Six-Month Exposure of Strain A/J Mice to Cigarette Sidestream Smoke: Cell Kinetics and Lung Tumor Data

Male strain A/J mice were exposed to sidestream smoke (SS) generatedfrom burning Kentucky 1R4F reference cigarettes. Chamber concentrationswere 4 mg/m³ of total suspended respirable particulate matter(TSP). Animals were exposed 6 hr a day, 5 days a week. One-weekcumulative labeling indices were significantly increased inthe large intrapulmonary airways during the 1st week and inthe respiratory epithelium of the nasal and maxillar turbinatesduring the first 3 weeks of exposure and then returned to controlvalues. Subsequently, signs of increased cell proliferationwere again found in the nasal and maxillar turbinates duringthe 9th and 16th exposure weeks. The experiment was terminatedafter 6 months. The number of animals bearing lung tumors wasthe same in smoke-exposed as in filtered airexposed animalsas was the average number of tumors per lung. Analysis of theDNA of individual tumors obtained from exposed and control micefor K-ras mutations suggested that exon 2 might be a specifictarget for SS. It was concluded that (1) duration of exposurewas too short or (2) concentration of TSP was too low to reveala possible carcinogenic potential of SS in strain A/J mice orthat (3) SS is not carcinogenic in strain A mice.     

Histological Alterations in Male A/J Mice Following Nose-Only Exposure to Tobacco Smoke

The incidence and multiplicity of grossly observed and microscopic lesions of the respiratory tract of A/J mice exposed nose-only to mainstream smoke (50, 200, or 400 mg total particulate matter/m³ from 2R4F cigarettes) was compared to those of filtered air controls. Animals were necropsied at the end of exposure (5 mo) or following 4 or 7 mo of recovery. Lungs were visually inspected for tumors at all necropsies and examined histopathologically at 9 and 12 mo. At 5 mo no tumors were recorded. No significant elevations in tumor incidence or multiplicity were recorded although at 9 mo multiplicity was elevated in the mid-exposure group (0.90 versus 0.55 tumors per animal for controls). At 12 mo, multiplicity was increased over the 9-mo necropsy at all exposures except 200 mg/m³; however, there were no dose-related trends in multiplicity or incidence. Histopathological alterations included hyperplasia, metaplasia, and inflammation of the nose and larynx and proliferative lesions of the lungs. At 9 mo, the multiplicity of focal lung lesions was 1.4 per animal in controls but averaged 1.0 among smoke-exposed groups. There was an inverse relation (p < .059) between smoke concentration and the percentage of hyperplastic lesions at 9 mo. At 12 mo the high-exposure group had slightly increased multiplicity of 2.3 lesions compared with 1.6 among controls, while the percentage of hyperplasic lesions was similar between groups. Nose-only inhalation of mainstream tobacco smoke resulted in chronic inflammatory changes of the respiratory tract yet failed to produce statistically significant changes in tumor incidence or multiplicity.     

Effect of Cigarette Smoke on UDP-Glucuronosyltransferase Activity and Cytochrome P450 Content in Liver,Lung and Kidney Microsomes in Mice

Abstract: The effect of cigarette smoke on the expression of several cytochromes P450 (CYP) and UDP-glucuronosyltransferases (UGT) was studied in mice. The animals were exposed to cigarette smoke for 4 to 30 days. Enzymatic activities supported by CYP1A1, 1A2, 2B, 2E1 and the glucuronidation activity toward phenols were measured in lung, liver and kidney microsomes. Cigarette smoke induced several CYPs, especially in lung. CYP2E1 was more induced than CYP1A1 in this organ. The expression of CYP2E1 was also increased in kidney (5.6 times after 30 days). The glucuronidation in kidney was non-sensitive to the treatment whatever substrate used. In contrast, this activity was enhanced in liver and particularly in lung, in which the glucuronidation of 1-naphthol and 2-hydroxybiphenyl was increased by 122 and 180%, respectively. Interestingly, the times of induction differed according to the substrate used, thus suggesting the presence of different UGTs active toward phenols that were differentially affected by cigarette smoke. The LIGT activities toward phenols were low in lung, when compared with those measured in liver or kidney. In conclusion, cigarette smoke greatly affected both glucuronidation activity and the hydroxylation reactions supported by CYPs in mouse liver and lung.     

