ABSTRACT
1. Oxidative metabolism of the bladder carcinogens FANFT/ANFT was examined in vitro in guinea pig (resistant species) relative to rat (susceptible species). 2. The total rate of ANFT hepatic metabolism by guinea pig (soluble metabolites plus protein bound, 354 pmol/min per mg protein) was approx. 4 times that in rat. 3. The total rate of FANFT metabolism was similar in both species and approx. one-quarter that for ANFT in guinea pig. In rat, the rate of total metabolism of FANFT and ANFT was similar. 4. Cytochrome P450 inhibitors, 2,4-dichloro-6-phenylphenoxyethylamine, 7,8-benzoflavone, and n-octylamine largely inhibited metabolism in guinea pig, but had little effect in rat. 5. H.p.l.c. analysis of ANFT metabolites indicated distinctly different products in guinea pig compared to rat. 7,8-Benzoflavone decreased metabolite formation by 80% in guinea pig, but only 30% in rat. 6. Flavin-dependent monooxygenases may participate in metabolism of these carcinogens in rat, but not guinea pig. 7. Because ANFT is thought to be a more proximate carcinogen than FANFT, the increased rate of ANFT metabolism and the formation of different products in guinea pig compared to rat may partially explain the resistance of guinea pig to FANFT-induced bladder cancer.
Subject(s)
Carcinogens/metabolism , FANFT/analogs & derivatives , FANFT/metabolism , Liver/metabolism , NADP/metabolism , Animals , DNA/metabolism , Enzyme Inhibitors/pharmacology , Guinea Pigs , Kidney/metabolism , Liver/enzymology , Male , Microsomes, Liver/enzymology , Organ Specificity , Oxidation-Reduction , Protein Binding/drug effects , Rats , Rats, Inbred F344 , Urinary Bladder Neoplasms/chemically inducedABSTRACT
All-female hybrids of the killifishes Fundulus heteroclitus and Fundulus diaphanus, known from two sites in Nova Scotia, Canada, are shown to reproduce clonally. Isozyme analysis of crosses between female hybrids and male F. heteroclitus reveals that their progeny are genetically identical and show no evidence of recombination or paternal inheritance. Flow cytometric measurement of DNA content shows the hybrids to be diploid, with DNA values intermediate to those of the parental species. Because they are related to F. heteroclitus, a fish used widely as a model organism in experimental biology, the clonal hybrids are potentially valuable for experimental studies requiring subjects with a constant genetic background. In addition, the discovery of unisexuality and cloning in a fish whose reproductive physiology and development are so well characterized provides a unique opportunity to examine the underlying causes of clonal reproduction in vertebrates.
Subject(s)
Crosses, Genetic , Killifishes/genetics , Alleles , Animals , Female , Genetic Carrier Screening , Genetic Variation , Hybridization, Genetic , In Vitro Techniques , Isoenzymes/genetics , Isoenzymes/isolation & purificationABSTRACT
The metabolism and disposition of N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide (FANFT) and 2-amino-4-(5-nitro-2-furyl)thiazole (ANFT) were studied in rat and guinea pig. Rat is susceptible whereas guinea pig is resistant to FANFT-induced bladder cancer. Rats and guinea pigs were p.o. administered either 2-[14C]ANFT or 2-[14C]FANFT (100 mg/kg), and 18-h urine and feces were collected. Tissue distribution of radiolabel was determined. In both species, the highest concentrations of radioactivity expressed as nmol/g tissue were observed in the urine and intestines. Urinary metabolites were separated by high-performance liquid chromatography and radioactivity determined by radioanalytical detection. FANFT was not detected in urine from either species under any experimental condition. More ANFT was observed in urine following FANFT than ANFT administration. This deformylation-dependent excretion of FANFT was demonstrated in both species and has been previously described as renal metabolic/excretory coupling. Less ANFT, the carcinogen more proximate than FANFT, is excreted in guinea pigs compared with rats. A unique ANFT metabolite was identified in guinea pig but not rat urine. This metabolite represented 80 and 18% of radioactivity recovered in guinea pig urine following ANFT and FANFT administration, respectively. A metabolite produced by guinea pig liver and kidney microsomes in the presence of uridine-5'-diphosphoglucuronic acid coeluted with this unique metabolite. The urinary metabolite was characterized using hydrolytic enzymes, acid hydrolysis, and mass spectrometry and identified as an ANFT-N-glucuronide. A unique UDP-glucuronosyl-transferase appears to be responsible, at least in part, for the reduced amount of free ANFT excreted by guinea pigs compared with rats. Reduced levels of urinary ANFT observed in guinea pigs may partially explain the resistance of this species to FANFT-induced bladder cancer.
Subject(s)
Carcinogens/metabolism , FANFT/analogs & derivatives , FANFT/metabolism , Urinary Bladder Neoplasms/chemically induced , Animals , Carbon Radioisotopes , Disease Susceptibility , FANFT/pharmacokinetics , Glucuronosyltransferase/metabolism , Guinea Pigs , Male , Mass Spectrometry , Rats , Rats, Inbred F344 , Species Specificity , Tissue DistributionABSTRACT
An in vivo urinary metabolite of the bladder carcinogen 2-amino-4-(5-nitro-2-furyl) thiazole was isolated from guinea pig urine and was identified by direct analysis using thermospray mass spectrometry/high-performance liquid chromatography as 1-(2-amino-4-(5-nitro-2-furyl)-2-thiazolyl)-1-deoxy-beta-D-glucopyran uronic acid. The structure of this metabolite was also established by chemical synthesis. Both positive and negative ion thermospray mass spectrometry of the conjugate showed fragment ions resulting from cleavage across the pyran ring of the glucuronic acid comprising of aglycone moiety. These characteristic fragment ions may be diagnostic for identification of N-glucuronides from O-glucuronides.
