DNA strand breakage repair in ataxia telangiectasia fibroblast-like cells

Energy Technology Data Exchange (ETDEWEB)

Vincent, Jr, R A; Sheridan, III, R B; Huang, P C [Johns Hopkins Univ., Baltimore, Md. (USA). Dept. of Environmental and Biophysical Sciences

1975-12-01

Human diploid fibroblast-like cells derived from four patients with the genetic disease ataxia telangiectasia and from two non-mutant donors were examined for the repair of x-ray induced strand breaks in DNA. The ataxia telangiectasia cultures showed no significant differences from the non-mutant cultures in the kinetics and extent of strand repair. This suggests that the increased spontaneous and x-ray induced chromatid aberrations observed in ataxia telangiectasia cells are not caused by a defect in the repair of single strand breaks as might be suspected from a general model of aberration production.

125I-induced DNA double strand breaks: use in calibration of the neutral filter elution technique and comparison with X-ray induced breaks

International Nuclear Information System (INIS)

Radford, I.R.; Hodgson, G.S.

1985-01-01

The neutral filter elution assay, for measurement of DNA double strand breakage, has been calibrated using mouse L cells and Chinese hamster V79 cells labelled with [ 125 I]dUrd and then held at liquid nitrogen temperature to accumulate decays. The basis of the calibration is the observation that each 125 I decay, occurring in DNA, produces a DNA double strand break. Linear relationships between 125 I decays per cell and lethal lesions per cell (minus natural logarithm survival) and the level of elution, were found. Using the calibration data, it was calculated that the yield of DNA double strand breaks after X-irradiation of both cell types was from 6 to 9 x 10 -12 DNA double strand breaks per Gy per dalton of DNA, for doses greater than 6 Gy. Neutral filter elution and survival data for X-irradiated and 125 I-labelled cells suggested that the relationships between lethal lesions and DNA double strand breakage were significantly different for both cell types. An attempt was made to study the repair kinetics for 125 I-induced DNA double strand breaks, but was frustrated by the rapid DNA degradation which occurs in cells that have been killed by the freezing-thawing process. (author)

Multiple pathways of DNA double-strand break processing in a mutant Indian muntjac cell line

International Nuclear Information System (INIS)

Bouffler, S.D.; Jha, B.; Johnson, R.T.

1990-01-01

DNA break processing is compared in the Indian muntjac cell lines, SVM and DM. The initial frequencies and resealing of X-ray generated single- and double-strand breaks are similar in the two cell lines. Inhibiting the repair of UV damage leads to greater double-strand breakage in SVM than in DM, and some of these breaks are not repaired; however, repair-associated single-strand breakage and resealing are normal. Dimethylsulfate also induces excess double-strand breakage in SVM, and these breaks are irreparable. Restricted plasmids are reconstituted correctly in SVM at approximately 30% of the frequency observed in DM. Thus SVM has a reduced capacity to repair certain types of double-strand break. This defect is not due to a DNA ligase deficiency. We conclude that DNA double-strand breaks are repaired by a variety of pathways within mammalian cells and that the structure of the break or its mode of formation determines its subsequent fate

Fragmentation in DNA double-strand breaks

International Nuclear Information System (INIS)

Wei Zhiyong; Suzhou Univ., Suzhou; Zhang Lihui; Li Ming; Fan Wo; Xu Yujie

2005-01-01

DNA double strand breaks are important lesions induced by irradiations. Random breakage model or quantification supported by this concept is suitable to analyze DNA double strand break data induced by low LET radiation, but deviation from random breakage model is more evident in high LET radiation data analysis. In this work we develop a new method, statistical fragmentation model, to analyze the fragmentation process of DNA double strand breaks. After charged particles enter the biological cell, they produce ionizations along their tracks, and transfer their energies to the cells and break the cellular DNA strands into fragments. The probable distribution of the fragments is obtained under the condition in which the entropy is maximum. Under the approximation E≅E 0 + E 1 l + E 2 l 2 , the distribution functions are obtained as exp(αl + βl 2 ). There are two components, the one proportional to exp(βl 2 ), mainly contributes to the low mass fragment yields, the other component, proportional to exp(αl), decreases slowly as the mass of the fragments increases. Numerical solution of the constraint equations provides parameters α and β. Experimental data, especially when the energy deposition is higher, support the statistical fragmentation model. (authors)

The sensitivity of active and inactive chromatin to ionizing radiation-induced DNA strand breakage

International Nuclear Information System (INIS)

Chiu, S.-M.; Oleinick, N.L.

1982-01-01

The sensitivity of DNA in actively transcribing and inactive states has been compared with regard to γ-radiation-induced single-strand break (SSB) induction. The results indicate that chromatin organization is important in the determination of the sensitivity of cellular DNA toward γ-radiation: Not only the yield but also the rate of repair of SSB is greater in the actively transcribing genes than in the total nuclear DNA. (author)

Formation of DNA single-strand breaks by near-ultraviolet and gamma-rays in normal and Bloom's syndrome skin fibroblasts

International Nuclear Information System (INIS)

Hirschi, M.; Netrawali, M.S.; Remsen, J.F.; Cerutti, P.A.

1981-01-01

The formation of single-strand breaks by near-ultraviolet light at 313 nm and by aerobic gamma-rays was compared for skin fibroblast monolayer cultures from 4 normal donors (NF) and 8 patients with Bloom's syndrome (BS) by the alkaline elution method. In 6 of 8 BS strains, the number of breaks induced by near-ultraviolet light, 2.25 kJ/sq m, at 0 degrees was comparable to NF, while elevated breakage was observed in BS strains HG 369 and HG 916. Breakage frequencies were increased substantially in 6 of 8 BS strains relative to NF when the near-ultraviolet light exposure was at 37 degrees. BS strain GM 2520 represents an exception since normal breakage frequencies were induced both at 0 degrees and 37 degrees. Aerobic gamma-rays (75 R) induced comparable numbers of single-strand breaks in BS and NF strains at 0 degrees. The breakage frequencies were reduced an average of 17% in NF when the same dose was given at 30 degrees followed by 6 min incubation. Under the same conditions, the breakage frequencies were on the average reduced by 42% relative to 0 degrees in the BS strains, indicating that they possess normal or possibly slightly increased capacities for the rejoining of gamma-ray-induced breaks

Increased rate of repair of ultraviolet-induced DNA strand breaks in mitogen stimulated lymphocytes

Energy Technology Data Exchange (ETDEWEB)

Hamlet, S.M.; Lavin, M.F.; Jennings, P.A. (Queensland Univ., St. Lucia (Australia). Dept. of Biochemistry; Queensland Univ., St. Lucia (Australia). Dept. of Veterinary Pathology; Queensland Univ. St. Lucia (Australia). Dept. of Public Health)

1982-05-01

Previous results have shown that phytohaemagglutinin-stimulated bovine lymphocytes exhibit a peak of ultraviolet-induced DNA repair synthesis 3 to 4 days after addition of mitogen. The level of repair synthesis was approximately tenfold higher than that in unstimulated lymphocytes. These studies have been extended to examine the rate of repair of strand breaks in U.V.-irradiated bovine lymphocytes. The extent of breakage of DNA was shown to be the same in mitogen-stimulated and unstimulated lymphocytes from two breeds of cattle, when determined by sedimentation of nucleoids on sucrose gradients. However, in mitogen-stimulated cells the time taken to repair DNA strand breaks was 6 hours compared with 12 hours in stationary phase lymphocytes after a U.V. dose of 5 J/m/sup 2/. These results suggest that the increased rate of repair of strand breaks is due to the induction of enzymes involved at the post-incision stage of DNA repair. Thus the increased level of repair synthesis observed in earlier work correlates with an increased rate of repair of DNA strand breaks in phytohaemagglutinin-stimulated bovine lymphocytes.

Increased rate of repair of ultraviolet-induced DNA strand breaks in mitogen stimulated lymphocytes

International Nuclear Information System (INIS)

Hamlet, S.M.; Lavin, M.F.; Jennings, P.A.; Queensland Univ., St. Lucia; Queensland Univ. St. Lucia

1982-01-01

Previous results have shown that phytohaemagglutinin-stimulated bovine lymphocytes exhibit a peak of ultraviolet-induced DNA repair synthesis 3 to 4 days after addition of mitogen. The level of repair synthesis was approximately tenfold higher than that in unstimulated lymphocytes. These studies have been extended to examine the rate of repair of strand breaks in U.V.-irradiated bovine lymphocytes. The extent of breakage of DNA was shown to be the same in mitogen-stimulated and unstimulated lymphocytes from two breeds of cattle, when determined by sedimentation of nucleoids on sucrose gradients. However, in mitogen-stimulated cells the time taken to repair DNA strand breaks was 6 hours compared with 12 hours in stationary phase lymphocytes after a U.V. dose of 5 J/m 2 . These results suggest that the increased rate of repair of strand breaks is due to the induction of enzymes involved at the post-incision stage of DNA repair. Thus the increased level of repair synthesis observed in earlier work correlates with an increased rate of repair of DNA strand breaks in phytohaemagglutinin-stimulated bovine lymphocytes. (author)

Aphidicolin synchronization of mouse L cells perturbs the relationship between cell killing and DNA double-strand breakage after X-irradiation

International Nuclear Information System (INIS)

Radford, I.R.; Broadhurst, S.

1988-01-01

The relationship between X-ray-induced cell killing and DNA double-strand breakage was examined for synchronized mouse L cells that had entered S-phase, G2-phase, mitosis, and G1-phase following release from aphidicolin and compared to asynchronous culture response. Aphidicolin-synchronized cells showed cycle phase-dependent changes in dose-responses for both killing and DNA dsb. However, on the basis of DNA dsb per unit length of DNA required to produce a lethal lesion, aphidicolin-synchronized cells were more sensitive to X-rays than asynchronous cultures. This sensitivity peaked 2 h after release from aphidicolin treatment, and then progressively declined towards the asynchronous culture value. It is argued that results are due to deregulation of the temporal order of DNA replication following aphidicolin treatment, and can be incorporated into the critical DNA target size model by postulating that the targets for radiation action in mammalian cells are DNA-associated with potentially transcriptionally active proto-oncogenes or constitutive fragile sites. (author)

Monophosphate end groups produced in radiation induced strand breakage in DNA

International Nuclear Information System (INIS)

Kay, E.; Ward, J.F.

1976-01-01

A solution of DNA was gamma-irradiated and treated with monophosphatase for studies on the amount of inorganic phosphate released as a function of time. Studies were also conducted on: effect of alkali on yield of monophosphate end groups; induction of DNA strand breaks by treatment with DNAase; initial G values for monophosphate termini; and effect of alkali on radioinduced DNA damage

Adaptive responses on chromosome aberration and DNA breakage of peripheral lymphocytes from workers exposed to thorium and rare earth mixed dust in Baotou steel plant

International Nuclear Information System (INIS)

Liu Qingjie; Feng Jiangbing; Lu Xue; Chen Deqing; Lv Huimin; Su Xu; Liu Yufei; Jia Kejun

2008-01-01

Objective: To explore if the occupational exposure to low dose thorium could induce adaptive response in peripheral lymphocytes. Methods: 40 individuals, who exposed to thorium and rare earth mixed dust (exposure group) or control in Baotou Steel Plant, were selected, and chromosome aberrations were analyzed. Then the peripheral blood samples were irradiated in vitro with 2 Gy 60 Co γ-rays, and unstable chromosome aberration or DNA stand breakage analysis using single cell gel electrophoresis was performed. Results: The dicentrics before 2 Gy exposure in exposure group was higher than that in control (P>0.05). But the dicentrics after 2 Gy exposure in exposure group was lower than that in control, but not significantly (P >0.05). The tricentrics in exposure group was significantly lower than that in control (U=3.1622, 0.001< P<0.002). The DNA strand breakage in control group was significantly higher than that in exposure group (t=25, P<0.001). Conclusions: Occupational exposure to low dose thorium could induce the adaptive response on chromosome aberration and DNA strand breakage in peripheral lymphocytes. (authors)

Resveratrol protects mouse embryonic stem cells from ionizing radiation by accelerating recovery from DNA strand breakage.

Science.gov (United States)

Denissova, Natalia G; Nasello, Cara M; Yeung, Percy L; Tischfield, Jay A; Brenneman, Mark A

2012-01-01

Resveratrol has elicited many provocative anticancer effects in laboratory animals and cultured cells, including reduced levels of oxidative DNA damage, inhibition of tumor initiation and progression and induction of apoptosis in tumor cells. Use of resveratrol as a cancer-preventive agent in humans will require that its anticancer effects not be accompanied by damage to normal tissue stem or progenitor cells. In mouse embryonic stem cells (mESC) or early mouse embryos exposed to ethanol, resveratrol has been shown to suppress apoptosis and promote survival. However, in cells exposed to genotoxic stress, survival may come at the expense of genome stability. To learn whether resveratrol can protect stem cells from DNA damage and to study its effects on genomic integrity, we exposed mESC pretreated with resveratrol to ionizing radiation (IR). Forty-eight hours pretreatment with a comparatively low concentration of resveratrol (10 μM) improved survival of mESC >2-fold after exposure to 5 Gy of X-rays. Cells pretreated with resveratrol sustained the same levels of reactive oxygen species and DNA strand breakage after IR as mock-treated controls, but repaired DNA damage more rapidly and resumed cell division sooner. Frequencies of IR-induced mutation at a chromosomal reporter locus were not increased in cells pretreated with resveratrol as compared with controls, indicating that resveratrol can improve viability in mESC after DNA damage without compromising genomic integrity.

Amifostine Protection Against Mitomycin-induced Chromosomal Breakage in Fanconi Anaemia Lymphocytes

Directory of Open Access Journals (Sweden)

Miriam T. P. Lopes

2008-08-01

Full Text Available Fanconi anaemia (FA is a rare genetic chromosomal instability syndrome caused by impairment of DNA repair and reactive oxygen species (ROS imbalance. This disease is also related to bone marrow failure and cancer. Treatment of these complications with radiation and alkylating agents may enhance chromosomal breakage. We have evaluated the effect of amifostine (AMF on basal and mitomycin C (MMC-induced chromosomal breakage in FA blood cells using the micronucleus assay. The basal micronuclei count was higher among FA patients than healthy subjects. Pre-treatment with AMF significantly inhibited micronucleation induced by MMC in healthy subjects (23.4 ± 4.0 – MMC vs 12.3 ± 2.9 – AMF →MMC MN/1000CB, p < 0.01, one way ANOVA as well as in FA patients (80.0 ± 5.8 – MMC vs 40.1 ± 5.8 – AMF →MMC MN/1000CB, p < 0.01, ANOVA. Release of ROS by peripheral blood mononuclear cells treated with AMF →MMC and measured by chemoluminometry showed that AMF-protection was statistically higher among FA patients than in healthy individuals. Based on these results we suggest that AMF prevents chromosomal breakage induced by MMC, probably by its antioxidant effect.

Investigations into the molecular mechanism of chromatid breakage in the G2-phase of mammalian cells

International Nuclear Information System (INIS)

Bryant, P.E.; Armstrong, G.N.; Gray, L.; Frankenberg, D.; Mozdarani, H.

2003-01-01

Chromatid breakage following irradiation of cells in the G2-phase of the cell cycle results from the induction of DNA double-strand breaks (dsb). The conversion of dsb into chromatid breaks (cb) has a genetic basis, seemingly different from that of dsb rejoining. The variation in extent of this conversion is exemplified by the stiking variation in frequency of cb in irradiated cycling T-lymphocytes between different normal individuals. Elevated cb frequency in lymphocytes of around 40% of breast cancer patients and their first-degree relatives suggests the presence of mutations in low penetrance cancer predisposing genes that also affect conversion of dsb to cb. Investigation of the mechanism of chromatid radiosensitivity using genetically engineered rodent cell lines containing unique dsb break sites indicate that a single isolated dsb is sufficient to cause a cb. The single-event nature of chromatid breakage is confirmed by the fact that cb are induced as a linear function of radiation dose. Moreover, we have recently shown that ultrasoft carbon-K X-rays also induce chromatid breakage. In this case the energy of the secondary electrons produced by carbon-K X-rays is too low to span more than one DNA double helix, thus further supporting our conclusion that a single dsb is responsible for the formation of a cb. Chromatid breakage is thought to involve a rearrangement between DNA strands at the crossover points of chromatin loop(s) triggered by the presence of a dsb within the loop structure. The occasional observation of 'looped-out' sections of chromatin at cb sites supports this hypothesis. The occurrence of 'colour-switches' between FPG stained chromatids at a proportion of break sites (e.g. about 16% in CHO cells) shows that a significant proportion of cb definitely result from chromatin rearrangements. Measurements of altered colour-switch ratio (csr) in mutant rodent and human cells (irs1 and AT cells respectively) also indicate a genetic basis for the

Phleomycin-induced lethality and DNA degradation in Escherichia coli K12

Energy Technology Data Exchange (ETDEWEB)

Nakayama, H

1975-01-01

The cell lethality and DNA fragmentation caused by phleomycin (PM) were studied in E. coli K12 strains with special reference to the effects of repair or recombination deficiencies and metabolic inhibitors. Unlike excision-defective derivatives of E. coli B, uvrA, uvrB, and uvrC mutants of strain K12 showed no peculiarities compared with wild type in regard to cell survival. Likewise, mutant alleles at uvrD and polA loci had no effect. In contrast, rec mutants were more sensitive to PM-killing than were rec/sup +/ strains. PM-induced strand breakage in DNA was observed in all strains tested including the above-mentioned mutants. There was no significant distinction between the uvr mutants and the wild type strain, indicating that the uvr-endonuclease was not responsible for the strand breaks. Involvement of endonuclease I was also ruled out. At least some of the PM-induced strand breaks were repairable. PM-induced lethality and strand breakage were totally dependent on energy supply. Inhibition of protein synthesis resulted in a partial and parallel suppression of the two effects. Our results suggest that the lethality is due to DNA strand breakage and the repair of such damage is postulated to be controlled by rec genes.

Chromosomal instability can be induced by the formation of breakage-prone chromosome rearrangement junctions

International Nuclear Information System (INIS)

Allen, R.N.; Ritter, L.; Moore, S.R.; Grosovsky, A.J.

2003-01-01

Full text: Studies in our lab have led to the hypothesis that chromosomal rearrangements can generate novel breakage-prone sites, resulting in chromosomal instability acting predominantly in cis. For example, specific breakage of large blocks of centromeric region heterochromatin on chromosome 16q by treatment with 2,6-diaminopurine (DAP) is associated with repeated rearrangement of chromosome 16q during outgrowth of DAP-treated clones, thereby establishing a link between the initial site of damage and the occurrence of persistent chromosomal instability. Similarly, karyotypic analysis of gamma ray induced instability demonstrated that chromosomal rearrangements in sub-clones were significantly clustered near the site of previously identified chromosomal rearrangement junctions in unstable parental clones. This study investigates the hypothesis that integration of transfected sequences into host chromosomes could create breakage-prone junction regions and persistent genomic instability without exposure to DNA-damage agents. These junctions may mimic the unstable chromosomal rearrangements induced by DAP or radiation, and thus provide a test of the broader hypothesis that instability can to some extent be attributed to the formation of novel chromosomal breakage hot spots. These experiments were performed using human-hamster hybrid AL cells containing a single human chromosome 11, which was used to monitor instability in a chromosomal painting assay. AL cells were transfected with a 2.5 Kb fragment containing multiple copies of the 180 bp human alpha heterochromatic repeat, which resulted in chromosomal instability in 41% of the transfected clones. Parallel exposure to gamma-radiation resulted in a similar level of chromosomal instability, although control transfections with plasmid alone did not lead to karyotypic instability. Chromosomal instability induced by integration of alpha heterochromatic repeats was also frequently associated with delayed reproductive

Chromosomal Instability and Molecular Defects in Induced Pluripotent Stem Cells from Nijmegen Breakage Syndrome Patients

Directory of Open Access Journals (Sweden)

Tomer Halevy

2016-08-01

Full Text Available Nijmegen breakage syndrome (NBS results from the absence of the NBS1 protein, responsible for detection of DNA double-strand breaks (DSBs. NBS is characterized by microcephaly, growth retardation, immunodeficiency, and cancer predisposition. Here, we show successful reprogramming of NBS fibroblasts into induced pluripotent stem cells (NBS-iPSCs. Our data suggest a strong selection for karyotypically normal fibroblasts to go through the reprogramming process. NBS-iPSCs then acquire numerous chromosomal aberrations and show a delayed response to DSB induction. Furthermore, NBS-iPSCs display slower growth, mitotic inhibition, a reduced apoptotic response to stress, and abnormal cell-cycle-related gene expression. Importantly, NBS neural progenitor cells (NBS-NPCs show downregulation of neural developmental genes, which seems to be mediated by P53. Our results demonstrate the importance of NBS1 in early human development, shed light on the molecular mechanisms underlying this severe syndrome, and further expand our knowledge of the genomic stress cells experience during the reprogramming process.

A novel setup for the determination of absolute cross sections for low-energy electron induced strand breaks in oligonucleotides - The effect of the radiosensitizer 5-fluorouracil

International Nuclear Information System (INIS)

Rackwitz, J.; Rankovic, M.L.; Milosavljevic, A.R.; Bald, I.

2017-01-01

Low-energy electrons (LEEs) play an important role in DNA radiation damage. Here we present a method to quantify LEE induced strand breakage in well-defined oligonucleotide single strands in terms of absolute cross sections. An LEE irradiation setup covering electron energies <500 eV is constructed and optimized to irradiate DNA origami triangles carrying well-defined oligonucleotide target strands. Measurements are presented for 10.0 and 5.5 eV for different oligonucleotide targets. The determination of absolute strand break cross sections is performed by atomic force microscopy analysis. An accurate fluence determination ensures small margins of error of the determined absolute single strand break cross sections σ SSB . In this way, the influence of sequence modification with the radiosensitive 5-Fluorouracil ( 5F U) is studied using an absolute and relative data analysis. We demonstrate an increase in the strand break yields of 5F U containing oligonucleotides by a factor of 1.5 to 1.6 compared with non-modified oligonucleotide sequences when irradiated with 10 eV electrons. (authors)

Environmental and chemotherapeutic agents induce breakage at genes involved in leukemia-causing gene rearrangements in human hematopoietic stem/progenitor cells

Energy Technology Data Exchange (ETDEWEB)

Thys, Ryan G., E-mail: rthys@wakehealth.edu [Department of Cancer Biology, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157-1016 (United States); Lehman, Christine E., E-mail: clehman@wakehealth.edu [Department of Cancer Biology, Wake Forest School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157-1016 (United States); Pierce, Levi C.T., E-mail: Levipierce@gmail.com [Human Longevity, Inc., San Diego, California 92121 (United States); Wang, Yuh-Hwa, E-mail: yw4b@virginia.edu [Department of Biochemistry and Molecular Genetics, University of Virginia, 1340 Jefferson Park Avenue, Charlottesville, VA 22908-0733 (United States)

2015-09-15

Highlights: • Environmental/chemotherapeutic agents cause DNA breakage in MLL and CBFB in HSPCs. • Diethylnitrosamine-induced DNA breakage at MLL and CBFB shown for the first time. • Chemical-induced DNA breakage occurs at topoisomerase II cleavage sites. • Chemical-induced DNA breaks display a pattern similar to those in leukemia patients. • Long-term exposures suggested to generate DNA breakage at leukemia-related genes. - Abstract: Hematopoietic stem and progenitor cells (HSPCs) give rise to all of the cells that make up the hematopoietic system in the human body, making their stability and resilience especially important. Damage to these cells can severely impact cell development and has the potential to cause diseases, such as leukemia. Leukemia-causing chromosomal rearrangements have largely been studied in the context of radiation exposure and are formed by a multi-step process, including an initial DNA breakage and fusion of the free DNA ends. However, the mechanism for DNA breakage in patients without previous radiation exposure is unclear. Here, we investigate the role of non-cytotoxic levels of environmental factors, benzene, and diethylnitrosamine (DEN), and chemotherapeutic agents, etoposide, and doxorubicin, in generating DNA breakage at the patient breakpoint hotspots of the MLL and CBFB genes in human HSPCs. These conditions represent exposure to chemicals encountered daily or residual doses from chemotherapeutic drugs. Exposure of HSPCs to non-cytotoxic levels of environmental chemicals or chemotherapeutic agents causes DNA breakage at preferential sites in the human genome, including the leukemia-related genes MLL and CBFB. Though benzene, etoposide, and doxorubicin have previously been linked to leukemia formation, this is the first study to demonstrate a role for DEN in the generation of DNA breakage at leukemia-specific sites. These chemical-induced DNA breakpoints coincide with sites of predicted topoisomerase II cleavage. The

Environmental and chemotherapeutic agents induce breakage at genes involved in leukemia-causing gene rearrangements in human hematopoietic stem/progenitor cells

International Nuclear Information System (INIS)

Thys, Ryan G.; Lehman, Christine E.; Pierce, Levi C.T.; Wang, Yuh-Hwa

2015-01-01

Highlights: • Environmental/chemotherapeutic agents cause DNA breakage in MLL and CBFB in HSPCs. • Diethylnitrosamine-induced DNA breakage at MLL and CBFB shown for the first time. • Chemical-induced DNA breakage occurs at topoisomerase II cleavage sites. • Chemical-induced DNA breaks display a pattern similar to those in leukemia patients. • Long-term exposures suggested to generate DNA breakage at leukemia-related genes. - Abstract: Hematopoietic stem and progenitor cells (HSPCs) give rise to all of the cells that make up the hematopoietic system in the human body, making their stability and resilience especially important. Damage to these cells can severely impact cell development and has the potential to cause diseases, such as leukemia. Leukemia-causing chromosomal rearrangements have largely been studied in the context of radiation exposure and are formed by a multi-step process, including an initial DNA breakage and fusion of the free DNA ends. However, the mechanism for DNA breakage in patients without previous radiation exposure is unclear. Here, we investigate the role of non-cytotoxic levels of environmental factors, benzene, and diethylnitrosamine (DEN), and chemotherapeutic agents, etoposide, and doxorubicin, in generating DNA breakage at the patient breakpoint hotspots of the MLL and CBFB genes in human HSPCs. These conditions represent exposure to chemicals encountered daily or residual doses from chemotherapeutic drugs. Exposure of HSPCs to non-cytotoxic levels of environmental chemicals or chemotherapeutic agents causes DNA breakage at preferential sites in the human genome, including the leukemia-related genes MLL and CBFB. Though benzene, etoposide, and doxorubicin have previously been linked to leukemia formation, this is the first study to demonstrate a role for DEN in the generation of DNA breakage at leukemia-specific sites. These chemical-induced DNA breakpoints coincide with sites of predicted topoisomerase II cleavage. The

X-ray-induced breakage and rejoining of human interphase chromosomes

International Nuclear Information System (INIS)

Cornforth, M.N.; Bedford, J.S.

1983-01-01

A method was developed for the high-resolution measurement of breaks in prematurely condensed chromosomes at the G 1 phase of the cell cycle. The dose response for fragments (breaks) produced immediately after x-irradiation of confluent cultures of normal human cells was linear down to 10.9 rad (0.109 Gy) and extrapolated to zero effect at zero dose. The curve had a slope of 0.063 breaks per cell per rad, which is at least an order of magnitude greater than that for breaks scored in the same cells after they have progressed to mitosis following subculture. When incubated at 37 0 C half of the breaks disappeared in 2 hours. A slower, perhaps nonrejoining component was apparent at later incubation times. The initial rate of break rejoining was similar to the rate of increase in survival after incubation because of the repair of potentially lethal damage and is also in close agreement with recently reported values for the rejoining of double-strand breakage in DNA

Inhibition of radiation-induced DNA strand breaks by hoechst 33258: OH-radical scavenging and DNA radical quenching

International Nuclear Information System (INIS)

Adhikary, A.; Bothe, E.; Von Sonntag, C.; Adhikary, A.

1997-01-01

The minor-groove-binding dye Hoechst 33258 has been found to protect pBR322 DNA in aqueous solution against radiation-induced single-strand breaks (ssb). This protective effect has been assumed to be largely due to the scavenging of the strand-break-generating OH radicals by Hoechst. From D 37 values for ssb at different Hoechst concentrations the value of the OH radical scavenging constant of DNA-bound Hoechst has been estimated at k Ho/DNA = 2.7 * 10 11 dm 3 mol -1 . This unexpectedly high value has led us to study the reactions of OH radicals with Hoechst in the absence and in the presence of double-stranded calf thymus DNA (ds DNA) by pulse radiolysis, and the formation of radiation-induced ssb by low angle laser light scattering. The D 37 /D 37 0 values at different Hoechst concentrations agree with the values obtained by Martin and al. and demonstrate the protection. However, this protection cannot be explained on the basis of OH radical scavenging alone using the above rate constants. There must, in addition, be some quenching of DNA radicals. Hoechst radicals are formed in the later ms time range, i.e a long time after the disappearance of the OH radicals. This delayed Hoechst radical formation has been assigned to a a reaction of DNA radicals with Hoechst, thereby inhibiting strand breakage. In confirmation, pulse radiolysis of aqueous solution of nucleotides in the presence of Hoechst yields a similar delayed Hoechst radical formation. The data indicate that in DNA the cross-section of this quenching has a diameter of 3 to 4 base pairs per Hoechst molecule. (N.C.)

Iodine-125 induced DNA strand breakage: Contributions of different physical and chemical radiation action mechanisms

International Nuclear Information System (INIS)

Li, W.

2002-01-01

The decay of the radioisotope 125 I into 125 Te is typically followed by the emission of two groups of approximately 10 electrons each. In deoxyribonucleic acid (DNA) with 125 I incorporated, these electrons produce various types of damage to DNA, e.g. single and double strand breaks. They occur through direct actions of physical tracks, or indirect actions of radicals produced in water. Among the direct actions one should consider not only the excitation and ionization of DNA by electrons but also the neutralization of highly charged 125m Te ions with electrons from neighboring molecules. The present work begins with a detailed description of electron tracks with the use of the PARTRAC code, compares results with recent experiments, and concludes with a firm assessment of the contribution to the strand break yields from the neutralization effect. (orig.)

Telomere healing following DNA polymerase arrest-induced breakages is likely the main mechanism generating chromosome 4p terminal deletions.

Science.gov (United States)

Hannes, Femke; Van Houdt, Jeroen; Quarrell, Oliver W; Poot, Martin; Hochstenbach, Ron; Fryns, Jean-Pierre; Vermeesch, Joris R

2010-12-01

Constitutional developmental disorders are frequently caused by terminal chromosomal deletions. The mechanisms and/or architectural features that might underlie those chromosome breakages remain largely unexplored. Because telomeres are the vital DNA protein complexes stabilizing linear chromosomes against chromosome degradation, fusion, and incomplete replication, those terminal-deleted chromosomes acquired new telomeres either by telomere healing or by telomere capture. To unravel the mechanisms leading to chromosomal breakage and healing, we sequenced nine chromosome 4p terminal deletion boundaries. A computational analysis of the breakpoint flanking region, including 12 previously published pure terminal breakage sites, was performed in order to identify architectural features that might be involved in this process. All terminal 4p truncations were likely stabilized by telomerase-mediated telomere healing. In the majority of breakpoints multiple genetic elements have a potential to induce secondary structures and an enrichment in replication stalling site motifs were identified. These findings suggest DNA replication stalling-induced chromosome breakage during early development is the first mechanistic step leading toward terminal deletion syndromes. © 2010 Wiley-Liss, Inc.

Suppressing effect of antimutagenic flavorings on chromosome aberrations induced by UV-light or X-rays in cultured Chinese hamster cells

International Nuclear Information System (INIS)

Sasaki, Yu.F.; Imanishi, Hisako; Watanabe, Mie; Ohta, Toshihiro; Shirasu

1990-01-01

Chromosome aberrations induces by UV-light or X-rays were suppressed by the post-treatment with antimutagenic flavorings, such as anisaldehyde, cinnamaldehyde, coumarin, and vanillin. UV- or X-ray-irradiated surviving cells increased in the presence of each flavouring. X-ray-induced breakage-type and exchange-type chromosome aberrations were suppressed by the vanillin treatment in the G 1 phase of the cell cycle and a greater decrease in the number of X-ray-induced chromosome aberrations during G 1 holding was observed in the presence of vanillin. Furthermore, a greater decrease in the number of X-ray-induced DNA single-strand breaks was observed in the presence of vanillin. Treatment with vanillin in the G 2 phase suppressed UV-and X-ray-induced breakage-type but not exchange-type chromosome aberrations. The suppression of breakage-type aberrations was assumed to be due to a modification of the capability of the post-replicational repair of DNA double-strand breaks. (author). 28 refs.; 5 figs.; 6 tabs

Analytical model of the critical current of a bent Nb3Sn strand

International Nuclear Information System (INIS)

Koizumi, Norikiyo; Murakami, Haruyuki; Hemmi, Tsutomu; Nakajima, Hideo

2011-01-01

The critical current performance of a large Nb 3 Sn cable-in-conduit conductor (CICC) was degraded by periodic bending of strands due to a large transverse electromagnetic force. The degradation of each strand due to this bending should be evaluated in calculations of the critical current of a CICC, but a suitable model has not been developed yet. Therefore, the authors have developed a new analytical model which takes into account plastic deformation of copper and bronze and filament breakage. The calculated results were compared with test results for uniformly bent Nb 3 Sn bronze-route strands. The calculated results assuming a high transverse resistance model (HTRM) show good agreement with the test results, a finding which confirms the validity of the model. Because of a much shorter calculation time than for numerical simulation, the developed model seems much more practical for use in calculating the critical current performance of a Nb 3 Sn CICC. In addition, simulation results show that since the neutral axis of a bent strand shifts to the compressive side due to plastic deformation of the copper and bronze, and/or filament breakage, the strand is elongated by bending. This elongation may enhance the strand's critical current performance. Moreover, the calculated results indicate that the dependence of the critical current on the bending strain is affected by the bending history if the strand is excessively bent, especially when filaments are broken. In a real magnet, since a strand in a CICC is normally subject to the maximum electromagnetic force prior to an evaluation of its performance at a lower electromagnetic force, the effect of over-bending should be taken into account in calculations of its critical current performance, especially when filament breakage occurs.

Radioprotective action of WR-1065 on radiation-induced DNA strand breaks in cultured Chinese hamster ovary cells

International Nuclear Information System (INIS)

Murray, D.; VanAnkeren, S.C.; Milas, L.; Meyn, R.E.

1988-01-01

We have examined the radioprotective effect of WR-1065 on cultured Chinese hamster ovary cells. The effects of the drug on the induction and rejoining of gamma-ray-induced DNA single-strand breaks (SSBs) and double-strand breaks (DSBs) were measured using alkaline (pH 12.1) and neutral (pH 7.0) elution, respectively. Molecular protection factors (PFs) calculated from these data allowed us to determine whether the degree of modification of strand breakage accurately predicted the PFs measured using the biological end point of cell survival. The drug did protect against the induction of both SSBs and DSBs, although to an extent that did not appear to fully account for the degree of radioprotection in terms of cell killing measured under identical conditions. It is therefore unlikely that radioprotection by WR-1065 occurs simply as a consequence of a general lowering of all types of gamma-ray-induced DNA lesions, and it is possible that the drug could differentially protect against the induction of subsets of these DNA lesions. The rate of SSB rejoining was retarded following preirradiation treatment of cells with WR-1065, but there was no effect on DSB rejoining. Postirradiation treatment with WR-1065 also appeared to retard SSB rejoining but without an accompanying effect on either DSB rejoining or cell survival; however, this effect was largely reversed by the addition of catalase and was, therefore, probably a result of H 2 O 2 generated by autoxidation of the drug. Based on these observations, it would appear that the molecular actions of aminothiol radioprotective compounds that lead to reduced cell killing are much more complex than previously thought

Enzymatic induction of DNA double-strand breaks in γ-irradiated Escherichia coli K-12

International Nuclear Information System (INIS)

Bonura, T.; Smith, K.C.; Kaplan, H.S.

1975-01-01

The polA1 mutation increases the sensitivity of E. coli K-12 to killing by γ-irradiation in air by a factor of 2.9 and increases the yield of DNA double-strand breaks by a factor of 2.5. These additional DNA double-strand breaks appear to be due to the action of nucleases in the polA1 strain rather than to the rejoining of radiation-induced double-strand breaks in the pol + strain. This conclusion is based upon the observation that γ-irradiation at 3 0 did not affect the yield of DNA double-strand breaks in the pol + strain, but decreased the yield in the polA1 strain by a factor of 2.2. Irradiation of the polA1 strain at 3 0 followed by incubation at 3 0 for 20 min before plating resulted in approximately a 1.5-fold increase in the D 0 . The yield of DNA double-strand breaks was reduced by a factor of 1.5. The pol + strain, however, did not show the protective effect of the low temperature incubation upon either survival or DNA double-strand breakage. We suggest that the increased yield of DNA double-strand breaks in the polA 1 strain may be the result of the unsuccessful excision repair of ionizing radiation-induced dna base damage

Paravertebral block catheter breakage by electrocautery during thoracic surgery.

Science.gov (United States)

Saeki, Noboru; Sugimoto, Yuki; Mori, Yoko; Kato, Takahiro; Miyoshi, Hirotsugu; Nakamura, Ryuji; Koga, Tomomichi

2017-06-01

Advantages of thoracic paravertebral analgesia (TPA) include placement of the catheter closer to the surgical field; however, the catheter can become damaged during the operation. We experienced a case of intraoperative TPA catheter breakage that prompted us to perform an experiment to investigate possible causes. A 50-year-old male underwent a thoracoscopic lower lobectomy under general anesthesia with TPA via an intercostal approach. Following surgery, it was discovered that the catheter had become occluded, as well as cut and fused, so we reopened the incision and removed the residual catheter. From that experience, we performed an experiment to examine electrocautery-induced damage in normal (Portex™, Smith's Medical), radiopaque (Perifix SoftTip™, BBraun), and reinforced (Perifix FX™, BBraun) epidural catheters (n = 8 each). Chicken meat was penetrated by each catheter and then cut by electrocautery. In the normal group, breakage occurred in 8 and occlusion in 6 of the catheters, and in the radiopaque group breakage occurred in 8 and occlusion in 7. In contrast, breakage occurred in only 3 and occlusion in none in the reinforced group, with the 5 without breakage remaining connected only by the spring coil. Furthermore, in 7 of the reinforced catheters, electric arc-induced thermal damage was observed at the tip of the catheter. A TPA catheter for thoracic surgery should be inserted via the median approach, or it should be inserted after surgery to avoid catheter damage during surgery.

Induction and repair of double- and single-strand DNA breaks in bacteriophage lambda superinfecting Escherichia coli

International Nuclear Information System (INIS)

Boye, E.; Krisch, R.E.

1980-01-01

Induction and repair of double-and single-strand DNA breaks have been measured after decays of 125 I and 3 H incorporated into the DNA and after external irradiation with 4 MeV electrons. For the decay experiments, cells of wild type Escherichia coli K-12 were superinfected with bacteriophage lambda DNA labelled with 5'-( 125 I)iodo-2'-deoxyuridine or with (methyl- 3 H)thymidine and frozen in liquid nitrogen. Aliquots were thawed at intervals and lysed at neutral pH, and the phage DNA was assayed for double- and single-strand breakage by neutral sucrose gradient centrifugation. The gradients used allowed measurements of both kinds of breaks in the same gradient. Decays of 125 I induced 0.39 single-strand breaks per double-strand break. No repair of either break type could be detected. Each 3 H disintegration caused 0.20 single-strand breaks and very few double-strand breaks. The single-strand breaks were rapidly rejoined after the cells were thawed. For irradiation with 4 MeV electrons, cells of wild type E. coli K-12 were superinfected with phage lambda and suspended in growth medium. Irradiation induced 42 single-strand breaks per double-strand break. The rates of break induction were 6.75 x 10 -14 (double-strand breaks) and 2.82 x 10 -12 (single-strand breaks) per rad and per dalton. The single-strand breaks were rapidly repaired upon incubation whereas the double-strand breaks seemed to remain unrepaired. It is concluded that double-strand breaks in superinfecting bacteriophage lambda DNA are repaired to a very small extent, if at all. (Author)

Correlation between γ-ray-induced DNA double-strand breakage and cell killing after biologically relevant doses: analysis by pulsed-field gel electrophoresis

International Nuclear Information System (INIS)

Murray, D.

1994-01-01

We examined the degree of correlation between γ-ray-induced lethality and DNA double-strand breaks (dsbs) after biologically relevant doses of radiation. Radiation lethality was modified by treating 14 C-labelled Chinese hamster ovary cells with either of two aminothiols (WR-1065 or WR-255591) and the associated effect on dsb induction was determined by pulsed-field gel electrophoresis (PFGE). The use of phosphorimaging to analyse the distribution of 14 C-activity in the gel greatly improved the low-dose resolution of the PFGE assay. Both WR-1065 and WR-255591 protected against dsb induction and lethality to a similar extent after low doses of radiation. although this correlation broke down when supralethal doses were used to induce dsbs. Thus, the level of dsbs induced in these cells as measured by PFGE after survival-curve doses of γ-radiation is consistently predictive of the degree of lethality obtained, implying a cause-effect relationship between these two parameters and confirming previous results obtained using the neutral filter elution assay for dsbs. (author)

Mechanical behaviors of multi-filament twist superconducting strand under tensile and cyclic loading

Science.gov (United States)

Wang, Xu; Li, Yingxu; Gao, Yuanwen

2016-01-01

The superconducting strand, serving as the basic unit cell of the cable-in-conduit-conductors (CICCs), is a typical multi-filament twist composite which is always subjected to a cyclic loading under the operating condition. Meanwhile, the superconducting material Nb3Sn in the strand is sensitive to strain frequently relating to the performance degradation of the superconductivity. Therefore, a comprehensive study on the mechanical behavior of the strand helps understanding the superconducting performance of the strained Nb3Sn strands. To address this issue, taking the LMI (internal tin) strand as an example, a three-dimensional structural finite element model, named as the Multi-filament twist model, of the strand with the real configuration of the LMI strand is built to study the influences of the plasticity of the component materials, the twist of the filament bundle, the initial thermal residual stress and the breakage and its evolution of the filaments on the mechanical behaviors of the strand. The effective properties of superconducting filament bundle with random filament breakage and its evolution versus strain are obtained based on the damage theory of fiber-reinforced composite materials proposed by Curtin and Zhou. From the calculation results of this model, we find that the occurrence of the hysteresis loop in the cyclic loading curve is determined by the reverse yielding of the elastic-plastic materials in the strand. Both the initial thermal residual stress in the strand and the pitch length of the filaments have significant impacts on the axial and hysteretic behaviors of the strand. The damage of the filaments also affects the axial mechanical behavior of the strand remarkably at large axial strain. The critical current of the strand is calculated by the scaling law with the results of the Multi-filament twist model. The predicted results of the Multi-filament twist model show an acceptable agreement with the experiment.

Responding to chromosomal breakage during M-phase: insights from a cell-free system

Directory of Open Access Journals (Sweden)

Costanzo Vincenzo

2009-07-01

Full Text Available Abstract DNA double strand breaks (DSBs activate ATM and ATR dependent checkpoints that prevent the onset of mitosis. However, how cells react to DSBs occurring when they are already in mitosis is poorly understood. The Xenopus egg extract has been utilized to study cell cycle progression and DNA damage checkpoints. Recently this system has been successfully used to uncover an ATM and ATR dependent checkpoint affecting centrosome driven spindle assembly. These studies have led to the identification of XCEP63 as major target of this pathway. XCEP63 is a coiled-coil rich protein localized at centrosome essential for proper spindle assembly. ATM and ATR directly phosphorylate XCEP63 on serine 560 inducing its delocalization from centrosome, which in turn delays spindle assembly. This pathway might contribute to regulate DNA repair or mitotic cell survival in the presence of chromosome breakage.

The production and repair of double strand breaks in cells from normal humans and patients with ataxia telangiectasia

International Nuclear Information System (INIS)

Lehman, A.R.; Stevens, S.

1977-01-01

The production and repair of double strand breaks induced by γ-rays in the DNA of human fibroblasts have been measured by sedimentation in sucrose gradients under non-denaturing conditions. Unirradiated DNA formed a rapidly sedimenting gel. Low doses of radiation released freely sedimenting DNA molecules from this gel. Higher doses reduced the rate of sedimentation of the free DNA due to the introduction of double strand breaks. The breakage efficiency was 1 break/1.3x10 10 daltons of DNA/krad. Postirradiation incubation after a high dose of radiation resulted in an increase in molecular weight of the free DNA molecules, and after a low dose the rapidly-sedimenting gel was reformed. These data suggest that double strand breaks are repaired in human fibroblasts. No significant differences were found between fibroblasts from two normal donors and four patients with the radiosensitive disorder, ataxia telangiectasia, in either the production or repair of double strand breaks

Measurement of DNA breakage and breakage repair in mice spleen cells induced by ionizing radiation

International Nuclear Information System (INIS)

Wang Qin; Xue Jingying; Li Jin; Mu Chuanjie; Fan Feiyue

2007-01-01

Objective: To investigate the radioresistance mechanism of IBM-2 mice through measuring DNA single-strand break(SSB) and double-strands break (DSB) as well as their repair. Methods: Pulsed-field gel electrophoresis was used to measure DSB and SSB in IRM-2 mice and their parental mice ICR/JCL and 615 mice after exposure to different doses of γ-ray at different postirradiation time. Results: The initial DNA damages, ie the quantities of DSB and SSB in unirradiation IRM-2 mice were less serious than that of their parental mice ICR/JCL and 615 alice(P<0.01). The percent- age of DSB and SSB in IBM -2 mice was significantly lower than that of ICB/JCL and 615 mice after exposure to various doses of γ-ray(P<0.01 and P<0.05). There were not statistic differences in DSB and SSB repair between IRM-2 mice and their parental mice after exposure to 2Gy radiation. The DNA damage repair rate induced by 4Gy and 8Gy radiation in IRM - 2 mice was rapid, ie the repair rate of SSB and DSB after 0.5h and 1h postirradiation in IRM-2 mice was higher than that of their' parental mice (P<0.01 and P<0.05). And remaining damages after repair in IRM-2 mice were lower than that of ICR/JCL and 615 mice. Conclusion: The DNA damages in IBM-2 mice were lower than that of their parental mice after exposure to ionizing radiation. Moreover, the repair rate of SSB and DSB was higher than that of their parental mice, which perhaps were the radioresistance causes of IBM-2 mice. Therefore IRM-2 mice are naturally resistant to DNA damages induced by ionizing radiation. (authors)

A single-strand specific lesion drives MMS-induced hyper-mutability at a double-strand break in yeast.

Science.gov (United States)

Yang, Yong; Gordenin, Dmitry A; Resnick, Michael A

2010-08-05

Localized hyper-mutability (LHM) can be important in evolution, immunity, and genetic diseases. We previously reported that single-strand DNA (ssDNA) can be an important source of damage-induced LHM in yeast. Here, we establish that the generation of LHM by methyl methanesulfonate (MMS) during repair of a chromosomal double-strand break (DSB) can result in over 0.2 mutations/kb, which is approximately 20,000-fold higher than the MMS-induced mutation density without a DSB. The MMS-induced mutations associated with DSB repair were primarily due to substitutions via translesion DNA synthesis at damaged cytosines, even though there are nearly 10 times more MMS-induced lesions at other bases. Based on this mutation bias, the promutagenic lesion dominating LHM is likely 3-methylcytosine, which is single-strand specific. Thus, the dramatic increase in mutagenesis at a DSB is concluded to result primarily from the generation of non-repairable lesions in ssDNA associated with DSB repair along with efficient induction of highly mutagenic ssDNA-specific lesions. These findings with MMS-induced LHM have broad biological implications for unrepaired damage generated in ssDNA and possibly ssRNA. Published by Elsevier B.V.

Chirality induction and protonation-induced molecular motions in helical molecular strands.

Science.gov (United States)

Kolomiets, Elena; Berl, Volker; Lehn, Jean-Marie

2007-01-01

The long oligopyridinedicarboxamide strand 9, containing 15 heterocyclic rings has been synthesized and its helical structure determined by X-ray crystallography. It was shown that the shorter analogue 6 displays induced circular dichroism and amplification of induced chirality upon dissolution in an optically active solvent, diethyl-L-tartrate. A novel class of helical foldamers was prepared, strands 14-16, based on two oligopyridine carboxamide segments linked through a L-tartaric acid derived spacer. These tartro strands display internal chirality induction as well as chirality amplification. NMR spectroscopy (on 8 and 9) and circular dichroism (on 16) studies show that the oligopyridine carboxamide strands undergo reversible unfolding/folding upon protonation. The protonation-induced unfolding has been confirmed by X-ray crystallographic determination of the molecular structure of the extended protonated heptameric form 8(+). The molecular-scale mechano-chemical motions of the protonation-induced structural switching consist of a change of the length of the molecule, from 6 angstroms (6, coiled form) to 29 angstroms (8(+), uncoiled form) for the heptamer and from 12.5 angstroms (9, coiled form, X-ray structure) to 57 angstroms (9(+), uncoiled form, from modeling) for the pentadecamer. Similar unfolding/folding motional processes take place in the L-tartro strands 15 and 16 upon protonation/deprotonation, with loss of helicity-induced circular dichroism on unfolding as shown for the protonated form 16(+).

[Correlation of single-cell gel electrophoresis and mitomycin C-induced chromosomal breakage for chromosomal instabiligy in children with Fanconi anemia].

Science.gov (United States)

Zhang, Li; Liu, Qiang; Zou, Yao; Liu, Xiao-ming; Zhang, Jia-yuan; Wang, Shu-chun; Chen, Xiao-juan; Guo, Ye; Yang, Wen-yu; Ruan, Min; Liu, Tian-feng; Liu, Fang; Cai, Xiao-jin; Chen, Yu-mei; Zhu, Xiao-fan

2013-02-01

Fanconi anemia (FA) is characterized by bone marrow failure, congenital abnormalities and predisposition to neoplasia. Hypersensitivity of FA cells to the clastogenic effect of mitomycin C (MMC) provides a unique marker for the diagnosis before the beginning of hematological manifestations. The aim of this study was to evaluate the relationship between Single-Cell Gel Electrophoresis (SCGE) and mitomycin C-induced chromosomal breakage in children with FA. Between January 2007 and June 2011, 248 children (results of the two methods and compared with each other. The receiver operating characteristic (ROC) curve was used to evaluate the parameters in SCGE. Seventeen patients were diagnosed as FA and 231 as non-FA. Chromosomal breakage was found to be significantly higher in FA patients [(32.2 ± 4.8)%] than non-FA [(19.9 ± 3.0)%] and controls[(21.6 ± 4.8)%] when induced by MMC 80 ng/ml. The parameters of SCGE were significantly different between FA patients and non-FA or controls. All the parameters were rectilinearly correlated with MMC (P = 0.000). The most closely correlated parameter was the rate of comet cell (r = 0.848, P = 0.000). The results of ROC curves suggested the comet cell rate (0.999) was more important. SCGE might be used to discriminate between FA and non-FA individuals. The relationship between SCGE and MMC-induced chromosomal breakage was significant. The rate of comet cell was the important parameter.

The effect of mitotic inhibitors on DNA strand size and radiation-associated break repair in Down syndrome fibroblasts

International Nuclear Information System (INIS)

Woods, W.G.; Steiner, M.E.; Kalvonjian, S.L.

1985-01-01

The effect of mitotic inhibitors on formation and repair of DNA breaks was studied in cultured fibroblasts from patients with Down syndrome in order to investigate the hypothesis that the karyotyping procedure itself may play a role in the increased chromosome breakage seen in these cells after gamma radiation exposure. Using the nondenaturing elution and alkaline elution techniques to examine fibroblasts from Down syndrome patients and from controls, no specific abnormalities in Down syndrome cells could be detected after exposure to mitotic inhibitors, including rate and extent of elution of DNA from filters as well as repair of radiation-induced DNA breaks. In both normal and Down syndrome cell strains, however, exposure to mitotic inhibitors was associated with a decrease in cellular DNA strand size, suggesting the presence of drug-induced DNA strand breaks. The mechanism of increased chromosome sensitivity of Down syndrome cells to gamma radiation remains unknown. (orig.)

Flavonoids with DNA strand-scission activity from Rhus javanica var. roxburghiana.

Science.gov (United States)

Lin, Chun-Nan; Chen, Hui-Ling; Yen, Ming-Hong

2008-01-01

The flavonoids isolated from the stems of Rhus javanica var. roxburghiana, taxifolin (1), fisetin (2), fustin (3), 3,7,4'-trihydroxyflavanone (4) and 3,7,4'-trihydroxyflavone (5) caused breakage of supercoiled plasmid pBR322 DNA in the presence of Cu(II). Cu(I) was shown to be an essential intermediate by using the Cu(I)-specific sequestering reagent neocuproine. The Cu(II)-mediated DNA scissions induced by 1, 2, 3 and 5 were inhibited by the addition of catalase and exhibited DNA strand break by the addition of KI and superoxide dimutase (SOD), while in the Cu(II)-mediated DNA scissions induced by 4 was inhibited by the addition of KI, SOD, and catalase. It is concluded that 1, 2, 3, and 5 can induce H2O2 and superoxide anion, while 4 can induce OH* and H2O2 and subsequent oxidative damage of DNA in the presence of Cu(II).

Protection of free-radical induced DNA strand breaks in vitro by flavonoids

International Nuclear Information System (INIS)

Fisher, L.; Anderson, R.F.

1998-01-01

Full text: We have used both plasmid and cosmid test systems to assay the effect of antioxidant flavonoids (AO) on DNA strand breakage in supercoiled closed circular DNA (DNA SC ) following the formation oxidative radical damage on DNA (DNA OXID + . ) in aqueous solution. Single strand breaks in DNA SC result in the formation of the relaxed circular form (DNA RC ) and double strand breaks give linear DNA (DNA L ). Dose response curves were constructed for the log of the loss of [DNA S C] against dose (0-600 Gy). The D 37 (dose for 37% unchanged DNA SC ) values determined in the presence of increasing amounts of flavonoids were compared as ratios to the D 37 control value to give dose modification factor (DMF). Irradiations were carried out under 'constant scavenging' conditions to separate out the effect of direct radical scavenging from the possible electron transfer reaction. Control irradiation experiments, were performed in aerated TRIS buffer, concentration 10 mM, which has a scavenging capacity, k s (defined as the summation of the rate constants for the reaction of OH radicals with all species in solution, multiplied by their concentrations) of 1.5 x 10 7 s -1 . The concentration of TRIS was reduced upon addition of AO to maintain k s at this level. Data will be presented for examples from all four major types of flavonoids (flavonols, isoflavones, flavones and flavon-3-ols) showing DMF values plateau at near 2.0 even at low concentrations (ca. 20 μM) of the flavonoids. Increased DNA strand breaks following post irradiation incubation with endo III protein was unaffected by having the flavonoids present at the time of irradiation. This result suggests that the protection afforded by the flavonoids is unlikely to be in repairing radical damage on pyrimidine bases that are precursors of DNA strand breaks. Overall these studies provide evidence for an additional mechanism of antioxidant activity

DNA and chromosome breaks induced by 123I-estrogen in CHO cells

International Nuclear Information System (INIS)

Schwartz, J.L.

1997-01-01

The effects of the Auger electron-emitting isotope I-123, covalently bound to estrogen, on DNA single- and double-strand breakage and on chromosome breakage was determined in estrogen positive Chinese hamster ovary (CHO-ER) cells. Exposure to the 123 I-estrogen induced both single- and double-strand breaks with a ratio of single- to double-strand breaks of 2.2. The corresponding ratio with 60 Co gamma rays was 15.6. The dose-response was biphasic suggesting that either receptor sites are saturated at high does, or that there is a nonrandom distribution of breaks induced by the 123 I-estrogen. The 123 I-estrogen treatment induced chromosome aberrations with an efficiency of about 1 aberration for each 1,000 disintegrations per cell. This corresponds to the mean lethal dose of 123 I-estrogen for these cells suggesting that the lethal event induced by the Auger electron emitter bound to estrogen is a chromosome aberration. Most of the chromosome-type aberrations were dicentrics and rings, suggesting that 123 I-estrogen-induced chromosome breaks are rejoined. The F-ratio, the ratio of dicentrics to centric rings, was 5.8 ± 1.7, which is similar to that seen with high LET radiations. Their results suggest that I-123 bound to estrogen is an efficient clastogenic agent, that the cytotoxic damage produced by I-123 bound to estrogen is very like high LET-induced damage, and the I-123 in the estrogen-receptor-DNA complex is probably in close proximity to the sugar-phosphate backbone of the DNA

Turbulent breakage of ductile aggregates.

Science.gov (United States)

Marchioli, Cristian; Soldati, Alfredo

2015-05-01

In this paper we study breakage rate statistics of small colloidal aggregates in nonhomogeneous anisotropic turbulence. We use pseudospectral direct numerical simulation of turbulent channel flow and Lagrangian tracking to follow the motion of the aggregates, modeled as sub-Kolmogorov massless particles. We focus specifically on the effects produced by ductile rupture: This rupture is initially activated when fluctuating hydrodynamic stresses exceed a critical value, σ>σ(cr), and is brought to completion when the energy absorbed by the aggregate meets the critical breakage value. We show that ductile rupture breakage rates are significantly reduced with respect to the case of instantaneous brittle rupture (i.e., breakage occurs as soon as σ>σ(cr)). These discrepancies are due to the different energy values at play as well as to the statistical features of energy distribution in the anisotropic turbulence case examined.

Diagnosis of Fanconi Anemia: Chromosomal Breakage Analysis

Science.gov (United States)

Oostra, Anneke B.; Nieuwint, Aggie W. M.; Joenje, Hans; de Winter, Johan P.

2012-01-01

Fanconi anemia (FA) is a rare inherited syndrome with diverse clinical symptoms including developmental defects, short stature, bone marrow failure, and a high risk of malignancies. Fifteen genetic subtypes have been distinguished so far. The mode of inheritance for all subtypes is autosomal recessive, except for FA-B, which is X-linked. Cells derived from FA patients are—by definition—hypersensitive to DNA cross-linking agents, such as mitomycin C, diepoxybutane, or cisplatinum, which becomes manifest as excessive growth inhibition, cell cycle arrest, and chromosomal breakage upon cellular exposure to these drugs. Here we provide a detailed laboratory protocol for the accurate assessment of the FA diagnosis as based on mitomycin C-induced chromosomal breakage analysis in whole-blood cultures. The method also enables a quantitative estimate of the degree of mosaicism in the lymphocyte compartment of the patient. PMID:22693659

Effects of hyperthermia on radiation-induced chromosome breakage and loss in excision repair deficient Drosophila melanogaster

International Nuclear Information System (INIS)

Mittler, S.

1986-01-01

Hyperthermia increased radiosensitivity with respect to γ-ray induced chromosome loss and breakage in all stages of spermatogenesis in the wild type Oregon R strain of Drosophila melanogaster, whereas hyperthermia increased radiosensitivity to a lesser extent in cn mus(2) 201sup(D1), an excision repair mutant with 0 per cent excision capacity and in mus(3) 308sup(D1), a strain with 24 per cent excision capacity. The differences in hyperthermia-induced radiation sensitivity between the excision repair mutants and the wild strain may be due to the hyperthermia affecting the excision repair mechanism, suggesting that one of the possible mechanisms involved in hyperthermia-increased radiosensitivity is an effect on excision repair. (author)

Enzymatic quantification of strand breaks of DNA induced by vacuum-UV radiation

International Nuclear Information System (INIS)

Ito, Takashi

1986-01-01

Hind3 digested plasmid DNA dried on an aluminum plate was irradiated by vacuum-UV at 160 and 195 nm using a synchrotron irradiation system. A change induced in the DNA, presumably a single strand break, was quantified by the aid of the strand break-derived stimulation of poly(ADP-ribose) synthetase activity. The end group of strand breaks so induced was recognized by the enzyme as effectively as that by DNase 1 treatment, suggesting a nicking as the major lesion inflicted on the DNA. The fluence (UV) dependent stimulation of poly(ADP-ribose) synthetase activity was much higher upon 160 nm irradiation than upon 195 nm irradiation. (Auth.)

Photolysis of phosphodiester bonds in plasmid DNA by high intensity UV laser irradiation

International Nuclear Information System (INIS)

Croke, D.T.; Blau, Werner; OhUigin, Colm; Kelly, J.M.; McConnell, D.J.

1988-01-01

The cleavage of phosphodiester bonds in DNA exposed to high intensity UV laser pulses in aerated aqueous solution has been investigated using a krypton fluoride excimer laser (248 nm) and bacterial plasmid DNA. The dependence of strand breakage on fluence and intensity has been studied in detail and shows that the process is non-linear with respect to intensity. The relationship between the quantum yield for strand breakage and intensity shows that the strand breakage reaction involves two-photon excitation of DNA bases. The quantum yield rises with intensity from a lower value of 7 x 10 -5 until a maximum value of 4.5 x 10 -4 is attained at intensities of 10 11 W m -2 and above. This value is approximately fifty-fold higher than the quantum yield for strand breakage induced by exposure to low density UV irradiation (254 nm, 12 W m -2 ). DNA sequencing experiments have shown that strand breakage occurs by the specific cleavage of the phosphodiester bond which lies immediately 3' to guanine residues in the DNA, leaving some alkali-labile remnant attached to the terminal phosphate. A mechanism for DNA strand breakage which involves the generation of guanine radical cations is proposed. (author)

Increased chromosomal breakage in Tourette syndrome predicts the possibility of variable multiple gene involvement in spectrum phenotypes: Preliminary findings and hypothesis

Energy Technology Data Exchange (ETDEWEB)

Gericke, G.S.; Simonic, I.; Cloete, E.; Buckle, C. [Univ. of Pretoria (South Africa)] [and others

1995-10-09

Increased chromosomal breakage was found in 12 patients with DSM-IV Tourette syndrome (TS) as compared with 10 non-TS control individuals with respect to untreated, modified RPM1-, and BrdU treated lymphocyte cultures (P < 0.001 in each category). A hypothesis is proposed that a major TS gene is probably connected to genetic instability, and associated chromosomal marker sites may be indicative of the localization of secondary genes whose altered expression could be responsible for associated comorbid conditions. This concept implies that genes influencing higher brain functions may be situated at or near highly recombigenic areas allowing enhanced amplification, duplication and recombination following chromosomal strand breakage. Further studies on a larger sample size are required to confirm the findings relating to chromosomal breakage and to analyze the possible implications for a paradigmatic shift in linkage strategy for complex disorders by focusing on areas at or near unstable chromosomal marker sites. 32 refs., 1 tab.

Effect of radiomodifying agents on the ratios of X-ray-induced lesions in cellular DNA: use in lethal lesion determination

International Nuclear Information System (INIS)

Radford, I.R.

1986-01-01

The effect of three radiomodifying agents, cysteamine, hyperthermia, and hypoxia, on the induction of the major classes of X-ray-induced DNA lesions, was studied using mouse L cells and Chinese hamster V79 cells. The use of filter elution techniques allowed most of these studies to be conducted at X-ray doses within the survival-curve range. Cysteamine was found to protect against DNA single-strand breakage (ssb), DNA base damage, and DNA-protein crosslinkage. Hyperthermia had no effect on the level of DNA ssb or DNA base damage, but in L cells (but not in V79 cells) it increased the level of DNA-protein crosslinkage relative to DNA ssb. Hypoxia protected against DNA ssb, had no significant effect on the level of DNA base damage, and enhanced the level of DNA-protein crosslinkage relative to DNA ssb. These results support the previous suggestion that the X-ray-induced lethal lesion is DNA double-strand breakage. Implications of these findings for the mechanisms of formation of X-ray-induced DNA lesions are also discussed. (author)

Monitoring of DNA breakage in embryonic stages of the African catfish Clarias gariepinus (Burchell, 1822) after exposure to lead nitrate using alkaline comet assay.

Science.gov (United States)

Osman, Alaa G M; Mekkawy, Imam A; Verreth, Johan; Wuertz, Sven; Kloas, Werner; Kirschbaum, Frank

2008-12-01

Increasing lead contamination in Egyptian ecosystems and high lead concentrations in food items have raised concern for human health and stimulated studies on monitoring ecotoxicological impact of lead-caused genotoxicity. In this work, the alkaline comet assay was modified for monitoring DNA strand breakage in sensitive early life stages of the African catfish Clarias gariepinus. Following exposure to 100, 300, and 500 microg/L lead nitrate, DNA strand breakage was quantified in embryos at 30, 48, 96, 144, and 168 h post-fertilization (PFS). For quantitative analysis, four commonly used parameters (tail % DNA, %TDNA; head % DNA, %HDNA; tail length, TL; tail moment, TM) were analyzed in 96 nuclei (in triplicates) at each sampling point. The parameter %TDNA revealed highest resolution and lowest variation. A strong correlation between lead concentration, time of exposure, and DNA strand breakage was observed. Here, genotoxicity detected by comet assay preceded the manifested malformations assessed with conventional histology. Qualitative evaluation was carried out using five categories are as follows: undamaged (%TDNA 75%) nuclei confirming a dose and time-dependent shift towards increased frequencies of highly and extremely damaged nuclei. A protective capacity provided by a hardened chorion is a an interesting finding in this study as DNA damage in the prehatching stages 30 h-PFS and 48 h-PFS was low in all treatments (qualitative and quantitative analyses). These results clearly show that the comet assay is a sensitive tool for the detection of genotoxicity in vulnerable early life stages of the African catfish and is a method more sensitive than histological parameters for monitoring genotoxic effects. 2008 Wiley Periodicals, Inc.

Electron attachment to DNA single strands: gas phase and aqueous solution.

Science.gov (United States)

Gu, Jiande; Xie, Yaoming; Schaefer, Henry F

2007-01-01

such as glycosidic bond breakage. However, the radical anions of the pyrimidine nucleotides with high VDE are expected to be electronically stable. Thus the base-centered radical anions of the pyrimidine nucleotides might be the possible intermediates for DNA single-strand breakage.

Summary of the performance of strand produced for the 1990 SSC dipole program

International Nuclear Information System (INIS)

Christopherson, D.; Capone, D. II; Seuntjens, J.; Pollock, D.; Hannaford, C.R.

1991-01-01

In 1990 and at the beginning of 1991, more than 4 million feet of wire was delivered to support the SSC Dipole Program. This wire was fabricated to meet specification SSC-MAG-M-4141, and test results and various statistics are compiled here. Certain strengths and weaknesses in the performance of the delivered strand are discussed, including analysis of strand breakage in certain billets. Test results of cable manufactured for 40 mm dipole magnets and 50 mm dipole magnets are reported, and a brief overview of the 1991-1992 Conductor Program is included

Coke breakage behaviour in relation to its structure

Energy Technology Data Exchange (ETDEWEB)

Peirce, T J; Horton, A E; Tucker, J

1980-06-14

The relationship between coke macrostructure and volumetric (coarse) breakage behaviour, and the manner in which both features are affected by the coal charge composition are discussed. Coke pieces which are extensively fissured fail spontaneously after only a few revolutions in the drum; coke pieces which are less fissured require an extended treatment in the drum and finally fail in a manner consistent with that of fatigue crack growth induced by repeated impact. This heterogeneity in breakage behaviour was demonstrated in terms of an abrupt change in gradient of the Weibull distribution function for the coke. Heterogeneity may be removed by fully stabilising the coke or by the use of a more energetic test method. The role of fatigue crack growth in coke breakage was studied by cutting fissures of defined depth in full stabilised coke and assessing the rate of crack growth as a function of fissure depth and fracture toughness. The form of the results was consistent with that given by the Paris-Erdogan law and the rate of crack growth (for a given stress intensity level) was shown to increase with reducing coke toughness. Explanations are suggested to account for the role of blend additives in modifying the fissured properties and fracture toughness of coke made from high-volatile coal.