Exposure of Human Lung Cells to Inhalable Substances: A Novel Test Strategy Involving Clean Air Exposure Periods Using Whole Diluted Cigarette Mainstream Smoke

An experimental approach was established for the validation of an in vitro test system for complex environmental test atmospheres consisting of both gaseous substances and particulates. Smoke from two different cigarette types (generated by an automatic cigarette-smoking machine) was employed to assess both the sensitivity and the specificity of the system. The smoke was diluted with synthetic air and used to expose human lung cells grown on microporous membranes. Cells were exposed alternately to diluted cigarette smoke and pure synthetic air. The effect of diluted smoke was assessed without humidification, addition of CO 2, or any other physical or chemical modification of the smoke. The experimental setup included online monitoring of the gas phase (by analysis of CO concentration) and particulate phase (by light-scattering photometry). Replicate experiments confirmed a reproducible generation and dilution of the smoke and a smoke age of about 7 s at the time it came into contact with the cells. Experiments using human lung cells revealed that smoke from the two different cigarette types induced different levels of dose-dependent toxicity. A cell exposure of 6 min using 6 alternating smoke and synthetic air periods was sufficient to cause different effects as measured by intracellular glutathione content. The fact that the system could differentiate between two different types of cigarette smoke demonstrated its high sensitivity and specificity. The system offers new ways to test native complex gaseous and aerosol mixtures in vitro using short exposure times and very small amounts of test substances.     

Cytochrome P-450 enzymes and FMO3 contribute to the disposition of the antipsychotic drug perazine in vitro

RATIONALE: Perazine (PER) is a phenothiazine antipsychotic drug frequently used in Germany that undergoes extensive metabolism. OBJECTIVES AND METHODS: To anticipate metabolic drug interactions and to explore the relevance of polymorphisms of metabolic enzymes, perazine-N-demethylation and perazine-N-oxidation were investigated in vitro using human liver microsomes and cDNA expressed enzymes. RESULTS: CYP3A4 and CYP2C9 were identified as the major enzymes mediating PER-N-demethylation. At 10 microM PER, a concentration consistent with anticipated in vivo liver concentrations, CYP3A4 and CYP2C9 contributed 50% and 35%, respectively, to PER-N-demethylation. With increasing PER concentrations, contribution of CYP2C9 decreased and CYP3A4 became more important. In human liver microsomes, PER-N-demethylation was inhibited by ketoconazole (>40%) and sulfaphenazole (16%). Allelic variants of recombinant CYP2C9 showed differences in PER-N-demethylase activity. The wild type allele CYP2C9*1 was the most active variant. Maximal activities of CYP2C9*2 and CYP2C9*3 were 88% and 18%, respectively, compared to the wild type activity. Perazine-N-oxidation was mainly mediated by FMO3. In the absence of NADPH, heat treatment of microsomes abolished PER-N-oxidase activity. Methimazole inhibited PER-N-oxidation, while CYP specific inhibitors had no inhibitory effect. Perazine is a potent inhibitor of dextromethorphan-O-demethylase, S-mephenytoin-hydroxylase, alprazolam-4-hydroxylase, phenacetin-O-deethylase and tolbutamide-hydroxylase activity in human liver microsomes. CONCLUSIONS: Alterations in the activity of CYP3A4, CYP2C9 and FMO3 through genetic polymorphisms, enzyme induction or inhibition bear the potential to cause clinically significant changes in perazine clearance. PER may alter the clearance of coadministered compounds metabolized by CYP2D6, CYP2C19, CYP2C9, CYP3A4 and CYP1A2.     

Cytochrome P450 2A6 and other human P450 enzymes in the oxidation of flavone and flavanone

1.?We previously reported that flavone and flavanone interact spectrally with cytochrome P450 (P450 or CYP) 2A6 and 2A13 and other human P450s and inhibit catalytic activities of these P450 enzymes. In this study, we studied abilities of CYP1A1, 1A2, 1B1, 2A6, 2A13, 2C9 and 3A4 to oxidize flavone and flavanone.

