摘要:

AbstractWe have established a comprehensive procedure based on the polymerase chain reaction (PCR) to analyze the molecular spectrum of mutations at the hypoxanthine‐guanine phosphoribosyltransferase (hprt) locus in Chinese hamster cells. The procedure includes direct sequencing of PCR‐amplifiedhprtcDNA for locating point mutations in the expressed coding sequences, multiplex PCR amplification of thehprtexons for screening large deletions, and direct sequencing of PCR‐amplifiedhprtexons and their flanking regions for detecting intronic mutations resulting in mRNA splicing errors. Using this procedure, we have identified different types of mutations among a representative collection of spontaneous and induced HPRT‐deficient Chinese hamster cell mutants. This procedure is simple, rapid, accurate, and practical for a comprehensive study of the mutation spectrum in a large number of HPRT‐deficient Chinese hamster cel

ISSN:0893-6692    

DOI:10.1002/em.2850190402

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractA prospective, longitudinal study was performed to test the hypothesis that environmental factors (e.g., diet or cigarette smoking) modulate genetic damage caused by treatment for breast cancer and render these women more susceptible to developing second malignancies. A total of 107 women (49 with breast cancer, 52 with benign breast masses, and 6 normal women) were enrolled. This report describes initial studies at the time of enrollment and disease presentation. Mutant frequency at thehprtlocus and cloning efficiency of peripheral blood lymphocytes did not differ significantly among the 3 groups. Mutant frequency increased with age, with a history of cigarette smoking, and with the number of years that current smokers used cigarettes. There was no correlation in women with benign masses between mutant frequency and the incidence of chromosome aberrations (28 women) or sister chromatid exchanges (23 women). A maternal history of breast cancer did not influence mutant frequency. There was no significant relationship between dietary intake of vitamins A, B12, C and E, folacin, selenium, calcium, caffeine, or multivitamin pills, and mutant frequency. Serum folate levels in the deficient range were associated (P = 0.02) with elevated mutant frequencies, whereas SCE rates inversely correlated with serum vitamin B12 levels. These results confirm the importance of age and, less so, cigarette smoking as factors that influence mutant frequency and suggest that a micronutrient, folic acid, may modify genetic damage at thehprtlocus. To the extent that somatic mutation contributes to carcinogenesis, these environmental factors may enhance the risk of developing malignant transformation.

ISSN:0893-6692    

DOI:10.1002/em.2850190403

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractSUP4‐o, a suppressor tRNA gene, is the target in a system for characterizing mutational specificity in the yeastSaccharomyces cerevisiae.To date, screening for loss of suppression has detected 172 of the 267 base‐pair substitutions possible in the exons and intron of theSUP4‐o gene. Although many of the remaining 95 changes might not be detected by this screen, some might occur spontaneously, or be induced, only at very low frequencies. For the purpose of analyzing mutational specificity, it would be valuable to determine which of these substitutions can be detected with the genetic screen employed in this system and which cannot. Thus we used in vitro mutagenesis to generate the 95 substitutions not yet detected inSUP4‐o in vivo. Assessment of the phenotypes conferred by these 95 directed mutations revealed that 50 would completely escape detection and only 45 would pass through the first stage of the screen. Of these 45 substitutions, 2 are detectable but have not yet been found among more than 5,000 characterizedSUP4‐o mutations that arose in vivo. In addition, 4 others should be detected by slightly relaxing the current criteria for selection ofSUP4‐o mutants. The results indicate that with these modifications the system can detect 174/225 substitutions possible in theSUP4‐o exons and 4/42

