Mutational inactivation of the xeroderma pigmentosum group C gene confers predisposition to 2-acetylaminofluorene-induced liver and lung cancer and to spontaneous testicular cancer in Trp53-/- mice.
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Mice that are genetically engineered to mimic the human hereditary cancer-prone DNA repair-defective disease xeroderma pigmentosum (XP) are highly predisposed to UV radiation-induced skin cancer. It is not clear, however, whether XP mice or humans are predisposed to cancers in other tissues associated with exposure to environmental carcinogens. To test the importance of nucleotide excision repair in protection against chemical carcinogenesis in internal organs, we treated XPC mutant (XPC-/-) mice with 2-acetylaminofluorene and NOH-2-acetylaminofluorene. We observed a significantly higher incidence of chemically induced liver and lung tumors in XPC-/- mice compared with normal and heterozygous littermates In addition, the progression of liver tumors in XPC-/- Trp53+/- mice is accelerated compared with XPC-/- Trp53+/+ animals. Finally, we demonstrate a higher incidence of spontaneous testicular tumors in XPC-/- TrpS3-/- double mutant mice compared with XPC+/+ Trp53-/- mice. (+info)
Pseudoenzymatic reduction of N-hydroxy-2-acetylaminofluorene to 2-acetylaminofluorene mediated by cytochrome P450.
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N-hydroxy-2-acetylaminofluorene (N-OH-AAF) was reduced to 2-acetylaminofluorene by rat liver microsomes in the presence of both NAD(P)H and FAD under anaerobic conditions. The microsomal reduction proceeds as if it were an enzymatic reaction. However, when the microsomes were boiled, the activity was not abolished, but was enhanced. The activity was also observed with cytochrome P450 2B1 alone, without NADPH-cytochrome P450 reductase, in the presence of these cofactors. Hematin also exhibited a significant reducing activity in the presence of both a reduced pyridine nucleotide and FAD. The activities of microsomes, cytochrome P450 2B1 and hematin were also observed upon the addition of photochemically reduced FAD instead of both NAD(P)H and FAD. The microsomal reduction of N-OH-AAF appears to be a non-enzymatic reaction by the reduced flavin, catalyzed by the heme group of cytochrome P450. (+info)
Effect of detergents on the N-and ring-hydroxylation of 2-acetamidofluorene by hamster liver microsomal preparations.
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Effects of detergents such as cholate, deoxycholate and Triton X-100 were studied on N-and ring-hydroxylation of 2-acetamidofluorene by reconstituted and unresolved microsomal systems from livers of hamsters pretreated with 3-methylcholanthrene. Triton X-100 (2.5 mg/nmol of cytochrome P-448) inhibited N-and ring-hydroxylation by wholemicrosomal preparations by 40 and 90% respectively Deoxycholate at the same concentration inhibited both hydroxylations completely, whereas cholate inhibited N-and ring-hydroxylation by 40 and 50% respectively. In reconstitution studies, the presence of Triton X-100(0.5-1.0mg/nmol of cytochrome P-448) along with unsolubilized cytochrome P-448 fraction and solubilized reductase fraction increased N-hydroxylation to an appreciable extent compared with ring-hydroxylation. Both cholate and deoxycholate at 0.5-1.0 mg concentrations had a greater stimulatory effect on ring-than on N-hydroxylation activity in such a reconstituted system. (+info)
Bone marrow as a potential source of hepatic oval cells.
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Bone marrow stem cells develop into hematopoietic and mesenchymal lineages but have not been known to participate in production of hepatocytes, biliary cells, or oval cells during liver regeneration. Cross-sex or cross-strain bone marrow and whole liver transplantation were used to trace the origin of the repopulating liver cells. Transplanted rats were treated with 2-acetylaminofluorene, to block hepatocyte proliferation, and then hepatic injury, to induce oval cell proliferation. Markers for Y chromosome, dipeptidyl peptidase IV enzyme, and L21-6 antigen were used to identify liver cells of bone marrow origin. From these cells, a proportion of the regenerated hepatic cells were shown to be donor-derived. Thus, a stem cell associated with the bone marrow has epithelial cell lineage capability. (+info)
Resistance to the promotion of glutathione S-transferase 7-7-positive liver lesions in Copenhagen rats.
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Previously, we have shown that Copenhagen (Cop) rats are highly resistant to the induction of putative preneoplastic, glutathione S-transferase 7-7 (GST 7-7)-positive liver lesions following treatment with a modified resistant hepatocyte protocol. The objective of the current study was to establish the time course for the development of resistance and examine potential resistance mechanisms in Cop rats using F344 rats as susceptible controls. Male Cop and F344 rats (n = 25), 7-8 weeks of age, were initiated with diethylnitrosamine (200 mg/kg) and promoted 3 weeks later with four doses of 2-acetylaminofluorene (20 mg/kg) and a 2/3 partial hepatectomy (PH). Groups of rats from each strain were killed on days 2, 4, 7, 14 and 21 post-PH, 2 h after receiving bromodeoxyuridine. Cop livers contained similar numbers of GST 7-7-positive lesions to F344 livers on days 2 and 4 post-PH. The percent volume of liver occupied by these lesions did not differ between the strains on days 2, 4 and 7 post-PH. On day 14, however, approximately 29% of the liver volume in F344 rats was occupied by lesions, whereas in Cop rats this was significantly less (approximately 9%, P < 0.001). On day 21, lesions occupied approximately 58% of F344 rat livers and only approximately 6% of Cop livers. Despite these differences, the labeling index of hepatocytes was not significantly different between the strains at any time point, either within lesions or within surrounding normal liver. Furthermore, the apoptotic indices were not different between the strains at any time. However, differences were found in the extent of lesion remodeling (redifferentiation) and in the pattern of oval cell response following PH in Cop livers. By day 14 post-PH, approximately 76% of Cop liver lesions showed evidence of remodeling, compared with only approximately 14% of F344 lesions. The oval cell response to PH was equivalent in the two strains up to day 4 post-PH but by day 7, in F344 livers there was extensive migration of these cells into the liver parenchyma, whereas in Cop livers, the response remained localized to the portal regions. These results suggest that Cop resistance occurs at the promotion stage and not the initiation stage of carcinogenesis. Resistance appears not to be due to a lower proliferation rate nor to a higher apoptotic rate within Cop lesions. Precocious remodeling and/or a diminished oval cell response, however, may contribute to the resistance of Cop rats to the growth of GST 7-7-positive hepatic lesions. (+info)
Analysis of loss of heterozygosity in neoplastic nodules induced by diethylnitrosamine in the resistant BFF1 rat strain.