2.?Human P450s oxidized flavone to 6- and 5-hydroxylated flavones, seven uncharacterized mono-hydroxylated flavones, and five di-hydroxylated flavones. CYP2A6 was most active in forming 6-hydroxy- and 5-hydroxyflavones and several mono- and di-hydroxylated products.

3.?CYP2A6 was also very active in catalyzing flavanone to form 2′- and 6-hydroxyflavanones, the major products, at turnover rates of 4.8?min^?1 and 1.3?min^?1, respectively. Other flavanone metabolites were 4′-, 3′- and 7-hydroxyflavanone, three uncharacterized mono-hydroxylated flavanones and five mono-hydroxylated flavones, including 6-hydroxyflavone. CYP2A6 catalyzed flavanone to produce flavone at a turnover rate of 0.72?min^?1 that was ～3-fold higher than that catalyzed by CYP2A13 (0.29?min^?1).

4.?These results indicate that CYP2A6 and other human P450s have important roles in metabolizing flavone and flavanone, two unsubstituted flavonoids, present in dietary foods. Chemical mechanisms of P450-catalyzed desaturation of flavanone to form flavone are discussed.     

Effect of Dietary Polyphenon E and EGCG on Lung Tumorigenesis in A/J Mice

Purpose  

To compare the chemopreventive efficacy of Polyphenon E (Poly E), (−)-epigallocatechin-3-gallate (EGCG) and Polyphenon E without EGCG (Poly E-EGCG) on the development of benzo(a)pyrene (B(a)P)-induced lung tumors in A/J mice.     

Immunohistochemical Detection of Apoptotic Proteins, p53/Bax and JNK/FasL Cascade, in the Lung of Rats Exposed to Cigarette Smoke

Lung disease is the leading and second-leading cause of death in women and men in Taiwan, respectively. Epidemiological studies conducted in Taiwan have shown that cigarette smoking is the principal risk factor of lung disease, but little is known about the association between apoptosis and cigarette smoke (CS)-induced lung pathogenesis. We designed an animal exposure system to study signal proteins involved in the process of apoptosis induced by smoking in rat terminal bronchiole. Rats were exposed to CS in doses of 5, 10, and 15 cigarettes, respectively, and the exposure lasted for 30 min, twice a day, 6 days a week for 1 month. Following which the rats were sacrificed and the lung tissues were analyzed by histopathological methods. The terminal bronchioles revealed mild to severe inflammation according to the doses of CS and marked lipid peroxidation, lymphocyte infiltration, congestion, and epithelial emphysema of alveolar spaces were also noted. Using an in situ cell death detection kit (TA300), the association of CS with apoptosis was determined in a concentration-dependent manner. Immunohistochemical evaluation showed that CS treatment produced an increase in the cellular levels of Bax, t-Bid, cleaved caspase-3, phospho-p53, phospho-JNK, and FasL but a decline in Bcl-2 and Mcl-1 (p<0.001 for all) in rat terminal bronchioles. The results provided evidences suggesting that exposure to CS not only induced apoptosis, but also involved p53/Bax and JNK/FasL cascade pathway.     

Heterogeneous Binding of Sigma Radioligands in the Rat Brain and Liver: Possible Relationship to Subforms of Cytochrome P-450

