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Biomedical subjects

E Dybing

Publications and source records attributed to E Dybing.

At least 109 records · Page 6Linked to original sources
PubMed1983

Modulation of aromatic amine mutagenicity in Salmonella typhimurium with rat-liver 9000 g supernatant or monolayers of rat hepatocytes as an activation system.

2-Aminofluorene (AF), 2-acetylaminofluorene (AAF) and N-hydroxy-2-acetylaminofluorene (N-OH-AAF) were studied for mutagenic activity in S. typhimurium and either liver 9000 g supernatant fractions (S9) or monolayer cultures of hepatocytes isolated from Wistar rats were used as an activation system. All 3 compounds were converted into mutagens excreted into the incubation medium by the cell-culture system, with N-OH-AAF greater than AF greater than AAF. Cultures used 24 h after plating were less efficient in promutagen conversion than were cultures used after 2 h. Phenobarbital, but not 3-methylcholanthrene, pretreatment of the rats caused similar effects on AF, AAF and N-OH-AAF mutagenicity with both S9 and hepatocyte cultures. The mutagenicities of AF and AAF were reduced by the cytochrome-P-450 inhibitors metyrapone and alpha-naphthoflavone, whereas the mutagenicity of N-OH-AAF was increased by using both inhibitors. Further, the microsomal deacetylase inhibitor paraoxon caused only a moderate reduction in N-OH-AAF mutagenicity, but a total inhibition of AAF mutagenicity. No significant effect of paraoxon on AF mutagenicity was seen. With the S9 system, no effect of ascorbate on the mutagenicity of AF, AAF or N-OH-AAF was observed. In contrast, the mutagenicity of all 3 compounds was increased by ascorbate when hepatocyte cultures were used as activation system. Incubation of hepatocyte monolayers in a sulfate-free medium did not change the mutagenicity of AF, AAF or N-OH-AAF. Galactosamine, an inhibitor of glucuronidation in cells, increased the mutagenicity of AF, AAF and N-OH-AAF with hepatocyte cultures. The addition of cofactor for glucuronidation in the S9 system, however, had no effect. A reduction in mutagenicity of AF and AAF, but not that of N-OH-AAF, was observed with the addition of glutathione (GSH) in both the S9 and the hepatocyte systems. On the other hand, no effect of cellular GSH depletion was seen on aromatic-amine mutagenicity in the hepatocyte system. The data indicate that the hepatocyte culture system offers advantages over the conventional liver-sub-fraction activation system as a model, in vivo, for the metabolism of the aromatic amine mutagens/carcinogens.

2-Acetylaminofluorene↗
PubMed1983

Comparative genotoxicity studies of the flame retardant tris(2,3-dibromopropyl)phosphate and possible metabolites.

Tris(2,3-dibromopropyl)phosphate (Tris-BP) was activated to mutagens in the Salmonella/microsome quantitative test system. Liver microsomes from rats pretreated with phenobarbital (PB) increased the mutagenicity of 0.05 mM Tris-BP to 186% of the activity obtained with liver microsomes from untreated rats. The addition of 0.02 mM Tris-BP to V79 Chinese hamster cells co-incubated with liver microsomes from PB-pretreated rats increased the number of mutants by a factor of 9.7. Tris-BP also caused genotoxic and cytotoxic responses in primary monolayers of rat hepatocytes. The relative increase in unscheduled DNA synthesis after treatment with 0.05 mM Tris-BP was 2.3-fold as measured by scintillation counting of radiolabelled thymidine incorporated into DNA of isolated nuclei. The use of hepatocytes isolated from PB-pretreated rats reduced the increases in DNA repair synthesis relatively to that in control cells. Monolayers of hepatocytes from untreated rats co-cultured with Salmonella typhimurium TA100 activated Tris-BP to mutagenic intermediates which were released into the culture medium. The studies with the V79 and liver-cell systems indicate that the reactive intermediates formed from Tris-BP are sufficiently stable and lipophilic to traverse the various membranes from the site of generation to the respective cellular targets. The relative degree of genotoxic responses of bis(2,3-dibromopropyl)phosphate, 2,3-dibromopropylphosphate, tris(2,3-bromopropyl)phosphate, tris(2-bromopropyl)phosphate and 2,3-dibromopropanol in the systems studied did not indicate that these compounds were proximate or ultimate reactive metabolites of Tris-BP in liver-derived activation systems.

Animals↗
PubMed1983

Drug metabolism activities of isolated rat hepatocytes in monolayer culture.

