Tuimorigenic activity of fluoranthene, 2-methylfluoranthene and 3-methylfluoranthene in newborn CD-I mice

Fluoranthene (FA) is frequently among the more abundant componentsdetected in environmental mixtures of polycyclic aromatic hydrocarbons.Several methylated fluoranthenes, although less prevalent thanFA, have also been detected as environmental pollutants. WhileFA is inactive as a tumorigenic agent on mouse skin, it doesinduce lung and liver tumors in newborn mice. Among the fiveisomers of methylfluoranthene, only 2-methylfluoranthene (2-MeFA)and 3-methylfluoranthene (3-MeFA) are active as tumor initiatorson mouse skin. A comparative bioassay was performed to determinethe relative tumorigenic activity of FA, 2-MeFA and 3-MeFA innewborn CD-1 mice. All three compounds were assayed at dosesof 3.46 and 17.3 µmol. The bioassay was terminated whenmice were 1 year old. At a dose of 17.3 µmol, FA and 2-MeFAinduced a similar incidence of lung tumors (65–96%) inboth male and female mice. However, tumor multiplicity in thelung was different between FA and 2-MeFA. At a dose of 17.3µmol, the multiplicity of lung tumors observed for miceadministered 2-MeFA ranged from 3.04 to 3.94 tumors per mouse.In contrast, animals treated with FA developed only an averageof 1.12–2.45 tumors per mouse. 3-MeFA did not induce astatistically significant incidence of lung tumors in eithermale or female mice. All three compounds when administered tonewborn mice did induce a significant incidence of liver tumorsamong male mice. The relative tumorigenic potency observed wasFA 5     

The role of acetylation in the mutagenicity of the antitumor agent, batracylin

The role of acetylation in the genotoxicity of the heterocyclicamine, batracylin, was evaluated in Salmonella typhimurium strainsexpressing various levels of N-and O-acetyltransferase activity.A significant correlation was observed between batracylin-inducedmutagenicity and bacterial N-acetyltransferase activity. Strainswith the greatest capacity for N-acetylating batracylin (YG1012 and YG 1024) were the most sensitive to the mutagenic effectsof the drug. The number of revertants/nmol batracylin and theformation of acetylbatracylin were     

5-Methylchrysene metabolism in mouse epidermis in vivo, diol epoxide--DNA adduct persistence, and diol epoxide reactivity with DNA as potential factors influencing the predominance of 5-methylchrysene-1,2-diol-3,4-epoxide--DNA adducts in mouse epidermis

5-Methylcbrysene (5-MeC) can form two bay region dihydrodiolepoxides: 1,2-dihydroxy-3-4-epoxy-1,2,3,4-tetrahydro-5-methylchrysene(DE-I) which has the methyl group and the epoxide ring in thesame bay region, and 7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydro-5-methylchrysene(DE-II). In a previous study, we observed that the ratio ofDE-I:DNA adduds to DE-II:DNA adducts in mouse epidermis, 24h after application of [³H]5-MeC metabolites was 2.7 to 1. Toinvestigate the basis for this observation we have now studied:(i) the formation of [³H]5-MeC in mouse epidermis in vivo atvarious time intervals from 0.33 to 24 h; (ii) the persistenceof DE-I:DNA adducts and DE-II:DNA adducts in mouse epidermisat 4–48 h after application of [³H]5-MeC and (iii) thereactions of DE-I and DE-II with calf thymus DNA in vitro. Incontrast to results obtained with mouse liver 9000 g supernatant,the dihydrodiol precursors of DE-I and DE-II were present inequivalent quantities in mouse epidermis in vivo at every timepoint studied. The ratio of DE-I:DNA adducts to DE-II:DNA adductsin mouse epidermis was constant throughout the time period studied.However, the extent of formation of DE-I:DNA adducts was greaterthan that of DE-II:DNA adducts upon reaction of DE-I or DE-IIwith calf thymus DNA in vitro. These results suggest that differencesin reactivity with DNA of DE-I and DE-II may be responsiblefor the higher levels in mouse epidermis of DE-I:DNA adductscompared with DE-II:DNA adducts and provide a possible basisfor the observed enhancing effect of a bay region methyl groupon the carcinogenicity of polynuclear aromatic hydrocarbons.     

The effects of antioxidant vitamin supplementation on resistance exercise induced lipid peroxidation in trained and untrained participants

Background  

The theoretical benefits of using antioxidant vitamin supplements to quench oxygen free radicals appear large. High intensity aerobic-type exercise produces oxygen free radicals that can cause damage to lipid membranes (lipid peroxidation) that may lead to many problems such as the inactivation of cell membrane enzymes, the progression of degenerative diseases (cardiovascular disease and cancer) and lessening of the effectiveness of the immune system. The major function of vitamin E is to work as a chain-breaking antioxidant in a fat soluble environment. Little research has examined lipid peroxidation associated with high intensity resistance exercise or possible protective effects of antioxidant supplementation or the effects of training state.     

