Clinical experience of laboratory follow-up with noninvasive prenatal testing using cell-free DNA and positive microdeletion results in 349 cases.

OBJECTIVE: Screening via noninvasive prenatal testing (NIPT) involving the analysis of cell-free DNA (cfDNA) from plasma has become readily available to screen for chromosomal and DNA aberrations through maternal blood. This report reviews a laboratory's experience with follow-up of positive NIPT screens for microdeletions. METHODS: Patients that were screened positive by NIPT for a microdeletion involving 1p, 4p, 5p, 15q, or 22q who underwent diagnostic studies by either chorionic villus sampling or amniocentesis were evaluated. RESULTS: The overall positive predictive value for 349 patients was 9.2%. When a microdeletion was confirmed, 39.3% of the cases had additional abnormal microarray findings. Unrelated abnormal microarray findings were detected in 11.8% of the patients in whom the screen positive microdeletion was not confirmed. Stretches of homozygosity in the microdeletion were frequently associated with a false positive cfDNA microdeletion result. CONCLUSIONS: Overall, this report reveals that while cfDNA analysis will screen for microdeletions, the positive predictive value is low; in our series it is 9.2%. Therefore, the patient should be counseled accordingly. Confirmatory diagnostic microarray studies are imperative because of the high percentage of false positives and the frequent additional abnormalities not delineated by cfDNA analysis.

Iron deposition in rabbit cerebellum after exposure to generated and mobile GSM electromagnetic fields.

BACKGROUND: Mobile phone application may cause structural, functional changes and accumulation of toxic elements in brain. OBJECTIVES: The aim of this study was to investigate iron accumulation in rabbit cerebellum after exposure to RF EMF with light and scanning electron microscopy. MATERIALS AND METHODS: Histochemical analysis of iron distribution by light and electron microscopy with energy-dispersive microanalysis was used. RESULTS: Light microscopy revealed dystrophic changes of Purkinje cells in irradiated groups and iron deposits located in various parts of cerebellum. Deposits consists of C, O, Na, Mg, Al, Si, P, S, Cl, Ca and Fe. CONCLUSION: Our experiment revealed structural changes of Purkinje cells and iron and aluminium accumulations in stratum granulosum of rabbit's cerebellum after exposure to RF EMF (Fig. 6, Ref. 33).

Electromagnetic fields - do they pose a cardiovascular risk?

Mobile wireless communication technologies have now become an everyday part of our lives, 24 hours a day, 7 days a week. Monitoring the autonomous system under exposition to electromagnetic fields may play an important role in broading of our still limited knowledge on their effect on human body. Thus, we studied the interaction of the high frequency electromagnetic field (HF EMF) with living body and its effect on the autonomic control of heart rate using Heart Rate Variability (HRV) linear and nonlinear analyses in healthy volunteers. A group of young healthy probands (n=30, age mean: 24.2 ± 3.5 years) without any symptoms of disease was exposed to EMF with f=2400 MHz (Wi Fi), and f=2600 MHz (4G) for 5 minutes applied on the chest area. The short-term heart rate variability (HRV) metrics were used as an indicator of complex cardiac autonomic control. The evaluated HRV parameters: RR interval (ms), high frequency spectral power (HF-HRV in [ln(ms2)]) as an index of cardiovagal control, and a symbolic dynamic index of 0V %, indicating cardiac sympathetic activity. The cardiac-linked parasympathetic index HF-HRV was significantly reduced (p =0.036) and sympathetically mediated HRV index 0V % was significantly higher (p=0.002) during EMF exposure at 2400 MHz (Wi-Fi), compared to simulated 4G frequency 2600 MHz. No significant differences were found in the RR intervals. Our results revealed a shift in cardiac autonomic regulation towards sympathetic overactivity and parasympathetic underactivity indexed by HRV parameters during EMF exposure in young healthy persons. It seems that HF EMF exposure results in abnormal complex cardiac autonomic regulatory integrity which may be associated with higher risk of later cardiovascular complications already in healthy probands.

Hepatic DNA adducts and production of mutagenic urine in 2,6-dinitrotoluene-treated B6C3F1 male mice.

