The direct spectrophotometric observation of benzo(a)pyrene phenol formation by liver microsomes.
Optical spectral repetitive scan analysis during the oxidative metabolism of benzo(a)pyrene by liver microsomal suspensions reveals the time-dependent formation of an intermediate(s) of which the visible spectra resemble those of several benzo(a)pyrene phenols. Liver microsomes from 3-methylcholanthrene-treated rats showed a greater rate of formation of the phenols than did microsomes from control animals; the rate of formation catalyzed by liver microsomes from phenobarbital-pretreated rats was intermediate. When 3-hydroxybenzo(a)pyrene was used as a standard for comparison of activity, the rates of formation of phenols were compared when measured by fluorometric, spectrophotometric, or high-pressure liquid chromatographic analytical techniques. An epoxide hydrase inhibitor, 1,1,1-trichloropropene-2,3-oxide, enhanced phenol formation regardless of the source of liver microsomes, and 7,8-benzoflavone inhibited control and 3-methylcholanthrene-induced microsomal metabolism of benzo(a)pyrene, 7,8-Benzoflavone did not effect benzo(a)pyrene metabolism by liver microsomes from phenobarbital-pretreated rats. The effect of inhibitors on the spectrophotometric assay correlates well with the results obtained from benzo(a)pyrene metabolite analysis using high-pressure liquid chromatography. (+info)
Long-term transplantability and morphological stability of three experimentally induced urinary bladder carcinomas in rats.
Three transitional cell carcinomas induced in Fischer 344 rats by a methylcholanthrene pellet or a foreign body inserted locally into the bladder have been serially transplanted in the syngeneic strain for up to 6.5 years. There have been no changes in the individual morphological characteristics of the tumors during this time. Cells cultured in vitro for varying numbers of passages reproduce regularly the morphology of each tumor when they are injected back into the animals and results from a microcytotoxicity assay for cellular immunity indicate that they retain a common, bladder tumor-specific antigen. These tumors are useful for research in turmo biology and are offered to other scientists seeking transplantable carcinomas for experimentation. (+info)
Formation in isolated rat liver microsomes and nuclei of benzo(a)pyrene metabolites that bind to DNA.
The hepatic nuclear fraction isolated from 3-methylcholanthrene (MC)-treated rats contained enhanced levels of cytochrome P-450 and aryl hydrocarbon hydroxylase [benzo(a)pyrene (BP) monooxygenase], whereas the activities of epoxide hydrase and reduced nicotinamide adenine dinucleotide phosphate-cytochrome c reductase and the concentration of cytochrome b5 were not altered. The metabolite pattern of BP was investigated by using high-pressure liquid chromatography and was found to be similar in nuclei and microsomes from MC-treated rats. After incubation of the nuclear fraction with [3H]BP and reduced nicotinamide adenine dinculeotide phosphate, radioactivity was found to be associated with nuclear DNA and the extent of binding was markedly enhanced by pretreatment of the animals with MC. Binding was strongly inhibited by a-napthoflavone but was not influenced by 1,1,1-trichloropropene-2,3-oxide, an inhibitor of epoxide hydrase. In the presence of microsomes from MC-treated rats, increased binding of BP to DNA was observed in nuclei from both control and MC-treated rats; moreover, when the nuclear DNA was replaced by a corresponding amount of calf thymus DNA, the extent of binding was severalfold enhanced. In contrast to nuclei from control rats, the nuclear fraction from MC-treated rats showed an increase in bound radioactivity when incubated with a microsome-free supernatant, obtained by incubating microsomes from MC-treated rats with [3H]BP. The increase in extent of binding was eliminated in the presence of menadione or alpha-naphthoflavone. It is suggested that under the conditions used here the following different processes may have contributed to the total incorporation of BP products into nuclear DNA: (a) formation of DNA-binding products derived from BP by nuclear aryl hydrocarbon hydroxylase; (b) formation of DNA-binding products from microsomal BP metabolites by nuclear aryl hydrocarbon hydroxylase; and (c) direct transfer of reactive microsomal metabolites to nuclear DNA. (+info)
Phospholipid requirement for dimethylnitrosamine demethylation by hamster hepatic microsomal cytochrome P-450 enzyme system.
