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G B Nelson

Publications and source records attributed to G B Nelson.

17 recordsLinked to original sources
PubMed2001

An evaluation of the mutagenicity, metabolism, and DNA adduct formation of 5-nitrobenzo[b]naphtho[2,1-d]thiophene.

Thioarenes, sulfur-containing polycyclic aromatic compounds, are environmental contaminants suspected of posing human health risks. In this study, 5-nitrobenzo[b]naphtho[2,1-d]thiophene (5-nitro-BNT), a nitrated-thioarene, was examined for its mutagenicity, metabolism and subsequent formation of DNA adducts. 5-Nitro-BNT was weakly mutagenic in Salmonella typhimurium strains TA98 and TA100 without Aroclor-1254-induced rat liver S9 (S9), and its activity was increased in the presence of S9. Anaerobic metabolism of 5-nitro-BNT by S9 or xanthine oxidase (XO) produced one major metabolite, identified as 5-amino-BNT by NMR, MS, and UV spectroscopy and by comparison with an authentic standard. Aerobic S9 metabolism of 5-nitro-BNT produced a major metabolite, identified as trans-9,10-dihydroxy-9,10-dihydro-5-nitro-BNT (5-nitro-BNT-9,10-diol). Also present was a minor amount of 5-amino-BNT and trans-9,10-dihydroxy-9,10-dihydro-5-amino-BNT (5-amino-BNT-9,10-diol). DNA adduct analyses were performed using the (32)P-postlabeling assay and reversed-phase HPLC. Three major XO-derived calf thymus DNA adducts were detected. On the basis of their chromatographic mobilities, two adducts were identified as reaction products of 5-nitro-BNT with 2'-deoxyguanosine and one adduct with 2'-deoxyadenosine. Incorporation of allopurinol (a specific XO inhibitor) in the incubation mixture resulted in loss of all three adducts, confirming enzymatic mediation by XO. Aerobic S9 activation of 5-nitro-BNT with calf thymus DNA produced three adducts. On the basis of their chromatographic mobilities, two were identified as reaction products of 5-nitro-BNT with 2'-deoxyguanosine and one with 2'-deoxyadenosine. Incorporation of 1-aminobenzotriazole (a P450 inhibitor) in the incubation mixture resulted in a loss of these adducts, confirming enzymatic mediation by P450. Aerobic S9-catalyzed metabolism of 5-nitro-BNT-9,10-diol produced the same DNA adducts as observed with 5-nitro-BNT. Aerobic S9-catalyzed metabolism of 5-amino-BNT-9,10-diol produced the same deoxyadenosine-derived DNA adducts as observed with 5-nitro-BNT and 5-nitro-BNT-9,10-diol. These results provide additional information that both ring oxidation and nitroreduction are involved in the metabolism, DNA adduct formation and mutagenicity of 5-nitro-BNT.

Animals↗
PubMed2000

Biotransformation of trans-4,5-dihydroxy-4,5-dihydrobenzo[a]pyrene to benzo[a]pyrene bis-diols and DNA adducts by induced rat liver microsomes.

The biotransformation of (+/-)-trans-4,5-dihydroxy-4, 5-dihydrobenzo[a]pyrene (trans-B[a]P-4,5-diol), the K-region dihydrodiol of B[a]P, by beta-naphthoflavone (BNF)-induced rat liver microsomes was studied. trans-B[a]P-4,5-diol was metabolized to six major products as characterized by NMR, MS, and UV spectroscopy, and all were identified as bis-diols: two diastereomers of trans,trans-4, 5:7,8-tetrahydroxy-4,5:7,8-tetrahydrobenzo[a]pyrene (trans, trans-B[a]P-4,5:7,8-bis-diol), two diastereomers of trans,trans-4, 5:9,10-tetrahydroxy-4,5:9,10-tetrahydrobenzo[a]pyrene (trans, trans-B[a]P-4,5:9,10-bis-diol), and two diastereomers of the somewhat unusual trans,trans-1,2:4,5-tetrahydroxy-1,2:4, 5-tetrahydrobenzo[a]pyrene (trans,trans-B[a]P-1,2:4,5-bis-diol). BNF-induced rat liver microsomes also metabolized B[a]P to the same trans-B[a]P-4,5-diol-derived bis-diols. The ability of trans-B[a]P-4, 5-diol to form DNA adducts was investigated using (32)P-postlabeling techniques specifically designed to detect stable polar DNA adducts. Four DNA adducts were detected after microsomal activation of trans-B[a]P-4,5-diol with calf thymus DNA. Further analyses indicated that each of these stable polar DNA adducts was derived from the further metabolic activation of the trans,trans-B[a]P-4,5:7, 8-bis-diols. We conclude that trans-B[a]P-4,5-diol can be metabolized to a series of B[a]P-bis-diols, and can also be metabolically activated to form stable polar DNA adducts. The trans, trans-B[a]P-4,5:7,8-bis-diols were shown to be metabolic intermediates in the formation of these DNA adducts.