The hepatocarcinogen 2,6-dinitrotoluene (2,6-DNT) is an intermediate in the chemical synthesis of 2,4,6-trinitrotoluene and polyurethane products and can contaminate the waste stream emitted by these industries. In this study, the production of mutagenic urine metabolites and the formation of hepatic DNA adducts is examined in the B6C3F1 male mouse. Animals were administered 50 mg/kg 2,6-DNT by gavage for 3 consecutive days. No body or liver weight effects were observed in treated animals. Following sacrifice, the livers were excised and DNA isolated for examination of 2,6-DNT-derived DNA adducts. During 2,6-DNT treatment, urine was collected, concentrated, and tested for mutagenicity in the Salmonella reversion bioassay. Mutagenic urine metabolites (469+/-53 revertants/ml urine) were excreted from B6C3F1 mice treated with 2,6-DNT and were comparable to results obtained for CD-1 mice and Fischer 344 rats. Two distinct hepatic DNA adducts (0.8+/-0.1 and 0.6+/-0.1 RAL/10(8) nucleotides) were detected in B6C3F1 mice by (32)P-postlabeling and thin layer chromatography which differed from the four adducts observed in hepatic DNA from 2,6-DNT-treated Fischer 344 rats.

Mutagenicity of 1-nitropyrene metabolites from lung S9.

The mutagenicity of 1-nitropyrene metabolites from rabbit lung S9 incubates was evaluated using the Salmonella typhimurium plate incorporation assay with strain TA98, with and without Aroclor-induced rat liver S9. The following metabolites were isolated, identified and quantitated by HPLC: 1-nitropyrene -4,5- or -9,10-dihydrodiol (K-DHD), N-acetyl-1-aminopyrene ( NAAP ), 1-aminopyrene (1-AMP), 10-hydroxy-1-nitropyrene, 4-, 5-, 6-, 8- or 9-monohydroxy-1-nitropyrene (phenols) and 3-hydroxy-1-nitropyrene. The predominant metabolites formed by lung S9 incubates were K-DHD, 3-OH-1-nitropyrene and phenols. All of the metabolites were mutagenic in the absence of the exogenous rat liver S9 metabolic activation system, and several, including two unidentified metabolites were more potent than the parent 1-nitropyrene. The mutagenicity of 3 of the metabolites ( NAAP , 10-OH-1-nitropyrene and phenols) were enhanced by S9 while most of the other metabolites were less mutagenic in the presence of S9. These results indicate that lung tissue is capable of both oxidative and reductive metabolism which produced mutagenic metabolites, several of which were more potent than the parent compound, 1-NP.

Role of the intestinal microbiota in the activation of the promutagen 2,6-dinitrotoluene to mutagenic urine metabolites and comparison of GI enzyme activities in germ-free and conventionalized male Fischer 344 rats.

After male germ-free and conventionalized Fischer 344 rats were administered per os (p.o.) 75 mg/kg 2,6-DNT, intestinal nitroreductase, beta-glucuronidase, and azo reductase activities were lower in the cecum and large intestine of germ-free animals. However, there was no significant difference in the small intestinal nitroreductase and azo reductase compared to the conventionalized counterparts. This indicated a potential mucosal source for the enzymes. Urines from germ-free rats (1144 +/- 64 revertants/ml) were less mutagenic than those from conventionalized animals (1467 +/- 171 revertants/ml) in Salmonella typhimurium strain TA98 without S9. In the presence of S9, urine from conventionalized animals (894 +/- 56 revertants/ml) was more mutagenic than that from germ-free rats (686 +/- 60 revertants/ml). The presence of the intestinal flora plays an important role in the activation of 2,6-DNT but other metabolic pathways, such as the small intestinal mucosal and/or hepatic enzymes, are present that can generate excreted genotoxicants.

Comparative gastrointestinal enzyme activity and activation of the promutagen 2,6-dinitrotoluene in male CD-1 mice and male Fischer 344 rats.

