R. Dean Blevins

Inhibition of DNA repair in ultraviolet-irradiated human cells by hydroxyurea

The effect on DNA repair in ultraviolet-irradiated human skin fibroblasts by hydroxyurea has been examined in this study using three independent methods for measuring DNA repair:the 5-bromodeoxyuridine photolysis assay which measures DNA repair replication, chromatographic measurement of thymine-containing dimers, and measurement of specific ultraviolet-endonuclease-sensitive sites in irradiated DNA. Little effect of hydroxyurea was observed at the concentration of 2 mM, which is often used to inhibit semiconservative DNA synthesis; however, 10 mM hydroxyurea resulted in marked inhibition (65--70%) of excision repair. This inhibition was accompanied by a possible doubling in the size of the repaired region. The accumulation of large numbers of single-strand breaks following ultraviolet irradiation and hydroxyurea incubation seen by other investigators was not observed with the normal skin fibroblasts used in this study. A comparison of hydroxyurea effects on the different DNA repair assays indicates inhibition of one step in DNA repair also results in varying degrees of inhibition of other steps as well.

Mutagenicity screening of five methyl carbamate insecticides and their nitroso derivatives using mutants of Salmonella typhimurium LT2

The mutagenic activity of five methyl carbamate insecticides-carbaryl, baygon, BUX-Ten, landrin and methomyl-and their nitroso derivatives were investigated using histidine auxotrophs-his TA98, his TA100, his TA1535, his TA1537 and his TA1538--of Salmonella typhimurium LT2 derived by Ames. The methyl carbamate insecticides did not cause a signficant increase in the number of revertant colonies in any of the strains used. In contrast, the nitroso derivatives of the carbamate insecticides greatly increased the number of colonies on plates inoculated with strains his TA100 and his TA1535. We conclude that the nitroso derivatives of the tested methyl carbamate insecticides are potent mutagens; whereas, the parent insecticides are non-mutagenic.

Nitrosated methylcarbamate insecticides: Effect on the DNA of human cells ☆

Normal human skin cells were treated with six insecticide esters of N-methylcarbamic acid or their N-nitroso derivatives. The DNA of the cells was sedimented in alkaline sucrose gradients at various times after treatment. The insecticides used were aldicarb, baygon, BUX-TEN, carbofuran, landrin, and methomyl. Numerous single-strand breaks were apparent in the DNA of all the nitroso derivative-treated cells but not in the DNA of those treated with the parent insecticides. Since the effect of the nitroso derivatives on the DNA could be observed for at least 20 h after removal of the chemical from the cultures, the DNA repairing events normally occurring in human cells after damage initiated by these chemical agents was not repaired as UV-type DNA damage or ionizing-type DNA damage in human cells. These observations suggest that the human cellular DNA in vivo is irreversibly altered by nitrosated N-methyl carbamate insecticides resulting in numerous alkali-sensitive bonds.

Metal concentrations in muscle of fish from aquatic systems in east Tennessee, U.S.A.

Heavy metal residues (i.e., As, Cd, Cu, Pb, Mn, Hg, and Zn) were determined in striated muscle of 268 fish specimens harvested during a 5-yr period (1980–1984) from several aquatic systems in east Tennessee (U.S.A.). Elevated concentrations of Hg, Mn, and Cd were found in the muscle of fish from several of the aquatic systems studied; concentrations of Hg exceeded the U.S. Food and Drug Administration action level of 1.0 ppm for food intended for human consumption. In general, the concentrations of the other metals in fish muscle were low. Moreover, muscle metal content did not vary among the three fish groups (i.e., game fish, catfish, and rough fish) investigated at any one of the nine sampling stations established. The results of this study are in agreement with the 1978–1979 pilot survey of Young and Blevins (1981) conducted at the same sampling stations. It appears that, in this region of Tennessee, heavy metal contamination of fish tissues has neither improved nor deteriorated during the last 5 yr.

2,3,7,8 Tetrachlorodibenzodioxin in fish from the Pigeon River of eastern Tennessee, USA: its toxicity and mutagenicity as revealed by the Ames Salmonella assay.