Investigations of radiation-induced strand breaks of poly(U) in aqueous solutions

International Nuclear Information System (INIS)

Lemaire, D.G.E.

1984-01-01

DNA strand breaks induced by γ irradiation were studied in polyuridylic acid (Poly(U)), a single-strand model substance with a single base. Poly(U) in diluted, aqueous solution was irradiated in a Co-γ source, and the 100 eV yields of strand breaks (Cr values) were determined on the basis of the loss of molecular weight. The molecular weight was determined by small-angle laser light scattering. (orig./PW) [de

TALEN-Induced Double-Strand Break Repair of CTG Trinucleotide Repeats

Directory of Open Access Journals (Sweden)

Valentine Mosbach

2018-02-01

Full Text Available Trinucleotide repeat expansions involving CTG/CAG triplets are responsible for several neurodegenerative disorders, including myotonic dystrophy and Huntington’s disease. Because expansions trigger the disease, contracting repeat length could be a possible approach to gene therapy for these disorders. Here, we show that a TALEN-induced double-strand break was very efficient at contracting expanded CTG repeats in yeast. We show that RAD51, POL32, and DNL4 are dispensable for double-strand break repair within CTG repeats, the only required genes being RAD50, SAE2, and RAD52. Resection was totally abolished in the absence of RAD50 on both sides of the break, whereas it was reduced in a sae2Δ mutant on the side of the break containing the longest repeat tract, suggesting that secondary structures at double-strand break ends must be removed by the Mre11-Rad50 complex and Sae2. Following the TALEN double-strand break, single-strand annealing occurred between both sides of the repeat tract, leading to repeat contraction.

Repair of endogenous and ionizing radiation-induced DNA damages: mechanisms and biological functions

International Nuclear Information System (INIS)

Boiteux, S.

2002-01-01

The cellular DNA is continuously exposed to endogenous and exogenous stress. Oxidative stress due to cellular metabolism is the major cause of endogenous DNA damage. On the other hand, ionizing radiation (IR) is an important exogenous stress. Both induce similar DNA damages: damaged bases, abasic sites and strand breakage. Most of these lesions are lethal and/or mutagenic. The survival of the cell is managed by efficient and accurate DNA repair mechanisms that remove lesions before their replication or transcription. DNA repair pathways involved in the removal of IR-induced lesions are briefly described. Base excision repair (BER) is mostly involved in the removal of base damage, abasic sites and single strand breaks. In contrast, DNA double strand breaks are mostly repaired by non-homologous end joining (NHEJ) or homologous recombination (HR). How DNA repair pathways prevent cancer process is also discussed. (author)

Genetics Home Reference: Warsaw breakage syndrome

Science.gov (United States)

... Oostra AB, Stumm M, Zdzienicka MZ, Joenje H, de Winter JP. Warsaw breakage syndrome, a cohesinopathy associated with mutations in the XPD ... Lelij P, Oostra AB, Rooimans MA, Joenje H, de Winter JP. Diagnostic Overlap ... Roberts Syndrome and Warsaw Breakage Syndrome. Anemia. 2010;2010:565268. ...

Repair of X-ray-induced single-strand breaks by a cell-free system

International Nuclear Information System (INIS)

Seki, Shuji; Ikeda, Shogo; Tsutui, Ken; Teraoka, Hirobumi

1990-01-01

Repair of X-ray-induced single-strand breaks of DNA was studied in vitro using an exonuclease purified from mouse ascites sarcoma (SR-C3H/He) cells. X-ray-dose-dependent unscheduled DNA synthesis was primed by the exonuclease. Repair of X-ray-induced single-strand breaks in pUC19 plasmid DNA was demonstrated by agarose gel electrophoresis after incubating the damaged DNA with the exonuclease, DNA polymerase (Klenow fragment of DNA polymerase I or DNA polymerase β purified from SR-C3H/He cells), four deoxynucleoside triphosphates, ATP and DNA ligase (T4 DNA ligase or DNA ligase I purified from calf thymus). The present results suggested that the exonuclease is involved in the initiation of repair of X-ray-induced single-strand breaks in removing 3' ends of X-ray-damaged DNA. (author)

Investigation of factors influencing the attrition breakage of coal

Energy Technology Data Exchange (ETDEWEB)

Oberholzer, V.; van der Walt, J. [North West University, Mmabatho (South Africa). School of Chemical & Minerals Engineering

2009-04-15

The presence of fines (particles smaller than 6 mm in diameter) causes hydrodynamic problems in gasifiers and therefore it is of great importance to minimize the amount of fine coal in the feed. This serves as motivation for understanding coal's breakage mechanisms, which could lead to the ability to predict the generation of fines. The aim of this project was to simulate the pulsating effect of the conveyor belt in order to investigate factors influencing the breakage. Results indicated that an increased initial particle size had an increasing effect in the amount of fines generated. Weathering had an insignificant effect on the breakage of coal. To conclude, a breakage model was developed to describe the rate of breakage out of the top size when a combination of two breakage modes is present.

Summary of the performance of strand produced for the 1990 SSC Dipole Program

International Nuclear Information System (INIS)

Christopherson, D.; Capone, D.; Seuntjens, J.; Pollock, D.; Hannaford, C.R.

1991-03-01

In 1990 and at the beginning of 1991, more than 4 million feet of wire was delivered to support the SSC Dipole Program. This wire was fabricated to meet specification SSC-MAG-M-4141, and test results and various statistics are compiled here. Certain strengths and weaknesses in the performance of the delivered strand are discussed, including analysis of strand breakage in certain billets. Test results of cable manufactured for 40 mm dipole magnets and 50 mm dipole magnets are reported, and a brief overview of the 1991--1992 Conductor Program is included. 4 refs., 5 figs., 4 tabs

Quantitation of ultraviolet-induced single-strand breaks using oligonucleotide chip

International Nuclear Information System (INIS)

Pal, Sukdeb; Kim, Min Jung; Choo, Jaebum; Kang, Seong Ho; Lee, Kyeong-Hee; Song, Joon Myong

2008-01-01

A simple, accurate and robust methodology was established for the direct quantification of ultraviolet (UV)-induced single-strand break (SSB) using oligonucleotide chip. Oligonucleotide chips were fabricated by covalently anchoring the fluorescent-labeled ssDNAs onto silicon dioxide chip surfaces. Assuming that the possibility of more than one UV-induced SSB to be generated in a small oligonucleotide is extremely low, SSB formation was investigated quantifying the endpoint probe density by fluorescence measurement upon UV irradiation. The SSB yields obtained based on the highly sensitive laser-induced fluorometric determination of fluorophore-labeled oligonucleotides were found to coincide well with that predicted from a theoretical extrapolation of the results obtained for plasmid DNAs using conventional agarose gel electrophoresis. The developed method has the potential to serve as a high throughput, sample-thrifty, and time saving tool to realize more realistic, and direct quantification of radiation and chemical-induced strand breaks. It will be especially useful for determining the frequency of SSBs or lesions convertible to SSBs by specific cleaving reagents or enzymes

Stem breakage of salt marsh vegetation under wave forcing: A field and model study

Science.gov (United States)

Vuik, Vincent; Suh Heo, Hannah Y.; Zhu, Zhenchang; Borsje, Bas W.; Jonkman, Sebastiaan N.

2018-01-01

One of the services provided by coastal ecosystems is wave attenuation by vegetation, and subsequent reduction of wave loads on flood defense structures. Therefore, stability of vegetation under wave forcing is an important factor to consider. This paper presents a model which determines the wave load that plant stems can withstand before they break or fold. This occurs when wave-induced bending stresses exceed the flexural strength of stems. Flexural strength was determined by means of three-point-bending tests, which were carried out for two common salt marsh species: Spartina anglica (common cord-grass) and Scirpus maritimus (sea club-rush), at different stages in the seasonal cycle. Plant stability is expressed in terms of a critical orbital velocity, which combines factors that contribute to stability: high flexural strength, large stem diameter, low vegetation height, high flexibility and a low drag coefficient. In order to include stem breakage in the computation of wave attenuation by vegetation, the stem breakage model was implemented in a wave energy balance. A model parameter was calibrated so that the predicted stem breakage corresponded with the wave-induced loss of biomass that occurred in the field. The stability of Spartina is significantly higher than that of Scirpus, because of its higher strength, shorter stems, and greater flexibility. The model is validated by applying wave flume tests of Elymus athericus (sea couch), which produced reasonable results with regards to the threshold of folding and overall stem breakage percentage, despite the high flexibility of this species. Application of the stem breakage model will lead to a more realistic assessment of the role of vegetation for coastal protection.

Effects of hyperthermia on repair of radiation-induced DNA strand breaks

International Nuclear Information System (INIS)

Mills, M.D.; Meyn, R.E.

1981-01-01

Previous reports have suggested a relationship between the heat-induced changes in nucleoprotein and the hyperthermic enhancement of radiation sensitivity. In an effort to further understand these relationships, we measured the level of initial DNA strand break damage and the DNA strand break rejoining kinetics in Chinese hamster ovary cells following combined hyperthermia and ionizing radiation treatments. The amount of protein associated with DNA measured as the ratio of [ 3 H)leucine to [ 14 C]thymidine was also compared in chromatin isolated from both heated and unheated cells. The results of these experiments show that the initial level of radiation-induced DNA strand breaks is significantly enhanced by a prior hyperthermia treatment of 43 0 C for 30 min. Treatments at higher temperatures and longer treatments at the same temperature magnified this effect. Hyperthermia was also shown to cause a substantial inhibition of the DNA strand break rejoining after irradiation. Both the initial level of DNA damage and the rejoining kinetics recovered to normal levels with incubation at 37 0 C between the hyperthermia and radiation treatments. Recovery of these parameters coincided with the return of the amount of protein associated with DNA to normal values, further suggesting a relationship between the changes in nucleoprotein and the hyperthermic enhancement of radiation sensivivity

Induction of DNA strand breaks by RSU-1069, a nitroimidazole-aziridine radiosensitizer

International Nuclear Information System (INIS)

Silver, A.R.J.; O'Neill, P.; Jenkins, T.C.

1985-01-01

[2- 14 C]-RSU-1069 [1-(2-nitro-1-imidazolyl)-3-(1-aziridino)-2-propanol], either as a parent or following radiation reduction, binds to calf thymus DNA in vitro. Radiation-reduced RSU-1069 binds to a greater extent and more rapidly than the parent compound. RSU-1137, a non-aziridino analogue of RSU-1069, binds following radiation reduction. Radiation-reduced misonidazole exhibits binding ratios a thousand-fold less than those of reduced RSU-1069. Both parent and reduced RSU-1069 cause single strand breaks (ssbs) in pSV2 gpt plasmid DNA with the reduced compound causing a greater number of breaks. Parent and reduced RSU-1137 and misonidazole do not cause ssbs. It is inferred that the aziridine moiety present in both parent and reduced RSU-1069 is required for ssb production. RSU-1069 reacts with inorganic phosphate probably via nucleophilic ring-opening of the aziridine fragment. Incubation of plasmid DNA with reduced RSU-1069 in the presence of either phosphate or deoxyribose-5-phosphate at concentrations greater than 0.35 mol dm -3 prevents strand breakage, whereas 1.2 mol dm -3 deoxyribose does not protect against strand breakage formation. It is proposed that the observed binding to DNA occurs via the aziridine and the reduced nitro group of RSU-1069 and that these two have different target sites. Binding to DNA via the reduced nitro group may serve to increase aziridine attack due to localization at or near its target. (author)

Repair and gamma radiation-induced single- and double-strand breaks in DNA of Escherichia coli

International Nuclear Information System (INIS)

Petrov, S.I.

1981-01-01

Studies in the kinetics of repair of γ-radiation-induced single- and double-strand breaks in DNA of E. coli cells showed that double-strand DNA breaks are rejoined by the following two ways. The first way is conditioned by repair of single-strand breaks and represents the repair of ''oblique'' double-strand breaks in DNA, whereas the second way is conditioned by functioning of the recombination mechanisms and, to all appearance, represents the repair of ''direct'' double-strand breaks in DNA

Pebble breakage in gravel

International Nuclear Information System (INIS)

Tuitz, C.

2012-01-01

The spatial clustering of broken pebbles in gravel layers of a Miocene sedimentary succession was investigated. Field observations suggested that the occurrence of broken pebbles could be related with gravel hosted shear deformation bands, which were the result of extensional regional deformation. Several different methods were used in this work to elucidate these observations. These methods include basic field work, measurements of physical pebble and gravel properties and, the application of different numerical modelling schemes. In particular, the finite element method in 2D and the discrete element method in 2D and 3D were used in order to quantify mechanisms of pebble deformation. The main objective of this work was to identify potential mechanisms that control particle breakage in fluvial gravel, which could explain the clustered spatial distribution of broken pebbles. The results of 2D finite element stress analysis indicated that the breakage load of differently located and oriented diametrical loading axes on a pebble varies and, that the weakest loading configuration coincides with the smallest principal axis of the pebble. The 3D discrete element method was applied to study the contact load distribution on pebbles in gravel deposits and the influence of different degrees of particle imbrication and orientation. The results showed that an increase of the number of imbricated particles leads to a significant load transfer from the rim to the centre of the oblate sides of the ellipsoidal particles. The findings of these pebble-scale investigations provided the basis for outcropscale modelling, where simulated gravel layers were subjected to layer-parallel extension. These outcrop-scale models revealed the existence of a particle breakage enhancing mechanism that becomes active during early stages of shear band formation. The interaction of such shear bands with the less deformed host material results in particle stress concentrations and subsequently

Lead-induced DNA damage in Vicia faba root cells: Potential involvement of oxidative stress

OpenAIRE

Pourrut, Bertrand; Jean, Séverine; Silvestre, Jérôme; Pinelli, Eric

2011-01-01

Genotoxic effects of lead (0–20 µM) were investigated in whole-plant roots of Vicia faba L., grown hydroponically under controlled conditions. Lead-induced DNA damage in V. faba roots was evaluated by use of the comet assay, which allowed the detection of DNA strand-breakage and with the V. faba micronucleus test, which revealed chromosome aberrations. The results clearly indicate that lead induced DNA fragmentation in a dose-dependant manner with a maximum effect at 10 µM. In addition, at th...

X-ray induced DNA double strand break production and repair in mammalian cells as measured by neutral filter elution

Energy Technology Data Exchange (ETDEWEB)

Bradley, M O; Kohn, K W [National Institutes of Health, Bethesda, MD (USA)

1979-10-01

A neutral filter elution method was used for detecting DNA double strand breaks in mouse L1210 cells after X-ray. The assay detected the number of double strand breaks induced by as little as 1000 rad of X-ray. The rate of DNA elution through the filters under neutral conditions increased with X-ray dose. Certain conditions for deproteinization, pH, and filter type were shown to increase the assay's sensitivity. Hydrogen peroxide and Bleomycin also induced apparent DNA double strand breaks, although the ratios of double to single strand breaks varied from those produced by X-ray. The introduction of double strand cuts by HpA I restriction endonuclease in DNA lysed on filters resulted in a rapid rate of elution under neutral conditions, implying that the method can detect double strand breaks if they exist in the DNA. The eluted DNA banded with a double stranded DNA marker in cesium chloride. This evidence suggested that the assay detected DNA double strand breaks. L1210 cells were shown to rejoin most of the DNA double strand breaks induced by 5-10 krad of X-ray with a half-time of about 40 minutes. (author).

Coke degradation by surface breakage in a modified tumble drum

Energy Technology Data Exchange (ETDEWEB)

Litster, J D

1987-01-01

The surface breakage rate constant for three Australian battery cokes was measured in a specially modified tumble drum using a previously developed technique. The effect of experimental test parameters - coke size, sample mass, drum speed, lifter height and lifter number - on the surface breakage rate constant was examined. The motion of coke particles within a tumble drum was filmed in a simulation experiment with a 0.31 m diameter drum. Particles were raised on the lifters, fell and collided with the bottom of the drum. These collisions were the main source of fines (minus 1 mm) production rather than true abrasion as depicted by a rubbing, rolling action. Hence the term 'surface breakage' is more appropriate than 'abrasion' to describe the breakage process. By measuring the volume of coke carried by each lifter and the height of fall of the coke, the effect of drum speed, sample mass, lifter height and number on the rate of surface breakage was successfully explained. The surface breakage rate constant was found to be proportional to particle size to the power 0.33 for the three cokes studied. A normalized surface breakage rate constant was derived which allowed comparison of cokes with different size distributions. This parameter characterises the coke surface breakage resistance.

Titanium dioxide nanoparticles induce oxidative stress and DNA-adduct formation but not DNA-breakage in human lung cells

Directory of Open Access Journals (Sweden)

Schins Roel PF

2009-06-01

Full Text Available Abstract Titanium dioxide (TiO2, also known as titanium (IV oxide or anatase, is the naturally occurring oxide of titanium. It is also one of the most commercially used form. To date, no parameter has been set for the average ambient air concentration of TiO2 nanoparticles (NP by any regulatory agency. Previously conducted studies had established these nanoparticles to be mainly non-cyto- and -genotoxic, although they had been found to generate free radicals both acellularly (specially through photocatalytic activity and intracellularly. The present study determines the role of TiO2-NP (anatase, ∅ in vitro. For comparison, iron containing nanoparticles (hematite, Fe2O3, ∅ 2-NP did not induce DNA-breakage measured by the Comet-assay in both cell types. Generation of reactive oxygen species (ROS was measured acellularly (without any photocatalytic activity as well as intracellularly for both types of particles, however, the iron-containing NP needed special reducing conditions before pronounced radical generation. A high level of DNA adduct formation (8-OHdG was observed in IMR-90 cells exposed to TiO2-NP, but not in cells exposed to hematite NP. Our study demonstrates different modes of action for TiO2- and Fe2O3-NP. Whereas TiO2-NP were able to generate elevated amounts of free radicals, which induced indirect genotoxicity mainly by DNA-adduct formation, Fe2O3-NP were clastogenic (induction of DNA-breakage and required reducing conditions for radical formation.

Correlations of DNA strand breaks and their repair with cell survival following acute exposure to mercury(II) and X-rays

International Nuclear Information System (INIS)

Cantoni, O.; Costa, M.

1983-01-01

Alkaline elution analysis demonstrates that both HgCl 2 and X-rays result in a rapid induction of DNA single-strand breaks at acutely cytotoxic doses (HgCl 2 , 25-100 microM for 60 min; X-rays, 150-600 rads) in cultured Chinese hamster ovary cells. Cytotoxicity, as measured by cell-plating efficiency, correlates linearly with the level of DNA breakage induced by both agents (HgCl 2 , r . 0.97; X-rays, r . 0.99), although a substantial difference in axis intercepts of the two linear regression lines indicates that a higher level of DNA damage was required by X-rays as compared with HgCl 2 to produce an equivalent level of cell killing. DNA damage induced by X-rays was rapidly repaired such that within 1 hr following treatment the elution rate of DNA from treated cells resembled that obtained in untreated cultures. In contrast, DNA damage after Hg 2+ insult was not repaired, and further damage was evident following a similar 1-hr recovery period. Addition of noncytotoxic, non-DNA-damaging concentrations of HgCl 2 (10 microM) to cells 15-45 min following treatment with X-rays greatly inhibited the repair of the DNA strand breaks. Thus, although both HgCl 2 and X-rays induce rapid and striking single-strand breaks in the DNA, persistence of Hg 2+ in the cell can inhibit the repair of these breaks. The inhibition of DNA repair by HgCl 2 may explain why this agent is not severely mutagenic or carcinogenic despite its ability to induce an X-ray-like DNA damage and why a lower level of mercury-induced DNA damage, compared with that induced by X-rays, was required to produce an equivalent level of cell death

DNA damage induced in mouse peritoneal exudate cells after in vivo administration of chemical and physical agents as determined by alkaline elution

International Nuclear Information System (INIS)

Nishi, Yoshisuke; Miyanaga, Kumiko; Sato, Sei-ichi; Inui, Naomichi

1990-01-01

The alkaline elution technique for detecting DNA strand breaks has been applied to the study of DNA damage in mouse peritoneal exudate cells resulting from the in vivo administration of chemical and physical agents. The direct methylating agents methyl methanesulphonate and N-methyl-N-nitrosourea induced extensive breakage in samples taken 2 h after administration. The direct ethylating agents ethyl methanesulphonate and N-ethyl-N-nitrosourea also induced DNA strand breaks, but to a lesser extent than the methylating agents. The indirect methylating agent dimethylnitrosamine showed hardly any effect in this system. A weak but positive response was observed upon treatment with the anti-neoplastic alkylating agent procarbazine hydrochloride. The whole-body irradiation of mice with 60 Co γ-rays also induced DNA strand breaks. The elution profiles for γ-ray irradiation were different from those of alkylating agents, and indicate that alkylating agents produce many more secondary lesions leading to DNA strand breaks than γ-rays. N-methyl-N-nitrosourea produced slightly more DNA strand breaks in mutagen-sensitive mice, which are derived from the CD-1 strain, than in ICR mice. (Author)

DNA damage induced in mouse peritoneal exudate cells after in vivo administration of chemical and physical agents as determined by alkaline elution

Energy Technology Data Exchange (ETDEWEB)

Nishi, Yoshisuke (Japan Tobacco Inc., Yokohama (Japan). Central Research Inst.); Miyanaga, Kumiko; Sato, Sei-ichi (Japan Tobacco Inc., Hatano, Kanagawa (Japan). Toxicology Research Lab.); Inui, Naomichi (Japan Tobacco Inc., Yokohama, Kanagawa (Japan). Pharmaceutical Research Labs.)

1990-01-01

The alkaline elution technique for detecting DNA strand breaks has been applied to the study of DNA damage in mouse peritoneal exudate cells resulting from the in vivo administration of chemical and physical agents. The direct methylating agents methyl methanesulphonate and N-methyl-N-nitrosourea induced extensive breakage in samples taken 2 h after administration. The direct ethylating agents ethyl methanesulphonate and N-ethyl-N-nitrosourea also induced DNA strand breaks, but to a lesser extent than the methylating agents. The indirect methylating agent dimethylnitrosamine showed hardly any effect in this system. A weak but positive response was observed upon treatment with the anti-neoplastic alkylating agent procarbazine hydrochloride. The whole-body irradiation of mice with {sup 60}Co {gamma}-rays also induced DNA strand breaks. The elution profiles for {gamma}-ray irradiation were different from those of alkylating agents, and indicate that alkylating agents produce many more secondary lesions leading to DNA strand breaks than {gamma}-rays. N-methyl-N-nitrosourea produced slightly more DNA strand breaks in mutagen-sensitive mice, which are derived from the CD-1 strain, than in ICR mice. (Author).

Distinct genetic control of homologous recombination repair of Cas9-induced double-strand breaks, nicks and paired nicks

NARCIS (Netherlands)

Vriend, Lianne E. M.; Prakash, Rohit; Chen, Chun-Chin; Vanoli, Fabio; Cavallo, Francesca; Zhang, Yu; Jasin, Maria; Krawczyk, Przemek M.

2016-01-01

DNA double-strand breaks (DSBs) are known to be powerful inducers of homologous recombination (HR), but single-strand breaks (nicks) have also been shown to trigger HR. Both DSB- and nick-induced HR ((nick)HR) are exploited in advanced genome-engineering approaches based on the bacterial RNA-guided

Breaks in plasmid DNA strand induced by laser radiation at a wavelength of 193 nm

International Nuclear Information System (INIS)

Gurzadyan, G.G.; Shul'te Frolinde, D.

1996-01-01

DNA of plasmid pB322 irradiated with laser at a wavelength of 193 nm was treated with an extract containing proteins from E.coli K12 AB1157 (wild-type). The enzymes were found to produce single- and double-strand DNA breaks, which was interpreted as a transformation of a portion of cyclobutane pyrimidine dimers and (6-4) photoproducts into nonrepairable single-strand DNA breaks. The products resulted from ionization of DNA, in particular, single-strand breaks, transform to double-strand breaks. A comparison of these data with the data on survival of plasmid upon transformation of E.coli K12 AB1157 enables one to assess the biological significance of single- and double-strand breaks. The inactivation of the plasmid is mainly determined by the number of directly formed laser-induced single-strand breaks. 26 refs.; 2 figs

Normal formation and repair of γ-radiation-induced single and double strand DNA breaks in Down syndrome fibroblasts

International Nuclear Information System (INIS)

Steiner, M.E.; Woods, W.G.

1982-01-01

Fibroblasts from patients with Down syndrome (Trisomy 21) were examined for repair capability of γ-radiation-induced single strand and double strand DNA breaks. Formation and repair of DNA breaks were determined by DNA alkaline and non-denaturing elution techniques. Down syndrome fibroblasts were found to repair single strand and double strand breaks as well as fibroblasts from normal controls. (orig.)

Combined quantum-mechanics/molecular-mechanics dynamics simulation of A-DNA double strands irradiated by ultra-low-energy carbon ions

Energy Technology Data Exchange (ETDEWEB)

Ngaojampa, C.; Nimmanpipug, P. [Computer Simulation and Modeling Laboratory (CSML), Department of Chemistry and Center for Innovation Chemistry, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Yu, L.D., E-mail: yuld@fnrf.science.cmu.ac.t [Plasma and Beam Physics Research Facility, Department of Physics and Materials Science, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Thailand Center of Excellence in Physics, Commission on Higher Education, 328 Si Ayutthaya Road, Bangkok 10400 (Thailand); Anuntalabhochai, S. [Molecular Biology Laboratory, Department of Biology, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Lee, V.S., E-mail: vannajan@gmail.co [Computer Simulation and Modeling Laboratory (CSML), Department of Chemistry and Center for Innovation Chemistry, Faculty of Science, Chiang Mai University, Chiang Mai 50200 (Thailand); Thailand Center of Excellence in Physics, Commission on Higher Education, 328 Si Ayutthaya Road, Bangkok 10400 (Thailand)

2011-02-15

In order to promote understanding of the fundamentals of ultra-low-energy ion interaction with DNA, molecular dynamics simulations using combined quantum-mechanics/molecular-mechanics of poly-AT and poly-GC A-DNA double strands irradiated by <200 eV carbon ions were performed to investigate the molecular implications of mutation bias. The simulations were focused on the responses of the DNA backbones and nitrogenous bases to irradiation. Analyses of the root mean square displacements of the backbones and non-hydrogen atoms of base rings of the simulated DNA structure after irradiation revealed a potential preference of DNA double strand separation, dependent on the irradiating energy. The results show that for the backbones, the large difference in the displacement between poly-GC and poly-AT in the initial time period could be the reason for the backbone breakage; for the nitrogenous base pairs, A-T is 30% more sensitive or vulnerable to ion irradiation than G-C, demonstrating a preferential, instead of random, effect of irradiation-induced mutation.

Combined quantum-mechanics/molecular-mechanics dynamics simulation of A-DNA double strands irradiated by ultra-low-energy carbon ions

International Nuclear Information System (INIS)

Ngaojampa, C.; Nimmanpipug, P.; Yu, L.D.; Anuntalabhochai, S.; Lee, V.S.

2011-01-01

In order to promote understanding of the fundamentals of ultra-low-energy ion interaction with DNA, molecular dynamics simulations using combined quantum-mechanics/molecular-mechanics of poly-AT and poly-GC A-DNA double strands irradiated by <200 eV carbon ions were performed to investigate the molecular implications of mutation bias. The simulations were focused on the responses of the DNA backbones and nitrogenous bases to irradiation. Analyses of the root mean square displacements of the backbones and non-hydrogen atoms of base rings of the simulated DNA structure after irradiation revealed a potential preference of DNA double strand separation, dependent on the irradiating energy. The results show that for the backbones, the large difference in the displacement between poly-GC and poly-AT in the initial time period could be the reason for the backbone breakage; for the nitrogenous base pairs, A-T is 30% more sensitive or vulnerable to ion irradiation than G-C, demonstrating a preferential, instead of random, effect of irradiation-induced mutation.

Radiation sensitization by an iodine-labelled DNA ligand

Energy Technology Data Exchange (ETDEWEB)

Martin, R F; Murray, V; D' Cunha, G; Pardee, M; Haigh, A; Hodgson, G S [Peter MacCallum Cancer Inst., Melbourne (Australia); Kampouris, E; Kelly, D P [Melbourne Univ., Parkville (Australia)

1990-05-01

An iodinated DNA ligand, iodoHoechst 33258, which binds in the minor groove of DNA, enhances DNA strand breakage and cell killing by UV-A irradiation. The sites of UV-induced strand breaks reflect the known sequence specificity of the ligand. (author).

Acute hypoxia and hypoxic exercise induce DNA strand breaks and oxidative DNA damage in humans

DEFF Research Database (Denmark)

Møller, P; Loft, S; Lundby, C

2001-01-01

; lymphocytes were isolated for analysis of DNA strand breaks and oxidatively altered nucleotides, detected by endonuclease III and formamidipyridine glycosylase (FPG) enzymes. Urine was collected for 24 h periods for analysis of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), a marker of oxidative DNA damage...... oxygen species, generated by leakage of the mitochondrial respiration or during a hypoxia-induced inflammation. Furthermore, the presence of DNA strand breaks may play an important role in maintaining hypoxia-induced inflammation processes. Hypoxia seems to deplete the antioxidant system of its capacity...

Photosensitization by iodinated DNA minor groove binding ligands: Evaluation of DNA double-strand break induction and repair.

Science.gov (United States)

Briggs, Benjamin; Ververis, Katherine; Rodd, Annabelle L; Foong, Laura J L; Silva, Fernando M Da; Karagiannis, Tom C

2011-05-03

Iodinated DNA minor groove binding bibenzimidazoles represent a unique class of UVA photosensitizer and their extreme photopotency has been previously characterized. Earlier studies have included a comparison of three isomers, referred to as ortho-, meta- and para-iodoHoechst, which differ only in the location of the iodine substituent in the phenyl ring of the bibenzimidazole. DNA breakage and clonogenic survival studies in human erythroleukemic K562 cells have highlighted the higher photo-efficiency of the ortho-isomer (subsequently designated UV(A)Sens) compared to the meta- and para-isomers. In this study, the aim was to compare the induction and repair of DNA double-strand breaks induced by the three isomers in K562 cells. Further, we examined the effects of the prototypical broad-spectrum histone deacetylase inhibitor, Trichostatin A, on ortho-iodoHoechst/UVA-induced double-strand breaks in K562 cells. Using γH2AX as a molecular marker of the DNA lesions, our findings indicate a disparity in the induction and particularly, in the repair kinetics of double-strand breaks for the three isomers. The accumulation of γH2AX foci induced by the meta- and para-isomers returned to background levels within 24 and 48 h, respectively; the number of γH2AX foci induced by ortho-iodoHoechst remained elevated even after incubation for 96 h post-irradiation. These findings provide further evidence that the extreme photopotency of ortho-iodoHoechst is due to not only to the high quantum yield of dehalogenation, but also to the severity of the DNA lesions which are not readily repaired. Finally, our findings which indicate that Trichostatin A has a remarkable potentiating effect on ortho-iodoHoechst/UVA-induced DNA lesions are encouraging, particularly in the context of cutaneous T-cell lymphoma, for which a histone deacetylase inhibitor is already approved for therapy. This finding prompts further evaluation of the potential of combination therapies. Copyright © 2011

Effect of particle breakage on cyclic densification of ballast: A DEM approach

International Nuclear Information System (INIS)

Thakur, P K; Vinod, J S; Indraratna, B

2010-01-01

In this paper, an attempt has been made to investigate the effect of particle breakage on densification behaviour of ballast under cyclic loading using Discrete Element Method (DEM). Numerical simulations using PFC 2D have been carried out on an assembly of angular particles with and without incorporation of particle breakage. Two-dimensional projection of angular ballast particles were simulated using clusters of bonded circular particles. Degradation of the bonds within a cluster was considered to represent particle breakage. Clump logic was used to make the cluster of particles unbreakable. DEM simulation results highlight that the particle breakage has a profound influence on the cyclic densification behaviour of ballast. The deformation behaviour exhibited by the assembly with breakage is in good agreement with the laboratory experiments. In addition, the evolution of particle displacement vectors clearly explains the breakage mechanism and associated deformations during cyclic loading.

Repair pathways for heavy ion-induced complex DNA double strand breaks

International Nuclear Information System (INIS)

Yajima, Hirohiko; Nakajima, Nakako; Hirakawa, Hirokazu; Murakami, Takeshi; Okayasu, Ryuichi; Fujimori, Akira

2012-01-01

DNA double strand break (DSB) induced by ionizing radiation (IR) is a deleterious damage leading to cell death and genome instability if not properly repaired. It is well known that DSB is repaired by two major pathways, non-homologous end-joining (NHEJ) and homologous recombination (HR). It is also known that NHEJ is dominant throughout the cell cycle after X- or gamma-ray irradiation in mammalian cells, Meanwhile, it is thought that heavy-ion radiation (e.g., carbon-ions, iron-ions) gives rise to clustered DNA damages consisting of not only strand breaks but also aberrant bases in the vicinity of DSBs (complex DSBs). Our previous work suggested that the efficiency of NHEJ is diminished for repair of complex DSBs induced by heavy-ion radiation. We thought that this difficulty in NHEJ process associated with heavy ion induced complex DNA damage might be extended to HR process in cells exposed to heavy ions. In order to find out if this notion is true or not, exposed human cells to X-rays and heavy-ions, and studied HR associated processes at the molecular level. Our result indicates that complex DSBs induced by heavy ions effectively evoke DNA end resection activity during the HR process. Together with our results, a relevant recent progress in the field of DNA DSB repair will be discussed. (author)

Determinants of condom breakage among female sex workers in Karnataka, India

Directory of Open Access Journals (Sweden)

Bradley Janet

2011-12-01

Full Text Available Abstract Background Condoms are effective in preventing the transmission of HIV and other sexually transmitted infections, when properly used. However, recent data from surveys of female sex workers (FSWs in Karnataka in south India, suggest that condom breakage rates may be quite high. It is important therefore to quantify condom breakage rates, and examine what factors might precipitate condom breakage, so that programmers can identify those at risk, and develop appropriate interventions. Methods We explored determinants of reported condom breakage in the previous month among 1,928 female sex workers in four districts of Karnataka using data from cross-sectional surveys undertaken from July 2008 to February 2009. Using stepwise multivariate logistic regression, we examined the possible determinants of condom breakage, controlling for several independent variables including the district and client load. Results Overall, 11.4% of FSWs reported at least one condom break in the previous month. FSWs were much more likely to report breakage if under 20 years of age (AOR 3.43, p = 0.005; if divorced/ separated/widowed (AOR 1.52, p = 0.012; if they were regular alcohol users (AOR 1.63, p = 0.005; if they mostly entertained clients in lodges/rented rooms (AOR 2.99, p = 0.029 or brothels (AOR 4.77, p = 0.003, compared to street based sex workers; if they had ever had anal sex (AOR 2.03, p = 0.006; if the sex worker herself (as opposed to the client applied the condom at last use (AOR 1.90, p Conclusions The reported incidence of condom breakage was high in this study, and this is a major concern for HIV/STI prevention programs, for which condom use is a key prevention tool. Younger and more marginalized female sex workers were most vulnerable to condom breakage. Special effort is therefore required to seek out such women and to provide information and skills on correct condom use. More research is also needed on what specific situational parameters

The role of glutathione in DNA damage by potassium bromate in vitro.

Science.gov (United States)

Parsons, J L; Chipman, J K

2000-07-01

We have investigated the role of reduced glutathione (GSH) in the genetic toxicity of the rodent renal carcinogen potassium bromate (KBrO(3)). A statistically significant increase in the concentration of 8-oxodeoxyguanosine (8-oxodG) relative to deoxyguanosine was measured following incubation of calf thymus DNA with KBrO(3) and GSH or N-acetylcysteine (NACys). This was dependent on these thiols and was associated with the loss of GSH and production of oxidized glutathione. A short-lived (potassium chlorate (KClO(3)) or potassium iodate (KIO(3)) were used instead of KBrO(3), though GSH depletion also occurred with KIO(3), but not with KClO(3). Other reductants and thiols in combination with KBrO(3) did not cause a significant increase in DNA oxidation. DNA strand breakage was also induced by KBrO(3) in human white blood cells (5 mM) and rat kidney epithelial cells (NRK-52E, 1.5 mM). This was associated with an apparent small depletion of thiols in NRK-52E cells at 15 min and with an elevation of 8-oxodG at a delayed time of 24 h. Depletion of intra-cellular GSH by diethylmaleate in human lymphocytes decreased the amount of strand breakage induced by KBrO(3). Extracellular GSH, however, protected against DNA strand breakage by KBrO(3), possibly due to the inability of the reactive product to enter the cell. In contrast, membrane-permeant NACys enhanced KBrO(3)-induced DNA strand breakage in these cells. DNA damage by KBrO(3) is therefore largely dependent on access to intracellular GSH.

Electrophoresis examination of strand breaks in plasmid DNA induced by low-energy nitrogen ion irradiation

International Nuclear Information System (INIS)

Zhao Yong; Tan Zheng; Du Yanhua; Qiu Guanying

2003-01-01

To study the effect on plasmid DNA of heavy ion in the energy range of keV where nuclear stopping interaction becomes more important or even predominant, thin film of plasmid pGEM-3Zf(-) DNA was prepared on aluminum surface and irradiated in vacuum ( -3 Pa) by low-energy nitrogen ions with energy of 30 keV (LET=285 keV/μm) at various fluence ranging from 2 x 10 10 to 8.2 x 10 13 ions/cm 2 . DNA strand breaks were analyzed by neutral electrophoresis followed by quantification with image analysis software. Low-energy nitrogen ion irradiation induced single-, double- and multiple double-strand breaks (DSB) and multiple DSB as the dominating form of DNA damages. Moreover, the linear fluence-response relationship at a low fluence range suggests that DSBs are induced predominantly by single ion track. However, strand break production is limited to a short range in the irradiated samples

Theoretical analysis on ac loss properties of two-strand parallel conductors composed of superconducting multifilamentary strands

CERN Document Server

Iwakuma, M; Funaki, K

2002-01-01

The ac loss properties of two-strand parallel conductors composed of superconducting multifilamentary strands were theoretically investigated. The constituent strands generally need to be insulated and transposed for the sake of uniform current distribution and low ac loss. In case the transposition points deviate from the optimum ones, shielding current is induced according to the interlinkage magnetic flux of the twisted loop enclosed by the insulated strands and the contact resistances at the terminals. It produces an additional ac loss. Supposing a simple situation where a two-strand parallel conductor with one-point transposition is exposed to a uniform ac magnetic field, the basic equations for the magnetic field were proposed and the theoretical expressions of the additional ac losses derived. As a result, the following features were shown. The additional ac loss in the non-saturation case, where the induced shielding current is less than the critical current of a strand, is proportional to the square ...

Empirical Formulae for Breakage of Dolosse and Tetrapods

OpenAIRE

Burcharth, H. F.; d'Angremond, K.; Meer, W. van der; Liu, Z.

2000-01-01

The slender, complex types of armour units, such as Tetrapods and Dolosse are widely used for rubble mound breakwaters. Many failures of such breakwaters were caused by unforeseen early breakage of the units, thus revealing an inbalance between the strength (structural integrity) of the units and the hydraulic stability (resistance to displacements) of the armour layers. Breakage occurs when the stresses from the static, pulsating and impact loads exceeds the tensile strength of the concrete....

An increase in pectin methyl esterase activity accompanies dormancy breakage and germination of yellow cedar seeds.

Science.gov (United States)

Ren, C; Kermode, A R

2000-09-01

Pectin methyl esterase (PME) (EC 3.1.1.11) catalyzes the hydrolysis of methylester groups of cell wall pectins. We investigated the role of this enzyme in dormancy termination and germination of yellow cedar (Chamaecyparis nootkatensis [D. Don] Spach) seeds. PME activity was not detected in dormant seeds of yellow cedar but was induced and gradually increased during moist chilling; high activity coincided with dormancy breakage and germination. PME activity was positively correlated to the degree of dormancy breakage of yellow cedar seeds. The enzyme produced in different seed parts and in seeds at different times during moist chilling, germination, and early post-germinative growth consisted of two isoforms, both basic with isoelectric points of 8.7 and 8.9 and the same molecular mass of 62 kD. The pH optimum for the enzyme was between 7.4 and 8.4. In intact yellow cedar seeds, activities of the two basic isoforms of PME that were induced in embryos and in megagametophytes following dormancy breakage were significantly suppressed by abscisic acid. Gibberellic acid had a stimulatory effect on the activities of these isoforms in embryos and megagametophytes of intact seeds at the germinative stage. We hypothesize that PME plays a role in weakening of the megagametophyte, allowing radicle emergence and the completion of germination.

Determinants of condom breakage among female sex workers in Karnataka, India.

Science.gov (United States)

Bradley, Janet; Rajaram, S; Alary, Michel; Isac, Shajy; Washington, Reynold; Moses, Stephen; Ramesh, B M

2011-12-29

Condoms are effective in preventing the transmission of HIV and other sexually transmitted infections, when properly used. However, recent data from surveys of female sex workers (FSWs) in Karnataka in south India, suggest that condom breakage rates may be quite high. It is important therefore to quantify condom breakage rates, and examine what factors might precipitate condom breakage, so that programmers can identify those at risk, and develop appropriate interventions. We explored determinants of reported condom breakage in the previous month among 1,928 female sex workers in four districts of Karnataka using data from cross-sectional surveys undertaken from July 2008 to February 2009. Using stepwise multivariate logistic regression, we examined the possible determinants of condom breakage, controlling for several independent variables including the district and client load. Overall, 11.4% of FSWs reported at least one condom break in the previous month. FSWs were much more likely to report breakage if under 20 years of age (AOR 3.43, p = 0.005); if divorced/ separated/widowed (AOR 1.52, p = 0.012); if they were regular alcohol users (AOR 1.63, p = 0.005); if they mostly entertained clients in lodges/rented rooms (AOR 2.99, p = 0.029) or brothels (AOR 4.77, p = 0.003), compared to street based sex workers; if they had ever had anal sex (AOR 2.03, p = 0.006); if the sex worker herself (as opposed to the client) applied the condom at last use (AOR 1.90, p < 0.001); if they were inconsistent condom users (AOR 2.77, p < 0.001); and if they had never seen a condom demonstration (AOR 2.37, p < 0.001). The reported incidence of condom breakage was high in this study, and this is a major concern for HIV/STI prevention programs, for which condom use is a key prevention tool. Younger and more marginalized female sex workers were most vulnerable to condom breakage. Special effort is therefore required to seek out such women and to provide information and skills on correct

Quantitation of the repair of gamma-radiation-induced double-strand DNA breaks in human fibroblasts

International Nuclear Information System (INIS)

Woods, W.G.

1981-01-01

The quantitation and repair of double-strand DNA breaks in human fibroblasts has been determined using a method involving the nondenaturing elution of DNA from a filter. DNA from cells from two human fibroblast lines exposed to γ-radiation from 0 to 10000 rad showed increasing retention on a filter with decreasing radiation dose, and the data suggest a linear relationship between double-strand breaks induced and radiation dose. The ability of normal human fibroblasts to repair double-strand breaks with various doses of radiation was demonstrated, with a tsub(1/2) of 10 min for repair of 5000 rad exposure and 39 min for repair of 10000 rad damage. The kinetics of the DNA rejoining were not linear and suggest that, as in the repair of single-strand breaks, both an initial fast and a later slow mechanism may be involved. (Auth.)

Fiber breakage phenomena in long fiber reinforced plastic preparation

International Nuclear Information System (INIS)

Huang, Chao-Tsai; Tseng, Huan-Chang; Chang, Rong-Yeu; Vlcek, Jiri

2015-01-01

Due to the high demand of smart green, the lightweight technologies have become the driving force for the development of automotives and other industries in recent years. Among those technologies, using short and long fiber-reinforced plastics (FRP) to replace some metal components can reduce the weight of an automotive significantly. However, the microstructures of fibers inside plastic matrix are too complicated to manage and control during the injection molding through the screw, the runner, the gate, and then into the cavity. This study focuses on the fiber breakage phenomena during the screw plastification. Results show that fiber breakage is strongly dependent on screw design and operation. When the screw geometry changes, the fiber breakage could be larger even with lower compression ratio. (paper)

Non-homologous end joining is the responsible pathway for the repair of fludarabine-induced DNA double strand breaks in mammalian cells

International Nuclear Information System (INIS)

Campos-Nebel, Marcelo de; Larripa, Irene; Gonzalez-Cid, Marcela

2008-01-01

Fludarabine (FLU), an analogue of adenosine, interferes with DNA synthesis and inhibits the chain elongation leading to replication arrest and DNA double strand break (DSB) formation. Mammalian cells use two main pathways of DSB repair to maintain genomic stability: homologous recombination (HR) and non-homologous end joining (NHEJ). The aim of the present work was to evaluate the repair pathways employed in the restoration of DSB formed following replication arrest induced by FLU in mammalian cells. Replication inhibition was induced in human lymphocytes and fibroblasts by FLU. DSB occurred in a dose-dependent manner on early/middle S-phase cells, as detected by γH2AX foci formation. To test whether conservative HR participates in FLU-induced DSB repair, we measured the kinetics of Rad51 nuclear foci formation in human fibroblasts. There was no significant induction of Rad51 foci after FLU treatment. To further confirm these results, we analyzed the frequency of sister chromatid exchanges (SCE) in both human cells. We did not find increased frequencies of SCE after FLU treatment. To assess the participation of NHEJ pathway in the repair of FLU-induced damage, we used two chemical inhibitors of the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs), vanillin and wortmannin. Human fibroblasts pretreated with DNA-PKcs inhibitors showed increased levels of chromosome breakages and became more sensitive to cell death. An active role of NHEJ pathway was also suggested from the analysis of Chinese hamster cell lines. XR-C1 (DNA-PKcs-deficient) and XR-V15B (Ku80-deficient) cells showed hypersensitivity to FLU as evidenced by the increased frequency of chromosome aberrations, decreased mitotic index and impaired survival rates. In contrast, CL-V4B (Rad51C-deficient) and V-C8 (Brca2-deficient) cell lines displayed a FLU-resistant phenotype. Together, our results suggest a major role for NHEJ repair in the preservation of genome integrity against FLU-induced DSB

Non-homologous end joining is the responsible pathway for the repair of fludarabine-induced DNA double strand breaks in mammalian cells

Energy Technology Data Exchange (ETDEWEB)

Campos-Nebel, Marcelo de [Departamento de Genetica, Instituto de Investigaciones Hematologicas Mariano R. Castex, Academia Nacional de Medicina, Buenos Aires (Argentina)], E-mail: mnebel@hematologia.anm.edu.ar; Larripa, Irene; Gonzalez-Cid, Marcela [Departamento de Genetica, Instituto de Investigaciones Hematologicas Mariano R. Castex, Academia Nacional de Medicina, Buenos Aires (Argentina)

2008-11-10

Fludarabine (FLU), an analogue of adenosine, interferes with DNA synthesis and inhibits the chain elongation leading to replication arrest and DNA double strand break (DSB) formation. Mammalian cells use two main pathways of DSB repair to maintain genomic stability: homologous recombination (HR) and non-homologous end joining (NHEJ). The aim of the present work was to evaluate the repair pathways employed in the restoration of DSB formed following replication arrest induced by FLU in mammalian cells. Replication inhibition was induced in human lymphocytes and fibroblasts by FLU. DSB occurred in a dose-dependent manner on early/middle S-phase cells, as detected by {gamma}H2AX foci formation. To test whether conservative HR participates in FLU-induced DSB repair, we measured the kinetics of Rad51 nuclear foci formation in human fibroblasts. There was no significant induction of Rad51 foci after FLU treatment. To further confirm these results, we analyzed the frequency of sister chromatid exchanges (SCE) in both human cells. We did not find increased frequencies of SCE after FLU treatment. To assess the participation of NHEJ pathway in the repair of FLU-induced damage, we used two chemical inhibitors of the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs), vanillin and wortmannin. Human fibroblasts pretreated with DNA-PKcs inhibitors showed increased levels of chromosome breakages and became more sensitive to cell death. An active role of NHEJ pathway was also suggested from the analysis of Chinese hamster cell lines. XR-C1 (DNA-PKcs-deficient) and XR-V15B (Ku80-deficient) cells showed hypersensitivity to FLU as evidenced by the increased frequency of chromosome aberrations, decreased mitotic index and impaired survival rates. In contrast, CL-V4B (Rad51C-deficient) and V-C8 (Brca2-deficient) cell lines displayed a FLU-resistant phenotype. Together, our results suggest a major role for NHEJ repair in the preservation of genome integrity against FLU-induced

Nick translation detection in situ of cellular DNA strand break induced by radiation

International Nuclear Information System (INIS)

Maehara, Y.; Anai, H.; Kusumoto, T.; Sakaguchi, Y.; Sugimachi, K.

1989-01-01

DNA strand break in HeLa cells induced by radiation was detected using the in situ nick translation method. The cells were exposed to radiation of 3, 6, 12, 18, and 24 Gy in Lab-Tek tissue culture chamber/slides and were fixed with ethanol/acetic acid on the slide glass. The break sites in DNA were translated artificially in the presence of Escherichia coli DNA polymerase I and [ 3 H]-labeled dTTP. Autoradiographic observation was made of the level of break sites in the DNA. The DNA strand break appeared even with a 3 Gy exposure, increased 8.6 times at 24 Gy compared with the control cells, and this level correlated reciprocally to change in cell viability. This nick translation method provides a rapid in situ assay for determining radiation-induced DNA damage of cultured cells, in a semi-quantitative manner

Interrelating the breakage and composition of mined and drill core coal

Science.gov (United States)

Wilson, Terril Edward

Particle size distribution of coal is important if the coal is to be beneficiated, or if a coal sales contract includes particle size specifications. An exploration bore core sample of coal ought to be reduced from its original cylindrical form to a particle size distribution and particle composition that reflects, insofar as possible, a process stream of raw coal it represents. Often, coal cores are reduced with a laboratory crushing machine, the product of which does not match the raw coal size distribution. This study proceeds from work in coal bore core reduction by Australian investigators. In this study, as differentiated from the Australian work, drop-shatter impact breakage followed by dry batch tumbling in steel cylinder rotated about its transverse axis are employed to characterize the core material in terms of first-order and zeroth-order breakage rate constants, which are indices of the propensity of the coal to degrade during excavation and handling. Initial drop-shatter and dry tumbling calibrations were done with synthetic cores composed of controlled low-strength concrete incorporating fly ash (as a partial substitute for Portland cement) in order to reduce material variables and conserve difficult-to-obtain coal cores. Cores of three different coalbeds--Illinois No. 6, Upper Freeport, and Pocahontas No. 5 were subjected to drop-shatter and dry batch tumbling tests to determine breakage response. First-order breakage, characterized by a first-order breakage index for each coal, occurred in the drop-shatter tests. First- and zeroth-order breakage occurred in dry batch tumbling; disappearance of coarse particles and creation of fine particles occurred in a systematic way that could be represented mathematically. Certain of the coal cores available for testing were dry and friable. Comparison of coal preparation plant feed with a crushed bore core and a bore core prepared by drop-shatter and tumbling (all from the same Illinois No.6 coal mining

Dependence of rates of breakage on fines content in wet ball mill grinding

Science.gov (United States)

Bhattacharyya, Anirban

The following research fundamentally deals with the cause and implications of nonlinearities in breakage rates of materials in wet grinding systems. The innate dependence of such nonlinearities on fines content and the milling environment during wet grinding operations is also tested and observed. Preferential breakage of coarser size fractions as compared to the finer size fractions in a particle population were observed and discussed. The classification action of the pulp was deemed to be the probable cause for such a peculiarity. Ores with varying degrees of hardness and brittleness were used for wet grinding experiments, primarily to test the variations in specific breakage rates as a function of varying hardness. For this research, limestone, quartzite, and gold ore were used. The degree of hardness is of the order of: limestone, quartzite, gold ore. Selection and breakage function parameters were determined in the course of this research. Functional forms of these expressions were used to compare experimentally derived parameter estimates. Force-fitting of parameters was not done in order to examine the realtime behavior of particle populations in wet grinding systems. Breakage functions were established as being invariant with respect to such operating variables like ball load, mill speed, particle load, and particle size distribution of the mill. It was also determined that specific selection functions were inherently dependent on the particle size distribution in wet grinding systems. Also, they were consistent with inputs of specific energy, according to grind time. Nonlinearity trends were observed for 1st order specific selection functions which illustrated variations in breakage rates with incremental inputs of grind time and specific energy. A mean particle size called the fulcrum was noted below which the nonlinearities in the breakage trends were observed. This magnitude of the fulcrum value varied with percent solids and slurry filling, indicating

Base damage within single-strand DNA underlies in vivo hypermutability induced by a ubiquitous environmental agent.

Directory of Open Access Journals (Sweden)

Kin Chan

Full Text Available Chromosomal DNA must be in single-strand form for important transactions such as replication, transcription, and recombination to occur. The single-strand DNA (ssDNA is more prone to damage than double-strand DNA (dsDNA, due to greater exposure of chemically reactive moieties in the nitrogenous bases. Thus, there can be agents that damage regions of ssDNA in vivo while being inert toward dsDNA. To assess the potential hazard posed by such agents, we devised an ssDNA-specific mutagenesis reporter system in budding yeast. The reporter strains bear the cdc13-1 temperature-sensitive mutation, such that shifting to 37°C results in telomere uncapping and ensuing 5' to 3' enzymatic resection. This exposes the reporter region, containing three closely-spaced reporter genes, as a long 3' ssDNA overhang. We validated the ability of the system to detect mutagenic damage within ssDNA by expressing a modified human single-strand specific cytosine deaminase, APOBEC3G. APOBEC3G induced a high density of substitutions at cytosines in the ssDNA overhang strand, resulting in frequent, simultaneous inactivation of two reporter genes. We then examined the mutagenicity of sulfites, a class of reactive sulfur oxides to which humans are exposed frequently via respiration and food intake. Sulfites, at a concentration similar to that found in some foods, induced a high density of mutations, almost always as substitutions at cytosines in the ssDNA overhang strand, resulting in simultaneous inactivation of at least two reporter genes. Furthermore, sulfites formed a long-lived adducted 2'-deoxyuracil intermediate in DNA that was resistant to excision by uracil-DNA N-glycosylase. This intermediate was bypassed by error-prone translesion DNA synthesis, frequently involving Pol ζ, during repair synthesis. Our results suggest that sulfite-induced lesions in DNA can be particularly deleterious, since cells might not possess the means to repair or bypass such lesions

Binding of Multiple Rap1 Proteins Stimulates Chromosome Breakage Induction during DNA Replication.

Directory of Open Access Journals (Sweden)

Greicy H Goto

2015-08-01

Full Text Available Telomeres, the ends of linear eukaryotic chromosomes, have a specialized chromatin structure that provides a stable chromosomal terminus. In budding yeast Rap1 protein binds to telomeric TG repeat and negatively regulates telomere length. Here we show that binding of multiple Rap1 proteins stimulates DNA double-stranded break (DSB induction at both telomeric and non-telomeric regions. Consistent with the role of DSB induction, Rap1 stimulates nearby recombination events in a dosage-dependent manner. Rap1 recruits Rif1 and Rif2 to telomeres, but neither Rif1 nor Rif2 is required for DSB induction. Rap1-mediated DSB induction involves replication fork progression but inactivation of checkpoint kinase Mec1 does not affect DSB induction. Rap1 tethering shortens artificially elongated telomeres in parallel with telomerase inhibition, and this telomere shortening does not require homologous recombination. These results suggest that Rap1 contributes to telomere homeostasis by promoting chromosome breakage.

X-ray- and TEM-induced mitotic recombination in Drosophila melanogaster: Unequal and sister-strand recombination

International Nuclear Information System (INIS)

Becker, H.J.

1975-01-01

Twin mosaic spots of dark-apricot and light-apricot ommatidia were found in the eyes of wsup(a)/wsup(a) females, of wsup(a) males, of females homozygous for In(1)sc 4 , wsup(a) and of attached-X females homozygous for wsup(a). The flies were raised from larvae which had been treated with 1,630 R of X-rays at the age of 48-52 hours. An additional group of wsup(a)/wsup(a) females and wsup(a) males came from larvae that had been fed with triethylene melamine (TEM) at the age of 22-24 hours. The twin spots apparently were the result of induced unequal mitotic recombination, i.e. from unequal sister-strand recombination in the males and from unequal sister-strand recombination as well as, possibly, unequal recombination between homologous strands in the females. That is, a duplication resulted in wsup(a)Dpwsup(a)/wsup(a) dark-apricto ommatidia and the corresponding deficiency in an adjacent area of wsup(a)/Dfwsup(a) light-apricot ommatidia. In an additional experiment sister-strand mitotic recombination in the ring-X chromosome of ring-X/rod-X females heterozygous for w and wsup(co) is believed to be the cause for X-ray induced single mosaic spots that show the phenotype of the rod-X marker. (orig.) [de

Heavy ion induced DNA strand breaks and their repair in diploid cells of the epithelium of the lens

International Nuclear Information System (INIS)

Heilman, J.

1987-11-01

This diploma thesis investigates by means of alkaline unwinding and neutral elution the induction of DNA strand breaks and of rejoining processes as an effect of irradiation with very heavy, accelerated ions. It is found that: The effectiveness of very heavy ions (Z > 18) increases per particle with higher ordinal number, and with increasing velocities. The relative biological effectiveness increases with higher particle masses and lower velocities. The effects of very heavy ions are determined both by the LET and by the particle track extension (specific energy) of the various particles. Heavy ions are much more effective than X-rays with regard to inducing double strand breaks, as compared to DNA single strand breaks induced. Rejoining processes induced by heavy ions have been found to be delayed and incomplete, as compared to the X-ray effects. The number of rejoining processes decreases with rising ordinal number. The experiments indicate that the irradiation with lead or uranium ions most probably makes rejoining impossible. (orig./MG) [de

Implementation of a method to visualize noise-induced hearing loss in mass stranded cetaceans

Science.gov (United States)

Morell, Maria; Brownlow, Andrew; McGovern, Barry; Raverty, Stephen A.; Shadwick, Robert E.; André, Michel

2017-02-01

Assessment of the impact of noise over-exposure in stranded cetaceans is challenging, as the lesions that lead to hearing loss occur at the cellular level and inner ear cells are very sensitive to autolysis. Distinguishing ante-mortem pathology from post-mortem change has been a major constraint in diagnosing potential impact. Here, we outline a methodology applicable to the detection of noise-induced hearing loss in stranded cetaceans. Inner ears from two mass strandings of long-finned pilot whales in Scotland were processed for scanning electron microscopy observation. In one case, a juvenile animal, whose ears were fixed within 4 hours of death, revealed that many sensory cells at the apex of the cochlear spiral were missing. In this case, the absence of outer hair cells would be compatible with overexposure to underwater noise, affecting the region which transduces the lowest frequencies of the pilot whales hearing spectrum. Perfusion of cochlea with fixative greatly improved preservation and enabled diagnostic imaging of the organ of Corti, even 30 hours after death. This finding supports adopting a routine protocol to detect the pathological legacy of noise overexposure in mass stranded cetaceans as a key to understanding the complex processes and implications that lie behind such stranding events.

A microhomology-mediated break-induced replication model for the origin of human copy number variation.

Directory of Open Access Journals (Sweden)

P J Hastings

2009-01-01

Full Text Available Chromosome structural changes with nonrecurrent endpoints associated with genomic disorders offer windows into the mechanism of origin of copy number variation (CNV. A recent report of nonrecurrent duplications associated with Pelizaeus-Merzbacher disease identified three distinctive characteristics. First, the majority of events can be seen to be complex, showing discontinuous duplications mixed with deletions, inverted duplications, and triplications. Second, junctions at endpoints show microhomology of 2-5 base pairs (bp. Third, endpoints occur near pre-existing low copy repeats (LCRs. Using these observations and evidence from DNA repair in other organisms, we derive a model of microhomology-mediated break-induced replication (MMBIR for the origin of CNV and, ultimately, of LCRs. We propose that breakage of replication forks in stressed cells that are deficient in homologous recombination induces an aberrant repair process with features of break-induced replication (BIR. Under these circumstances, single-strand 3' tails from broken replication forks will anneal with microhomology on any single-stranded DNA nearby, priming low-processivity polymerization with multiple template switches generating complex rearrangements, and eventual re-establishment of processive replication.

Protection against {sup 131}I-induced Double Strand DNA Breaks in Thyroid Cells

Energy Technology Data Exchange (ETDEWEB)

Hershman, J.M.; Okunyan, A.; Cannon, S.; Hogen, V. [Endocrinology, UCLA-VA, Los Angeles (United States); Rivina, Y. [Radiation Biology, UCLA, Los Angeles (United States)

2012-07-01

Radioiodine-131 (I{sup 131}) released from nuclear reactor accidents has dramatically increased the incidence of papillary thyroid cancer in exposed individuals, especially young children. The accepted measure for prevention of radiation-induced thyroid cancer is potassium iodide tablets that contain 100 mg iodide taken daily to block thyroid uptake of I{sup 131}. The deposition of ionizing radiation in cells results in double-strand DNA breaks (DSB) at fragile sites, and this early event can generate oncogenic rearrangements that eventually cause the cancer. We have developed a thyroid cell model to quantify the mitogenic effect of I{sup 131}. I{sup 131} causes double strand DNA breaks in FRTL-5 cells detected by 53BP1 or gamma H2AX and had no effect on cells that do not transport iodide. Perchlorate, iodide, and thiocyanate protect against DSB induced by I{sup 131}. Preincubation with the anion or radioprotective compounds prevents DSB; delayed addition of the anion is much less effective. These data provide a basis for studies of radioprotection against DSB induced by I{sup 131} in animals in order to refine the prevention of thyroid cancer resulting from nuclear fallout

UVA-induced DNA double-strand breaks result from the repair of clustered oxidative DNA damages

Science.gov (United States)

Greinert, R.; Volkmer, B.; Henning, S.; Breitbart, E. W.; Greulich, K. O.; Cardoso, M. C.; Rapp, Alexander

2012-01-01

UVA (320–400 nm) represents the main spectral component of solar UV radiation, induces pre-mutagenic DNA lesions and is classified as Class I carcinogen. Recently, discussion arose whether UVA induces DNA double-strand breaks (dsbs). Only few reports link the induction of dsbs to UVA exposure and the underlying mechanisms are poorly understood. Using the Comet-assay and γH2AX as markers for dsb formation, we demonstrate the dose-dependent dsb induction by UVA in G1-synchronized human keratinocytes (HaCaT) and primary human skin fibroblasts. The number of γH2AX foci increases when a UVA dose is applied in fractions (split dose), with a 2-h recovery period between fractions. The presence of the anti-oxidant Naringin reduces dsb formation significantly. Using an FPG-modified Comet-assay as well as warm and cold repair incubation, we show that dsbs arise partially during repair of bi-stranded, oxidative, clustered DNA lesions. We also demonstrate that on stretched chromatin fibres, 8-oxo-G and abasic sites occur in clusters. This suggests a replication-independent formation of UVA-induced dsbs through clustered single-strand breaks via locally generated reactive oxygen species. Since UVA is the main component of solar UV exposure and is used for artificial UV exposure, our results shine new light on the aetiology of skin cancer. PMID:22941639

Alterations in the nuclear matrix protein mass correlate with heat-induced inhibition of DNA single-strand-break repair

International Nuclear Information System (INIS)

Warters, R.L.; Brizgys, L.M.; Lyons, B.W.

1987-01-01

The total protein mass co-isolating with the nuclear matrix or nucleoid from Chinese hamster ovary (CHO) cells was observed to increase in heated cells as a function of increasing exposure temperature between 43 0 C and 45 0 C or of exposure time at any temperature. The sedimentation distance of the CHO cell nucleoid in sucrose gradients increased with increasing exposure time at 45 0 C. Both these nuclear alterations correlated in a log-linear manner with heat-induced inhibition of DNA strand break repair. A two-fold threshold increase in nuclear matrix protein mass preceded any substantial inhibition of repair of DNA single-strand breaks. When preheated cells were incubated at 37 0 C the nuclear matrix protein mass and nucleoid sedimentation recovered with a half-time of about 5 h, while DNA single-strand-break repair recovered with a half-time of about 2 h. When preheated cells were placed at 41 0 C a further increase was observed in the nuclear matrix protein mass and the half-time of DNA strand break repair, while nucleoid sedimentation recovered toward control values. These results implicate alterations in the protein mass of the nuclear matrix in heat-induced inhibition of repair of DNA single-strand breaks. (author)

Formation of plasmid DNA strand breaks induced by low-energy ion beam: indication of nuclear stopping effects

International Nuclear Information System (INIS)

Chen Yu; Jiang Bingyao; Chen Youshan; Ding Xingzhao; Liu Xianghuai; Chen Ceshi; Guo Xinyou; Yin Guanglin

1998-01-01

Plasmid pGEM 3zf(+) was irradiated by nitrogen ion beam with energies between 20 and 100 keV and the fluence kept as 1 x 10 12 ions/cm 2 . The irradiated plasmid was assayed by neutral electrophoresis and quantified by densitometry. The yields of DNA with single-strand and double-strand breaks first increased then decreased with increasing ion energy. There was a maximal yield value in the range of 20-100 keV. The relationship between DNA double-strand breaks (DSB) cross-section and linear energy transfer (LET) also showed a peak-shaped distribution. To understand the physical process during DNA strand breaks, a Monte Carlo calculation code known as TRIM (Transport of Ions in Matter) was used to simulate energy losses due to nuclear stopping and to electronic stopping. It can be assumed that nuclear stopping plays a more important role in DNA strand breaks than electronic stopping in this energy range. The physical mechanisms of DNA strand breaks induced by a low-energy ion beam are also discussed. (orig.)

Compound Poisson Processes and Clustered Damage of Radiation Induced DNA Double Strand Breaks

International Nuclear Information System (INIS)

Gudowska-Nowak, E.; Ritter, S.; Taucher-Scholz, G.; Kraft, G.