(6/450)
Loss of heterozygosity (LOH) at specific chromosomal regions is a frequent event in poorly differentiated human hepatocellular carcinomas (HCCs), but rare in mouse HCCs. This behavior could depend on interspecies differences in mechanisms of hepatocarcinogenesis or in developmental stage of lesions. To verify if LOH is involved in rat hepatocarcinogenesis, we studied LOH frequency in slowly growing neoplastic nodules induced by Solt-Farber model in diethylnitrosamine-initiated BFF1 rats. We analyzed, with microsatellites, markers at 67 rat loci dispersed over all chromosomes, corresponding to regions homologous to those lost in human HCCs or containing hepatocellular susceptibility (Hcs) or resistance (Hcr) loci in rat and mouse. In agreement with previous findings with mouse HCCs, but at variance with human HCCs, no detectable LOH was found at any locus in rats, suggesting rare LOH involvement in neoplastic nodules, with low tendency to progress to full malignancy, of BFF1 rats. (+info)
Development of resistance during the early stages of experimental liver carcinogenesis.
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The present study was designed to determine whether the resistant phenotype is acquired at the initiated cell stage itself or requires further exposure to a promoting regimen to express resistance. Male Fischer 344 rats were initiated with diethylnitrosamine (DENA) (200 mg/kg i.p.) and were subjected to either no further treatment or to the resistant hepatocyte (RH) model of liver tumor promotion. Six weeks later, the resistance of the focal lesions generated in these two groups to the mitoinhibitory effects of 2-acetylaminofluorene (2-AAF) was determined by subjecting the rats to two-thirds partial hepatectomy (PH) in the presence of a mitoinhibitory dose of 2-AAF (5 mg/kg i.p.) given at the time of PH. Labeling index was determined by administering multiple injections of [(3)H]thymidine. All rats were killed 48 h post-PH. While only a small percentage (23%) of the glutathione S-transferase-positive foci generated by DENA in the absence of an exogenous liver tumor promoting regimen were resistant to the mitoinhibitory effects of 2-AAF, a majority (85%) of the foci became resistant to 2-AAF following exposure to the RH model of liver tumor promotion. Further, initiated rats exposed to either 2-AAF or to CCl(4) alone, the two components of the RH model, resulted in 71% of the foci being resistant to the mitoinhibitory effects of 2-AAF. Similar patterns of results were obtained when the resistance of the foci to the mitoinhibitory effects of orotic acid, a liver tumor promoter and an inhibitor of DNA synthesis in normal hepatocytes, was monitored. These results suggest that the majority of initiated hepatocytes are not of resistant phenotype, however, they have acquired a unique ability to express resistance upon exposure to certain agents such as 2-AAF and CCl(4) or to a promoting regimen such as the RH model of liver tumor promotion. (+info)
Modulation of the gene network connected to interferon-gamma in liver regeneration from oval cells.
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Suppression subtractive hybridization was used to clone genes associated with proliferation of oval cells in rat liver regenerating after a 70% partial hepatectomy combined with the feeding of 2-acetylaminofluorene. A subset of the identified genes comprised interferon-gamma receptor alpha subunit (IFN-gammaRalpha), gp91phox, interleukin-1beta (IL-1beta), lymphocyte function-associated molecule-1alpha (LFA-1), eukaryotic initiation factor-2-associated 67-kd protein (eIF-2-associated 67-kd protein), and alpha-fetoprotein, which constitute part of the cellular program modulated by IFN-gamma. Therefore, expression analysis performed by Northern blotting and immunohistochemistry were extended to include IFN-gamma, the IFN-gamma receptor beta subunit (IFN-gammaRbeta), three secondary response genes induced by interaction of IFN-gamma with IFN-gamma receptor complexes, ie, IL-1beta-converting enzyme (ICE), intercellular adhesion molecule-1 (ICAM-1), and urokinase-type plasminogen activator receptor (uPAR), and a cytokine inducing IFN-gamma expression, ie, interleukin-18 (IL-18). The Northern blot analysis showed that all examined genes were modulated when progenitor-like oval cells were activated and recruited for liver regeneration. Immunohistochemistry localized the subunits of the IFN-gamma receptor complex, IFN-gammaRalpha and IFN-gammaRbeta, the secondary response genes uPAR and ICAM-1, the IFN-gamma-inducing factor IL-18, and ICE to the ductular structures of oval cells. In contrast, during liver regeneration after a 70% partial hepatectomy, only modulation of IL-1beta and ICE was observed. Our results, therefore, indicate that IFN-gamma-mediated events may be particularly important when cells in the bile ductules must respond to liver damage by production of ductular oval cells. (+info)