Levels of 2,3,7,8 tetrachlorodibenzodioxin (TCDD) were determined in both striated muscle (fillets) and whole body extracts of fish specimens harvested during a two-year period (1987–1989) from the Pigeon River (between Hartford and Newport) of Eastern Tennessee (USA). Whole body (wet weight) fish extract levels as high as 117 μg/kg body weight and composite fish fillet (wet weight) extract levels as high as 87 μg/kg fillet weight were observed. Pure TCDD was found to be highly toxic to theSalmonella typhimurium strains TA97, TA98, and TA100 at dosages which exceeded 825 ng TCDD/ml in the top agar of the Ames Salmonella assay. An 825 ng/ml TCDD dosage was not mutagenic to any of the tested Salmonella strains, either with or without metabolic activation (S9 mix). However, when acidic fish extracts from the Pigeon River were tested for mutagenicity, most of the fish extracts were mutagenic to Salmonella strains TA97, TA98, and TA100. These mutagenic extracts also demonstrated mutagenic dose-response curves. Other chemicals within the extracts as well as synergistic effects may account for the mutagenicity.

Fate of mutagenic activity during conventional treatment of municipal wastewater sludge

The mutagenic activity of wastewater was followed during conventional activated sludge treatment at a municipal plant. Raw wastewater was initially screened for mutagenic potential, using the AmesSalmonella/mammalian microsomal test and employer tester strains. The combined raw wastewater produced dose-related mutagenic responses in the presence of S9 metabolic activation. Raw wastewater from domestic sources alone was not mutagenic. Mutagenic activity was observed throughout the treatment process. Activated sludge prior to chlorination contained the highest specific mutagenic activity. Chlorination decreased the specific mutagenic activity at pH 11. Mutagenic activity in municipal wastewater containing industrial discharges is not removed by conventional treatment processes and can be enhanced by activated sludge treatment.

Chemically-induced histone modification as a predictor of carcinogenicity

The interaction between carcinogens and DNA is believed to initiate neoplastic transformation, but evidence suggests that epigenetic mechanisms may also be of importance. Because the histone proteins have important roles in chromatin structure and cellular function, they provide a reasonably well understood epigenetically-based system for the detection of carcinogens. In this study, human foreskin fibroblastic cells were exposed to one of several mutagens and/or carcinogens for 3, 12, or 24 h to determine if induced histone modification may be a means of predicting chemical carcinogenicity.Butyric acid (5 mM), known to result in acetylation of histones H3 and H4, and 12-O-tetradecanoylphorbol-13-acetate (3 μM), known to result in phosphorylated histone H1, were tested initially. Electrophoresis of the histone fractions was capable of resolving multiple forms of histones H1, H3, and H4.Propane sultone (0.1 mM) induced a broadening of the H2A and H2B bands after a 24 h exposure and carbon tetrachloride (1 mM) induced the formation of new histone forms in the H1 fraction after 24 h and in the H3 fraction after 3 h. Experimental variability limited the statistically significant modifications to carbon tetrachloride and propane sultone, two known carcinogens, where new forms of modified histone were detected. Therefore, the histone modification assay, with further experimentation, may be an alternate method of detecting carcinogens, especially when conventional genotoxic tests prove unreliable.

Organochlorine pesticides in gamebirds of Eastern Tennessee

Muscular tissue specimens of adult grouse, quail and woodcock from the rural counties of upper East Tennessee have been analyzed by gas chromatography for the organochlorine pesticides lindane, heptachlor, endrin, aldrin, dieldrin, DDT (total) and methoxychlor. Procedures for the analyses are described. The pesticidal levels in the muscle of all gamebirds exceeded the acceptable levels shown in governmental sources and professional journals. The observed concentrations constitute a serious threat to the wildlife of the region and permit accumulation and magnification even in man, who eats gamebirds as a part of his diet.

Bioaccumulation of mutagens and promutagens in fish taken from four sites in Northeast Tennessee

Abstract The purpose of this study was to identify the accumulation of mutagens and/or promutagens in fish using the Ames Salmenella/mammalian microsome assay. The fish were taken from four different collection sites in Northeast Tennessee area: Watauga Lake, Boone Lake, the Nolichucky River, and the Holston River. A gross autopsy vas performed on each fish; this autopsy consisted of recording weight, length, age, sex, and the physical condition of the various fish organs. Each fish (whole body) was extracted using ultra‐pure Z‐propanol as the solvent. Extracts were then screened for mutageniclty using bacterial Salmonella typhlmurlum tester strains TA97, TA98, TA100, and TA102. Two fish extracts showed mutagenicity (i.e., mutagenicity ratio of > 2.5) ‐ one from Boone Lake and the other from the Nollchucky River. These two sample extracts were retested at several doses. Results indicated that only the fish sample from the Nolichucky River displayed a dose response when using either strains TA97 or strain ...