Carcinogenic potential and genomic instability of beryllium sulphate in BALB/c-3T3 cells.

Occupational exposure to beryllium (Be) and Be compounds occurs in a wide range of industrial processes. A large number of workers are potentially exposed to this metal during manufacturing and processing, so there is a concern regarding the potential carcinogenic hazard of Be. Studies were performed to determine the carcinogenic potential of beryllium sulfate (BeSO4) in cultured mammalian cells. BALB/c-3T3 cells were treated with varying concentrations of BeSO4 for 72 h and the transformation frequency was determined after 4 weeks of culturing. Concentrations from 50-200 microg BeSO4/ml, caused a concentration-dependent increase (9-41 fold) in transformation frequency. Non-transformed BALB/c-3T3 cells and cells from transformed foci induced by BeSO4 were injected into both axillary regions of nude mice. All ten Be-induced transformed cell lines injected into nude mice produced fibrosarcomas within 50 days after cell injection. No tumors were found in nude mice receiving non-transformed BALB/c-3T3 cells 90 days post-injection. Gene amplification was investigated in K-ras, c-myc, c-fos, c-jun, c-sis, erb-B2 and p53 using differential PCR while random amplified polymorphic DNA fingerprinting was employed to detect genomic instability. Gene amplification was found in K-ras and c-jun, however no change in gene expression or protein level was observed in any of the genes by Western blotting. Five of the 10 transformed cell lines showed genetic instability using different random primers. In conclusion, these results indicate that BeSO4 is capable of inducing morphological cell transformation in mammalian cells and that transformed cells induced by BeSO4 are potentially tumorigenic. Also, cell transformation induced by BeSO4 may be attributed, in part, to the gene amplification of K-ras and c-jun and some BeSO4-induced transformed cells possess neoplastic potential resulting from genomic instability.

Transforming and carcinogenic potential of cadmium chloride in BALB/c-3T3 cells.

A large number of workers are potentially exposed to cadmium during mining and processing. Therefore, there is a concern regarding the potential carcinogenic hazards of cadmium to exposed workers. Studies have been performed to determine if cadmium chloride (CdCl(2)) can induce morphological cell transformation, DNA from CdCl(2)-induced transformed cells can transform other mammalian cells, and the transformed cells induced by CdCl(2) can form tumors in nude mice. BALB/c-3T3 cells were treated with different concentrations of CdCl(2) for 72 h. The frequency of transformed foci from each treatment was determined after cells were cultured for 4 to 5 weeks. DNAs from five CdCl(2)-induced transformed cell lines were isolated and gene transfection assay was performed using NIH-3T3 cells. Non-transformed BALB/c-3T3 cells and cells from 10 transformed cell lines induced by CdCl(2) were injected into both axillary regions of nude mice. Mice were screened once a week for the appearance and size of tumors. CdCl(2) caused a statistically significant, concentration-related increase in the transformation frequency. DNA from all five CdCl(2)-induced transformed cell lines tested was found to induce varying degrees of transfection-mediated transformation in NIH-3T3 cells. All 10 CdCl(2)-induced transformed cell lines formed fibrosarcomas in nude mice within 39 days of inoculation. Within this time period, no tumors were found in nude mice injected with non-transformed BALB/c-3T3 cells. These results indicate that CdCl(2) is capable of inducing morphological cell transformation and that the transformed cells induced by CdCl(2) are potentially tumorigenic.

Occupational exposure to genotoxic agents.

Millions of workers in the United States are potentially exposed each year to hazardous chemicals, dusts, or fibers in occupational settings. Some of these agents are genotoxic and may cause genetic alterations in the somatic or germ cells of exposed workers. Such alterations, if they occur in proto-oncogenes or tumor suppressor genes, which are involved in controlling cell growth or differentiation, may lead to the development of cancer. Genetic alterations in germ cells may also lead to reproductive failure or genetic disorders in subsequent generations. It has been estimated that occupational exposure accounts for 4% of all human cancers and up to 30% of cancer among blue-collar workers. Approximately 20,000 cancer deaths each year are attributable to occupational exposure in the United States. Occupational cancer and reproductive abnormalities have been listed on the National Occupational Research Agenda master list of research priorities as major occupational diseases and injuries.