Animals↗
PubMed1997

Dibenzo[a,l]pyrene-induced DNA adduction, tumorigenicity, and Ki-ras oncogene mutations in strain A/J mouse lung.

Dibenzo[a,l]pyrene (DB[a,l]P), an environmental polycyclic aromatic hydrocarbon, is the most potent carcinogen ever tested in mouse skin and rat mammary gland. In this study, DB[a,l]P was examined for DNA adduction, tumorigenicity, and induction of Ki-ras oncogene mutations in tumor DNA in strain A/J mouse lung. Groups of mice received a single i.p. injection of 0.3, 1.5, 3.0, or 6.0 mg/kg DB[a,l]P in tricaprylin. Following treatment, DNA adducts were measured at times between 1 and 28 days, while tumors were counted at 250 days and analyzed for the occurrence of point mutations in codons 12 and 61 of the Ki-ras oncogene. DB[a,l]P in strain A/J mouse lung induced six major and four minor DNA adducts. Maximal levels of adduction occurred between 5 and 10 days after injection followed by a gradual decrease. DB[a,l]P-DNA adducts in lung tissue were derived from both anti- and syn-11,12-dihydroxy-13,14-epoxy- 11,12,13,14-tetrahydrodibenzo[a,l]pyrene (DB[a,l]PDE) and both deoxyadenosine (dAdo) and deoxyguanosine (dGuo) residues in DNA as revealed by cochromatography. The major adduct was identified as a product of the reaction of an anti-DB[a,l]PDE with dAdo in DNA. DB[a,l]P induced significant numbers of lung adenomas in a dose-dependent manner, with the highest dose (6.0 mg/kg) yielding 16.1 adenomas/mouse. In tricaprylin-treated control animals, there were 0.67 adenomas/mouse. Based on the administered dose, DB[a,l]P was more active than other environmental carcinogens including benzo[a]pyrene. As a function of time-integrated DNA adduct levels, DB[a,l]P induced lung adenomas with about the same potency as other PAHs, suggesting that the adducts formed by DB[a,l]P are similar in carcinogenic potency to other PAHs in the strain A/J mouse lung model. Analysis of the Ki-ras mutation spectrum in DB[a,l]P-induced lung tumors revealed the predominant mutations to be G-->T transversions in the first base of codon 12, A-->G transitions in the second base of codon 12, and A-->T transversions in the second or third base of codon 61, concordant with the DNA adduct profile.

Adenoma↗
PubMed1995

Adenomas induced by polycyclic aromatic hydrocarbons in strain A/J mouse lung correlate with time-integrated DNA adduct levels.