2000-01-01

Recent experimental data have demonstrated that DNA damage induced by densely ionizing radiation in mammalian cells is distributed along the DNA molecule in the form of clusters. The principal constituent of DNA damage are double-strand breaks (DSB) which are formed when the breaks occur in both DNA strands and are directly opposite or separated by only a few base pairs. DSBs are believed to be most important lesions produced in chromosomes by radiation; interaction between DSBs can lead to cell killing, mutation or carcinogenesis. The paper discusses a model of clustered DSB formation viewed in terms of compound Poisson process along with the predictive essay of the formalism in application to experimental data. (author)

ATR Prohibits Replication Catastrophe by Preventing Global Exhaustion of RPA

DEFF Research Database (Denmark)

Toledo Lazaro, Luis Ignacio; Altmeyer, Matthias; Rask, Maj-Britt

2013-01-01

origin firing generates an excess of single-stranded DNA that exhausts the nuclear pool of RPA. Partial reduction of RPA accelerated fork breakage, and forced elevation of RPA was sufficient to delay such "replication catastrophe" even in the absence of ATR activity. Conversely, unscheduled origin firing...... induced breakage of stalled forks even in cells with active ATR. Thus, ATR-mediated suppression of dormant origins shields active forks against irreversible breakage via preventing exhaustion of nuclear RPA. This study elucidates how replicating genomes avoid destabilizing DNA damage. Because cancer cells...

Theoretical Modeling of Rock Breakage by Hydraulic and Mechanical Tool

Directory of Open Access Journals (Sweden)

Hongxiang Jiang

2014-01-01

Full Text Available Rock breakage by coupled mechanical and hydraulic action has been developed over the past several decades, but theoretical study on rock fragmentation by mechanical tool with water pressure assistance was still lacking. The theoretical model of rock breakage by mechanical tool was developed based on the rock fracture mechanics and the solution of Boussinesq’s problem, and it could explain the process of rock fragmentation as well as predicating the peak reacting force. The theoretical model of rock breakage by coupled mechanical and hydraulic action was developed according to the superposition principle of intensity factors at the crack tip, and the reacting force of mechanical tool assisted by hydraulic action could be reduced obviously if the crack with a critical length could be produced by mechanical or hydraulic impact. The experimental results indicated that the peak reacting force could be reduced about 15% assisted by medium water pressure, and quick reduction of reacting force after peak value decreased the specific energy consumption of rock fragmentation by mechanical tool. The crack formation by mechanical or hydraulic impact was the prerequisite to improvement of the ability of combined breakage.

The shape, stability and breakage of pendant liquid bridges

Science.gov (United States)

Padday, J. F.; Pétré, G.; Rusu, C. G.; Gamero, J.; Wozniak, G.

1997-12-01

Pendant liquid bridges are defined as pendant drops supporting a solid axisymmetric endplate at their lower end. The stability and shape properties of such bridges are defined in terms of the capillary properties of the system and of the mass and radius of the lower free-floating endplate. The forces acting in the pendant liquid bridge are defined exactly and expressed in dimensionless form. Numerical analysis has been used to derive the properties of a given bridge and it is shown that as the bridge grows by adding more liquid to the system a maximum volume is reached. At this maximum volume, the pendant bridge becomes unstable with the length of the bridge increasing spontaneously and irreversibly at constant volume. Finally the bridge breaks with the formation of a satellite drop or an extended thread. The bifurcation and breakage processes have been recorded using a high-speed video camera with a digital recording rate of up to 6000 frames per second. The details of the shape of the bridge bifurcation and breakage for many pendant bridge systems have been recorded and it is shown that satellite drop formation after rupture is not always viscosity dependent. Bifurcation and breakage in simulated low gravity demonstrated that breakage was very nearly symmetrical about a plane through the middle of the pendant bridge.

Molecular targets, DNA breakage, DNA repair: Their roles in mutation induction in mammalian germ cells

International Nuclear Information System (INIS)

Sega, G.A.

1989-01-01

Variability in genetic sensitivity among different germ-cell stages in the mammal to various mutagens could be the result of how much chemical reaches the different stages, what molecular targets may be affected in the different stages and whether or not repair of lesions occurs. Several chemicals have been found to bind very strongly to protamine in late-spermatid and early-spermatozoa stages in the mouse. The chemicals also produce their greatest genetic damage in these same germ-cell stages. While chemical binding to DNA has not been correlated with the level of induced genetic damage, DNA breakage in the sensitive stages has been shown to increase. This DNA breakage is believed to indirectly result from chemical binding to sulfhydryl groups in protamine which prevents normal chromatin condensation within the sperm nucleus. 22 refs., 5 figs

Breakage of cephalomedullary nailing in operative treatment of trochanteric and subtrochanteric femoral fractures.

Science.gov (United States)

von Rüden, Christian; Hungerer, Sven; Augat, Peter; Trapp, Oliver; Bühren, Volker; Hierholzer, Christian

2015-02-01

Mechanical breakage of cephalomedullary nail osteosynthesis is a rare complication attributed to delayed fracture union or nonunion. This study presents a series of cases of breakage and secondary lag screw dislocation after cephalomedullary nailing. The aim of this study was to identify factors that contribute to cephalomedullary nail breakage. In a retrospective case series review between 02/2005 and 12/2013, we analyzed 453 patients with trochanteric and subtrochanteric fracture who had been treated by cephalomedullary nailing. Fractures were classified according to AO/OTA classification. 13 patients with cephalomedullary nail breakage were included (failure rate 2.9 %). Seven patients were women, and six men with a mean age of 72 years (range 35-94). Implant breakage occurred 6 months postoperatively (range 1-19 months). In ten cases, breakage was secondary to delayed or nonunion, which was thought to be mainly due to insufficient reduction of the fracture, and in two cases due to loss of the lag screw because of missing set screw. In one case, breakage was apparent during elective metal removal following complete fracture healing. Short-term outcome was evaluated 6 months after operative revision using Harris hip score in 11 out of 13 patients showing a mean score of 84 %. Complete radiological fracture healing has been found in 11 patients available for follow-up within 6 months after revision surgery. Breakage of cephalomedullary nail osteosynthesis of trochanteric fractures is a severe complication. The results of our study demonstrate that revision surgery provides good clinical and radiological short-term results. Predominately, failures of trochanteric fractures are related to lack of surgeon performance. Therefore, application of the implant requires accurate preoperative planning, advanced surgical experience to evaluate the patient and the fracture classification, and precise surgical technique including attention to detail and anatomical

Genetics of x-ray induced double strand break repair in saccharomyces cerevisiae

International Nuclear Information System (INIS)

Budd, M.E.

1982-07-01

The possible fates of x-ray-induced double-strand breaks in Saccharomyces cerevisiae were examined. One possible pathway which breaks can follow, the repair pathway, was studied by assaying strains with mutations in the RAD51, RAD54, and RAD57 loci for double-strand break repair. In order of increasing radiation sensitivity one finds: rad57-1(23 0 )> rad51-1(30 0 )> rad54-3(36 0 ). At 36 0 , rad54-3 cells cannot repair double-strand breaks, while 23 0 , they can. Strains with the rad57-1 mutation can rejoin broken chromosomes at both temperatures. However, the low survival at 36 0 shows that the assay is not distinguishing large DNA fragments which allow cell survival from those which cause cell death. A rad51-1 strain could also rejoin broken chromosomes, and was thus capable of incomplete repair. The data can be explained with the hypothesis that rad54-3 cells are blocked in an early step of repair, while rad51-1 and rad57-1 strains are blocked in a later step of repair. The fate of double-strand breaks when they are left unrepaired was investigated with the rad54-3 mutation. If breaks are prevented from entering the RAD54 repair pathway they become uncommitted lesions. These lesions are repaired slower than the original breaks. One possible fate for an uncommitted lesion is conversion into a fixed lesion, which is likely to be an unrepairable or misrepaired double-strand break. The presence of protein synthesis after irradiation increases the probability that a break will enter the repair pathway. Evidence shows that increased probability of repair results from enhanced synthesis of repair proteins shortly after radiation

Chromatin condensation and differential sensitivity of mammalian and insect cells to DNA strand breaks induced by bleomycin

International Nuclear Information System (INIS)

Lopez-Larraza, Daniel M.; Padron, Juan; Ronci, Natalia E.; Vidal Rioja, Lidia A.

2006-01-01

Bleomycin (BLM) induces DNA damage in living cells. In this report we analyzed the role of chromatin compactness in the differential response of mosquito (ATC-15) and mammalian (CHO) cells to DNA strand breaks induced by BLM. We used cells unexposed and exposed to sodium butyrate (NaB), which induces chromatin decondensation. By nucleoid sedimentation assay and digestions of nuclei with DNAse I, untreated mosquito cells (no BLM; no NaB) were shown to have more chromatin condensation than untreated CHO cells. By alkaline unwinding ATC-15 cells treated with NaB showed more BLM-induced DNA strand breaks than NaB-untreated CHO cells. The time-course of BLM-induced DNA damage to nuclear DNA was similar for NaB-untreated mammalian and insect cells, but with mosquito cells showing less DNA strand breaks, both at physiological temperatures and at 4 o C. However, when DNA repair was inhibited by low temperatures and chromatin was decondensed by NaB treatments, differences in BLM-induced DNA damage between these cells lines were no longer observed. In both cell lines, NaB did not affect BLM action on cell growth and viability. On the other hand, the low sensitivity of ATC-15 cells to BLM was reflected in their better growth efficiency. These cells exhibited a satisfactory growth at BLM doses that produced a permanent arrest of growth in CHO cells. The data suggest that mosquito cells might have linker DNAs shorter than those of mammalian cells, which would result in the observed both greater chromatin condensation and greater resistance to DNA damage induced by BLM as compared to CHO cells

Chromatin condensation and differential sensitivity of mammalian and insect cells to DNA strand breaks induced by bleomycin

Energy Technology Data Exchange (ETDEWEB)

Lopez-Larraza, Daniel M. [IMBICE, C.C. 403, 1900 La Plata (Argentina)]. E-mail: danielop@imbice.org.ar; Padron, Juan [IMBICE, C.C. 403, 1900 La Plata (Argentina); Ronci, Natalia E. [IMBICE, C.C. 403, 1900 La Plata (Argentina); Vidal Rioja, Lidia A. [IMBICE, C.C. 403, 1900 La Plata (Argentina)

2006-08-30

Bleomycin (BLM) induces DNA damage in living cells. In this report we analyzed the role of chromatin compactness in the differential response of mosquito (ATC-15) and mammalian (CHO) cells to DNA strand breaks induced by BLM. We used cells unexposed and exposed to sodium butyrate (NaB), which induces chromatin decondensation. By nucleoid sedimentation assay and digestions of nuclei with DNAse I, untreated mosquito cells (no BLM; no NaB) were shown to have more chromatin condensation than untreated CHO cells. By alkaline unwinding ATC-15 cells treated with NaB showed more BLM-induced DNA strand breaks than NaB-untreated CHO cells. The time-course of BLM-induced DNA damage to nuclear DNA was similar for NaB-untreated mammalian and insect cells, but with mosquito cells showing less DNA strand breaks, both at physiological temperatures and at 4 {sup o}C. However, when DNA repair was inhibited by low temperatures and chromatin was decondensed by NaB treatments, differences in BLM-induced DNA damage between these cells lines were no longer observed. In both cell lines, NaB did not affect BLM action on cell growth and viability. On the other hand, the low sensitivity of ATC-15 cells to BLM was reflected in their better growth efficiency. These cells exhibited a satisfactory growth at BLM doses that produced a permanent arrest of growth in CHO cells. The data suggest that mosquito cells might have linker DNAs shorter than those of mammalian cells, which would result in the observed both greater chromatin condensation and greater resistance to DNA damage induced by BLM as compared to CHO cells.

Trapping and breaking of in vivo nicked DNA during pulsed-field gel electrophoresis

Science.gov (United States)

Khan, Sharik R.; Kuzminov, Andrei

2013-01-01

Pulsed field gel electrophoresis (PFGE) offers a high-resolution approach to quantify chromosomal fragmentation in bacteria, measured as percent of chromosomal DNA entering the gel. The degree of separation in PFG depends upon the size of DNA, as well as various conditions of electrophoresis, such as electric field strength (FS), time of electrophoresis, switch time and buffer composition. Here we describe a new parameter, the structural integrity of the sample DNA itself, that influences its migration through PFGs. We show that sub-chromosomal fragments containing both spontaneous and DNA damage-induced nicks are prone to breakage during PFGE. Such breakage at single strand interruptions results in artefactual decrease in molecular weight of linear DNA making accurate determination of the number of double strand breaks difficult. While breakage of nicked sub-chromosomal fragments is FS-independent, some high molecular weight sub-chromosomal fragments are also trapped within wells under the standard PFGE conditions. This trapping can be minimized by lowering the field strength and increasing the time of electrophoresis. We discuss how breakage of nicked DNA may be mechanistically linked to trapping. Our results suggest how to optimize conditions for PFGE when quantifying chromosomal fragmentation induced by DNA damage. PMID:23770235

Mathematical modelling of the automated FADU assay for the quantification of DNA strand breaks and their repair in human peripheral mononuclear blood cells

International Nuclear Information System (INIS)

Junk, Michael; Salzwedel, Judy; Sindlinger, Thilo; Bürkle, Alexander; Moreno-Villanueva, Maria

2014-01-01

Cells continuously undergo DNA damage from exogenous agents like irradiation or genotoxic chemicals or from endogenous radicals produced by normal cellular metabolic activities. DNA strand breaks are one of the most common genotoxic lesions and they can also arise as intermediates of DNA repair activity. Unrepaired DNA damage can lead to genomic instability, which can massively compromise the health status of organisms. Therefore it is important to measure and quantify DNA damage and its repair. We have previously published an automated method for measuring DNA strand breaks based on fluorimetric detection of alkaline DNA unwinding [1], and here we present a mathematical model of the FADU assay, which enables to an analytic expression for the relation between measured fluorescence and the number of strand breaks. Assessment of the formation and also the repair of DNA strand breaks is a crucial functional parameter to investigate genotoxicity in living cells. A reliable and convenient method to quantify DNA strand breakage is therefore of significant importance for a wide variety of scientific fields, e.g. toxicology, pharmacology, epidemiology and medical sciences

Cellular DNA breakage by soy isoflavone genistein and its methylated structural analogue biochanin A.

Science.gov (United States)

Ullah, Mohd Fahad; Shamim, Uzma; Hanif, Sarmad; Azmi, Asfar S; Hadi, Sheikh M

2009-11-01

Epidemiological studies have indicated that populations with high isoflavone intake through soy consumption have lower rates of breast, prostate, and colon cancer. The isoflavone polyphenol genistein in soybean is considered to be a potent chemopreventive agent against cancer. In order to explore the chemical basis of chemopreventive activity of genistein, in this paper we have examined the structure-activity relationship between genistein and its structural analogue biochanin A. We show that both genistein and its methylated derivative biochanin A are able to mobilize nuclear copper in human lymphocyte, leading to degradation of cellular DNA. However, the relative rate of DNA breakage was greater in the case of genistein. Further, the cellular DNA degradation was inhibited by copper chelator (neocuproine/bathocuproine) but not by compounds that specifically bind iron and zinc (desferrioxamine mesylate and histidine, respectively). We also compared the antioxidant activity of the two isoflavones against tert-butylhydroperoxide-induced oxidative breakage in lymphocytes. Again genistein was found to be more effective than biochanin A in providing protection against oxidative stress induced by tert-butylhydroperoxide. It would therefore appear that the structural features of isoflavones that are important for antioxidant properties are also the ones that contribute to their pro-oxidant action through a mechanism that involves redox cycling of chromatin-bound nuclear copper.

The influence of chromatin structure on the frequency of radiation-induced DNA strand breaks: a study using nuclear and nucleoid monolayers

International Nuclear Information System (INIS)

Ljungman, M.

1991-01-01

To assess the influence of chromatin structure on the frequency of radiation-induced DNA strand breaks, the alkaline unwinding technique was applied to nuclear and nucleoid monolayers. These chromatin substrates were prepared by treating human fibroblasts grown as monolayers with the nonionic detergent Triton X-100 and varying concentrations of cations. The chromatin structure was modified either by a stepwise removal of DNA-bound proteins by extraction in increasing concentrations of monovalent salt, or by the addition or deletion of mono- and divalent cations to condense or decondense the chromatin, respectively. It was found that the stepwise removal of DNA-bound proteins from the chromatin dramatically increased the frequency of radiation-induced DNA strand breaks. The DNA-bound proteins showed a qualitative difference in their ability to protect the DNA where proteins removed by salt concentrations above 1.0 M exerted the greatest protection. Furthermore, the frequency of radiation-induced DNA strand breaks was found to be 6 times lower in condensed chromatin than in decondensed chromatin and about 80 times lower than in protein-depleted chromatin. It is concluded that the presence of DNA-bound proteins and the folding of the chromatin into higher-order structures protect the DNA against radiation-induced strand breaks

Protective effects of pulmonary epithelial lining fluid on oxidative stress and DNA single-strand breaks caused by ultrafine carbon black, ferrous sulphate and organic extract of diesel exhaust particles

Energy Technology Data Exchange (ETDEWEB)

Chuang, Hsiao-Chi [School of Respiratory Therapy, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Division of Pulmonary Medicine, Department of Internal Medicine, Shuang Ho Hospital, Taipei Medical University, Taipei, Taiwan (China); Cheng, Yi-Ling; Lei, Yu-Chen [Institute of Occupational Medicine and Industrial Hygiene, College of Public Health, National Taiwan University, Taipei, Taiwan (China); Chang, Hui-Hsien [Institute of Environmental Health, College of Public Health, National Taiwan University, Taipei, Taiwan (China); Cheng, Tsun-Jen, E-mail: tcheng@ntu.edu.tw [Institute of Occupational Medicine and Industrial Hygiene, College of Public Health, National Taiwan University, Taipei, Taiwan (China); Department of Public Health, College of Public Health, National Taiwan University, Taipei, Taiwan (China)

2013-02-01

Pulmonary epithelial lining fluid (ELF) is the first substance to make contact with inhaled particulate matter (PM) and interacts chemically with PM components. The objective of this study was to determine the role of ELF in oxidative stress, DNA damage and the production of proinflammatory cytokines following physicochemical exposure to PM. Ultrafine carbon black (ufCB, 15 nm; a model carbonaceous core), ferrous sulphate (FeSO{sub 4}; a model transition metal) and a diesel exhaust particle (DEP) extract (a model organic compound) were used to examine the acellular oxidative potential of synthetic ELF and non-ELF systems. We compared the effects of exposure to ufCB, FeSO{sub 4} and DEP extract on human alveolar epithelial Type II (A549) cells to determine the levels of oxidative stress, DNA single-strand breaks and interleukin-8 (IL-8) production in ELF and non-ELF systems. The effects of ufCB and FeSO{sub 4} on the acellular oxidative potential, cellular oxidative stress and DNA single-strand breakage were mitigated significantly by the addition of ELF, whereas there was no decrease following treatment with the DEP extract. There was no significant effect on IL-8 production following exposure to samples that were suspended in ELF/non-ELF systems. The results of the present study indicate that ELF plays an important role in the initial defence against PM in the pulmonary environment. Experimental components, such as ufCB and FeSO{sub 4}, induced the production of oxidative stress and led to DNA single-strand breaks, which were moderately prevented by the addition of ELF. These findings suggest that ELF plays a protective role against PM-driven oxidative stress and DNA damage. -- Highlights: ► To determine the role of ELF in ROS, DNA damage and IL-8 after exposure to PM. ► ufCB, FeSO{sub 4} and DEP extract were used to examine the protective effects of ELF. ► PM-driven oxidative stress and DNA single-strand breakage were mitigated by ELF. ► The findings

Determination and analysis of site-specific 125I decay-induced DNA double-strand break end-group structures.

Science.gov (United States)

Datta, Kamal; Weinfeld, Michael; Neumann, Ronald D; Winters, Thomas A

2007-02-01

End groups contribute to the structural complexity of radiation-induced DNA double-strand breaks (DSBs). As such, end-group structures may affect a cell's ability to repair DSBs. The 3'-end groups of strand breaks caused by gamma radiation, or oxidative processes, under oxygenated aqueous conditions have been shown to be distributed primarily between 3'-phosphoglycolate and 3'-phosphate, with 5'-phosphate ends in both cases. In this study, end groups of the high-LET-like DSBs caused by 125I decay were investigated. Site-specific DNA double-strand breaks were produced in plasmid pTC27 in the presence or absence of 2 M DMSO by 125I-labeled triplex-forming oligonucleotide targeting. End-group structure was assessed enzymatically as a function of the DSB end to serve as a substrate for ligation and various forms of end labeling. Using this approach, we have demonstrated 3'-hydroxyl (3'-OH) and 3'-phosphate (3'-P) end groups and 5'-ends (> or = 42%) terminated by phosphate. A 32P postlabeling assay failed to detect 3'-phosphoglycolate in a restriction fragment terminated by the 125I-induced DNA double-strand break, and this is likely due to restricted oxygen diffusion during irradiation as a frozen aqueous solution. Even so, end-group structure and relative distribution varied as a function of the free radical scavenging capacity of the irradiation buffer.

Nijmegen breakage syndrome (NBS

Directory of Open Access Journals (Sweden)

Chrzanowska Krystyna H

2012-02-01

Full Text Available Abstract Nijmegen breakage syndrome (NBS is a rare autosomal recessive syndrome of chromosomal instability mainly characterized by microcephaly at birth, combined immunodeficiency and predisposition to malignancies. Due to a founder mutation in the underlying NBN gene (c.657_661del5 the disease is encountered most frequently among Slavic populations. The principal clinical manifestations of the syndrome are: microcephaly, present at birth and progressive with age, dysmorphic facial features, mild growth retardation, mild-to-moderate intellectual disability, and, in females, hypergonadotropic hypogonadism. Combined cellular and humoral immunodeficiency with recurrent sinopulmonary infections, a strong predisposition to develop malignancies (predominantly of lymphoid origin and radiosensitivity are other integral manifestations of the syndrome. The NBN gene codes for nibrin which, as part of a DNA repair complex, plays a critical nuclear role wherever double-stranded DNA ends occur, either physiologically or as a result of mutagenic exposure. Laboratory findings include: (1 spontaneous chromosomal breakage in peripheral T lymphocytes with rearrangements preferentially involving chromosomes 7 and 14, (2 sensitivity to ionizing radiation or radiomimetics as demonstrated in vitro by cytogenetic methods or by colony survival assay, (3 radioresistant DNA synthesis, (4 biallelic hypomorphic mutations in the NBN gene, and (5 absence of full-length nibrin protein. Microcephaly and immunodeficiency are common to DNA ligase IV deficiency (LIG4 syndrome and severe combined immunodeficiency with microcephaly, growth retardation, and sensitivity to ionizing radiation due to NHEJ1 deficiency (NHEJ1 syndrome. In fact, NBS was most commonly confused with Fanconi anaemia and LIG4 syndrome. Genetic counselling should inform parents of an affected child of the 25% risk for further children to be affected. Prenatal molecular genetic diagnosis is possible if disease

Impact of polymeric membrane breakage on drinking water quality and an online detection method of the breakage.

Science.gov (United States)

Wu, Qilong; Zhang, Zhenghua; Cao, Guodong; Zhang, Xihui

2017-10-15

Polymeric membrane has been widely used for the treatment of drinking water in China, and the total treating capacity has reached up to 3.8 million m 3 /d. However, the membrane breakage found in the membrane modules in many water treatment plants resulted in an increase in turbidity and bacterial amount in the membrane permeate. In this study, a membrane module running for 3 years in a full-scale application was examined in terms of the breaking positions and the numbers of the broken fibers. It was found that most of the breaking positions were mainly on the outlet side of the module and that the distance from these points to the outlet was about 1/10-2/10 length of the membrane module. The lab-scale tests showed that the increase of the numbers of the breaking fibers in the membrane module (the breaking fibers were from 1 to 4 of 75 fibers) resulted in the increase in turbidity, particle count and the amount of total bacteria and coliform bacteria. Meanwhile, the water quality after the filtration with broken membrane fibers was similar to the quality of the raw water, which indicated that once the membrane fiber breakage occurred in the membrane module, the quality of drinking water after membrane filtration was significantly affected. Furthermore, the breaking position closer to the outlet side of the membrane module exposed much higher microbiological risk than those in the middle or near the bottom side. A pilot scale test was conducted by using a membrane module with 6600 fibers, and the effect of the membrane breakage (1-4 broken fibers) on water quality was also investigated. The results indicated that periodical backwashing caused drastic fluctuation of turbidity, particle count and the bacterial amount in the permeate water, which might be due to the washing force and self-blocking action inside the hollow fibers. Moreover, there is a good quantitative relationship (R 2 = 0.945) between particle count and the bacterial amount, which indicated that an

The radiation hypersensitivity of cells at mitosis.

Science.gov (United States)

Stobbe, C C; Park, S J; Chapman, J D

2002-12-01

Mitotic cells are hypersensitive to ionizing radiation, exhibiting single-hit inactivation coefficients near to those of repair deficient cell lines and lymphocytes. To elucidate possible mechanisms for this hypersensitivity, the kinetics of oxygen radiosensitization, the proportion of indirect effect by OH radicals and the kinetics of radiation-induced DNA strand breakage in the chromatin of mitotic cells were investigated. Synchronized populations of >90% mitotic HT-29 cells were obtained by the mitotic shake-off method. Cells were irradiated at indirect effect of OH radicals was investigated with the radical scavenger, DMSO. DNA strand breakage was measured by the comet assay. Mitotic HT-29 cell inactivation is well described by a single-hit inactivation coefficient (alpha) of 1.14 +/- 0.06 Gy(-1). The oxygen enhancement ratio of mitotic cells (at 10% survival) was found to be approximately 2.0, significantly lower than the value of 2.8 measured for interphase (asynchronous) cells. More than 60% of mitotic cell killing was eliminated when the media contained 2 M DMSO, indicating that indirect effect is as important in the killing of mitotic cells as it is for interphase cells. The chromatin in mitotic cells was found to be ~2.8 times more sensitive to radiation-induced DNA single-strand breakage than the chromatin of interphase cells. The alpha-inactivation coefficient of mitotic HT-29 cells was ~30 times larger than that of interphase cells. Mitotic cell chromatin appears to contain intrinsic DNA breaks that are not lethal. In addition, chromatin in mitotic cells was found to be more susceptible to radiation-induced DNA strand-breakage than the dispersed chromatin of interphase cells. How the enhanced production of these simple DNA lesions (that are usually reparable) translates into the lethal (non-reparable) events associated with alpha-inactivation is not known. The compaction/dispersion status of DNA throughout the cell cycle appears to be an important

Radiation sensitivity of organisms of different organization level: an approach including DNA strand breakage

International Nuclear Information System (INIS)

Kampf, G.

1983-01-01

The mean numbers of DNA double-strand breaks (DSB) suggested to be necessary to lead to the loss of reproductive capacity are compared with bacteriophages, bacteria, and cells of the Chinese hamster after the influence of several radiation qualities. The results suggest that the critical target for the inactivating action of radiations may not be the entire DNA of all organisms but a structure unit of it designed as membrane-attached super structure unit. With organisms having only one of these structures (bacteria) the inactivation probability of one DSB will be near unity, with their multiplication in higher cells it will become lower. This means, eukaryotic cells are able to tolerate more DSB before being inactivated than organisms of a lower organization level, and consequently are more ''lesion resistant''. This behavior represents an evolutionary stabilization of higher cells towards the lethal action of severe DNA lesions such as DSB. (author)

Benzoyl peroxide-induced damage to DNA and its components

DEFF Research Database (Denmark)

Hazlewood, C; Davies, Michael Jonathan

1996-01-01

of base adducts, though the exact identity of the species detected in these cases could not be determined due to the complexity of the spectra. Hydrogen abstraction at the sugar-phosphate backbone is also believed to occur with these substrates as strand breakage is observed; the extent of the latter...

Induction of double-strand breaks in DNA of prokaryotes and eukaryotes and their repair. 1. Application of elastoviscosimetry for studying double-strand breaks in DNA of Escherichia coli induced by. gamma. -irradiation

Energy Technology Data Exchange (ETDEWEB)

Bresler, S E; Noskin, L A; Suslov, A V [AN SSSR, Leningrad. Inst. Yadernoj Fiziki

1980-11-01

It is shown that the method of elastoviscosimetry gives a possibility to record the formation of DNA double-strand breaks in Escherichia coli cells induced by ..gamma.. irradiation at doses close to D/sub 37/. The dependence of changes of elastoviscosity parameter on the dose (tau/sub 0/) passes through the maximum. It is shown that the ascending section of this curve (at minimum ..gamma.. irradiation doses) characterizes the relaxation process of the superspiralised chromosome in nucleotide of the E. coli. This relaxation is observed due to ..gamma.. induced damages which are not double-strand breaks. By the maximum position one can judge on a dose yield of the first DNA double-strand break, the descending part of the dose curve describes the kinetics of accumulation of breaks with the dose increase. The analysis of the data obtained gives the possibility to come to the conclusion that when applying a usual technique of irradiation and lysis of cells not providing for special measures on inhibition of endo-and exonuclease activity in ..gamma.. irradiated cells, the dose yield of double-strand breaks noticeably increases (by 4.2 times). In the case of an essential, though incomplete, inhibition of nuclease activities in ..gamma.. irradiated cells the dose yield of breaks approximately corresponds to the dose curve of inactivation of these cells (D/sub 37/12.5+-3.0 krad, the first double-strand break -at 14.5+-2.4 krad).

Induction of double-strand breaks in DNA of prokaryotes and eukaryotes and their repair. 1. Application of elastoviscosimetry for studying double-strand breaks in DNA of Escherichia coli induced by γ-irradiation

International Nuclear Information System (INIS)

Bresler, S.E.; Noskin, L.A.; Suslov, A.V.

1980-01-01

It is shown that the method of elastoviscosimetry gives a possibility to record the formation of DNA double-strand breaks in Escherichia coli cells induced by γ irradiation at doses close to D 37 . The dependence of changes of elastoviscosity parameter on the dose (tau 0 ) passes through the maximum. It is shown that the ascending section of this curve (at minimum γ irradiation doses) characterizes the relaxation process of the superspiralised chromosome in nucleotide of the E. coli. This relaxation is observed due to γ induced damages which are not double-strand breaks. By the maximum position one can judge on a dose yield of the first DNA double-strand break, the descending part of the dose curve describes the kinetics of accumulation of breaks with the dose increase. The analysis of the data obtained gives the possibility to come to the conclusion that when applying a usual technique of irradiation and lysis of cells not providing for special measures on inhibition of endo-and exonuclease activity in γ irradiated cells, the dose yield of double-strand breaks noticeably increases (by 4.2 times). In the case of an essential, though incomplete, inhibition of nuclease activities in γ irradiated cells the dose yield of breaks approximately corresponds to the dose curve of inactivation of these cells (D 37 12.5+-3.0 krad, the first double-strand break -at 14.5+-2.4 krad)

The risk of capsular breakage from phacoemulsification needle contact with the lens capsule: a laboratory study.

Science.gov (United States)

Meyer, Jay J; Kuo, Annie F; Olson, Randall J

2010-06-01

To determine capsular breakage risk from contact by phacoemulsification needles by machine and tip type. Experimental laboratory investigation. Infiniti (Alcon, Inc.) with Intrepid cartridges and Signature (Abbott Medical Optics, Inc.) phacoemulsification machines were tested using 19- and 20-gauge sharp and rounded tips. Actual and unoccluded flow vacuum were determined at 550 mm Hg, bottle height of 75 cm, and machine-indicated flow rate of 60 mL/minute. Breakage from brief tip contact with a capsular surrogate and human cadaveric lenses was calculated. Nineteen-gauge tips had more flow and less unoccluded flow vacuum than 20-gauge tips for both machines, with highest unoccluded flow vacuum in the Infiniti. The 19-gauge sharp tip was more likely than the 20-gauge sharp tip to cause surrogate breakage for Signature with micropulse and Ellips (Abbott Medical Optics, Inc.) ultrasound at 100% power. For Infiniti using OZil (Alcon, Inc.) ultrasound, 20-gauge sharp tips were more likely than 19-gauge sharp tips to break the membrane. For cadaveric lenses, using rounded 20-gauge tips at 100% power, breakage rates were micropulse (2.3%), Ellips (2.3%), OZil (5.3%). Breakage rates for sharp 20-gauge Ellips tips were higher than for rounded tips. Factors influencing capsular breakage may include active vacuum at the tip, flow rate, needle gauge, and sharpness. Nineteen-gauge sharp tips were more likely than 20-gauge tips to cause breakage in lower vacuum methods. For higher-vacuum methods, breakage is more likely with 20-gauge than with 19-gauge tips. Rounded-edge tips are less likely than sharp-edged tips to cause breakage. Copyright 2010 Elsevier Inc. All rights reserved.

Cranial MRI in the Nijmegen breakage syndrome

International Nuclear Information System (INIS)

Bekiesinska-Figatowska, M.; Chrzanowska, K.H.; Krajewska-Walasek, M.; Sikorska, J.; Walecki, J.; Jozwiak, S.; Kleijer, W.J.

2000-01-01

We present the results of MRI examinations in ten patients with documented Nijmegen breakage syndrome (NBS), aged 1.75-19 years. T1-, Proton-Density- and T2-weighted spin-echo sequences were performed in three planes. All patients showed microcephaly with decreased size of the frontal lobes and narrow frontal horns. In four patients agenesis of the posterior part of the corpus callosum was found, with colpocephaly and temporal horns dilatation. In one patient callosal hypoplasia was accompanied by abnormal cerebrospinal fluid spaces and wide cerebral cortex, suspicious of pachygyria. Sinusitis was present in all ten patients, as a result of primary immunodeficiency. As in ataxia teleangiectasia and other breakage syndromes, patients with NBS show an inherited susceptibility to malignancy and hypersensitivity to X- and γ-radiation. CT is therefore contraindicated in these patients and MRI should be the method of choice for diagnostic imaging. (orig.)

Medium from X-rayed cultures induces DNA strand-breaks in non-irradiated HeLa cells

International Nuclear Information System (INIS)

Ikushima, T.; Okuyama, K.; Tanizaki, Y.

2002-01-01

There is growing evidence to indicate that several types of responses are induced by ionizing radiation in non-irradiated cells. Such bystander effects include the killing of non-irradiated cells, the induction of sister chromatid exchanges and chromosomal aberrations, and the induction of gene mutations and chromosomal instability and enhanced cell growth. In the present study, we assessed whether the medium from irradiated cultures can induce DNA strand-breaks in non-irradiated cells, using single-cell gel electrophoresis assay (comet assay). HeLa cells in culture were irradiated with 0.5 to 8 Gy of 140 kVp X-rays and one hour later, the medium was taken from the irradiated culture, passed through a filter and transferred to the parallel culture of non-irradiated HeLa cells as non-target cells. After incubation for 30 min, the comet assay was performed under alkaline and neutral conditions. Such treatments resulted in a dose-dependent increase in tail moment under either alkaline or neutral condition, indicating the induction of DNA single- or double-strand breaks, respectively. It was also shown that the clonogenic survival was reduced in the cells cultured in the medium from irradiated cultures. Such a change was not detected at all when medium alone was irradiated. These results provided disputed evidence that irradiated cells released certain genotoxic factor(s) into the culture medium that can induce DNA strand breaks leading to cell death. Our results suggest that physical contact between irradiated and non-irradiated cells may not be necessary for the bystander effects observed in this study. It appears that bystander responses may be mediated by multiple mechanisms

Preferential repair of ionizing radiation-induced damage in the transcribed strand of an active human gene is defective in Cockayne syndrome

International Nuclear Information System (INIS)

Leadon, S.A.; Copper, P.K.

1993-01-01

Cells from patients with Cockayne syndrome (CS), which are sensitive to killing by UV although overall damage removal appears normal, are specifically defective in repair of UV damage in actively transcribe genes. Because several CS strains display cross-sensitivity to killing by ionizing radiation, the authors examined whether ionizing radiation-induced damage in active genes is preferentially repaired by normal cells and whether the radiosensitivity of CS cells can be explained by a defect in this process. They found that ionizing radiation-induced damage was repaired more rapidly in the transcriptionally active metallothionein IIA (MTIIA) gene than in the inactive MTIIB gene or in the genome overall in normal cells as a result of faster repair on the transcribed strand of MTIIA. Cells of the radiosensitive CS strain CS1AN are completely defective in this strand-selective repair of ionizing radiation-induced damage, although their overall repair rate appears normal. CS3BE cells, which are intermediate in radiosensitivity, do exhibit more rapid repair of the transcribed strand but at a reduced rate compared to normal cells. Xeroderma pigmentosum complementation group A cells, which are hypersensitive to UV light because of a defect in the nucleotide excision repair pathway but do not show increased sensitivity to ionizing radiation, preferentially repair ionizing radiation-induced damage on the transcribed strand of MTIIA. Thus, the ability to rapidly repair ionizing radiation-induced damage in actively transcribing genes correlates with cell survival. The results extend the generality of preferential repair in active genes to include damage other than bulky lesions

Breakage of Agglomerates : Attrition, Abrasion and Compression

NARCIS (Netherlands)

Van Laarhoven, B.

2010-01-01

In many industries particulate solids are handled in different forms. When producing particles breakage is an important wanted, in the case of grinding, or unwanted phenomenon. Granules often consist of more than one component and multiple phases. This means that granules are strongly anisotropic

Strand breaks and lethal damage in plasmid DNA subjected to 60CO-γirradiation

International Nuclear Information System (INIS)

Klimczak, U.

1992-01-01

Experiments with calf thymus DNA subjected to extracellular irradiation yield information on the role of direct and indirect effects in single-strand breakage, if this is evaluated with reference to the scavenger activity in respect of OH radicals. The role of the two processes in the occurrence of double-stand breaks and further damage leading to cell decay has so far remained largely obscure. It was the aim of the study described here to contribute to research in this field by performing in vitro experiments on biologically active DNA. For this purpose, DNA from pBR322 plasmids was irradiated in the presence of OH-radical scavengers. The number of single-strand and double-strand breaks was determined on the basis of the system's ability to eliminate OH radicals. In order to asses the influence of irradiation processes on the biological activity of DNA, investigations were carried out in E. coli for transformations caused by irradiated plasmid DNA. The results were interpreted in the light of theories about inhomogenous reaction kinetics put forward by Mark et al. (1989). It was finally discussed, which of the gamma-irradiation injuries occurring in DNA was to be held responsible for the inactivation of plasmid DNA and which enzymatic processes were additionally at work here. (orig./MG) [de

Torsional regulation of hRPA-induced unwinding of double-stranded DNA

NARCIS (Netherlands)

De Vlaminck, I.; Vidic, I.; Van Loenhout, M.T.J.; Kanaar, R.; Lebbink, J.H.G.; Dekker, C.

2010-01-01

All cellular single-stranded (ss) DNA is rapidly bound and stabilized by single stranded DNA-binding proteins (SSBs). Replication protein A, the main eukaryotic SSB, is able to unwind double-stranded (ds) DNA by binding and stabilizing transiently forming bubbles of ssDNA. Here, we study the

The role of nibrin in doxorubicin-induced apoptosis and cell senescence in Nijmegen Breakage Syndrome patients lymphocytes.

Directory of Open Access Journals (Sweden)

Olga Alster

Full Text Available Nibrin plays an important role in the DNA damage response (DDR and DNA repair. DDR is a crucial signaling pathway in apoptosis and senescence. To verify whether truncated nibrin (p70, causing Nijmegen Breakage Syndrome (NBS, is involved in DDR and cell fate upon DNA damage, we used two (S4 and S3R spontaneously immortalized T cell lines from NBS patients, with the founding mutation and a control cell line (L5. S4 and S3R cells have the same level of p70 nibrin, however p70 from S4 cells was able to form more complexes with ATM and BRCA1. Doxorubicin-induced DDR followed by cell senescence could only be observed in L5 and S4 cells, but not in the S3R ones. Furthermore the S3R cells only underwent cell death, but not senescence after doxorubicin treatment. In contrary to doxorubicin treatment, cells from all three cell lines were able to activate the DDR pathway after being exposed to γ-radiation. Downregulation of nibrin in normal human vascular smooth muscle cells (VSMCs did not prevent the activation of DDR and induction of senescence. Our results indicate that a substantially reduced level of nibrin or its truncated p70 form is sufficient to induce DNA-damage dependent senescence in VSMCs and S4 cells, respectively. In doxorubicin-treated S3R cells DDR activation was severely impaired, thus preventing the induction of senescence.

Destabilized SMC5/6 complex leads to chromosome breakage syndrome with severe lung disease

NARCIS (Netherlands)

van der Crabben, Saskia N; Hennus, Marije P; McGregor, Grant A; Ritter, Deborah I; Nagamani, Sandesh C S; Wells, Owen S; Harakalova, Magdalena; Chinn, Ivan K; Alt, Aaron; Vondrova, Lucie; Hochstenbach, Ron; van Montfrans, Joris M; Terheggen-Lagro, Suzanne W; van Lieshout, Stef; van Roosmalen, Markus J; Renkens, Ivo; Duran, Karen; Nijman, Isaäc J.; Kloosterman, Wigard P; Hennekam, Eric; Orange, Jordan S; van Hasselt, Peter M; Wheeler, David A; Palecek, Jan J; Lehmann, Alan R; Oliver, Antony W; Pearl, Laurence H; Plon, Sharon E; Murray, Johanne M; van Haaften, Gijs

The structural maintenance of chromosomes (SMC) family of proteins supports mitotic proliferation, meiosis, and DNA repair to control genomic stability. Impairments in chromosome maintenance are linked to rare chromosome breakage disorders. Here, we have identified a chromosome breakage syndrome

Carbon ion induced DNA double-strand breaks in melanophore B16

International Nuclear Information System (INIS)

Wei Zengquan; Zhou Guangming; Wang Jufang; He Jing; Li Qiang; Li Wenjian; Xie Hongmei; Cai Xichen; Tao Huang; Dang Bingrong; Han Guangwu

1997-01-01

DNA double-strand breaks (DSBs) in melanophore B 16 induced by plateau and extended Bragg peak of 75 MeV/u 12 C 6+ ions were studied by using a technique of inverse pulsed-field gel electrophoresis (PIGE). DNA fragment lengths were distributed in two ranges: the larger in 1.4 Mbp-3.2 Mbp and the smaller in less than 1.2 Mbp. It indicates that distribution of DNA fragments induced by heavy ion irradiation is not stochastic and there probably are sensitive sites to heavy ions in DNA molecules of B 16 . Percentage of DNA released from plug (PR) increased and trended towards a quasi-plateau ∝85% as dose increased. Content of the larger fragments decreased and flattened with increasing dose while content of the smaller ones increased and trended towards saturation. (orig.)

Grain breakage under uniaxial compression, through 3D DEM modelling

Directory of Open Access Journals (Sweden)

Nader François

2017-01-01

Full Text Available A breakable grain model is presented, using the concept of particles assembly. Grains of polyhedral shapes are generated, formed by joining together tetrahedral subgrains using cohesive bonds. Single grain crushing simulations are performed for multiple values of the intra-granular cohesion to study the effect on the grain’s strength. The same effect of intra-granular cohesion is studied under oedometric compression on samples of around 800 grains, which allows the evaluation of grain breakage model on the macroscopic behaviour. Grain size distribution curves and grain breakage ratios are monitored throughout the simulations.

RPA Stabilization of Single-Stranded DNA Is Critical for Break-Induced Replication.

Science.gov (United States)

Ruff, Patrick; Donnianni, Roberto A; Glancy, Eleanor; Oh, Julyun; Symington, Lorraine S

2016-12-20

DNA double-strand breaks (DSBs) are cytotoxic lesions that must be accurately repaired to maintain genome stability. Replication protein A (RPA) plays an important role in homology-dependent repair of DSBs by protecting the single-stranded DNA (ssDNA) intermediates formed by end resection and by facilitating Rad51 loading. We found that hypomorphic mutants of RFA1 that support intra-chromosomal homologous recombination are profoundly defective for repair processes involving long tracts of DNA synthesis, in particular break-induced replication (BIR). The BIR defects of the rfa1 mutants could be partially suppressed by eliminating the Sgs1-Dna2 resection pathway, suggesting that Dna2 nuclease attacks the ssDNA formed during end resection when not fully protected by RPA. Overexpression of Rad51 was also found to suppress the rfa1 BIR defects. We suggest that Rad51 binding to the ssDNA formed by excessive end resection and during D-loop migration can partially compensate for dysfunctional RPA. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

Development of yarn breakage detection software system based on machine vision

Science.gov (United States)

Wang, Wenyuan; Zhou, Ping; Lin, Xiangyu

2017-10-01

For questions spinning mills and yarn breakage cannot be detected in a timely manner, and save the cost of textile enterprises. This paper presents a software system based on computer vision for real-time detection of yarn breakage. The system and Windows8.1 system Tablet PC, cloud server to complete the yarn breakage detection and management. Running on the Tablet PC software system is designed to collect yarn and location information for analysis and processing. And will be processed after the information through the Wi-Fi and http protocol sent to the cloud server to store in the Microsoft SQL2008 database. In order to follow up on the yarn break information query and management. Finally sent to the local display on time display, and remind the operator to deal with broken yarn. The experimental results show that the system of missed test rate not more than 5%o, and no error detection.

Inhibition of peroxynitrite-mediated DNA strand cleavage and hydroxyl radical formation by aspirin at pharmacologically relevant concentrations: Implications for cancer intervention

Energy Technology Data Exchange (ETDEWEB)

Chen, Wei [Division of Biomedical Sciences, Edward Via Virginia College of Osteopathic Medicine, Virginia Tech Corporate Research Center, Blacksburg, VA 24060 (United States); College of Food Science and Biotechnology, Zhejiang Gongshang University, Hangzhou 310035 (China); Department of Food Science and Technology, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061 (United States); Zhu, Hong; Jia, Zhenquan [Division of Biomedical Sciences, Edward Via Virginia College of Osteopathic Medicine, Virginia Tech Corporate Research Center, Blacksburg, VA 24060 (United States); Li, Jianrong [College of Food Science and Biotechnology, Zhejiang Gongshang University, Hangzhou 310035 (China); Misra, Hara P. [Division of Biomedical Sciences, Edward Via Virginia College of Osteopathic Medicine, Virginia Tech Corporate Research Center, Blacksburg, VA 24060 (United States); Zhou, Kequan, E-mail: kzhou@wayne.edu [Department of Nutrition and Food Science, Wayne State University, Detroit, MI 48202 (United States); Li, Yunbo, E-mail: yli@vcom.vt.edu [Division of Biomedical Sciences, Edward Via Virginia College of Osteopathic Medicine, Virginia Tech Corporate Research Center, Blacksburg, VA 24060 (United States)

2009-12-04

Epidemiological studies have suggested that the long-term use of aspirin is associated with a decreased incidence of human malignancies, especially colorectal cancer. Since accumulating evidence indicates that peroxynitrite is critically involved in multistage carcinogenesis, this study was undertaken to investigate the ability of aspirin to inhibit peroxynitrite-mediated DNA damage. Peroxynitrite and its generator 3-morpholinosydnonimine (SIN-1) were used to cause DNA strand breaks in {phi}X-174 plasmid DNA. We demonstrated that the presence of aspirin at concentrations (0.25-2 mM) compatible with amounts in plasma during chronic anti-inflammatory therapy resulted in a significant inhibition of DNA cleavage induced by both peroxynitrite and SIN-1. Moreover, the consumption of oxygen caused by 250 {mu}M SIN-1 was found to be decreased in the presence of aspirin, indicating that aspirin might affect the auto-oxidation of SIN-1. Furthermore, EPR spectroscopy using 5,5-dimethylpyrroline-N-oxide (DMPO) as a spin trap demonstrated the formation of DMPO-hydroxyl radical adduct (DMPO-OH) from authentic peroxynitrite, and that aspirin at 0.25-2 mM potently diminished the radical adduct formation in a concentration-dependent manner. Taken together, these results demonstrate for the first time that aspirin at pharmacologically relevant concentrations can inhibit peroxynitrite-mediated DNA strand breakage and hydroxyl radical formation. These results may have implications for cancer intervention by aspirin.

Inhibition of peroxynitrite-mediated DNA strand cleavage and hydroxyl radical formation by aspirin at pharmacologically relevant concentrations: Implications for cancer intervention

International Nuclear Information System (INIS)

Chen, Wei; Zhu, Hong; Jia, Zhenquan; Li, Jianrong; Misra, Hara P.; Zhou, Kequan; Li, Yunbo

2009-01-01

Epidemiological studies have suggested that the long-term use of aspirin is associated with a decreased incidence of human malignancies, especially colorectal cancer. Since accumulating evidence indicates that peroxynitrite is critically involved in multistage carcinogenesis, this study was undertaken to investigate the ability of aspirin to inhibit peroxynitrite-mediated DNA damage. Peroxynitrite and its generator 3-morpholinosydnonimine (SIN-1) were used to cause DNA strand breaks in φX-174 plasmid DNA. We demonstrated that the presence of aspirin at concentrations (0.25-2 mM) compatible with amounts in plasma during chronic anti-inflammatory therapy resulted in a significant inhibition of DNA cleavage induced by both peroxynitrite and SIN-1. Moreover, the consumption of oxygen caused by 250 μM SIN-1 was found to be decreased in the presence of aspirin, indicating that aspirin might affect the auto-oxidation of SIN-1. Furthermore, EPR spectroscopy using 5,5-dimethylpyrroline-N-oxide (DMPO) as a spin trap demonstrated the formation of DMPO-hydroxyl radical adduct (DMPO-OH) from authentic peroxynitrite, and that aspirin at 0.25-2 mM potently diminished the radical adduct formation in a concentration-dependent manner. Taken together, these results demonstrate for the first time that aspirin at pharmacologically relevant concentrations can inhibit peroxynitrite-mediated DNA strand breakage and hydroxyl radical formation. These results may have implications for cancer intervention by aspirin.

Perspectives on condom breakage: a qualitative study of female sex workers in Bangalore, India.

Science.gov (United States)

Gurav, Kaveri; Bradley, Janet; Chandrashekhar Gowda, G; Alary, Michel

2014-01-01

A qualitative study was conducted to obtain a detailed understanding of two key determinants of condom breakage - 'rough sex' and poor condom fit - identified in a recent telephone survey of female sex workers, in Bangalore, India. Transcripts from six focus-group discussions involving 35 female sex workers who reported condom breakage during the telephone survey were analysed. Rough sex in different forms, from over-exuberance to violence, was often described by sex workers as a result of clients' inebriation and use of sexual stimulants, which, they report, cause tumescence, excessive thrusting and sex that lasts longer than usual, thereby increasing the risk of condom breakage. Condom breakage in this setting is the result of a complex set of social situations involving client behaviours and power dynamics that has the potential to put the health and personal lives of sex workers at risk. These findings and their implications for programme development are discussed.

Single and double strand breaks induced by 3H incorporated in DNA of cultured human kidney cells

International Nuclear Information System (INIS)

Tisljar-Lentulis, G.; Henneberg, P.; Mielke, T.; Feinendegen, L.E.

1978-01-01

In the course of the investigations of the biological effects of radionuclides incorporated in DNA single (SSB) and double strand breaks (DSB) caused tritium-decay were measured and compared with respective data resulting from 125 I. Tritium bound to thymidine and iododeoxyuridine seems to be more effective than tritium bound to other DNA-precursors. On the basis of decay, methyl- 3 H thymidine appears to be more effective with regard to the production of strand breaks than 3 H in position 6 of the pyrimidine ring. Based on the numbers of strand-breaks per rad, position 6 is more effective in accordance with data obtained by F. Krasin et al. The ratio of SSBs to DSBs per tritium decay appears to be approximately 8 in mammlian cells. Not only SSBs but also DSBs induced by 3 H in mammalian cells are reapairable. (orig./AJ) [de

Repair of single-strand breaks induced in the DNA of Proteus mirabilis by excision repair after UV-irradiation

International Nuclear Information System (INIS)

Stoerl, K.; Mund, C.

1977-01-01

Single-strand breaks have been produced in the DNA of P. mirabilis after UV-irradiation in dependence on the incident UV-doses. It has been found that there exists a discrepancy between the single-strand breaks estimated from sedimentation in alkaline sucrose gradients and the expected single-strand breaks approximated from measurements of dimer excision. The low number in incision breaks observed by sedimentation experiments is an indication that the cells are able to repair the excision-induced breaks as fast as they are formed. Toluenized cells have been used for investigation of the incision step independently of subsequent repair processes. In presence of NMN the appearance of more single-strand breaks in the DNA has been observed. Furthermore, the number of incision breaks in toluenized cells increased in presence of exogenous ATP. The completion of the excision repair process has been investigated by observing the rejoining of incision breaks. After irradiation with UV-doses higher than approximately 240 erg/mm 2 the number of single-strand breaks remaining unrepaired in the DNA increased. Studies of the influence of nutrition conditions on the repair process have shown approximately the same capacity for repair of single-strand breaks in growth medium as well as in buffer. Progress in the excision repair was also followed by investigation of the DNA synthesized at the template-DNA containing the pyrimidine dimers. In comparison with E. coli, P. mirabilis showed a somewhat lower efficiency for the repair of single-strand breaks during the excision repair. (author)

Force deficits and breakage rates after single lengthening contractions of single fast fibers from unconditioned and conditioned muscles of young and old rats.

Science.gov (United States)

Lynch, Gordon S; Faulkner, John A; Brooks, Susan V

2008-07-01

The deficit in force generation is a measure of the magnitude of damage to sarcomeres caused by lengthening contractions of either single fibers or whole muscles. In addition, permeabilized single fibers may suffer breakages. Our goal was to understand the interaction between breakages and force deficits in "young" and "old" permeabilized single fibers from control muscles of young and old rats and "conditioned" fibers from muscles that completed a 6-wk program of in vivo lengthening contractions. Following single lengthening contractions of old-control fibers compared with young-control fibers, the twofold greater force deficits at a 10% strain support the concept of an age-related increase in the susceptibility of fibers to mechanical damage. In addition, the much higher breakage rates for old fibers at all strains tested indicate an increase with aging in the number of fibers at risk of being severely injured during any given stretch. Following the 6-wk program of lengthening contractions, young-conditioned fibers and old-conditioned fibers were not different with respect to force deficit or the frequency of breakages. A potential mechanism for the increased resistance to stretch-induced damage of old-conditioned fibers is that, through intracellular damage and subsequent degeneration and regeneration, weaker sarcomeres were replaced by stronger sarcomeres. These data indicate that, despite the association of high fiber breakage rates and large force deficits with aging, the detrimental characteristics of old fibers were improved by a conditioning program that altered both sarcomeric characteristics as well as the overall structural integrity of the fibers.

Breakage of a Third Generation Gamma Nail: A Case Report and Review of the Literature

Directory of Open Access Journals (Sweden)

Takashi Iwakura

2013-01-01

Full Text Available The use of intramedullary nails to treat trochanteric fractures of the femur has increased with the increasing size of the elderly population. The third generation Gamma nail is currently one of the most popular devices for the treatment of trochanteric fractures. Nail breakage is a rare complication, possibly resulting from fatigue fracture of the implant. We present the first reported case of breakage of a third generation Gamma nail that was not used to treat a pathological fracture. An 83-year-old woman with an unstable trochanteric fracture of the femur was treated using a third generation Gamma nail. She was referred to our hospital 14 months postoperatively with nail breakage at the opening for the lag screw. The breakage was secondary to nonunion, which was thought to be mainly due to insufficient reduction of the fracture. The broken nail was removed, and the patient underwent cemented bipolar hemiarthroplasty. At followup 18 months later, she was mobile with a walker and asymptomatic with no complications. This case shows that inadequate operation such as insufficient reduction of the trochanteric fracture may result in nonunion and implant breakage, even when using a high-strength, well-designed implant.

Empirical Formulae for Breakage of Dolosse and Tetrapods

DEFF Research Database (Denmark)

Burcharth, H. F.; d'Angremond, K.; Meer, W. van der

2000-01-01

The slender, complex types of armour units, such as Tetrapods and Dolosse are widely used for rubble mound breakwaters. Many failures of such breakwaters were caused by unforeseen early breakage of the units, thus revealing an inbalance between the strength (structural integrity) of the units...... and the hydraulic stability (resistance to displacements) of the armour layers. Breakage occurs when the stresses from the static, pulsating and impact loads exceeds the tensile strength of the concrete. While the hydraulic stability can be studied in Froude-scale hydraulic model tests, it is not possible to study...... armour unit stresses in small scale models. This is partly because the strain in model armour units are too small to be recorded, and partly because the scaling law for impact load generated stresses is nonlinear. The paper discusses the scaling laws related to type of stresses and presents a method...

FBH1 co-operates with MUS81 in inducing DNA double-strand breaks and cell death following replication stress

DEFF Research Database (Denmark)

Fugger, Kasper; Chu, Wai Kit; Haahr, Peter

2013-01-01

The molecular events occurring following the disruption of DNA replication forks are poorly characterized, despite extensive use of replication inhibitors such as hydroxyurea in the treatment of malignancies. Here, we identify a key role for the FBH1 helicase in mediating DNA double-strand break...... formation following replication inhibition. We show that FBH1-deficient cells are resistant to killing by hydroxyurea, and exhibit impaired activation of the pro-apoptotic factor p53, consistent with decreased DNA double-strand break formation. Similar findings were obtained in murine ES cells carrying...... of replication stress. Our data suggest that FBH1 helicase activity is required to eliminate cells with excessive replication stress through the generation of MUS81-induced DNA double-strand breaks....

Connecting localized DNA strand displacement reactions

Science.gov (United States)

Mullor Ruiz, Ismael; Arbona, Jean-Michel; Lad, Amitkumar; Mendoza, Oscar; Aimé, Jean-Pierre; Elezgaray, Juan

2015-07-01

Logic circuits based on DNA strand displacement reactions have been shown to be versatile enough to compute the square root of four-bit numbers. The implementation of these circuits as a set of bulk reactions faces difficulties which include leaky reactions and intrinsically slow, diffusion-limited reaction rates. In this paper, we consider simple examples of these circuits when they are attached to platforms (DNA origamis). As expected, constraining distances between DNA strands leads to faster reaction rates. However, it also induces side-effects that are not detectable in the solution-phase version of this circuitry. Appropriate design of the system, including protection and asymmetry between input and fuel strands, leads to a reproducible behaviour, at least one order of magnitude faster than the one observed under bulk conditions.Logic circuits based on DNA strand displacement reactions have been shown to be versatile enough to compute the square root of four-bit numbers. The implementation of these circuits as a set of bulk reactions faces difficulties which include leaky reactions and intrinsically slow, diffusion-limited reaction rates. In this paper, we consider simple examples of these circuits when they are attached to platforms (DNA origamis). As expected, constraining distances between DNA strands leads to faster reaction rates. However, it also induces side-effects that are not detectable in the solution-phase version of this circuitry. Appropriate design of the system, including protection and asymmetry between input and fuel strands, leads to a reproducible behaviour, at least one order of magnitude faster than the one observed under bulk conditions. Electronic supplementary information (ESI) available. See DOI: 10.1039/C5NR02434J

Radiation-induced DNA single-strand scission and its rejoining in spermatogonia and spermatozoa of mouse

International Nuclear Information System (INIS)

Ono, T.; Okada, S.

1977-01-01

Gamma-ray-induced DNA single-strand scissions and the ability to repair the scissions in spermatogonia from young mice and in spermatozoa from adult mice were studied quantitatively by an alkaline sucrose density-gradient centrifugation method. The average size of DNAs in non-irradiated spermatogonia was 2.6-3.0xx10 8 daltons, similar to those of a spermatid-rich population, and the size of DNA in non-irradiated spermatozoa was 1.2x10 8 daltons. In spermatogonia, the radiosensitivity of DNA was 0.42 single-strand breaks/10 12 daltons of DNA/rad in oxic conditions and only 0.24 under anoxic conditions. In spermatozoa the break efficiency of DNA was 0.22 single-strand breaks/10 12 daltons of DNA/rad under oxic conditions and altered little under anoxic irradiation. The DNA scissions were efficiently repaired in spermatogonia within 10 min, whereas the breaks in spermatozoa were not rejoined at all even after two days of post-irradiation time. The radiosensitivities of DNA, repair capability and non- and/or slowreparable DNA scissions were compared in spermatogonium-rich, spermatid-rich and spermatozoanrich populations

Electronic cigarettes induce DNA strand breaks and cell death independently of nicotine in cell lines.

Science.gov (United States)

Yu, Vicky; Rahimy, Mehran; Korrapati, Avinaash; Xuan, Yinan; Zou, Angela E; Krishnan, Aswini R; Tsui, Tzuhan; Aguilera, Joseph A; Advani, Sunil; Crotty Alexander, Laura E; Brumund, Kevin T; Wang-Rodriguez, Jessica; Ongkeko, Weg M

2016-01-01

Evaluate the cytotoxicity and genotoxicity of short- and long-term e-cigarette vapor exposure on a panel of normal epithelial and head and neck squamous cell carcinoma (HNSCC) cell lines. HaCaT, UMSCC10B, and HN30 were treated with nicotine-containing and nicotine-free vapor extract from two popular e-cigarette brands for periods ranging from 48 h to 8 weeks. Cytotoxicity was assessed using Annexin V flow cytometric analysis, trypan blue exclusion, and clonogenic assays. Genotoxicity in the form of DNA strand breaks was quantified using the neutral comet assay and γ-H2AX immunostaining. E-cigarette-exposed cells showed significantly reduced cell viability and clonogenic survival, along with increased rates of apoptosis and necrosis, regardless of e-cigarette vapor nicotine content. They also exhibited significantly increased comet tail length and accumulation of γ-H2AX foci, demonstrating increased DNA strand breaks. E-cigarette vapor, both with and without nicotine, is cytotoxic to epithelial cell lines and is a DNA strand break-inducing agent. Further assessment of the potential carcinogenic effects of e-cigarette vapor is urgently needed. Copyright © 2015 Elsevier Ltd. All rights reserved.

Molecular characterization of a complex site-specific radiation-induced DNA double-strand break

International Nuclear Information System (INIS)

Datta, K.; Dizdaroglu, M.; Jaruga, P.; Neumann, R.D.; Winters, T.A.

2003-01-01

Radiation lethality is a function of radiation-induced DNA double-strand breaks (DSB). Current models propose the lethality of a DSB to be a function of its structural complexity. We present here for the first time a map of damage associated with a site-specific double-strand break produced by decay of 125 I in a plasmid bound by a 125 I-labeled triplex forming oligonucleotide ( 125 I-TFO). The E. coli DNA repair enzymes, endonuclease IV (endo IV), endonuclease III (endo III), and formamidopyrimidine-DNA glycosylase (Fpg), which recognize AP sites, and pyrimidine and purine base damage respectively, were used as probes in this study. 125 I-TFO bound plasmid was incubated with and without DMSO at -80 deg C for 1 month. No significant difference in DSB yield was observed under these conditions. A 32 base pair fragment from the upstream side of the decay site was isolated by restriction digestion and enzymatically probed to identify damage sites. Endo IV treatment of the 5'-end labeled upper strand indicated clustering of AP sites within 3 bases downstream and 7 bases upstream of the targeted base. Also, repeated experiments consistently detected an AP site 4 bases upstream of the 125 Itarget base. This was further supported by complementary results with the 3'-end labeled upper strand. Endo IV analysis of the lower strand also shows clustering of AP sites near the DSB end. Endo III and Fpg probing demonstrated that base damage is also clustered near the targeted break site. DSBs produced in the absence of DMSO displayed a different pattern of enzyme sensitive damage than those produced in the presence of DMSO. Identification of specific base damage types within the restriction fragment containing the DSB end was achieved with GC/MS. Base damage consisted of 8-hydroguanine, 8-hydroxyadenine, and 5-hydroxycytosine. These lesions were observed at relative yields of 8-hydroguanine and 5-hydroxycytosine to 8-hydroxyadenine of 7.4:1 and 4.7:1, respectively, in the absence

Differences in heavy-ion-induced DNA double-strand breaks in a mouse DNA repair-deficient mutant cell line (SL3-147) before and after chromatin proteolysis

International Nuclear Information System (INIS)

Murakami, Masahiro; Eguchi-Kasai, Kiyomi; Sato, Koki; Minohara, Shinichi; Kanai, Tatsuaki; Yatagai, Fumio.

1995-01-01

DNA double-strand breaks induced by X- or neon beam-irradiation in a DNA double-strand break-repair-deficient mutant cell line (SL3-147) were examined. The increase in the number of DNA double-strand breaks was dose-depend after irradiation with X-rays and neon beams and was enhanced by chromatin-proteolysis treatment before irradiation. These results suggest that the induction of DNA double-strand breaks by ionizing radiation, including heavy-ions, is influenced by the chromatin structure. (author)

Visual Inspection for Breakage of Micro-milling Cutter

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WANG Lei

2014-11-01

Full Text Available In order to realize visual inspection for breakage of micro-milling cutter, a developed image acquisition method of the surface of a micro-milling cutter was constructed and a classification method based on multilayer neural network was proposed in this article. While the milling cutter was rotating at a constant speed, a camera was triggered by a rotary encoder to capture a series of images. And the developed image of milling cutter was created by image mosaic algorithms. The moment of regional feature as well as the gray feature of the tooth edge was extracted as the input vector of neural network. The feature vector includes moment of inertia, geometric central moment, three-dimensional invariants moment and the gray value of the projection on two principal axis directions of the tooth region. By designing a proper neural network, breakage defects can be detected 100 %. And the false discovery rate is 0.5 %.

Carbon ion induced DNA double-strand breaks in melanophore B{sub 16}

Energy Technology Data Exchange (ETDEWEB)

Zengquan, Wei; Guangming, Zhou; Jufang, Wang; Jing, He; Qiang, Li; Wenjian, Li; Hongmei, Xie; Xichen, Cai; Huang, Tao; Bingrong, Dang; Guangwu, Han [Chinese Academy of Sciences, Lanzhou (China). Inst. of Modern Physics; Qingxiang, Gao [Lanzhou Univ. (China)

1997-09-01

DNA double-strand breaks (DSBs) in melanophore B{sub 16} induced by plateau and extended Bragg peak of 75 MeV/u {sup 12}C{sup 6+} ions were studied by using a technique of inverse pulsed-field gel electrophoresis (PIGE). DNA fragment lengths were distributed in two ranges: the larger in 1.4 Mbp-3.2 Mbp and the smaller in less than 1.2 Mbp. It indicates that distribution of DNA fragments induced by heavy ion irradiation is not stochastic and there probably are sensitive sites to heavy ions in DNA molecules of B{sub 16}. Percentage of DNA released from plug (PR) increased and trended towards a quasi-plateau {proportional_to}85% as dose increased. Content of the larger fragments decreased and flattened with increasing dose while content of the smaller ones increased and trended towards saturation. (orig.)