Comparative intestinal nitroreductase, azo reductase, beta-glucuronidase, dechlorinase and dehydrochlorinase activities in young male Fischer 344 rats and young male CD-1 mice were measured in vitro while the comparative biotransformation of 2,6-dinitrotoluene to mutagenic metabolites was determined in vivo. The mice, which exhibit a high spontaneous incidence of hepatomas, had markedly greater nitroreductase activity and metabolized significantly more 2,6-dinitrotoluene to mutagenic metabolites than did Fischer 344 rats, which show a low incidence of liver tumors. Results of this study indicate that species differences in the incidence of hepatomas may be influenced by microbial flora and/or the biotransformation of xenobiotics in the G.I. tract.

Detection and comparison of DNA adducts after in vitro and in vivo diesel emission exposures.

Development of methods to evaluate certain classes of polycyclic aromatic compounds (PAC) detected in complex mixtures to which humans are exposed would greatly improve the diagnostic potential of 32P-postlabeling analysis. Identification of DNA adduct patterns or specific exposure-related marker adducts would strengthen associations between observed DNA adducts and exposures to different environmental pollutants (e.g., kerosene, cigarette smoke, coke oven, and diesel). We have compared diesel-modified DNA adduct patterns in various in vitro and in vivo rodent model systems and compared them to DNA reactive oxidative and reductive metabolites of 1-nitropyrene. The formation of nitrated polycyclic aromatic hydrocarbon (nitrated PAH) DNA adducts, derived from the metabolism of diesel extract constituents, was enhanced relative to other PAH-derived DNA adducts via xanthine oxidase-catalyzed nitroreduction. These adducts were detectable only by the butanol extraction version of the postlabeling analysis. Five major DNA adducts were detected in human lymphocytes treated in vitro with diesel extract. A major adduct detected in human lymphocytes treated in vitro with diesel extract comigrated with a major adduct detected in lymphocyte DNA treated with benzo[a]pyrene (BaP) alone. Other adducts that co-migrated with the major BaP-derived adducts were detected in skin and lung DNA isolated from rodents topically treated with (50 mg) diesel extract and the major adduct detected in calf thymus DNA treated with rat liver S9 and diesel particle extract. Postlabeling of lung DNA isolated from rodents exposed via lung inhalation for 24 months to diesel combustion emissions resulted in the formation of a major nuclease-P1-sensitive DNA adduct that did not co-migrate with the major BaP-diol epoxide adduct.(ABSTRACT TRUNCATED AT 250 WORDS)

In vitro characterization of DNA adducts formed by foundry air particulate matter.

This study is part of an ongoing investigation of biomarkers in iron foundry workers exposed to polycyclic aromatic compounds. Foundry workers with the highest exposures had elevated levels of DNA adducts in their white blood cells in previous studies. The purpose of this study was to characterize the nature of DNA reactive chemicals in foundry air samples through incubating the foundry filter extract with DNA and activation enzymes. Calf thymus DNA was incubated with foundry filter extract and activated by either rat liver activation mixture (S9 mix) or xanthine oxidase. A complex pattern of adducts was observed on thin-layer chromatography (TLC) by the 32P-postlabeling assay. Two selected polycyclic aromatic hydrocarbons (PAHs)--1-NP-and anti(+/-)benzo[a]pyrene-trans-7,8-dihydrodiol-9,10-epoxide [anti(+/-) BPDE]-DNA adducts--were used as marker compounds in characterizing the postlabeled DNA adducts by TLC combined with high-performance liquid chromatography (HPLC). After an initial separation of DNA adducts by TLC, individual spots were isolated and separated further on HPLC. HPLC analysis and spiking with anti(+/-)BPDE-DNA standard confirmed the co-migration of the anti(+/-)BPDE-DNA standard with one PAH adduct formed by the S9 mix-activated DCM extract in calf thymus DNA.

Effect of pentachlorophenol on the activation of 2,6-dinitrotoluene to genotoxic urinary metabolites in CD-1 mice: a comparison of GI enzyme activities and urine mutagenicity.