Abstract: The binding of four sigma receptor ligands, ³H-(+)-N-allyl-N-normetazocine (³H-(+)-SKF 10,047), ³H-(+)-3-(3-hydroxyphenyl)-N-(1-propyl)piperidine (³H-(+)-3-PPP), ³H-haloperidol and ³H-N,N'-di(o-totyl)guanidine (³H-DTG), and the cytochrome P450IID6 ligand and dopamine uptake inhibitor ³H-1-[2-(diphenylmethoxy)ethyl]-4-(3-phenylpropyl)piperazine (³H-GBR 12935) to membranal preparations of rat liver or whole rat brain was examined regarding kinetical properties and inhibition by various compounds with affinity for sigma binding sites or cytochrome P-450. In rat brain the density of binding sites was increased in order (+)-SKF 10,047 <(+)-3-PPP<DTG<<GBR 12935. In liver the corresponding order was (+)-SKF 10,047 <DTG<haloperidol<(+)-3-PPP<GBR 12935. The inhibition pattern of each ligand was similar in brain and liver, indicating that the binding sites were similar in the two tissues. With the exception of ³H-(+)-SKF 10,047 which appears to bind to a homogeneous haloperidol-sensitive site, there were quite marked differences between the ligands studied, suggesting heterogeneous binding sites. For instance, (+)-SKF 10,047 and progesterone were potent inhibitors of the binding of ³H-(+)-SKF 10,047, ³H-(+)-3-PPP and ³H-haloperidol but inhibited only a minor fraction of the binding of ³H-DTG to the brain and liver preparations. Multiple binding sites were also indicated by the low Hill coefficients found for most of the compounds studied. It was found that the cytochrome P-450 inhibitor proadifen (SKF 525A), like haloperidol, was a potent inhibitor of the binding of ³H-(+)-SKF 10,047, ³H-(+)-3-PPP and ³H-haloperidol to the liver and brain preparations, less active in inhibiting the binding of ³H-DTG and least effective on the binding of ³H-GBR 12935. Another cytochrome P-450 inhibitor, L-lobeline, was particularly potent in inhibiting the binding of ³H-DTG but was also quite potent inhibitor of the binding of the other sigma ligands. It was less potent in inhibiting the binding of ³H-GBR 12935. The binding of the latter ligand was potently inhibited by the analogous compound GBR 12909 but of the other compounds examined only L-lobeline, proadifen, haloperidol, DTG and (+)-3-PPP had IC50 values below 10 μM. The possibility that the sigma binding sites are identical with some subforms of cytochrome P-450 is discussed.     

Phenotypic Analysis of Spleen, Thymus, and Peripheral Blood Cells in Aged C57BI/6 Mice Following Long-Term Exposure to 2,3,7,8-Tetrachlorodibenzo-p-Dioxin

A mouse model was used to identify potential biomarkers of exposureto the environmental contaminant 2,3,7,8-tetrachiorodibenzo-p-dioxin(TCDD). Female C57B1/6 mice were treated weekly with 0.2 µgTCDD/kg body weight or vehicle for 14–15 months. Phenotypicanalysis by flow cytometry identified the major cell subpopulationsin the spleen, thymus, and peripheral blood as defined by theexpression of CD4, CD8, B220, and Mac-1 molecules. These subpopulationswere further characterized for the expression of I-A, Pgp-1,CD45RB, and/or T cell receptor antigens (CD3, ß ).A group of young (4 months old) mice was evaluated concurrentlyto document immunophenotype alterations associated with aging.Results showed several age-related changes in phenotype distributionin the spleen and blood, but not in the thymus, despite significantage-dependent thymic involution. The age-dependent changes insplenic phenotypes included a decreased frequency of CD4⁺ cellsand a major shift in the frequency distribution from naive Tcells to effector and memory T cells as defined by Pgp-1 andCD45RB expression. These phenotypic changes in the spleen dueto aging correlated with similar changes in the blood, providingpreliminary support for the use of spleen cells as surrogatesfor blood in the development of biomarkers of immunotoxicity.Long-term exposure to a total cumulative dose 12–13 µgTCDD/kg body weight resulted in no overt toxicity, a 16-foldelevation of hepatic ethoxyresorufin-O-deethylase activity,and residue levels of 1.27 ± 0.16 ng TCDD/g abdominalfat. In comparison to the effects of aging, TCDD treatment producedrelatively subtle changes in immunophenotypes. In the TCDD-treatedthymus, the proportion of CD4^–CD8^– cells was increasedas was the proportion of ⁺ thymocytes. These effects were verysmall but of interest in that similar thymic effects have beenpreviously reported following prenatal exposure to TCDD. Inthe spleen, TCDD exposure did not alter the frequency of CD4⁺or CD8⁺ T cells, B cells, or macrophages but significantly alteredfunctionally discrete subpopulations within the T cell compartment.The most definitive change in TCDD-treated mice was a decreasein the frequency of memory T helper cells, defined as CD4⁺ Pgp-1^hiCD45RB^lo, with a concomitant increase in the proportion ofnaive T helper cells identified as CD4⁺Pgp-1^loCD45RB^hi. Thesechanges are consistent with the known immunosuppressive activityof TCDD. Thus, these results identify Pgp-1 and CD45RB as potentialbiomarkers of TCDD immunotoxicity.     