Radiation-chemical discussion on inverse dose-rate effect observed in radiation-induced strand breaks of plasmid DNA

International Nuclear Information System (INIS)

Masuda, Takahiro

1994-01-01

Experimental results of inverse dose-rate effect, so-called Kada Effects, which was published by Takakura and her coworkers on radiation-induced strand breaks of plasmid DNA in aerated aqueous solution, have been kinetically analyzed and discussed on the basis of radiation chemistry. the kinetic analysis indicates that there are two possible mechanisms; 1) equilibrium mixture of O 2 - and HO 2 is responsible for strand breaks of DNA, and 2) peroxyl radical produced from citrate is effective for the strand breaks. However, the detailed kinetic analysis revealed that the latter is improbable because unimolecular decay of the peroxyl radical must be assumed to be negligible for its participation despite fast decay of analogous organic peroxyl radicals. The analysis has also given 9.93±0.10 dm 3 mol -1 s -1 per nucleotide unit, which corresponds to 7.62 x 10 4 dm 3 mol -1 s -1 per DNA molecule, as the rate constant for the reaction of the equilibrium mixture with plasmid pBR 322 DNA. Furthermore the probability that the reaction of the mixture with a nucleotide unit of DNA leads to strand breaks was obtained to be 3.36 x 10 -3 for gamma-irradiated system and 1.98 x 10 -3 for beta-irradiated system, respectively. (author)

Experimental Research on the Specific Energy Consumption of Rock Breakage Using Different Waterjet-Assisted Cutting Heads

Directory of Open Access Journals (Sweden)

Hongxiang Jiang

2018-01-01

Full Text Available To investigate the specific energy consumption (SE of rock breakage by cutting heads assisted by different types of waterjet and to identify optimal waterjet parameters and assistance types, rock cutting with and without waterjets was carried on a rock fragmentation test bed. SE is a comprehensive evaluation index and was developed according to the applied load on cutting head, and the SE under different cutting conditions was compared and analyzed. The results show that the SE of rock breakage without waterjet assistance increased with the increasing of rock compressive strength (RCS but that the limited drilling depth decreased. The effect of the waterjet pressure on the SE of rock breakage by the cutting head I was marked, and SE decreased by 30∼40% when the ratio between RCS and waterjet pressure was less than 1. However, the function of the waterjet assistance was poor; therefore, a ratio of 1 could be used to distinguish the rock breakage effect of cutting head I. For cutting head II, the rock damage from the waterjet impact was limited due to the large waterjet standoff distance; thus the rock breakage performance of cutting head II was also limited. The waterjet impacting at the tip of the conical pick using cutting head III could enter into the cracks caused by the mechanical pick and fracture the rock. Therefore, the rock breakage performance of cutting head III was better than that of cutting head II.

Protective role of 3-nitrotyrosine against gamma radiation-induced DNA strand breaks: A comparison study with tyrosine

International Nuclear Information System (INIS)

Shi Weiqun; Ni Meinan; Kong Fuquan; Sui Li; Hu Jia; Xu Diandou; Li Yanmei

2008-01-01

3-Nitrotyrosine(3-NY) has been reported as a potential source of reactive oxygen species (ROSs). In this work, plasmid pBR322 DNA was irradiated by gamma rays in aqueous solution in presence and absence of 3-NY, DNA strand breaks were analyzed by neutral electrophoresis followed by quantification with image analysis software. It was found that the presence of 3-NY could effectively reduce radiation-induced DNA strand breaks. A side-by-side comparison was performed between 3-NY and tyrosine, the results showed that the protective role 3-NY was comparable with tyrosine, which might imply that protein tyrosine nitration might not significantly decrease its ability as a free radical scavenger

Induction of DNA strand breaks in 14C-labelled cells

International Nuclear Information System (INIS)

Sundell-Bergman, S.; Johanson, K.J.

1979-01-01

Chinese hamster cells grown in vitro were labelled with 14 C-thymidine for 18 hours and after 3 hours in non-radioactive medium they were stored at 0 0 C for various periods ( 1 to 12 hours). During this treatment a number of DNA strand breaks were induced by 14 C decay which were not repaired at 0 0 C. The number of DNA strand breaks was determined using the DNA unwinding technique. At 0.5-1 dpm per cell a detectable number of DNA strand breaks were found. Treatment for six hours (1 dpm per cell) reduced the percentage of double-stranded DNA from 80 to 70%, corresponding to about 750 DNA strand breaks per cell. The rejoining of DNA strand breaks was studied after treatment for 12 hours at 0 0 C followed by incubation of the cells for various periods at 37 0 C. Most of the DNA strand breaks induced by 14 C decay at 0 0 C were repaired after incubation at 37 0 C for 15 minutes. Assuming an absorbed dose of 1.8 mGy per 14 C decay to the cell nucleus an RBE value close to 1 was found for internal irradiation from 14 C decay as compared with 60 Co-gamma irradiation. (author)

Tool wear and breakage monitoring in machining

International Nuclear Information System (INIS)

Madl, J.

1992-01-01

Risk minimization of metal cutting operations is one of the main problems of metal cutting technology. This paper describes some aspects in monitoring and control of machining processes. Tool monitoring is the fokus of machining process monitoring. Tool breakage and tool life recognition are the main problems of tool monitoring. All problems of this type of monitoring have not yet been fully solved. (orig.)

Radiation-induced DNA damage in tumors and normal tissues. III. Oxygen dependence of the formation of strand breaks and DNA-protein crosslinks

International Nuclear Information System (INIS)

Zhang, H.; Wallen, C.A.; Wheeler, K.T.; Joch, C.J.

1995-01-01

Results from several laboratories, including ours, have suggested that measurements of radiation-induced DNA strand breaks and DNA-protein crosslinks (DPCs) may be used to estimate the hypoxic fraction or fractional hypoxic volume of tumors and normal tissues. This suggestion has been predicated on both published and nonpublished information that (1) the oxygen dependence of the formation of strand breaks in irradiated mammalian cells is similar to the oxygen dependence of radiation-produced cell killing, and (2) the oxygen dependence of the formation of DPCs in irradiated mammalian cells is the mirror image of the oxygen dependence of radiation-induced cell killing. However, the published studies that attempted to determine the relationship between the oxygen dependence of the formation of strand breaks and the radiation sensitivity of mammalian cells were not performed at 37 degrees C, the exact oxygen concentrations were not always known, and the results were conflicting. In addition, most of the data on the oxygen dependence of the formation of DPCs are unpublished. Consequently, we have undertaken a comprehensive investigation of one cell line, 9L/Ro rat brain tumor cells, to determine if the shape of the oxygen dependence curve and the K m value for radiation-induced strand breaks and DPCs were similar when 9L cells were irradiated under both ideal gas-liquid equilibrium conditions at 4 degrees C and nonideal gas-liquid equilibrium conditions at 37 degrees C. At 4 degrees C under ideal gas-liquid equilibrium conditions, the K m for the formation of strand breaks was approximately 0.0045 mM, and Km for radiation sensitivity was approximately 0.005mM. A similar comparison for the formation of DPCs at 4 degrees C could not be made, because the efficiency of the formation of DPC was much lower at 4 degrees C than at 37 degrees C. 30 refs., 3 figs

Explanation for excessive DNA single-strand breaks and endogenous repair foci in pluripotent mouse embryonic stem cells.

Science.gov (United States)

Banáth, J P; Bañuelos, C A; Klokov, D; MacPhail, S M; Lansdorp, P M; Olive, P L

2009-05-01

Pluripotent mouse embryonic stem cells (mES cells) exhibit approximately 100 large gammaH2AX repair foci in the absence of measurable numbers of DNA double-strand breaks. Many of these cells also show excessive numbers of DNA single-strand breaks (>10,000 per cell) when analyzed using the alkaline comet assay. To understand the reasons for these unexpected observations, various methods for detecting DNA strand breaks were applied to wild-type mES cells and to mES cells lacking H2AX, ATM, or DNA-PKcs. H2AX phosphorylation and expression of other repair complexes were measured using flow and image analysis of antibody-stained cells. Results indicate that high numbers of endogenous gammaH2AX foci and single-strand breaks in pluripotent mES cells do not require ATM or DNA-PK kinase activity and appear to be associated with global chromatin decondensation rather than pre-existing DNA damage. This will limit applications of gammaH2AX foci analysis in mES cells to relatively high levels of initial or residual DNA damage. Excessive numbers of single-strand breaks in the alkaline comet assay can be explained by the vulnerability of replicating chromatin in mES cells to osmotic shock. This suggests that caution is needed in interpreting results with the alkaline comet assay when applied to certain cell types or after treatment with agents that make chromatin vulnerable to osmotic changes. Differentiation of mES cells caused a reduction in histone acetylation, gammaH2AX foci intensity, and DNA single-strand breakage, providing a link between chromatin structural organization, excessive gammaH2AX foci, and sensitivity of replicating mES cell chromatin to osmotic shock.

Breakage of internal maxillary distractor: considerable complication of maxillary distraction osteogenesis.

Science.gov (United States)

Aikawa, Tomonao; Iida, Seiji; Isomura, Emiko T; Namikawa, Mari; Matsuoka, Yudai; Yamada, Chiaki; Yamamoto, Taku; Takigawa, Yoko

2008-07-01

Maxillary distraction osteogenesis using intraoral distractors is now one of the standard treatments of maxillary retrusion. This report shows 2 cases of breakage of this internal maxillary distractor in patients with cleft lip and palate; one was observed during the distraction period and the other was during the retention period. The first case required a rotational movement of the distraction segment, and this movement caused the laterally dislocation of the posterior part of the distractor, where the distractor suffered some mechanical forces by mouth opening. In the latter case, breakage of distractor was observed on the radiographs taken 3 months after distraction and this complication may have been caused by mechanical force by occlusion and mastication. Both breakages were found at the joint of the anchorage plate and the extension rod, which has some flexibility for adjusting the plate to the bone surface. Therefore, surgeons should pay special attention for this mechanical weak area in this distractor not only during the advancement period, but also during the retention period and should avoid unnecessary frequent bending for adopting the bone surface, which directly weakens the joint.

Mechanical Properties of Photovoltaic Silicon in Relation to Wafer Breakage

Science.gov (United States)

Kulshreshtha, Prashant Kumar

This thesis focuses on the fundamental understanding of stress-modified crack-propagation in photovoltaic (PV) silicon in relation to the critical issue of PV silicon "wafer breakage". The interactions between a propagating crack and impurities/defects/residual stresses have been evaluated for consequential fracture path in a thin PV Si wafer. To investigate the mechanism of brittle fracture in silicon, the phase transformations induced by elastic energy released at a propagating crack-tip have been evaluated by locally stressing the diamond cubic Si lattice using a rigid Berkovich nanoindenter tip (radius ≈50 nm). Unique pressure induced phase transformations and hardness variations have been then related to the distribution of precipitates (O, Cu, Fe etc.), and the local stresses in the wafer. This research demonstrates for the first time the "ductile-like fracture" in almost circular crack path that significantly deviates from its energetically favorable crystallographic [110](111) system. These large diameter (≈ 200 mm) Si wafers were sliced to less than 180 microm thickness from a Czochralski (CZ) ingot that was grown at faster than normal growth rates. The vacancy (vSi) driven precipitation of oxygen at enhanced thermal gradients in the wafer core develops large localized stresses (upto 100 MPa) which we evaluated using Raman spectral analysis. Additional micro-FTIR mapping and microscopic etch pit measurements in the wafer core have related the observed crack path deviations to the presence of concentric ring-like distributions of oxygen precipitates (OPs). To replicate these "real-world" breakage scenarios and provide better insight on crack-propagation, several new and innovative tools/devices/methods have been developed in this study. An accurate quantitative profiling of local stress, phase changes and load-carrying ability of Si lattice has been performed in the vicinity of the controlled micro-cracks created using micro-indentations to represent

Influence of the potential well on the breakage rate of colloidal aggregates in simple shear and uniaxial extensional flows.

Science.gov (United States)

Ren, Zhiqiang; Harshe, Yogesh M; Lattuada, Marco

2015-06-02

In this work we build on our previous paper (Harshe, Y. M.; Lattuada, M. Langmuir 2012, 28, 283-292) and compute the breakage rate of colloidal aggregates under the effect of shear forces by means of Stokesian dynamics simulations. A library of clusters made of identical spherical particles covering a broad range of masses and fractal dimension values (from 1.8 to 3.0) was generated by means of a combination of several Monte Carlo methods. DLVO theory has been used to describe the interparticle interactions, and contact forces have been introduced by means of the discrete element method. The aggregate breakage process was investigated by exposing them to well-defined shear forces, generated under both simple shear and uniaxial extensional flow conditions, and by recording the time required to reach the first breakage event. It has been found that the breakage rate of clusters was controlled by the potential well between particles as described by DLVO theory. A semiempirical Arrhenius-type exponential equation that relates the potential well to the breakage rate has been used to fit the simulation results. The dependence of the breakage process on the radius of gyration, on the external shear strength, and on the fractal dimension has been obtained, providing a very general relationship for the breakage rate of clusters. It was also found that the fragment mass distribution is insensitive to the presence of electrostatic repulsive interactions. We also clarify the physical reason for the large difference in the breakage rate of clusters between simple shear and the uniaxial extensional flow using a criterion based on the energy dissipation rate. Finally, in order to answer the question of the minimum cluster size that can break under simple shear conditions, a critical rotation number has been introduced, expressing the maximum number of rotations that a cluster exposed to simple shear could sustain before breakage.

Epidermal growth factor stimulating reparation of γ-ray-induced single-strand breaks predominantly in untranscribed DNA of HeLa cells

International Nuclear Information System (INIS)

Igusheva, O.A.; Bil'din, V.N.; Zhestyanikov, V.D.

1994-01-01

Considerable evidence suggest that genomic DNA undergoes reparation unevenly because of different transcription activities of its particular sequence. It is highly probably that transcriptional factors are necessary for postion stages of excision reparation and for reparation of single-strand DNA breaks caused by ionizing radiation. There is evidence suggesting that DNA lesions inflicted by γ-radiation is preferentially initiated in transcribed rather than in untranscribed DNA species. This paper looks at the relationship between stimulatory effect of epidermal growth factor (EGF) on reparation of single-strand DNA breaks and reparation of the damage done to active and inert fragments of chromatin. The results show that EGF stimulates reparation of single-strand DNA breaks induced by γ-radiation more effectively in untranscribed than in transcribed DNA. 13 refs., 1 fig., 1 tab

Hydraulic breakage of tanks for the transport of uranium hexafluoride (UF6)

International Nuclear Information System (INIS)

Biaggio, A.L.; Lee Gonzales, H.M.; Lopez Vietri, J.R.; Novo, R.G.

1987-01-01

To begin with, the tank models that are proposed by the international norms for the transport and storage of hexafluoride of uranium (UF 6 ) are briefly described. The operations related to the transport in its different forms are also described, particularly those that can produce the hydraulic breakage of tanks during its course or in later stages, when incorrectly performed. With reference to those operations, the most important physicochemical properties of UF 6 as for safety are analyzed. A quantitative evaluation of the deviations of parameters that are controlled during the heating of tanks, comparing them with the normative nominal values, is performed. Adopting some simplifying hypothesis, a general study, applicable to all tank models proposed by norms, is carried out to determine the temperature at which the hydraulic breakage takes place when they are heated in closed-valve conditions. A curve is obtained by plotting the hydraulic breakage temperature against the filling degree. To conclude, the values obtained are compared with the results of other theoretical studies on this subject. (Author)

Eliminating a Major Cause of Wire Drawing Breakage in A-15 High-Field Superconductors

International Nuclear Information System (INIS)

Austen, Alfred R.

2003-01-01

Eliminating a Major Cause of Wire Drawing Breakage in A-15 High-Field Superconductors Phase 1 Summary Purpose of the research: The Phase 1 goal was to make a significant improvement in the wire drawing technology used for difficult to draw superconductor precursor composites. Many ductile Nb-Al and Nb-Sn precursor wire composites have experienced the onset of wire drawing breakage at about 1.5 mm diameter. Phase 1 focused on evaluating the role that precision rigid guidance of the wire into the drawing die and the hydrostatic stress state at the die entrance played in preventing wire breakage. Research carried out: The research performed depended upon the construction of both a mechanical wire guide and a hydrostatic pressure stiffened wire guidance system. Innovare constructed the two wire guidance systems and tested them for their ability to reduce wire drawing breakage. One set of hardware provided rigid alignment of the wires to their wire drawing die axes within 0.35 degrees using ''hydrostatic pressure stiffening'' to enable the precision guidance strategy to be implemented for these highly flexible small diameter wires. This apparatus was compared to a guide arrangement that used short span mechanical guide alignment with a misalignment limit of about 0.75 degrees. Four A-15 composite wires with breakage histories were drawn to evaluate the use of these wire guiding systems to reduce and/or eliminate wire breakage. Research findings and results: In Phase 1, a breakthrough in wire drawing technology for A-15 superconductor composites was achieved by dramatically limiting or eliminating breakage in four different A-15 composite precursor wire designs during the drawing of these very desirable composites that previously could not be drawn to near final size. Research results showed that the proposed Phase 1 mechanical wire guides were sufficiently effective and successful in eliminating breakage when used along with other advanced wire drawing technology to

Effects of DNA double-strand and single-strand breaks on intrachromosomal recombination events in cell-cycle-arrested yeast cells

International Nuclear Information System (INIS)

Galli, A.; Schiestl, R.H.

1998-01-01

Intrachromosomal recombination between repeated elements can result in deletion (DEL recombination) events. We investigated the inducibility of such intrachromosomal recombination events at different stages of the cell cycle and the nature of the primary DNA lesions capable of initiating these events. Two genetic systems were constructed in Saccharomyces cerevisiae that select for DEL recombination events between duplicated alleles of CDC28 and TUB2. We determined effects of double-strand breaks (DSBs) and single-strand breaks (SSBs) between the duplicated alleles on DEL recombination when induced in dividing cells or cells arrested in G1 or G2. Site-specific DSBs and SSBs were produced by overexpression of the I-Sce I endonuclease and the gene II protein (gIIp), respectively. I-Sce I-induced DSBs caused an increase in DEL recombination frequencies in both dividing and cell-cycle-arrested cells, indicating that G1- and G2-arrested cells are capable of completing DSB repair. In contrast, gIIp-induced SSBs caused an increase in DEL recombination frequency only in dividing cells. To further examine these phenomena we used both γ-irradiation, inducing DSBs as its most relevant lesion, and UV, inducing other forms of DNA damage. UV irradiation did not increase DEL recombination frequencies in G1 or G2, whereas γ-rays increased DEL recombination frequencies in both phases. Both forms of radiation, however, induced DEL recombination in dividing cells. The results suggest that DSBsbut not SSBs induce DEL recombination, probably via the single-strand annealing pathway. Further, DSBs in dividing cells may result from the replication of a UV or SSB-damaged template. Alternatively, UV induced events may occur by replication slippage after DNA polymerase pausing in front of the damage. (author)

The effects of radioprotective agents on the radiation-induced DNA single strand breaks

International Nuclear Information System (INIS)

Rhiu, Sung Ryul; Ko, Kyung Hwan; Jung, In Yong; Cho, Chul Ku; Kim, Tae Hwan; Park, Woo Wiun; Kim, Sung Ho; Ji, Young Hoon; Kim, Kyung Jung; Bang, Hio Chang; Jung, Young Suk; Choi, Moon Sik

1992-04-01

With the increased use of atomic energy in science, industry, medicine and public power production, the probability of nuclear accidents certainly appears to be on the increase. Therefore, early medical diagnosis and first-aid are needed urgently to establish an efficient treatment. We carried out the studies of radiation protector such as DDC, MEA, WR-2721 and variety of decontaminator with a view to establishing the protective measure and diagnostic standards for safety of worker and neighbors living around the radiation area in case of occurring the accidental contamination. In this experiment, we examined radiation-induced DNA single strand breaks as one of the study on molecular biology of the response of cells to radiation because an understanding of the radiation-induced damage in molecular level would add to our knowledge of radiation protection and treatment. (Author)

A novel technique using DNA denaturation to detect multiply induced single-strand breaks in a hydrated plasmid DNA molecule by X-ray and 4He2+ ion irradiation

International Nuclear Information System (INIS)

Yokoya, A.; Shikazono, N.; Fujii, K.; Noguchi, M.; Urushibara, A.

2011-01-01

To detect multiple single-strand breaks (SSBs) produced in plasmid DNA molecules by direct energy deposition from radiation tracks, we have developed a novel technique using DNA denaturation by which irradiated DNA is analysed as single-strand DNA (SS-DNA). The multiple SSBs that arise in both strands of DNA, but do not induce a double-strand break, are quantified as loss of SS-DNA using agarose gel electrophoresis. We have applied this method to X-ray and 4 He 2+ ion-irradiated samples of fully hydrated pUC18 plasmid DNA. The fractions of both SS-DNA and closed circular DNA (CC-DNA) exponentially decrease with the increasing dose of X rays and 4 He 2+ ions. The efficiency of the loss of SS-DNA was half that of CC-DNA for both types of irradiation, indicating that one of two strands in DNA is not broken when one SSB is produced in CC-DNA by irradiation. Contrary to our initial expectation, these results indicate that SSBs are not multiply induced even by high linear energy transfer radiation distributed in both strands. (authors)

Nucleic Acid Analogue Induced Transcription of Double Stranded DNA

DEFF Research Database (Denmark)

1998-01-01

RNA is transcribed from a double stranded DNA template by forming a complex by hybridizing to the template at a desired transcription initiation site one or more oligonucleic acid analogues of the PNA type capable of forming a transcription initiation site with the DNA and exposing the complex...... to the action of a DNA dependant RNA polymerase in the presence of nucleoside triphosphates. Equal length transcripts may be obtained by placing a block to transcription downstream from the initiation site or by cutting the template at such a selected location. The initiation site is formed by displacement...... of one strand of the DNA locally by the PNA hybridization....

Parametric Study on Responses of a Self-Anchored Suspension Bridge to Sudden Breakage of a Hanger

Directory of Open Access Journals (Sweden)

Wenliang Qiu

2014-01-01

Full Text Available The girder of self-anchored suspension bridge is subjected to large compression force applied by main cables. So, serious damage of the girder due to breakage of hangers may cause the collapse of the whole bridge. With the time increasing, the hangers may break suddenly for their resistance capacities decrease due to corrosion. Using nonlinear static and dynamic analysis methods and adopting 3D finite element model, the responses of an actual self-anchored suspension bridge to sudden breakage of hangers are studied in this paper. The results show that the sudden breakage of a hanger causes violent vibration and large changes in internal forces of the bridge. In the process of the vibration, the maximum tension of hanger produced by breakage of a hanger exceeds 2.22 times its initial value, and the reaction forces of the bearings increase by more than 1.86 times the tension of the broken hanger. Based on the actual bridge, the influences of some factors including flexural stiffness of girder, torsion stiffness of girder, flexural stiffness of main cable, weight of girder, weight of main cable, span to sag ratio of main cable, distance of hangers, span length, and breakage time of hanger on the dynamic responses are studied in detail, and the influencing extent of the factors is presented.

Parametric study on responses of a self-anchored suspension bridge to sudden breakage of a hanger.

Science.gov (United States)

Qiu, Wenliang; Jiang, Meng; Huang, Cailiang

2014-01-01

The girder of self-anchored suspension bridge is subjected to large compression force applied by main cables. So, serious damage of the girder due to breakage of hangers may cause the collapse of the whole bridge. With the time increasing, the hangers may break suddenly for their resistance capacities decrease due to corrosion. Using nonlinear static and dynamic analysis methods and adopting 3D finite element model, the responses of an actual self-anchored suspension bridge to sudden breakage of hangers are studied in this paper. The results show that the sudden breakage of a hanger causes violent vibration and large changes in internal forces of the bridge. In the process of the vibration, the maximum tension of hanger produced by breakage of a hanger exceeds 2.22 times its initial value, and the reaction forces of the bearings increase by more than 1.86 times the tension of the broken hanger. Based on the actual bridge, the influences of some factors including flexural stiffness of girder, torsion stiffness of girder, flexural stiffness of main cable, weight of girder, weight of main cable, span to sag ratio of main cable, distance of hangers, span length, and breakage time of hanger on the dynamic responses are studied in detail, and the influencing extent of the factors is presented.

The adsorption-desorption transition of double-stranded DNA interacting with an oppositely charged dendrimer induced by multivalent anions.

Science.gov (United States)

Jiang, Yangwei; Zhang, Dong; Zhang, Yaoyang; Deng, Zhenyu; Zhang, Linxi

2014-05-28

The adsorption-desorption transition of DNA in DNA-dendrimer solutions is observed when high-valence anions, such as hexavalent anions, are added to the DNA-dendrimer solutions. In the DNA-dendrimer solutions with low-valence anions, dendrimers bind tightly with the V-shaped double-stranded DNA. When high-valence anions, such as pentavalent or hexavalent anions, are added to the DNA-dendrimer solutions, the double-stranded DNA chains can be stretched straightly and the dendrimers are released from the double-stranded DNA chains. In fact, adding high-valence anions to the solutions can change the charge spatial distribution in the DNA-dendrimer solutions, and weaken the electrostatic interactions between the positively charged dendrimers and the oppositely charged DNA chains. Adsorption-desorption transition of DNA is induced by the overcharging of dendrimers. This investigation is capable of helping us understand how to control effectively the release of DNA in gene/drug delivery because an effective gene delivery for dendrimers includes non-covalent DNA-dendrimer binding and the effective release of DNA in gene therapy.

Investigation of DNA strand breaks induced by 7Li and 12C ions

International Nuclear Information System (INIS)

Sui Li; Zhao Kui; Ni Meinan; Guo Jiyu; Luo Hongbing; Mei Junping; Lu Xiuqin; Zhou Ping

2004-01-01

Deoxyribonucleic acid (DNA) is an important biomacromolecule. It is a carrier of genetic information and a critical target for radiobiological effects. Numerous lesions have been identified in irradiated DNA. DNA double strand breaks (DSBs) are considered as the most important initial damage of all biological effects induced by ionizing radiation. In this experiment, DNA DSBs induced by heavy ions in the early period were investigated with atomic force microscopy (AFM). Choosing 7 Li and 12 C heavy ions with different LET values delivered by HI-13 tandem accelerator respectively, purified plasmid DNA samples in aqueous solution were irradiated at different doses. AFM was used for nanometer-level-structure analysis of DNA damage induced by these two kinds of heavy ions. Measurement of the DNA fragment lengths was accomplished with the Scion Image analyzed soft-ware. Change laws of three forms of DNA, supercoils, open circular and linear form as dose increased were obtained. Distributed function of DNA fragment length was also obtained, and fitted with Tsallis entropy statistical theory. (author)

Single-strand breaks induced in Bacillus subtilis DNA by ultraviolet light: action spectrum and properties

International Nuclear Information System (INIS)

Peak, M.J.; Peak, J.G.

1982-01-01

The induction of single-strand breaks (alkali-labile bonds plus frank breaks) in the DNA of Bacillus subtilis irradiated in vivo by monochromatic UV light at wavelengths from 254 to 434nm was measured. The spectrum consists of a major far-UV (below 320nm) component and a minor near-UV shoulder. A mutant deficient in DNA polymerase I accumulates breaks caused by near-UV (above 320nm) wavelengths faster than the wild-type strain proficient in polymerase I. Measurable breaks in extracted DNA are induced at a higher frequency than those induced in vivo. Anoxia, glycerol, and diazobicyclo (2.2.2.) octane inhibit break formation in extracted DNA. Alkali-labile bonds induced by 365-nm UV radiation are largely (78%) covalent bond chain breaks, the remainder consists of true alkali-labile bonds, probably apurinic and apyrimidinic sites. (author)

Chromosome breakage in peripheral lymphocytes of thorium workers

International Nuclear Information System (INIS)

Hoegerman, S.F.; Cummins, H.T.

1979-01-01

Cytogenic analysis of 21 thorium workers and 3 controls has not shown a significant elevation in the level of chromosome breakage in the workers' peripheral lymphocytes. The observation of a single dicentric chromosome in 100-cell samples from each of two workers with relatively long periods of occupational exposure and relatively high body burdens suggests, however, that such exposure might result in increases in chromosome aberration frequency

Review on Parameters Influencing the Rice Breakage and Rubber Roll Wear in Sheller

Directory of Open Access Journals (Sweden)

Prabhakaran P.

2017-09-01

Full Text Available The present review deals with parameters influencing the rice breakage during rice milling operations and the effect of rubber roll Sheller in rice husk removal process. The main objective of rice milling system is to remove the husk and bran layer to produce the white rice. In this process, rubber roll sheller is used to remove husk from the grains by friction process. If the rubber material is too soft, there may not be sufficient shear force to husk the paddy. Wear will be minimum for rubber material with high hardness but indeed it pronounce the breakage of rice. Hence, for efficient husking the rubber roll material should possess the balance of physico-mechanical properties. Rice breakage depends on several other parameters like the type of harvest, drying temperature, drying methods, physical characteristics of paddy, husking characteristics, paddy moisture content, rubber roller speed, rubber roll pressure, paddy feed rate and fissures. Rubber roll wear depends on the type of rubber material attached to the roller, feed rate, roller speed, pressure etc.

Video-supported analysis of Beggiatoa filament growth, breakage, and movement

DEFF Research Database (Denmark)

Kamp, Anja; Røy, Hans; Schulz-Vogt, Heide N.

2008-01-01

A marine Beggiatoa sp. was cultured in semi-solid agar with opposing oxygen-sulfide gradients. Growth pattern, breakage of filaments for multiplication, and movement directions of Beggiatoa filaments in the transparent agar were investigated by time-lapse video recording. The initial doubling time...

DNA double-strand breaks & poptosis in the testis

NARCIS (Netherlands)

Hamer, Geert

2003-01-01

During spermatogenesis, DNA damage is a naturally occurring event. At a certain stage, during the first meiotic prophase, DNA breaks are endogenously induced and even required for meiotic recombination. We studied these DNA breaks but also used ionizing radiation (IR) to induce DNA double-strand

Yield of radiation-induced DNA single-strand breaks in Escherichia coli and superinfecting phage lambda at different dose rates. Repair of strand breaks in different buffers

International Nuclear Information System (INIS)

Boye, E.; Johansen, I.; Brustad, T.

1976-01-01

Cells of E. coli K-12 strain AB 1886 were irradiated in oxygenated phosphate buffered saline at 2 0 C with electrons from a 4-MeV linear accelerator. The yield of DNA single-strand breaks was determined as a function of the dose rate between 2.5 and 21,000 krad/min. For dose rates over 100 krad/min the yield was found to be constant. Below 10 krad/min the yield of breaks decreases drastically. This is explained by rejoining of breaks during irradiation. Twenty percent of the breaks induced by acute exposure are repaired within 3 min at 2 0 C. Superinfecting phage lambda DNA is repaired at the same rate as chromosomal DNA. In contrast to the results obtained with phosphate-buffered saline, an increase in the number of breaks after irradiation is observed when the bacteria are suspended in tris buffer. It is suggested that buffers of low ionic strength facilitate the leakage through the membrane of a small-molecular-weight component(s) necessary for DNA strand rejoining

Determination of critical breakage conditions for double glazing in fire

International Nuclear Information System (INIS)

Wang, Yu; Li, Ke; Su, Yanfei; Lu, Wei; Wang, Qingsong; Sun, Jinhua; He, Linghui; Liew, K.M.

2017-01-01

Highlights: • Critical heat fluxes of exposed and ambient panes are 6 kW/m"2 and 25 kW/m"2. • Critical temperature difference of fire side pane is around 60 °C. • The ambient pane survives three times longer due to radiation filter and air gap. • Heat transfer in double glazing is revealed by a heat flux based theoretical model. - Abstract: Double glazing unit normally demonstrates better fire resistance than single glazing, but the knowledge on its thermal behavior and heat transfer mechanism during fire is limited. In this work, nine double glazing units were heated by a 500 × 500 mm"2 pool fire. The incident heat flux, temperature on four surfaces, breakage time and cracking behavior were obtained. The critical breakage conditions for interior and exterior panes were determined through gradually decreasing the glass-burner distance from 750 mm to 450 mm. It is established that in double glazing the pane at ambient side can withstand significantly more time than the pane exposed to fire. The critical temperature difference for interior pane is 60 °C; the critical temperature of exterior pane breakage is much higher due to no frame-covered area. In addition, the heat flux at the time of crack initiation is 6 kW/m"2 for the pane at fire side, while more than 25 kW/m"2 for ambient side pane. To reveal the heat transfer mechanism in glazing-air-glazing, theoretical and numerical investigations are also performed, which agrees well with the experimental results.

Needle breakage during an inferior alveolar nerve block in a child with KBG syndrome: A case report.

Science.gov (United States)

Bagattoni, S; D'Alessandro, G; Marzo, G; Piana, G

2018-04-01

Needle breakage during the administration of dental analgesia is an extremely rare event. A case of needle breakage during the administration of an inferior alveolar nerve block occurred in a child with KBG syndrome. During the injection, a sudden movement of the child caused the breakage of the needle. The next day, the retrieval of the needle was performed surgically under general analgesia. Three months after the surgery the healing was good. Two years later the child underwent a dental extraction with the aid of nitrous oxide/oxygen analgesia/anxiolysis. Needle fracture is a possible event during the administration of dental analgesia in children.

DNA double-strand breaks induced by high-energy neon and iron ions in human fibroblasts. I. Pulsed-field gel electrophoresis method

International Nuclear Information System (INIS)

Rydberg, B.; Loebrich, M.; Cooper, P.K.

1994-01-01

The relative effectiveness of high-energy neon and iron ions for the production of DNA double-strand breaks was measured in one transformed and one nontransformed human fibroblast cell line using pulsed-field gel electrophoresis. The DNA released from the gel plug (fraction of activity released: FAR) as well as the size distribution of the DNA entering the gel were used to compare the effects of the heavy-ion exposure with X-ray exposure. Both methods gave similar results, indicating similar distributions of breaks over megabase-pair distances for the heavy ions and the X rays. The relative biological effectiveness (RBE) compared to 225 kVp X rays of initially induced DNA double-strand breaks was found to be 0.85 for 425 MeV/u neon ions (LET 32 keV/μm) and 0.42-0.55 for 250-600 MeV/u iron ions (LET 190-350 keV/μm). Postirradiation incubation showed less efficient repair of breaks induced by the neon ions and the 600 MeV/u iron ions compared to X rays. Survival experiments demonstrated RBE values larger than one for cell killing by the heavy ions in parallel experiments (neon: RBE = 1.2, iron: RBE = 2.3-3.0, based on D 10 values). It is concluded that either the initial yield of DNA double-strand breaks induced by the high-energy particles is lower than the yield for X rays, or the breaks induced by heavy ions are present in clusters that cannot be resolved with the technique used. These results are confirmed in the accompanying paper. 48 refs., 5 figs., 2 tabs

Use of M-FISH analysis of α-particle-induced chromosome aberrations for the assessment of chromosomal breakpoint distribution and complex aberration formation

International Nuclear Information System (INIS)

Anderson, R.M.; Sumption, N.D.; Papworth, D.G.; Goodhead, D.T.

2003-01-01

Double strand breaks (dsb) of varying complexity are an important class of damage induced after exposure to ionising radiation and are considered to be the critical lesion for the formation of radiation-induced chromosome aberrations. Assuming the basic principles of the 'Breakage and Reunion' theory, dsb represent 'breakage' and aberrations are produced from the illegitimate repair (reunion) of the resulting dsb free-'ends'. Numerous questions relate to this process, in particular, (1) do chromosomal breakpoint 'hot-spots' that represent sensitive sites for breakage and/or regions of preferential repair/mis-repair, exist? (2) Considering that individual chromosomes and chromosome regions occupy discrete territories in the interphase nucleus, could rearrangements between specific chromosomes reflect domain organisation at the time of damage? (3) Assuming the topological constraints imposed on chromatin are not dramatically influenced by the presence of dsb, then how do multiple 'ends' from different chromosomes proximally associate for mis-repair as complex chromosome aberrations? To address these questions, we have analysed the chromosome aberrations induced in peripheral blood lymphocytes after exposure to 0.5 Gy α -particles (mean of 1 α -particle/cell) using the technique of M-FISH. This technique 'paints' all the human chromosomes (excluding homologues) uniquely, allowing chromosomal mis-repair to be visualised as differential colour-junctions and in addition, enhanced DAPI banding enables gross breakpoint assignation of these colour junctions. To test for non-randomness, we are comparing the frequency of occurrence of breakpoints obtained up to now with the F98 glioma model our knowledbased on chromosome length. Similarly, the involvement of each chromosome relative to other chromosomes within individual rearrangements can be determined by assuming the volume of chromosome domains is also proportional to their length. The current data to be presented will

ATR prohibits replication catastrophe by preventing global exhaustion of RPA.

Science.gov (United States)

Toledo, Luis Ignacio; Altmeyer, Matthias; Rask, Maj-Britt; Lukas, Claudia; Larsen, Dorthe Helena; Povlsen, Lou Klitgaard; Bekker-Jensen, Simon; Mailand, Niels; Bartek, Jiri; Lukas, Jiri

2013-11-21

ATR, activated by replication stress, protects replication forks locally and suppresses origin firing globally. Here, we show that these functions of ATR are mechanistically coupled. Although initially stable, stalled forks in ATR-deficient cells undergo nucleus-wide breakage after unscheduled origin firing generates an excess of single-stranded DNA that exhausts the nuclear pool of RPA. Partial reduction of RPA accelerated fork breakage, and forced elevation of RPA was sufficient to delay such "replication catastrophe" even in the absence of ATR activity. Conversely, unscheduled origin firing induced breakage of stalled forks even in cells with active ATR. Thus, ATR-mediated suppression of dormant origins shields active forks against irreversible breakage via preventing exhaustion of nuclear RPA. This study elucidates how replicating genomes avoid destabilizing DNA damage. Because cancer cells commonly feature intrinsically high replication stress, this study also provides a molecular rationale for their hypersensitivity to ATR inhibitors. Copyright © 2013 Elsevier Inc. All rights reserved.

A quantitative model of the major pathways for radiation-induced DNA double-strand break repair

International Nuclear Information System (INIS)

Belov, O.V.; Krasavin, E.A.; Lyashko, M.S.; Batmunkh, M.; Sweilam, N.H.

2014-01-01

We have developed a model approach to simulate the major pathways of DNA double-strand break (DSB) repair in mammalian and human cells. The proposed model shows a possible mechanistic explanation of the basic regularities of DSB processing through the nonhomologous end-joining (NHEJ), homologous recombination (HR), and single-strand annealing (SSA). It reconstructs the time-courses of radiation-induced foci specific to particular repair processes including the major intermediate stages. The model is validated for ionizing radiations of a wide range of linear energy transfer (0.2-236 keV/μm) including a relatively broad spectrum of heavy ions. The appropriate set of reaction rate constants was suggested to satisfy the kinetics of DSB rejoining for the considered types of exposure. The simultaneous assessment of three repair pathways allows one to describe their possible biological relations in response to radiation. With the help of the proposed approach, we reproduce several experimental data sets on γ-H2AX foci remaining in different types of cells including those defective in NHEJ, HR, or SSA functions.

Influence of the power law index on the fiber breakage during injection molding by numerical simulations

Science.gov (United States)

Desplentere, Frederik; Six, Wim; Bonte, Hilde; Debrabandere, Eric

2013-04-01

In predictive engineering for polymer processes, the proper prediction of material microstructure from known processing conditions and constituent material properties is a critical step forward properly predicting bulk properties in the finished composite. Operating within the context of long-fiber thermoplastics (LFT, length > 15mm) this investigation concentrates on the influence of the power law index on the final fiber length distribution within the injection molded part. To realize this, the Autodesk Simulation Moldflow Insight Scandium 2013 software has been used. In this software, a fiber breakage algorithm is available from this release on. Using virtual material data with realistic viscosity levels allows to separate the influence of the power law index on the fiber breakage from the other material and process parameters. Applying standard settings for the fiber breakage parameters results in an obvious influence on the fiber length distribution through the thickness of the part and also as function of position in the part. Finally, the influence of the shear rate constant within the fiber breakage model has been investigated illustrating the possibility to fit the virtual fiber length distribution to the possible experimentally available data.

Xrcc1-dependent and Ku-dependent DNA double-strand break repair kinetics in Arabidopsis plants.

Science.gov (United States)

Charbonnel, Cyril; Gallego, Maria E; White, Charles I

2010-10-01

Double-strand breakage (DSB) of DNA involves loss of information on the two strands of the DNA fibre and thus cannot be repaired by simple copying of the complementary strand which is possible with single-strand DNA damage. Homologous recombination (HR) can precisely repair DSB using another copy of the genome as template and non-homologous recombination (NHR) permits repair of DSB with little or no dependence on DNA sequence homology. In addition to the well-characterised Ku-dependent non-homologous end-joining (NHEJ) pathway, much recent attention has been focused on Ku-independent NHR. The complex interrelationships and regulation of NHR pathways remain poorly understood, even more so in the case of plants, and we present here an analysis of Ku-dependent and Ku-independent repair of DSB in Arabidopsis thaliana. We have characterised an Arabidopsis xrcc1 mutant and developed quantitative analysis of the kinetics of appearance and loss of γ-H2AX foci as a tool to measure DSB repair in dividing root tip cells of γ-irradiated plants in vivo. This approach has permitted determination of DSB repair kinetics in planta following a short pulse of γ-irradiation, establishing the existence of a Ku-independent, Xrcc1-dependent DSB repair pathway. Furthermore, our data show a role for Ku80 during the first minutes post-irradiation and that Xrcc1 also plays such a role, but only in the absence of Ku. The importance of Xrcc1 is, however, clearly visible at later times in the presence of Ku, showing that alternative end-joining plays an important role in DSB repair even in the presence of active NHEJ. © 2010 The Authors. Journal compilation © 2010 Blackwell Publishing Ltd.

Chromosomal aberration

International Nuclear Information System (INIS)

Ishii, Yutaka

1988-01-01

Chromosomal aberrations are classified into two types, chromosome-type and chromatid-type. Chromosom-type aberrations include terminal deletion, dicentric, ring and interstitial deletion, and chromatid-type aberrations include achromatic lesion, chromatid deletion, isochromatid deletion and chromatid exchange. Clastogens which induce chromosomal aberration are divided into ''S-dependent'' agents and ''S-independent''. It might mean whether they can induce double strand breaks independent of the S phase or not. Double strand breaks may be the ultimate lesions to induce chromosomal aberrations. Caffeine added even in the G 2 phase appeared to modify the frequency of chromatid aberrations induced by X-rays and mitomycin C. Those might suggest that the G 2 phase involves in the chromatid aberration formation. The double strand breaks might be repaired by ''G 2 repair system'', the error of which might yield breakage types of chromatid aberrations and the by-pass of which might yield chromatid exchanges. Chromosome-type aberrations might be formed in the G 1 phase. (author)

Sensitivity to chromosomal breakage as risk factor in young adults with oral squamous cell carcinoma.

Science.gov (United States)

Braakhuis, Boudewijn J M; Nieuwint, Aggie W M; Oostra, Anneke B; Joenje, Hans; Flach, Géke B; Graveland, A Peggy; Brakenhoff, Ruud H; Leemans, C René

2016-03-01

Oral squamous cell carcinoma (OSCC) may develop in young adults. In contrast to older patients, the well-known etiological factors, exposure to tobacco and alcohol, play a minor role in the carcinogenesis in this patient group. It has been suggested that an intrinsic susceptibility to environmental genotoxic exposures plays a role in the development of OSCC in these patients. The hypothesis was tested whether young OSCC patients have an increased sensitivity to induced chromosomal damage. Fourteen OSCC patients with an average age of 32 years (range 20-42) were selected. Peripheral blood lymphocytes and skin fibroblasts of patients and 14 healthy controls were subjected to the chromosome breakage test with Mitomycin C. This test is routinely used to identify Fanconi anemia patients, who are well-known for their inherited high sensitivity to this type of DNA damage, but also for the high risk to develop OSCC. Human papilloma virus status of the carcinomas was also determined. None of the 14 young patients with OSCC had an increased response in the MMC-chromosomal breakage test. All tumors tested negative for human papilloma virus. No evidence was obtained for the existence of a constitutional hypersensitivity to DNA chromosomal damage as a potential risk factor for OSCC in young adults. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

A new cable-in-conduit conductor magnet with insulated strands

International Nuclear Information System (INIS)

Yamaguchi, Satarou; Yamamoto, Junya; Motojima, Osamu.

1995-09-01

Many studies have used cable-in-conduit conductor (CICC) coils in trying to develop an AC superconducting magnet because of its enormous potential if AC losses were low and insulation voltage was high. The strands in the most recent CICC magnets are coated with chromium or another metal with high electrical resistance to order to induce current re-distribution among the strands and to avoid a quench caused by a current imbalance. Current re-distribution is highly complex and very difficult to analyze because the conditions of the strand surfaces and the contact areas vary greatly with the operation of the conductor. If, however, the cable currents were well-balanced, insulating the strands would be the best way to reduce AC losses. We propose a new CICC magnet structure featuring a current lead that balances the strand currents via its resistance. Having calculated current balances, we find that strand currents are well within the present parameters for nuclear fusion experiments and superconducting magnet energy storages. (author)

Effects of particle size and confining pressure on breakage factor of rockfill materials using medium triaxial test

Directory of Open Access Journals (Sweden)

Ashok Kumar Gupta

2016-06-01

Full Text Available Rockfill dams are mostly constructed using blasted rockfill materials obtained by blasting rocks or alluvial rockfill materials collected from the riverbeds. Behaviors of rockfill materials and their characterization significantly depend on breakage factor observed during triaxial loading. In this paper, two modeled rockfill materials are investigated by using medium triaxial cell. Drained triaxial tests are conducted on various sizes of modeled rockfill materials used in the two dams, and test data are analyzed accordingly. Breakage factor of rockfill material is studied and the effects of particle size and confining pressure on breakage factor are investigated using medium triaxial cell as many researchers have already conducted investigation using large triaxial cell.

Accelerated heavy ions induced DNA double-strand breaks in yeast cells

International Nuclear Information System (INIS)

Akpa, T.C.

1993-01-01

Yeast cells of strain cerevisiae, were irradiated with monoenergetic heavy ions, X-rays and α particles and assayed for DNA double-strand breaks and cell survival. The method of neutral sucrose gradient velocity sedimentation was used for all heavy-ion experiments because it is a well established technique.The method of pulsed-field gel electrophoresis was used for X-rays, α particles and argon ions. Results show that within the range of LET of the particles used (300 - 10 5 KeV/μm) the induction cross-section for DNA double-strand break is constant between 300 and around 7000 KeV/μm and increases at higher LET values. The inactivation cross-section follow the same trend. The DSB-induction and inactivation cross-section was shown to be linearly related with a slope of (1.01±0.15)/109 gmol-i. The RBE for DSB -induced decreases with LET and tails off at high LET values also. These results when compared with results from literature shows that the trend of induction is first monotonic rise of rate of DSB-induction up to 100keV/μm, followed by a plateau and a further rise which is due to increased effect of energetic γ-rays formed as shown for survival studies and predicted is possible to separate the cell DNA contents into 13 to 15 chromosome bands. The relative decrease in DNA content of the first band as determined by ethidium bromide-UV fluorescence decreases exponentially. The cross-section for DSB-induction determined by this method are (9.8±0.01)dsb/10 12 gmol - 1 Gy - 1, for 80 kV X-rays in haploid 211 yeast strain; (0.04+0.003)dsb/109gmol - 1μm 2 for Am-radioisotope α particles in haploid cells, (0.184±0.034) dsb/10 9 gmol - 1μm 2 in diploid 211*B cells and (0.55±0.04) dsb/10 9 gmol - 1μm 2 for 7MeV Argon ion in the diploid cells. The values are comparable to those obtained with velocity sedimentation technique. However, the reason for the low value obtained for a particle induced DSB in haploid cells is not clear

DNA damages induced by Ar F laser

Energy Technology Data Exchange (ETDEWEB)

Chapel, C.; Rose, S.; Chevrier, L.; Cordier, E.; Courant, D. [CEA Fontenay-aux-Roses, 92 (France). Dept. de Radiobiologie et de Radiopathologie

2006-07-01

The photo ablation process used in corneal refractive surgery by the Argon Fluoride (Ar F) laser emitting in ultraviolet C at 193 nm, exposes viable cells round the irradiated zone to sub ablative doses (< 400 joules.m -2). Despite that DNA absorption is higher at 193 nm than 254 nm, cytotoxicity of 193 nm laser radiation is lower than radiation emitted by 254 nm UV-C lamps. In situ, DNA could be protected of laser radiation by cellular components. Consequently, some authors consider that this radiation does not induce genotoxic effect whereas others suspect it to be mutagenic. These lasers are used for fifteen years but many questions remain concerning the long term effects on adjacent cells to irradiated area. The purpose of this study is to describe the effect of 193 nm laser radiation on DNA of stromal keratocytes which are responsible of the corneal structure. The 193 nm laser irradiation induces directly DNA breakage in keratocytes as it has been shown by the comet assay under alkaline conditions. Two hours post irradiation, damages caused by the highest exposure (150 J.m-2) are not repaired as it has been measured with the Olive Tail Moment (product of tail length and tail DNA content). They give partly evidence of induction of an apoptotic process in cells where DNA could be too damaged. In order to characterize specifically double strand breaks, a comparative analysis by immunofluorescence of the H2 Ax histone phosphorylation (H2 Ax) has been performed on irradiated keratocytes and unirradiated keratocytes. Results show a dose dependent increase of the number of H2 Ax positive cells. Consequences of unrepaired DNA lesions could be observed by the generation of micronuclei in cells. Results show again an increase of micronuclei in laser irradiated cells. Chromosomal aberrations have been pointed out by cytogenetic methods 30 mn after irradiation. These aberrations are dose dependent (from 10 to 150 J.m-2). The number of breakage decreases in the long run

Diagnostic Overlap between Fanconi Anemia and the Cohesinopathies: Roberts Syndrome and Warsaw Breakage Syndrome

Directory of Open Access Journals (Sweden)

Petra van der Lelij

2010-01-01

Full Text Available Fanconi anemia (FA is a recessively inherited disease characterized by multiple symptoms including growth retardation, skeletal abnormalities, and bone marrow failure. The FA diagnosis is complicated due to the fact that the clinical manifestations are both diverse and variable. A chromosomal breakage test using a DNA cross-linking agent, in which cells from an FA patient typically exhibit an extraordinarily sensitive response, has been considered the gold standard for the ultimate diagnosis of FA. In the majority of FA patients the test results are unambiguous, although in some cases the presence of hematopoietic mosaicism may complicate interpretation of the data. However, some diagnostic overlap with other syndromes has previously been noted in cases with Nijmegen breakage syndrome. Here we present results showing that misdiagnosis may also occur with patients suffering from two of the three currently known cohesinopathies, that is, Roberts syndrome (RBS and Warsaw breakage syndrome (WABS. This complication may be avoided by scoring metaphase chromosomes—in addition to chromosomal breakage—for spontaneously occurring premature centromere division, which is characteristic for RBS and WABS, but not for FA.

Destabilized SMC5/6 complex leads to chromosome breakage syndrome with severe lung disease

Science.gov (United States)

van der Crabben, Saskia N.; Hennus, Marije P.; McGregor, Grant A.; Ritter, Deborah I.; Nagamani, Sandesh C.S.; Wells, Owen S.; Harakalova, Magdalena; Chinn, Ivan K.; Alt, Aaron; Vondrova, Lucie; Hochstenbach, Ron; van Montfrans, Joris M.; Terheggen-Lagro, Suzanne W.; van Lieshout, Stef; van Roosmalen, Markus J.; Renkens, Ivo; Duran, Karen; Nijman, Isaac J.; Kloosterman, Wigard P.; Hennekam, Eric; van Hasselt, Peter M.; Wheeler, David A.; Palecek, Jan J.; Lehmann, Alan R.; Oliver, Antony W.; Pearl, Laurence H.; Plon, Sharon E.; Murray, Johanne M.

2016-01-01

The structural maintenance of chromosomes (SMC) family of proteins supports mitotic proliferation, meiosis, and DNA repair to control genomic stability. Impairments in chromosome maintenance are linked to rare chromosome breakage disorders. Here, we have identified a chromosome breakage syndrome associated with severe lung disease in early childhood. Four children from two unrelated kindreds died of severe pulmonary disease during infancy following viral pneumonia with evidence of combined T and B cell immunodeficiency. Whole exome sequencing revealed biallelic missense mutations in the NSMCE3 (also known as NDNL2) gene, which encodes a subunit of the SMC5/6 complex that is essential for DNA damage response and chromosome segregation. The NSMCE3 mutations disrupted interactions within the SMC5/6 complex, leading to destabilization of the complex. Patient cells showed chromosome rearrangements, micronuclei, sensitivity to replication stress and DNA damage, and defective homologous recombination. This work associates missense mutations in NSMCE3 with an autosomal recessive chromosome breakage syndrome that leads to defective T and B cell function and acute respiratory distress syndrome in early childhood. PMID:27427983

A Critique of a Phenomenological Fiber Breakage Model for Stress Rupture of Composite Materials

Science.gov (United States)

Reeder, James R.

2010-01-01

Stress rupture is not a critical failure mode for most composite structures, but there are a few applications where it can be critical. One application where stress rupture can be a critical design issue is in Composite Overwrapped Pressure Vessels (COPV's), where the composite material is highly and uniformly loaded for long periods of time and where very high reliability is required. COPV's are normally required to be proof loaded before being put into service to insure strength, but it is feared that the proof load may cause damage that reduces the stress rupture reliability. Recently, a fiber breakage model was proposed specifically to estimate a reduced reliability due to proof loading. The fiber breakage model attempts to model physics believed to occur at the microscopic scale, but validation of the model has not occurred. In this paper, the fiber breakage model is re-derived while highlighting assumptions that were made during the derivation. Some of the assumptions are examined to assess their effect on the final predicted reliability.

Study of the effects of radiation of nucleic acids and related compounds. Progress report, August 15, 1975--August 14, 1976

International Nuclear Information System (INIS)

Wang, S.Y.

1976-04-01

Ionizing radiation produces genetic effects in biological systems. Since genetic effects are usually the result of modifications of DNA or sometimes of RNA, interest is being centered on the chemical and physical nature of radiation-induced lesions to nucleic acids and their components. These investigations have revealed the enormous complexity of chemical events and the possible degradation of nucleic acids by strand breakage. Therefore, work in the ionization radiation of nucleic acids has proceeded along a dual course. On the one hand, molecular changes have been characterized for a number of primary radiation products. On the other hand, strand breakage has been investigated intensively as a direct primary event. Both of these aspects were emphasized in our research last year. We succeeded in improving the synthesis of 5-hydroxy-methyl thymine (α-TOOH). α-TOOH was found to be much more effective than cis-5,6-dihydro-6-hydroperoxy-5-hydroxy thymine (6-TOOH) in the inactivation of transforming DNA of H. influenzae cells although α-TOOH is much less reactive chemically than 6-TOOH. 6-TOOH causes inactivation and acts as an inhibitor of DNA synthesis in mammalian cells. In addition, evidence may indicate that 6-TOOH does not induce strand breaks directly in DNA although we showed that 6-TOOH is a clastogenic compound

Protection for low current superconducting coils wound with insulated strand cable

International Nuclear Information System (INIS)

Satti, J.

1980-09-01

The insulated strand cable concept for winding of low current superconducting coil leads to an ideal quench protection by induction coupling. A superconducting secondary loop was made within a cable of a 6.2 Henry dipole coil. When quenching occurred, current was induced in the secondary strand above the critical value. The normal strand quenched the whole cable due to good thermal contact. The secondary loop works as a heater turned on as the wire becomes normal throughout the coil. With a well spread quench, the energy dissipation density is decreased thus preventing local burnout. The mechanism is possible because of close coupling that is present in the insulated cable as in bifilar winding. For the coil tested a 12 strand cable was used, thus a favorable 11 to 1 turn ratio was obtained for the primary to secondary. The superconductor in the secondary had a lower resistance until the critical current was achieved. A theoretical explanation is described for a simplified circuit. Test on the dipole coil with four individual shells showed that the one shell protected with the induced coupling heater always had a more rapid reduction of current. The induced coupling heater tested and explained in this paper works automatically and does not rely on mechanical or electrical devices

Chromosomes of older humans are more prone to aminopterine-induced breakage

International Nuclear Information System (INIS)

Esposito, D.; Fassina, G.; Szabo, P.; Weksler, M.; De Angelis, P.; Siniscalco, M.; Rodgers, L.

1989-01-01

The authors have adopted a simplified version of the cell hybrid cotransfer method to test the hypothesis that human lymphocytes derived from elderly individuals have a higher chromosome instability. Peripheral blood lymphocytes from old male individuals and young controls were fused with a Chinese hamster cell line (CHO-YH21), yielding 10 HAT-resistant rodent-human clones from the old propositi and 22 from the young controls. Both series of hybrid clones were analyzed with respect to the retention of the enzyme glucose-6-phosphate dehydrogenase and the surface antigen MIC2 identified by monoclonal antibody 12E7, two human X chromosome-linked markers located at opposite ends of the X chromosome. Cell hybrid clones with an X chromosome from a young control retained both markers in about 70% of the cells. In contrast, cell hybrid clones with an X chromosome from an old donor retained the MIC2 marker in only 30% of their cells. Slot-blot hybridization studies have established that the observed loss of the MIC2 marker is due to loss of the coding gene, not to suppression of its expression. T lymphocytes from old donors were also found to have an LD 50 for aminopterine significantly lower than the concentration of this drug in the HAT medium used to grow the hybrids. They speculate that the higher rate of chromosomal breakage and of marker loss observed along the old-age X chromosomes could be the result of molecular scars accumulated with aging at sites of constitutive chromosomal fragility

Timely Endocytosis of Cytokinetic Enzymes Prevents Premature Spindle Breakage during Mitotic Exit.

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Cheen Fei Chin

2016-07-01

Full Text Available Cytokinesis requires the spatio-temporal coordination of membrane deposition and primary septum (PS formation at the division site to drive acto-myosin ring (AMR constriction. It has been demonstrated that AMR constriction invariably occurs only after the mitotic spindle disassembly. It has also been established that Chitin Synthase II (Chs2p neck localization precedes mitotic spindle disassembly during mitotic exit. As AMR constriction depends upon PS formation, the question arises as to how chitin deposition is regulated so as to prevent premature AMR constriction and mitotic spindle breakage. In this study, we propose that cells regulate the coordination between spindle disassembly and AMR constriction via timely endocytosis of cytokinetic enzymes, Chs2p, Chs3p, and Fks1p. Inhibition of endocytosis leads to over accumulation of cytokinetic enzymes during mitotic exit, which accelerates the constriction of the AMR, and causes spindle breakage that eventually could contribute to monopolar spindle formation in the subsequent round of cell division. Intriguingly, the mitotic spindle breakage observed in endocytosis mutants can be rescued either by deleting or inhibiting the activities of, CHS2, CHS3 and FKS1, which are involved in septum formation. The findings from our study highlight the importance of timely endocytosis of cytokinetic enzymes at the division site in safeguarding mitotic spindle integrity during mitotic exit.

A theoretical investigation of the feasibility of Tannor-Rice type control: Application to selective bond breakage in gas-phase dihalomethanes

DEFF Research Database (Denmark)

Shu, Chuan-Cun; Rozgonyi, Tamas; Gonzalez, Leticia

2012-01-01

Within the math absorption band of CH2BrCl, we theoretically analyze the laser-induced control of the Br/Cl branching ratio, Br + CH2Cl ← CH2BrCl → CH2Br + Cl, with CH2BrCl initially in its vibrational ground state. For weak-field excitation, the Br/Cl branching ratio increases as a function of w......-Rice type laser control mechanism for selective bond breakage in CH2BrCl cannot take place without accompanying photoionization....

Study on DNA damages induced by UV radiation

International Nuclear Information System (INIS)

Doan Hong Van; Dinh Ba Tuan; Tran Tuan Anh; Nguyen Thuy Ngan; Ta Bich Thuan; Vo Thi Thuong Lan; Tran Minh Quynh; Nguyen Thi Thom

2015-01-01

DNA damages in Escherichia coli (E. coli) exposed to UV radiation have been investigated. After 30 min of exposure to UV radiation of 5 mJ/cm"2, the growth of E. coli in LB broth medium was about only 10% in compared with non-irradiated one. This results suggested that the UV radiation caused the damages for E. coli genome resulted in reduction in its growth and survival, and those lesions can be somewhat recovered. For both solutions of plasmid DNAs and E. coli cells containing plasmid DNA, this dose also caused the breakage on single and double strands of DNA, shifted the morphology of DNA plasmid from supercoiled to circular and linear forms. The formation of pyrimidine dimers upon UV radiation significantly reduced when the DNA was irradiated in the presence of Ganoderma lucidum extract. Thus, studies on UV-induced DNA damage at molecular level are very essential to determine the UV radiation doses corresponding to the DNA damages, especially for creation and selection of useful radiation-induced mutants, as well as elucidation the protective effects of the specific compounds against UV light. (author)

Early models of DNA damage formation

International Nuclear Information System (INIS)

Śmiałek, Małgorzata A

2012-01-01

Quantification of DNA damage, induced by various types of incident radiation as well as chemical agents, has been the subject of many theoretical and experimental studies, supporting the development of modern cancer therapy. The primary observations showed that many factors can lead to damage of DNA molecules. It became clear that the development of experimental techniques for exploring this phenomenon is required. Another problem was simultaneously dealt with, anticipating on how the damage is distributed within the double helix of the DNA molecule and how the single strand break formation and accumulation can influence the lethal double strand break formation. In this work the most important probabilistic models for DNA strand breakage and damage propagation are summarized and compared.

Monte Carlo simulation of ionizing radiation induced DNA strand breaks utilizing coarse grained high-order chromatin structures.

Science.gov (United States)

Liang, Ying; Yang, Gen; Liu, Feng; Wang, Yugang

2016-01-07

Ionizing radiation threatens genome integrity by causing DNA damage. Monte Carlo simulation of the interaction of a radiation track structure with DNA provides a powerful tool for investigating the mechanisms of the biological effects. However, the more or less oversimplification of the indirect effect and the inadequate consideration of high-order chromatin structures in current models usually results in discrepancies between simulations and experiments, which undermine the predictive role of the models. Here we present a biophysical model taking into consideration factors that influence indirect effect to simulate radiation-induced DNA strand breaks in eukaryotic cells with high-order chromatin structures. The calculated yields of single-strand breaks and double-strand breaks (DSBs) for photons are in good agreement with the experimental measurements. The calculated yields of DSB for protons and α particles are consistent with simulations by the PARTRAC code, whereas an overestimation is seen compared with the experimental results. The simulated fragment size distributions for (60)Co γ irradiation and α particle irradiation are compared with the measurements accordingly. The excellent agreement with (60)Co irradiation validates our model in simulating photon irradiation. The general agreement found in α particle irradiation encourages model applicability in the high linear energy transfer range. Moreover, we demonstrate the importance of chromatin high-order structures in shaping the spectrum of initial damage.

Monte Carlo simulation of ionizing radiation induced DNA strand breaks utilizing coarse grained high-order chromatin structures

International Nuclear Information System (INIS)

Liang, Ying; Yang, Gen; Liu, Feng; Wang, Yugang

2016-01-01

Ionizing radiation threatens genome integrity by causing DNA damage. Monte Carlo simulation of the interaction of a radiation track structure with DNA provides a powerful tool for investigating the mechanisms of the biological effects. However, the more or less oversimplification of the indirect effect and the inadequate consideration of high-order chromatin structures in current models usually results in discrepancies between simulations and experiments, which undermine the predictive role of the models. Here we present a biophysical model taking into consideration factors that influence indirect effect to simulate radiation-induced DNA strand breaks in eukaryotic cells with high-order chromatin structures. The calculated yields of single-strand breaks and double-strand breaks (DSBs) for photons are in good agreement with the experimental measurements. The calculated yields of DSB for protons and α particles are consistent with simulations by the PARTRAC code, whereas an overestimation is seen compared with the experimental results. The simulated fragment size distributions for 60 Co γ irradiation and α particle irradiation are compared with the measurements accordingly. The excellent agreement with 60 Co irradiation validates our model in simulating photon irradiation. The general agreement found in α particle irradiation encourages model applicability in the high linear energy transfer range. Moreover, we demonstrate the importance of chromatin high-order structures in shaping the spectrum of initial damage. (paper)

Tree species traits but not diversity mitigate stem breakage in a subtropical forest following a rare and extreme ice storm.

Directory of Open Access Journals (Sweden)

Karin Nadrowski

Full Text Available Future climates are likely to include extreme events, which in turn have great impacts on ecological systems. In this study, we investigated possible effects that could mitigate stem breakage caused by a rare and extreme ice storm in a Chinese subtropical forest across a gradient of forest diversity. We used Bayesian modeling to correct stem breakage for tree size and variance components analysis to quantify the influence of taxon, leaf and wood functional traits, and stand level properties on the probability of stem breakage. We show that the taxon explained four times more variance in individual stem breakage than did stand level properties; trees with higher specific leaf area (SLA were less susceptible to breakage. However, a large part of the variation at the taxon scale remained unexplained, implying that unmeasured or undefined traits could be used to predict damage caused by ice storms. When aggregated at the plot level, functional diversity and wood density increased after the ice storm. We suggest that for the adaption of forest management to climate change, much can still be learned from looking at functional traits at the taxon level.

Poly(ADP-ribose polymerase (PARP-1 is not involved in DNA double-strand break recovery

Directory of Open Access Journals (Sweden)

Fernet Marie

2003-07-01

Full Text Available Abstract Background The cytotoxicity and the rejoining of DNA double-strand breaks induced by γ-rays, H2O2 and neocarzinostatin, were investigated in normal and PARP-1 knockout mouse 3T3 fibroblasts to determine the role of poly(ADP-ribose polymerase (PARP-1 in DNA double-strand break repair. Results PARP-1-/- were considerably more sensitive than PARP-1+/+ 3T3s to induced cell kill by γ-rays and H2O2. However, the two cell lines did not show any significant difference in the susceptibility to neocarzinostatin below 1.5 nM drug. Restoration of PARP-1 expression in PARP-1-/- 3T3s by retroviral transfection of the full PARP-1 cDNA did not induce any change in neocarzinostatin response. Moreover the incidence and the rejoining kinetics of neocarzinostatin-induced DNA double-strand breaks were identical in PARP-1+/+ and PARP-1-/- 3T3s. Poly(ADP-ribose synthesis following γ-rays and H2O2 was observed in PARP-1-proficient cells only. In contrast neocarzinostatin, even at supra-lethal concentration, was unable to initiate PARP-1 activation yet it induced H2AX histone phosphorylation in both PARP1+/+ and PARP-1-/- 3T3s as efficiently as γ-rays and H2O2. Conclusions The results show that PARP-1 is not a major determinant of DNA double-strand break recovery with either strand break rejoining or cell survival as an endpoint. Even though both PARP-1 and ATM activation are major determinants of the cell response to γ-rays and H2O2, data suggest that PARP-1-dependent poly(ADP-ribose synthesis and ATM-dependent H2AX phosphorylation, are not inter-related in the repair pathway of neocarzinostatin-induced DNA double-strand breaks.

Misrepair of overlapping daughter strand gaps as a possible mechanism for UV induced mutagenesis in uvr strains of Escherichia coli: a general model for induced mutagenesis by misrepair (SOS repair) of closely spaced DNA lesions

International Nuclear Information System (INIS)

Sedgwick, S.G.