2,6-Dinitrotoluene (2,6-DNT) and pentachlorophenol (PCP) are used for industrial purposes and are found in the environment as hazardous contaminants. Because concurrent exposure to both compounds can occur, it is of interest to determine if organochlorine compounds potentiate the effect of nitroaromatic chemicals. CD-1 mice were treated with PCP (42.8 mg/kg) for 4 weeks. On weeks 1, 2, and 4 after the initial PCP dose, mice were treated p.o. with 2,6-DNT (75 mg/kg) and 24 hr urines were collected. After concentration, the urines were tested for their mutagenic activity in Salmonella typhimurium strain TA98 without metabolic activation in a microsuspension bioassay. A significant increase (P less than .05) in mutagenicity was observed in urines from mice treated with 2,6-DNT alone and in combination with PCP. By week 4, mice that received both 2,6-DNT and PCP excreted urine that was more mutagenic than that from animals which received only 2,6-DNT. At weeks 2 and 4, mice were sacrificed and intestinal enzyme activities (nitroreductase, azo reductase, beta-glucuronidase, dechlorinase, and dehydrochlorinase) were quantitated. The enhanced genotoxicity observed in urines from 2,6-DNT/PCP-treated mice coincided with a decrease in nitroreductase and an increase in beta-glucuronidase activities in the small intestine.

Atrazine treatment potentiates excretion of mutagenic urine in 2,6-dinitrotoluene-treated Fischer 344 rats.

Atrazine (ATZ), an s-triazine herbicide, is a widespread environmental contaminant. The hepatocarcinogenic component of technical grade dinitrotoluene, 2,6-dinitrotoluene (2,6-DNT, 19.5%), is a byproduct of trinitrotoluene synthesis and is found at production sites. This study explores the effect of ATZ treatment on the bioactivation of the promutagen, 2,6-DNT. Male Fischer 344 rats (5 weeks old) were administered 50 mg/kg of ATZ by gavage for 5 weeks. At 1, 3, and 5 weeks, both DMSO-control and ATZ-pretreated rats were treated p.o. with 75 mg/kg of 2,6-DNT and were housed in metabolism cages for urine collection. Sulfatase- and beta-glucuronidase-treated, concentrated urine was bioassayed for urinary mutagens in a microsuspension modification of the Salmonella assay with and without metabolic activation. No significant change in mutagen excretion was observed in ATZ-treated rats; however, an elevation in direct-acting urine mutagens from rats receiving ATZ and 2,6-DNT at weeks 1 (359 +/- 68 vs. 621 +/- 96 revertants/ml) and 5 (278 +/- 46 vs. 667 +/- 109 revertants/ml) of treatment was observed. The increase in production of urinary mutagens was accompanied by an elevation in small intestinal nitroreductase activity. Increases in large intestinal nitroreductase and beta-glucuronidase were observed after 5 weeks. There was no apparent effect of ATZ following 5 weeks of treatment on the production of 2,6-DNT-derived hepatic DNA adducts. ATZ treatment modifies intestinal enzymes responsible for promutagen bioactivation, and potentiates the excretion of mutagenic urine in 2,6-DNT-treated animals.

Modulation of 2,6-dinitrotoluene genotoxicity by alachlor treatment of Fischer 344 rats.