The induction of DNA adducts and adenomas in the lungs of strain A/J mice has been investigated following the single i.p. administration of each of the following polycyclic aromatic hydrocarbons (PAH): pyrene, dibenz[a,h]anthracene, benzo[a]pyrene, benzo[b]fluoranthene, 5-methylchrysene, and cyclopenta[c,d]pyrene. DNA adducts were measured by 32P-postlabeling at times between 1 and 21 days following injection, while adenomas were counted at 240 days after treatment. Pyrene did not induce either DNA adducts or lung adenomas at any of the doses examined. Each of the remaining PAH induced both adenomas and DNA adducts in a dose-dependent manner, with dibenz[a,h]anthracene > 5-methylchrysene > cyclopenta[c,d]pyrene > benzo[a]pyrene > benzo[b]fluoranthene. DNA adducts reached maximal levels between 3 and 9 days after injection, followed by a gradual decrease. The time-integrated DNA adduct level (TIDAL) was calculated by numerically integrating the areas under the adduct persistence curves extrapolated to 240 days for each PAH at each dose level. This value represents the effective total molecular dose of PAH that was delivered to the lung DNA over the entire course of tumorigenesis. A strong correlation of lung adenoma induction with the TIDAL values was observed for each PAH. The slopes of the tumors versus TIDAL value relationships were essentially identical for 5-methylchrysene, cyclopenta[cd]pyrene, benzo[a]pyrene, and benzo[b]fluoranthene. The slope of this relationship for dibenz[a,h]anthracene was markedly greater. The essentially identical induction of adenomas as a function of TIDAL values for these PAH suggests that the formation and persistence of DNA adducts determines their carcinogenic potency.

Adenoma↗
PubMed1994

Tumor multiplicity, DNA adducts and K-ras mutation pattern of 5-methylchrysene in strain A/J mouse lung.

This study was undertaken to evaluate the carcinogenic potential of 5-methylchrysene (5-MeC) in strain A/J mouse lung and to correlate the 5-MeC-DNA adduct profile in lung tissue with the mutation spectrum in the K-ras gene of lung tumors. Strain A/J mice received a single i.p. injection of 5-MeC at doses of 10, 50, 100 and 200 mg/kg and after 24, 48 and 72 h their lungs were collected for DNA adduct analysis. Eight months later, lungs from the remaining mice were harvested and the lung tumors counted and collected for subsequent mutational analysis of the K-ras gene. 5-MeC was found to be a potent lung carcinogen in strain A/J mice, inducing more than 100 tumors/mouse at a concentration of 200 mg/kg. Six 5-MeC-DNA adducts were observed; one adduct comigrated with the standard N2-deoxyguanosine adduct of 5-MeC-diol-epoxide I [1R,2S,3S-trihydroxy-4R-(N2-deoxy-guanosyl-3'-phosphate)- 1,2,3,4-tetrahydro-5-methyl-chrysene], derived from the bay-region diol-epoxide of 5-MeC. DNAs isolated from 5-MeC-induced lung tumors were evaluated for activating mutations in the K-ras gene by polymerase chain reaction-single strand conformation polymorphism and direct DNA sequencing analysis. Mutations were detected in 44 of 49 (90%) 5-MeC-induced tumors and the mutations were GGT-->TGT (50%), GGT-->GTT (23%) and GGT-->CGT (27%) in codon 12 of the gene. These results suggest that the N2-deoxyguanosine adduct of 5-MeC-diol-epoxide I may be one of the promutagenic adducts of 5-MeC in strain A/J mouse lung.

Animals↗
PubMed1992

Relationship between DNA damage, DNA repair, metabolic state and cell lethality.

Induction of unrepairable DNA damage, accumulation of misrepaired DNA damage, and generation of imbalances in competing biochemical and/or metabolic processes have been proposed to explain the relationship between radiation-induced DNA damage and cell lethality. Theoretically, the temperature dependence of the critical DNA repair process(es) should be 1) either independent of or identical to the temperature dependence of cell killing if the first two hypotheses are correct, and 2) different if the third hypothesis is correct. To test this, exponentially growing rat 9L brain tumor cells were left at 37 degrees C or equilibrated for 3-14 h at 20 degrees C before irradiation. Cells were irradiated and allowed to repair at either 20 degrees C or 37 degrees C. Alternatively, the cells were irradiated at one of these temperatures and immediately shifted to the other temperature for repair. DNA damage was assessed by the alkaline elution technique; cell kill was assessed by a clonogenic assay. 9L cells maintained at 20 degrees C or 37 degrees C sustained the same amount of DNA damage as measured by alkaline elution. DNA repair instantaneously assumed the rate characteristic of the postirradiation temperature. For 9L cells equilibrated, irradiated, and repaired at 20 degrees C, the half-time of the fast phase of the DNA repair decreased by a factor of approximately 2 and the half-time of the slow phase decreased by a factor of approximately 5 over that measured in cells incubated, irradiated and repaired at 37 degrees C. Although the rate of DNA repair decreased substantially at 20 degrees C, the survival of 9L cells that were equilibrated and irradiated at 20 degrees C was greater (p less than 10(-4)) than those incubated and irradiated at 37 degrees C, when assayed by an immediate plating protocol. In addition, the survival of 9L cells equilibrated and irradiated at 20 degrees C and then shifted to 37 degrees C immediately after irradiation was greater (p less than 10(-2)) than that obtained with any other delayed plating protocol. Thus, the temperature dependence of the DNA repair processes measured by alkaline elution was different from the temperature dependence of cell killing measured either by an immediate or delayed plating protocol. These data support the hypothesis that many irradiated 9L tumor cells die because of imbalances in sets of competing biochemical and/or metabolic processes.