Microglial activation precedes dopamine terminal pathology in methamphetamine-induced neurotoxicity

Previous studies have demonstrated methamphetamine (METH)-induced toxicity to dopaminergic and serotonergic axons in rat striatum. Although several studies have identified the nature of reactive astrogliosis in this lesion model, the response of microglia has not been examined in detail. In this investigation, we characterized the temporal relationship of reactive microgliosis to neuropathological alterations of dopaminergic axons in striatum following exposure to methamphetamine. Adult male Sprague-Dawley rats were administered a neurotoxic regimen of methamphetamine and survived 12 h, or 1, 2, 4, and 6 days after treatment. Immunohistochemical methods were used to evaluate reactive changes in microglia throughout the brain of methamphetamine-treated rats, with a particular focus upon striatum. Pronounced morphological changes, indicative of reactive microgliosis, were evident in the brains of all methamphetamine-treated animals and were absent in saline-treated control animals. These included hyperplastic changes in cell morphology that substantially increased the size and staining intensity of reactive microglia. Quantitative analysis of reactive microglial changes in striatum demonstrated that these changes were most robust within the ventrolateral region and were maximal 2 days after methamphetamine administration. Analysis of tissue also revealed that microglial activation preceded the appearance of pathological changes in striatal dopamine fibers. Reactive microgliosis was also observed in extra-striatal regions (somatosensory and piriform cortices, and periaqueductal gray). These data demonstrate a consistent, robust, and selective activation of microglia in response to methamphetamine administration that, at least in striatum, precedes the appearance of morphological indicators of axon pathology. These observations raise the possibility that activated microglia may contribute to methamphetamine-induced neurotoxicity.     

Functional gamma-secretase complex assembly in Golgi/trans-Golgi network: interactions among presenilin, nicastrin, Aph1, Pen-2, and gamma-secretase substrates

Gamma-secretase is a proteolytic complex whose substrates include Notch, beta-amyloid precursor protein (APP), and several other type I transmembrane proteins. Presenilin (PS) and nicastrin are known components of this high-molecular-weight complex, and recent genetic screens in invertebrates have identified two additional gene products, Aph1 and Pen-2, as key factors in gamma-secretase activity. Here, we examined the interaction of the components of the gamma-secretase complex in Chinese hamster ovary cells stably expressing human forms of APP, PS1, Aph1, and Pen-2. Subcellular fractionation of membrane vesicles and subsequent coimmunoprecipitation of individual gamma-secretase components revealed that interactions among all proteins occurred in the Golgi/trans-Golgi network (TGN) compartments. Furthermore, incubation of the Golgi/TGN-enriched vesicles resulted in de novo generation of amyloid beta-protein and APP intracellular domain. Immunofluorescent staining of the individual gamma-secretase components supported our biochemical evidence that the gamma-secretase components assemble into the proteolytically active gamma-secretase complex in the Golgi/TGN compartment.     

Fluorine probes for investigating the mechanism of activation of indeno[1,2,3-cd]pyrene to a tumorigenic agent

Indeno[1,2,3-cd]pyrene (IP) is a non-alternant polycyclic aromatic hydrocarbon that has tumor-initiating activity on mouse skin and is carcinogenic in newborn mice and in rat lungs. Previous studies have shown that 8- and 9-hydroxyIP and IP-1,2-diol are major metabolites formed in vivo in mouse skin. 8-HydroxyIP-1,2-diol and 9-hydroxyIP-1,2-diol are also observed as in vivo metabolites of IP. Although 8-hydroxyIP had marginal tumor-initiating activity on mouse skin, IP-1,2-diol and its epoxide precursor, IP-1,2-oxide, had similar tumorigenic activity as IP. In the present study fluorine probes have been employed to investigate the contribution of metabolic activation at the 1,2 and 7-10 positions of IP. At a total initiating dose of 4.0 mumol, 2-fluoroIP induced skin tumors in 76% of the treated animals with an average of 3.9 tumors/mouse. At the same dose, IP induced a 72% incidence of tumor-bearing mice with 2.1 tumors/mouse. In contrast, 8,9-difluoroIP elicited a tumorigenic response in 40% of the treated animals with 0.6 tumors/animal. Five mice from each experimental group were killed at the conclusion of the initiation phase of the bioassay and DNA was isolated from the treated areas of skin. 32P-Postlabeling analysis of the hydrolyzed DNA indicated that IP forms one major detectable DNA adduct that migrates close to the origin. This adduct is absent in mice treated with 8,9-difluoroIP. In contrast, 2-fluoroIP forms one major adduct spot with different retention behavior as compared with the adduct formed from IP. DNA from mice treated topically with IP-1,2-diol and IP-1,2-oxide was subjected to 32P-postlabeling analysis. IP-1,2-diol forms one major DNA adduct spot with mobility similar to that observed for the IP-DNA adduct. IP-1,2-oxide displayed an intense pattern of DNA adducts centered around the location of the IP-DNA adduct. No adducts were detected which had mobility similar to that formed from 2-fluoroIP. These results are consistent with IP undergoing metabolic activation at positions 7-10 either alone or in conjunction with dihydrodiol formation at the 1,2 position.     