Due to its widespread use as a preemergent herbicide, alachlor has been detected as a groundwater contaminant. The procarcinogen, 2,6-dinitrotoluene (DNT), a by-product of the munitions industry and a precursor to polyurethane production, is found in the manufacturing waste stream. This study explores the effect of alachlor treatment on the bioactivation of DNT by examining urine mutagenicity, intestinal enzymes, and hepatic DNA adducts to detect changes in metabolism. Five-week-old male rats were treated daily by gavage with 50 mg/kg of alachlor for up to 5 weeks while control animals received an equal volume of peanut oil. At 1, 3, and 5 weeks following the initial alachlor dose, animals were administered p.o. 75 mg/kg DNT or DMSO. Urine was collected for 24 hr in metabolism cages. Following incubation with sulfatase and beta-glucuronidase, urines were individually concentrated by C-18 solid phase extraction, dried under N2, and prepared for bioassay in Salmonella typhimurium strain TA98 with and without metabolic activation. Urine from peanut oil- and alachlor-treated rots was not mutagenic. Even though calf thymus DNA-alachlor adducts formed in vitro, no hepatic DNA adducts were detected in vivo in these two treatment groups. Interestingly, a significant increase in excretion of mutagenic urine from DNT-treated rats was observed following 3 weeks of alachlor treatment in the absence of S9 (690 +/- 130 vs. 339 +/- 28 revertants/ml) which corresponded to increased DNT-related hepatic DNA adduct formation (5.90 +/- 0.88 adducts/10(8) nucleotides vs. 10.56 x +/- 0.59 adducts/10(8) nucleotides [relative adduct level (RAL)]). Elevation in the production of mutagenic urine from control and treated animals was linked to increases in intestinal nitroreductase and beta-glucuronidase activities; however, the only significant alachlor-related effects were an increase in small intestinal 1-week beta-glucuronidase and 5-week dehydrochlorinase activities. The increased urine mutagenicity and hepatic DNA adduct formation indicates that alachlor has a transient effect on DNT bioactivation that apparently is unrelated to intestinal bioactivation.

Mutagenic screening of marker grenade dyes by the Salmonella reversion assay, L5178Y/TK+/- mouse lymphoma assay, and in vivo sister chromatid exchange analysis in mice.

Two dyes (C.I. Solvent Yellow No. 33 and a mixture of C.I. Solvent Yellow No. 33 and C.I. Solvent Green No. 3) were tested for mutagenicity in the Salmonella reversion assay and the L5178Y/TK+/- mouse lymphoma assay, and also for sister chromatid exchange (SCE) induction in vivo in C57B1/6J mice. In addition, a greater than 99.9% pure sample of the yellow dye [2-(2'-quinolyl)-1,3-indandione] was tested with and without exogenous activation in the Salmonella reversion assay and the L5178Y/TK+/- mouse lymphoma assay. Neither C.I. Solvent Yellow No. 33 nor the C.I. Solvent Yellow No. 33 and Solvent Green No. 3 mixture was positive for inducing SCEs in vivo. All three dyes were tested in the standard plate incorporation test in seven Salmonella strains TA98, TA100, TA102, TA104, TA1535, TA1537, and TA1538. The dyes were negative with and without exogenous activation in TA98, TA1535, and TA1538. One test with TA1537 was positive with the greater than 99.9% purified yellow dye. All three dyes gave weakly positive results (less than a twofold increase) with S-9 in TA100 and were clearly positive in TA102 and TA104 both with and without S-9. They also induced mutation at the thymidine kinase locus in mouse lymphoma cells, produced both large- and small-colony trifluorothymidine-resistant mutants, and were clastogenic. The purified yellow dye was further tested for SCE induction in mouse lymphoma cells and was determined to give a slightly positive response in the presence of S-9.

Mutagenicity of HPLC-fractionated urinary metabolites from 2,4,6-trinitrotoluene-treated Fischer 344 rats.

The production and storage of explosives has resulted in the environmental accumulation of the mutagen 2,4,6-trinitrotoluene (TNT). In order to characterize the production of mutagenic urinary metabolites, 6-week old male Fischer 344 rats were administered 75 mg of TNT/kg or DMSO vehicle by gavage. The animals were placed into metabolism cages, and urine was collected for 24 hr. Following filtration, metabolites in the urine were deconjugated with sulfatase and beta-glucuronidase and concentrated by solid phase extraction. The eluate was fractionated by reverse-phase high-performance liquid chromatography (HPLC) using acetonitrile/water, and the fractions, were solvent exchanged in DMSO by nitrogen evaporation. Each HPLC fraction was bioassayed in strains TA98, TA98NR, TA100, and TA100NR without metabolic activation using a microsuspension modification of the Salmonella histidine reversion assay. Fractions 3, 5-18, 21, 22, and 24-26 contained mutagens detected by strain TA98. In the nitroreductase-deficient strain TA98NR, some mutagenic activity was lost; however, fractions 3, 6, 9-11, 15, and 25 clearly contained direct-acting mutagens. Fewer fractions were positive in strain TA100 (9-16, 19, 20, and 25) with less activity observed in the nitroreductase deficient strain TA100NR (fractions 3, 12, 14, 15, and 25). Although some mutagenic activity coeluted with known TNT metabolite standards, there were still many unidentified mutagenic peaks.