A study of the effect of chrysotile fiber surface composition on genotoxicity in vitro.

Chrysotile fibers (NIEHS intermediate length) were treated with ultrapure HCl to alter the fiber surface chemistry without substantially changing fiber morphology or dimensions. The objective of the study was to determine whether fiber surface chemistry is an important variable in fiber genotoxicity in vitro. The modified fibers, along with native chrysotile fibers, were used to challenge Chinese hamster lung fibroblasts (V79) in vitro using the micronucleus induction genotoxicity assay. Fiber dimensions were assessed using scanning electron microscopy by measuring the distribution of fiber lengths in 3 length ranges: less than 3 microm, 3-10 microm, and greater than 10 microm. For both treated and native fiber samples, 500 fibers were examined. Results indicate that acid-treated fibers were about 20% shorter than untreated chrysotile. Surface chemistry alterations were verified by zeta-potential reversal, x-ray photoelectron spectroscopy (XPS), and scanning electron microscopy/energy-dispersive x-ray spectroscopy (SEM-EDS) elemental analysis. Scanning Auger spectrometry indicated the presence of Mg, O, and Si in both treated and native chrysotile samples, which confirmed the surface purity of both fiber samples. Both XPS and SEM-EDS analysis demonstrated substantial depletion of Mg from fiber surfaces. Results of the micronucleus assay showed a positive concentration-related response for both samples, with toxicity evident only at the highest concentration. No significant difference was found for the treated and untreated chrysotile samples. These results indicate that the surface chemistry is not an important variable in the in vitro genotoxicity of chrysotile asbestos in V79 cells as detected by the micronucleus assay under the conditions used in this study, and support a model of chemically nonspecific chromosomal and spindle damage effects.

Genomic instability in silica- and cadmium chloride-transformed BALB/c-3T3 and tumor cell lines by random amplified polymorphic DNA analysis.

Our earlier studies using random amplified polymorphic DNA (RAPD) analysis have shown genetic instability in human lung cancer tissues. Here we have investigated the potential for genetic instability in silica- and cadmium chloride (CdCl2)-transformed BALB/c-3T3 cell lines. Non-transformed, transformed BALB/c-3T3 cells, and tumor cell lines (obtained by injecting nude mice with transformed cell lines) were analyzed for genomic changes. DNAs from 10 different transformed clones and their corresponding tumor cell lines were amplified individually by RAPD analysis using 10 arbitrary primers. DNA from non-transformed BALB/c-3T3 cells was used as a control to compare genetic alterations, if any, between non-transformed, transformed and tumor cell populations. PCR products from RAPD were electrophoretically separated on agarose gels and the banding profiles were visualized by ethidium bromide staining. Five of the 10 primers tested revealed genomic changes in silica-transformed cell lines when compared to non-transformed BALB/c-3T3 cells. Comparison of all 10 transformed and tumor cell lines showed varied degrees of genomic changes using all 10 primers. CdCl2-transformed cell lines displayed fewer genomic changes, only three of 10 primers showed a positive result. CdCl2-transformed cells and their corresponding tumor cell lines showed specific banding pattern differences in six of the 10 samples tested with six of the 10 primers. Changes in band intensity were the most commonly observed changes both in silica- and CdCl2-transformed and tumor cell lines. The results seem to indicate a progressive change in genomic rearrangements which may directly or indirectly be associated with progression of tumorigenesis.

Protective effect of vanillin on radiation-induced micronuclei and chromosomal aberrations in V79 cells.