The levels of cytochrome P-450 in hepatocytes cultured as monolayers for 22 hrs in Dulbecco's modified Eagle medium supplemented with serum and insulin was reduced to approximately 40% of initial values of freshly isolated hepatocytes. In correspondence with this the activities of the cytochrome P-450 monooxygenases aryl hydrocarbon (benzo(a)pyrene) hydroxylase (AHH) and ethylmorphine (EM) N-demethylase were reduced to 40 and 22% of their initial activities, respectively. Modifying the culture medium through omission of cysteine and cystine, and adding dexamethazone and delta-amino levulinic acid, increased the content of cytochrome P-450 to 59% and EM N-demethylase to 46% of initial values, but was without effect on AHH activity. However, further modifications by adding high concentrations of asparagine and leucine increased AHH activity to 62% of initial values, but did not further enhance the total content of cytochrome P-450 or the EM N-demethylase activity. The activities of cytochrome P-450 reductase, flavin containing monooxygenase, epoxide hydrolase and glutathione S-transferase decreased less (to about 70-80% of initial values) than cytochrome P-450 associated monooxygenase activities, whereas UDP-glucuronyl transferase decreased to about 50% of initial values. In contrast to what was observed regarding cytochrome P-450 and associated monooxygenase activities, modification of the incubation conditions did not affect the non-cytochrome P-450 enzymatic activities.

Animals↗
PubMed1983

No effect of prolonged fluoride exposure on cytochrome P-450 and associated monooxygenases or on the level of polyamines in the rat.

When exposing rats to drinking water containing 100 p.p.m. fluoride for 8 weeks, no effect could be detected in biochemical parameters of the liver, such as the concentrations of the polyamines putrescine, spermidine and spermine; the levels of microsomal protein and cytochrome P-450; or the activities of two associated monooxygenases, aryl hydrocarbon hydroxylase and ethylmorphine N-demethylase. Neither was there any increase in plasma glutamic-oxalacetic transaminase indicative of liver damage.

Animals↗
PubMed1983

Increased cytochrome P-450 independent drug metabolism and mutagen activation in rat liver by octachlorostyrene.

Single intraperitoneal injections of 200 mg/kg octachlorostyrene (OCS) increased the activities of flavin-containing monooxygenase, epoxide hydrolase and glutathione S-transferase in the livers of male Wistar rats. UDP-glucuronyl transferase activities measured with aglycones increased by methylcholanthrene or phenobarbital treatment, were both slightly increased by OCS treatment. A liver 9,000 X g supernatant fraction from OCS pretreated rats increased the bacterial mutagenicity of 2-acetylaminofluorene and 2-aminofluorene compared to controls, while insignificant or only minor effects were seen on N-hydroxy 2-acetylaminofluorene and benzo(a)pyrene mutagenicity. The effect of OCS on mutagen activation was similar to that seen after phenobarbital treatment. The use of monolayers of hepatocytes instead of 9,000 X g subfractions did not reveal any qualitative differences in mutagen activation.

Animals↗
PubMed1982

Induction of liver microsomal cytochrome P-450 and associated monooxygenases by octachlorostyrene in inbred strains of mice. Lack of correlation with the murine Ah locus.

Single intraperitoneal injections of octachlorostyrene (OCS) and hexachlorobenzene in genetically polycyclic aromatic hydrocarbon "responsive' C57/BL/6 (B6) mice led to a time- and dose-dependent increase in the levels of liver microsomal cytochromes P-450 and b5 as well as in the activities of NADPH cytochrome P-450 (cytochrome c) reductase, ethylmorphine (EM) N-demethylase, 4-nitroanisole (PNA) O-demethylase and acetanilide 4-hydroxylase (AcA hydroxylase). No, or only a very moderate, increase in the activity of aryl hydrocarbon hydroxylase was seen after OCS and HCB, respectively. Pretreatments with phenobarbital (PB) or 3-methylcholanthrene (MC) both increased AcA hydroxylase activity to a similar degree, whereas pretreatment with polychlorinated biphenyls (Aroclor 1254) had an effect equal to the sum of PB and MC. Judged from sodium dodecylsulfate polyacrylamide gel electrophoresis studies, OCS and HCB predominantly increased a microsomal polypeptide of apparent mol. wt 52,000, similar to PB. A reduced response was seen after OCS or HCB treatment of aromatic hydrocarbon "non-responsive' DBA/2 (D2) mice compared to B6 mice, both with respect to AcA hydroxylase as well as EM demethylase and PNA demethylase activities. OCS treatment of B6D2F1 mice resulted in a doubling of AcA hydroxylase activity, but in mice of the (B6D2)D2 backcross no distinct subgrouping of individual AcA hydroxylase activities were apparent. These results demonstrate that OCS is an inducer of the PB-type in mice and that induction of AcA hydroxylase by OCS is not regulated by the Ah locus.