Cytochrome P-450-mediated activation of procarcinogens and promutagens to DNA-damaging products by measuring expression of umu gene in Salmonella typhimurium TA1535/pSK1002

A simple and sensitive procedure for the determination of cytochrome P-450 (P-450)-mediated activation of chemical procarcinogens and promutagens to DNA-damaging products has been developed using a method measuring the expression of the umu gene in Salmonella typhimurium TA1535/pSK1002, which is based upon the initial procedures as described by Oda et al. [Mutation Res. 147, 219 (1985)]. The chemicals examined were a variety of potent carcinogenic and mutagenic compounds including heterocyclic aromatic amines, aromatic amines, polycyclic aromatic hydrocarbons and aflatoxin B1. These chemicals were incubated with rat liver microsomes or a reconstituted monooxygenase system containing three forms of purified P-450 in the presence of a bacterial tester strain, and the induced-umu gene expression was determined by measuring the beta-galactosidase activity produced by fusion gene in the cells. The activity was increased linearly for at least 2 hr with an initial lag time of 30 min and was dependent on the concentrations of P-450 in the reaction mixture. Thus, the metabolic activation of these compounds by P-450 could be compared on a basis of the specific beta-galactosidase activity/min/nmol P-450. Among three forms of P-450, two isozymes induced by 3-methylcholanthrene were found to be more active in catalyzing the metabolic activation of most of the chemicals examined than a form of P-450 which is induced by phenobarbital. Data also showed that a high spin form of P-450 isolated from 3-methylcholanthrene-treated rats had a profound role in the activation of procarcinogens and promutagens. This conclusion was based on the results of catalytic activities by three forms of P-450 in a reconstituted monooxygenase system, and on the effects of specific antibodies against these P-450s on the reactions catalyzed by liver microsomes.     

Relationship Between Activities of Cytochrome P-450 Monooxygenases in Human Placental Microsomes and Binding of Benzo(A)Pyrene Metabolites to Calf Thymus DNA

ABSTRACT

Benzo(a)pyrene metabolism in human placental microsomes from smokers was studied. Benzo(a)pyrene metabolites were separated using high pressure liquid chromatographic technique. Reaction of benzo(a)pyrene with a microsomal fraction of placenta from individuals who smoke cigarettes during pregnency yields 7,8 dihydroxy benzo(a)pyrene as a major metabolite, while 3′-hydroxy benzo(a)pyrene, 4,5 dihydroxy benzo(a)pyrene and quinones constitute minor metabolites. The activities of arylhydrocarbon hydroxylase and 7-ethoxycoumarine deethylase exhibited much higher activities in smokers than in nonsmokers. Examination of specific binding of monoclonal antibodies to cytochrome P-450 isozymes in placental microsomes revealed that cigaratte smoking specifically enhanced the level of cytochrome P-450 c and d isozymes in human placental microsomes. Coincubation of ³H-benzo(a)pyrene and calf thymus DNA with placental microsomes yielded acid insoluble ³H-B(a)P from smokers, suggesting that cigarette smoking may induce placental enzymes which convert benzo(a)pyrene into ultimate metabolites to form carcinogen-DNA adducts.     

Lung tumor production and tissue metal distribution after exposure to manual metal ARC-stainless steel welding fume in A/J and C57BL/6J mice

Stainless steel welding produces fumes that contain carcinogenic metals. Therefore, welders may be at risk for the development of lung cancer, but animal data are inadequate in this regard. Our main objective was to examine lung tumor production and histopathological alterations in lung-tumor-susceptible (A/J) and -resistant C57BL/6J (B6) mice exposed to manual metal arc-stainless steel (MMA-SS) welding fume. Male mice were exposed to vehicle or MMA-SS welding fume (20 mg/kg) by pharyngeal aspiration once per month for 4 mo. At 78 wk postexposure, gross tumor counts and histopathological changes were assessed and metal analysis was done on extrapulmonary tissue (aorta, heart, kidney, and liver). At 78 wk postexposure, gross lung tumor multiplicity and incidence were unremarkable in mice exposed to MMA-SS welding fume. Histopathology revealed that only the exposed A/J mice contained minimal amounts of MMA-SS welding fume in the lung and statistically increased lymphoid infiltrates and alveolar macrophages. A significant increase in tumor multiplicity in the A/J strain was observed at 78 wk. Metal analysis of extrapulmonary tissue showed that only the MMA-SS-exposed A/J mice had elevated levels of Cr, Cu, Mn, and Zn in kidney and Cr in liver. In conclusion, this study further supports that MMA-SS welding fume does not produce a significant tumorigenic response in an animal model, but may induce a chronic lung immune response. In addition, long-term extrapulmonary tissue alterations in metals in the susceptible A/J mouse suggest that the adverse effects of this fume might be cumulative.     