Potentiation of 2,6-dinitrotoluene genotoxicity in Fischer-344 rats by pretreatment with Aroclor 1254.

Pretreatment of Fischer 344 rats for 5 weeks with Aroclor 1254, a commercial mixture of polychlorinated biphenyls, potentiated the genotoxicity of 2,6-dinitrotoluene (DNT), a component of an industrial chemical used in the production of polyurethane foams. This interaction resulted from Aroclor 1254-mediated bioactivation of DNT to markedly greater levels of the genotoxic metabolites, that were excreted in urine and formed DNA adducts in the liver. A significant increase in the excretion of mutagenic urinary DNT metabolites was observed after the first week of Aroclor 1254 treatment, peaked at week 2 and then declined by nearly 25% at week 4. Nevertheless, by week 5, there was almost a 4-fold increase in the formation of hepatic DNA adducts. Significantly elevated hepatic metabolism and increased beta-glucuronidase in the small intestine and cecum, at 4 weeks, may account for the increased adducts and decreased urinary mutagens. Altered nitroreductase activity, reduced pH, and changes in the microfloral population may also play a role in the effect of Aroclor 1254 on the bioactivation of DNT. Such chemical interactions could be important to predictive risk assessment because the overall cancer risk of the mixture would exceed that determined by the current guidelines for chemical mixtures.

Mutagenicity of diesel-exhaust particle extract, 1-nitropyrene, and 2,7-dinitrofluorenone in Salmonella typhimurium under various metabolic activation conditions.

The mutagenic activities of 1-nitropyrene (1-NP), 2,7-dinitrofluorenone (2,7-DNF), and a diesel-exhaust extract were compared using the Salmonella typhimurium plate-incorporation assay. Each sample was tested with and without a 9000 X g liver homogenate (S9), both with and without an NADPH-generating system. The samples were also treated with the microsome fraction of S9, cytosol fraction of S9, boiled S9, bovine serum albumin (BSA), and boiled BSA. Salmonella tester strains TA98 and TA98FR1 were used in all treatments; TA98/1,8DNP6 was used to test mutagenic activity without activation. Without the NADPH-generating system, the samples generally had less mutagenic activity than samples treated with the NADPH-generating system. The addition of the NADPH-generating system resulted in marked increases in mutagenic activity of 1-NP in the microsome and S9 treatments, and of all 3 samples in the cytosol fraction treatment. These results indicate that although protein binding reduced the mutagenic activity of diesel-exhaust extract and 1-NP, microsomal activation increased the mutagenic activity of 1-NP. Because 1-NP and 2,7-DNF contributed less than 1.5% of the mutagenic activity of the diesel-exhaust extract, the response to diesel exhaust was not typified by these compounds.

Bacterial mutagenicity of aceanthrylene: a novel cyclopenta-fused polycyclic aromatic hydrocarbon of low molecular weight.

Aceanthrylene, a non-alternant cyclopenta-fused hydrocarbon, was shown to be weakly mutagenic without S9 and strongly mutagenic with S9 in the Ames Salmonella plate incorporation assay. The compound was most active in strain TA100 (35 revertants/nmole in the presence of 0.3 mg of S9 protein), and less active in strains TA98, TA1537 and TA1538 (20, 10 and 3.1 rev/nmole respectively, + S9). Strain TA1535 was unresponsive, suggesting that this compound induces frameshift mutations rather than base-pair substitutions. The mutagenic potency of aceanthrylene is consistent with predictions of its activity based on the relatively large delocalization energy (delta E deloc/beta = 0.931) of the carbonium ion which would result from oxirane ring opening of the 1,2-epoxide, a potential active metabolite.

Mutagenicity of derivatives and metabolites of 1-nitropyrene: activation by rat liver S9 and bacterial enzymes.