Vanillin (VA), an anticlastogen, has been demonstrated to inhibit gene mutations in both bacterial and mammalian cells. However, the data on its effect against radiation-induced cytogenetic damage are limited. The aim of this study was to investigate the protective effect of VA on radiation-induced chromosomal damage in V79 cells. Exponentially growing cells were exposed to five doses of X-rays (1-12 Gy) and UV radiation (50-800 microJ x 10(2) cm-2 and posttreated with 3 concentrations of VA (5, 50 or 100 micrograms ml-1 for 16 h for micronucleus (MN) and 18 h for structural chromosomal aberration (SCA) analyses. MN and SCA assays were performed concurrently according to standard procedures. Results indicate that there was a dose related increase in the percent of micronucleated binucleated cells (MNBN) (5.6 to 79.6) and percent of aberrant cells (Abs) (12 to 98) with X-ray treatment alone. Inhibition studies showed that the addition of VA at 100 micrograms ml-1 significantly reduced the percent of MNBN (21 to 48) induced by X-ray at 1, 2, and 4 Gy. There was a slight decrease in percent MNBN at 5 and 50 micrograms VA ml-1. All three concentrations of VA decreased percent Abs (15.7 to 57.1) induced by X-rays at all doses. UV radiation alone significantly increased percent MNBN (3.5 to 14.8) and percent Abs (17 to 29). Addition of 50 or 100 micrograms VA ml-1, significantly decreased percent MNBN (31.7 to 86.2) and percent Abs (54.5 to 90.9) at all doses of UV radiation. A decrease in percent MNBN (2.8 to 72.4) and percent Abs (34.8 to 66.7) was also noted at 5 micrograms VA ml-1. These data clearly indicate the protective effect of VA on radiation-induced chromosomal damage, suggesting that VA is an anticlastogenic agent.

Inhibition of methotrexate-induced chromosomal damage by folinic acid in V79 cells.

Methotrexate (MTX), an anticancer compound, is widely used in the treatment of leukemia. It induces cytogenetic damage as well as cytostatic effects on a variety of cell systems. Folinic acid (Leucovorin) is generally administered along with MTX as a rescue agent to decrease MTX-induced toxicity. However, information regarding the inhibitory effect of folinic acid against cytogenetic damage caused by MTX is limited. This study was conducted to assess the cytogenetic effect of MTX and its inhibition by folinic acid (FA) using the micronucleus and chromosomal aberration assays concurrently. Exponentially growing V79 cells were treated with MTX at five different concentrations (5-100 micrograms ml-1) with S9 microsomal fraction for 6 h and post-treated with two concentrations of FA (5 or 50 micrograms) for 40 h. Results indicate that MTX alone induced a concentration-related increase in % micronucleated binucleated cells (MNBN) and % aberrant cells (Abs). There was a decrease in nuclear division index (NDI) with increase in MTX concentration. Similarly, the mitotic index (MI) also decreased in all concentrations of MTX tested. The addition of FA at 50 micrograms ml-1 significantly reduced % MNBN (40-68%) and % Abs (36-77%). Inhibition was also seen at 5 micrograms FA (12 to 54% for MNBN and 20 to 61% for Abs). These results indicate that FA is capable of reducing the cytogenetic damage induced by MTX and appears to be an anticlastogenic agent.

Detection of genomic instability in lung cancer tissues by random amplified polymorphic DNA analysis.

Genomic instability resulting in multiple mutations is believed to be a driving force in the carcinogenic process. In this study, the random amplified polymorphic DNA (RAPD) technique, a simple PCR-based DNA polymorphism assay system, was used for detecting genomic instability in lung cancer tissues. DNAs from 20 lung cancer (18 non-small cell lung cancers and two small cell lung cancers) and their corresponding normal tissues were amplified individually by RAPD with seven different 10-base arbitrary primers. PCR products from RAPD were electrophoretically separated in agarose gels and banding profiles were visualized by ethidium bromide staining. The ability to detect genomic instability in 20 cancer tissues by each single primer ranged from 15 to 75%. DNA changes were detected by at least one primer in 19 (95%) cancer tissues. These results seem to indicate that genomic rearrangement is associated with lung carcinogenesis and that RAPD analysis is useful for the detection of genomic instability in lung cancer tissues.