Animals↗
PubMed1982

Induction of liver microsomal cytochrome P-450 and associated monooxygenases by octachlorostyrene in the rat.

The inducing effects of octachlorostyrene (OCS) and hexachlorobenzene (HCB), chlorinated hydrocarbon contaminants found in fish, on the microsomal cytochrome p-450 system were studied in rats. Administration of single doses of OCS and HCB at 50mg/kg intraperitoneally and higher led to increases in microsomal protein an cytochrome p-450 content, cytochrome p-450 reductase, ethylmorphine N-demethylase, 4-nitroanisole O-demethylase, acetanilide 4-hydroxylase, but not in benzo(a)pyrene hydroxylase activities. Similar patterns of induction were seen after 14 days repeated intraperitoneal or oral dosing. Siodium dodecyl sulfate polyacrylamide gel electrophoresis of microsomes from OCS and HCB treated rats showed increases in proteins similar to those found after phenobarbital (PB) induction. Under the present experimental conditions OCS and HCB were found to be PB-type inducers.

Animals↗
PubMed1982

Cytotoxic effects of N-hydroxyparacetamol in suspsensions of isolated rat hepatocytes.

The cytotoxicity of N-hydroxyparacetamol (N-OH-pHAA), a postulated proximate metabolite of the hepatotoxic and nephrotoxic analgesic paracetamol, was studied in suspensions of hepatocytes isolated by collagen-perfusion of livers of male rats. Incubation of cells with 0.25-2.0 mM N-OH-pHAA led after 3-5 hours to increased cell permeability measured by increased trypan blue uptake, increased NADH penetration or leakage of prelabelled 51Cr. N-OH-pHAA rapidly depleted cellular glutathione, 16% of initial levels were seen after 30 min. incubation. 3H-N-OH-pHAA bound covalently to cellular proteins in a time- and concentration-dependent manner, considerably higher binding rates were seen with boiled cells compared to intact cells. Pretreatment of animals with the cytochrome P-450 inducer phenobarbital did not affect N-OH-pHAA cytotoxicity or covalent binding, whereas the cytochrome P-450 inhibitor metyrapone inhibited both cytotoxicity and binding. Lipid peroxidation in hepatocytes could be seen as a limited range of N-OH-pHAA concentrations. In contrast, lipid peroxidation was an early event in cells exposed to carbon tetrachloride. A minimal exposure time of 30 min. of the hepatocytes to N-OH-pHAA was sufficient to elicit cellular damage occurring after 3-5 hours.

Acetaminophen↗
PubMed1982

Modulation of N-hydroxyparacetamol cytotoxicity in suspensions of isolated rat hepatocytes.

The effect of inhibitors of toxicity of N-hydroxyparacetamol (N-OH-pHAA), a postulated proximate metabolite of paracetamol, was studied in isolated rat hepatocytes. Additions of ascorbate, menadione, thiol-containing amino acids and glutathione (GSH) led to an increased stability of N-OH-pHAA, reduced the covalent binding of N-OH-pHAA to cellular protein and decreased GSH depletion caused by N-OH-pHAA. Two to three hours elapsed after a 30 min. exposure of the cells to N-OH-pHAA before the cells responded with increased cell permeability. Ascorbate, acetylcysteine, GSH and promethazine were capable of inhibiting this second phase of N-OH-pHAA cytotoxicity in addition to their effects during the initial exposure phase. In contrast, the anti-oxidant tocopherole and phenacetin wee only effective during the second phase. Increasing the incubation medium pH during the second phase of N-OH-pHAA mediated cellular damage resulted in decreases in cytotoxicity. Lipid peroxidation, as measured by accumulation of thiobarbituric acid reactive metabolites, did not seem to be directly correlated with cytotoxicity, since cysteamine or higher concentrations of N-OH-pHAA inhibited lipid peroxidation without decreasing cellular damage.

Acetaminophen↗
PubMed1981

In vitro and in vivo covalent binding of the kidney toxicant and carcinogen tris(2,3-dibromopropyl)-phosphate.