The mutagenicity and activation requirements of purified synthetic derivatives and potential metabolites of 1-nitropyrene have been characterized in the Ames plate incorporation assay with the Salmonella tester strains TA98, TA98NR and TA98/1,8-DNP6, in the presence or absence of exogenous metabolic activation provided by Aroclor-induced rat liver S9. All the compounds tested (1-aminopyrene, N-acetyl-1-aminopyrene, N-hydroxy-N-acetyl-1-aminopyrene, 3-hydroxy-1-nitropyrene, 6-hydroxy-1-nitropyrene, and 8-hydroxy-1-nitropyrene) exhibited mutagenic activity under one or more assay conditions. 1-Nitropyrene was metabolized to 3-hydroxy-1-nitropyrene, 6- or 8-hydroxy-1-nitropyrene, 1-aminopyrene, N-acetyl-1-aminopyrene and other unidentified products (including some bound to protein) by an S9 preparation analogous to that used for exogenous metabolic activation in the Ames assay. 1-Nitropyrene and 3-hydroxy-1-nitropyrene were activated primarily by the 'classical' nitroreductase, while the other compounds, particularly in the presence of S9 metabolic activation, were dependent on transesterification for expression of their mutagenicity.

1-Chloromethylpyrene: a reference skin sensitizer and genotoxin.

1-Chloromethylpyrene (1-CMP) has been evaluated as a model mutagen and toxin related to the ultimate electrophiles derived from benzo[a]pyrene and 1-nitropyrene. It was mutagenic to Salmonella (greater than 100 pg/plate) and exceptionally reactive to DNA when assessed by the 32P-postlabelling technique. 1-CMP was inactive in a mouse bone micronucleus assay when administered by gavage, probably due to hydrolysis, whose kinetics have been studied (t1/2 approximately 23 min at 37 degrees C). However, as expected, it was a potent skin toxin as determined by its activity as a mitogen to mouse skin and its contact allergenicity, as determined using the local lymph node proliferative assay. It is concluded that 1-CMP will probably be a potent human skin carcinogen and contact allergen.

Mutagenicity in lung of big Blue((R)) mice and induction of tandem-base substitutions in Salmonella by the air pollutant peroxyacetyl nitrate (PAN): predicted formation of intrastrand cross-links.

Peroxyacetyl nitrate (PAN) is a ubiquitous air pollutant formed from NO(2) reacting with acetoxy radicals generated from ambient aldehydes in the presence of sunlight and ozone. It contributes to eye irritation associated with photochemical smog and is present in most urban air. PAN was generated in a chamber containing open petri dishes of Salmonella TA100 (gas-phase exposure). After subtraction of the background mutation spectrum, the spectrum of PAN-induced mutants selected at 3.1-fold above the background mutant yield was 59% GC-->TA, 29% GC-->AT, 2% GC-->CG, and 10% multiple mutations - primarily GG-->TT tandem-base substitutions. Using computational molecular modeling methods, a mechanism was developed for producing this unusual tandem-base substitution. The mechanism depends on the protonation of PAN near the polyanionic DNA to release NO(2)(+) resulting in intrastrand dimer formation. Insertion of AA opposite the dimerized GG would account for the tandem GG-->TT transversions. Nose-only exposure of Big Blue((R)) mice to PAN at 78ppm (near the MTD) was mutagenic at the lacI gene in the lung (mutant frequency +/-S.E. of 6.16+/-0.58/10(5) for controls versus 8.24+/-0.30/10(5) for PAN, P=0.016). No tandem-base mutations were detected among the 40 lacI mutants sequenced. Dosimetry with 3H-PAN showed that 24h after exposure, 3.9% of the radiolabel was in the nasal tissue, and only 0.3% was in the lung. However, based on the molecular modeling considerations, the labeled portion of the molecule would not have been expected to have been bound covalently to DNA. Our results indicate that PAN is weakly mutagenic in the lungs of mice and in Salmonella and that PAN produces a unique signature mutation (a tandem GG-->TT transversion) in Salmonella that is likely due to a GG intrastrand cross-link. Thus, PAN may pose a mutagenic and possible carcinogenic risk to humans, especially at the high concentrations at which it is present in some urban environments.