1976-01-01

It has been previously reported that an inducible form of post-replication repair appeared to be required for UV induced mutagenesis in an uvrA strain of Escherichia coli. It is shown here that the numbers of daughter strand gaps requiring inducible repair were similar to the numbers calculated to be overlapping one another in opposite daughter chromosomes. An estimation of survival with no repair of these gaps resembled the survival predicted with mutagenesis. It is thus proposed that inducible post-replication repair causes mutagenesis by the repair of overlapping daughter strand gaps. A general model for induced mutagenesis is presented. It is proposed that (a) some DNA lesions introduced by any DNA damaging agent may be close enough to interfere with constitutive repair replication of each other, (b) these lesions induce a repair system (SOS repair) which involves the recA + . lexA + and polC + genes (c) repair, and noncomitant mutagenesis occurs during repair replication by the insertion of mismatched bases oppposite the noncoding DNA lesions

Mobilization of Copper ions by Flavonoids in Human Peripheral Lymphocytes Leads to Oxidative DNA Breakage: A Structure Activity Study

Science.gov (United States)

Arif, Hussain; Rehmani, Nida; Farhan, Mohd; Ahmad, Aamir; Hadi, Sheikh Mumtaz

2015-01-01

Epidemiological studies have linked dietary consumption of plant polyphenols with lower incidence of various cancers. In particular, flavonoids (present in onion, tomato and other plant sources) induce apoptosis and cytotoxicity in cancer cells. These can therefore be used as lead compounds for the synthesis of novel anticancer drugs with greater bioavailability. In the present study, we examined the chemical basis of cytotoxicity of flavonoids by studying the structure–activity relationship of myricetin (MN), fisetin (FN), quercetin (QN), kaempferol (KL) and galangin (GN). Using single cell alkaline gel electrophoresis (comet assay), we established the relative efficiency of cellular DNA breakage as MN > FN > QN > KL > GN. Also, we determined that the cellular DNA breakage was the result of mobilization of chromatin-bound copper ions and the generation of reactive oxygen species. The relative DNA binding affinity order was further confirmed using molecular docking and thermodynamic studies through the interaction of flavonoids with calf thymus DNA. Our results suggest that novel anti-cancer molecules should have ortho-dihydroxy groups in B-ring and hydroxyl groups at positions 3 and 5 in the A-ring system. Additional hydroxyl groups at other positions further enhance the cellular cytotoxicity of the flavonoids. PMID:26569217

Radiation induced DNA double-strand breaks in radiology; Strahleninduzierte DNA-Doppelstrangbrueche in der Radiologie

Energy Technology Data Exchange (ETDEWEB)

Kuefner, M.A. [Dornbirn Hospital (Austria). Dept. of Radiology; Brand, M.; Engert, C.; Uder, M. [Erlangen University Hospital (Germany). Dept. of Radiology; Schwab, S.A. [Radiologis, Oberasbach (Germany)

2015-10-15

Shortly after the discovery of X-rays, their damaging effect on biological tissues was observed. The determination of radiation exposure in diagnostic and interventional radiology is usually based on physical measurements or mathematical algorithms with standardized dose simulations. γ-H2AX immunofluorescence microscopy is a reliable and sensitive method for the quantification of radiation induced DNA double-strand breaks (DSB) in blood lymphocytes. The detectable amount of these DNA damages correlates well with the dose received. However, the biological radiation damage depends not only on dose but also on other individual factors like radiation sensitivity and DNA repair capacity. Iodinated contrast agents can enhance the x-ray induced DNA damage level. After their induction DSB are quickly repaired. A protective effect of antioxidants has been postulated in experimental studies. This review explains the principle of the γ-H2AX technique and provides an overview on studies evaluating DSB in radiologic examinations.

Detection of heavy ion induced DNA double-strand breaks using static-field gel electrophoresis

International Nuclear Information System (INIS)

Taucher-Scholz, G.; Heilmann, J.; Schneider, G.; Kraft, G.

1994-11-01

Radiation induced DNA double-strand breaks (DSBs) were measured in Chinese hamster ovary cells (CHO-K1) using an experimental protocol involving static-field gel electrophoresis following exposure to various accelerated ions. Dose-effect curves were set up and relative biological efficiencies (RBEs) for DSB induction were determined for different radiation qualities. RBEs around 1 were obtained for low energy deuterons (6-7 keV/μm), while for high energy oxygen ions (20 keV/μm) an RBE value slightly greater than 1 was determined. Low energetic oxygen ions (LET ∼ 250 keV/μm) were found to show RBEs substantially below unity, and for higher LET particles (≥ 250 keV/μm) RBEs for DSB induction were generally found to be smaller than 1. The data presented here are in line with the generally accepted view that not induced DSBs, but misrepaired or unrepaired DNA-lesions are related to cellular inactivation. (orig.)

Some aspects of the mechanism of bacteriophage function. Final progress report

International Nuclear Information System (INIS)

Freifelder, D.

1977-01-01

Data are summarized from a ten-year study on the radiobiology of phages. The results showed that: phages are inactivated principally by damage to DNA; DNA damage is of two types, base damage and double-strand breakage; double-strand breakage may be lethal because of interruption within a gene, however in phage systems the damage is more fundamental in that only a single DNA fragment is injected into the host; E. coli phage T4 is relatively resistant to inactivation by x-rays; and the rate of production of strand breaks and base damage is nearly the same in bacteriophage and bacteria

Thymocyte apoptosis induced by p53-dependent and independent pathways

International Nuclear Information System (INIS)

Clarke, A.R.; Purdie, C.A.; Harrison, D.J.; Morris, R.G.; Bird, C.C.; Hooper, M.L.; Wyllie, A.H.

1993-01-01

The authors studied the dependence of apoptosis on p53 expression in cells from the thymus cortex. Short-term thymocyte cultures were prepared from mice constitutively heterozygous or homozygous for a deletion in the p53 gene introduced into the germ line after gene targeting. Wild-type thymocytes readily undergo apoptosis after treatment with ionizing radiation, the glucocorticoid methylprednisolone, or etoposide (an inhibitor of topoisomerase II), or after Ca 2+ -dependent activation by phorbol ester and a calcium ionophore. In contrast, homozygous null p53 thymocytes are resistant to induction of apoptosis by radiation or etoposide, but retain normal sensitivity to glucocorticoid and calcium. The time-dependent apoptosis that occurs in untreated cultures is unaffected by p53 status. Cells heterozygous for p53 deletion are partially resistant to radiation and etoposide. Results show that p53 exerts a significant and dose-dependent effect in the initiation of apoptosis, but only when it is induced by agents that cause DNA-strand breakage. (Author)

Modification of the radiation sensitivity of human tumour cells by a bis-benzimidazole derivative

Energy Technology Data Exchange (ETDEWEB)

Smith, P J; Anderson, C O [Medical Research Council, Cambridge (UK)

1984-10-01

A comparison was made of the ability of either X-radiation or a DNA-specific ligand (the vital bis-benzimidazole dye; Hoechst 33342) to induce: cell killing, inhibition of de novo DNA synthesis, DNA strand breakage and the delay of cell division in human colon adenocarcinoma cells in vitro. Unlike radiation-induced cell killing, ligand-induced cytotoxicity appeared to be positively correlated with the extent of inhibition of de novo DNA synthesis-a feature consistent with the persistent binding of ligand molecules to nuclear DNA. Ligand-induced DNA strand-breaks disappeared slowly although ligand-treated cells retained apparently normal capacities to repair discrete radiogenic DNA strand-breaks. Pre-treatment of cells with Hoechst 33342 resulted in a dose-modifying enhancement of radiation resistance not associated with altered dosimetry for strand-break induction. However, radioresistance was accompanied by the protracted retention of cells in the G/sub 2/ phase of the cell cycle. We suggest that the results provide direct evidence that the retention of cells in G/sub 2/ phase is a sparing phenomenon and is triggered by the responses of chromatin domains to the presence of DNA damage.

Application of Laser Micro-irradiation for Examination of Single and Double Strand Break Repair in Mammalian Cells.

Science.gov (United States)

Holton, Nathaniel W; Andrews, Joel F; Gassman, Natalie R

2017-09-05

Highly coordinated DNA repair pathways exist to detect, excise and replace damaged DNA bases, and coordinate repair of DNA strand breaks. While molecular biology techniques have clarified structure, enzymatic functions, and kinetics of repair proteins, there is still a need to understand how repair is coordinated within the nucleus. Laser micro-irradiation offers a powerful tool for inducing DNA damage and monitoring the recruitment of repair proteins. Induction of DNA damage by laser micro-irradiation can occur with a range of wavelengths, and users can reliably induce single strand breaks, base lesions and double strand breaks with a range of doses. Here, laser micro-irradiation is used to examine repair of single and double strand breaks induced by two common confocal laser wavelengths, 355 nm and 405 nm. Further, proper characterization of the applied laser dose for inducing specific damage mixtures is described, so users can reproducibly perform laser micro-irradiation data acquisition and analysis.

On the linearity of the dose-effect relationship of DNA double strand breaks

International Nuclear Information System (INIS)

Chadwick, K.H.; Leenhouts, H.P.

1994-01-01

Most radiation biologists believe that DNA double-strand breaks are induced linearly with radiation dose for all types of radiation. Since 1985, with the advent of elution and gel electrophoresis techniques which permit the measurement of DNA double-strand breaks induced in mammalian cells at doses having radiobiological relevance, the true nature of the dose-effect relationship has been brought into some doubt. Many investigators measured curvilinear dose-effect relationships and a few found good correlations between the induction of the DNA double-strand breaks and cell survival. We approach the problem pragmatically by assuming that the induction of DNA double-strand breaks by 125 I Auger electron emitters incorporated into the DNA of the cells is a linear function of the number of 125 I decays, and by comparing the dose-effect relationship for sparsely ionizing radiation against this standard. The conclusion drawn that the curvilinear dose-effect relationships and the correlations with survival are real. (Author)

A switch from high-fidelity to error-prone DNA double-strand break repair underlies stress-induced mutation.

Science.gov (United States)

Ponder, Rebecca G; Fonville, Natalie C; Rosenberg, Susan M

2005-09-16

Special mechanisms of mutation are induced in microbes under growth-limiting stress causing genetic instability, including occasional adaptive mutations that may speed evolution. Both the mutation mechanisms and their control by stress have remained elusive. We provide evidence that the molecular basis for stress-induced mutagenesis in an E. coli model is error-prone DNA double-strand break repair (DSBR). I-SceI-endonuclease-induced DSBs strongly activate stress-induced mutations near the DSB, but not globally. The same proteins are required as for cells without induced DSBs: DSBR proteins, DinB-error-prone polymerase, and the RpoS starvation-stress-response regulator. Mutation is promoted by homology between cut and uncut DNA molecules, supporting a homology-mediated DSBR mechanism. DSBs also promote gene amplification. Finally, DSBs activate mutation only during stationary phase/starvation but will during exponential growth if RpoS is expressed. Our findings reveal an RpoS-controlled switch from high-fidelity to mutagenic DSBR under stress. This limits genetic instability both in time and to localized genome regions, potentially important evolutionary strategies.

Flap Endonuclease 1 Limits Telomere Fragility on the Leading Strand*

Science.gov (United States)

Teasley, Daniel C.; Parajuli, Shankar; Nguyen, Mai; Moore, Hayley R.; Alspach, Elise; Lock, Ying Jie; Honaker, Yuchi; Saharia, Abhishek; Piwnica-Worms, Helen; Stewart, Sheila A.

2015-01-01

The existence of redundant replication and repair systems that ensure genome stability underscores the importance of faithful DNA replication. Nowhere is this complexity more evident than in challenging DNA templates, including highly repetitive or transcribed sequences. Here, we demonstrate that flap endonuclease 1 (FEN1), a canonical lagging strand DNA replication protein, is required for normal, complete leading strand replication at telomeres. We find that the loss of FEN1 nuclease activity, but not DNA repair activities, results in leading strand-specific telomere fragility. Furthermore, we show that FEN1 depletion-induced telomere fragility is increased by RNA polymerase II inhibition and is rescued by ectopic RNase H1 expression. These data suggest that FEN1 limits leading strand-specific telomere fragility by processing RNA:DNA hybrid/flap intermediates that arise from co-directional collisions occurring between the replisome and RNA polymerase. Our data reveal the first molecular mechanism for leading strand-specific telomere fragility and the first known role for FEN1 in leading strand DNA replication. Because FEN1 mutations have been identified in human cancers, our findings raise the possibility that unresolved RNA:DNA hybrid structures contribute to the genomic instability associated with cancer. PMID:25922071

A Stress Measurement Method for Steel Strands Based on LC Oscillation

Directory of Open Access Journals (Sweden)

Dongjun Chen

2018-01-01

Full Text Available The prestress loss is one of the main factors affecting the safety of prestressed concrete structure. While the detecting signals like sound and light are difficult to spread in steel strands, there is no effective method for prestress detection of the bonded prestressed steel strands in existing structures yet. In this paper, taking into consideration that the electromagnetic oscillation characteristic can make the signal propagate effectively on the bonded prestressed steel strands, a nondestructive prestress detection method based on the electromagnetic effect to detect oscillation frequency is proposed. In a detection circuit, the steel strands are simulated as an inductance component, in which an induced electromagnetic signal passes through the steel strands to form resonance. And then, a frequency meter is used to detect the oscillation frequency of the resonant circuit. The oscillation frequency is supposed to have relationship with the prestress loading on the steel strands. A section of steel strands with a length of 1.2 m is adopted to test the correlation of stress and oscillation frequency. Both the theoretical and experimental results show that the resonant frequency of the circuit decreases with the increase of the stress of the strand and is linear in a certain range.

DNA strand breaks induced by soft X-ray pulses from a compact laser plasma source

Science.gov (United States)

Adjei, Daniel; Wiechec, Anna; Wachulak, Przemyslaw; Ayele, Mesfin Getachew; Lekki, Janusz; Kwiatek, Wojciech M.; Bartnik, Andrzej; Davídková, Marie; Vyšín, Luděk; Juha, Libor; Pina, Ladislav; Fiedorowicz, Henryk

2016-03-01

Application of a compact laser plasma source of soft X-rays in radiobiology studies is demonstrated. The source is based on a laser produced plasma as a result of irradiation of a double-stream gas puff target with nanosecond laser pulses from a commercially available Nd:YAG laser. The source allows irradiation of samples with soft X-ray pulses in the "water window" spectral range (wavelength: 2.3-4.4 nm; photon energy: 280-560 eV) in vacuum or a helium atmosphere at very high-dose rates and doses exceeding the kGy level. Single-strand breaks (SSB) and double-strand breaks (DBS) induced in DNA plasmids pBR322 and pUC19 have been measured. The different conformations of the plasmid DNA were separated by agarose gel electrophoresis. An exponential decrease in the supercoiled form with an increase in linear and relaxed forms of the plasmids has been observed as a function of increasing photon fluence. Significant difference between SSB and DSB in case of wet and dry samples was observed that is connected with the production of free radicals in the wet sample by soft X-ray photons and subsequent affecting the plasmid DNA. Therefore, the new source was validated to be useful for radiobiology experiments.

Macromolecule oxidation and DNA repair in mussel (Mytilus edulis L.) gill following exposure to Cd and Cr(VI)

International Nuclear Information System (INIS)

Emmanouil, C.; Sheehan, T.M.T.; Chipman, J.K.

2007-01-01

The oxidation of DNA and lipid was analysed in the marine mussel (Mytilus edulis) in response to exposure (10 μg/l and 200 μg/l) to cadmium (Cd) and chromium [Cr(VI)]. Concentration dependent uptake of both metals into mussel tissues was established and levels of gill ATP were not depleted at these exposure levels. DNA strand breakage in gill cells (analysed by the comet assay) was elevated by both metals, however, DNA oxidation [measured by DNA strand breakage induced by the DNA repair enzyme formamidopyrimidine glycosylase (FPG)] was not elevated. This was despite a statistically significant increase in both malondialdehyde and 4-hydroxynonenal - indicative of lipid peroxidation - following treatment with Cd. In contrast, both frank DNA stand breaks and FPG-induced DNA strand breaks (indicative of DNA oxidation) were increased following injection of mussels with sodium dichromate (10.4 μg Cr(VI)/mussel). The metals also showed differential inhibitory potential towards DNA repair enzyme activity with Cd exhibiting inhibition of DNA cutting activity towards an oligonucleotide containing 8-oxo-7,8-dihydro-2'-deoxyguanosine and Cr(VI) showing inhibition of such activity towards an oligonucleotide containing ethenoadenosine, both at 200 μg/l. The metals thus show DNA damage activity in mussel gill with distinct mechanisms involving both direct and indirect (oxidative) DNA damage, as well as impairing different DNA repair capacities. A combination of these activities can contribute to adverse effects in these organisms

Linear scans of hair strands for trace elements by proton induced x-ray emission

International Nuclear Information System (INIS)

Jolly, R.K.; Pehrson, G.R.; Gupta, S.K.; Buckle, D.C.; Aceto, H. Jr.

1974-01-01

Hair strands obtained from school children in the 10 to 12 year age group were analyzed for trace element concentration as a function of distance from the root by proton-induced x-ray emission to study the history of exposure of the donors to toxic trace metals. These samples were collected from the vicinity of a copper smelter where high levels of As, Cd, Sb, and Pb have been noted. Scans show a continual build-up of Pb as a function of distance from the root, while As shows a reproducible and distinct maximum approximately 10 cm from the root. The concentration of Zn was found to be constant in all samples (without exception) to within the uncertainties of our measurements

Detection and Repair of Ionizing Radiation-Induced DNA Double Strand Breaks: New Developments in Nonhomologous End Joining

International Nuclear Information System (INIS)

Wang, Chen; Lees-Miller, Susan P.

2013-01-01

DNA damage can occur as a result of endogenous metabolic reactions and replication stress or from exogenous sources such as radiation therapy and chemotherapy. DNA double strand breaks are the most cytotoxic form of DNA damage, and defects in their repair can result in genome instability, a hallmark of cancer. The major pathway for the repair of ionizing radiation-induced DSBs in human cells is nonhomologous end joining. Here we review recent advances on the mechanism of nonhomologous end joining, as well as new findings on its component proteins and regulation

Detection and Repair of Ionizing Radiation-Induced DNA Double Strand Breaks: New Developments in Nonhomologous End Joining

Energy Technology Data Exchange (ETDEWEB)

Wang, Chen [Departments of Biochemistry and Molecular Biology and Oncology, and Southern Alberta Cancer Research Institute, University of Calgary, Calgary (Canada); Lees-Miller, Susan P., E-mail: leesmill@ucalgary.ca [Departments of Biochemistry and Molecular Biology and Oncology, and Southern Alberta Cancer Research Institute, University of Calgary, Calgary (Canada)

2013-07-01

DNA damage can occur as a result of endogenous metabolic reactions and replication stress or from exogenous sources such as radiation therapy and chemotherapy. DNA double strand breaks are the most cytotoxic form of DNA damage, and defects in their repair can result in genome instability, a hallmark of cancer. The major pathway for the repair of ionizing radiation-induced DSBs in human cells is nonhomologous end joining. Here we review recent advances on the mechanism of nonhomologous end joining, as well as new findings on its component proteins and regulation.

Mobilization of Copper ions by Flavonoids in Human Peripheral Lymphocytes Leads to Oxidative DNA Breakage: A Structure Activity Study

Directory of Open Access Journals (Sweden)

Hussain Arif

2015-11-01

Full Text Available Epidemiological studies have linked dietary consumption of plant polyphenols with lower incidence of various cancers. In particular, flavonoids (present in onion, tomato and other plant sources induce apoptosis and cytotoxicity in cancer cells. These can therefore be used as lead compounds for the synthesis of novel anticancer drugs with greater bioavailability. In the present study, we examined the chemical basis of cytotoxicity of flavonoids by studying the structure–activity relationship of myricetin (MN, fisetin (FN, quercetin (QN, kaempferol (KL and galangin (GN. Using single cell alkaline gel electrophoresis (comet assay, we established the relative efficiency of cellular DNA breakage as MN > FN > QN > KL > GN. Also, we determined that the cellular DNA breakage was the result of mobilization of chromatin-bound copper ions and the generation of reactive oxygen species. The relative DNA binding affinity order was further confirmed using molecular docking and thermodynamic studies through the interaction of flavonoids with calf thymus DNA. Our results suggest that novel anti-cancer molecules should have ortho-dihydroxy groups in B-ring and hydroxyl groups at positions 3 and 5 in the A-ring system. Additional hydroxyl groups at other positions further enhance the cellular cytotoxicity of the flavonoids.

DNA damage and mutagenesis of lambda phage induced by gamma-rays

International Nuclear Information System (INIS)

Bertram, Heidi

1988-01-01

Lambda phage DNA was gamma irradiated in aqueous solution and strand breakage determined. Twice as much minor structural damage per lethal hit was found in this DNA compared with DNA from irradiated phage suspensions. The in vitro irradiated DNA was repackaged into infectious particles. Induction of mutations in the cI or cII cistron was scored using SOS-induced host cells. In vitro prepared particles were found to have second-order kinetics for mutagenesis induced by gamma rays indicating two pre-mutational events were necessary to produce a mutation, but bacteria-free phage suspensions ('lys-phage') showed single hit kinetics for mutagenesis after irradiation. Increase in the mutation rate in the phage particles was mainly due to minor lesions, i.e. ssb, als and unidentified base damage. In lys-phage, mutagenesis might be enhanced by clustered DNA damage - configuration not existing in pack-phage. Loss of infectivity was analysed in comparison with structural damage. All lesions contributed to biological inactivation. Minor lesions were tolerated by lambda phage to a limited extent. Major lesions (e.g. dsb) contributed most to infectivity loss and were considered lethal events. (U.K.)

Ionizing-radiation induced DNA double-strand breaks: A direct and indirect lighting up

International Nuclear Information System (INIS)

Vignard, Julien; Mirey, Gladys; Salles, Bernard

2013-01-01

The occurrence of DNA double-strand breaks (DSBs) induced by ionizing radiation has been extensively studied by biochemical or cell imaging techniques. Cell imaging development relies on technical advances as well as our knowledge of the cell DNA damage response (DDR) process. The DDR involves a complex network of proteins that initiate and coordinate DNA damage signaling and repair activities. As some DDR proteins assemble at DSBs in an established spatio-temporal pattern, visible nuclear foci are produced. In addition, post-translational modifications are important for the signaling and the recruitment of specific partners at damaged chromatin foci. We briefly review here the most widely used methods to study DSBs. We also discuss the development of indirect methods, using reporter expression or intra-nuclear antibodies, to follow the production of DSBs in real time and in living cells

Flap Endonuclease 1 Limits Telomere Fragility on the Leading Strand.

Science.gov (United States)

Teasley, Daniel C; Parajuli, Shankar; Nguyen, Mai; Moore, Hayley R; Alspach, Elise; Lock, Ying Jie; Honaker, Yuchi; Saharia, Abhishek; Piwnica-Worms, Helen; Stewart, Sheila A

2015-06-12

The existence of redundant replication and repair systems that ensure genome stability underscores the importance of faithful DNA replication. Nowhere is this complexity more evident than in challenging DNA templates, including highly repetitive or transcribed sequences. Here, we demonstrate that flap endonuclease 1 (FEN1), a canonical lagging strand DNA replication protein, is required for normal, complete leading strand replication at telomeres. We find that the loss of FEN1 nuclease activity, but not DNA repair activities, results in leading strand-specific telomere fragility. Furthermore, we show that FEN1 depletion-induced telomere fragility is increased by RNA polymerase II inhibition and is rescued by ectopic RNase H1 expression. These data suggest that FEN1 limits leading strand-specific telomere fragility by processing RNA:DNA hybrid/flap intermediates that arise from co-directional collisions occurring between the replisome and RNA polymerase. Our data reveal the first molecular mechanism for leading strand-specific telomere fragility and the first known role for FEN1 in leading strand DNA replication. Because FEN1 mutations have been identified in human cancers, our findings raise the possibility that unresolved RNA:DNA hybrid structures contribute to the genomic instability associated with cancer. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Single-strand breaks in supercoiled DNA induced by vacuum-UV radiation in aqueous solution

Energy Technology Data Exchange (ETDEWEB)

Takakura, Kaoru; Ishikawa, Mitsuo; Hieda, Kotaro; Kobayashi, Katsumi; Ito, Atsushi; Ito, Takashi

1986-09-01

The induction of single-strand breaks in the DNA of plasmid pBR 322 by vacuum-UV radiation above 145 nm in aqueous solutions was studied in relation to the production of OH-radicals in water. The similarity and dissimilarity were examined on the wavelength dependence between the two effects. The maximum of single strand breaks at 150 nm could be explained by the action of OH-radicals derived from direct water photolysis: the maximum at 180 nm remains unexplained. There was no indication that the direct absorption of photon by the DNA molecule plays an important role in the production of single-strand breaks.

Single-strand breaks in supercoiled DNA induced by vacuum-UV radiation in aqueous solution

International Nuclear Information System (INIS)

Takakura, Kaoru; Ishikawa, Mitsuo; Hieda, Kotaro; Kobayashi, Katsumi; Ito, Atsushi; Ito, Takashi

1986-01-01

The induction of single-strand breaks in the DNA of plasmid pBR 322 by vacuum-UV radiation above 145 nm in aqueous solutions was studied in relation to the production of OH-radicals in water. The similarity and dissimilarity were examined on the wavelength dependence between the two effects. The maximum of single strand breaks at 150 nm could be explained by the action of OH-radicals derived from direct water photolysis: the maximum at 180 nm remains unexplained. There was no indication that the direct absorption of photon by the DNA molecule plays an important role in the production of single-strand breaks. (author)

Mechanical properties of rubberwood oriented strand lumber (OSL: The effect of strand length

Directory of Open Access Journals (Sweden)

Buhnnum Kyokong

2005-09-01

Full Text Available Effect of strand length on mechanical properties (tension, compression and bending of oriented strand lumber (OSL made of rubberwood (Hevea brasiliensis Muell. Arg. was reported. Three strand lengths of 50 mm, 100 mm, and 150 mm with 1 mm thickness and 15 mm width were used. The strands were mixed with 5% pMDI glue (weight basis in a tumble mixer. The OSL specimens were formed by hot pressing process of unidirectionally aligned strands. Average specific gravity and moisture content were 0.76 and 8.34%, respectively. Tension and compression tests were carried out for directions both parallel and perpendicular to grain while bending test was performed only in parallel direction. Ultimate stresses and moduli of elasticity were examined from the stress-strain curves. It was found that for the parallel-to-grain direction, the longer strand OSL gave higher strength. The role of the strand length did not appear for the direction normal to the grain. The relationship between the mechanical properties of OSL and strand length was well described by the modified Hankinson formula.

Effect of Twist Pitch in the Strands on the Saturation and Losses in the Nb3Sn Strands for the ITER TF CICC

International Nuclear Information System (INIS)

Martovetsky, N.N.

2007-01-01

ITER TF coils will see a significant longitudinal magnetic field in the event of the plasma disruption. This abrupt change of magnetic fields results in the appearance of an additional electrical field in the strands. The mechanism of this electrical field is the induced currents that expel the flux from the strands. This effect was known since the late 1970's [1-3] and most of the details necessary for the analyses given in this report are presented in [4]. Let's assume for simplicity a zero transport current in the strand. When a longitudinal pulsed field is applied, the outer filaments will carry an induced current repelling the change of flux. The current density of this current is 'critical' in the simplification of Bean's critical state model, where superconducting transition is represented as j=j c at any non-zero electrical field and zero where the electrical field has not penetrated. In reality, since the current density is roughly logarithmic with the electrical field, E=E c *exp[(j-j c )/jo], Bean's model is just a simplification, and current density is slightly nonuniform in the outer filament and more so for the interior strands. The inner portion of the filaments will carry a current of the opposite sign. Even in the Bean's model it is not uniform, but the assumption that it is uniform and less than critical simplifies mathematics significantly and does not deviate far from the real current density distribution. In certain circumstances, the average electrical field in the strands will be high enough to exceed the take-off electrical field averaged across the cross section. In this case, the multifilamentary strand will become unstable and will experience transition to the normal state. With zero transport current, it will eventually recover, of course. This phenomenon is analogous to the flux jump. If the strand carries a transport current, the situation becomes more complicated. If it goes unstable and the transport current is higher than the

Structure-spectrophotometric selectivity relationship in interactions of quercetin related flavonoids with double stranded and single stranded RNA

Science.gov (United States)

Piantanida, Ivo; Mašić, Lozika; Rusak, Gordana

2009-04-01

Interactions of five flavonoids with dsRNA and single stranded ssRNA were studied by UV/vis titrations. The results obtained supported the intercalative binding mode as a dominant interaction of studied flavonoids with dsRNA as well as major interaction with ssRNA. Furthermore, changes of the UV/vis spectra of flavonoids induced by addition of poly G or poly C, respectively, are significantly stronger than changes induced by double stranded poly G-poly C, pointing to essential role of the free poly G or poly C sequence (not hydrogen bonded in double helix). Exclusively poly G caused significant batochromic shift of the UV/vis maxima of all studied flavonoids, whereby the intensity of batochromic shift is nicely correlated to the number of OH groups of flavonoid. Unlikely to poly G, addition of poly A and poly U induced measurable changes only in the UV/vis spectra of flavonoids characterised by no OH (galangin) or three OH groups (myricetin) on the phenyl part of the molecule. Consequently, flavonoids with one- or two-OH groups on the phenyl part of the molecule (luteolin, fisetin, kaempferol) specifically differentiate between poly A, poly U (negligible changes in the UV/Vis spectra) and poly G (strong changes in the UV/Vis spectra) as well as poly C (moderate changes in the UV/Vis spectra).

Mechanical behavior of Nb{sub 3}Sn strands under transverse electromagnetic loads

Energy Technology Data Exchange (ETDEWEB)

Ru, Yanyun; Yong, Huadong, E-mail: yonghd@lzu.edu.cn; Zhou, Youhe

2016-11-15

Highlights: • The contact forces between the strands under transverse electromagnetic load are considered. • The indentation depth for two-dimensional contact problem is calculated. • The strand damage for different layers is presented. • We discuss the effect of bending wavelength on the Nb{sub 3}Sn strand. - Abstract: The performance of Nb{sub 3}Sn cable-in-conduit-conductor (CICC) shows a significant degradation with increasing electromagnetic load in the International Thermonuclear Experimental Reactor (ITER). As the strand is under the compression caused by adjacent strands, the strand damage may occur for high contact force. In this paper, we present a 3D helix model based on the TEMLOP and FEMCAM to simulate the contact force among the strands under transverse load. The maximum linear strain and indentation depth induced by contact stress are calculated with two-dimensional contact model of cylinder. The numerical results are compared with the experimental values. Finally, the indentation depth is discussed for different layers and cross angles based on two models. With the increasing of layer number, the degradation of performance will occur. The short bending wavelength leads to small indentation depth.

Repair of chromosome damage induced by X-irradiation during G2 phase in a line of normal human fibroblasts and its malignant derivative

International Nuclear Information System (INIS)

Parshad, R.; Gantt, R.; Sanford, K.K.; Jones, G.M.; Tarone, R.E.

1982-01-01

A line of normal human skin fibroblasts (KD) differed from its malignant derivative (HUT-14) in the extent of cytogenetic damage induced by X-irradiation during G2 phase. Malignant cells had significantly more chromatid breaks and gaps after exposure to 25, 50, or 100 rad. The gaps may represent single-strand breaks. Results from alkaline elution of cellular DNA immediately after irradiation showed that the normal and malignant cells in asynchronous population were equally sensitive to DNA single-strand breakage by X-irradiation. Caffeine or beta-cytosine arabinoside (ara-C), inhibitors of DNA repair, when added directly following G2 phase exposure, significantly increased the incidence of radiation-induced chromatid damage in the normal cells. In contrast, similar treatment of the malignant cells had little influence. Ara-C differed from caffeine in its effects; whereas both agents increased the frequency of chromatid breaks and gaps, only ara-C increased the frequency of gaps to the level observed in the irradiated malignant cells. Addition of catalase, a scavenger of the derivative free hydroxyl radical (.OH), to the cultures of malignant cells before, during, and following irradiation significantly reduced the chromatid damage; and catalase prevented formation of chromatid gaps. The DNA damage induced by X-ray during G2 phase in the normal KD cells was apparently repaired by a caffeine- and ara-C-sensitive mechanism(s) that was deficient or absent in their malignant derivatives

Extremely Low Frequency Magnetic Fields Induce Spermatogenic Germ Cell Apoptosis: Possible Mechanism

Directory of Open Access Journals (Sweden)

Sang-Kon Lee

2014-01-01

Full Text Available The energy generated by an extremely low frequency electromagnetic field (ELF-EMF is too weak to directly induce genotoxicity. However, it is reported that an extremely low frequency magnetic field (ELF-MF is related to DNA strand breakage and apoptosis. The testes that conduct spermatogenesis through a dynamic cellular process involving meiosis and mitosis seem vulnerable to external stress such as heat, MF exposure, and chemical or physical agents. Nevertheless the results regarding adverse effects of ELF-EMF on human or animal reproductive functions are inconclusive. According to the guideline of the International Commission on Non-Ionizing Radiation Protection (ICNIRP; 2010 for limiting exposure to time-varying MF (1 Hz to 100 kHz, overall conclusion of epidemiologic studies has not consistently shown an association between human adverse reproductive outcomes and maternal or paternal exposure to low frequency fields. In animal studies there is no compelling evidence of causal relationship between prenatal development and ELF-MF exposure. However there is increasing evidence that EL-EMF exposure is involved with germ cell apoptosis in testes. Biophysical mechanism by which ELF-MF induces germ cell apoptosis has not been established. This review proposes the possible mechanism of germ cell apoptosis in testes induced by ELF-MF.

Base substitutions, frameshifts, and small deletions constitute ionizing radiation-induced point mutations in mammalian cells

International Nuclear Information System (INIS)

Grosovsky, A.J.; de Boer, J.G.; de Jong, P.J.; Drobetsky, E.A.; Glickman, B.W.

1988-01-01

The relative role of point mutations and large genomic rearrangements in ionizing radiation-induced mutagenesis has been an issue of long-standing interest. Recent studies using Southern blotting analysis permit the partitioning of ionizing radiation-induced mutagenesis in mammalian cells into detectable deletions and major genomic rearrangements and into point mutations. The molecular nature of these point mutations has been left unresolved; they may include base substitutions as well as small deletions, insertions, and frame-shifts below the level of resolution of Southern blotting analysis. In this investigation, we have characterized a collection of ionizing radiation-induced point mutations at the endogenous adenine phosphoribosyltransferase (aprt) locus of Chinese hamster ovary cells at the DNA sequence level. Base substitutions represented approximately equal to 2/3 of the point mutations analyzed. Although the collection of mutants is relatively small, every possible type of base substitution event has been recovered. These mutations are well distributed throughout the coding sequence with only one multiple occurrence. Small deletions represented the remainder of characterized mutants; no insertions have been observed. Sequence-directed mechanisms mediated by direct repeats could account for some of the observed deletions, while others appear to be directly attributable to radiation-induced strand breakage

Nbs1 ChIP-Seq Identifies Off-Target DNA Double-Strand Breaks Induced by AID in Activated Splenic B Cells.

Directory of Open Access Journals (Sweden)

Lyne Khair

2015-08-01

Full Text Available Activation-induced cytidine deaminase (AID is required for initiation of Ig class switch recombination (CSR and somatic hypermutation (SHM of antibody genes during immune responses. AID has also been shown to induce chromosomal translocations, mutations, and DNA double-strand breaks (DSBs involving non-Ig genes in activated B cells. To determine what makes a DNA site a target for AID-induced DSBs, we identify off-target DSBs induced by AID by performing chromatin immunoprecipitation (ChIP for Nbs1, a protein that binds DSBs, followed by deep sequencing (ChIP-Seq. We detect and characterize hundreds of off-target AID-dependent DSBs. Two types of tandem repeats are highly enriched within the Nbs1-binding sites: long CA repeats, which can form Z-DNA, and tandem pentamers containing the AID target hotspot WGCW. These tandem repeats are not nearly as enriched at AID-independent DSBs, which we also identified. Msh2, a component of the mismatch repair pathway and important for genome stability, increases off-target DSBs, similar to its effect on Ig switch region DSBs, which are required intermediates during CSR. Most of the off-target DSBs are two-ended, consistent with generation during G1 phase, similar to DSBs in Ig switch regions. However, a minority are one-ended, presumably due to conversion of single-strand breaks to DSBs during replication. One-ended DSBs are repaired by processes involving homologous recombination, including break-induced replication repair, which can lead to genome instability. Off-target DSBs, especially those present during S phase, can lead to chromosomal translocations, deletions and gene amplifications, resulting in the high frequency of B cell lymphomas derived from cells that express or have expressed AID.

Fragile DNA Motifs Trigger Mutagenesis at Distant Chromosomal Loci in Saccharomyces cerevisiae

Science.gov (United States)

Saini, Natalie; Zhang, Yu; Nishida, Yuri; Sheng, Ziwei; Choudhury, Shilpa; Mieczkowski, Piotr; Lobachev, Kirill S.

2013-01-01

DNA sequences capable of adopting non-canonical secondary structures have been associated with gross-chromosomal rearrangements in humans and model organisms. Previously, we have shown that long inverted repeats that form hairpin and cruciform structures and triplex-forming GAA/TTC repeats induce the formation of double-strand breaks which trigger genome instability in yeast. In this study, we demonstrate that breakage at both inverted repeats and GAA/TTC repeats is augmented by defects in DNA replication. Increased fragility is associated with increased mutation levels in the reporter genes located as far as 8 kb from both sides of the repeats. The increase in mutations was dependent on the presence of inverted or GAA/TTC repeats and activity of the translesion polymerase Polζ. Mutagenesis induced by inverted repeats also required Sae2 which opens hairpin-capped breaks and initiates end resection. The amount of breakage at the repeats is an important determinant of mutations as a perfect palindromic sequence with inherently increased fragility was also found to elevate mutation rates even in replication-proficient strains. We hypothesize that the underlying mechanism for mutagenesis induced by fragile motifs involves the formation of long single-stranded regions in the broken chromosome, invasion of the undamaged sister chromatid for repair, and faulty DNA synthesis employing Polζ. These data demonstrate that repeat-mediated breaks pose a dual threat to eukaryotic genome integrity by inducing chromosomal aberrations as well as mutations in flanking genes. PMID:23785298

Oxygen-independent direct deoxyribonucleic acid backbone breakage caused by rose bengal and visible light

Energy Technology Data Exchange (ETDEWEB)

Peak, M J; Peak, J G; Foote, C S; Krinsky, N I

1984-01-01

An oxygen enhancement ratio of 10 for the induction of backbone single-strand breaks (SSBs) in purified deoxyribonucleic acid (DNA) by monochromatic 365 nm UV radiation was obtained. Similarly, a dose reduction factor of 10 was observed when the DNA was irradiated in the presence of 0.1 M diazabicyclo(2.2.2)octane (DABCO). To determine whether this breakage of DNA was due to the action of a reactive oxygen species such as singlet oxygen, we used the photosensitizing dye Rose Bengal and visible light as a system for generating singlet oxygen. Treatment of the DNA with Rose Bengal and 545 nm monochromatic light enhanced the rate of induction of SSBs six times, compared with the rate we obtained when the light was used alone. Elimination of oxygen or addition of 0.1 M DABCO during the 545 nm irradiation in the presence of Rose Bengal did not alter the enhancement of SSBs in the DNA caused by Rose Bengal and 545 nm radiation. The induction of SSBs in the DNA caused by irradiation of the DNA by 545 nm light in the presence of Rose Bengal was not enhanced by the use of D/sub 2/O instead of H/sub 2/O as a solvent. The results indicate that Rose Bengal plus visible light can cause biological damage without the intermediacy of reactive oxygen species, i.e. Rose Bengal and visible light can react directly with biological material, in reactions that appear to be type I photosensitized processes, independent of singlet oxygen as an intermediate.

DNA replication and the repair of DNA strand breaks in nuclei of Physarum polycephalum. Progress report, September 1, 1977--July 31, 1978

International Nuclear Information System (INIS)

Brewer, E.N.; Nygaard, O.F.; Kuncio, G.

1978-01-01

Isolated nuclei and intact plasmodia of Physarum contain a heat-stable stimulator of nuclear DNA replication. This substance has been purified extensively and found to contain both protein and carbohydrate. The molecular weight, estimated by gel filtration, is ca. 30,000 d. The purified material does not exhibit DNA polymerase or DNase activity, and does not stimulate DNA polymerase activity per se. In the presence of the stimulatory factor, DNA chain elongation occurs at an elevated rate, and continues for a longer time than in its absence, but G 2 nuclei are not stimulated to initiate DNA synthesis. Double-strand breaks in nuclear DNA of irradiated plasmodia are repaired in vitro to a greater extent following nuclear isolation during G 2 , and the DNA of unirradiated plasmodia is less susceptible to double-strand breakage during cell-free nuclear incubation, than is the DNA of S-phase nuclei. This correlation suggests a common basis for both observations, for example an increase in deoxyribonuclease activity or a decrease in DNA ligase activity during the S period. This, in turn, may account for the cell cycle-dependent sensitivity of this organism, in terms of mitotic delay, to ionizing radiation

Age-dependent decline in rejoining of X-ray-induced DNA double-strand breaks in normal human lymphocytes

International Nuclear Information System (INIS)

Mayer, P.J.; Lange, C.S.; Bradley, M.O.; Nichols, W.W.

1989-01-01

Unstimulated human peripheral bloodlymphocytes (HPBL), separated by density centrifugation from anticoagulated whole blood, were X-irradiated on ice and incubated in medium at 37 0 C for repair times of 15, 30 and 120 min. Blood donors were 18 normotensive, non-smoking Caucasians aged 23-78, free from overt pathology and not taking any medications. Neutral filter elution was used to assay DNA double-strand break (DSB) induction and completeness of DSB rejoining. After 30 or 120 min repair incubation, the percentage of DSBs rejoined by cells from oder donors was less than half the percentage of DSBs rejoined by cells from younger donors. When data from the 3 age groups were pooled, the age-related decline in percent DSBs rejoined was significant for repair times 30 min and 120 min but not for 15 min. These age-related declines were observed even though DNA from older donors sustained fewer strand breaks as demonstrated by the negative correlation between donor age and DSB induction. These results suggest that the efficacy of X-ray-induced DSB repair diminishes with in vivo age in unstimulated HPBL. (author). 38 refs.; 2 figs.; 1 tab

Stranded costs and exit fees

International Nuclear Information System (INIS)

2002-01-01

The New Brunswick Market Design Committee has been directed to examine the issue of stranded costs since it is a major component of restructuring within the electricity sector. When regulated monopolies are faced with competition, they could find that some of their embedded costs cannot be recovered. These costs are referred to as stranded costs. Common sources include large capital investments in uneconomic plants or expensive power purchase contracts or fuel supply contracts. In general, stranded costs do not include gains or losses associated with normal business risks experienced by regulated utilities. This report presents recommendations for mitigation of stranded costs, valuation methodologies and cost-recovery mechanisms. It also presents a summary of experience with stranded costs in other jurisdictions such as California, Rhode Island, Pennsylvania and Ontario. Stranded costs are often recovered through an obligatory charge on all customers, particularly in jurisdictions where retail competition exists. In the New Brunswick market, however, the only customers who can create stranded costs are those eligible to choose their own suppliers. It is argued that since most customers will not have a choice of electricity suppliers, they cannot generate stranded costs and therefore, should not have to pay costs stranded by others. A method to quantify stranded costs is presented, along with a review of transmission-related stranded costs in New Brunswick. Expansion of self-generation in New Brunswick could strand transmission assets. Currently, self-generators only contribute a small amount to fixed charges of the transmission system. However, under new recommended tariffs, the amount could increase. It is likely that the net amount of stranded transmission costs will not be large. 2 refs., 1 fig

Controlled Folding, Motional, and Constitutional Dynamic Processes of Polyheterocyclic Molecular Strands.

Science.gov (United States)

Barboiu, Mihail; Stadler, Adrian-Mihail; Lehn, Jean-Marie

2016-03-18

General design principles have been developed for the control of the structural features of polyheterocyclic strands and their effector-modulated shape changes. Induced defined molecular motions permit designed enforcement of helical as well as linear molecular shapes. The ability of such molecular strands to bind metal cations allows the generation of coiling/uncoiling processes between helically folded and extended linear states. Large molecular motions are produced on coordination of metal ions, which may be made reversible by competition with an ancillary complexing agent and fueled by sequential acid/base neutralization energy. The introduction of hydrazone units into the strands confers upon them constitutional dynamics, whereby interconversion between different strand compositions is achieved through component exchange. These features have relevance for nanomechanical devices. We present a morphological and functional analysis of such systems developed in our laboratories. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Radiation damage to DNA-protein complexes

Czech Academy of Sciences Publication Activity Database

Spotheim-Maurizot, M.; Davídková, Marie

2011-01-01

Roč. 261, zima (2011), s. 1-10 ISSN 1742-6588. [COST Chemistry CM0603-MELUSYN Joint Meeting Damages Induced in Biomolecules by Low and High Energy Radiations. Paříž, 09.03.2010-12.03.2010] R&D Projects: GA AV ČR IAA1048103; GA AV ČR KJB4048401; GA MŠk 1P05OC085; GA MŠk OC09012; GA AV ČR IAB1048901 Institutional research plan: CEZ:AV0Z10480505 Keywords : radiolysis * molecular-dynamics simulation * hydroxyl radical attack * induced strand breakage Subject RIV: BO - Biophysics

Radiation and biophysical studies on cells and viruses. Progress report, February 29, 1974--March 31, 1975

International Nuclear Information System (INIS)

Cole, A.

Progress is reported on the following research projects: sedimentation analysis of chromosome components from interphase and mitotic chromosomes; electron microscopy of mitotic chromosomes; radiosensitive site analysis using short range particle beams; studies on nucleoproteins and DNA; RBE and OER for double strand breaks and single strand breaks of DNA irradiated with 241 Am alpha particles; use of 241 Am alpha particle track-ends to study the location of radiosensitive sites; gamma irradiation of nucleoprotein model systems; assembly of new equipment for the analysis of DNA size distributions; cell rejoining of DNA breaks induced by various radiations; studies on cell transformation induced by gamma radiation; localization of cellular sites for DNA breakage using labeled specific antibodies; and semiconductor properties of melanins related to preferential killing of melanoma cells. (U.S.)

The double-stranded RNA-activated protein kinase mediates viral-induced encephalitis

International Nuclear Information System (INIS)

Scheuner, Donalyn; Gromeier, Matthias; Davies, Monique V.; Dorner, Andrew J.; Song Benbo; Patel, Rupali V.; Wimmer, Eckard J.; McLendon, Roger E.; Kaufman, Randal J.

2003-01-01

The double-stranded (ds) RNA-activated protein kinase (PKR) plays an important role in control of viral infections and cell growth. We have studied the role of PKR in viral infection in mice that are defective in the PKR signaling pathway. Transgenic mice were derived that constitutively express a trans-dominant-negative kinase-defective mutant PKR under control of the β-actin promoter. The trans-dominant-negative PKR mutant expressing transgenic mice do not have a detectable phenotype, similar to observations with PKR knock-out mice. The requirement for PKR in viral pathogenesis was studied by intracerebral infection of mice with a mouse-adapted poliovirus. Histopathological analysis revealed diffuse encephalomyelitis with severe inflammatory lesions throughout the central nervous system (CNS) in infected wild-type mice. In contrast, histopathological evaluation of virus-injected trans-dominant-negative PKR transgenic mice as well as PKR knock-out mice yielded no signs of tissue damage associated with inflammatory host responses. However, the virus did replicate in both models of PKR-deficient mice at a level equal to that observed in wild-type infected mice. Although the results indicate a clear difference in susceptibility to poliovirus-induced encephalitis, this difference manifests clinically as a slight delay in fatal neuropathy in trans-dominant-negative PKR transgenic and PKR knock-out animals. Our observations support the finding that viral-induced PKR activation may play a significant role in pathogenesis by mediating the host response to viral CNS infection. They support PKR to be an effective target to control tissue damage due to deleterious host responses to viral infection

Xeroderma pigmentosum complementation group C cells remove pyrimidine dimers selectively from the transcribed strand of active genes

International Nuclear Information System (INIS)

Venema, J.; van Hoffen, A.; Karcagi, V.; Natarajan, A.T.; van Zeeland, A.A.; Mullenders, L.H.

1991-01-01

The authors have measured the removal of UV-induced pyrimidine dimers from DNA fragments of the adenosine deaminase (ADA) and dihydrofolate reductase (DHFR) genes in primary normal human and xeroderma pigmentosum complementation group C (XP-C) cells. Using strand-specific probes, we show that in normal cells, preferential repair of the 5' part of the ADA gene is due to the rapid and efficient repair of the transcribed strand. Within 8 h after irradiation with UV at 10 J m-2, 70% of the pyrimidine dimers in this strand are removed. The nontranscribed strand is repaired at a much slower rate, with 30% dimers removed after 8 h. Repair of the transcribed strand in XP-C cells occurs at a rate indistinguishable from that in normal cells, but the nontranscribed strand is not repaired significantly in these cells. Similar results were obtained for the DHFR gene. In the 3' part of the ADA gene, however, both normal and XP-C cells perform fast and efficient repair of either strand, which is likely to be caused by the presence of transcription units on both strands. The factor defective in XP-C cells is apparently involved in the processing of DNA damage in inactive parts of the genome, including nontranscribed strands of active genes. These findings have important implications for the understanding of the mechanism of UV-induced excision repair and mutagenesis in mammalian cells

Strand Displacement by DNA Polymerase III Occurs through a τ-ψ-χ Link to Single-stranded DNA-binding Protein Coating the Lagging Strand Template*

OpenAIRE

Yuan, Quan; McHenry, Charles S.

2009-01-01

In addition to the well characterized processive replication reaction catalyzed by the DNA polymerase III holoenzyme on single-stranded DNA templates, the enzyme possesses an intrinsic strand displacement activity on flapped templates. The strand displacement activity is distinguished from the single-stranded DNA-templated reaction by a high dependence upon single-stranded DNA binding protein and an inability of γ-complex to support the reaction in the absence of τ. However, if γ-complex is p...

Double-strand break repair-adox: Restoration of suppressed double-strand break repair during mitosis induces genomic instability.

Science.gov (United States)

Terasawa, Masahiro; Shinohara, Akira; Shinohara, Miki

2014-12-01

Double-strand breaks (DSBs) are one of the severest types of DNA damage. Unrepaired DSBs easily induce cell death and chromosome aberrations. To maintain genomic stability, cells have checkpoint and DSB repair systems to respond to DNA damage throughout most of the cell cycle. The failure of this process often results in apoptosis or genomic instability, such as aneuploidy, deletion, or translocation. Therefore, DSB repair is essential for maintenance of genomic stability. During mitosis, however, cells seem to suppress the DNA damage response and proceed to the next G1 phase, even if there are unrepaired DSBs. The biological significance of this suppression is not known. In this review, we summarize recent studies of mitotic DSB repair and discuss the mechanisms of suppression of DSB repair during mitosis. DSB repair, which maintains genomic integrity in other phases of the cell cycle, is rather toxic to cells during mitosis, often resulting in chromosome missegregation and aberration. Cells have multiple safeguards to prevent genomic instability during mitosis: inhibition of 53BP1 or BRCA1 localization to DSB sites, which is important to promote non-homologous end joining or homologous recombination, respectively, and also modulation of the non-homologous end joining core complex to inhibit DSB repair. We discuss how DSBs during mitosis are toxic and the multiple safeguard systems that suppress genomic instability. © 2014 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

DNA-incorporated 125I induces more than one double-strand break per decay in mammalian cells.

Science.gov (United States)

Elmroth, Kecke; Stenerlöw, Bo

2005-04-01

The Auger-electron emitter 125I releases cascades of 20 electrons per decay that deposit a great amount of local energy, and for DNA-incorporated 125I, approximately one DNA double-strand break (DSB) is produced close to the decay site. To investigate the potential of 125I to induce additional DSBs within adjacent chromatin structures in mammalian cells, we applied DNA fragment-size analysis based on pulsed-field gel electrophoresis (PFGE) of hamster V79-379A cells exposed to DNA-incorporated 125IdU. After accumulation of decays at -70 degrees C in the presence of 10% DMSO, there was a non-random distribution of DNA fragments with an excess of fragments even higher. In contrast, using a conventional low-resolution assay without measurement of smaller DNA fragments, the yield was close to one DSB/decay. We conclude that a large fraction of the DSBs induced by DNA-incorporated 125I are nonrandomly distributed and that significantly more than one DSB/decay is induced in an intact cell. Thus, in addition to DSBs produced close to the decay site, DSBs may also be induced within neighboring chromatin fibers, releasing smaller DNA fragments that are not detected by conventional DSB assays.

Strand displacement by DNA polymerase III occurs through a tau-psi-chi link to single-stranded DNA-binding protein coating the lagging strand template.

Science.gov (United States)

Yuan, Quan; McHenry, Charles S

2009-11-13

In addition to the well characterized processive replication reaction catalyzed by the DNA polymerase III holoenzyme on single-stranded DNA templates, the enzyme possesses an intrinsic strand displacement activity on flapped templates. The strand displacement activity is distinguished from the single-stranded DNA-templated reaction by a high dependence upon single-stranded DNA binding protein and an inability of gamma-complex to support the reaction in the absence of tau. However, if gamma-complex is present to load beta(2), a truncated tau protein containing only domains III-V will suffice. This truncated protein is sufficient to bind both the alpha subunit of DNA polymerase (Pol) III and chipsi. This is reminiscent of the minimal requirements for Pol III to replicate short single-stranded DNA-binding protein (SSB)-coated templates where tau is only required to serve as a scaffold to hold Pol III and chi in the same complex (Glover, B., and McHenry, C. (1998) J. Biol. Chem. 273, 23476-23484). We propose a model in which strand displacement by DNA polymerase III holoenzyme depends upon a Pol III-tau-psi-chi-SSB binding network, where SSB is bound to the displaced strand, stabilizing the Pol III-template interaction. The same interaction network is probably important for stabilizing the leading strand polymerase interactions with authentic replication forks. The specificity constant (k(cat)/K(m)) for the strand displacement reaction is approximately 300-fold less favorable than reactions on single-stranded templates and proceeds with a slower rate (150 nucleotides/s) and only moderate processivity (approximately 300 nucleotides). PriA, the initiator of replication restart on collapsed or misassembled replication forks, blocks the strand displacement reaction, even if added to an ongoing reaction.

The consequences of granulate heterogeneity towards breakage and attrition upon fluid-bed drying

NARCIS (Netherlands)

Nieuwmeyer, Florentine; Maarschalk, Kees van der Voort; Vromans, Herman

High-shear granulated lactose granulates were dried in it fluid-bed dryer at various conditions. Granules were characterized by water content and size analysis. It is shown that the drying process is very dynamic in terms of growth and breakage phenomena. Granular size heterogeneity, composition and

Repair-induced DNA double strand breaks after ultraviolet-light and either aphidocolin or 1-β-D-arabinofuranosylcytosine/hydroxyurea

International Nuclear Information System (INIS)

Bradley, M.O.; Taylor, V.I.

1983-01-01

A study was performed to determine whether 'repair-induced double strand breaks' (RDSBs) occur in IMR-90 cells at low u.v. doses and whether the RDSBs are themselves repairable by holding open the excision-repair induced gaps by inhibiting nucleotide polymerization after u.v. light with hydroxyurea/ara C or aphidocolin. The results show as little as 2.5 J.m -2 of u.v. light induces RDSBs during repair incubation when repair inhibitors are present. This suggests that 'hot spots' of high lesion frequency occur and the overlapping excision in these areas will produce RDSBs. Removing aphidocolin showed that RDSBs are only partially repairable with between 15 and 40% of the breaks unrepaired at 24 h. Because the lesions are partially repairable they should not always cause toxicity and may be involved in processes such as mutation, transformation, and chromosome or chromatid type aberrations of the sort associated with human tumors. (author)

DNA Strand Breaks in Mitotic Germ Cells of Caenorhabditis elegans Evaluated by Comet Assay

Science.gov (United States)

Park, Sojin; Choi, Seoyun; Ahn, Byungchan

2016-01-01

DNA damage responses are important for the maintenance of genome stability and the survival of organisms. Such responses are activated in the presence of DNA damage and lead to cell cycle arrest, apoptosis, and DNA repair. In Caenorhabditis elegans, double-strand breaks induced by DNA damaging agents have been detected indirectly by antibodies against DSB recognizing proteins. In this study we used a comet assay to detect DNA strand breaks and to measure the elimination of DNA strand breaks in mitotic germline nuclei of C. elegans. We found that C. elegans brc-1 mutants were more sensitive to ionizing radiation and camptothecin than the N2 wild-type strain and repaired DNA strand breaks less efficiently than N2. This study is the first demonstration of direct measurement of DNA strand breaks in mitotic germline nuclei of C. elegans. This newly developed assay can be applied to detect DNA strand breaks in different C. elegans mutants that are sensitive to DNA damaging agents. PMID:26903030

Radiation-induced DNA Double Strand Breaks and Their Modulations by Treatments with Moringa oleifera Lam. Leaf Extracts: A Cancer Cell Culture Model

International Nuclear Information System (INIS)

Boonsirichai, K.; Jetawattana, S.

2014-01-01

Gamma radiation brings deleterious effects upon human cells by inducing oxidative stress and DNA damages. Antioxidants have been shown to confer protective effects on irradiated normal cells. Moringa oleifera Lam. is a widely used nutritional supplement with antioxidant activities. This report showed that antioxidant-containing supplements, in addition to protecting normal cells, could protect cancer cells against genotoxic effects of gamma radiation. γ-H2AX immunofluorescent foci were utilized as an indicator of radiation-induced DNA double strand breaks. MCF-7 human breast adenocarcinoma cells were irradiated with 2-8 Gy gamma radiation. A linear relationship between the formation of γ-H2AX foci and radiation dose was observed with an average of 10 foci per cell per Gy. A 30-minute pretreatment of the cells with either the aqueous or the ethanolic extract of M. oleifera leaves could partially protect the cells from radiation-induced DNA double strand breaks. A pretreatment with 500 µg/mL aqueous extract reduced the number of foci formed by 15% when assayed at 30 minutes post-irradiation. The ethanolic extract was more effective; 500 µg/mL of its concentration reduced the number of foci among irradiated cells by 30%. The results indicated that irradiated cancer cells responded similarly to nutritional supplements containing antioxidants as irradiated normal cells. These natural antioxidants could confer protective effects upon cancer cells against gamma radiation. (author)

Kazinol Q from Broussonetia kazinoki Enhances Cell Death Induced by Cu(ll through Increased Reactive Oxygen Species

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Hsue-Yin Hsu

2011-04-01

Full Text Available The ability of the flavan kazinol Q (KQ to induce DNA breakage in the presence of Cu(II was examined by agarose gel electrophoresis using supercoiled plasmid DNA. In KQ-mediated DNA breakage reaction, the involvement of reactive oxygen species (ROS, H2O2 and O2 - was established by the inhibition of DNA breakage by catalase and revealed DNA breakage by superoxide dismutase (SOD. The cell viability of gastric carcinoma SCM-1 cells treated with various concentrations of KQ was significantly decreased by cotreatment with Cu(II. Treatment of SCM-1 cells with 300 μM Cu(II enhanced the necrosis induced by 100 μM KQ. Treatment of SCM-1 cells with 100 mM KQ in the presence of 300 mM Cu(II increased the generation of H2O2. Taken together, the above finding suggested that KQ cotreatment with Cu(II produced increased amounts of H2O2, thus enhancing subsequent cell death due to necrosis.

The foci of DNA double strand break-recognition proteins localize with γH2AX after heat treatment

International Nuclear Information System (INIS)

Takahashi, Akihisa; Mori, Eiichiro; Ohnishi, Takeo

2010-01-01

Recently, there have been many reports concerning proteins which can recognize DNA double strand break (DSBs), and such proteins include histone H2AX phosphorylated at serine 139 (γH2AX), ataxia telangiectasia mutated (ATM) phospho-serine 1981, DNA-dependent protein kinase catalytic subunit (DNA-PKcs) phospho-threonine 2609, Nijmegen breakage syndrome 1 (NBS1) phospho-serine 343, checkpoint kinase 2 (CHK2), phospho-threonine 68, and structural maintenance of chromosomes 1 (SMC1) phospho-serine 966. Thus, it should be possible to follow the formation of DSBs and their repair using immunohistochemical methods with multiple antibodies to detect these proteins. When normal human fibroblasts (AG1522 cells) were exposed to 3 Gy of X-rays as a control, clearly discernable foci for these proteins were detected, and these foci localized with γH2AX foci. After heat treatment at 45.5 deg C for 20 min, these proteins are partially localized with γH2AX foci. Here we show that there were slight differences in the localization pattern among these proteins, such as a disappearance from the nucleus (phospho-ATM) and translocation to the cytoplasm (phospho-NBS1) at 30 min after heat treatment, and some foci (phospho-DNA-PKcs and phospho-CHK2) appeared at 8 h after heat treatment. These results are discussed from perspectives of heat-induced denaturation of proteins and formation of DSBs. (author)

Detection of DNA strand breaks in mammalian cells using the radioresistant bacterium PprA protein

International Nuclear Information System (INIS)

Satoh, Katsuya; Wada, Seiichi; Narumi, Issay; Kikuchi, Masahiro; Funayama, Tomoo; Kobayashi, Yasuhiko

2003-01-01

We have previously found that the PprA protein from Deinococcus radiodurans possesses ability to recognize DNA carrying strand breaks. In the present study, we attempted to visualize radiation-induced DNA strand breaks with PprA protein using immunofluorescence technique to elucidate the DNA damage response mechanism in mammalian cultured cells. As a result, colocalization of Cy2 and DAPI fluorescent signals was observed. This observation suggests that DNA strand breaks in the nucleus of CHO-K1 cells were effectively detected using the PprA protein. The amount of DNA strand breaks (integrated density of Cy2 fluorescent signals) was increased with the increase in the radiation dose. (author)

Simulation on breakage of heterogeneous materials caused by detonative loading; Bakugo shogeki ni yoru fukinshitsu zairyo no hakai gensho no simulation

Energy Technology Data Exchange (ETDEWEB)

Sassa, K; Watanabe, T; Ashida, Y [Kyoto University, Kyoto (Japan). Faculty of Engineering

1996-05-01

Investigations were conducted by simulation of breakage of inhomogeneous materials (rock) attributable to detonative loading, which simulation used the Days-2 Code. During the simulation, one-free-face blastings were used for testing a homogeneous structure, horizontal 2-layer structure, and horizontal 3-layer structure. Property values were assigned to the rocks on the assumption that they were sedimentary rocks such as sandstone or mudstone or hard rocks such as granite. As the result, it was found that a detonative loading resulted in shear failure in a sphere near the focus that was followed by radially developed cracks due to tension breakage, that more area is damaged in a soft rock than in a hard rock, that cracks due to breakage are produced by the overlapping of waves directly from the focus and those reflected from the free face in case of one-free-face blastings, that such cracks propagated along the soft rock layer in case there is a soft rock layer in a hard rock, but that breakage does not extend beyond the soft rock layer. 6 refs., 6 figs., 1 tab.

RDE-1 slicer activity is required only for passenger-strand cleavage during RNAi in Caenorhabditis elegans.

NARCIS (Netherlands)

Steiner, F.A.; Okihara, K.L.; Hoogstrate, S.W.; Sijen, T.; Ketting, R.F.

2009-01-01

RNA interference (RNAi) is a process in which double-stranded RNA is cleaved into small interfering RNAs (siRNAs) that induce the destruction of homologous single-stranded mRNAs. Argonaute proteins are essential components of this silencing process; they bind siRNAs directly and can cleave RNA

Viral single-strand DNA induces p53-dependent apoptosis in human embryonic stem cells.

Science.gov (United States)

Hirsch, Matthew L; Fagan, B Matthew; Dumitru, Raluca; Bower, Jacquelyn J; Yadav, Swati; Porteus, Matthew H; Pevny, Larysa H; Samulski, R Jude

2011-01-01

Human embryonic stem cells (hESCs) are primed for rapid apoptosis following mild forms of genotoxic stress. A natural form of such cellular stress occurs in response to recombinant adeno-associated virus (rAAV) single-strand DNA genomes, which exploit the host DNA damage response for replication and genome persistence. Herein, we discovered a unique DNA damage response induced by rAAV transduction specific to pluripotent hESCs. Within hours following rAAV transduction, host DNA damage signaling was elicited as measured by increased gamma-H2AX, ser15-p53 phosphorylation, and subsequent p53-dependent transcriptional activation. Nucleotide incorporation assays demonstrated that rAAV transduced cells accumulated in early S-phase followed by the induction of apoptosis. This lethal signaling sequalae required p53 in a manner independent of transcriptional induction of Puma, Bax and Bcl-2 and was not evident in cells differentiated towards a neural lineage. Consistent with a lethal DNA damage response induced upon rAAV transduction of hESCs, empty AAV protein capsids demonstrated no toxicity. In contrast, DNA microinjections demonstrated that the minimal AAV origin of replication and, in particular, a 40 nucleotide G-rich tetrad repeat sequence, was sufficient for hESC apoptosis. Our data support a model in which rAAV transduction of hESCs induces a p53-dependent lethal response that is elicited by a telomeric sequence within the AAV origin of replication.

Viral single-strand DNA induces p53-dependent apoptosis in human embryonic stem cells.

Directory of Open Access Journals (Sweden)

Matthew L Hirsch

Full Text Available Human embryonic stem cells (hESCs are primed for rapid apoptosis following mild forms of genotoxic stress. A natural form of such cellular stress occurs in response to recombinant adeno-associated virus (rAAV single-strand DNA genomes, which exploit the host DNA damage response for replication and genome persistence. Herein, we discovered a unique DNA damage response induced by rAAV transduction specific to pluripotent hESCs. Within hours following rAAV transduction, host DNA damage signaling was elicited as measured by increased gamma-H2AX, ser15-p53 phosphorylation, and subsequent p53-dependent transcriptional activation. Nucleotide incorporation assays demonstrated that rAAV transduced cells accumulated in early S-phase followed by the induction of apoptosis. This lethal signaling sequalae required p53 in a manner independent of transcriptional induction of Puma, Bax and Bcl-2 and was not evident in cells differentiated towards a neural lineage. Consistent with a lethal DNA damage response induced upon rAAV transduction of hESCs, empty AAV protein capsids demonstrated no toxicity. In contrast, DNA microinjections demonstrated that the minimal AAV origin of replication and, in particular, a 40 nucleotide G-rich tetrad repeat sequence, was sufficient for hESC apoptosis. Our data support a model in which rAAV transduction of hESCs induces a p53-dependent lethal response that is elicited by a telomeric sequence within the AAV origin of replication.

Depletion of the type 1 IGF receptor delays repair of radiation-induced DNA double strand breaks

International Nuclear Information System (INIS)

Turney, Benjamin W.; Kerr, Martin; Chitnis, Meenali M.; Lodhia, Kunal; Wang, Yong; Riedemann, Johann; Rochester, Mark; Protheroe, Andrew S.; Brewster, Simon F.; Macaulay, Valentine M.