Examining the Validity of Self-reported Primary and Secondary Exposure to Cigarette Smoke in Adolescent Girls: The Utility of Salivary Cotinine as a Biomarker

Background: Studies of cigarette use and exposure often rely on either self-report or cotinine assay. In adolescence it is not clear how well assays and self-report correspond, or what effect estrogen exposure has on cotinine. Objectives: This study sought to identify optimal cut-points for salivary cotinine thresholds for girls with primary, secondary, and no smoke exposure, and whether menarche and hormone contraceptive use are important for interpreting salivary cotinine. Methods: This longitudinal prospective study recruited 262 healthy adolescent girls who participated in three annual interviews across 24 months. Salivary cotinine assays and self-report of primary and secondary smoke exposure, menarcheal status, and hormone contraceptive use were collected. Results: No adolescents reported primary smoke exposure without secondary exposure. Optimal cut-points for distinguishing primary smoke exposure from secondary-only and no smoke exposure were 1.05 and 3.01 ng/ml, respectively based on receiver operator curves (ROC); no reliable cut-point for secondary-only versus no smoke exposure was identified. The ideal salivary cotinine cut-point to distinguish primary smoke exposure varied by hormone contraceptive use and was 2.14 ng/ml for those using progesterone contraceptives, higher than that of girls using estrogen contraceptives and those not using hormone contraceptives. Conclusions: This study is the first to examine variance in salivary cotinine cut-points based on hormone exposure for adolescent girls, with findings indicating that hormone contraceptive use in particular may be a key consideration when identifying adolescent smoking. The use of previously recommended salivary cotinine cut-points of 3.85 ng/ml or higher may overestimate nonsmokers.     

Comparative Ability of Various PCBs, PCDFs, and TCDD to Induce Cytochrome P450 1A1 and 1A2 Activity Following 4 Weeks of Treatment

Toxic equivalency factors (TEFs) have been proposed for di benzo-p-dioxins,dibenzofurans, and polyhalogenated biphenyls. The proposed toxicequivalency factors (TEFs), which are presently being evaluatedin our laboratory, are currently used to estimate the potentialhealth risk associated with exposure to complex mixtures containingthese chemicals. In preliminary studies, equally potent doses,based on the published TEEs and relative enzyme-inducing potency,of 2,3,7,8-tetrachioro-dibenzo-p-dioxin (TCDD), 2,3,7,8-tetrachlorodibenzofuran,1,2,3,7,8-pentachlorodibenzofuran, 1,2,3,4,6,7,8,9-octachloro-dibenzofuran,3,4,3',4'-tetrachlorobiphenyl, 2,3,4,3',4'-pentachlo-robiphenyl,3,4,5,3',4'-pentachlorobiphenyl, 2,3,4,3',4',5'-hex-achlorobiphenyl,2,3,4,5,3',4'-hexachlorobiphenyl, and 3,4,5,3',4',5'-hexachlorobiphenylwere administered to female B6C3F1 mice 5 days a week over a4-week period. Hepatic, skin, and lung cytochrome P450 1A1 andhepatic 1A2 activities were determined for all chemicals testedand compared to those from TCDD-treated mice. These initialstudies indicate that the present TEFs do not reliably predictinduction potency for many of the chemicals. Furthermore, ourdata suggest that the relative inductive potency of these chemicalsmay be tissue specific and that estimates of TEFs based on hepaticethoxyresorufin O-de ethylase activity may not accurately reflectthe potency of these chemicals in nonhepatic tissue. The TEFsproposed for the "dioxin-like" polychlorinated biphenyls (PCBs)overestimate the potency of these compounds by factors of 10–1000.The present study indicates that more experimental data arerequired before TEFs for PCBs should be used in regulatory decisionmaking.     

