area of glutathione S-transferase-placental form (GST-P)-positive foci and GST-P mRNA levels in (F344xDRH)F(2) rat livers induced by feeding 3'-Me-DAB for 8 weeks. It is unclear, however, whether these QTLs affecting pre-neoplastic lesions are also the determinants of the later stage hepatocarcinogenesis, and whether there are any additional QTLs affecting hepatocarcinogenesis in the progression stage. To answer these questions, we analyzed QTL parameters for liver tumors in 99 (F344xDRH)F(2) rats induced by feeding 3'-Me-DAB for 20 weeks. The QTL parameters examined were GST-P mRNA, ornithine decarboxylase activity, and the number and total area of HCC/nodules macroscopically detectable on the liver surface. In composite interval mapping, we observed two major QTL peaks overlapping on the map positions of Drh1 on rat chromosome 1 (RNO1) and Drh2 on RNO4, respectively. The newly mapped QTL on RNO1 affected the GST-P mRNA level at 20 weeks of 3'-Me-DAB feeding, but did not affect the number and size of tumors. The primary effect of Drh1 is, therefore, to inhibit GST-P induction and to prevent enzyme altered foci (EAF) formation. On the other hand, the QTLs on RNO4, co-mapped to Drh2, affected all parameters of liver tumors examined except for the level of GST-P mRNA. The latter QTLs influenced not only the induction of GST-P and formation of EAF but also the progression of tumors in the later stage of hepatocarcinogenesis. The GST-P induction is differentially controlled by stages of hepatocarcinogenesis and the DRH resistance to carcinogenesis is principally attributed to the QTLs on RNO4 out of two resistance QTLs identified in the pre-neoplastic stage. (+info)
Expressions of c-fos and c-myc genes during 3'-methyl-4-dimethylaminoazobenzene (3'-MeDAB)-induced rat hepatocarcinoma.
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We investigated the expression of the growth-related nuclear proto-oncogenes, c-fos and c-myc, in early preneoplastic regions and tumor nodules of 3'-MeDAB induced rat hepatocarcinoma. To amplify the levels of these transcripts, we gave cycloheximide (100 mg/kg B.W. i.p.) to each group of rats. The elevated levels of the 2.2 kb c-fos and 2.4 kb c-myc transcripts appeared as early as the 2nd week after feeding on the 3'-MeDAB diet and lasted through the 4th; 6th weeks and tumor. Southern blot analysis indicated that gross amplification or rearrangements were not observed in DNA of the preneoplastic livers and hepatoma nodules. We also measured the rate of the incorporation of [3H] thymidine into hepatic DNA in order to monitor the rate of cell proliferation occurring at the early preneoplastic periods. We have found that the rate of [3H] thymidine incorporation corresponds to the elevated levels of c-fos and c-myc transcripts in the precancerous stages. This finding suggests that the elevated expressions of c-fos and c-myc may result from the continuous cell proliferative stimuli generated in the carcinogen altered cells, which is essential to the initiation and promotion of chemical hepatocarcinogenesis. (+info)
RNA sulfurtransferase activity in rat liver and chemically induced hepatomas.
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RNA sulfurtransferase activity has been detected in rat liver and in hepatomas from rats fed a diet containing 0.06% 3'-methyl-4-dimethylaminoazobenzene for 14 to 18 weeks. The reaction measured was the transfer of sulfur from cysteine to acceptor sites in Escherichia coli B transfer RNA (tRNA). Specific activities of the enzymes in liver and hepatoma supernatant fractions were similar, as were the rates and extents of sulfur transfer to tRNA. DEAE-cellulose chromatography of digests of the [35S]tRNA reaction products revealed 3 peaks associated with nucleotide material, the amounts of these peaks differing in tRNA from liver and hepatoma systems. This may suggest differences in specific sulfurtransferases in these tissues. (+info)
Binding of the azocarcinogen 3'-methyl-p-dimethylaminoazobenzene to cellular components of normal rat liver and azocarcinogen-induced hepatomas.
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The location of binding sites for 3'-methyl-p-dimethylaminoazobenzene (3'-Me-DAB) or metabolites on components of rat liver cells and hepatoma cells in tumors induced by this carcinogen was determined at 2 stages during the induction of tumors in rats: (a) in normal liver immediately following the application of a massive dose of the azocarcinogen by intragastric feeding, and (b) in liver and tumor after hepatomas had developed following repeated exposures to the carcinogen by s.c. injections. Bound 3'-Me-DAB or metabolites were detected by the use of rabbit antisera directed against either p-azoazobenzene or p'-azo-p-dimethylaminoazobenzene in an indirect fluorescent antibody technique. Soon after massive intragastric doses of 3'-Me-DAB, the staining observed when the anti-p-azoazobenzene antiserum was used was principally on cytoplasmic components of liver cells with some staining of the intranuclear components. When the second antiserum, anti-p'-azo-p-dimethylaminoazobenzene antiserum, was used, the most intense fluorescent staining was on the nuclear membranes, although there was some cytoplasmic and intranuclear staining as well. (+info)
Effects of gomisin A on hepatocarcinogenesis by 3'-methyl-4-dimethylaminoazobenzene in rats.
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We examined the effects of gomisin A on tumor promotion in the liver after a short-term feeding of 3'-methyl-4-dimethylaminoazobenzene (3'-MeDAB) to rats, compared with the effects of phenobarbital. Male Donryu rats were fed ad libitum a diet containing 0.06% 3'-MeDAB and 0.03% or 0.01% gomisin A or water containing 0.05% phenobarbital. Gomisin A and phenobarbital did not cause any proliferative and neoplastic lesions by themselves in 40 weeks of feeding. Altered foci in the liver increased with a peak at 12 weeks after the rats were fed 3'-MeDAB. Gomisin A decreased the number of hepatic altered foci such as the clear cell and basophilic cell type foci in the early stages. Phenobarbital enhanced neoplastic alterations so that the number and size of the foci were much larger in the phenobarbital-combined group than in the 3'-MeDAB-control group. Thus, phenobarbital acted as a promoter of cells initiated by 3'-MeDAB; on the other hand, gomisin A showed a weak suppressive effect on tumor promotion. (+info)