ISSN:0893-6692    

DOI:10.1002/em.2850190404

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractIn this work we report on the isolation of anEscherichia coliK‐12 mutation, which confers a high sensitivity to bacteria cells to mutagenesis by simple monofunctional alkylating agents. The mutation emerged spontaneously from a bacterial strain that already proved useful in various mutagenicity studies. By monitoring the influence of such a mutation on the frequency of induced mutation by ethylating (EMS, DES, ENU, ENNG) vs. methylating (MMS, DMS, MNU, MNNG) compounds, and on the in vivo repair capacity for different alkyl‐DNA lesions (O6‐alkG, N7‐alkG, N3‐meA), we conclude that the mutation should affect the gene (ogt) that encodes constitutive DNA repair alkyltransferase (ATase). Thus in the presence ofada, differences in mutagenicity were observed only with ethylating agents; the sensitization of cells to both the ethylating and methylating partners requiring, by contrast, the absence of theadaprotein. These results support the reported in vitro substrate specificities for bothogtandadaATases. The parental cells exhibited biphasic dose‐response curves in accordance with the idea of low basal level saturation attributed to the uninducibleogtATase. Deficient bacterial derivatives showed, by contrast, linear mutation induction responses. The in vivo removal of alkylated bases from DNA was measured in bacterial strains deficient in the excision repair pathway (ΔuvrB) and unable to induce the adaptive response (ada::Tn10). The very low initial levels for O6‐meG and O6‐etG (1.1 and 0.2 molecules per cell, respectively) were readily repaired by the parental cells but remained unchanged in the hypermutable derivatives. This result suggests that in the absence of nucleotide excision repair and of the adaptive response, no alternative pathway, other thanogt, is available for the repair of the major mutagenic lesion, O6‐alkG, at least during the first 4 hours after alkylation. Comparatively, no differences were found in the capacity to repair the major lethal adduct, N3‐meA, in agreement with the fact that no effect on cell survival was detected. In conclusion, we propose that the biological significance of theogtprotein relies mainly on its ability to prevent mutagenesis by low levels of bulkier ethylation products (especially in the absence ofuvrexcision repair). This is in accordance with the relative inefficiency of these agents as inducers of the adaptive response, as well as the relative inefficiency of theadaATase in dealing with alkylation products longer than methyl adducts. Theogtprotein provided additional resistance to mutation by low doses of methylating agents, as it was observed inadadeficient bacteria. Thus the broad substrate specificity ofogtATase would appear to be also of

ISSN:0893-6692    

DOI:10.1002/em.2850190405

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractFluorescence‐activated cell sorting (FACS) was used to establish clonal populations of a human lymphoblastoid cell strain that contain spontaneously occurring and N‐methyl‐N'‐nitro‐N‐nitrosoguanidine‐induced mutations in the lambda immunoglobulin gene. Multiple rounds of FACS using a monoclonal antibody specific for the membrane‐expressed human lambda immunoglobulin were used to enrich the population fraction of cells lacking a wild‐type lambda immunoglobulin on the cell surface. Approximately 20% of the clonal populations established after five rounds of FACS‐mediated enrichment did not express the lambda immunoglublin epitope recognized by the monoclonal antibody used for selection. However, evaluation of the FACS‐selected mutant clonal populations with a polyclonal antilambda antibody, or a monoclonal antibody directed against a different epitope on the lambda immunoglobulin made by the T5‐1 cells, indicated that the mutant clonal populations expressed lambda immunoglobulin on their cell surfaces. Additionally, the presence of lambda mRNA and of both secreted and cytoplasmic lambda protein confirmed the transcription and translation of the lambda immunoglobulin gene. These data suggest that FACS‐mediated selection employing epitope‐specific monoclonal antibodies provides a powerful technique for isolation of cell populations that express mutations within the coding region of the

ISSN:0893-6692    

DOI:10.1002/em.2850190406

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractHuman cytomegalovirus (HCMV) has been shown to increase the frequency of chromosome aberrations, primarily chromatid‐type, in human peripheral blood lymphocytes (PBLs). Because HCMV persists in most humans, pathologically activates cells, and may perturb the cell cycle, we investigated the possibility that HCMV‐infected cells have a modified sensitivity to chromosome damage induced by genotoxic chemicals. Uninfected PBLs exposed to bleomycin (3 to 100 μg/ml) demonstrated a linear increase in the frequency of chromosome aberrations. HCMV infection of PBLs at an intensity that did not cause detectable damage followed by exposure to the same concentrations of bleomycin resulted in a significant enhancement (p<0.01) in the frequency of chromosome aberrations relative to the effect of bleomycin alone. A more than additive enhancement of the frequency of chromosome aberrations was also noted in HCMV‐infected PBLs exposed to 4‐hydroxyaminoquinoline‐1‐oxide (4‐HAQO; 0.1 to 0.3 μg/ml) relative to uninfected cells treated with 4‐HAQO alone. No increase in the percentage of aberrant cells or the frequency of chromosome aberrations was observed in HCMV‐infected cells treated with 4‐nitroquinoline‐1‐oxide (4‐NQO) relative to similarly treated uninfected PBLs. These results suggest that HCMV can potentiate the induction of chromosome aberrations in human PBLs caused by