2012-01-01

Background and purpose: IGF-1R depletion sensitizes prostate cancer cells to ionizing radiation and DNA-damaging cytotoxic drugs. This study investigated the hypothesis that IGF-1R regulates DNA double strand break (DSB) repair. Methods: We tested effects of IGF-1R siRNA transfection on the repair of radiation-induced DSBs by immunoblotting and immunofluorescence for γH2AX, and pulsed-field gel electrophoresis. Homologous recombination (HR) was quantified by reporter assays, and cell cycle distribution by flow cytometry. Results: We confirmed that IGF-1R depletion sensitized DU145 and PC3 prostate cancer cells to ionizing radiation. DU145 control transfectants resolved radiation-induced DSBs within 24 h, while IGF-1R depleted cells contained 30–40% unrepaired breaks at 24 h. IGF-1R depletion induced significant reduction in DSB repair by HR, although the magnitude of the repair defect suggests additional contributory factors. Radiation-induced G2-M arrest was attenuated by IGF-1R depletion, potentially suppressing cell cycle-dependent processes required for HR. In contrast, IGF-1R depletion induced only minor radiosensitization in LNCaP cells, and did not influence repair. Cell cycle profiles were similar to DU145, so were unlikely to account for differences in repair responses. Conclusions: These data indicate a role for IGF-1R in DSB repair, at least in part via HR, and support use of IGF-1R inhibitors with DNA damaging cancer treatments.

Depletion of the type 1 IGF receptor delays repair of radiation-induced DNA double strand breaks.

Science.gov (United States)

Turney, Benjamin W; Kerr, Martin; Chitnis, Meenali M; Lodhia, Kunal; Wang, Yong; Riedemann, Johann; Rochester, Mark; Protheroe, Andrew S; Brewster, Simon F; Macaulay, Valentine M

2012-06-01

IGF-1R depletion sensitizes prostate cancer cells to ionizing radiation and DNA-damaging cytotoxic drugs. This study investigated the hypothesis that IGF-1R regulates DNA double strand break (DSB) repair. We tested effects of IGF-1R siRNA transfection on the repair of radiation-induced DSBs by immunoblotting and immunofluorescence for γH2AX, and pulsed-field gel electrophoresis. Homologous recombination (HR) was quantified by reporter assays, and cell cycle distribution by flow cytometry. We confirmed that IGF-1R depletion sensitized DU145 and PC3 prostate cancer cells to ionizing radiation. DU145 control transfectants resolved radiation-induced DSBs within 24 h, while IGF-1R depleted cells contained 30-40% unrepaired breaks at 24 h. IGF-1R depletion induced significant reduction in DSB repair by HR, although the magnitude of the repair defect suggests additional contributory factors. Radiation-induced G2-M arrest was attenuated by IGF-1R depletion, potentially suppressing cell cycle-dependent processes required for HR. In contrast, IGF-1R depletion induced only minor radiosensitization in LNCaP cells, and did not influence repair. Cell cycle profiles were similar to DU145, so were unlikely to account for differences in repair responses. These data indicate a role for IGF-1R in DSB repair, at least in part via HR, and support use of IGF-1R inhibitors with DNA damaging cancer treatments. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Nitric oxide mediated DNA double strand breaks induced in proliferating bystander cells after {alpha}-particle irradiation

Energy Technology Data Exchange (ETDEWEB)

Han Wei [Department of Physics and Materials Science, City University of Hong Kong, Tat Chee Avenue, Kowloon Tong (Hong Kong); Chen Shaopeng [Department of Physics and Materials Science, City University of Hong Kong, Tat Chee Avenue, Kowloon Tong (Hong Kong); Key Laboratory of Ion Beam Bioengineering, Institute of Plasma Physics, Chinese Academy of Sciences, Hefei 230031 (China); Yu, K.N., E-mail: peter.yu@cityu.edu.hk [Department of Physics and Materials Science, City University of Hong Kong, Tat Chee Avenue, Kowloon Tong (Hong Kong); Wu Lijun [Key Laboratory of Ion Beam Bioengineering, Institute of Plasma Physics, Chinese Academy of Sciences, Hefei 230031 (China)

2010-02-03

Low-dose {alpha}-particle exposures comprise 55% of the environmental dose to the human population and have been shown to induce bystander responses. Previous studies showed that bystander effect could induce stimulated cell growth or genotoxicity, such as excessive DNA double strand breaks (DSBs), micronuclei (MN), mutation and decreased cell viability, in the bystander cell population. In the present study, the stimulated cell growth, detected with flow cytometry (FCM), and the increased MN and DSB, detected with p53 binding protein 1 (53BP1) immunofluorescence, were observed simultaneously in the bystander cell population, which were co-cultured with cells irradiated by low-dose {alpha}-particles (1-10 cGy) in a mixed system. Further studies indicated that nitric oxide (NO) and transforming growth factor {beta}1 (TGF-{beta}1) played very important roles in mediating cell proliferation and inducing MN and DSB in the bystander population through treatments with NO scavenger and TGF-{beta}1 antibody. Low-concentrations of NO, generated by spermidine, were proved to induce cell proliferation, DSB and MN simultaneously. The proliferation or shortened cell cycle in bystander cells gave them insufficient time to repair DSBs. The increased cell division might increase the probability of carcinogenesis in bystander cells since cell proliferation increased the probability of mutation from the mis-repaired or un-repaired DSBs.

Nitric oxide mediated DNA double strand breaks induced in proliferating bystander cells after α-particle irradiation

International Nuclear Information System (INIS)

Han Wei; Chen Shaopeng; Yu, K.N.; Wu Lijun

2010-01-01

Low-dose α-particle exposures comprise 55% of the environmental dose to the human population and have been shown to induce bystander responses. Previous studies showed that bystander effect could induce stimulated cell growth or genotoxicity, such as excessive DNA double strand breaks (DSBs), micronuclei (MN), mutation and decreased cell viability, in the bystander cell population. In the present study, the stimulated cell growth, detected with flow cytometry (FCM), and the increased MN and DSB, detected with p53 binding protein 1 (53BP1) immunofluorescence, were observed simultaneously in the bystander cell population, which were co-cultured with cells irradiated by low-dose α-particles (1-10 cGy) in a mixed system. Further studies indicated that nitric oxide (NO) and transforming growth factor β1 (TGF-β1) played very important roles in mediating cell proliferation and inducing MN and DSB in the bystander population through treatments with NO scavenger and TGF-β1 antibody. Low-concentrations of NO, generated by spermidine, were proved to induce cell proliferation, DSB and MN simultaneously. The proliferation or shortened cell cycle in bystander cells gave them insufficient time to repair DSBs. The increased cell division might increase the probability of carcinogenesis in bystander cells since cell proliferation increased the probability of mutation from the mis-repaired or un-repaired DSBs.

Chromatin- and temperature-dependent modulation of radiation-induced double-strand breaks.

Science.gov (United States)

Elmroth, K; Nygren, J; Stenerlöw, B; Hultborn, R

2003-10-01

To investigate the influence of chromatin organization and scavenging capacity in relation to irradiation temperature on the induction of double-strand breaks (DSB) in structures derived from human diploid fibroblasts. Agarose plugs with different chromatin structures (intact cells+/-wortmannin, permeabilized cells with condensed chromatin, nucleoids and DNA) were prepared and irradiated with X-rays at 2 or 37 degrees C and lysed using two different lysis protocols (new ice-cold lysis or standard lysis at 37 degrees C). Induction of DSB was determined by constant-field gel electrophoresis. The dose-modifying factor (DMF(temp)) for irradiation at 37 compared with 2 degrees C was 0.92 in intact cells (i.e. more DSB induced at 2 degrees C), but gradually increased to 1.5 in permeabilized cells, 2.2 in nucleoids and 2.6 in naked DNA, suggesting a role of chromatin organization for temperature modulation of DNA damage. In addition, DMF(temp) was influenced by the presence of 0.1 M DMSO or 30 mM glutathione, but not by post-irradiation temperature. The protective effect of low temperature was correlated to the indirect effects of ionizing radiation and was not dependent on post-irradiation temperature. Reasons for a dose modifying factor <1 in intact cells are discussed.

Radiobiological study on DNA strand breaks and repair using single cell gel electrophoresis

International Nuclear Information System (INIS)

Ikushima, Takaji

1994-01-01

Single cell gel electrophoresis (SCGE) provides a novel method to measure DNA damage in individual cells and more importantly, to assess heterogeneity in response within a mixed population of cells. Cells embedded in agarose are lysed, subjected to electrophoresis, stained with a fluorescent DNA-specific dye, and viewed under a fluorescence microscope. Damaged cells display 'comets', broken DNA migrating farther to the anode in the electric field. We have previously used this technique to quantify DNA damage induced by moderate doses of low and high LET radiations in cultured Chinese hamster cells. The assay has been optimized in terms of lysing and electrophoresis conditions, and applied to analyse the DNA strand breaks, their repair kinetics and heterogeneity in response in individual Chinese hamster cells exposed to gamma-rays, and to KUR thermal neutrons with and without 10 B or to KEK PF monochromatic soft X-rays as well as to a radio-mimetic agent, neocarzinostatin. The DNA double-strand breaks induced by boron-neutron captured reactions were repaired at a slower rate, but a heterogeneity in response might not contribute to the difference. The neocarzinostatin-induced DNA damage were efficiently repaired in a dose-dependent fashion. The initial amount of gamma-ray induced DNA double-strand breaks was not significantly altered in cells pre-exposed to very low adapting dose. (author)

Wear and breakage monitoring of cutting tools by an optical method: theory

Science.gov (United States)

Li, Jianfeng; Zhang, Yongqing; Chen, Fangrong; Tian, Zhiren; Wang, Yao

1996-10-01

An essential part of a machining system in the unmanned flexible manufacturing system, is the ability to automatically change out tools that are worn or damaged. An optoelectronic method for in situ monitoring of the flank wear and breakage of cutting tools is presented. A flank wear estimation system is implemented in a laboratory environment, and its performance is evaluated through turning experiments. The flank wear model parameters that need to be known a priori are determined through several preliminary experiments, or from data available in the literature. The resulting cutting conditions are typical of those used in finishing cutting operations. Through time and amplitude domain analysis of the cutting tool wear states and breakage states, it is found that the original signal digital specificity (sigma) 2x and the self correlation coefficient (rho) (m) can reflect the change regularity of the cutting tool wear and break are determined, but which is not enough due to the complexity of the wear and break procedure of cutting tools. Time series analysis and frequency spectrum analysis will be carried out, which will be described in the later papers.

Hydrogen peroxide-induced DNA damage is independent of nuclear calcium but dependent on redox-active ions.

Science.gov (United States)

Jornot, L; Petersen, H; Junod, A F

1998-01-01

In cells undergoing oxidative stress, DNA damage may result from attack by .OH radicals produced by the Fenton reaction, and/or by nucleases activated by nuclear calcium. In the present study, the participation of these two mechanisms was investigated in HeLa cells. Nuclear-targeted aequorin was used for selectively monitoring Ca2+ concentrations within the nuclei ([Ca2+]n), in conjunction with the cell-permeant calcium chelator bis-(o-aminophenoxy)ethane-N,N,N', N'-tetraacetic acid acetoxymethyl ester (BAPTA/AM), the lipid-soluble broad-spectrum metal chelator with low affinity for Ca2+ and Mg2+ N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN), and the high-affinity iron/copper chelator 1, 10-phenanthroline (PHE). In Ca2+-containing medium, H2O2 induced extensive DNA strand breaks and an increase in [Ca2+]n that was almost identical to that observed in the cytosol ([Ca2+]c). In cells bathed in Ca2+-free/EGTA medium, in which the increases in [Ca2+]n and [Ca2+]c due to H2O2 were significantly reduced, similar levels of DNA fragmentation also occurred. In cells preloaded with BAPTA/AM or TPEN, the small increase of [Ca2+]n normally elicited by H2O2 in Ca2+-free medium was completely buffered, and DNA damage was largely prevented. On the other hand, pretreatment with PHE did not affect the calcium response in the nuclei, but completely prevented DNA strand breakage induced by H2O2. Re-addition of 100 microM CuSO4 and 100 microM FeSO4 to TPEN- and PHE-treated cells prior to H2O2 challenge reversed the effect of TPEN and PHE, whereas 1 mM was necessary to negate the effect of BAPTA/AM. The levels of DNA strand breakage observed, however, did not correlate with the amounts of 8-hydroxy 2'-deoxyguanosine (8-OHdG): H2O2 did not produce 8-OHdG, whereas PHE alone slightly increased 8-OHdG levels. CuSO4 and FeSO4 enhanced the effects of PHE, particularly in the presence of H2O2. Exposure of cells to a mixture of CuSO4/FeSO4 also resulted in a significant increase in

Induction and removal of DNA interstrand cross-links in V-79 Chinese hamster cells measured by hydroxylapatite chromatography after treatments with bifunctional furocoumarins

International Nuclear Information System (INIS)

Dardalhon, M.; Averbeck, D.

1988-01-01

DNA interstrand crosslinks (CL) photoinduced by bifunctional furocoumarins in V-79 Chinese hamster cells were measured by alkaline denaturation and hydroxylapatite chromatography. Treatments with 5-methoxypsoralen (5-MOP), 8-methoxypsoralen (8-MOP) and 4,5',8-trimethylpsoralen (4,5',8-TMP) and 365 nm irradiation (UVA) confer a dose-dependent linear increase in the amount of double-stranded DNA indicating the induction of CL. Determination in alkaline sucrose gradients of the molecular weight of the DNA and estimation of drug-induced strand breakage allowed quantification of the CL induced. 5-MOP was found to be slightly more effective than 8-MOP whereas 4,5',8-TMP was 9 times more effective for the induction of CL. The fate of CL during post-treatment incubation was also followed. Cells in exponential growth phase were found to be efficient in the removal of CL. (Author)

Photosensitive interaction of RSU 1069 with DNA

Energy Technology Data Exchange (ETDEWEB)

Edwards, D.I.; Knox, R.J.; Skolimowski, I.M.; Zahoor, A.; Knight, R.C.

1984-08-01

RSU 1069 is a 2-nitroimidazole radiosensitizer with an aziridine-containing side chain. In light (360 nm) the absorbance maximum of the nitro group at 325 nm disappears, which is accompanied by expulsion of the nitro group as the nitrite ion. This photosensitive effect was used to determine separately the damage of DNA induced by the reduced nitro group and the alkylating property of the aziridine. The aziridine-induced DNA damage is maximized in the dark when the nitro group is either absent (electrolytically reduced prior to the addition of DNA) or non functional (unreduced). In the light, damage is reduced. Typical DNA damage includes helix disruption leading to single strand breaks and the release of thymidine. Alkaline filter elution studies show evidence only for strand breakage and none for cross-linking indicating the drug is capable of mono-functional alkylation only.

Photosensitive interaction of RSU 1069 with DNA

International Nuclear Information System (INIS)

Edwards, D.I.; Knox, R.J.; Skolimowski, I.M.; Zahoor, A.; Knight, R.C.

1984-01-01

RSU 1069 is a 2-nitroimidazole radiosensitizer with an aziridine-containing side chain. In light (360 nm) the absorbance maximum of the nitro group at 325 nm disappears, which is accompanied by expulsion of the nitro group as the nitrite ion. This photosensitive effect was used to determine separately the damage of DNA induced by the reduced nitro group and the alkylating property of the aziridine. The aziridine-induced DNA damage is maximized in the dark when the nitro group is either absent (electrolytically reduced prior to the addition of DNA) or non functional (unreduced). In the light, damage is reduced. Typical DNA damage includes helix disruption leading to single strand breaks and the release of thymidine. Alkaline filter elution studies show evidence only for strand breakage and none for cross-linking indicating the drug is capable of mono-functional alkylation only

ATM is required for the repair of Topotecan-induced replication-associated double-strand breaks

International Nuclear Information System (INIS)

Köcher, Sabrina; Spies-Naumann, Anja; Kriegs, Malte; Dahm-Daphi, Jochen; Dornreiter, Irena

2013-01-01

Purpose: DNA replication is a promising target for anti-cancer therapies. Therefore, the understanding of replication-associated DNA repair mechanisms is of great interest. One key factor of DNA double-strand break (DSB) repair is the PIK kinase Ataxia-Telangiectasia Mutated (ATM) but it is still unclear whether ATM is involved in the repair of replication-associated DSBs. Here, we focused on the involvement of ATM in homology-directed repair (HDR) of indirect DSBs associated with replication. Material and methods: Experiments were performed using ATM-deficient and -proficient human cells. Replication-associated DSBs were induced with Topotecan (TPT) and compared with γ-irradiation (IR). Cell survival was measured by clonogenic assay. Overall DSB repair and HDR were evaluated by detecting residual γH2AX/53BP1 and Rad51 foci, respectively. Cell cycle distribution was analysed by flow cytometry and protein expression by Western blot. Results: ATM-deficiency leads to enhanced numbers of residual DSBs, resulting in a pronounced S/G2-block and decreased survival upon TPT-treatment. In common with IR, persisting Rad51 foci were detected following TPT-treatment. Conclusions: These results demonstrate that ATM is essentially required for the completion of HR-mediated repair of TPT-induced DSBs formed indirectly at replication forks

Region-specific chromatin decondensation and micronucleus formation induced by 5-azacytidine in human TIG-7 cells.

Science.gov (United States)

Satoh, T; Yamamoto, K; Miura, K F; Sofuni, T

2004-01-01

A human diploid lung fibroblast cell strain, TIG-7, has a heteromorphic chromosome 15 with an extra short arm carrying a homogeneously staining region (15p+hsr). We demonstrated previously that the 15p+hsr consists of an inactive and G+C-rich rDNA cluster characterized by fluorescence in situ hybridization (FISH) and various chromosome banding techniques. Thus, it was suggested that the region could contain highly methylated DNA. To observe methylation status on the target region directly under the microscope, we used a demethylating agent, 5-azacytidine (5-azaC), to induce decondensation of the chromatin containing methylated DNA. At 24 h after treatment with 0.5 microM 5-azaC, marked decondensation of the 15p+hsr was observed in almost all of the metaphases. Furthermore, we observed micronuclei, which were equivalent to the rDNA of the 15p+hsr demonstrated by FISH in the same preparation. In contrast, the DNA cross-linking agent mitomycin C (MMC) preferentially induced 15p+hsr-negative micronuclei. These findings indicated that chromatin decondensation and subsequent DNA strand breakage induced by the demethylating effect of 5-azaC led specifically to 15p+hsr-positive micronuclei. Copyright 2003 S. Karger AG, Basel

Radiation-induced DNA Double Strand Breaks and Their Modulations by Treatments with Moringa oleifera Lam. Leaf Extracts: A Cancer Cell Culture Model

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K. Boonsirichai

2014-04-01

Full Text Available Gamma radiation brings deleterious effects upon human cells by inducing oxidative stress and DNA damages. Antioxidants have been shown to confer protective effects on irradiated normal cells. Moringa oleifera Lam. is a widely used nutritional supplement with antioxidant activities. This report showed that antioxidant-containing supplements, in addition to protecting normal cells, could protect cancer cells against genotoxic effects of gamma radiation. -H2AX immunofluorescent foci were utilized as an indicator of radiation-induced DNA double strand breaks. MCF-7 human breast adenocarcinoma cells were irradiated with 2-8 Gy gamma radiation. A linear relationship between the formation of -H2AX foci and radiation dose was observed with an average of 10 foci per cell per Gy. A 30-minute pretreatment of the cells with either the aqueous or the ethanolic extract of M. oleifera leaves could partially protect the cells from radiation-induced DNA double strand breaks. A pretreatment with 500 µg/mL aqueous extract reduced the number of foci formed by 15% when assayed at 30 minutes post-irradiation. The ethanolic extract was more effective; 500 µg/mL of its concentration reduced the number of foci among irradiated cells by 30%. The results indicated that irradiated cancer cells responded similarly to nutritional supplements containing antioxidants as irradiated normal cells. These natural antioxidants could confer protective effects upon cancer cells against gamma radiation

Incorrect condom use and frequent breakage among female sex workers and their clients.

Science.gov (United States)

Mukenge-Tshibaka, Léonard; Alary, Michel; Geraldo, Nassirou; Lowndes, Catherine M

2005-05-01

Our objective was to assess if female sex workers (FSWs) and their potential male clients in Cotonou, Benin, know how to use male condoms correctly. From April to June 2000, 314 FSWs and 208 men were interviewed, and asked to demonstrate on a wooden penis how they usually use male condoms. In all, 27.6% of both women and men tore the condom envelope on the notch; 89.3% of the women versus 75.4% of the men easily found the correct side; 17.3% of the women versus 28.3% of the men held the top of the condom to avoid air entering; 91.4% of the women versus 75.6% of the men correctly unrolled the condom. Taking all the four criteria together, only approximately 11% of participants performed a correct condom use demonstration. FSWs frequently reported condom breakage, which was significantly associated with incorrect condom demonstration (P = 0.04). Correct condom use is suboptimal in these heavy consumers of male condoms in Benin. Condom breakage is frequent and is associated with incorrect use.

KARAKTERISTIK ORIENTED STRAND BOARD DARI KAYU AKASIA DAN AFRIKA BERDASARKAN PENYUSUNAN ARAH STRAND

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Nurhaida

2008-04-01

Full Text Available The research objectives arc to evaluate physical and mechanical properties of OSB based on strands orientation; and to evaluate physical and mechanical properties of OSB made from akasia wood (Acacia mangium Wild and afrika wood (Maesopsis eminii Engl. Akasia and afrika wood are used for OSB strand material with phenol formaldehyde (PF as adhesives and addition of paraffin. OSB made in this research is consist of three plies whereas are differed into eight (8 strand orientations. In the making process, hot press was carried out at 160OC and pressure 25kg.cm-2 for 15 minutes. Determination of OSB physical and mechanical properties is referred to JIS A 5908-2003. Result showed that strand orientations has no affect to OSB physical properties except for linicr swelling 24h, but it significantly influence all mechanical properties of OSB. Wood species have an effect on mechanical properties of OSB in the dry test, wet MOE lengthwise test and OSB physical properties, particularly to OSB density and water absorbing capability at 2h and 24h. All of OSB physical properties arc meet JIS A 5908-2003 standard, but not all of the mechanical properties such as dry MOE lengthwise, dry MOE and MOR widthwise. The best physical and mechanical properties is presented by OSB made from akasia wood in strand orientation F, G, Band C whereas all parameters meet JIS A 5908-2003 standard. In comparation with strand orientation B that is frequent used in industry, strand orientation F and G arc proficient to raise the modulus elasticity value (MOE and strength (MOR as much as 167.81-231.65% and 89.73-109.87%, respectively; especially in widthwise board application. Furthermore, strand orientation F and G arc more flexible as structural components

Mechanism of resistance of noncycling mammalian cells to 4'-(9-acridinylamino)methanesulfon-m-anisidide: comparison of uptake, metabolism, and DNA breakage in log- and plateau-phase Chinese hamster fibroblast cell cultures

International Nuclear Information System (INIS)

Robbie, M.A.; Baguley, B.C.; Denny, W.A.; Gavin, J.B.; Wilson, W.R.

1988-01-01

Resistance of noncycling cells to amsacrine (m-AMSA) has been widely reported and may limit the activity of this drug against solid tumors. The biochemical mechanism(s) for this resistance have been investigated using spontaneously transformed Chinese hamster fibroblasts (AA8 cells, a subline of Chinese hamster ovary-cells) in log- and plateau-phase spinner cultures. In early plateau phase most cells entered a growth-arrested state with a G1-G0 DNA content and showed a marked decrease in sensitivity to cytotoxicity induced by a 1-h exposure to m-AMSA or to its solid tumor-active analogue, CI-921. Studies with radiolabeled m-AMSA established that similar levels of drug were accumulated by log- and plateau-phase cells and that there was no significant drug metabolism in either of these cultures after 1 h. However, marked differences in sensitivity to m-AMSA-induced DNA breakage were observed using a fluorescence assay for DNA unwinding. Changes in sensitivity to DNA breakage occurred in parallel with changes in sensitivity to m-AMSA-induced cell killing. DNA breaks disappeared rapidly after drug removal (half-time approximately 4 min), suggesting that these lesions were probably mediated by DNA topoisomerase II. Resistance to m-AMSA may therefore be associated with changes in topoisomerase II activity in noncycling cells

Fibre Length Reduction in Natural Fibre-Reinforced Polymers during Compounding and Injection Moulding—Experiments Versus Numerical Prediction of Fibre Breakage

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Katharina Albrecht

2018-03-01

Full Text Available To establish injection-moulded, natural fibre-reinforced polymers in the automotive industry, numerical simulations are important. To include the breakage behaviour of natural fibres in simulations, a profound understanding is necessary. In this study, the length and width reduction of flax and sisal fibre bundles were analysed experimentally during compounding and injection moulding. Further an optical analysis of the fibre breakage behaviour was performed via scanning electron microscopy and during fibre tensile testing with an ultra-high-speed camera. The fibre breakage of flax and sisal during injection moulding was modelled using a micromechanical model. The experimental and simulative results consistently show that during injection moulding the fibre length is not reduced further; the fibre length was already significantly reduced during compounding. For the mechanical properties of a fibre-reinforced composite it is important to overachieve the critical fibre length in the injection moulded component. The micromechanical model could be used to predict the necessary fibre length in the granules.

A soluble RecN homologue provides means for biochemical and genetic analysis of DNA double-strand break repair in Escherichia coli.

Science.gov (United States)

Grove, Jane I; Wood, Stuart R; Briggs, Geoffrey S; Oldham, Neil J; Lloyd, Robert G

2009-12-03

RecN is a highly conserved, SMC-like protein in bacteria. It plays an important role in the repair of DNA double-strand breaks and is therefore a key factor in maintaining genome integrity. The insolubility of Escherichia coli RecN has limited efforts to unravel its function. We overcame this limitation by replacing the resident coding sequence with that of Haemophilus influenzae RecN. The heterologous construct expresses Haemophilus RecN from the SOS-inducible E. coli promoter. The hybrid gene is fully functional, promoting survival after I-SceI induced DNA breakage, gamma irradiation or exposure to mitomycin C as effectively as the native gene, indicating that the repair activity is conserved between these two species. H. influenzae RecN is quite soluble, even when expressed at high levels, and is readily purified. Its analysis by ionisation-mass spectrometry, gel filtration and glutaraldehyde crosslinking indicates that it is probably a dimer under physiological conditions, although a higher multimer cannot be excluded. The purified protein displays a weak ATPase activity that is essential for its DNA repair function in vivo. However, no DNA-binding activity was detected, which contrasts with RecN from Bacillus subtilis. RecN proteins from Aquifex aeolicus and Bacteriodes fragilis also proved soluble. Neither binds DNA, but the Aquifex RecN has weak ATPase activity. Our findings support studies indicating that RecN, and the SOS response in general, behave differently in E. coli and B. subtilis. The hybrid recN reported provides new opportunities to study the genetics and biochemistry of how RecN operates in E. coli.

Structural features of single-stranded integron cassette attC sites and their role in strand selection.

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Marie Bouvier

2009-09-01

Full Text Available We recently showed that cassette integration and deletion in integron platforms were occurring through unconventional site-specific recombination reactions involving only the bottom strand of attC sites. The lack of sequence conservation among attC sites led us to hypothesize that sequence-independent structural recognition determinants must exist within attC sites. The structural data obtained from a synaptic complex of the Vibrio cholerae integrase with the bottom strand of an attC site has shown the importance of extra helical bases (EHB inside the stem-loop structure formed from the bottom strand. Here, we systematically determined the contribution of three structural elements common to all known single-stranded attC site recombination substrates (the EHBs, the unpaired central spacer (UCS, and the variable terminal structure (VTS to strand choice and recombination. Their roles have been evaluated in vivo in the attIxattC reaction context using the suicide conjugation assay we previously developed, but also in an attCxattC reaction using a deletion assay. Conjugation was used to deliver the attC sites in single-stranded form. Our results show that strand choice is primarily directed by the first EHB, but the presence of the two other EHBs also serves to increase this strand selection. We found that the structure of the central spacer is essential to achieve high level recombination of the bottom strand, suggesting a dual role for this structure in active site exclusion and for hindering the reverse reaction after the first strand exchange. Moreover, we have shown that the VTS has apparently no role in strand selectivity.

Radiation-induced DNA damage in halogenated pyrimidine incorporated cells and its correlation with radiosensitivity

International Nuclear Information System (INIS)

Watanabe, R.; Nikjoo, H.

2003-01-01

Cells with DNA containing 5-halogenated pyrimidines in place of thymidine show significant reductions of slope (Do) and shoulder (Dq) of their radiation survival curves. Similar radiosensitization has also been observed in the yield of DNA strand breaks. The purpose of this study is to obtain an insight into the mechanism of cell lethality by examining the relationship between the spectrum of DNA damage and the cell survival. In this study we estimated the enhancement of strand breaks due to incorporation of halogenated pyrimidine, the complexity of DNA damage and the probability of the initial DNA damage leading to cell inactivation. Monte Carlo track structure methods were used to model and simulate the induction of strand breakage by X-rays. The increase of DNA strand break was estimated by assuming the excess strand break was caused by the highly reactive uracil radicals at the halouracil substituted sites. The assumption of the enhancement mechanism of strand breaks was examined and verified by comparison with experimental data for induction of SSB and DSB. The calculated DNA damage spectrum shows the increase in complexity of strand breaks is due to incorporation of halogenated pyrimidines. The increase in the yield of DSB and cell lethality show similar trend at various degrees of halogenated pyrimidine substitution. We asked the question whether this agreement supports the hypothesis that DSB is responsible for cell lethality? The estimated number of lethal damage from the cell survival using a linear-quadratic model is much less than the initial yield of DSB. This work examines the correlation of cell lethality as a function of frequencies of complex form of double strand breaks

Human lymphocytes exposed to low doses of ionizing radiations become refractory to high doses of radiation as well as to chemical mutagens that induce double-strand breaks in DNA

International Nuclear Information System (INIS)

Wolff, Sheldon; Afzal, Veena; Wiencke, J.K.; Olivieri, G.; Michaeli, A.

1988-01-01

The results indicate that prior exposure to 0.01 Gy of X-rays reduces the number of chromosome breaks induced by double-strand breaks, and perhaps even by cross-links, in DNA, but has the opposite effect on breaks induced by the alkylating agent MMS. The results also show that the induced repair mechanism is different from that observed in the adaptive reponse that follows exposure to low doses of alkylating agents. (author)

Chemical and structural composition of Atlantic Canadian moose (Alces alces) incisors with patterns of high breakage

Energy Technology Data Exchange (ETDEWEB)

MacKenzie, Cynthia S. Kendall; Clough, Michael J. [Biology Department, Saint Mary' s University, Halifax, Nova Scotia, Canada B3H 3C3 (Canada); Broders, Hugh G., E-mail: hugh.broders@smu.ca [Biology Department, Saint Mary' s University, Halifax, Nova Scotia, Canada B3H 3C3 (Canada); Tubrett, Mike [Inco Innovation Centre, ICP-MS Facility, Memorial University of Newfoundland, P.O. Box 4200, St. John' s, Newfoundland, Canada A1C 5S7 (Canada)

2011-11-15

Analysis of mammalian teeth can provide information regarding local environmental conditions. For example, a high incidence of breakage and wear within a population may indicate poor food quality. Individuals consuming a diet causing high mechanical stress on their teeth, and/or lacking the appropriate minerals for proper development, could experience degradation of tooth condition. Previously, we documented a high rate of incisor tooth breakage, with age, in two genetically distinct moose populations in Atlantic Canada. In this study, multi-element ({sup 11}B, {sup 63}Cu, {sup 64}Zn, {sup 75}As, {sup 85}Rb, {sup 88}Sr, {sup 111}Cd, {sup 118}Sn, {sup 137}Ba, {sup 208}Pb, {sup 232}Th, and {sup 238}U) analyses using laser ablation ICP-MS were performed on moose incisors from multiple North American regions. The purpose was to determine whether the elemental composition of moose incisors varies among regions, and whether that variation is related to tooth degradation among Atlantic Canadian populations. A principal components analysis revealed that nearly 50% of the elemental variation in the inner enamel matrix of moose teeth was explained by three groupings of elements. The element groupings revealed differences among geographic regions, but did not explain the variation between incisors that were broken and those that were not. Regression models indicate that the elemental group which includes Cu, Pb, and Zn is related to decreases in incisal integrity. It is likely that other environmental factors contribute to the occurrence of increased incisor breakage in affected populations. The relationship between food resource quantity and quality, as a function of moose density, is hypothesized to explain loss of tooth integrity. - Highlights: {yields} Multi-element analysis of 1300 moose incisors using laser ablation ICP-MS. {yields} Three element groupings explain 50% of tooth composition variation among regions. {yields} Regional differences in tooth composition

DNA double-strand breaks in blood lymphocytes induced by two-day 99mTc-MIBI myocardial perfusion scintigraphy.

Science.gov (United States)

Rief, Matthias; Hartmann, Lisa; Geisel, Dominik; Richter, Felicitas; Brenner, Winfried; Dewey, Marc

2018-07-01

To investigate DNA double-strand breaks (DSBs) in blood lymphocytes induced by two-day 99m Tc-MIBI myocardial perfusion scintigraphy (MPS) using y-H2AX immunofluorescence microscopy and to correlate the results with 99m Tc activity in blood samples. Eleven patients who underwent two-day MPS were included. DSB blood sampling was performed before and 5min, 1h and 24h after the first and second radiotracer injections. 99m Tc activity was measured in each blood sample. For immunofluorescence microscopy, distinct foci representing DSBs were quantified in lymphocytes after staining for the phosphorylated histone variant y-H2AX. The 99m Tc-MIBI activity measured on days one and two was similar (254±25 and 258±27 MBq; p=0.594). Compared with baseline DSB foci (0.09±0.05/cell), a significant increase was found at 5min (0.19±0.04/cell) and 1h (0.18±0.04/cell) after the first injection and at 5min and 1h after the second injection (0.21±0.03 and 0.19±0.04/cell, respectively; p=0.003 for both). At 24h after the first and second injections, the number of DSB foci had returned to baseline (0.06±0.02 and 0.12±0.05/cell, respectively). 99m Tc activity levels in peripheral blood samples correlated well with DSB counts (r=0.451). DSB counts reflect 99m Tc-MIBI activity after injection for two-day MPS, and might allow individual monitoring of biological effects of cardiac nuclear imaging. • Myocardial perfusion scintigraphy using 99m Tc induces time-dependent double-strand breaks (DSBs) • γ-H2AX immunofluorescence microscopy shows DSB as an early response to radiotracer injection • Activity measurements of 99m Tc correlate well with detected DSB • DSB foci induced by 99m Tc return to baseline 24h after radiotracer injection.

Five-Strand versus Four-Strand Hamstring Tendon Graft Technique for Anterior Cruciate Ligament Reconstruction: A Biomechanical Comparison.

Science.gov (United States)

Vaillant, Eric R; Parks, Brent G; Camire, Lyn M; Hinton, Richard Y

2017-11-01

The aim of this article is to compare diameter and stiffness, displacement, and strain in a five-strand versus four-strand hamstring graft for anterior cruciate ligament reconstruction. Eight matched pairs of lower extremities underwent four-strand or five-strand hamstring graft reconstruction. Diameter was significantly higher in the five-strand versus the four-strand construct ( p  = 0.002). No significant difference was found between the groups in construct displacement or stiffness. Significantly higher strain was observed in the inner limb versus the outer limb in the four-strand construct ( p  = 0.001) and in the inner limb versus the fifth limb in the 5-strand construct ( p  = 0.004). A fifth limb added to a four-strand hamstring graft significantly increased graft diameter but did not significantly change stiffness or displacement, suggesting that attachment of additional graft material via suture did not provide for full incorporation of the added limb into the graft at time zero. The inner limb in both constructs absorbed significantly greater load than did other limbs. The use of suture to attach additional material to a four-strand hamstring graft may not contribute to improved biomechanical qualities of the graft at time zero. Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.

Modelling Toehold-Mediated RNA Strand Displacement

OpenAIRE

Šulc, Petr; Ouldridge, Thomas E.; Romano, Flavio; Doye, Jonathan P.K.; Louis, Ard A.

2015-01-01

We study the thermodynamics and kinetics of an RNA toehold-mediated strand displacement reaction with a recently developed coarse-grained model of RNA. Strand displacement, during which a single strand displaces a different strand previously bound to a complementary substrate strand, is an essential mechanism in active nucleic acid nanotechnology and has also been hypothesized to occur in vivo. We study the rate of displacement reactions as a function of the length of the toehold and temperat...

Excess single-stranded DNA inhibits meiotic double-strand break repair.

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Rebecca Johnson

2007-11-01

Full Text Available During meiosis, self-inflicted DNA double-strand breaks (DSBs are created by the protein Spo11 and repaired by homologous recombination leading to gene conversions and crossovers. Crossover formation is vital for the segregation of homologous chromosomes during the first meiotic division and requires the RecA orthologue, Dmc1. We analyzed repair during meiosis of site-specific DSBs created by another nuclease, VMA1-derived endonuclease (VDE, in cells lacking Dmc1 strand-exchange protein. Turnover and resection of the VDE-DSBs was assessed in two different reporter cassettes that can repair using flanking direct repeat sequences, thereby obviating the need for a Dmc1-dependent DNA strand invasion step. Access of the single-strand binding complex replication protein A, which is normally used in all modes of DSB repair, was checked in chromatin immunoprecipitation experiments, using antibody against Rfa1. Repair of the VDE-DSBs was severely inhibited in dmc1Delta cells, a defect that was associated with a reduction in the long tract resection required to initiate single-strand annealing between the flanking repeat sequences. Mutants that either reduce Spo11-DSB formation or abolish resection at Spo11-DSBs rescued the repair block. We also found that a replication protein A component, Rfa1, does not accumulate to expected levels at unrepaired single-stranded DNA (ssDNA in dmc1Delta cells. The requirement of Dmc1 for VDE-DSB repair using flanking repeats appears to be caused by the accumulation of large quantities of ssDNA that accumulate at Spo11-DSBs when Dmc1 is absent. We propose that these resected DSBs sequester both resection machinery and ssDNA binding proteins, which in wild-type cells would normally be recycled as Spo11-DSBs repair. The implication is that repair proteins are in limited supply, and this could reflect an underlying mechanism for regulating DSB repair in wild-type cells, providing protection from potentially harmful effects

Enhancing repair of radiation-induced strand breaks in cellular DNA as a radiotherapeutic potential

International Nuclear Information System (INIS)

Nair, C.K.K.

2014-01-01

Protection of mammalian organisms including man from deleterious effects of ionizing radiation is of paramount importance and development of effective approaches to combat radiation damages using non-toxic radioprotectors is of considerable interest for defence, nuclear industries, radiation accidents, space travels, etc., besides the protection of normal tissues during radiotherapy of tumours. Many synthetic as well as natural compounds have been investigated in the recent past for their efficacy to protect the biological systems from radiation induced damages. They include sulfhydryl compounds, antioxidants, plant extracts, immune-modulators, and other agents. However, the inherent toxicity of many of the synthetic agents at the effective radio-protective concentration warranted further search for safer and more effective radio-protectors. In this context, therapeutic radioprotectors which are effective on post irradiation administration are of special relevance. One of the property that can be applied while screening for such radiation protective therapeutics is their ability to enhance repair of radiation-induced lesions in cellular DNA in terms of cellular repair index based on the parameters of the DNA following comet assay. Post irradiation administration of some natural and synthetic agents have shown their potential to enhance repair of radiation-induced strand breaks in cellular DNA in mice. These include phytoceuticals such as gallic acid, sesamol etc., extracts of medicinal plants such as Andrographis panniculata, and a few synthetic compounds such as tocopherol-mono-glucoside. The talk will give an overview of the work on DNA repair enhancement by a few natural and synthetic agents. (author)

Observed mechanism for the breakup of small bundles of cellulose Iα and Iβ in ionic liquids from molecular dynamics simulations.

Science.gov (United States)

Rabideau, Brooks D; Agarwal, Animesh; Ismail, Ahmed E

2013-04-04

Explicit, all-atom molecular dynamics simulations are used to study the breakup of small bundles of cellulose Iα and Iβ in the ionic liquids [BMIM]Cl, [EMIM]Ac, and [DMIM]DMP. In all cases, significant breakup of the bundles is observed with the initial breakup following a common underlying mechanism. Anions bind strongly to the hydroxyl groups of the exterior strands of the bundle, forming negatively charged complexes. Binding also weakens the intrastrand hydrogen bonds present in the cellulose strands, providing greater strand flexibility. Cations then intercalate between the individual strands, likely due to charge imbalances, providing the bulk to push the individual moieties apart and initiating the separation. The peeling of an individual strand from the main bundle is observed in [EMIM]Ac with an analysis of its hydrogen bonds with other strands showing that the chain detaches glucan by glucan from the main bundle in discrete, rapid events. Further analysis shows that the intrastrand hydrogen bonds of each glucan tend to break for a sustained period of time before the interstrand hydrogen bonds break and strand detachment occurs. Examination of similar nonpeeling strands shows that, without this intrastrand hydrogen bond breakage, the structural rigidity of the individual unit can hinder its peeling despite interstrand hydrogen bond breakage.

R&D on dental implants breakage

Science.gov (United States)

Croitoru, Sorin Mihai; Popovici, Ion Alexandru

2017-09-01

Most used dental implants for human dental prostheses are of two steps type: first step means implantation and, after several months healing and osseointegration, second step is prosthesis fixture. For sure, dental implants and prostheses are meant to last for a lifetime. Still, there are unfortunate cases when dental implants break. This paper studies two steps dental implants breakage and proposes a set of instruments for replacement and restoration of the broken implant. First part of the paper sets the input data of the study: structure of the studied two steps dental implants based on two Romanian patents and values of the loading forces found in practice and specialty papers. In the second part of the paper, using DEFORM 2D™ FEM simulation software, worst case scenarios of loading dental implants are studied in order to determine which zones and components of the dental implant set are affected (broken). Last part of the paper is dedicated to design and presentation of a set for extracting and cutting tools used to restore the broken implant set.

PI3K-delta mediates double-stranded RNA-induced upregulation of B7-H1 in BEAS-2B airway epithelial cells

International Nuclear Information System (INIS)

Kan-o, Keiko; Matsumoto, Koichiro; Asai-Tajiri, Yukari; Fukuyama, Satoru; Hamano, Saaka; Seki, Nanae; Nakanishi, Yoichi; Inoue, Hiromasa

2013-01-01

Highlights: •Double-stranded RNA upregulates B7-H1 on BEAS-2B airway epithelial cells. •The upregulation of B7-H1 is attenuated by inhibition of PI3Kδ isoform. •PI3Kδ-mediated upregulation of B7-H1 is independent of NF-κB activation. •Inhibition of PI3Kδ may prevent persistent viral infection induced by B7-H1. -- Abstract: Airway viral infection disturbs the health-related quality of life. B7-H1 (also known as PD-L1) is a coinhibitory molecule associated with the escape of viruses from the mucosal immunity, leading to persistent infection. Most respiratory viruses generate double-stranded (ds) RNA during replication. The stimulation of cultured airway epithelial cells with an analog of viral dsRNA, polyinosinic-polycytidylic acid (poly IC) upregulates the expression of B7-H1 via activation of the nuclear factor κB(NF-κB). The mechanism of upregulation was investigated in association with phosphatidylinositol 3-kinases (PI3Ks). Poly IC-induced upregulation of B7-H1 was profoundly suppressed by a pan-PI3K inhibitor and partially by an inhibitor or a small interfering (si)RNA for PI3Kδ in BEAS-2B cells. Similar results were observed in the respiratory syncytial virus-infected cells. The expression of p110δ was detected by Western blot and suppressed by pretreatment with PI3Kδ siRNA. The activation of PI3Kδ is typically induced by oxidative stress. The generation of reactive oxygen species was increased by poly IC. Poly IC-induced upregulation of B7-H1 was attenuated by N-acetyl-L-cysteine, an antioxidant, or by oxypurinol, an inhibitor of xanthine oxidase. Poly IC-induced activation of NF-κB was suppressed by a pan-PI3K inhibitor but not by a PI3Kδ inhibitor. These results suggest that PI3Kδ mediates dsRNA-induced upregulation of B7-H1 without affecting the activation of NF-κB

Mutation inactivation of Nijmegen breakage syndrome gene (NBS1 in hepatocellular carcinoma and intrahepatic cholangiocarcinoma.

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Yan Wang

Full Text Available Nijmegen breakage syndrome (NBS with NBS1 germ-line mutation is a human autosomal recessive disease characterized by genomic instability and enhanced cancer predisposition. The NBS1 gene codes for a protein, Nbs1(p95/Nibrin, involved in the processing/repair of DNA double-strand breaks. Hepatocellular carcinoma (HCC is a complex and heterogeneous tumor with several genomic alterations. Recent studies have shown that heterozygous NBS1 mice exhibited a higher incidence of HCC than did wild-type mice. The objective of the present study is to assess whether NBS1 mutations play a role in the pathogenesis of human primary liver cancer, including HBV-associated HCC and intrahepatic cholangiocarcinoma (ICC. Eight missense NBS1 mutations were identified in six of 64 (9.4% HCCs and two of 18 (11.1% ICCs, whereas only one synonymous mutation was found in 89 control cases of cirrhosis and chronic hepatitis B. Analysis of the functional consequences of the identified NBS1 mutations in Mre11-binding domain showed loss of nuclear localization of Nbs1 partner Mre11, one of the hallmarks for Nbs1 deficiency, in one HCC and two ICCs with NBS1 mutations. Moreover, seven of the eight tumors with NBS1 mutations had at least one genetic alteration in the TP53 pathway, including TP53 mutation, MDM2 amplification, p14ARF homozygous deletion and promoter methylation, implying a synergistic effect of Nbs1 disruption and p53 inactivation. Our findings provide novel insight on the molecular pathogenesis of primary liver cancer characterized by mutation inactivation of NBS1, a DNA repair associated gene.

Coronavirus minus-strand RNA synthesis and effect of cycloheximide on coronavirus RNA synthesis

International Nuclear Information System (INIS)

Sawicki, S.G.; Sawicki, D.L.

1986-01-01

The temporal sequence of coronavirus plus-strand and minus-strand RNA synthesis was determined in 17CL1 cells infected with the A59 strain of mouse hepatitis virus (MHV). MHV-induced fusion was prevented by keeping the pH of the medium below pH 6.8. This had no effect on the MHV replication cycle, but gave 5- to 10-fold-greater titers of infectious virus and delayed the detachment of cells from the monolayer which permitted viral RNA synthesis to be studied conveniently until at least 10 h postinfection. Seven species of poly(A)-containing viral RNAs were synthesized at early and late times infection, in nonequal but constant ratios. MHV minus-strand RNA synthesis was first detected at about 3 h after infection and was found exclusively in the viral replicative intermediates and was not detected in 60S single-stranded form in infected cells. Early in the replication cycle, from 45 to 65% of the [ 3 H]uridine pulse-labeled RF core of purified MHV replicative intermediates was in minus-strand RNA. The rate of minus-strand synthesis peaked at 5 to 6 h postinfection and then declined to about 20% of the maximum rate. The addition of cycloheximide before 3 h postinfection prevented viral RNA synthesis, whereas the addition of cycloheximide after viral RNA synthesis had begun resulted in the inhibition of viral RNA synthesis. The synthesis of both genome and subgenomic mRNAs and of viral minus strands required continued protein synthesis, and minis-strand RNA synthesis was three- to fourfold more sensitive to inhibition of cycloheximide than was plus-strand synthesis

Immediate and repair induced DNA double strand breaks in mammalian cells

International Nuclear Information System (INIS)

Bryant, P.E.

1986-01-01

It seems logical to postulate that double strand breaks (dsb) arising both at the time of irradiation and via repair processes are potentially equally damaging for a cell in terms of the potential to induce chromosomal aberrations. However, in some cell systems the repair of double es or es-ssb sites may run concurrently with the incision so that these lesions do not remain open for long: hence the lack of accumulation of dsb during repair. The rate of incision will thus determine both the accumulation and the probability of exchanges leading to chromosomal aberrations between these and other frank dsb. Rapid incision leading to a large additional pool of dsb appears to be the case in Chinese hamster V79 cells. Some evidence also exists for the conversion of base damage, via dsb, into deletion type chromatid aberrations which accumulate in irradiated G2 human cells treated with ara C. A small fraction of dsb, probably arising both at the time of irradiation as well as enzymatically during repair of base or sugar damage, appears to be either left unrepaired, yielding deletion type chromosomal aberrations, or is misrepaired, yielding exchange aberrations. The induction of these aberrations appears to be of central importance in the biological effects of ionizing radiation such as mutations, oncogenic transformation, and cell death. 52 refs., 5 figs

Carboplatin enhances the production and persistence of radiation-induced DNA single-strand breaks

International Nuclear Information System (INIS)

Yang, L.; Douple, E.B.; O'Hara, J.A.; Wang, H.J.

1995-01-01

Fluorometric analysis of DNA unwinding and alkaline elution were used to investigate the production and persistence of DNA single-strand breaks (SSBs) in Chinese hamster V79 and xrs-5 cells treated with the chemotherapeutic agent carboplatin in combination with radiation. Carboplatin was administered to cells before irradiation in hypoxic conditions, or the drug was added immediately after irradiation during the postirradiation recovery period in air. The results of DNA unwinding studies suggest that carboplatin enhances the production of radiation-induced SSBs in hypoxic V79 cells and xrs-5 cells by a factor of 1.86 and 1.83, respectively, when combined with radiation compared to the SSBs produced by irradiation alone. Carboplatin alone did not produce a measureable number of SSBs. Alkaline elution profiles also indicated that the rate of elution of SSBs was higher in cells treated with the carboplatin is present after irradiation and during the postirradiation recovery period, the rejoining of radiation-induced SSBs by a factor of 1.46 in V79 cells with 20 Gy irradiation and by a factor of 2.02 in xrs-5 cells with 20 Gy irradiation. When carboplatin is present after irradiation and during the postirradiation recovery period, the rejoining of radiation-induced SSBs is inhibited during this postirradiation incubation period (radiopotentiation) with a relative inhibition factor at 1 h postirradiation of 1.25 in V79 cells and 1.15 in xrs-5 cells. An increased production and persistence of SSBs resulting from the interaction of carboplatin with radiation may be an important step in the mechanism responsible for the potentiated cell killing previously from studies in animal tumors and in cultured cells. 31 refs., 7 figs

Condom Use: Slippage, Breakage, and Steps for Proper Use among Adolescents in Alternative School Settings

Science.gov (United States)

Coyle, Karin K.; Franks, Heather M.; Glassman, Jill R.; Stanoff, Nicole M.

2012-01-01

Background: School-based human immunodeficiency virus (HIV)/sexually transmitted infection (STI), and pregnancy prevention programs often focus on consistent and correct condom use. Research on adolescents' experience using condoms, including condom slippage/breakage, is limited. This exploratory study examines proper condom use and the…

DNA strand breaks induced by electrons simulated with nanodosimetry Monte Carlo simulation code: NASIC

International Nuclear Information System (INIS)

Li, Junli; Qiu, Rui; Yan, Congchong; Xie, Wenzhang; Zeng, Zhi; Li, Chunyan; Wu, Zhen; Tung, Chuanjong

2015-01-01

The method of Monte Carlo simulation is a powerful tool to investigate the details of radiation biological damage at the molecular level. In this paper, a Monte Carlo code called NASIC (Nanodosimetry Monte Carlo Simulation Code) was developed. It includes physical module, pre-chemical module, chemical module, geometric module and DNA damage module. The physical module can simulate physical tracks of low-energy electrons in the liquid water event-by-event. More than one set of inelastic cross sections were calculated by applying the dielectric function method of Emfietzoglou's optical-data treatments, with different optical data sets and dispersion models. In the pre-chemical module, the ionised and excited water molecules undergo dissociation processes. In the chemical module, the produced radiolytic chemical species diffuse and react. In the geometric module, an atomic model of 46 chromatin fibres in a spherical nucleus of human lymphocyte was established. In the DNA damage module, the direct damages induced by the energy depositions of the electrons and the indirect damages induced by the radiolytic chemical species were calculated. The parameters should be adjusted to make the simulation results be agreed with the experimental results. In this paper, the influence study of the inelastic cross sections and vibrational excitation reaction on the parameters and the DNA strand break yields were studied. Further work of NASIC is underway (authors)

Comparison of DNA strand-break simulated with different DNA models

International Nuclear Information System (INIS)

Xie, Wenzhang; Li, Junli; Qiu, Rui; Yan, Congchong; Zeng, Zhi; Li, Chunyan

2013-01-01

Full text of the publication follows. In Monte Carlo simulation of DNA damage, the geometric model of DNA is of great importance. To study the influence of DNA model on the simulation of DNA damage, three DNA models were created in this paper. They were a volume model and two atomic models with different parameters. Direct DNA strand-break induced by low-energy electrons were simulated respectively with the three models. The results show that most of the energy depositions in the DNA segments do not lead to strand-breaks. The simple single strand-break (SSB) tends to be the predominant damage type, and the contribution of complex double strand-break (DSB) to the total DSB cannot be neglected. Among the yields of all the three DNA target models applied here, the yields of the volume model are the highest, the yields of the atomic model with double van der Waals radii (r) take the second place, whereas the yields of the atomic model with single r come last. On average, the ratios of SSB yields are approximately equivalent to the corresponding ratios of the models' volume. However, there seems to be no clear relationship between the DSB yields and the models' volume. (authors)

Inhibition of X-ray induced DNA strand break repair in polyamine-depleted HeLa cells

Energy Technology Data Exchange (ETDEWEB)

Snyder, R.D.

1989-05-01

Treatment of HeLa cells with the polyamine biosynthesis inhibitors, alpha-difluoromethylornithine (DFMO) or methylglyoxal bis(guanylhydrazone) (MGBG), results in, depending on the conditions, partial or complete depletion of the cellular polyamines: putrescine, spermidine and spermine. In this compromised state cells exhibited a distinct deficiency in repair of X-ray-induced DNA strand breaks. The half-time for return of normal DNA sedimentation following 1.6 Gy was 9.5 min for untreated control cells and 22, 32 and 50 min for cells treated with MGBG, DFMO+MGBG and DFMO, respectively. Normal repair kinetics were restored to these cells upon a short incubation in media containing all three polyamines. The rapid early phase of repair following higher X-ray doses (16 Gy) was also delayed in polyamine-depleted cells but later repair occurring 1-4 h post-irradiation, representing chromatin reconstitution, was apparently normal. (author).

Inhibition of X-ray induced DNA strand break repair in polyamine-depleted HeLa cells

International Nuclear Information System (INIS)

Snyder, R.D.

1989-01-01

Treatment of HeLa cells with the polyamine biosynthesis inhibitors, alpha-difluoromethylornithine (DFMO) or methylglyoxal bis(guanylhydrazone) (MGBG), results in, depending on the conditions, partial or complete depletion of the cellular polyamines: putrescine, spermidine and spermine. In this compromised state cells exhibited a distinct deficiency in repair of X-ray-induced DNA strand breaks. The half-time for return of normal DNA sedimentation following 1.6 Gy was 9.5 min for untreated control cells and 22, 32 and 50 min for cells treated with MGBG, DFMO+MGBG and DFMO, respectively. Normal repair kinetics were restored to these cells upon a short incubation in media containing all three polyamines. The rapid early phase of repair following higher X-ray doses (16 Gy) was also delayed in polyamine-depleted cells but later repair occurring 1-4 h post-irradiation, representing chromatin reconstitution, was apparently normal. (author)

Radiation-induced strand-breaks and DNA-protein crosslinks depend predominantly on the dose, oxygen concentration and repair time

International Nuclear Information System (INIS)

Wheeler, K.T.; Miyagi, Y.; Zhang, H.

1995-01-01

It has been known for many years that the DNA damage produced by ionizing radiation depends upon the oxygen concentration around the DNA. For example, the number of DNA strand-breaks (SBs) formed per unit dose decreases at low oxygen concentrations, and the number of DNA-protein crosslinks formed per unit dose increases at low oxygen concentrations. If radiation-induced SBs and DPCs are to be useful for detecting and/or quantifying hypoxic cells in solid tumors, the formation of these lesions must depend predominantly on the oxygen concentration around the DNA. All other physical, biological, and physiological factors must either be controllable or have little influence on the assay used to measure these lesions. This paper is a summary of the authors' recent experiments to determine if the radiation-induced SBs and DPCs measured by alkaline elution may be used to estimate the hypoxic fraction or fractional hypoxic volume of solid tumors

DNA strand breaks detected in embryos of the adult snails, Potamopyrgus antipodarum, and in neonates exposed to genotoxic chemicals

International Nuclear Information System (INIS)

Vincent-Hubert, Françoise; Revel, Messika; Garric, Jeanne

2012-01-01

We tested the freshwater mudsnail Potamopyrgus antipodarum, which is a species that has already been used for endocrine-disrupting compounds (EDCs) to determine whether early life stages of aquatic organisms are sensitive to genotoxic chemicals. For this purpose, we first developed the alkaline comet assay on adults, embryos, and neonates. The comet assay protocol was validated on both embryonic cells exposed in vitro to hydrogen peroxide and adult snails in the reproducing stage exposed to methyl methane sulfonate. During the latter experiment, DNA strand breaks were investigated on both embryonic cells and on adult gill cells. The second part of this study investigated the stability of DNA strand breaks in adult reproducing snails and neonates exposed to cadmium (Cd) and bisphenol A for 8 days. Hydrogen peroxide-induced DNA strand breaks in vitro in isolated embryonic cells. Exposure of adult reproducing snails to methyl methane sulfonate for 24 h induced DNA strand breaks in embryos. Bisphenol A induced a significant increase in the DNA strand-break level in whole embryonic cells and whole neonate cells. Cd was genotoxic for both embryos and neonates during the exposure time and also after 7 days of depuration, suggesting that Cd could inhibit DNA repair enzymes. These preliminary results on this original model have encouraged us to consider the impact of genotoxic environmental contaminants on the F1 generation.

DNA strand breaks detected in embryos of the adult snails, Potamopyrgus antipodarum, and in neonates exposed to genotoxic chemicals

Energy Technology Data Exchange (ETDEWEB)

Vincent-Hubert, Francoise, E-mail: francoise.vincent-hubert@irstea.fr [Unite de Recherche Hydrosystemes et Bioprocedes, equipe BELCA, IRSTEA/CEMAGREF, 1 rue Pierre-Gilles de Gennes, CS10030, 92761 Antony cedex, 92163 Antony (France); Revel, Messika [Unite de Recherche Hydrosystemes et Bioprocedes, equipe BELCA, IRSTEA/CEMAGREF, 1 rue Pierre-Gilles de Gennes, CS10030, 92761 Antony cedex, 92163 Antony (France); Garric, Jeanne [MALY Laboratoire d' ecotoxicologie, IRSTEA/CEMAGREF, 23 bis Quai Chauveau, 69006 Lyon (France)

2012-10-15

We tested the freshwater mudsnail Potamopyrgus antipodarum, which is a species that has already been used for endocrine-disrupting compounds (EDCs) to determine whether early life stages of aquatic organisms are sensitive to genotoxic chemicals. For this purpose, we first developed the alkaline comet assay on adults, embryos, and neonates. The comet assay protocol was validated on both embryonic cells exposed in vitro to hydrogen peroxide and adult snails in the reproducing stage exposed to methyl methane sulfonate. During the latter experiment, DNA strand breaks were investigated on both embryonic cells and on adult gill cells. The second part of this study investigated the stability of DNA strand breaks in adult reproducing snails and neonates exposed to cadmium (Cd) and bisphenol A for 8 days. Hydrogen peroxide-induced DNA strand breaks in vitro in isolated embryonic cells. Exposure of adult reproducing snails to methyl methane sulfonate for 24 h induced DNA strand breaks in embryos. Bisphenol A induced a significant increase in the DNA strand-break level in whole embryonic cells and whole neonate cells. Cd was genotoxic for both embryos and neonates during the exposure time and also after 7 days of depuration, suggesting that Cd could inhibit DNA repair enzymes. These preliminary results on this original model have encouraged us to consider the impact of genotoxic environmental contaminants on the F1 generation.

Alkaline gel electrophoresis assay to detect DNA strand breaks and repair mechanisms in Escherichia coli

International Nuclear Information System (INIS)

Mattos, Jose Carlos Pelielo de; Motta, Ellen Serri da; Oliveira, Marcia Betania Nunes de; Dantas, Flavio Jose da Silva; Araujo, Adriano Caldeira de

2008-01-01

Reactive oxygen species (ROS) can induce lesions in different cellular targets, including DNA. Stannous chloride (SnCl 2 ) is a ROS generator, leading to lethality in Escherichia coli (E. coli), with the base excision repair (BER) mechanism playing a role in this process. Many techniques have been developed to detect genotoxicity, as comet assay, in eukaryotic cells, and plasmid DNA agarose gel electrophoresis. In this study, an adaptation of the alkaline gel electrophoresis method was carried out to ascertain the induction of strand breaks by SnCl 2 in bacterial DNA, from E. coli BER mutants, and its repair pathway. Results obtained show that SnCl 2 was able to induce DNA strand breaks in all strains tested. Moreover, endonuclease IV and exonuclease III play a role in DNA repair. On the whole, data has shown that the alkaline gel electrophoresis assay could be used both for studying DNA strand breaks induction and for associated repair mechanisms. (author)

Alkaline gel electrophoresis assay to detect DNA strand breaks and repair mechanisms in Escherichia coli

Energy Technology Data Exchange (ETDEWEB)

Mattos, Jose Carlos Pelielo de; Motta, Ellen Serri da; Oliveira, Marcia Betania Nunes de; Dantas, Flavio Jose da Silva; Araujo, Adriano Caldeira de [Universidade do Estado do Rio de Janeiro (UERJ), RJ (Brazil). Dept. de Biofisica e Biometria. Lab. de Radio e Fotobiologia]. E-mail: jcmattos@uerj.br

2008-12-15

Reactive oxygen species (ROS) can induce lesions in different cellular targets, including DNA. Stannous chloride (SnCl{sub 2}) is a ROS generator, leading to lethality in Escherichia coli (E. coli), with the base excision repair (BER) mechanism playing a role in this process. Many techniques have been developed to detect genotoxicity, as comet assay, in eukaryotic cells, and plasmid DNA agarose gel electrophoresis. In this study, an adaptation of the alkaline gel electrophoresis method was carried out to ascertain the induction of strand breaks by SnCl{sub 2} in bacterial DNA, from E. coli BER mutants, and its repair pathway. Results obtained show that SnCl{sub 2} was able to induce DNA strand breaks in all strains tested. Moreover, endonuclease IV and exonuclease III play a role in DNA repair. On the whole, data has shown that the alkaline gel electrophoresis assay could be used both for studying DNA strand breaks induction and for associated repair mechanisms. (author)

Variation in normal and tumor tissue sensitivity of mice to ionizing radiation-induced DNA strand breaks in vivo

International Nuclear Information System (INIS)

Meyn, R.E.; Jenkins, W.T.

1983-01-01

The efficiency of DNA strand break formation in normal and tumor tissues of mice was measured using the technique of alkaline elution coupled with a microfluorometric determination of DNA. This methodology allowed measurement of the DNA strand breaks produced in tissues irradiated in vivo with doses of radiation comparable to those used in radiotherapy (i.e., 1.0 gray) without the necessity for the cells to be dividing and incorporating radioactive precursors to label the DNA. The results showed that substantial differences existed among various tissues in terms of the amount of DNA strand break damage produced for a given dose of radiation. Of the normal tissues, the most breaks were produced in bone marrow and the least were produced in gut. Furthermore, strand break production was relatively inefficient in the tumor compared to the normal tissues. The efficiency of DNA strand break formation measured in the cells from the tissues irradiated in vitro was much more uniform and considerably greater than that measured in vivo, suggesting that the normal tissues in the animal may be radiobiologically hypoxic

The opportunistic pathogen Pseudomonas aeruginosa activates the DNA double-strand break signaling and repair pathway in infected cells

International Nuclear Information System (INIS)

Elsen, S.; Collin-Faure, V.; Gidrol, X.; Lemercier, C.

2013-01-01

Highly hazardous DNA double-strand breaks can be induced in eukaryotic cells by a number of agents including pathogenic bacterial strains. We have investigated the genotoxic potential of Pseudomonas aeruginosa, an opportunistic pathogen causing devastating nosocomial infections in cystic fibrosis or immunocompromised patients. Our data revealed that infection of immune or epithelial cells by P. aeruginosa triggered DNA strand breaks and phosphorylation of histone H2AX (γH2AX), a marker of DNA double-strand breaks. Moreover, it induced formation of discrete nuclear repair foci similar to gamma-irradiation-induced foci, and containing γH2AX and 53BP1, an adaptor protein mediating the DNA-damage response pathway. Gene deletion, mutagenesis, and complementation in P. aeruginosa identified ExoS bacterial toxin as the major factor involved in γH2AX induction. Chemical inhibition of several kinases known to phosphorylate H2AX demonstrated that Ataxia Telangiectasia Mutated (ATM) was the principal kinase in P. aeruginosa-induced H2AX phosphorylation. Finally, infection led to ATM kinase activation by an auto-phosphorylation mechanism. Together, these data show for the first time that infection by P. aeruginosa activates the DNA double-strand break repair machinery of the host cells. This novel information sheds new light on the consequences of P. aeruginosa infection in mammalian cells. As pathogenic Escherichia coli or carcinogenic Helicobacter pylori can alter genome integrity through DNA double-strand breaks, leading to chromosomal instability and eventually cancer, our findings highlight possible new routes for further investigations of P. aeruginosa in cancer biology and they identify ATM as a potential target molecule for drug design. (authors)

Effect of vanillin on methylene blue plus light-induced single-strand breaks in plasmid pBR322 DNA.

Science.gov (United States)

Kumar, S S; Ghosh, A; Devasagayam, T P; Chauhan, P S

2000-09-20

The ability of vanillin (4-hydroxy-3-methoxybenzaldehyde), a naturally occurring food flavouring agent, in inhibiting photosensitization-induced single-strand breaks (ssbs) in plasmid pBR322 DNA has been examined in an in vitro system, independent of DNA repair/replication processes. Photosensitization of DNA with methylene blue, visible light and oxygen, induced ssbs resulting in the production of open circular form (OC form) in a concentration-dependent manner. The yield of OC form induced by photosensitization was increased several-fold by deuteration of the buffer and was found to be inhibited by sodium azide, a scavenger of singlet oxygen (1O(2)). Vanillin, per se, did not induce but inhibited photosensitization-induced ssbs in plasmid DNA, at millimolar concentrations. The inhibitory effect of vanillin was both concentration- and time-dependent. On a molar basis, vanillin was, however, less effective than trolox, a water-soluble analogue of alpha-tocopherol. Photosensitization by methylene blue system generates singlet oxygen, as one of the major components of ROS. Therefore, interaction of singlet oxygen with vanillin was investigated. The rate constant of vanillin with 1O(2) was estimated to be 5.93x10(7)M(-1)s(-1) and that of sodium azide as 2. 7x10(8)M(-1)s(-1). The present investigations show that vanillin can protect against photosensitization-induced ssbs in the plasmid pBR322 DNA, and this effect may partly be due to its ability to scavenge 1O(2).