Animals↗
PubMed1992

DNA adducts and induction of sister chromatid exchanges in the rat following benzo[b]fluoranthene administration.

Benzo[b]fluoranthene (B[b]F) was administered (100 mg/kg by i.p. injection) to male Sprague--Dawley rats. Lungs, livers and peripheral blood lymphocytes (PBLs) were harvested 1, 3, 5, 7, 14, 28 and 56 days after treatment. Several DNA adducts were observed in each tissue, with maximal levels occurring at approximately 7 days after treatment. Lung DNA exhibited consistently higher adduct levels than liver or PBL DNA. At 56 days after B[b]F administration, the adducts in liver and PBL DNA were present at < 10 amol/microgram DNA, while in lung there were 100 amoles/microgram DNA. No significant differences were observed between tissues in the types of adducts produced. Co-chromatography with synthetic standards showed that only a minor adduct produced in vivo is derived from trans-9,10-dihydro-9,10-dihydroxybenzo[b]fluoranthene-11,12-oxide. Sister chromatid exchanges (SCEs) from whole blood cultures were significantly increased relative to concurrent controls between 1 and 14 days after B[b]F administration, with maximum levels at 14 days. By 28 days after treatment, SCEs had essentially returned to control levels. SCE induction did not correlate with the amount of B[b]F--DNA adducts remaining in the PBLs at harvest time.

Animals↗
PubMed1991

Saturation of DNA repair measured by alkaline elution.

To determine whether the half-times (T1/2) of the DNA repair processes measured by alkaline elution increased in a dose-dependent manner, exponentially growing 9L/Ro rat brain tumor cells were irradiated with doses of 15-50 Gy, and their DNA repair kinetics was measured by alkaline elution. At 15 Gy, the DNA repair kinetics was biphasic with the fast phase having a T1/2 approximately 6 min and the slow phase having a T1/2 approximately 42 min. As the dose was increased to 50 Gy, the fast-phase T1/2 remained at approximately 6 min, but the slow-phase T1/2 increased to approximately 87 min. Although a dose-dependent increase in the T1/2 of the slow phase of DNA repair (saturation) was measured by alkaline elution, both the absolute value of the slow-phase T1/2 and the dependency of the slow-phase T1/2 on dose were less than those measured by alkaline sucrose gradient sedimentation in zonal rotors with slow reorienting gradient capability. Thus these two techniques appear either to depend on different hydrodynamic properties of the DNA or to have different coefficients of dependency for the same hydrodynamic properties of the DNA. The lower sensitivity for detection of the dose dependency of DNA repair makes it unlikely that the alkaline elution technique will be useful for quantitatively relating the shape of mammalian cell survival curves to the doses at which saturation of a DNA repair process occurs.

Animals↗
PubMed1990

Radiation-induced cytotoxicity, DNA damage and DNA repair: implications for cell survival theory.