Cytochrome P-450 3A and 2D6 catalyze ortho hydroxylation of 4-hydroxytamoxifen and 3-hydroxytamoxifen (droloxifene) yielding tamoxifen catechol: involvement of catechols in covalent binding to hepatic proteins.

Earlier study suggested that 3,4-dihydroxytamoxifen (tam catechol), a tamoxifen metabolite, is proximate to the reactive intermediate that binds covalently to proteins and possibly to DNA (). The current study demonstrates that rat and human hepatic cytochrome P-450s (CYPs) catalyze tam catechol formation from tamoxifen (tam), 3-hydroxy-tam (Droloxifene), and 4-hydroxy-tam (4-OH-tam). Higher levels of catechol were formed from 4-OH-tam and 3-hydroxy-tam than from tam. Evidence that human hepatic CYP3A4 and 2D6 catalyze the formation of tam catechol from 4-OH-tam and supportive data that the catechol is proximate to the reactive intermediate, was obtained: 1) There was a good correlation (r = 0.82; p 相似文献   

Two-Week, Repeated Inhalation Exposure of F344/N Rats and B6C3F Mice to Ferrocene

Two-Week, Repeated Inhalation Exposure of F344/N Rats and B6C3F¹Mice to Ferrocene. SUN, J. D., DAHL, A. R., GILLETT, N. A.,BARR, E. B., CREWS, M. L., EIDSON, A. F., BECHTOLD, W. E., BURT,D. G., DIETER, M. P., AND HOBBS, C. H. (1991). Fundam. ApplToxicol. 17, 150-158. Ferrocene (dicyclopentadienyl iron; CASNo. 102-54-5) is a relatively volatile, organometallic compoundused as a chemical intermediate, a catalyst, and as an antiknockadditive in gasoline. It is of particular interest because ofits structural similarities to other metallocenes that havebeen shown to be carcinogenic. F344/N rats and B6C3F, mice wereexposed to 0, 2.5, 5.0, 10, 20, and 40 mg ferrocene vapor/m3,6 hr/day for 2 weeks. During these exposures, there were nomortality and no observable clinical signs of ferrocene-relatedtoxicity in any of the animals. At the end of the exposures,male rats exposed to the highest level of ferrocene had decreasedbody-weight gains relative to the weight gained by filteredair-exposed control rats, while body-weight gains for all groupsof both ferrocene- and filtered air-exposed female rats weresimilar. Male mice exposed to the highest level of ferrocenealso had decreased body-weight gains, relative to controls,while female mice had relative decreases in body-weight gainsat the three highest exposure levels. Male rats had a slightdecrease in relative liver weight at the highest level of exposure,whereas no relative differences in organ weights were seen infemale rats. Male mice had exposure-relative decreases in liverand spleen weights, and an increase in thymus weights, relativeto controls. For female mice, relative decreases in organ weightswere seen for brain, liver, and spleen. No exposure-relatedgross lesions were seen in any of the rats or mice at necropsy.Histopathological examination was done only on the nasal turbinates,lungs, liver, and spleen. The only exposure-related findingwas histopathologic lesions in the nasal turbinates of bothspecies. These lesions were primarily centered in the olfactoryepithelium and were morphologically diagnosed as subacute, necrotizinginflammation. Nasal lesions were observed in all ferrocene-exposedanimals and differed only in severity, which was dependent onthe exposure concentration. In vitro metabolism studies of ferroceneshowed that nasal tissue, particularly the olfactory epithelium,had 10 times higher "ferrocene hydroxylating" activity thandid liver tissue from the same animals. These results suggestthat the mechanism of ferrocene toxicity may be the intracellularrelease of ferrous ion through ferrocene metabolism, followedby iron-catalyzed lipid peroxidalion of cellular membranes.     