Hair breakage as a presenting sign of early or occult central centrifugal cicatricial alopecia: clinicopathologic findings in 9 patients.

Science.gov (United States)

Callender, Valerie D; Wright, Dakara Rucker; Davis, Erica C; Sperling, Leonard C

2012-09-01

Central centrifugal cicatricial alopecia is the most common form of cicatricial alopecia in African American women. Treatment options are limited and mostly aimed at halting further hair loss but rarely result in hair regrowth. Therefore, it is important to recognize early clinical signs, perform a confirmatory biopsy, and begin treatment promptly. We have observed that hair breakage may be a key sign of early central centrifugal cicatricial alopecia, and this association is not clearly described in the literature. Nine patients with hair breakage on the vertex with or without scalp symptoms underwent scalp biopsies as part of their evaluation. Of these, 8 had histologic samples adequate for complete interpretation: 5 specimens (63%) showed histologic changes typical of central centrifugal cicatricial alopecia, with 1 of these showing advanced end-stage changes of cicatricial alopecia. Two (25%) revealed premature desquamation of the inner root sheath as the sole finding suggestive of early central centrifugal cicatricial alopecia and 1 (13%) was normal. Although hair breakage can have multiple causes, early central centrifugal cicatricial alopecia must be considered in the differential diagnosis, particularly in women of African ancestry. Histologic evaluation may reveal early or late findings that can help establish the diagnosis.

Exposure to 1800 MHz radiofrequency electromagnetic radiation induces oxidative DNA base damage in a mouse spermatocyte-derived cell line.

Science.gov (United States)

Liu, Chuan; Duan, Weixia; Xu, Shangcheng; Chen, Chunhai; He, Mindi; Zhang, Lei; Yu, Zhengping; Zhou, Zhou

2013-03-27

Whether exposure to radiofrequency electromagnetic radiation (RF-EMR) emitted from mobile phones can induce DNA damage in male germ cells remains unclear. In this study, we conducted a 24h intermittent exposure (5 min on and 10 min off) of a mouse spermatocyte-derived GC-2 cell line to 1800 MHz Global System for Mobile Communication (GSM) signals in GSM-Talk mode at specific absorption rates (SAR) of 1 W/kg, 2 W/kg or 4 W/kg. Subsequently, through the use of formamidopyrimidine DNA glycosylase (FPG) in a modified comet assay, we determined that the extent of DNA migration was significantly increased at a SAR of 4 W/kg. Flow cytometry analysis demonstrated that levels of the DNA adduct 8-oxoguanine (8-oxoG) were also increased at a SAR of 4 W/kg. These increases were concomitant with similar increases in the generation of reactive oxygen species (ROS); these phenomena were mitigated by co-treatment with the antioxidant α-tocopherol. However, no detectable DNA strand breakage was observed by the alkaline comet assay. Taking together, these findings may imply the novel possibility that RF-EMR with insufficient energy for the direct induction of DNA strand breaks may produce genotoxicity through oxidative DNA base damage in male germ cells. Crown Copyright © 2013. Published by Elsevier Ireland Ltd. All rights reserved.

Ca2+ improves organization of single-stranded DNA bases in human Rad51 filament, explaining stimulatory effect on gene recombination.

KAUST Repository

Fornander, Louise H

2012-02-22

Human RAD51 protein (HsRad51) catalyses the DNA strand exchange reaction for homologous recombination. To clarify the molecular mechanism of the reaction in vitro being more effective in the presence of Ca(2+) than of Mg(2+), we have investigated the effect of these ions on the structure of HsRad51 filament complexes with single- and double-stranded DNA, the reaction intermediates. Flow linear dichroism spectroscopy shows that the two ionic conditions induce significantly different structures in the HsRad51/single-stranded DNA complex, while the HsRad51/double-stranded DNA complex does not demonstrate this ionic dependence. In the HsRad51/single-stranded DNA filament, the primary intermediate of the strand exchange reaction, ATP/Ca(2+) induces an ordered conformation of DNA, with preferentially perpendicular orientation of nucleobases relative to the filament axis, while the presence of ATP/Mg(2+), ADP/Mg(2+) or ADP/Ca(2+) does not. A high strand exchange activity is observed for the filament formed with ATP/Ca(2+), whereas the other filaments exhibit lower activity. Molecular modelling suggests that the structural variation is caused by the divalent cation interfering with the L2 loop close to the DNA-binding site. It is proposed that the larger Ca(2+) stabilizes the loop conformation and thereby the protein-DNA interaction. A tight binding of DNA, with bases perpendicularly oriented, could facilitate strand exchange.

Folate deficiency facilitates recruitment of upstream binding factor to hot spots of DNA double-strand breaks of rRNA genes and promotes its transcription.

Science.gov (United States)

Xie, Qiu; Li, Caihua; Song, Xiaozhen; Wu, Lihua; Jiang, Qian; Qiu, Zhiyong; Cao, Haiyan; Yu, Kaihui; Wan, Chunlei; Li, Jianting; Yang, Feng; Huang, Zebing; Niu, Bo; Jiang, Zhengwen; Zhang, Ting

2017-03-17

The biogenesis of ribosomes in vivo is an essential process for cellular functions. Transcription of ribosomal RNA (rRNA) genes is the rate-limiting step in ribosome biogenesis controlled by environmental conditions. Here, we investigated the role of folate antagonist on changes of DNA double-strand breaks (DSBs) landscape in mouse embryonic stem cells. A significant DSB enhancement was detected in the genome of these cells and a large majority of these DSBs were found in rRNA genes. Furthermore, spontaneous DSBs in cells under folate deficiency conditions were located exclusively within the rRNA gene units, representing a H3K4me1 hallmark. Enrichment H3K4me1 at the hot spots of DSB regions enhanced the recruitment of upstream binding factor (UBF) to rRNA genes, resulting in the increment of rRNA genes transcription. Supplement of folate resulted in a restored UBF binding across DNA breakage sites of rRNA genes, and normal rRNA gene transcription. In samples from neural tube defects (NTDs) with low folate level, up-regulation of rRNA gene transcription was observed, along with aberrant UBF level. Our results present a new view by which alterations in folate levels affects DNA breakage through epigenetic control leading to the regulation of rRNA gene transcription during the early stage of development. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Phenotypic Analysis of ATM Protein Kinase in DNA Double-Strand Break Formation and Repair.

Science.gov (United States)

Mian, Elisabeth; Wiesmüller, Lisa

2017-01-01

Ataxia telangiectasia mutated (ATM) encodes a serine/threonine protein kinase, which is involved in various regulatory processes in mammalian cells. Its best-known role is apical activation of the DNA damage response following generation of DNA double-strand breaks (DSBs). When DSBs appear, sensor and mediator proteins are recruited, activating transducers such as ATM, which in turn relay a widespread signal to a multitude of downstream effectors. ATM mutation causes Ataxia telangiectasia (AT), whereby the disease phenotype shows differing characteristics depending on the underlying ATM mutation. However, all phenotypes share progressive neurodegeneration and marked predisposition to malignancies at the organismal level and sensitivity to ionizing radiation and chromosome aberrations at the cellular level. Expression and localization of the ATM protein can be determined via western blotting and immunofluorescence microscopy; however, detection of subtle alterations such as resulting from amino acid exchanges rather than truncating mutations requires functional testing. Previous studies on the role of ATM in DSB repair, which connects with radiosensitivity and chromosomal stability, gave at first sight contradictory results. To systematically explore the effects of clinically relevant ATM mutations on DSB repair, we engaged a series of lymphoblastoid cell lines (LCLs) derived from AT patients and controls. To examine DSB repair both in a quantitative and qualitative manners, we used an EGFP-based assay comprising different substrates for distinct DSB repair mechanisms. In this way, we demonstrated that particular signaling defects caused by individual ATM mutations led to specific DSB repair phenotypes. To explore the impact of ATM on carcinogenic chromosomal aberrations, we monitored chromosomal breakage at a breakpoint cluster region hotspot within the MLL gene that has been associated with therapy-related leukemia. PCR-based MLL-breakage analysis of HeLa cells

Genetic recombination induced by DNA double-strand break in bacteriophage T4: nature of the left/right bias.

Science.gov (United States)

Shcherbakov, Victor P; Shcherbakova, Tamara; Plugina, Lidiya; Sizova, Svetlana; Kudryashova, Elena; Granovsky, Igor

2008-06-01

The experimental system combining double-strand breaks (DSBs), produced site-specifically by SegC endonuclease, with the famous advantages of the bacteriophage T4 rII mutant recombination analysis was used here to elucidate the origin of the recombination bias on two sides of the DSB, especially pronounced in gene 39 (topoisomerase II) and gene 59 (41-helicase loader) mutants. Three sources were found to contribute to the bias: (1) the SegC endonuclease may remain bound to the end of the broken DNA and thus protect it from exonuclease degradation; (2) in heteroduplex heterozygotes (HHs), arising as the recombinant products in the left-hand crosses, the transcribed strands are of rII mutant phenotype, so they, in contrast to the right-hand HHs, do not produce plaques on the lawn of the lambda-lysogenic host; and (3) the intrinsic polarity of T4 chromosome, reflected in transcription, may be a cause for discrimination of promoter-proximal and promoter-distal DNA sequences. It is shown that the apparent recombination bias does not imply one-sidedness of the DSB repair but just reflects a different depth of the end processing. It is inferred that the cause, underlying the "intrinsic" bias, might be interference between strand exchange and transcription. Topoisomerase and helicase functions are necessary to turn the process in favor of strand exchange. The idea is substantiated that the double-stranded to single-stranded DNA transition edge (not ss-DNA tip) serves as an actual recombinogenic element.

DNA conformation of Chinese hamster V79 cells and sensitivity to ionizing radiation

International Nuclear Information System (INIS)

Olive, P.L.; Hilton, J.; Durand, R.E.

1986-01-01

Chinese hamster V79 cells grown for 20 h in suspension culture form small clusters of cells (spheroids) which are more resistant to killing by ionizing radiation than V79 cells grown as monolayers. This resistance appears to be due to the greater capacity of cells grown in contact to repair radiation damage. Attempts to relate this ''contact effect'' to differences in DNA susceptibility or DNA repair capacity have provided conflicting results. Two techniques, alkaline sucrose gradient sedimentation and alkaline elution, show no difference in the amounts of radiation-induced DNA single-strand breakage or its repair between suspension or monolayer cells. However, using the alkali-unwinding assay, the rate of DNA unwinding is much slower for suspension cells than for monolayer cells. Interestingly, a decrease in salt concentration or in pH of the unwinding solution eliminates these differences in DNA unwinding kinetics. A fourth assay, sedimentation of nucleoids on neutral sucrose gradients, also shows a significant decrease in radiation damage produced in suspension compared to monolayer cultures. It is believed that this assay measures differences in DNA conformation (supercoiling) as well as differences in DNA strand breakage. We conclude from these four assays that the same number of DNA strand breaks/Gy is produced in monolayer and spheroid cells. However, changes in DNA conformation or packaging occur when cells are grown as spheroids, and these changes are responsible for reducing DNA damage by ionizing radiation

Asymmetric strand segregation: epigenetic costs of genetic fidelity?

Directory of Open Access Journals (Sweden)

Diane P Genereux

2009-06-01

Full Text Available Asymmetric strand segregation has been proposed as a mechanism to minimize effective mutation rates in epithelial tissues. Under asymmetric strand segregation, the double-stranded molecule that contains the oldest DNA strand is preferentially targeted to the somatic stem cell after each round of DNA replication. This oldest DNA strand is expected to have fewer errors than younger strands because some of the errors that arise on daughter strands during their synthesis fail to be repaired. Empirical findings suggest the possibility of asymmetric strand segregation in a subset of mammalian cell lineages, indicating that it may indeed function to increase genetic fidelity. However, the implications of asymmetric strand segregation for the fidelity of epigenetic information remain unexplored. Here, I explore the impact of strand-segregation dynamics on epigenetic fidelity using a mathematical-modelling approach that draws on the known molecular mechanisms of DNA methylation and existing rate estimates from empirical methylation data. I find that, for a wide range of starting methylation densities, asymmetric -- but not symmetric -- strand segregation leads to systematic increases in methylation levels if parent strands are subject to de novo methylation events. I found that epigenetic fidelity can be compromised when enhanced genetic fidelity is achieved through asymmetric strand segregation. Strand segregation dynamics could thus explain the increased DNA methylation densities that are observed in structured cellular populations during aging and in disease.

Measurement of intracellular DNA double-strand break induction and rejoining along the track of carbon and neon particle beams in water

International Nuclear Information System (INIS)

Heilmann, Johannes; Taucher-Scholz, Gisela; Haberer, Thomas; Scholz, Michael; Kraft, Gerhard

1996-01-01

Purpose: The study was aimed at the measurement of effect-depth distributions of intracellularly induced DNA damage in water as tissue equivalent after heavy ion irradiation with therapy particle beams. Methods and Materials: An assay involving embedding of Chinese hamster ovary (CHO-K1) cells in large agarose plugs and electrophoretic elution of radiation induced DNA fragments by constant field gel electrophoresis was developed. Double-strand break production was quantified by densitometric analysis of DNA-fluorescence after staining with ethidium-bromide and determination of the fraction of DNA eluted out of the agarose plugs. Intracellular double-strand break induction and the effect of a 3 h rejoining incubation were investigated following irradiation with 250 kV x-rays and 190 MeV/u carbon- and 295 MeV/u neon-ions. Results and Conclusion: While the DNA damage induced by x-irradiation decreased continuously with penetration depth, a steady increase in the yield of double-strand breaks was observed for particle radiation, reaching distinct maxima at the position of the physical Bragg peaks. Beyond this, the extent of radiation damage dropped drastically. From comparison of DNA damage and calculated dose profiles, relative biological efficiencies (RBEs) for both double-strand break induction and unrejoined strand breaks after 3 h were determined. While RBE for the induction of DNA double-strand breaks decreased continuously with penetration depth, RBE maxima greater than unity were found with carbon- and neon-ions for double-strand break rejoining near the maximum range of the particles. The method presented here allows for a fast and accurate determination of depth profiles of relevant radiobiological effects for mixed particle fields in tissue equivalent

Method for detecting DNA strand breaks in mammalian cells using the Deinococcus radiodurans PprA protein

International Nuclear Information System (INIS)

Satoh, Katsuya; Wada, Seiichi; Kikuchi, Masahiro; Funayama, Tomoo; Narumi, Issay; Kobayashi, Yasuhiko

2006-01-01

In a previous study, we identified the novel protein PprA that plays a critical role in the radiation resistance of Deinococcus radiodurans. In this study, we focussed on the ability of PprA protein to recognize and bind to double-stranded DNA carrying strand breaks, and attempted to visualize radiation-induced DNA strand breaks in mammalian cultured cells by employing PprA protein using an immunofluorescence technique. Increased PprA protein binding to CHO-K1 nuclei immediately following irradiation suggests the protein is binding to DNA strand breaks. By altering the cell permeabilization conditions, PprA protein binding to CHO-K1 mitochondria, which is probably resulted from DNA strand break immediately following irradiation, was also detected. The method developed and detailed in this study will be useful in evaluating DNA damage responses in cultured cells, and could also be applicable to genotoxic tests in the environmental and pharmaceutical fields

INDUCTION OF DNA STRAND BREAKS BY TRIHALOMETHANES IN PRIMARY HUMAN LUNG EPITHELIAL CELLS

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Abstract Trihalomethanes (TEMs) are disinfection by-products and suspected human carcinogens present in chlorinated drinking water. Previous studies have shown that many THMs induce sister chromatid exchanges and DNA strand breaks in human peripheral blood lymphocyte...

The Degree of Radiation-Induced DNA Strand Breaks Is Altered by Acute Sleep Deprivation and Psychological Stress and Is Associated with Cognitive Performance in Humans.

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Moreno-Villanueva, Maria; von Scheven, Gudrun; Feiveson, Alan; Bürkle, Alexander; Wu, Honglu; Goel, Namni

2018-03-27

Sleep deprivation is associated with impaired immune responses, cancer, and morbidity and mortality, and can degrade cognitive performance, although individual differences exist in such responses. Sleep deprivation induces DNA strand breaks and DNA base oxidation in animals, and psychological stress is associated with increased DNA damage in humans. It remains unknown whether sleep deprivation or psychological stress in humans affects DNA damage response from environmental stressors, and whether these responses predict cognitive performance during sleep deprivation. Sixteen healthy adults (ages 29-52;mean age±SD, 36.4±7.1 years;7 women) participated in a 5-day experiment involving two 8 hour time-in-bed [TIB] baseline nights, followed by 39 hours total sleep deprivation (TSD), and two 8-10 hour TIB recovery nights. A modified Trier Social Stress Test was conducted on the day after TSD. Psychomotor Vigilance Tests measured behavioral attention. DNA damage was assessed in blood cells collected at 5 time points, and blood cells were irradiated ex-vivo. TSD, alone or in combination with psychological stress, did not induce significant increases in DNA damage. By contrast, radiation-induced DNA damage decreased significantly in response to TSD, but increased back to baseline when combined with psychological stress. Cognitively-vulnerable individuals had more radiation-induced DNA strand breaks before TSD, indicating their greater sensitivity to DNA damage from environmental stressors. Our results provide novel insights into the molecular consequences of sleep deprivation, psychological stress, and performance vulnerability. They are important for situations involving sleep loss, radiation exposure and cognitive deficits, including cancer therapy, environmental toxicology, and space medicine.

Evaluation of the neutral comet assay for detection of alpha-particle induced DNA-double-strand-breaks

International Nuclear Information System (INIS)

Hofbauer, Daniela

2010-01-01

Aim of this study was to differentiate DNA-double-strand-breaks from DNA-single-strand-breaks on a single cell level, using the comet assay after α- and γ-irradiation. Americium-241 was used as a alpha-irradiation-source, Caesium-137 was used for γ-irradiation. Because of technical problems with both the neutral and alkaline comet assay after irradiation of gastric cancer cells and human lymphocytes, no definite differentiation of DNA-damage was possible.

Persistence of Breakage in Specific Chromosome Bands 6 Years after Acute Exposure to Oil.

Directory of Open Access Journals (Sweden)

Alexandra Francés

Full Text Available The identification of breakpoints involved in chromosomal damage could help to detect genes involved in genetic disorders, most notably cancer. Until now, only one published study, carried out by our group, has identified chromosome bands affected by exposure to oil from an oil spill. In that study, which was performed two years after the initial oil exposure in individuals who had participated in clean-up tasks following the wreck of the Prestige, three chromosomal bands (2q21, 3q27, 5q31 were found to be especially prone to breakage. A recent follow-up study, performed on the same individuals, revealed that the genotoxic damage had persisted six years after oil exposure.To determine whether there exist chromosome bands which are especially prone to breakages and to know if there is some correlation with those detected in the previous study. In addition, to investigate if the DNA repair problems detected previously persist in the present study.Follow-up study performed six years after the Prestige oil spill.Fishermen cooperatives in coastal villages.Fishermen highly exposed to oil spill who participated in previous genotoxic study six years after the oil.Chromosome damage in peripheral lymphocytes. For accurate identification of the breakpoints involved in chromosome damage of circulating lymphocytes, a sequential stain/G-banding technique was employed. To determine the most break-prone chromosome bands, two statistical methods, the Fragile Site Multinomial and the chi-square tests (where the bands were corrected by their length were used. To compare the chromosome lesions, structural chromosome alterations and gaps/breaks between two groups of individuals we used the GEE test which takes into account a possible within-individual correlation. Dysfunctions in DNA repair mechanisms, expressed as chromosome damage, were assessed in cultures with aphidicolin by the GEE test.Cytogenetic analyses were performed in 47 exposed individuals. A total of

The oncogenic action of ionizing radiation on rat skin: Progress report, February 1, 1987-January 31, 1988

International Nuclear Information System (INIS)

Burns, F.J.

1987-01-01

The work outlined in this report includes: epidermal DNA strand breaks and radiation penetration; activation of oncogenes in radiation induced rat skin tumors; and rat skin carcinogenesis by neon ions. The effect of radiation penetration on DNA single strand breaks has been studied extensively in rat and mouse epidermis. The results show clearly that the number of strand breaks per unit dose in the epidermal DNA is reduced by 50% to 65% when the radiation penetration is reduced from 1.0 mm to 0.2 mm. This penetration effect on DNA strand breaks was not seen in mouse epidermal cell lines growing in plastic dishes. The results imply that DNA strand breakage in superficial cells is partially dependent on the radiation dose to underlying tissue. The phenomenon is not mediated by systemic interactions as it was observed in irradiated explanted skin. The oncogene activation pattern in the radiation-induced skin tumors was found to be tumor dependent. Either K-ras activation or c-myc amplification or both was observed in each tumor analyzed so far. Even benign fibromas exhibited c-myc amplification. The carcinogenicity of high penetration electrons (2.0 MeV) was determined in preparation for similar studies with a neon ion beam at the Berkeley Bevelac. The principal finding so far is a large excess of connective tissue tumors, fibromas (benign) and sarcomas (malignant). 59 refs., 1 tab

Fluorometric determination of nucleic acids based on the use of polydopamine nanotubes and target-induced strand displacement amplification.

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Ge, Jia; Bai, Dong-Mei; -Geng, Xin; Hu, Ya-Lei; Cai, Qi-Yong; Xing, Ke; Zhang, Lin; Li, Zhao-Hui

2018-01-10

The authors describe a fluorometric method for the quantitation of nucleic acids by combining (a) cycled strand displacement amplification, (b) the unique features of the DNA probe SYBR Green, and (c) polydopamine nanotubes. SYBR Green undergoes strong fluorescence enhancement upon intercalation into double-stranded DNA (dsDNA). The polydopamine nanotubes selectively adsorb single-stranded DNA (ssDNA) and molecular beacons. In the absence of target DNA, the molecular beacon, primer and SYBR Green are adsorbed on the surface of polydopamine nanotubes. This results in quenching of the fluorescence of SYBR Green, typically measured at excitation/emission wavelengths of 488/518 nm. Upon addition of analyte (target DNA) and polymerase, the stem of the molecular beacon is opened so that it can bind to the primer. This triggers target strand displacement polymerization, during which dsDNA is synthesized. The hybridized target is then displaced due to the strand displacement activity of the polymerase. The displaced target hybridizes with another molecular beacon. This triggers the next round of polymerization. Consequently, a large amount of dsDNA is formed which is detected by addition of SYBR Green. Thus, sensitive and selective fluorometric detection is realized. The fluorescent sensing strategy shows very good analytical performances towards DNA detection, such as a wide linear range from 0.05 to 25 nM with a low limit of detection of 20 pM. Graphical abstract Schematic of a fluorometric strategy for highly sensitive and selective determination of nucleic acids by combining strand displacement amplification and the unique features of SYBR Green I (SG) and polydopamine nanotubes.

Modelling toehold-mediated RNA strand displacement.

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Šulc, Petr; Ouldridge, Thomas E; Romano, Flavio; Doye, Jonathan P K; Louis, Ard A

2015-03-10

We study the thermodynamics and kinetics of an RNA toehold-mediated strand displacement reaction with a recently developed coarse-grained model of RNA. Strand displacement, during which a single strand displaces a different strand previously bound to a complementary substrate strand, is an essential mechanism in active nucleic acid nanotechnology and has also been hypothesized to occur in vivo. We study the rate of displacement reactions as a function of the length of the toehold and temperature and make two experimentally testable predictions: that the displacement is faster if the toehold is placed at the 5' end of the substrate; and that the displacement slows down with increasing temperature for longer toeholds. Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Second-strand cDNA synthesis: classical method

International Nuclear Information System (INIS)

Gubler, U.

1987-01-01

The classical scheme for the synthesis of double-stranded cDNA as it was reported in 1976 is described. Reverse transcription of mRNA with oligo(dT) as the primer generates first strands with a small loop at the 3' end of the cDNA (the end that corresponds to the 5' end of the mRNA). Subsequent removal of the mRNA by alkaline hydrolysis leaves single-stranded cDNA molecules again with a small 3' loop. This loop can be used by either reverse transcriptase or Klenow fragment of DNA polymerase I as a primer for second-strand synthesis. The resulting products are double-stranded cDNA molecules that are covalently closed at the end corresponding to the 5' end of the original mRNA. Subsequent cleavage of the short piece of single-stranded cDNA within the loop with the single-strand-specific S 1 nuclease generate open double-stranded molecules that can be used for molecular cloning in plasmids or in phage. Useful variations of this scheme have been described

Gametocidal chromosomes enhancing chromosome aberration in common wheat induced by 5-azacytidine.

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Su, W-Y; Cong, W-W; Shu, Y-J; Wang, D; Xu, G-H; Guo, C-H

2013-07-08

The gametocidal (Gc) chromosome from Aegilops spp induces chromosome mutation, which is introduced into common wheat as a tool of chromosome manipulation for genetic improvement. The Gc chromosome functions similar to a restriction-modification system in bacteria, in which DNA methylation is an important regulator. We treated root tips of wheat carrying Gc chromosomes with the hypomethylation agent 5-azacytidine; chromosome breakage and micronuclei were observed in these root tips. The frequency of aberrations differed in wheat containing different Gc chromosomes, suggesting different functions inducing chromosome breakage. Gc chromosome 3C caused the greatest degree of chromosome aberration, while Gc chromosome 3C(SAT) and 2C caused only slight chromosome aberration. Gc chromosome 3C induced different degrees of chromosome aberration in wheat varieties Triticum aestivum var. Chinese Spring and Norin 26, demonstrating an inhibition function in common wheat.

Purification, crystallization and preliminary X-ray diffraction experiments on the breakage-reunion domain of the DNA gyrase from Mycobacterium tuberculosis

International Nuclear Information System (INIS)

Piton, Jérémie; Matrat, Stéphanie; Petrella, Stéphanie; Jarlier, Vincent; Aubry, Alexandra; Mayer, Claudine

2009-01-01

The breakage-reunion domain of M. tuberculosis DNA gyrase was crystallized using the hanging-drop vapour-diffusion method. One of the four crystal forms obtained belonged to space group C2 and diffraction data were collected to a resolution of 2.7 Å. Mycobacterium tuberculosis DNA gyrase, a nanomachine that is involved in the regulation of DNA topology, is the only type II topoisomerase present in this organism and hence is the sole target for fluoroquinolone action. The breakage-reunion domain of the A subunit plays an essential role in DNA binding during the catalytic cycle. Two constructs of 53 and 57 kDa (termed GA53BK and GA57BK) corresponding to this domain have been overproduced, purified and crystallized. Diffraction data were collected from four crystal forms. The resolution limits ranged from 4.6 to 2.7 Å depending on the crystal form. The best diffracting crystals belonged to space group C2, with a biological dimer in the asymmetric unit. This is the first report of the crystallization and preliminary X-ray diffraction analysis of the breakage-reunion domain of DNA gyrase from a species containing one unique type II topoisomerase

The multiple personalities of Watson and Crick strands.

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Cartwright, Reed A; Graur, Dan

2011-02-08

In genetics it is customary to refer to double-stranded DNA as containing a "Watson strand" and a "Crick strand." However, there seems to be no consensus in the literature on the exact meaning of these two terms, and the many usages contradict one another as well as the original definition. Here, we review the history of the terminology and suggest retaining a single sense that is currently the most useful and consistent. The Saccharomyces Genome Database defines the Watson strand as the strand which has its 5'-end at the short-arm telomere and the Crick strand as its complement. The Watson strand is always used as the reference strand in their database. Using this as the basis of our standard, we recommend that Watson and Crick strand terminology only be used in the context of genomics. When possible, the centromere or other genomic feature should be used as a reference point, dividing the chromosome into two arms of unequal lengths. Under our proposal, the Watson strand is standardized as the strand whose 5'-end is on the short arm of the chromosome, and the Crick strand as the one whose 5'-end is on the long arm. Furthermore, the Watson strand should be retained as the reference (plus) strand in a genomic database. This usage not only makes the determination of Watson and Crick unambiguous, but also allows unambiguous selection of reference stands for genomics. This article was reviewed by John M. Logsdon, Igor B. Rogozin (nominated by Andrey Rzhetsky), and William Martin.

The multiple personalities of Watson and Crick strands

Directory of Open Access Journals (Sweden)

Graur Dan

2011-02-01

Full Text Available Abstract Background In genetics it is customary to refer to double-stranded DNA as containing a "Watson strand" and a "Crick strand." However, there seems to be no consensus in the literature on the exact meaning of these two terms, and the many usages contradict one another as well as the original definition. Here, we review the history of the terminology and suggest retaining a single sense that is currently the most useful and consistent. Proposal The Saccharomyces Genome Database defines the Watson strand as the strand which has its 5'-end at the short-arm telomere and the Crick strand as its complement. The Watson strand is always used as the reference strand in their database. Using this as the basis of our standard, we recommend that Watson and Crick strand terminology only be used in the context of genomics. When possible, the centromere or other genomic feature should be used as a reference point, dividing the chromosome into two arms of unequal lengths. Under our proposal, the Watson strand is standardized as the strand whose 5'-end is on the short arm of the chromosome, and the Crick strand as the one whose 5'-end is on the long arm. Furthermore, the Watson strand should be retained as the reference (plus strand in a genomic database. This usage not only makes the determination of Watson and Crick unambiguous, but also allows unambiguous selection of reference stands for genomics. Reviewers This article was reviewed by John M. Logsdon, Igor B. Rogozin (nominated by Andrey Rzhetsky, and William Martin.

Prooxidant action of furanone compounds: implication of reactive oxygen species in the metal-dependent strand breaks and the formation of 8-hydroxy-2'-deoxyguanosine in DNA.

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Murakami, K; Haneda, M; Makino, T; Yoshino, M

2007-07-01

Prooxidant properties of furanone compounds including 2,5-furanone (furaneol, 4-hydroxy-2,5-dimethyl-furan-3-one), 4,5-furanone (4,5-dimethyl-3-hydroxy-2(5H)-furanone) (sotolone) and cyclotene (2-hydroxy-3-methyl-2-cyclopenten-1-one) were analyzed in relation to the metal-reducing activity. Only 2.5-furanone known as a "strawberry or pineapple furanone" inactivated aconitase the most sensitive enzyme to active oxygen in the presence of ferrous sulfate, suggesting the furaneol/iron-mediated generation of reactive oxygen species. 2,5-Furanone caused strand scission of pBR322 DNA in the presence of copper. Treatment of calf thymus DNA with 2,5-furanone plus copper produced 8-hydroxy-2'-deoxyguanosine in DNA. 2,5-Furanone showed a potent copper-reducing activity, and thus, DNA strand breaks and the formation of 8-hydroxy-2'-deoxyguanosine by 2,5-furanone can be initiated by the production of superoxide radical through the reduction of cupric ion to cuprous ion, resulting in the conversion to hydrogen peroxide and hydroxyl radical. However, an isomer and analog of 2,5-furanone, 4,5-furanone and cyclotene, respectively, did not show an inactivation of aconitase, DNA injuries including strand breakage and the formation of 8-hydroxy-2'-deoxyguanosine, and copper-reducing activity. Cytotoxic effect of 2,5-furanone with hydroxyketone structure can be explained by its prooxidant properties: furaneol/transition metal complex generates reactive oxygen species causing the inactivation of aconitase and the formation of DNA base damage by hydroxyl radical.

Radiation dose response of strand breaks in SINPV-DNA

International Nuclear Information System (INIS)

Zhang Chunxiang; Luo Daling; Li Mianfeng; Liu Xiaowei; Zeng Rong; Wang Xunzhang

1995-01-01

The Spodoplera litura Nuclear Polyhedrosis Viruses (SINPV) is a kind of insectile virus with a simple structure, in which a double helix DNA is encapsulated in a protein coat and there is no function of enzymatic repair. The SINPV samples in dry powdered form held in sealed plastic tube were irradiated by 1-100 kGy gamma rays. The single strand breaks (SSB) and double strand breaks (DSB) induced in SINPV after irradiation were measured by neutral and alkaline agarose gel electrophoresis. A dose-response function combining the responses of one-hit and two-hit events was used to describe the SSB and DSB dose-response curves. It is shown that the SSB are one-hit events and the DSB are the combination of both one-hit, and two-hit events, and two-hit events are predominant in the DSB process

DNA strand breaking and rejoining in response to ultraviolet light in normal human and xeroderma pigmentosum cells

International Nuclear Information System (INIS)

Dingman, C.W.; Kakunaga, T.

1976-01-01

A description is given of a reproducible technique for measuring DNA strand breaking and rejoining in cells after treatment with U.V.-light. Results obtained with normal human cells, xeroderma pigmentosum cells (XP, complementation group A) and XP variant cells suggested that all three of these cell-types can carry out single-strand incision with equal rapidity. However, the breaks so induced appeared to be only slowly rejoined in the XP variant cells and rejoined not at all in XP complementation group A cells. Furthermore, parental strand rejoining was inhibited by caffeine in XP variant cells but not in normal cells. (author)

Tooth breakage in patients injected with 224Ra

International Nuclear Information System (INIS)

Sonnabend, E.; Spiess, H.; Mays, C.W.

1986-01-01

Tooth breakage has been common among the 224 Ra patients, especially those injected as teenagers. The fracture of 1 or more teeth was reported by 26% (8/31) of boys and 23% (7/30) of girls injected with 224 Ra at 16-20 years of age. Combining results from all age groups, the incidence of tooth fractures increased significantly with dose (P=0.01). Unlike the normal loss of permanent teeth by periodontal disease, in which the entire tooth is lost, the tooth loss following 224 Ra injection was primarily from tooth resorption near the gum line causing the tooth crowns to break off easily. Eventually the tooth roots may either become incorporated into the jawbone or are resorbed and replaced with bone. These tooth fractures resembled those observed in the U.S. radium dial painters and in dogs injected with bone-seeking α-emitters. (orig.)

Sea Turtle Stranding Network Reports

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National Oceanic and Atmospheric Administration, Department of Commerce — The Sea Turtle Stranding and Salvage Network (STSSN) was formally established in 1980 to collect information on and document the stranding of marine turtles along...

UV light-induced DNA synthesis arrest in HeLa cells is associated with changes in phosphorylation of human single-stranded DNA-binding protein

International Nuclear Information System (INIS)

Carty, M.P.; Zernik-Kobak, M.; McGrath, S.; Dixon, K.

1994-01-01

We show that DNA replication activity in extracts of human HeLa cells decreases following UV irradiation. Alterations in replication activity in vitro parallel the UV-induced block in cell cycle progression of these cells in culture. UV irradiation also induces specific changes in the pattern of phosphorylation of the 34 kDa subunit of a DNA replication protein, human single-stranded DNA-binding protein (hSSB). The appearance of a hyperphosphorylated form of hSSB correlates with reduced in vitro DNA replication activity in extracts of UV-irradiated cells. Replication activity can be restored to these extracts in vitro by addition of purified hSSB. These results suggest that UV-induced DNA synthesis arrest may be mediated in part through phosphorylation-related alterations in the activity of hSSB, an essential component of the DNA replication apparatus. (Author)

A Tool for Determining the Number of Contributors: Interpreting Complex, Compromised Low-Template Dna Samples

Science.gov (United States)

2017-09-28

Ph.D. Catherine Grgicak Phone: (617) 638-1968 STEM Degrees: STEM Participants: RPPR Final Report as of 17-Oct-2017 Agreement Number: W911NF-14-C...to degrade into increasingly smaller fragments over time. The mechanism inducing DNA damage can include strand breakage, formation of pyrimidine...in this example ⌊(7.8⁡10−4)(48⁡103) 63⁄ ⌋ = ⌊5.94⌋ = 5. Note that 48 µL stems from the knowledge that typically 2 of 50 µL of the extract is

Adaptive response of DNA strand breaks in lymphocytes to low dose and γ-rays

International Nuclear Information System (INIS)

Du Zeji; Su Liaoyuan; Kong Xiangrong; Tian Hailin

1996-01-01

Fluorometric analysis of DNA unwinding was used to study the adaptive response of DNA strand breaks induced by low dose γ-rays and the effect of pADPRT inhibitor-3-AB on the adaptive response. The results indicated that 0.5-4 cGy γ-rays could induce adaptive response of DNA strand breaks in lymphocytes, especially at the doses of 2.0 and 4.0 cGy. This response was not obvious after 8.0 cGy γ-rays irradiation. A challenge dose of 5-20 Gy could make the response expressed, 15 Gy was the best one and 30 Gy was too high to give an adaptive response . 0.5 mM 3-AB could inhibit the response vigorously. As the concentration increased, the adaptive response could be inhibited completely

Extensive and equivalent repair in both radiation-resistant and radiation-sensitive E. coli determined by a DNA-unwinding technique

International Nuclear Information System (INIS)

Ahnstroem, G.; George, A.M.; Cramp, W.A.

1978-01-01

The extent of strand breakage and repair in irradiated E. coli B/r and Bsub(s-l) was studied using a DNA-unwinding technique in denaturing conditions of weak alkali. Although these two strains showed widely different response to the lethal effects of ionizing radiation, they both had an equal capacity to repair radiation-induced breaks in DNA. Oxygen enhancement ratios for the killing of B/r and Bsub(s-l) were respectively 4 and 2; but after repair in non-nutrient or nutrient post-irradiation conditions, the oxygen enhancement values for the residual strand breaks were always the same for the two strains. The equal abilities of E.coli B/r and E.coli Bsub(s-l) to remove the strand breaks measured by this weak-alkali technqiue has led to the suggestion that some other type of damage to either DNA or another macromolecule may play a major role in determining whether or not the cells survive to proliferate. (author)

Extensive and equivalent repair in both radiation-resistant and radiation-sensitive E. coli determined by a DNA-unwinding technique

Energy Technology Data Exchange (ETDEWEB)

Ahnstroem, G [Stockholm Univ. (Sweden); George, A M; Cramp, W A

1978-10-01

The extent of strand breakage and repair in irradiated E. coli B/r and Bsub(s-l) was studied using a DNA-unwinding technique in denaturing conditions of weak alkali. Although these two strains showed widely different response to the lethal effects of ionizing radiation, they both had an equal capacity to repair radiation-induced breaks in DNA. Oxygen enhancement ratios for the killing of B/r and Bsub(s-l) were respectively 4 and 2; but after repair in non-nutrient or nutrient post-irradiation conditions, the oxygen enhancement values for the residual strand breaks were always the same for the two strains. The equal abilities of E.coli B/r and E.coli Bsub(s-l) to remove the strand breaks measured by this weak-alkali technqiue has led to the suggestion that some other type of damage to either DNA or another macromolecule may play a major role in determining whether or not the cells survive to proliferate.

Radiation-induced chromosome breakages in bread wheat (Triticum aestivum L.)

International Nuclear Information System (INIS)

Larik, A.S.

1975-01-01

Meiosis and pollen fertility were studied in the M 2 generation in four varieties of hexaploid wheat. Meiosis was characterized by the formation of interchange configurations, such as rings and chains of four chromosomes in several cells. Chromosomal aberrations showed linear relationship with gamma irradiation; 45 kR dose induced the highest chromosomal abnormalities. Most multivalents were interchange rings of four chromosomes. Translocations involving two pairs of homologous or nonhomologous chromosomes seemed to be higher in frequency than those involving more than two pairs of chromosomes. Anaphase abnormalities, such as laggards, bridges and fragments and unequal segregation of chromosomes, were frequently observed. Pollen fertility was considerably reduced in the M 2 plants arising form the treatments of higher doses of gamma rays because of the induced chromosome interchanges. (author)

A critical role for topoisomerase IIb and DNA double strand breaks in transcription.

Science.gov (United States)

Calderwood, Stuart K

2016-05-26

Recent studies have indicated a novel role for topoisomerase IIb in transcription. Transcription of heat shock genes, serum-induced immediate early genes and nuclear receptor-activated genes, each required DNA double strands generated by topoisomerase IIb. Such strand breaks seemed both necessary and sufficient for transcriptional activation. In addition, such transcription was associated with initiation of the DNA damage response pathways, including the activation of the enzymes: ataxia-telangiectasia mutated (ATM), DNA-dependent protein kinase and poly (ADP ribose) polymerase 1. DNA damage response signaling was involved both in transcription and in repair of DNA breaks generated by topoisomerase IIb.

Double Strand Break Repair, one mechanism can hide another: Alternative non-homologous end joining

International Nuclear Information System (INIS)

Rass, E.; Grabarz, A.; Bertrand, P.; Lopez, B.S.

2012-01-01

DNA double strand breaks are major cytotoxic lesions encountered by the cells. They can be induced by ionizing radiation or endogenous stress and can lead to genetic instability. Two mechanisms compete for the repair of DNA double strand breaks: homologous recombination and non-homologous end joining (NHEJ). Homologous recombination requires DNA sequences homology and is initiated by single strand resection. Recently, advances have been made concerning the major steps and proteins involved in resection. NHEJ, in contrast, does not require sequence homology. The existence of a DNA double strand break repair mechanism, independent of KU and ligase IV, the key proteins of the canonical non homologous end joining pathway, has been revealed lately and named alternative non homologous end joining. The hallmarks of this highly mutagenic pathway are deletions at repair junctions and frequent use of distal micro-homologies. This mechanism is also initiated by a single strand resection of the break. The aim of this review is firstly to present recent data on single strand resection, and secondly the alternative NHEJ pathway, including a discussion on the fidelity of NHEJ. Based on current knowledge, canonical NHEJ does not appear as an intrinsically mutagenic mechanism, but in contrast, as a conservative one. The structure of broken DNA ends actually dictates the quality repair of the alternative NHEJ and seems the actual responsible for the mutagenesis attributed beforehand to the canonical NHEJ. The existence of this novel DNA double strand breaks repair mechanism needs to be taken into account in the development of radiosensitizing strategies in order to optimise the efficiency of radiotherapy. (authors)

Breakage and drying behaviour of granules in a continuous fluid bed dryer: Influence of process parameters and wet granule transfer.

Science.gov (United States)

De Leersnyder, F; Vanhoorne, V; Bekaert, H; Vercruysse, J; Ghijs, M; Bostijn, N; Verstraeten, M; Cappuyns, P; Van Assche, I; Vander Heyden, Y; Ziemons, E; Remon, J P; Nopens, I; Vervaet, C; De Beer, T

2018-03-30

Although twin screw granulation has already been widely studied in recent years, only few studies addressed the subsequent continuous drying which is required after wet granulation and still suffers from a lack of detailed understanding. The latter is important for optimisation and control and, hence, a cost-effective practical implementation. Therefore, the aim of the current study is to increase understanding of the drying kinetics and the breakage and attrition phenomena during fluid bed drying after continuous twin screw granulation. Experiments were performed on a continuous manufacturing line consisting of a twin-screw granulator, a six-segmented fluid bed dryer, a mill, a lubricant blender and a tablet press. Granulation parameters were fixed in order to only examine the effect of drying parameters (filling time, drying time, air flow, drying air temperature) on the size distribution and moisture content of granules (both of the entire granulate and of size fractions). The wet granules were transferred either gravimetrically or pneumatically from the granulator exit to the fluid bed dryer. After a certain drying time, the moisture content reached an equilibrium. This drying time was found to depend on the applied airflow, drying air temperature and filling time. The moisture content of the granules decreased with an increasing drying time, airflow and drying temperature. Although smaller granules dried faster, the multimodal particle size distribution of the granules did not compromise uniform drying of the granules when the target moisture content was achieved. Extensive breakage of granules was observed during drying. Especially wet granules were prone to breakage and attrition during pneumatic transport, either in the wet transfer line or in the dry transfer line. Breakage and attrition of granules during transport and drying should be anticipated early on during process and formulation development by performing integrated experiments on the granulator

Single-strand breaks in oligodeoxyribonucleotides induced by fission neutrons and gamma radiation and measured by gel electrophoresis. Protective effects of aminothiols

International Nuclear Information System (INIS)

Swenberg, C.E.; Vaishnav, Y.N.; Li, Bin; Tsao, Hong; Mao, Bing; Geacintov, N.E.

1997-01-01

The technique of high-resolution gel electrophoresis using oligodeoxyribonucleotides of known composition as model systems, offers a simple quantitative estimate of DNA damage in aqueous solution induced by ionizing radiation. The fraction of damaged DNA can be quantitatively defined in terms of the increased electrophoretic mobilities of the damaged oligonucleotides, relative to the mobility of the unirradiated and intact oligonucleotides. The usual direct strand breaks can be observed at γ-ray dosages of 200 Gy. However, at a γ-ray dosage of 400 Gy, only a broad background, attributed to heterogeneously and multiply damaged oligonucleotide fragments with overlapping and varying electrophoretic mobilities, can be distinguished. On the other hand, individual bands due to resolvable DNA fragments are evident even at dosages as high as 400 Gy for fission neutrons. When double-stranded oligonucleotides are exposed to γ-ray dosages of 200 Gy, the fraction of damaged DNA approaches 30-40%. This damage can be almost completely suppressed (>99%) if the irradiations are conducted in aqueous solutions in the presence of 0.5-1.0 mM concentrations of the thiols cysteamine or 3-(3-methylaminopropylamino)propanethiol (WR-151326). The rate constant of reaction of OH·radicals with small double stranded oligonucleotides 16 base pairs long, K DNA , is found to be closer to the diffusion-controlled value (>3 x 10 9 M -1 s -1 ) than the magnitudes of K DNA for the higher molecular weight, native DNA reported in the literature. These observations suggest that oligonucleotides represent more simple model systems than native DNA in solutions for studying the mechanisms of radioprotection exerted by thiols of different structures. (author)

Normal rejoining of DNA strand breaks in ataxia telangiectasia fibroblast lines after low x-ray exposure

International Nuclear Information System (INIS)

Hariharan, P.V.; Eleczko, S.; Smith, B.P.; Paterson, M.C.

1981-01-01

The alkaline elution method was used to measure the enzymatic repair of x-ray-induced DNA strand breaks in skin fibroblasts derived from human subjects afflicted with ataxia telangiectasia (AT). Monolayer cultures of normal control and AT cell lines were exposed acutely to moderately lethal (250-rad) and highly lethal (1250-rad) doses of 250-kV x rays under aerobic conditions. Upon receiving 250 rad, the control fibroblasts from a clinically normal donor rejoined all detectable single-strand breaks (plus alkali-labile bonds) within 30 to 60 min of incubation. When challenged with 1250 rad the kinetics of strand rejoining by the normal control cells were biphasic. For both exposures, no significant difference in either the rate or the extent of strand rejoining was detected between the normal cell line (GM38) and three mutant cell lines (AT2BE, AT3BI, AT4BI) belonging to the three known genetic complementation groups in AT. It would thus appear that the enhanced radiosensitivity of cultured AT cells does not stem from faulty rejoining of radiogenic DNA strand breaks

REV7 counteracts DNA double-strand break resection and affects PARP inhibition

NARCIS (Netherlands)

Xu, Guotai; Chapman, J. Ross; Brandsma, Inger; Yuan, Jingsong; Mistrik, Martin; Bouwman, Peter; Bartkova, Jirina; Gogola, Ewa; Warmerdam, Daniël; Barazas, Marco; Jaspers, Janneke E.; Watanabe, Kenji; Pieterse, Mark; Kersbergen, Ariena; Sol, Wendy; Celie, Patrick H. N.; Schouten, Philip C.; van den Broek, Bram; Salman, Ahmed; Nieuwland, Marja; de Rink, Iris; de Ronde, Jorma; Jalink, Kees; Boulton, Simon J.; Chen, Junjie; van Gent, Dik C.; Bartek, Jiri; Jonkers, Jos; Borst, Piet; Rottenberg, Sven

2015-01-01

Error-free repair of DNA double-strand breaks (DSBs) is achieved by homologous recombination (HR), and BRCA1 is an important factor for this repair pathway(1). In the absence of BRCA1-mediated HR, the administration of PARP inhibitors induces synthetic lethality of tumour cells of patients with

DNA single-strand breaks during repair of uv damage in human fibroblasts and abnormalities of repair in xeroderma pigmentosum

International Nuclear Information System (INIS)

Fornace, A.J. Jr.; Kohn, K.W.; Kann, H.E. Jr.

1976-01-01

The method of DNA alkaline elution was applied to a study of the formation and resealing of DNA single-strand breaks after irradiation of human fibroblasts with ultraviolet light (UV). The general features of the results were consistent with current concepts of DNA excision repair, in that breaks appeared rapidly after uv, and resealed slowly in normal fibroblasts, whereas breaks did not appear in those cells of patients with xeroderma pigmentosum (XP) that are known to have defects in DNA repair synthesis. The appearance of breaks required a short post-uv incubation, consistent with the expected action of an endonuclease. Cells of the variant form of XP characterized by normal DNA repair synthesis exhibited normal production of breaks after uv, but were slower than normal cells in resealing these breaks. This difference was enhanced by caffeine. A model is proposed to relate this finding with a previously described defect in post-replication repair in these XP variant cells. DNA crosslinking appears to cause an underestimate in the measurement of DNA breakage after uv

Coincident resection at both ends of random, γ-induced double-strand breaks requires MRX (MRN, Sae2 (Ctp1, and Mre11-nuclease.

Directory of Open Access Journals (Sweden)

James W Westmoreland

2013-03-01

Full Text Available Resection is an early step in homology-directed recombinational repair (HDRR of DNA double-strand breaks (DSBs. Resection enables strand invasion as well as reannealing following DNA synthesis across a DSB to assure efficient HDRR. While resection of only one end could result in genome instability, it has not been feasible to address events at both ends of a DSB, or to distinguish 1- versus 2-end resections at random, radiation-induced "dirty" DSBs or even enzyme-induced "clean" DSBs. Previously, we quantitatively addressed resection and the role of Mre11/Rad50/Xrs2 complex (MRX at random DSBs in circular chromosomes within budding yeast based on reduced pulsed-field gel electrophoretic mobility ("PFGE-shift". Here, we extend PFGE analysis to a second dimension and demonstrate unique patterns associated with 0-, 1-, and 2-end resections at DSBs, providing opportunities to examine coincidence of resection. In G2-arrested WT, Δrad51 and Δrad52 cells deficient in late stages of HDRR, resection occurs at both ends of γ-DSBs. However, for radiation-induced and I-SceI-induced DSBs, 1-end resections predominate in MRX (MRN null mutants with or without Ku70. Surprisingly, Sae2 (Ctp1/CtIP and Mre11 nuclease-deficient mutants have similar responses, although there is less impact on repair. Thus, we provide direct molecular characterization of coincident resection at random, radiation-induced DSBs and show that rapid and coincident initiation of resection at γ-DSBs requires MRX, Sae2 protein, and Mre11 nuclease. Structural features of MRX complex are consistent with coincident resection being due to an ability to interact with both DSB ends to directly coordinate resection. Interestingly, coincident resection at clean I-SceI-induced breaks is much less dependent on Mre11 nuclease or Sae2, contrary to a strong dependence on MRX complex, suggesting different roles for these functions at "dirty" and clean DSB ends. These approaches apply to resection at

RDE-1 slicer activity is required only for passenger-strand cleavage during RNAi in Caenorhabditis elegans.

Science.gov (United States)

Steiner, Florian A; Okihara, Kristy L; Hoogstrate, Suzanne W; Sijen, Titia; Ketting, René F

2009-02-01

RNA interference (RNAi) is a process in which double-stranded RNA is cleaved into small interfering RNAs (siRNAs) that induce the destruction of homologous single-stranded mRNAs. Argonaute proteins are essential components of this silencing process; they bind siRNAs directly and can cleave RNA targets using a conserved RNase H motif. In Caenorhabditis elegans, the Argonaute protein RDE-1 has a central role in RNAi. In animals lacking RDE-1, the introduction of double-stranded RNA does not trigger any detectable level of RNAi. Here we show that RNase H activity of RDE-1 is required only for efficient removal of the passenger strand of the siRNA duplex and not for triggering the silencing response at the target-mRNA level. These results uncouple the role of the RDE-1 RNase H activity in small RNA maturation from its role in target-mRNA silencing in vivo.

Identification of cis-acting elements on positive-strand subgenomic mRNA required for the synthesis of negative-strand counterpart in bovine coronavirus.

Science.gov (United States)

Yeh, Po-Yuan; Wu, Hung-Yi

2014-07-30

It has been demonstrated that, in addition to genomic RNA, sgmRNA is able to serve as a template for the synthesis of the negative-strand [(-)-strand] complement. However, the cis-acting elements on the positive-strand [(+)-strand] sgmRNA required for (-)-strand sgmRNA synthesis have not yet been systematically identified. In this study, we employed real-time quantitative reverse transcription polymerase chain reaction to analyze the cis-acting elements on bovine coronavirus (BCoV) sgmRNA 7 required for the synthesis of its (-)-strand counterpart by deletion mutagenesis. The major findings are as follows. (1) Deletion of the 5'-terminal leader sequence on sgmRNA 7 decreased the synthesis of the (-)-strand sgmRNA complement. (2) Deletions of the 3' untranslated region (UTR) bulged stem-loop showed no effect on (-)-strand sgmRNA synthesis; however, deletion of the 3' UTR pseudoknot decreased the yield of (-)-strand sgmRNA. (3) Nucleotides positioned from -15 to -34 of the sgmRNA 7 3'-terminal region are required for efficient (-)-strand sgmRNA synthesis. (4) Nucleotide species at the 3'-most position (-1) of sgmRNA 7 is correlated to the efficiency of (-)-strand sgmRNA synthesis. These results together suggest, in principle, that the 5'- and 3'-terminal sequences on sgmRNA 7 harbor cis-acting elements are critical for efficient (-)-strand sgmRNA synthesis in BCoV.

Single-stranded γPNAs for in vivo site-specific genome editing via Watson-Crick recognition.

Science.gov (United States)

Bahal, Raman; Quijano, Elias; McNeer, Nicole A; Liu, Yanfeng; Bhunia, Dinesh C; Lopez-Giraldez, Francesco; Fields, Rachel J; Saltzman, William M; Ly, Danith H; Glazer, Peter M

2014-01-01

Triplex-forming peptide nucleic acids (PNAs) facilitate gene editing by stimulating recombination of donor DNAs within genomic DNA via site-specific formation of altered helical structures that further stimulate DNA repair. However, PNAs designed for triplex formation are sequence restricted to homopurine sites. Herein we describe a novel strategy where next generation single-stranded gamma PNAs (γPNAs) containing miniPEG substitutions at the gamma position can target genomic DNA in mouse bone marrow at mixed-sequence sites to induce targeted gene editing. In addition to enhanced binding, γPNAs confer increased solubility and improved formulation into poly(lactic-co-glycolic acid) (PLGA) nanoparticles for efficient intracellular delivery. Single-stranded γPNAs induce targeted gene editing at frequencies of 0.8% in mouse bone marrow cells treated ex vivo and 0.1% in vivo via IV injection, without detectable toxicity. These results suggest that γPNAs may provide a new tool for induced gene editing based on Watson-Crick recognition without sequence restriction.

Fair Exchange in Strand Spaces

Directory of Open Access Journals (Sweden)

Joshua D. Guttman

2009-10-01

Full Text Available Many cryptographic protocols are intended to coordinate state changes among principals. Exchange protocols coordinate delivery of new values to the participants, e.g. additions to the set of values they possess. An exchange protocol is fair if it ensures that delivery of new values is balanced: If one participant obtains a new possession via the protocol, then all other participants will, too. Fair exchange requires progress assumptions, unlike some other protocol properties. The strand space model is a framework for design and verification of cryptographic protocols. A strand is a local behavior of a single principal in a single session of a protocol. A bundle is a partially ordered global execution built from protocol strands and adversary activities. The strand space model needs two additions for fair exchange protocols. First, we regard the state as a multiset of facts, and we allow strands to cause changes in this state via multiset rewriting. Second, progress assumptions stipulate that some channels are resilient-and guaranteed to deliver messages-and some principals are assumed not to stop at certain critical steps. This method leads to proofs of correctness that cleanly separate protocol properties, such as authentication and confidentiality, from invariants governing state evolution. G. Wang's recent fair exchange protocol illustrates the approach.

Protected DNA strand displacement for enhanced single nucleotide discrimination in double-stranded DNA.

Science.gov (United States)

Khodakov, Dmitriy A; Khodakova, Anastasia S; Huang, David M; Linacre, Adrian; Ellis, Amanda V

2015-03-04

Single nucleotide polymorphisms (SNPs) are a prime source of genetic diversity. Discriminating between different SNPs provides an enormous leap towards the better understanding of the uniqueness of biological systems. Here we report on a new approach for SNP discrimination using toehold-mediated DNA strand displacement. The distinctiveness of the approach is based on the combination of both 3- and 4-way branch migration mechanisms, which allows for reliable discrimination of SNPs within double-stranded DNA generated from real-life human mitochondrial DNA samples. Aside from the potential diagnostic value, the current study represents an additional way to control the strand displacement reaction rate without altering other reaction parameters and provides new insights into the influence of single nucleotide substitutions on 3- and 4-way branch migration efficiency and kinetics.

Repair of single-strand breaks in normal and trisomic lymphocytes

International Nuclear Information System (INIS)

Leonard, J.C.; Merz, T.

1982-01-01

Recently, Athanasiou and colleagues (1981) reported a deficiency in the capacity of lymphocytes from persons with Down's syndrome to repair single-strand DNA breaks. They found that 1 h after exposure to 160 Gray, repair processes had restored the sedimentation profile of DNA from normal lymphocytes to control values, whereas the relative average molecular weight of DNA from irradiated lymphocytes from persons with Down's syndrome showed no increase during the repair interval. They have suggested that their data, in conjunction with the earlier data concerning the frequencies of induced chromosomal aberrations in lymphocytes from persons with Down's syndrome, reflect a decreased efficiency in some aspect of DNA repair in trisomic cells. However, for further studies of this hypothesis, it is more appropriate to study the rejoining of DNA single-strand breaks after doses comparable to those used in tests for chromosomal aberrations. (orig.)

Identification of Cis-Acting Elements on Positive-Strand Subgenomic mRNA Required for the Synthesis of Negative-Strand Counterpart in Bovine Coronavirus

Directory of Open Access Journals (Sweden)

Po-Yuan Yeh

2014-07-01

Full Text Available It has been demonstrated that, in addition to genomic RNA, sgmRNA is able to serve as a template for the synthesis of the negative-strand [(−-strand] complement. However, the cis-acting elements on the positive-strand [(+-strand] sgmRNA required for (−-strand sgmRNA synthesis have not yet been systematically identified. In this study, we employed real-time quantitative reverse transcription polymerase chain reaction to analyze the cis-acting elements on bovine coronavirus (BCoV sgmRNA 7 required for the synthesis of its (−-strand counterpart by deletion mutagenesis. The major findings are as follows. (1 Deletion of the 5'-terminal leader sequence on sgmRNA 7 decreased the synthesis of the (−-strand sgmRNA complement. (2 Deletions of the 3' untranslated region (UTR bulged stem-loop showed no effect on (−-strand sgmRNA synthesis; however, deletion of the 3' UTR pseudoknot decreased the yield of (−-strand sgmRNA. (3 Nucleotides positioned from −15 to −34 of the sgmRNA 7 3'-terminal region are required for efficient (−-strand sgmRNA synthesis. (4 Nucleotide species at the 3'-most position (−1 of sgmRNA 7 is correlated to the efficiency of (−-strand sgmRNA synthesis. These results together suggest, in principle, that the 5'- and 3'-terminal sequences on sgmRNA 7 harbor cis-acting elements are critical for efficient (−-strand sgmRNA synthesis in BCoV.

Cisplatin enhances the formation of DNA single- and double-strand breaks by hydrated electrons and hydroxyl radicals.

Science.gov (United States)

Rezaee, Mohammad; Sanche, Léon; Hunting, Darel J

2013-03-01

The synergistic interaction of cisplatin with ionizing radiation is the clinical rationale for the treatment of several cancers including head and neck, cervical and lung cancer. The underlying molecular mechanism of the synergy has not yet been identified, although both DNA damage and repair processes are likely involved. Here, we investigate the indirect effect of γ rays on strand break formation in a supercoiled plasmid DNA (pGEM-3Zf-) covalently modified by cisplatin. The yields of single- and double-strand breaks were determined by irradiation of DNA and cisplatin/DNA samples with (60)Co γ rays under four different scavenging conditions to examine the involvement of hydrated electrons and hydroxyl radicals in inducing the DNA damage. At 5 mM tris in an N2 atmosphere, the presence of an average of two cisplatins per plasmid increased the yields of single- and double-strand breaks by factors of 1.9 and 2.2, respectively, relative to the irradiated unmodified DNA samples. Given that each plasmid of 3,200 base pairs contained an average of two cisplatins, this represents an increase in radiosensitivity of 3,200-fold on a per base pair basis. When hydrated electrons were scavenged by saturating the samples with N2O, these enhancement factors decreased to 1.5 and 1.2, respectively, for single- and double-strand breaks. When hydroxyl radicals were scavenged using 200 mM tris, the respective enhancement factors were 1.2 and 1.6 for single- and double-strand breaks, respectively. Furthermore, no enhancement in DNA damage by cisplatin was observed after scavenging both hydroxyl radicals and hydrated electrons. These findings show that hydrated electrons can induce both single- and double-strand breaks in the platinated DNA, but not in unmodified DNA. In addition, cisplatin modification is clearly an extremely efficient means of increasing the formation of both single- and double-strand breaks by the hydrated electrons and hydroxyl radicals created by ionizing

The European Nb3Sn advanced strand development programme

International Nuclear Information System (INIS)

Vostner, A.; Salpietro, E.

2005-01-01

Strands relevant for fusion with high critical current densities and moderate hysteresis losses were developed and already produced on industrial scale. Based on these achievements EFDA-CSU Garching has launched a Nb 3 Sn strand development and procurement action inside Europe in order to assess the current status of the Nb 3 Sn strand production capability. All six addressed companies have replied positively to the strand R and D programme which includes the three major Nb 3 Sn production techniques namely the bronze, internal-tin and powder-in-tube (PIT) route. According to the strand requirements for the ITER TF conductor a critical current density of 800 A/mm 2 (at 12 T, 4.2 K and 10 μV/m) and overall strand hysteresis losses below 500 kJ/m 3 have been specified as the minimum guaranteed strand performance. The second major objective of this programme is to motivate the strand manufacturers to develop and design high performance Nb 3 Sn strands optimised for the ITER conductor. For this purpose, a target critical current density of 1100 A/mm 2 has been added to the specification. This paper describes the strategy behind the strand development programme, the actual status of the strand production as well as first preliminary results obtained from the strand suppliers

R.b.e. of 50 kVp X-rays and 660 keV γ-rays (137Cs) with respect to the production of DNA damage, repair and cell-killing in Escherichia coli K-12

International Nuclear Information System (INIS)

Bonura, T.; Youngs, D.A.; Smith, K.C.

1975-01-01

A comparison has been made of the efficiency of cell-killing, DNA single-strand breakage and double-strand breakage in an Escherichia coli K-12 wild-type strain after irradiation with soft X-rays (50 kVp) and hard γ-rays (660 keV) under aerobic conditions. Irradiation with 50 kVp X-rays resulted in 1.47 times more cell-killing than was observed with 137 Cs γ-rays based on a comparison of D 0 values evaluated from the survival curves. DNA sedimentation studies showed that, although 50 kVp X-rays were 1.93 times more effective than 137 Cs γ-rays in producing DNA double-strand breaks, there was no significant difference between the two qualities of radiation with respect to the initial number of single-strand breaks produced. When the cells were irradiated and allowed to repair maximally in minimal medium, 1.57 times more unrepaired DNA single-strand breaks remained per krad after irradiation with 50 kVp X-rays than with 137 Cs γ-rays. The increased yield of DNA double-strand breaks resulting from 50 kVp X-irradiation may account for most of these additional unrepaired single-strand breaks, since single- and double-strand breaks are indistinguishable on alkaline sucrose gradients. These results suggest that the greater r.b.e. of 50 kVp X-rays may be related to an increased effectiveness for producing DNA double-strand breaks compared with the higher energy 137 Cs γ-rays. (author)

Inducted circulation current in a conductor consisting of strands coated with a high resistive layer

International Nuclear Information System (INIS)

Koizumi, Norikiyo; Takahashi, Yoshikazu; Kato, Takashi; Tsuji, Hiroshi; Shimamoto, Susumu

2000-01-01

Nonuniform current distribution is generated in a conductor consisting of strands coated by a high resistive layer, such as chromium plating, as a result of superimposition of transport and induce circulation currents. The characteristics of the induced circulation current are analytically studied by using a distributed model circuit. The parameters mostly used in this calculation are those of US-DPC coil, which at first exhibited instability and so-called ramp rate limitation (RRL) because of current imbalance in the conductor consisting of chrome-plated strands. Thus the conductance along strands and the inductance of unit length loop and length of the conductor are mostly assumed to be 10 kS/m, 0.5 μH/m and 150 m, respectively. The analysis results indicate that the induced circulation current can be classified into the boundary and interstrand-induce circulation currents hereafter referred to as BICC an IICC. BICC is induced only across the joint at the ends of the conductor, resulting in a constant along the conductor axis, when the total leakage magnetic flux of the loop is not zero. Its decay time constant is quite long, more than a few hours. In contrast, when the leakage magnetic flux distributes along the conductor axis, IICC is induced among strands in the conductor to eliminate this flux. Since the leakage magnetic flux normally becomes largest where the magnetic field is highest, it becomes larger where the time variation of the magnetic field is larger. Its decay time contrast is much less than that of BICC. If the leakage magnetic flux linearly changes along the US-DPC conductor, it is evaluated to be about 10 s. This IICC therefore becomes dominate in a pulse charge, whose ramping tine is less than 10 s. Moreover, it is found that the variation of the leakage a magnetic flux with the relatively long cycle, such as more than a few 10-meter lengths, causes IICC with a decay-time constant of more than several hundred milliseconds. Such and IICC can

Targeting abnormal DNA double strand break repair in cancer

OpenAIRE

Rassool, Feyruz V.; Tomkinson, Alan E.

2010-01-01

A major challenge in cancer treatment is the development of therapies that target cancer cells with little or no toxicity to normal tissues and cells. Alterations in DNA double strand break (DSB) repair in cancer cells include both elevated and reduced levels of key repair proteins and changes in the relative contributions of the various DSB repair pathways. These differences can result in increased sensitivity to DSB-inducing agents and increased genomic instability. The development of agent...

Studies of the effects of ultraviolet radiation on the structural integrities of ribosomal RNA components of the Escherichia coli 50S ribosomal subunit

International Nuclear Information System (INIS)

Gorelic, L.; Parker, D.