Detection of mineral-dust-induced DNA damage in two mammalian cell lines using the alkaline single cell gel/comet assay.

It has been estimated that over three million workers in the USA are potentially exposed to silica or other mineral dusts. Results of epidemiological studies evaluating whether silica or glass fibers increase lung cancer risk to the exposed workers are inconclusive. Detection of DNA damage in cells exposed to genotoxic agents is being used to assess the carcinogenic potential of environmental agents. The alkaline (pH > 13) single cell gel/comet (SCG) assay was used to determine and compare DNA damage in cultured Chinese hamster lung fibroblasts (V79 cells) and human embryonic lung fibroblasts (Hel 299 cells) exposed to crystalline silica (Min-U-Sil 5), amorphous silica (Spherisorb), carbon black, and glass fibers (AAA-10). V79 or Hel 299 cells were exposed to these mineral dusts for 3 h at various concentrations. Min-U-Sil 5 and AAA-10, at almost all concentrations tested, caused a significant increase in DNA migration measured as tail length in both V79 and Hel 299 exposed cells. However, the increase was much higher in V79 then in Hel 299 cells for Min-U-Sil 5. Tail length was also increased relative to controls after amorphous silica treatment, but not to the same extent as that induced by crystalline silica. Exposure to carbon black did not induce DNA migration at any of the concentrations tested. These results indicate that silica and glass fibers, but not carbon black, can induce DNA damage in mammalian cells, and that crystalline silica has a higher DNA-damaging activity than amorphous silica. For glass fibers, induction of DNA damage in both V79 and Hel 299 cells was observed even at a concentration 10 times lower than silica and the response was similar in both cell lines. These results suggest that the SCG/comet assay is useful for the detection of DNA damage caused by occupationally related dusts/particles.

Quantitative and qualitative comparison of spontaneous and chemical-induced specific-locus mutation in the ad-3 region of heterokaryon 12 of Neurospora crassa.

The data from forward-mutation experiments to obtain specific-locus mutations at two closely linked loci in the adenine-3 (ad-3) region of heterokaryon 12 (H-12) of Neurospora crassa have been tabulated to determine the relative frequencies and mutational spectra of ad-3 mutants occurring spontaneously and those induced by 22 different chemical treatments. Previous studies have demonstrated that specific-locus mutations at these two loci result from 5 major genotypic classes, namely two classes of gene/point mutations (ad-3AR and ad-3BR), and 3 classes of multilocus deletion mutations ([ad-3A]IR, [ad-3B]IR and [ad-3A ad-3B]IR). In addition, prior studies have demonstrated that some chemical mutagens induced ad-3 mutants exclusively, or almost exclusively, by gene/point mutation and other chemical mutagens by gene/point mutation and multilocus deletion mutation. In the latter cases, there was wide variation in the percentages of ad-3 mutants in these 5 major genotypic classes. Two comparative methods of analysis that also were used to compare spontaneous and chemical-induced ad-3 mutational spectra included X2-tests on the numbers of ad-3 mutants resulting in the following two sets of ratios: (1) gene/point mutations and multilocus deletion mutations; and (2) complementing and non-complementing ad-3BR, mutants. Combination of the p-values from X2-tests for these two methods of comparison demonstrated that all 22 chemicals induce a spectrum of ad-3 mutants that is qualitatively different from that occurring spontaneously. In addition, these same two methods of comparison have been used to compare the mutagenic effects of each of the 22 chemical treatments with each other. Combination of the data from these two methods of comparison has demonstrated that 93.1% (215/231) of the pairwise combinations of these 22 chemicals were different from each other. The implication of these experimental data on the induction of specific-locus mutations in somatic cells of Neurospora for genetic risk assessment exercises is discussed.

Studies on the mutagenicity of mild gasification products of coal and their subfractions by the Salmonella/microsomal assay.