A Comparison of the Fate of Inhaled Methyl Chloroform (1,1,1-Trichloroethane) Following Single or Repeated Exposure in Rats and Mice

A Comparison of the Fate of Inhaled Methyl Chloroform (1,1,1-Trichloroethane)Following Single or Repeated Exposure in Rats and Mice. Schumann,A.M., Fox, T.R. and Watanabe, P.G. (1982). Fundam. Appl. Toxicol.2:27–32. Male Fischer 344 rats and B6C3F1 mice were exposedby inhalation to 1500 ppm of methyl chloroform (MC) 6 hours/day,5 days/week for approximately 16 months. On the last day ofrepeated exposure ¹⁴C-labeled MC was used. The fate of the ¹⁴C-MCin the repeatedly exposed animals was compared to a group ofrats and mice which had been exposed concurrently for 16 monthsto chamber air (age-matched controls) prior to receiving thesingle 6 hour exposure to 1500 ppm of ¹⁴C-MC. The routes ofexcretion and tissue concentration of ¹⁴C activity were similarbetween the singly and repeatedly exposed rats and mice. Themajor route of elimination of MC was exhalation of the parentchemical In the expired air and constituted approximately 97%of the total recovered radioactivity in rats and 92–94%in mice. The remaining radioactivity (3.9%) was recovered asmetabolized MC in the expired air (¹⁴CO₂) and as nonvolatileradioactivity in the urine, feces, carcass and cage wash. Micewere found to eliminate MC more rapidly via the pulmonary routeand to biotransform approximately 5-fold more MC on a body weightbasis than rats. Repeated exposure to MC did not significantlyaffect the disposition of MC compared to the singly exposedrats and mice. Thus even after long-term repeated exposure toMC, its biotransformation remains limited. Comparison of theresults of the present study to those obtained previously inyoung-adult rats and mice indicates that alterations in thepharmacokinetics of ¹⁴C-MC (increased body burden and decreasedrate of pulmonary elimination) occur with age but not priorrepeated exposure to MC.     

Thirteen-Week, Repeated Inhalation Exposure of F344/N Rats and B6C3F1 Mice to Ferrocene

Ferrocene (dicyclopentadienyl iron; CAS No. 102-54-5) is a relativelyvolatile compound used as a chemical intermediate, a catalyst,and an antiknock additive in gasoline. This organometallic chemicalis of particular interest because of its structural similaritiesto other metallocenes, some of which are carcinogenic. F344/Nrats and B6C3F₁ mice were exposed to 0, 3.0, 10, and 30 mg ferrocenevapor/m³, 6 hr/day, 5 days/week, for 13 weeks. During theseexposures, no rats or mice died, nor were any clinical signsof ferrocene-related toxicity observed. At the end of the exposures,male rats exposed to the lowest and highest level of ferrocenehad decreased body weight gains compared to filtered-air-exposedcontrol male rats, while body weight gains for all groups ofboth ferrocene- and filtered-air-exposed female rats were similar.Male mice exposed to ferrocene had no differences in body weightgains, compared to controls, but female mice had decreases inbody weight gains at the 10 and 30 mg/m³ exposure levels. Therewere exposure concentration- and exposure-time-related increasesin lung burdens of iron. The mean iron lung burden in rats exposedto 30 mg ferrocene vapor/m³ for 90 days was four times greaterthan the burden in control rats. No exposure-related changesin respiratory function, lung biochemistry, bronchoalveolarlavage cytology, total lung collagen, clinical chemistry, andhematology parameters were observed. This suggests that theaccumulations of iron in lung did not cause an inflammatoryresponse nor any functional impairment of the lung. There wereno indications of developing pulmonary fibrosis nor of any hematologictoxicity. No exposure-related gross lesions were seen in anyof the rats or mice at necropsy. Exposure-related histopathologicalterations, primarily pigment accumulations, were observedin the nose, larynx, trachea, lung, and liver of both species,and in the kidneys of mice. Lesions were most severe in thenasal olfactory epithelium where pigment accumulation, necrotizinginflammation, metaplasia, and epithelial regeneration occurred.Nasal lesions were observed in all ferrocene-exposed animalsand differed only in severity, which was dependent on the exposureconcentration. Histochernical stains of these target tissuesshowed the presence of iron ions. The results suggest that themechanism of ferrocene toxicity may be the intracellular releaseof ferrous ion through ferrocene metabolism, followed by eitheriron-catalyzed lipid peroxidation of cellular membranes or theiron-catalyzed Fenton reaction to form hydroxyl radicals thatdirectly react with other key cellular components, such as proteinor DNA.