Extraction with butan-1-ol of freeze-dried microsomal fractions from livers of 3-methyl-cholarthrene-pre-treated hamsters removed about 90% of the total lipid content, but the lipid remaining proved sufficient for the cytochrome P-450 enzyme system to retain about 15-40% of its original catalytic activity for dimethylnitrosamine demethylation. Addition of butan-1-ol-extracted total phospholipid or phosphatidylcholine could not restore any activity, whereas the addition of the synthetic phospholipid dilauroyl phosphatidylcholine was able to restore almost complete activity. Synthetic dipalmitoyl or distearoyl phosphatidylcholine was ineffective in restoring the activity in this reconstituted system. (+info)
Selective promoting activity of phorbol myristate acetate in experimental skin carcinogenesis.
Experiments were undertaken to study the effect of promotion treatment on epidermal tumour induction pattern in precancerous mouse skin. Swiss albino mice were given a single s.c. injection of 0-5 mg 20-methylcholanthrene in the right scapular region. Six weeks later, 1-83 nmol of phorbol myristate acetate (PMA) was applied biweekly on the reactive skin. Histopathology of the induced tumours showed early appearance of squamous cell carcinomas and rhabdomyosarcomas. Fibrosarcoma, the most common tumour type induced on MCA injection alone, was absent. Trichoepithelioma, a benign tumour, was induced in PMA-treated mice. This gives new evidence of the selective action of PMA, enhancing the induction of epithelial and muscle tumours, with concurrent inhibition of fibroblastic tumours. (+info)
Demonstration of functional heterogeneity of hepatic uridine diphosphate glucuronosyltransferase activities after administration of 3-methylcholanthrene and phenobarbital to rats.
After the administration of 3-methylcholanthrene to adult male rats, activities of hepatic UDP-glucuronosyltransferase towards six from a group of 12 substrates were stimulated by 250-350%. Activities towards the remaining six substrates were unaffected. Conversely, after phenobarbital administration, activities formerly stimulated by 3-methylcholanthrene remained unchanged, and the other six activities were stimulated by 160-280%. The relationship of these two groups of transferase activities to other evidence suggesting the same heterogeneity of the enzyme is discussed. (+info)
The role of biotransformation in chemical-induced liver injury.
The role of drug metabolism in chemical-induced liver injury is reviewed. Parameters for studying the formation of chemically reactive metabolites are discussed and the factors that alter the formation and covalent binding of reactive metabolites are selectively emphasized. Some of the experimental work that led to these concepts is discussed, especially the chemical toxicology of the hepatic injury produced by acetaminophen, bromobenzene, furosemide, isoniazid and iproniazid. (+info)
Primary chemically induced tumors induce profound immunosuppression concomitant with apoptosis and alterations in signal transduction in T cells and NK cells.
Whereas transplantable tumors can be readily cured with immunotherapeutic approaches, similar therapies in cancer patients have been less effective. This difference may be explained by an immunosuppression resulting from the presence of a slowly growing primary tumor in the patient, whereas the immune system in a mouse with a rapidly proliferating transplantable tumor would be less affected. As a more appropriate model to the immune dysfunction in patients, slowly progressing primary tumors were induced by the carcinogen methylcholanthrene (MC) in mice. Their ability to induce immunosuppression in T cells and natural killer (NK) cells was compared to that of rapidly growing transplanted MC-induced tumors. The results demonstrate that mice bearing primary MC tumors had significantly diminished T-cell and NK-cell functions, impaired capacity to produce Th1 cytokines, and markedly reduced levels of the signal-transducing zeta chain in T cells and NK cells, similar to that described in cancer patients. Moreover, a substantial number of CD8+ T cells in mice with large primary MC tumors were undergoing apoptosis, correlating with alterations in CD4/CD8 ratios. In contrast, T cells and NK cells from mice bearing rapidly growing transplanted tumors were only marginally affected. These findings could explain the apparent discrepancy between the consistent findings of a diminished immune response and alterations in signal transduction in cancer patients as compared to the less reproducible observations in murine transplantable tumors. In addition, they could explain the differences in the high efficacy of immunotherapy in mice with transplantable tumors and the low therapeutic results in cancer patients. (+info)