Mild gasification of coal is a technology being developed in the United States in order to upgrade lower rank coals and facilitate their use in coal-burning electric generation plants. Thirteen coal-derived mild gasification products from different coal sources and processing conditions have been examined for their potential biohazards. The mutagenicity of these samples was tested with the Ames Salmonella/microsomal assay. Two solvents, dimethyl sulfoxide (DMSO) and polyoxyethylene-sorbitan monooleate (Tween 80), were used to dissolve samples in a manner to facilitate their interaction with the test organisms. The results showed that 9 of the 13 samples displayed mutagenic activity in test strains TA98 and/or TA100 with or without metabolic activation, whether dissolved in Tween 80 or DMSO. Five mutagenic and two nonmutagenic samples were class-fractionated into basic, acidic, nonpolar, and polar neutral subfractions to examine their class-related mutagenic activities. Results of the testing of subfractions of the five mutagenic and one nonmutagenic samples showed mutagenic activity in at least the nonpolar neutral fraction. The subfractions of the another nonmutagenic sample did not display any mutagenic activity. Chemical characterization of the subfractions revealed the existence of aromatic hydrocarbons in certain subfractions, which may be responsible for the mutagenic activity of the coal-derived mild gasification products.

Inhibition of methotrexate-induced chromosomal damage by vanillin and chlorophyllin in V79 cells.

Methotrexate (MTX), a chemotherapeutic agent used to treat cancer, produces cytogenetic damage and has a cytostatic effect in a variety of test systems. Several antigenotoxic agents have been studied in various in vitro and in vivo systems. However, data are limited regarding their ability to modulate MTX-induced genotoxicity. In the present study, vanillin (VA) and chlorophyllin (CHL) were used as antigenotoxic agents to study their ability to minimize the DNA damage caused by MTX. Exponentially growing V79 Chinese hamster lung cells were treated with MTX at five different concentrations (5-100 micrograms/ml) with S9 activation for 6 h and post-treated with two concentrations of either VA (50 or 100 micrograms/ml) or CHL (50 or 100 micrograms/ml) for 40 h. Cytochalasin B was added for the micronucleus (MN) assay along with antigenotoxic agents to evaluate MN in binucleated cells. Chromosomal aberrations were also evaluated in parallel cultures. Results indicate that MTX alone induced a dose-dependent decrease in the nuclear division index (NDI) and the mitotic index (MI). A significant increase in percent micronucleated binucleated cells (MNBN) and percent aberrant cells (Abs) was observed. Studies using VA as an antigenotoxic agent showed a decrease in the number of MNBN (26.3-83.1%) and Abs (16.0-87.5%) with the addition of either 50 or 100 micrograms VA/ml. The addition of CHL also significantly reduced the number of MNBN (53.0-91.5%) at both concentrations tested. Chromosomal aberrations were also significantly reduced (41.0-83.0). These studies indicate that both VA and CHL are capable of effectively minimizing MTX-induced chromosomal damage.

Micronucleus formation in V79 cells treated with respirable silica dispersed in medium and in simulated pulmonary surfactant.

Chinese hamster lung fibroblasts (V79 cells) were challenged with respirable silica particles using an in vitro genotoxicity assay. Two particle sizes of crystalline quartz and a non-crystalline silica were assayed for induction of micronuclei (MN) in V79 cells. Some of the silica dusts used were pretreated with simulated pulmonary surfactant to model in vivo exposure conditions. The results showed that both crystalline and non-crystalline silica dispersed in medium (MEM) induced MN formation in a dose-dependent manner. Crystalline silica was more active in this assay than non-crystalline silica on a mass basis. The results also show that the frequency of micronucleated cells in cultures treated with surfactant-coated silica was not significantly different from that of the non-treated control cultures. These results seem to indicate that silica can cause chromosomal aberrations and/or aneuploidies in V79 cells; however, pretreatment of silica particles with simulated pulmonary surfactant reduces or delays genotoxicity in this assay.

Induction of chromosomal aberrations by 2,4-dichloro-6-aminophenol in cultured V79 cells.