The carcinogenicity of quinoline and benzoquinolines in newborn CD-1 mice

The environmental occurrence and mutagenic activity of quinoline and benzoquinolines are well-documented. In this study, the relative carcinogenic activities of quinoline, benzo[f]quinoline, benzo[h]quinoline, and phenanthridine were evaluated in newborn mice. Mice were injected intraperitoneally on the first, eighth, and fifteenth day of life with 0.25, 0.5, and 1.0 mumol of each of these aza-arenes. Quinoline induced a 71% incidence (P less than 0.005) of hepatic tumors among the male mice sacrificed at 52 weeks of age. None of the female mice treated with these aza-arenes developed hepatomas. Among the female mice treated with quinoline there was a significant development of leukemia or lymphoma (P less than 0.05) which was not evident among the female mice in any of the other experimental groups. Benzo[h]quinoline and phenanthridine were not carcinogenic under these assay conditions. Benzo[f]quinoline did induce an increase in the incidence of hepatomas among male mice (19% as compared to 5.9% among controls). This increase, however, was not statistically significant. These data indicate that quinoline has greater carcinogenic potential than any of these isomeric benzoquinolines in newborn mice.     

Identification of the mutagenic metabolites of fluoranthene, 2-methylfluoranthene, and 3-methylfluoranthene

The metabolites of fluoranthene, 2-methylfluoranthene, and 3-methylfluorantheneobtained upon incubation with liver homogenate from Aroclorpretreated rats were assayed for mutagenicity in Salmonellatyphimurium TA100. The mutagenic metabolites of fluorantheneand 2-methylfluoranthene were identified as 2,3-dihydro-2,3-dihydroxyfluorantheneand 4,5-dihydro-4,5-dihydroxy-2-methylfluoranthene, respectively.In contrast to these results, the major proximate mutagen detectedamong the in vitro metabolites of 3-methylfluoranthene was 3-hydroxymethylfluoranthene.Comparison of the mutagenic potential of 2-hydroxymethylfluor-anthenedemonstrated that the latter was a more powerful mutagen. Quantitativeanalyses of the metabolites of fluoranthene with that of 3-hydroxymethylfluoranthenedemonstrated that the latter was a more powerful mutagen. Quantitativeanalyses of the metabolites of fluoranthene, 2-methylfluoranthene,and 3-methylfluoranthene indicated that similar amounts of dihydrodiolswere formed. 4,5-Dihydro-4,5-dihydoxy-3-methylfluoranthene,however, was not found to be a potent mutagenic metabolite.These data suggest that the activation pathway to ultimate mutagensmay differ for 2- and 3-methylfluoranthene.     

Identification of metabolites of benzo[b]fluoranthene

The metabolism by rat liver 9000 x g supernatant of the environmentalcarcinogen benzo[b]fluoranthene was investigated. The majormetabolites were identified, by comparison to synthetic samples,as 5- and 6-hydroxybenzo[b]-fluoranthene and 4- or 7-hydroxybenzo[b]fluoranthene.The principal dihydrodiol metabolite formed under these conditionswas trans-11,12-dihydro-11,12-dihydroxybenzo[b]-fluoranthene,which was identified by comparison to the synthetic compound.1,2-Dihydro-1,2-dihydroxybenzo[b]fluoranthene was identified,by its mass spectrum and by comparison of its u.v. spectrumto that of a synthetic model compound, 11-ethylidene-11H-benzo[b]fluorene.No evidence was obtained for the formation of 7b,8-dihydro-7b,8-di-hydroxybenzo[b]fluorantheneor trans-9, 10-dihydro-9,10-dihydroxybenzo[b]fluoranthene. Thelatter would be the precursor to a bay region dihydrodiol epoxideof benzo[b]-fluoranthene.