ISSN:0893-6692    

DOI:10.1002/em.2850190407

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractSome 2‐bromo‐propanamides were prepared and tested for direct mutagenicity inSalmonella typhimuriumTA 100. Results confirm the mutagenic activity of 2‐bromo‐N‐benzyl‐propanamide and indicate that it is independent of enantiomeric configuration. A variation in the chemical structure, namely, the addition of a methyl group at the benzylic carbon, causes the four resulting diastereomers to be devoid of any activity. Conversely, some racemic ring‐substituted methoxy and/or hydroxy derivatives of the parent compound displayed mutagenic properties, causing an increase in the number ofhis+revertants up to 524 per millig

ISSN:0893-6692    

DOI:10.1002/em.2850190408

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractThe hibernating female Turkish hamster (Mesocricetus brandti) was utilized for a study of possible in vivo effects of cold on oocyte maturation. Such a physiologic model offered an opportunity to analyze the ability of oocytes exposed to prolonged periods of reduced core temperature and/or light to subsequently mature to Metaphase II. Detailed observations of core temperatures, torpor/arousal, serum estradiol, and ovarian histology were made. An average incidence of 37.7% binuclearity was found in the germinal vesicle, metaphase I and II occytes of this species. Maturation to Metaphase II of total chromosome complements did not vary significantly in the experimental groups compared with the control, but aneuploidy was detected in the oocytes of animals exposed to reduced temperature or light. An effect of in vivo reduced core temperatures on oocyte chromosome complements validates many of the published in vitro studies with temperature reduction. The model presents an excellent physiologic system for perturbing and analyzing many aspects of mammalian oocyte development.

ISSN:0893-6692    

DOI:10.1002/em.2850190409

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractFenfluramine, an amphetamine derivative used in the treatment of obesity, has been evaluated in vivo in the bone marrow cells of Swiss albino mice using two cytogenetic endpoints for assessing its genotoxic and clastogenic potentials. Concentrations of 0.75, 1.5, 3.0, and 5.0 mg/kg b.w. were administered orally for the study of sister chromatid exchange frequencies and chromosome aberrations (CA). SCE frequencies showed a positive dose response; 1.5 mg/kg being the minimum effective concentration. Fen caused a prolongation of cell cycle at all concentrations. Except for the minimum therapeutic dose (0.75 mg), all other doses (1.5, 3.0, and 5.0 mg) showed a significant increase in the percentage of damaged cells over that of the vehicle control. The degree of clastogenicity was directly proportional to the dosage used and inversely related with the duration of treatment. A gradual reduction of the clastogenic potential was observed after 12 and 24 hr of exposure, indicating that the maximum effect occurs at the middle or late synthetic phase of the cell cycle. This study, probably the first detailed screening of the drug for its genotoxicity, shows that Fen is moderately clastogenic and a DNA damaging agent in vivo.

ISSN:0893-6692    

DOI:10.1002/em.2850190410

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY

摘要:

AbstractThe phenoxyherbicide and peroxisome proliferator 2‐methyl‐4‐chlorophenoxyacetic acid (MCPA) was tested for its ability to induce sister‐chromatid exchanges (SCE) in chick embryos. Erbitox E30 (a commercial formulation containing 28% MCPA sodium potassium salt as active ingredient) was injected into the air chamber in concentrations of MCPA of 0, 0.35, 0.7, 1.4, 2.8, or 5.6 mg/egg on day 0 of incubation. Pure MCPA sodium salt was tested at 2.8 mg/egg. Neutral red at 0.25 mg/egg was the mutagenic reference compound (positive control group). Eggs were then incubated for 4 days. MCPA induced a slight but significant increase in SCE frequency (about 1.3 times base line) at 2.8 mg/egg. The dose of 5.6 mg/egg was toxic. No difference in genetic activity between the commercial formulation and the pure compound was found. A cell cycle delaying effect of MCPA was evident at all the dose levels tested. The mitotic index remained un

ISSN:0893-6692    

DOI:10.1002/em.2850190411

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1992

数据来源: WILEY