2,4-Dichloro-6-aminophenol (DCAP) is a major metabolite of the herbicide 2,4-dichloro-6-nitrophenol ammonium (DCNPA). In our previous studies this metabolite, but not the parent compound, was found to induce sister chromatid exchanges (SCEs) and micronucleus (MN) formation in Chinese hamster lung fibroblasts (V79 cells). Further studies were carried out to determine whether DCAP can also induce structural chromosomal aberrations (CAs) in cultured V79 cells. The assay was performed under three different treatment conditions: 1) 3 h treatment followed by 17 h post-treatment incubation. 2) 12 h treatment, and 3) 20 h treatment with multiple sampling times. Results showed that CAs were induced by DCAP in V79 cells. The aberrations were mainly chromatid types (gaps, breaks, and deletions), triradials, quadriradials, and complex rearrangements. However, induction of CAs appears to be dependent on the treatment condition. The 3 h treatment following a 17 h post-treatment incubation gave rise to 24% aberrant cells (excluding gaps) and a total of 44 aberrations per 100 metaphases, while the 12 h treatment only gave a borderline response and the 20 h treatment did not yield a significant increase of CAs at any concentration tested. It seems that a short-term treatment with higher concentrations is a better treatment protocol for genotoxicity studies of certain cytotoxic chemicals such as DCAP. Results of this study further indicate that the herbicide DCNPA is potentially hazardous to the exposed population.

Comparison of the mutagenicity and mutagen specificity of ethylenimine with triethylenemelamine in the ad-3 region of heterokaryon 12 of Neurospora crassa.

Studies have been performed to compare the mutagenicity and mutagenic specificity of the trifunctional alkylating agent, triethylenemelamine (TEM), and a closely related monofunctional agent, ethylenimine (EI), in the adenine-3 (ad-3) region of a 2-component heterokaryon (H-12) of Neurospora crassa. The primary objective of our studies was to characterize the genetic damage produced by each agent with regard to (1) mutagenic potency, and (2) the spectrum of specific-locus mutations induced in a lower eukaryotic organism. As in higher eukaryotes, specific-locus mutations in the ad-3 region of H-12 result from gene/point mutations, multilocus deletion mutations, and multiple-locus mutations. Specific-locus mutations resulting from gene/point mutation and multilocus deletion mutation can be detected in higher eukaryotes, but multiple-locus mutations can be detected only with difficulty or not at all. Our experiments with the ad-3 forward-mutation assay have demonstrated that TEM is a strong mutagen (maximum forward-mutation frequency between 100 and 1000 ad-3 mutations per 10(6) survivors) and EI is a moderate mutagen (maximum forward-mutation frequency between 10 and 100 ad-3 mutations per 10(6) survivors) for the induction of specific-locus mutations in the ad-3 region. Classical genetic tests were used to identify the different genotypic classes and subclasses among the EI- and TEM-induced ad-3 mutations from each experiment. The overall data base demonstrates that both EI- and TEM-induced ad-3 mutations result predominantly from gene/point mutations at the ad-3A and ad-3B loci (97.3% and 95.5%, respectively), and infrequently from multilocus deletion mutations (2.7% and 4.5%, respectively). Heterokaryon tests for allelic complementation on TEM- and EI-induced ad-3B mutations, however, have revealed a difference between the percentages showing allelic complementation (63.1% and 40.9%, respectively). Based on the specific revertibility of complementing and noncomplementing ad-3B mutations induced by other agents, this difference in the percentages of ad-3B mutations showing allelic complementation results from a difference between the spectrum of genetic alterations at the molecular level. In addition, comparison of the ratio of TEM-induced ad-3A and ad-3B mutations with those induced by EI has revealed a difference between the ad-3B/ad-3A ratios. Additional comparisons are made of the mutagenic effects of TEM and EI with those of other chemical mutagens and carcinogens in the ad-3 specific-locus assay in Neurospora.

[Mutagenicity study of eight diesel emission samples].

Mutagenicity of eight diesel emission samples were examined by the Microsuspension test. The results showed that four samples were positive, and the others were negative. We also compared the method with the standard Ames test. The data showed that the microsuspension test is sensitive and economical. The microsuspension test method is useful for determining mutagenicity of environmental mutagens.