The radiosensitivities and the kinetics for removal of radiation-induced DNA damage were compared for proliferative (P) and quiescent (Q) cells of the lines 66 and 67 derived from a mouse mammary adenocarcinoma. As determined from cell survival assays, the 66 and 67 Q cells were more radiosensitive than their 66 and 67 P counterparts. The rank order of their radiosensitivity was: 67 Q greater than 66 Q greater than or equal to 67 P greater than 66 P. Induction of radiation damage in the DNA of these cells, as measured by the alkaline elution technique, was identical for 66 and 67 P and Q cells. The repair of this DNA damage was biphasic for 66 and 67 P and Q cells. The half-times for the fast and slow repair phases in 66 Q cells were identical to those previously measured in 67 Q cells. The half-times of the fast and slow repair phases in 66 P cells were also identical to those previously measured in 67 P cells. However, the half-times for the fast and slow repair phases in 66 and 67 Q cells were longer than those measured in their 66 and 67 P counterparts. The 66 cell data are consistent with our previously published hypothesis that Q cells are more radiosensitive than their corresponding P cells because they repair their radiation-induced DNA damage slower. However, our results are not consistent with hypotheses that attempt to explain the radiosensitivity differences between lines 66 and 67 solely on the basis of measurable induction and repair of DNA damage.

Animals↗
PubMed1988

Regulation of DNA repair kinetics in proliferative and quiescent tumor cells.

The radiosensitivity and kinetics of repair of radiation-induced DNA damage were determined for proliferative (P) and quiescent (Q) cells of the mouse mammary adenocarcinoma line 67. 67 Q cells are more radiosensitive than 67 P cells. Radiation induced the same amount of DNA damage in both 67 P and 67 Q cells. Both 67 P and 67 Q cells repaired their DNA damage with biphasic kinetics, but the half-times for the fast and slow phase were longer in 67 Q cells. Q cell DNA appeared to be in a more compact or condensed chromatin structure and was less accessible to enzymatic digestion than P cell DNA. These data suggest that 67 Q cells are more sensitive to ionizing radiation than 67 P cells because they repair their radiation-induced DNA damage more slowly, perhaps as a result of their more condensed chromatin structure.

Adenocarcinoma↗
PubMed1988

Postirradiation alterations of neuronal chromatin structure.

Previous work from our laboratory suggested that neuronal chromatin structure may be altered immediately after exposure to ionizing radiation. In the present study, whole brains of 4-month-old male Fisher 344 rats were irradiated with a dose of 25 Gy. The kinetics of restoring the chromatin structure to its unirradiated state was investigated in rat cerebellar neurons using three different approaches: (1) measurement of changes in the DNA superhelical structure by the fluorescent halo assay, (2) measurement of changes in chromatin accessibility to digestion by micrococcal nuclease, and (3) measurement of changes in the accessibility of the nuclear-matrix-associated DNA to digestion by DNase I. Immediately after irradiation, the topological constraints on the DNA loops were altered, the chromatin was more accessible to m. nuclease digestion, and the DNA associated with the nuclear matrix was more resistant to digestion by DNase I. Return of the chromatin structure to its unirradiated state as measured by each of the three methods followed biphasic kinetics with the fast phase having a half-time of several minutes and the slow phase having a half-time of several hours. The kinetics are similar to that previously reported for repair of radiation-induced DNA damage in mammalian cells. Although the independent assays used in this study seemed to follow the same kinetics, their relationship at the molecular level remains to be determined.

Animals↗
PubMed1987

Saturation of a DNA repair process in dividing and nondividing mammalian cells.

We have reported that the half-time (T1/2) for the slow phase of repair of radiation-induced DNA lesions increased with dose from 12.5 to 50.0 Gy in both 9L rat brain tumor cells and cerebellar neurons. In this manuscript these studies have been extended to lower doses to determine at what dose this DNA repair process becomes unsaturated. Our alkaline sucrose sedimentation technique in zonal rotors limits determination of the slow phase T1/2 to doses of greater than or equal to 6 Gy for 9L tumor cells and greater than or equal to 4 Gy for cerebellar neurons. The slow phase T1/2 in cerebellar neurons appears constant at doses less than or equal to 6 Gy and then increases exponentially at higher doses; survival of whole brain irradiated rats does not begin to decrease until doses greater than 6 Gy. The slow phase T1/2 in 9L tumor cells is either saturated or just becoming saturated at 6 Gy; a dose just before the final slope of the 9L survival curve is reached. These data support the hypothesis that the shape of mammalian cell survival curves is related to the saturation of a DNA repair process.

Animals↗