The nephrotoxicant and nephrocarcinogen tris(2,3-dibromopropyl)-phosphate (Tris-BP) is activated to products which bind covalently to microsomal protein by a cytochrome P-450 dependent oxidation reaction. Binding to rat liver microsomes proceeds 15 times faster than with kidney microsomes. The binding in liver microsomes is markedly increased by phenobarbital pretreatment, the apparent Vmax of the reaction is 175 pmol/mg microsomal protein/min with control microsomes and 1053 pmol/mg protein/min with induced microsomes. Binding with kidney microsomes is doubled after pretreatment with polychlorinated biphenyls. 2,3-Dibromopropanol (2,3-DBP), a hydrolysis product of Tris-BP, is also activated to covalently protein-bound products, but at a much slower rate than Tris-BP. Administration of Tris-BP to rats leads to its covalent binding to proteins in liver and kidney, with 5 time higher binding levels in kidney than in liver, correlating with its relative organotoxic potential in single dose experiments. Binding to proteins in the kidney was increased by pretreatment of animals with polychlorinated biphenyls. A covalent interaction of Tris-BP could also be demonstrated to DNA, both when DNA was added to liver microsomal incubations in vitro and to DNA extracted from liver and kidney after administration of Tris-BP in vivo. The binding levels were 4 times higher to kidney DNA than to liver DNA.

Animals↗
PubMed1981

Genetic differences in dimethylnitrosamine mutagenicity in vitro associated with mouse hepatic aryl hydrocarbon hydroxylase activity induced by 3-methylcholanthrene.

The effects of 3-methylcholanthrene (MC) pretreatment on metabolism and mutagenic activation of dimethylnitrosamine (DMN) were studied with liver subfractions from two strains of mice differing genetically with respect to aromatic hydrocarbon responsiveness. Both mutagenic activation and DMN N-demethylase activity segregated with aryl hydrocarbon (benzo[a]pyrene) hydroxylase activity as a dominant trait in appropriate crosses between C57BL/6J (Ahb Ahb) and DBA/2J (Ahd Ahd) mice. DMN metabolism and mutagenicity were increased by MC-pretreatment in responsive Ahb Ahb and Ahb Ahd mice, but not in non-responsive Ahd Ahd mice. This indicates the involvement of the Ah locus in the genetic regulation of these activities in mice. Deuteration of DMN reduced mutagenicity and DMN N-demethylase activity by approximately 90 and 50 percent, respectively.

Animals↗
PubMed1981

Studies on mutagenic and carcinogenic N-substituted aryl compounds: cosmetics and drugs.

We aimed our studies toward gaining an understanding of some of the reactions and pathways involved in the metabolism and activation of the aromatic diamines used in hair dyes and of phenacetin used in analgesic mixtures. Comparison of the data obtained from human and animal tissues established that animal tissues can serve as suitable models for evaluation of the activity of these compounds in humans.

Adult↗
PubMed1980

Nephrotoxicity of the flame retardant tris(2,3-dibromopropyl)phosphate.

The flame retardant tris(2,3-dibromopropyl)phosphate (Tris-BP) is known to be activated to a potent mutagen in the Salmonella test system and to induce kidney tumors in long-term feeding studies in mice and rats. Administration of Tris-BP to rats leads to extensive tubular necrosis at doses of 250 mg/kg i.p. and higher. The histological lesion is present in most animals 24 h after administration. There is a close correlation between the increase in kidney weights, the degree of kidney damage and the increase in plasma urea levels. A continuous increase in kidney weights with respect to time is seen, 7 days after a dose of 250 mg/kg i.p. the kidney/body weight ratio is 192% of controls. The kidney damage is not altered by previous phenobarbital-treatment, whereas cobaltous chloride, an inhibitor of cytochrome P-450, slightly reduces the kidney damage.

Animals↗
PubMed1980

The role of ethyl and fluorine substitution in the 4'-position for N,N-diethyl-4-aminoazobenzene mutagenicity and azo reduction.

The mutagenicity and azo reduction rate of N,N-diethyl-4-aminoazobenzene (DEAB) were influenced by substitution in the 4'-position with a ethyl or a fluorine group. The parent dye (DEAB) was shown to be slightly mutagenic with Salmonella typhimurium TA98 using Aroclor 1254-pretreated 9000 x g supernatant fractions from rat liver. Introduction of a 4'-ethyl group in DEAB did not affect mutagenicity of the dye, but a 4-fluoro group markedly enhanced its mutagenicity. DEAB underwent azo reduction and its reduction rate was influenced by 4'-substituents. A 4'-fluoro group in DEAB increased its azo reduction rate, while a 4'-ethyl group abolished it. Inhibitors of cytochrome P-450 also inhibited 4'-fluoro-DEAB mutagenicity and azo reduction.

Animals↗