Induction of micronuclei in BALB/c-3T3 cells by selected chemicals and complex mixtures.

The genotoxicity of benzo[a]pyrene, cyclophosphamide, 2-aminoanthracene, 2-nitrofluorene, nitrosated coal-dust extracts, and cigarette-smoke condensate were tested with the micronucleus assay using an established mammalian cell line. The results showed that all chemicals and complex mixtures studied induced micronuclei in BALB/c-3T3 cells. These results indicate that BALB/c-3T3 cells are capable of activating certain promutagens and procarcinogens. It seems, therefore, that in addition to cell transformation, the micronucleus assay in BALB/c-3T3 cells without an exogenous activation system may be useful for in vitro studies to detect genotoxic chemicals and complex mixtures.

Micronucleus induction and phagocytosis in mammalian cells treated with diesel emission particles.

Micronucleus induction and phagocytosis in V79 and CHO cells treated with diesel emission particles (DEP) were studied. After separation of the sample into supernatant and sediment fractions, the genotoxic activity of DEP was shown to reside in the supernatant fraction for the DMSO-extracted sample, and in the sedimented fraction for the dipalmitoyl lecithin (DPL), a primary component of pulmonary surfactant, dispersed sample. More particles from DMSO sediment samples were phagocytized than DPL sediment by both types of cells. This had no effect, however, on micronucleus induction. CHO cells phagocytized fewer particles, but gave a higher number of micronuclei than V79 cells. CHO cells seem to be more sensitive to DEP. Evidently, micronucleus induction is not the result of phagocytosis per se, but is due to the different response of the indicator cells to the DEP sample tested. These results further indicate that most, if not all, genotoxic compounds associated with DEP can be extracted by DMSO and that genotoxic activity associated with DEP inhaled into the lung may also be expressed by dispersion of particles in pulmonary surfactant.

Antimutagenicity studies of chlorophyllin using the Salmonella arabinose-resistant assay system.

Studies with the arabinose-resistant Salmonella forward mutation assay system were performed to determine the antimutagenic activity of chlorophyllin against the mutagenic activity of aflatoxin B1 (AFB1), 2-aminoanthracene (2AA), benzo[a]pyrene (BaP), N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and solvent extracts of coal dust (CD), diesel emission particles (DE), airborne particles (AP), tobacco snuff (TS), black pepper (BP) and red wine (RW). Various concentrations of each chemical and complex mixture extract were assayed for mutagenic activity with and/or without S9 in a preincubation test. One concentration of each chemical and complex mixture extract was then tested with various concentrations of chlorophyllin. Results showed that chlorophyllin, at concentrations of 2.5 mg/plate or less, completely or almost completely inhibited the mutagenicity of 2AA, AFB1, BaP, MNNG and solvent extracts of CD, DE and RW. With concentrations from 1.25 to 5 mg/plate, chlorophyllin inhibited over 50% of the mutagenicity of AP, TS and BP extracts. These results further substantiate the antimutagenic efficacy of chlorophyllin against chemicals and complex mixtures.

Comparative study of micronucleus assay and chromosomal aberration analysis in V79 cells exposed to ethylene oxide.

Micronucleus formation and chromosomal aberration (CA) in V79 cells were compared for their sensitivity in response to ethylene oxide (EtO) treatment. The results indicate that EtO exposure for 30 min induced CAs in V79 cells at the concentration of 3,500 ppm or higher. A statistically significant difference (P less than 0.01) was found between treated and control groups at all concentrations tested based on percent aberrant cells. The increase was dose-dependent with a correlation coefficient of 0.88. The aberrations found include chromatid and isochromatid breaks, fragments, minutes, and exchanges. Results of the micronucleus assay both in mononucleated and binucleated cells showed a slight but not statistically significant increase in micronuclei with doses between 457 to 4115 ppm. At the highest concentration tested (12344 ppm) EtO caused a significant increase in the micronucleus frequency (P less than 0.05).