1978-01-01

The effects of 254-nm radiation on the structural integrities of free and 50S ribosome-bound 5S and 23S ribosomal ribonucleic acids (rRNA) have been elucidated. Irradiation of aqueous solutions of Escherichia coli 50S ribosomes with 253.7-nm radiation results in the formation of single-strand breaks in double-stranded regions of the 23S rRNA component, but not in rRNA chain scission, and destabilization of the secondary structure of the 23S rRNA toward denaturation. The minimum doses of 253.7-nm radiation required for the first detection of the two effects are 7 x 10 19 quanta for the production of single-strand breaks in double-stranded regions of the 23S rRNA, and 19 quanta for destabilization of the 23S rRNA secondary structure. Free 23S rRNA is resistant toward photoinduced chain breakage at doses of 253.7-nm radiation up to at least 2.3 x 10 20 and is much less sensitive toward destabilization of secondary structure than ribosome-bound 23S rRNA. In contrast to the photosensitivity of 50S ribosome-bound 23S rRNA toward chain breakage, 50S ribosome-bound 5S rRNA is resistant toward chain breakage at doses of 253.7-nm radiation up to at least 2.3 x 10 20 quanta. Ribosome-bound 5S and 23S rRNA are also not photosensitive toward intermolecular 5S/23S rRNA cross-linkage

Mechanism of replication of ultraviolet-irradiated single-stranded DNA by DNA polymerase III holoenzyme of Escherichia coli. Implications for SOS mutagenesis

International Nuclear Information System (INIS)

Livneh, Z.

1986-01-01

Replication of UV-irradiated oligodeoxynucleotide-primed single-stranded phi X174 DNA with Escherichia coli DNA polymerase III holoenzyme in the presence of single-stranded DNA-binding protein was investigated. The extent of initiation of replication on the primed single-stranded DNA was not altered by the presence of UV-induced lesions in the DNA. The elongation step exhibited similar kinetics when either unirradiated or UV-irradiated templates were used. Inhibition of the 3'----5' proofreading exonucleolytic activity of the polymerase by dGMP or by a mutD mutation did not increase bypass of pyrimidine photodimers, and neither did purified RecA protein influence the extent of photodimer bypass as judged by the fraction of full length DNA synthesized. Single-stranded DNA-binding protein stimulated bypass since in its absence the fraction of full length DNA decreased 5-fold. Termination of replication at putative pyrimidine dimers involved dissociation of the polymerase from the DNA, which could then reinitiate replication at other available primer templates. Based on these observations a model for SOS-induced UV mutagenesis is proposed

Study on radiogenotoxicological effect induced by retention of fission product 147Pm

International Nuclear Information System (INIS)

Zhu Shoupeng; Zheng Siying; Luo Zhongyu; Wang Liuyi; Zhao Xiuying

1988-01-01

Internal contamination of 147 Pm can be induced 4 chromatid breakage in one cell. This phenomenon might be due in part to nonuniform irradiation of bone marrow cells with local deposition of 147 Pm. In addition, among the types of aberrations induced by 147 Pm were predominantly of chromatid type, accompanied with a few chromosome breakage and translocation. A statistically significant elevation of SCEs was observed after 147 Pm injection. The number of SECs per cell in bone marrow cells were always higher in mice when the animals were maintained on the radioactive dose of 0.001 μCi/g level of 147 Pm. The results showed that obvious increase of mincronucleus rates could be induced by 147 Pm with low dose as 0.005 μCi/g. While the peak value appeared at 1.34% after injecting 5 μCu/g of 147 Pm for 24hr. In general, the induction of chromosomal alterations by 147 Pm to induce chromosome aberration was only 0.001 μGi/g. This value was lower than ALI of 147 Pm, as reported by ICRP publication

Characterizing the strand-specific distribution of non-CpG methylation in human pluripotent cells.

Science.gov (United States)

Guo, Weilong; Chung, Wen-Yu; Qian, Minping; Pellegrini, Matteo; Zhang, Michael Q

2014-03-01

DNA methylation is an important defense and regulatory mechanism. In mammals, most DNA methylation occurs at CpG sites, and asymmetric non-CpG methylation has only been detected at appreciable levels in a few cell types. We are the first to systematically study the strand-specific distribution of non-CpG methylation. With the divide-and-compare strategy, we show that CHG and CHH methylation are not intrinsically different in human embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). We also find that non-CpG methylation is skewed between the two strands in introns, especially at intron boundaries and in highly expressed genes. Controlling for the proximal sequences of non-CpG sites, we show that the skew of non-CpG methylation in introns is mainly guided by sequence skew. By studying subgroups of transposable elements, we also found that non-CpG methylation is distributed in a strand-specific manner in both short interspersed nuclear elements (SINE) and long interspersed nuclear elements (LINE), but not in long terminal repeats (LTR). Finally, we show that on the antisense strand of Alus, a non-CpG site just downstream of the A-box is highly methylated. Together, the divide-and-compare strategy leads us to identify regions with strand-specific distributions of non-CpG methylation in humans.

Increased sister chromatid cohesion and DNA damage response factor localization at an enzyme-induced DNA double-strand break in vertebrate cells.

LENUS (Irish Health Repository)

Dodson, Helen

2009-10-01

The response to DNA damage in vertebrate cells involves successive recruitment of DNA signalling and repair factors. We used light microscopy to monitor the genetic dependencies of such localization to a single, induced DNA double strand break (DSB) in vertebrate cells. We used an inducible version of the rare-cutting I-SceI endonuclease to cut a chromosomally integrated I-SceI site beside a Tet operator array that was visualized by binding a Tet repressor-GFP fusion. Formation of gamma-H2AX foci at a single DSB was independent of ATM or Ku70. ATM-deficient cells showed normal kinetics of 53Bp1 recruitment to DSBs, but Rad51 localization was retarded. 53Bp1 and Rad51 foci formation at a single DSB was greatly reduced in H2AX-null DT40 cells. We also observed decreased inter-sister chromatid distances after DSB induction, suggesting that cohesin loading at DSBs causes elevated sister chromatid cohesion. Loss of ATM reduced DSB-induced cohesion, consistent with cohesin being an ATM target in the DSB response. These data show that the same genetic pathways control how cells respond to single DSBs and to multiple lesions induced by whole-cell DNA damage.

Down-Regulation of p53 by Double-Stranded RNA Modulates the Antiviral Response

OpenAIRE

Marques, Joao T.; Rebouillat, Dominique; Ramana, Chilakamarti V.; Murakami, Junko; Hill, Jason E.; Gudkov, Andrei; Silverman, Robert H.; Stark, George R.; Williams, Bryan R. G.

2005-01-01

p53 has been well characterized as a tumor suppressor gene, but its role in antiviral defense remains unclear. A recent report has demonstrated that p53 can be induced by interferons and is activated after vesicular stomatitis virus (VSV) infection. We observed that different nononcogenic viruses, including encephalomyocarditis virus (EMCV) and human parainfluenza virus type 3 (HPIV3), induced down-regulation of p53 in infected cells. Double-stranded RNA (dsRNA) and a mutant vaccinia virus la...

DNA cleavage agents from Schisandra propinqua var. sinensis

African Journals Online (AJOL)

STORAGESEVER

2009-09-15

Sep 15, 2009 ... 2Engineering Research Center of Sustainable Development and Utilization of Biomass Energy, Ministry of ... DNA strand breakage process is involved in various bio- ..... Bioactive prenylated flavonoids from the stem bark.

Ionisation and dissociation of water induced by swift multicharged ions

International Nuclear Information System (INIS)

Legendre, S.

2006-02-01

Ionization and dissociation of water molecules and water clusters induced by 11.7 MeV/A Ni 25+ ions were carried out by imaging techniques. Branching ratios, ionisation cross sections and Kinetic Energy Released distributions have been measured together with fragmentation dynamics studies. Multiple ionization represents approximately 30% of the ionizing events. Double ionization produces in significant way atomic oxygen, considered as a possible precursor of the large production of HO 2 radical in liquid water radiolysis by ions of high Linear Energy Transfer. We evidence a strong selectivity of bond breakage in the case of ion-induced HOD fragmentation. Once the molecule doubly ionized, the breakage of the O-H bond is found 6.5 times more probable than that of the O-D bond. A semi-classical calculation simulating the fragmentation dynamics on the potential energy surface of the ground-state of di-cation H 2 O 2+ makes possible to as well reproduce the preferential nature of the breakage of the O-H bond as the position and the shift of the kinetic energy distributions. First results concerning interaction with water clusters are also reported. Measurements in coincidence are carried out giving access to correlation, with the distributions in energy and angle of the emitted fragments. Mass spectrum points fast intra-cluster proton transfer, leading to the emission of protonated clusters. (author)

Caffeine inhibits homology-directed repair of I-SceI-induced DNA double-strand breaks.

Science.gov (United States)

Wang, Huichen; Boecker, Wilfried; Wang, Hongyan; Wang, Xiang; Guan, Jun; Thompson, Larry H; Nickoloff, Jac A; Iliakis, George

2004-01-22

We recently reported that two Chinese hamster mutants deficient in the RAD51 paralogs XRCC2 and XRCC3 show reduced radiosensitization after treatment with caffeine, thus implicating homology-directed repair (HDR) of DNA double-strand breaks (DSBs) in the mechanism of caffeine radiosensitization. Here, we investigate directly the effect of caffeine on HDR initiated by DSBs induced by a rare cutting endonuclease (I-SceI) into one of two direct DNA repeats. The results demonstrate a strong inhibition by caffeine of HDR in wild-type cells, and a substantial reduction of this effect in HDR-deficient XRCC3 mutant cells. Inhibition of HDR and cell radiosensitization to killing shows similar dependence on caffeine concentration suggesting a cause-effect relationship between these effects. UCN-01, a kinase inhibitor that effectively abrogates checkpoint activation in irradiated cells, has only a small effect on HDR, indicating that similar to radiosensitization, inhibition of checkpoint signaling is not sufficient for HDR inhibition. Recombination events occurring during treatment with caffeine are characterized by rearrangements reminiscent to those previously reported for the XRCC3 mutant, and immunofluorescence microscopy demonstrates significantly reduced formation of IR-specific RAD51 foci after caffeine treatment. In summary, our results identify inhibition of HDR as a significant contributor to caffeine radiosensitization.

Molecular mechanisms of induced mutagenesis. Replication in vivo of bacteriophage phiX174 single-stranded, ultraviolet light-irradiated DNA in intact and irradiated host cells

Energy Technology Data Exchange (ETDEWEB)

Caillet-Fauquet, P; Defais, M; Radman, M [Brussels Univ. (Belgium)

1977-11-25

Genetic analysis has revealed that radiation and many chemical mutagens induce in bacteria an error-prone DNA repair process which is responsible for their mutagenic effect. The biochemical mechanism of this inducible error-prone repair has been studied by analysis of the first round of DNA synthesis on ultraviolet light-irradiated phiX174 DNA in both intact and ultraviolet light-irradiated host cells. Intracellular phiX174 DNA was extracted, subjected to isopycnic CsCl density-gradient analysis, hydroxylapatite chromatography and digestion by single-strand-specific endonuclease S/sub 1/. Ultraviolet light-induced photolesions in viral DNA cause a permanent blockage of DNA synthesis in intact Escherichia coli cells. However, when host cells were irradiated and incubated to induce fully the error-prone repair system, a significant fraction of irradiated phiX174 DNA molecules can be fully replicated. Thus, inducible error-prone repair in E.coli is manifested by an increased capacity for DNA synthesis on damaged phiX174 DNA. Chloramphenicol (100 ..mu.. g/ml), which is an inhibitor of the inducible error-prone DNA repair, is also an inhibitor of this particular inducible DNA synthesis.

A label-free ultrasensitive fluorescence detection of viable Salmonella enteritidis using enzyme-induced cascade two-stage toehold strand-displacement-driven assembly of G-quadruplex DNA.

Science.gov (United States)

Zhang, Peng; Liu, Hui; Ma, Suzhen; Men, Shuai; Li, Qingzhou; Yang, Xin; Wang, Hongning; Zhang, Anyun

2016-06-15

The harm of Salmonella enteritidis (S. enteritidis ) to public health mainly by contaminating fresh food and water emphasizes the urgent need for rapid detection techniques to help control the spread of the pathogen. In this assay, an newly designed capture probe complex that contained specific S. enteritidis-aptamer and hybridized signal target sequence was used for viable S. enteritidis recognition directly. In the presence of the target S. enteritidis, single-stranded target sequences were liberated and initiated the replication-cleavage reaction, producing numerous G-quadruplex structures with a linker on the 3'-end. And then, the sensing system took innovative advantage of quadratic linker-induced strand-displacement for the first time to release target sequence in succession, leading to the cyclic reuse of the target sequences and cascade signal amplification, thereby achieving the successive production of G-quadruplex structures. The fluorescent dye, N-Methyl mesoporphyrin IX, binded to these G-quadruplex structures and generated significantly enhanced fluorescent signals to achieve highly sensitive detection of S. enteritidis down to 60 CFU/mL with a linear range from 10(2) to 10(7)CFU/mL. By coupling the cascade two-stage target sequences-recyclable toehold strand-displacement with aptamer-based target recognition successfully, it is the first report on a novel non-label, modification-free and DNA extraction-free ultrasensitive fluorescence biosensor for detecting viable S. enteritidis directly, which can discriminate from dead S. enteritidis. Copyright © 2016 Elsevier B.V. All rights reserved.

Prediction of guide strand of microRNAs from its sequence and secondary structure

Directory of Open Access Journals (Sweden)

Ahmed Firoz

2009-04-01

Full Text Available Abstract Background MicroRNAs (miRNAs are produced by the sequential processing of a long hairpin RNA transcript by Drosha and Dicer, an RNase III enzymes, and form transitory small RNA duplexes. One strand of the duplex, which incorporates into RNA-induced silencing complex (RISC and silences the gene expression is called guide strand, or miRNA; while the other strand of duplex is degraded and called the passenger strand, or miRNA*. Predicting the guide strand of miRNA is important for better understanding the RNA interference pathways. Results This paper describes support vector machine (SVM models developed for predicting the guide strands of miRNAs. All models were trained and tested on a dataset consisting of 329 miRNA and 329 miRNA* pairs using five fold cross validation technique. Firstly, models were developed using mono-, di-, and tri-nucleotide composition of miRNA strands and achieved the highest accuracies of 0.588, 0.638 and 0.596 respectively. Secondly, models were developed using split nucleotide composition and achieved maximum accuracies of 0.553, 0.641 and 0.602 for mono-, di-, and tri-nucleotide respectively. Thirdly, models were developed using binary pattern and achieved the highest accuracy of 0.708. Furthermore, when integrating the secondary structure features with binary pattern, an accuracy of 0.719 was seen. Finally, hybrid models were developed by combining various features and achieved maximum accuracy of 0.799 with sensitivity 0.781 and specificity 0.818. Moreover, the performance of this model was tested on an independent dataset that achieved an accuracy of 0.80. In addition, we also compared the performance of our method with various siRNA-designing methods on miRNA and siRNA datasets. Conclusion In this study, first time a method has been developed to predict guide miRNA strands, of miRNA duplex. This study demonstrates that guide and passenger strand of miRNA precursors can be distinguished using their

Defective processing of methylated single-stranded DNA by E. coli alkB mutants

Science.gov (United States)

Dinglay, Suneet; Trewick, Sarah C.; Lindahl, Tomas; Sedgwick, Barbara

2000-01-01

Escherichia coli alkB mutants are very sensitive to DNA methylating agents. Despite these mutants being the subject of many studies, no DNA repair or other function has been assigned to the AlkB protein or to its human homolog. Here, we report that reactivation of methylmethanesulfonate (MMS)-treated single-stranded DNA phages, M13, f1, and G4, was decreased dramatically in alkB mutants. No such decrease occurred when using methylated λ phage or M13 duplex DNA. These data show that alkB mutants have a marked defect in processing methylation damage in single-stranded DNA. Recombinant AlkB protein bound more efficiently to single- than double-stranded DNA. The single-strand damage processed by AlkB was primarily cytotoxic and not mutagenic and was induced by SN2 methylating agents, MMS, DMS, and MeI but not by SN1 agent N-methyl-N-nitrosourea or by γ irradiation. Strains lacking other DNA repair activities, alkA tag, xth nfo, uvrA, mutS, and umuC, were not defective in reactivation of methylated M13 phage and did not enhance the defect of an alkB mutant. A recA mutation caused a small but additive defect. Thus, AlkB functions in a novel pathway independent of these activities. We propose that AlkB acts on alkylated single-stranded DNA in replication forks or at transcribed regions. Consistent with this theory, stationary phase alkB cells were less MMS sensitive than rapidly growing cells. PMID:10950872

Force-induced rupture of double-stranded DNA in the absence and presence of covalently bonded anti-tumor drugs: Insights from molecular dynamics simulations

Science.gov (United States)

Upadhyaya, Anurag; Nath, Shesh; Kumar, Sanjay

2018-06-01

DNA intra-strand cross-link (ICL) agents are widely used in the treatment of cancer. ICLs are thought to form a link between the same strand (intra-strand) or complimentary strand (inter-strand) and thereby increase the stability of DNA, which forbids the processes like replication and transcription. As a result, cell death occurs. In this work, we have studied the enhanced stability of a double stranded DNA in the presence of ICLs and compared our findings with the results obtained in the absence of these links. Using atomistic simulations with explicit solvent, a force is applied along and perpendicular to the direction of the helix and we measured the rupture force and the unzipping force of DNA-ICL complexes. Our results show that the rupture and the unzipping forces increase significantly in the presence of these links. The ICLs bind to the minor groove of DNA, which enhance the DNA stabilisation. Such information may be used to design alternative drugs that can stall replication and transcription that are critical to a growing number of anticancer drug discovery efforts.

A correlation between residual radiation-induced DNA double-strand breaks in cultured fibroblasts and late radiotherapy reactions in breast cancer patients

International Nuclear Information System (INIS)

Kiltie, A.E.; Ryan, A.J.; Swindell, R.; Barber, J.B.P.; West, C.M.L.; Magee, B.; Hendry, J.H.

1999-01-01

Background and purpose: Prediction of late normal tissue reactions to radiotherapy would permit tailoring of dosage to each patient. Measurement of residual DNA double strand breaks using pulsed field gel electrophoresis (PFGE) shows promise in this field. The aim of this study was to test the predictive potential of PFGE in a group of retrospectively studied breast cancer patients.Materials and methods: Thirty nine patients, treated uniformly for breast cancer 9-15 years previously, with excision of the tumour and radiotherapy to the breast and drainage areas, were assessed clinically using the LENT SOMA scale, and a 5-mm punch biopsy taken from the buttock. Fibroblast cell strains were established and used to study residual DNA double strand breaks, using PFGE.Results: There were significant correlations between the DNA assay results and the fibrosis score (r s =0.46; P=0.003), the combined fibrosis and retraction score (r s =0.45, P=0.004) and the overall LENT score (r s =0.43; P=0.006). Using polychotomous logistic regression, the fibroblast DNA assay result was an independent prognostic factor for fibrosis severity.Conclusions: There is a relationship between residual radiation-induced DNA damage in fibroblasts and the severity of the late normal tissue damage seen in the patients from whom the cells were cultured. (Copyright (c) 1999 Elsevier Science B.V., Amsterdam. All rights reserved.)

When, where and how the bridge breaks: anaphase bridge breakage plays a crucial role in gene amplification and HSR generation

International Nuclear Information System (INIS)

Shimizu, Noriaki; Shingaki, Kenta; Kaneko-Sasaguri, Yukiko; Hashizume, Toshihiko; Kanda, Teru

2005-01-01

Amplified genes are frequently localized on extrachromosomal double minutes (DMs) or in chromosomal homogenously staining regions (HSRs). We previously showed that a plasmid bearing a mammalian replication initiation region could efficiently generate DMs and HSRs after transfection into human tumor cell lines. The Breakage-Fusion-Bridge (BFB) cycle model, a classical model that explains how HSRs form, could also be used to explain how the transfected plasmids generate HSRs. The BFB cycle model involves anaphase bridge formation due to the presence of dicentric chromosomes, followed by the breakage of the bridge. In this study, we used our plasmid-based model system to analyze how anaphase bridges break during mitosis. Dual-color fluorescence in situ hybridization analyses revealed that anaphase bridges were most frequently severed in their middle irrespective of their lengths, which suggests that a structurally fragile site exists in the middle of the anaphase bridge. Breakage of the chromosomal bridges occurred prior to nuclear membrane reformation and the completion of cytokinesis, which indicates that mechanical tension rather than cytokinesis is primarily responsible for severing anaphase bridges. Time-lapse observation of living cells revealed that the bridges rapidly shrink after being severed. If HSR length was extended too far, the bridge could no longer be resolved and became tangled depending on the tension. The unbroken bridge appeared to inhibit the completion of cytokinesis. These observations strongly suggest that anaphase bridges are highly elastic and that the length of the spindle axis determines the maximal HSR length

DNA strand breaks, repair, and survival in x-irradiated mammalian cells

International Nuclear Information System (INIS)

Dugle, D.L.; Gillespie, C.J.; Chapman, J.D.

1976-01-01

The yields of unrepairable single- and double-strand breaks in the DNA of x-irradiated Chinese hamster cells were measured by low-speed neutral and alkaline sucrose density gradient sedimentation in order to investigate the relation between these lesions and reproductive death. After maximal single-strand rejoining, at all doses, the number of residual single-strand breaks was twice the number of residual double-strand breaks. Both double-strand and unrepairable single-strand breaks were proportional to the square of absorbed dose, in the range 10-50 krad. No rejoining of double-strand breaks was observed. These observations suggest that, in mammalian cells, most double-strand breaks are not repairable, while all single-strand breaks are repaired except those that are sufficiently close on complementary strands to constitute double-strand breaks. Comparison with cell survival measurements at much lower doses suggests that loss of reproductive capacity corresponds to induction of approximately one double-strand break

Ultraviolet light induces double-strand breaks in DNA of cultured human P3 cells as measured by neutral filter elution

International Nuclear Information System (INIS)

Peak, J.G.; Peak, M.J.

1990-01-01

Neutral filter elution at pH 7.2 and 9.6 was used to measure the induction of DNA lesions in human P3 teratocarcinoma cells by monochromatic 254-, 270-, 313-, 334-, 334-,365-, and 405-nm radiation and by 60 gamma rays. In this assay DNA double-strand breaks (dsb) increase the rate of elution of DNA from cell lysates on a filter. Yields of dsb as measured by this procedure were determined by using a calibration of the assay that correlates elution parameters with number of dsb caused by disintegration of 125 I incorporated into the DNA. Analysis of fluence responses obtained by using the calibrated assay indicated that the number of dsb induced per dalton of DNA as measured by this assay is proportional to the square of the fluence at all the energies of radiation studied, implying that the induction of these lesions may be a two-hit event. Analysis of the relative efficiencies for the induction of dsb by ultraviolet radiation, corrected for quantum efficiency, revealed a spectrum that coincided closely with that for the induction of single-strand breaks (ssb) in the same cells, having a close fit with the spectrum of nucleic acid in the UVC and UVB region below 313 nm, and a shoulder in the UVA region. It was calculated, however, that there may be too few ssb for dsb to result from randomly distributed closely opposed ssb. (author)

Simulating mechanisms for dispersal, production and stranding of small forage fish in temporary wetland habitats

Science.gov (United States)

Yurek, Simeon; DeAngelis, Donald L.; Trexler, Joel C.; Jopp, Fred; Donalson, Douglas D.

2013-01-01

Movement strategies of small forage fish (wetland habitats affect their overall population growth and biomass concentrations, i.e., availability to predators. These fish are often the key energy link between primary producers and top predators, such as wading birds, which require high concentrations of stranded fish in accessible depths. Expansion and contraction of seasonal wetlands induce a sequential alternation between rapid biomass growth and concentration, creating the conditions for local stranding of small fish as they move in response to varying water levels. To better understand how landscape topography, hydrology, and fish behavior interact to create high densities of stranded fish, we first simulated population dynamics of small fish, within a dynamic food web, with different traits for movement strategy and growth rate, across an artificial, spatially explicit, heterogeneous, two-dimensional marsh slough landscape, using hydrologic variability as the driver for movement. Model output showed that fish with the highest tendency to invade newly flooded marsh areas built up the largest populations over long time periods with stable hydrologic patterns. A higher probability to become stranded had negative effects on long-term population size, and offset the contribution of that species to stranded biomass. The model was next applied to the topography of a 10 km × 10 km area of Everglades landscape. The details of the topography were highly important in channeling fish movements and creating spatiotemporal patterns of fish movement and stranding. This output provides data that can be compared in the future with observed locations of fish biomass concentrations, or such surrogates as phosphorus ‘hotspots’ in the marsh.

Calibration of pulsed field gel electrophoresis for measurement of DNA double-strand breaks

International Nuclear Information System (INIS)

Ager, D.D.; Dewey, W.C.

1990-01-01

Pulsed field gel electrophoresis (PFGE) assay was calibrated for the measurement of X-ray induced DNA double-strand breaks in Chinese hamster ovary (CHO) cells. Calibration was conducted by incorporating [ 125 I] deoxyuridine into DNA, which induces one double-strand break for every disintegration that occurs in frozen cells. Based on the percentage of DNA migrating into the gel, the number of breaks/dalton/Gy was estimated to be (9.3±1.0) x 10 -12 . This value is close to (10 to 12) x 10 -12 determined by neutral filter elution using similar cell lysis procedures at 24 o C and at pH8.0. The estimate is in good agreement with the value of (11.7±2) x 10 -12 breaks/dalton/Gy as measured in Ehrlich ascites tumour cells using the neutral sucrose gradient method (Bloecher 1988), and (6 to 9) x 10 -12 breaks/dalton/Gy as measured in mouse L and Chinese hamster V79 cells using neutral filter elution (Radford and Hodgson 1985). (author)

Morphology and mechanical properties of multi-stranded amyloid fibrils probed by atomistic and coarse-grained simulations

International Nuclear Information System (INIS)

Yoon, Gwonchan; Lee, Myeongsang; Kim, Kyungwoo; In Kim, Jae; Joon Chang, Hyun; Baek, Inchul; Na, Sungsoo; Eom, Kilho

2015-01-01

Amyloid fibrils are responsible for pathogenesis of various diseases and exhibit the structural feature of an ordered, hierarchical structure such as multi-stranded helical structure. As the multi-strandedness of amyloid fibrils has recently been found to be highly correlated with their toxicity and infectivity, it is necessary to study how the hierarchical (i.e. multi-stranded) structure of amyloid fibril is formed. Moreover, although it has recently been reported that the nanomechanics of amyloid proteins plays a key role on the amyloid-induced pathogenesis, a critical role that the multi-stranded helical structure of the fibrils plays in their nanomechanical properties has not fully characterized. In this work, we characterize the morphology and mechanical properties of multi-stranded amyloid fibrils by using equilibrium molecular dynamics simulation and elastic network model. It is shown that the helical pitch of multi-stranded amyloid fibril is linearly proportional to the number of filaments comprising the amyloid fibril, and that multi-strandedness gives rise to improving the bending rigidity of the fibril. Moreover, we have also studied the morphology and mechanical properties of a single protofilament (filament) in order to understand the effect of cross-β structure and mutation on the structures and mechanical properties of amyloid fibrils. Our study sheds light on the underlying design principles showing how the multi-stranded amyloid fibril is formed and how the structure of amyloid fibrils governs their nanomechanical properties. (paper)

Acid polypeptides as inhibitors of the repair of double-strand DNA breaks induced by γ-irradiation of Hela cells

International Nuclear Information System (INIS)

Medvedev, A.I.; Revina, G.I.; Kuzin, A.M.

1990-01-01

The effect of natural modificator's synthetic analogue -polyaspartylglytamate (AG) - on the repair of radiation-induced double-strand DNA breaks is studies. The radiation and modificator effects were determined by the criterion of the formation of chromosome recombinations and reproductive death of cells on Hela cell culture and in Chinese hamsters. It is shown that the incubation of Hela cells with AG doubles and triples the degradation effect of rdiation at 50 and 10 Gy doses. When radiation dose equals 1 Gy and repair time is G-22 h, 1.5 - 3 time - increased yield of chromotide and chromosome abberations is detected in Chinese hamster cells in the presence of the modificator during all periods of cell fixation. The effect of radiation mutagenic action enhancement by the modificator is not observed during the incubation of cells with AG 30-45 min after irradiation

Effect of nalidixic acid on repair of single-strand breaks in DNA induced by ionizing irradiation in Escherichia coli

Energy Technology Data Exchange (ETDEWEB)

Francia, I [Debreceni Orvostudomanyi Egyetem (Hungary); Okos, A; Hernadi, F J [Institute of Pharmacology, Debrecen (Hungary)

1978-09-30

The incidence of DNA single-strand breaks induced by /sup 60/Co irradiation and their repair in E.coli K12 (AB 1157) rec/sup +/ cells were studied by the alkaline sucrose gradient sedimentation method described by McGrath and Williams. For the quantitative analysis of sedimentation profiles we used the s 1/2 values described by Veatch and Okada. The s 1/2 value of non-irradiated controls was 22.4, and after 20 krads irradiation it was found to be 11.7. A postirradiation incubation at 37 /sup 0/C for 60 min increasedthe s 1/2 value from 11.7 to 22.1. Nalidixic acid at low concentration (20-50 ..mu..g/ml) did not block, but at 100 ..mu..g/ml extensively inhibited the above repair process, exhibiting an s 1/2 value of 14.4.

Effects of 3-Deoxyadenosine (Cordycepin) on the repair of X-ray-induced DNA single- and double-strand breaks in chinese hamster V79 cells

International Nuclear Information System (INIS)

Hiraoka, Wakako; Kuwabara, Mikinori; Sato, Fumiaki

1990-01-01

The ability of cordycepin to inhibit the repair of DNA strand breaks was examined with X-irradiated Chinese hamster V79 cells in log-phase culture. A filter elution technique revealed that 70 μM cordycepin did not inhibit the repair of single-strand breaks but inhibited the repair of double-strand breaks. These findings confirmed the fact that the increase in the lethality of cordycepin in X-irradiated cultured mammalian cells was attributable to unrepaired DNA double-strand breaks. (author)

Detection of bacteriophage phi 6 minus-strand RNA and novel mRNA isoconformers synthesized in vivo and in vitro, by strand-separating agarose gels

International Nuclear Information System (INIS)

Pagratis, N.; Revel, H.R.

1990-01-01

Two urea-free agarose gel protocols that resolve the six individual strands of bacteriophage phi 6 dsRNA were developed and used to analyze phage RNA synthesis in vivo and in vitro. Citrate gels separate strands of the large and medium chromosomes while Tris-borate-EDTA (TBE) gels resolve the medium and small dsRNA segments. Minus strands migrate faster than plus strands on citrate gels but are retarded on TBE gels. A study of electrophoretic conditions showed that pH affects strand resolution on citrate gels, and that voltage gradient, agarose concentration, and ethidium bromide significantly alter strand migration on TBE gels. Analysis of native phi 6 RNA synthesized in vivo and in vitro showed that the large and medium message RNAs comigrate with the corresponding plus strands of denatured virion dsRNA. The small messenger RNA is exceptional. Native small mRNA was detected as three isoconformers in vivo and in vitro. The isoconformers were converted by heat denaturation to a single RNA species that comigrates with the virion s+ strand. Minus strands labeled in vivo were detected only after heat denaturation. Minus strand synthesis was detected also in heat-denatured samples from in vitro phi 6 nucleocapsid RNA polymerase reactions at pH values suboptimal for transcription

Analysis of DNA polymerase activity in Petunia protoplasts treated with clastogenic agents

International Nuclear Information System (INIS)

Benediktsson, I.; Spampinato, C.P.; Andreo, C.S.; Schieder, O.

1994-01-01

Clastogenic agents, i.e. agents that can induce chromosome or DNA breakage, have been shown to enhance the role of direct gene transfer to protoplasts. The effect was analysed at the enzymatic level using protoplast homogenates as well as intact protoplasts. For that purpose existing procedures were modified to enable measurement of DNA polymerase in vivo. In the system used, external DNA was able to enter the cells without the addition of membrane-permeabilizing compounds. When comparing total DNA polymerase activity of protoplasts irradiated with X-rays or UV-light with that of untreated cells we did not observe significant differences. Incubation of protoplasts with high doses of bleomycin affected total DNA polymerase activity negatively. but dideoxythymidine triphosphate-sensitive activity was not influenced. We conclude that the DNA strand-breaks induced by low doses of X-rays. UV-light or bleomycin do not increase the total or the repair-DNA polymerase activity and. therefore. that the increase in the transformation rates after DNA strand-breaking is not preceded by enhanced DNA polymerase activity. (author)

Dynamics of Leading-strand Lesion Skipping by the Replisome

Science.gov (United States)

Yeeles, Joseph T.P.; Marians, Kenneth J.

2013-01-01

SUMMARY The E. coli replisome stalls transiently when it encounters a lesion in the leading-strand template, skipping over the damage by reinitiating replication at a new primer synthesized downstream by the primase. We report here that template unwinding and lagging-strand synthesis continue downstream of the lesion at a reduced rate after replisome stalling, that one replisome is capable of skipping multiple lesions, and that the rate limiting steps of replication restart involve the synthesis and activation of the new primer downstream. We also find little support for the concept that polymerase uncoupling, where extensive lagging-strand synthesis proceeds downstream in the absence of leading-strand synthesis, involves physical separation of the leading-strand polymerase from the replisome. Instead, our data indicate that extensive uncoupled replication likely results from a failure of the leading-strand polymerase still associated with the DNA helicase and the lagging-strand polymerase that are proceeding downstream to reinitiate synthesis. PMID:24268579

Southeast US Historical Marine Mammal Stranding Database

Data.gov (United States)

National Oceanic and Atmospheric Administration, Department of Commerce — Data on marine mammal strandings are collected by the Southeast Marine Mammal Stranding Network. Basic data on the location, species identification, animal...

High incidence of non-random template strand segregation and asymmetric fate determination in dividing stem cells and their progeny.

Science.gov (United States)

Conboy, Michael J; Karasov, Ariela O; Rando, Thomas A

2007-05-01

Decades ago, the "immortal strand hypothesis" was proposed as a means by which stem cells might limit acquiring mutations that could give rise to cancer, while continuing to proliferate for the life of an organism. Originally based on observations in embryonic cells, and later studied in terms of stem cell self-renewal, this hypothesis has remained largely unaccepted because of few additional reports, the rarity of the cells displaying template strand segregation, and alternative interpretations of experiments involving single labels or different types of labels to follow template strands. Using sequential pulses of halogenated thymidine analogs (bromodeoxyuridine [BrdU], chlorodeoxyuridine [CldU], and iododeoxyuridine [IdU]), and analyzing stem cell progeny during induced regeneration in vivo, we observed extraordinarily high frequencies of segregation of older and younger template strands during a period of proliferative expansion of muscle stem cells. Furthermore, template strand co-segregation was strongly associated with asymmetric cell divisions yielding daughters with divergent fates. Daughter cells inheriting the older templates retained the more immature phenotype, whereas daughters inheriting the newer templates acquired a more differentiated phenotype. These data provide compelling evidence of template strand co-segregation based on template age and associated with cell fate determination, suggest that template strand age is monitored during stem cell lineage progression, and raise important caveats for the interpretation of label-retaining cells.

High incidence of non-random template strand segregation and asymmetric fate determination in dividing stem cells and their progeny.

Directory of Open Access Journals (Sweden)

Michael J Conboy

2007-05-01

Full Text Available Decades ago, the "immortal strand hypothesis" was proposed as a means by which stem cells might limit acquiring mutations that could give rise to cancer, while continuing to proliferate for the life of an organism. Originally based on observations in embryonic cells, and later studied in terms of stem cell self-renewal, this hypothesis has remained largely unaccepted because of few additional reports, the rarity of the cells displaying template strand segregation, and alternative interpretations of experiments involving single labels or different types of labels to follow template strands. Using sequential pulses of halogenated thymidine analogs (bromodeoxyuridine [BrdU], chlorodeoxyuridine [CldU], and iododeoxyuridine [IdU], and analyzing stem cell progeny during induced regeneration in vivo, we observed extraordinarily high frequencies of segregation of older and younger template strands during a period of proliferative expansion of muscle stem cells. Furthermore, template strand co-segregation was strongly associated with asymmetric cell divisions yielding daughters with divergent fates. Daughter cells inheriting the older templates retained the more immature phenotype, whereas daughters inheriting the newer templates acquired a more differentiated phenotype. These data provide compelling evidence of template strand co-segregation based on template age and associated with cell fate determination, suggest that template strand age is monitored during stem cell lineage progression, and raise important caveats for the interpretation of label-retaining cells.

The stranding anomaly as population indicator

DEFF Research Database (Denmark)

Peltier, Helene; Baagøe, Hans J.; Camphuysen, Kees C. J.

2013-01-01

Ecological indicators for monitoring strategies are expected to combine three major characteristics: ecological significance, statistical credibility, and cost-effectiveness. Strategies based on stranding networks rank highly in cost-effectiveness, but their ecological significance and statistica...... surveys, mostly SCANS surveys (1994 and 2005). This new indicator could be applied to cetacean populations across the world and more widely to marine megafauna....... credibility are disputed. Our present goal is to improve the value of stranding data as population indicator as part of monitoring strategies by constructing the spatial and temporal null hypothesis for strandings. The null hypothesis is defined as: small cetacean distribution and mortality are uniform...

Formation of double-strand breaks in DNA of γ-irradiated bacteria depending on the function of fast repair processes of DNA single-strand breaks

International Nuclear Information System (INIS)

Petrov, S.I.; Gaziev, A.I.

1980-01-01

The formation of double-strand breaks in DNA of γ-irradiated ( 60 Co)Ex coli bacteria depending on the function of fast repair processes of DNA single-strand breaks, is investigated. The profiles of sedimentation of DNA Ex coli cells, irradiated at 0-2 deg C in the salt medium and in EDTA-borate buffer, are presented. It is shown that when irradiating cells in EDTA-borate buffer, the output of single- and double strand breaks in DNA is much higher than in the case of their irradiation in the minimum salt medium. The dependence of output of single-strand and double-strand breaks depending on the radiatier doze of E coli cells in the salt medium and EDTA-borate buffer, is studied. The supposition is made on the presence of a regulative interaction between the accumulation of DNA single-breaks and their repair with the formation of double-strand breaks. The functionating of fast and superfast repair processes considerably affects the formation of double-strand breaks in DNA of a bacterium cell. A considerable amount of double-breaks registered immediately after irradiation forms due to a close position of single-strand breaks on the opposite DNA strands

Cytokine overproduction and crosslinker hypersensitivity are unlinked in Fanconi anemia macrophages.

Science.gov (United States)

Garbati, Michael R; Hays, Laura E; Rathbun, R Keaney; Jillette, Nathaniel; Chin, Kathy; Al-Dhalimy, Muhsen; Agarwal, Anupriya; Newell, Amy E Hanlon; Olson, Susan B; Bagby, Grover C

2016-03-01

The Fanconi anemia proteins participate in a canonical pathway that repairs cross-linking agent-induced DNA damage. Cells with inactivated Fanconi anemia genes are universally hypersensitive to such agents. Fanconi anemia-deficient hematopoietic stem cells are also hypersensitive to inflammatory cytokines, and, as importantly, Fanconi anemia macrophages overproduce such cytokines in response to TLR4 and TLR7/8 agonists. We questioned whether TLR-induced DNA damage is the primary cause of aberrantly regulated cytokine production in Fanconi anemia macrophages by quantifying TLR agonist-induced TNF-α production, DNA strand breaks, crosslinker-induced chromosomal breakage, and Fanconi anemia core complex function in Fanconi anemia complementation group C-deficient human and murine macrophages. Although both M1 and M2 polarized Fanconi anemia cells were predictably hypersensitive to mitomycin C, only M1 macrophages overproduced TNF-α in response to TLR-activating signals. DNA damaging agents alone did not induce TNF-α production in the absence of TLR agonists in wild-type or Fanconi anemia macrophages, and mitomycin C did not enhance TLR responses in either normal or Fanconi anemia cells. TLR4 and TLR7/8 activation induced cytokine overproduction in Fanconi anemia macrophages. Also, although TLR4 activation was associated with induced double strand breaks, TLR7/8 activation was not. That DNA strand breaks and chromosome breaks are neither necessary nor sufficient to account for the overproduction of inflammatory cytokines by Fanconi anemia cells suggests that noncanonical anti-inflammatory functions of Fanconi anemia complementation group C contribute to the aberrant macrophage phenotype and suggests that suppression of macrophage/TLR hyperreactivity might prevent cytokine-induced stem cell attrition in Fanconi anemia. © Society for Leukocyte Biology.

UPregulated single-stranded DNA-binding protein 1 induces cell chemoresistance to cisplatin in lung cancer cell lines.

Science.gov (United States)

Zhao, Xiang; He, Rong; Liu, Yu; Wu, Yongkai; Kang, Leitao

2017-07-01

Cisplatin and its analogues are widely used as anti-tumor drugs in lung cancer but many cisplatin-resistant lung cancer cases have been identified in recent years. Single-stranded DNA-binding protein 1 (SSDBP1) can effectively induce H69 cell resistance to cisplatin in our previous identification; thus, it is necessary to explore the mechanism underlying the effects of SSDBP1-induced resistance to cisplatin. First, SSDBP1-overexpressed or silent cell line was constructed and used to analyze the effects of SSDBP1 on chemoresistance of lung cancer cells to cisplatin. SSDBP1 expression was assayed by real-time PCR and Western blot. Next, the effects of SSDBP1 on cisplatin sensitivity, proliferation, and apoptosis of lung cancer cell lines were assayed by MTT and flow cytometry, respectively; ABC transporters, apoptosis-related genes, and cell cycle-related genes by real-time PCR, and DNA wound repair by comet assay. Low expression of SSDBP1 was observed in H69 cells, while increased expression in cisplatin-resistant H69 cells. Upregulated expression of SSDBP1 in H69AR cells was identified to promote proliferation and cisplatin resistance and inhibit apoptosis, while downregulation of SSDBP1 to inhibit cisplatin resistance and proliferation and promoted apoptosis. Moreover, SSDBP1 promoted the expression of P2gp, MRP1, Cyclin D1, and CDK4 and inhibited the expression of caspase 3 and caspase 9. Furthermore, SSDBP1 promoted the DNA wound repair. These results indicated that SSDBP1 may induce cell chemoresistance of cisplatin through promoting DNA repair, resistance-related gene expression, cell proliferation, and inhibiting apoptosis.

Wnt11b is involved in cilia-mediated symmetry breakage during Xenopus left-right development.

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Peter Walentek

Full Text Available Breakage of bilateral symmetry in amphibian embryos depends on the development of a ciliated epithelium at the gastrocoel roof during early neurulation. Motile cilia at the gastrocoel roof plate (GRP give rise to leftward flow of extracellular fluids. Flow is required for asymmetric gene expression and organ morphogenesis. Wnt signaling has previously been involved in two steps, Wnt/ß-catenin mediated induction of Foxj1, a regulator of motile cilia, and Wnt/planar cell polarity (PCP dependent cilia polarization to the posterior pole of cells. We have studied Wnt11b in the context of laterality determination, as this ligand was reported to activate canonical and non-canonical Wnt signaling. Wnt11b was found to be expressed in the so-called superficial mesoderm (SM, from which the GRP derives. Surprisingly, Foxj1 was only marginally affected in loss-of-function experiments, indicating that another ligand acts in this early step of laterality specification. Wnt11b was required, however, for polarization of GRP cilia and GRP morphogenesis, in line with the known function of Wnt/PCP in cilia-driven leftward flow. In addition Xnr1 and Coco expression in the lateral-most GRP cells, which sense flow and generate the first asymmetric signal, was attenuated in morphants, involving Wnt signaling in yet another process related to symmetry breakage in Xenopus.

The importance of becoming double-stranded: Innate immunity and the kinetic model of HIV-1 central plus strand synthesis

International Nuclear Information System (INIS)

Poeschla, Eric

2013-01-01

Central initiation of plus strand synthesis is a conserved feature of lentiviruses and certain other retroelements. This complication of the standard reverse transcription mechanism produces a transient “central DNA flap” in the viral cDNA, which has been proposed to mediate its subsequent nuclear import. This model has assumed that the important feature is the flapped DNA structure itself rather than the process that produces it. Recently, an alternative kinetic model was proposed. It posits that central plus strand synthesis functions to accelerate conversion to the double-stranded state, thereby helping HIV-1 to evade single-strand DNA-targeting antiviral restrictions such as APOBEC3 proteins, and perhaps to avoid innate immune sensor mechanisms. The model is consistent with evidence that lentiviruses must often synthesize their cDNAs when dNTP concentrations are limiting and with data linking reverse transcription and uncoating. There may be additional kinetic advantages for the artificial genomes of lentiviral gene therapy vectors. - Highlights: • Two main functional models for HIV central plus strand synthesis have been proposed. • In one, a transient central DNA flap in the viral cDNA mediates HIV-1 nuclear import. • In the other, multiple kinetic consequences are emphasized. • One is defense against APOBEC3G, which deaminates single-stranded DNA. • Future questions pertain to antiviral restriction, uncoating and nuclear import

Alaska Beluga Strandings

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National Oceanic and Atmospheric Administration, Department of Commerce — NMFS is mandated by the Marine Mammal Protection Act (MMPA) to collect reports of marine mammal stranding events. This particular catalogue item is specific to the...

A comparative investigation of DNA strand breaks, sister chromatid exchanges and K-ras gene mutations induced by cadmium salts in cultured human cells

International Nuclear Information System (INIS)

Mouron, Silvana Andrea; Grillo, Claudia Alejandra; Dulout, Fernando Noel; Golijow, Carlos Daniel

2004-01-01

Cadmium (Cd) is a toxic heavy metal of continuing occupational and environmental concern with a wide variety of adverse effects. Several studies have shown that cadmium produces DNA strand breaks, DNA-protein cross-links, oxidative DNA damage, chromosomal aberrations, dysregulation of gene expression resulting in enhanced proliferation, depressed apoptosis and/or altered DNA repair. This study was undertaken to investigate the ability of cadmium chloride (CdCl 2 ) and cadmium sulphate (CdSO 4 ) to induce point mutations in codon 12 of the K-ras protooncogene assessed by polymerase chain reaction-single strand conformation polymorphisms (PCR-SSCP) and RFLP-enriched PCR methods. Also their genotoxic effects were analyzed by the comet assay and sister chromatid exchanges test. The human lung fibroblast cell line MRC-5 was used for the experiments. Sister chromatid exchanges assay (SCEs) frequencies were significantly increased in cells exposed to cadmium salts in relation to controls (p < 0.001). Despite the slow increment observed in the three comet parameters considered when cells were treated with cadmium chloride, significant differences between groups were only found in the variable comet moment (CM) (p < 0.005). On the other hand, when cells were exposed to cadmium sulphate, the Kruskal-Wallis test showed highly significant differences between groups for migration, tail moment and comet moment parameters (p < 0.001). Nevertheless, a null or weak point mutation induction in K-ras protooncogene was detected using polymerase chain reaction-low ionic strength-single strand conformation polymorphisms (PCR-LIS-SSCP) and RFLP-enriched PCR methods when cells were treated with cadmium salts. Thus, inorganic cadmium produces genotoxicity in human lung fibroblast MRC-5 cells, in the absence of significant point mutation of the K-ras gene

Assessment of DNA double-strand breaks induced by intravascular iodinated contrast media following in vitro irradiation and in vivo, during paediatric cardiac catheterization.

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Gould, Richard; McFadden, Sonyia L; Horn, Simon; Prise, Kevin M; Doyle, Philip; Hughes, Ciara M

2016-01-01

Paediatric cardiac catheterizations may result in the administration of substantial amounts of iodinated contrast media and ionizing radiation. The aim of this work was to investigate the effect of iodinated contrast media in combination with in vitro and in vivo X-ray radiation on lymphocyte DNA. Six concentrations of iodine (15, 17.5, 30, 35, 45, and 52.5 mg of iodine per mL blood) represented volumes of iodinated contrast media used in the clinical setting. Blood obtained from healthy volunteers was mixed with iodinated contrast media and exposed to radiation doses commonly used in paediatric cardiac catheterizations (0 mGy, 70 mGy, 140 mGy, 250 mGy and 450 mGy). Control samples contained no iodine. For in vivo experimentation, pre and post blood samples were collected from children undergoing cardiac catheterization, receiving iodine concentrations of up to 51 mg of iodine per mL blood and radiation doses of up to 400 mGy. Fluorescence microscopy was performed to assess γH2AX-foci induction, which corresponded to the number of DNA double-strand breaks. The presence of iodine in vitro resulted in significant increases of DNA double-strand breaks beyond that induced by radiation for ≥ 17.5 mg/mL iodine to blood. The in vivo effects of contrast media on children undergoing cardiac catheterization resulted in a 19% increase in DNA double-strand breaks in children receiving an average concentration of 19 mg/mL iodine to blood. A larger investigation is required to provide further information of the potential benefit of lowering the amount of iodinated contrast media received during X-ray radiation investigations. Copyright © 2015 John Wiley & Sons, Ltd.

DN2 Thymocytes Activate a Specific Robust DNA Damage Response to Ionizing Radiation-Induced DNA Double-Strand Breaks

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Irene Calvo-Asensio

2018-06-01

Full Text Available For successful bone marrow transplantation (BMT, a preconditioning regime involving chemo and radiotherapy is used that results in DNA damage to both hematopoietic and stromal elements. Following radiation exposure, it is well recognized that a single wave of host-derived thymocytes reconstitutes the irradiated thymus, with donor-derived thymocytes appearing about 7 days post BMT. Our previous studies have demonstrated that, in the presence of donor hematopoietic cells lacking T lineage potential, these host-derived thymocytes are able to generate a polyclonal cohort of functionally mature peripheral T cells numerically comprising ~25% of the peripheral T cell pool of euthymic mice. Importantly, we demonstrated that radioresistant CD44+ CD25+ CD117+ DN2 progenitors were responsible for this thymic auto-reconstitution. Until recently, the mechanisms underlying the radioresistance of DN2 progenitors were unknown. Herein, we have used the in vitro “Plastic Thymus” culture system to perform a detailed investigation of the mechanisms responsible for the high radioresistance of DN2 cells compared with radiosensitive hematopoietic stem cells. Our results indicate that several aspects of DN2 biology, such as (i rapid DNA damage response (DDR activation in response to ionizing radiation-induced DNA damage, (ii efficient repair of DNA double-strand breaks, and (iii induction of a protective G1/S checkpoint contribute to promoting DN2 cell survival post-irradiation. We have previously shown that hypoxia increases the radioresistance of bone marrow stromal cells in vitro, at least in part by enhancing their DNA double-strand break (DNA DSB repair capacity. Since the thymus is also a hypoxic environment, we investigated the potential effects of hypoxia on the DDR of DN2 thymocytes. Finally, we demonstrate for the first time that de novo DN2 thymocytes are able to rapidly repair DNA DSBs following thymic irradiation in vivo.

Study of transport and micro-structural properties of magnesium di-boride strand under react and bend mode and bend and react mode

International Nuclear Information System (INIS)

Kundu, Ananya; Das, Subrat Kumar; Bano, Anees; Pradhan, Subrata

2015-01-01

I-V characterization of commercial multi-filamentary Magnesium Di-Boride (MgB 2 ) wire of diameter 0.83 mm were studied in cryocooler based self-field characterization system under both react and bent mode and bent and react mode for a range of temperature 6 K - 25 K. This study is of practical technical relevance where the heat treatment of the superconducting wire makes the sample less flexible for winding in magnet and in other applications. There are limited reported data, available on degradation of MgB 2 wire with bending induced strain in react and wind and wind and react method. In the present work the bending diameter were varied from 80 mm to 20 mm in the interval of 10 mm change of bending diameter and for each case critical current (Ic) of the strand is measured for the above range of temperature. An ETP copper made customized sample holder for mounting the MgB 2 strand was fabricated and is thermally anchored to the cooling stage of the cryocooler. It is seen from the experimental data that in react and bent mode the critical current degrades from 105 A to 87 A corresponding to bending diameter of 80 mm and 20 mm respectively. The corresponding bending strain was analytically estimated and compared with the simulation result. It is also observed that in react and bent mode, the degradation of the transport property of the strand is less as compared to react and bent mode. For bent and react mode in the same sample, the critical current (Ic) was measured to be ∼145 A at 15 K for bending diameter of 20 mm. Apart from studying the bending induced strain on MgB 2 strand, the tensile test of the strand at RT was carried out. The electrical characterizations of the samples were accompanied by the microstructure analyses of the bent strand to examine the bending induced degradation in the grain structure of the strand. All these experimental findings are expected to be used as input to fabricate prototype MgB 2 based magnet. (author)

Low-residue euthanasia of stranded mysticetes.

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Harms, Craig A; McLellan, William A; Moore, Michael J; Barco, Susan G; Clarke, Elsburgh O; Thayer, Victoria G; Rowles, Teresa K

2014-01-01

Euthanasia of stranded large whales poses logistic, safety, pharmaceutical, delivery, public relations, and disposal challenges. Reasonable arguments may be made for allowing a stranded whale to expire naturally. However, slow cardiovascular collapse from gravitational effects outside of neutral buoyancy, often combined with severely debilitating conditions, motivate humane efforts to end the animal's suffering. The size of the animal and prevailing environmental conditions often pose safety concerns for stranding personnel, which take priority over other considerations. When considering chemical euthanasia, the size of the animal also necessitates large quantities of euthanasia agents. Drug residues are a concern for relay toxicity to scavengers, particularly for pentobarbital-containing euthanasia solutions. Pentobarbital is also an environmental concern because of its stability and long persistence in aquatic environments. We describe a euthanasia technique for stranded mysticetes using readily available, relatively inexpensive, preanesthetic and anesthetic drugs (midazolam, acepromazine, xylazine) followed by saturated KCl delivered via custom-made needles and a low-cost, basic, pressurized canister. This method provides effective euthanasia while moderating personnel exposure to hazardous situations and minimizing drug residues of concern for relay toxicity.

DNA double-strand breaks induced by cavitational mechanical effects of ultrasound in cancer cell lines.

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Yukihiro Furusawa

Full Text Available Ultrasonic technologies pervade the medical field: as a long established imaging modality in clinical diagnostics; and, with the emergence of targeted high intensity focused ultrasound, as a means of thermally ablating tumours. In parallel, the potential of [non-thermal] intermediate intensity ultrasound as a minimally invasive therapy is also being rigorously assessed. Here, induction of apoptosis in cancer cells has been observed, although definitive identification of the underlying mechanism has thus far remained elusive. A likely candidate process has been suggested to involve sonochemical activity, where reactive oxygen species (ROS mediate the generation of DNA single-strand breaks. Here however, we provide compelling new evidence that strongly supports a purely mechanical mechanism. Moreover, by a combination of specific assays (neutral comet tail and staining for γH2AX foci formation we demonstrate for the first time that US exposure at even moderate intensities exhibits genotoxic potential, through its facility to generate DNA damage across multiple cancer lines. Notably, colocalization assays highlight that ionizing radiation and ultrasound have distinctly different signatures to their respective γH2AX foci formation patterns, likely reflecting the different stress distributions that initiated damage formation. Furthermore, parallel immuno-blotting suggests that DNA-PKcs have a preferential role in the repair of ultrasound-induced damage.

Alkali-labile sites and post-irradiation effects in single-stranded DNA induced by H radicals

International Nuclear Information System (INIS)

Lafleur, M.V.M.; Heuvel, N. van; Woldhuis, J.; Loman, H.

1978-01-01

Single-stranded phiX174 DNA in aqueous solutions has been irradiated in the absence of oxygen, under conditions in which H radicals react with the DNA. It was shown that H radical reactions result in breaks, which contribute approximately 10 per cent inactivation. Further, two types of alkali-labile sites were formed. One was lethal and gave rise to single-strand breaks by alkali and was most probably identical with post-irradiation heat damage and contributed about 33 per cent to the inactivation mentioned above. The other consisted of non-lethal damage, partly dihydropyrimidine derivatives, and was converted to lethal damage by alkali. This followed from experiments in which the DNA was treated with osmium-tetroxide, which oxidized thymine to 5,6-dihydroxydihydrothymine. Treatment with alkali of this DNA gave the same temperature dependence as found for the non-lethal alkali-labile sites in irradiated DNA. A similar temperature dependence was found for dihydrothymine and irradiated pyrimidines with alkali. (author)

Stranded Specimen Data

Data.gov (United States)

National Oceanic and Atmospheric Administration, Department of Commerce — The Marine Mammal and Turtle Division collects data about individual cetaceans and sea turtles that come ashore, or strand on the beach. The date and location of...

scid mutation in mice confers hypersensitivity to ionizing radiation and a deficiency in DNA double-strand break repair

International Nuclear Information System (INIS)

Biedermann, K.A.; Sun, J.R.; Giaccia, A.J.; Tosto, L.M.; Brown, J.M.

1991-01-01

C.B-17 severe combined immunodeficient (scid) mice carry the scid mutation and are severely deficient in both T cell- and B cell-mediated immunity, apparently as a result of defective V(D)J joining of the immunoglobulin and T-cell receptor gene elements. In the present studies, we have defined the tissue, cellular, and molecular basis of another characteristic of these mice: their hypersensitivity to ionizing radiation. Bone marrow stem cells, intestinal crypt cells, and epithelial skin cells from scid mice are 2- to 3-fold more sensitive when irradiated in situ than are congenic BALB/c or C.B-17 controls. Two independently isolated embryo fibroblastic scid mouse cell lines display similar hypersensitivities to gamma-rays. In addition, these cell lines are sensitive to cell killing by bleomycin, which also produces DNA strand breaks, but not by the DNA crosslinking agent mitomycin C or UV irradiation. Measurement of the rejoining of gamma-ray-induced DNA double-strand breaks by pulsed-field gel electrophoresis indicates that these animals are defective in this repair system. This suggests that the gamma-ray sensitivity of the scid mouse fibroblasts could be the result of reduced repair of DNA double-strand breaks. Therefore, a common factor may participate in both the repair of DNA double-strand breaks as well as V(D)J rejoining during lymphocyte development. This murine autosomal recessive mutation should prove extremely useful in fundamental studies of radiation-induced DNA damage and repair

Further delineation of the Nijmegen breakage syndrome

International Nuclear Information System (INIS)

Taalman, R.D.; Hustinx, T.W.; Weemaes, C.M.; Seemanova, E.; Schmidt, A.; Passarge, E.; Scheres, J.M.

1989-01-01

We report on five independent families with a chromosome instability disorder that earlier had been called the Nijmegen breakage syndrome (NBS). These families, two from the Netherlands and three from Czechoslovakia, had a total of eight patients, five of whom are still alive. The main clinical manifestations were microcephaly, short stature, a ''bird-like'' face, immunological defects involving both the humoral and cellular system. In four of the five living patients it has been possible to study the chromosomes of cultured lymphocytes. The basic karyotype in these patients were normal, but in 17% to 35% of the metaphases rearrangements were found, preferentially involving chromosomes 7 and/or 14 at the sites 7p13, 7q34, and 14q11. The chromosomes of all five living patients were very sensitive to ionizing radiation. In addition, the DNA synthesis in their cultured lymphocytes and fibroblasts was more resistant to X-rays than in cells from controls. The NBS shares a number of important features with ataxia telangiectasia (AT). Both syndromes are characterized by the occurrence of typical rearrangements of chromosomes 7 and/or 14, cellular and chromosomal hypersensitivity to X-irradiation, radioresistance of DNA replication and immunodeficiency. However, there are also obvious differences: NBS patients have microcephaly but neither ataxia nor telangiectasia, and in contrast to the situation in AT the alpha-fetoprotein level in their serum is normal

Binding of radiation-induced phenylalanine radicals to DNA

International Nuclear Information System (INIS)

Schans, G.P. van der; Rijn, C.J.S. van; Bleichrodt, J.F.

1975-11-01

When an aqueous solution of double-stranded DNA of bacteriophage PM2 containing phenylalanine and saturated with N 2 O is irradiated with γ-rays, radiation-induced phenylalanine radicals are bound covalently. Under the conditions used about 25 phenylalanine molecules may be bound per lethal hit. Also for single-stranded PM2 DNA, most of the phenylalanine radicals bound are non-lethal. Evidence is presented that in double-stranded DNA an appreciable fraction of the single-strand breaks is induced by phenylalanine radicals. Radiation products of phenylalanine and the phenylalanine bound to the DNA decrease the sensitivity of the DNA to the induction of single-strand breaks. There are indications that the high efficiency of protection by radiation products of phenylalanine is due to their positive charge, which will result in a relatively high concentration of these compounds in the vicinity of the negatively charged DNA molecules

Purification of Single-Stranded cDNA Based on RNA Degradation Treatment and Adsorption Chromatography.

Science.gov (United States)

Trujillo-Esquivel, Elías; Franco, Bernardo; Flores-Martínez, Alberto; Ponce-Noyola, Patricia; Mora-Montes, Héctor M

2016-08-02

Analysis of gene expression is a common research tool to study networks controlling gene expression, the role of genes with unknown function, and environmentally induced responses of organisms. Most of the analytical tools used to analyze gene expression rely on accurate cDNA synthesis and quantification to obtain reproducible and quantifiable results. Thus far, most commercial kits for isolation and purification of cDNA target double-stranded molecules, which do not accurately represent the abundance of transcripts. In the present report, we provide a simple and fast method to purify single-stranded cDNA, exhibiting high purity and yield. This method is based on the treatment with RNase H and RNase A after cDNA synthesis, followed by separation in silica spin-columns and ethanol precipitation. In addition, our method avoids the use of DNase I to eliminate genomic DNA from RNA preparations, which improves cDNA yield. As a case report, our method proved to be useful in the purification of single-stranded cDNA from the pathogenic fungus Sporothrix schenckii.

A model treating the DNA double-strand break repair inhibition by damage clustering

International Nuclear Information System (INIS)

Rosemann, M.; Abel, H.; Regel, K.

1992-01-01

A microdosimetric model for the interpretation of radiation induced irreparable DNA double-strand breaks was applied to the biological endpoint of chromosomal aberrations. The model explains irreparable DNA double-strand breaks in terms of break clustering in DNA subunits. The model predicts quite good chromosomal aberrations in gamma- and X-ray irradiated V79 cells and human lymphocytes. In the case of α-particle irradiation the presumption had to be made, that only the cells with indirect events in the nucleus (due to delta-electrons) reach the metaphase and are analysed. With the help of this model we are able to explain the peculiar effectiveness of ultrasoft C-X-rays in human lymphocytes. In addition, an interpretation of experiments with accelerated and spatially correlated particles is given. (author)

The use of caspase inhibitors in pulsed-field gel electrophoresis may improve the estimation of radiation-induced DNA repair and apoptosis

International Nuclear Information System (INIS)

Balart, Josep; Pueyo, Gemma; Llobet, Lara I de; Baro, Marta; Sole, Xavi; Marin, Susanna; Casanovas, Oriol; Mesia, Ricard; Capella, Gabriel

2011-01-01

Radiation-induced DNA double-strand break (DSB) repair can be tested by using pulsed-field gel electrophoresis (PFGE) in agarose-encapsulated cells. However, previous studies have reported that this assay is impaired by the spontaneous DNA breakage in this medium. We investigated the mechanisms of this fragmentation with the principal aim of eliminating it in order to improve the estimation of radiation-induced DNA repair. Samples from cancer cell cultures or xenografted tumours were encapsulated in agarose plugs. The cell plugs were then irradiated, incubated to allow them to repair, and evaluated by PFGE, caspase-3, and histone H2AX activation (γH2AX). In addition, apoptosis inhibition was evaluated through chemical caspase inhibitors. We confirmed that spontaneous DNA fragmentation was associated with the process of encapsulation, regardless of whether cells were irradiated or not. This DNA fragmentation was also correlated to apoptosis activation in a fraction of the cells encapsulated in agarose, while non-apoptotic cell fraction could rejoin DNA fragments as was measured by γH2AX decrease and PFGE data. We were able to eliminate interference of apoptosis by applying specific caspase inhibitors, and improve the estimation of DNA repair, and apoptosis itself. The estimation of radiation-induced DNA repair by PFGE may be improved by the use of apoptosis inhibitors. The ability to simultaneously determine DNA repair and apoptosis, which are involved in cell fate, provides new insights for using the PFGE methodology as functional assay

Reconstitution of DNA strand exchange mediated by Rhp51 recombinase and two mediators.

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Yumiko Kurokawa

2008-04-01

Full Text Available In the fission yeast Schizosaccharomyces pombe, genetic evidence suggests that two mediators, Rad22 (the S. pombe Rad52 homolog and the Swi5-Sfr1 complex, participate in a common pathway of Rhp51 (the S. pombe Rad51 homolog-mediated homologous recombination (HR and HR repair. Here, we have demonstrated an in vitro reconstitution of the central step of DNA strand exchange during HR. Our system consists entirely of homogeneously purified proteins, including Rhp51, the two mediators, and replication protein A (RPA, which reflects genetic requirements in vivo. Using this system, we present the first robust biochemical evidence that concerted action of the two mediators directs the loading of Rhp51 onto single-stranded DNA (ssDNA precoated with RPA. Dissection of the reaction reveals that Rad22 overcomes the inhibitory effect of RPA on Rhp51-Swi5-Sfr1-mediated strand exchange. In addition, Rad22 negates the requirement for a strict order of protein addition to the in vitro system. However, despite the presence of Rad22, Swi5-Sfr1 is still essential for strand exchange. Importantly, Rhp51, but neither Rad22 nor the Swi5-Sfr1 mediator, is the factor that displaces RPA from ssDNA. Swi5-Sfr1 stabilizes Rhp51-ssDNA filaments in an ATP-dependent manner, and this stabilization is correlated with activation of Rhp51 for the strand exchange reaction. Rad22 alone cannot activate the Rhp51 presynaptic filament. AMP-PNP, a nonhydrolyzable ATP analog, induces a similar stabilization of Rhp51, but this stabilization is independent of Swi5-Sfr1. However, hydrolysis of ATP is required for processive strand transfer, which results in the formation of a long heteroduplex. Our in vitro reconstitution system has revealed that the two mediators have indispensable, but distinct, roles for mediating Rhp51 loading onto RPA-precoated ssDNA.

Current redistribution in cables made of insulated, soldered, or oxidized strands

International Nuclear Information System (INIS)

Turck, B.

1979-07-01

Current redistributions are compared in cables made of insulated strands, soldered, or oxidized strands and insulated strands with periodic joints. After discussing the different current redistributions in the cases of a rapidly changing current and a dc current, several particular situations are investigated: what happens if a strand is broken, or if a local normal zone appears that does not affect all the strands equally, the detection of this normal zone, and the influence of short circuits between strands

Concrete Cracking Prediction Including the Filling Proportion of Strand Corrosion Products

Science.gov (United States)

Wang, Lei; Dai, Lizhao; Zhang, Xuhui; Zhang, Jianren

2016-01-01

The filling of strand corrosion products during concrete crack propagation is investigated experimentally in the present paper. The effects of stirrups on the filling of corrosion products and concrete cracking are clarified. A prediction model of crack width is developed incorporating the filling proportion of corrosion products and the twisting shape of the strand. Experimental data on cracking angle, crack width, and corrosion loss obtained from accelerated corrosion tests of concrete beams are presented. The proposed model is verified by experimental data. Results show that the filling extent of corrosion products varies with crack propagation. The rust filling extent increases with the propagating crack until a critical width. Beyond the critical width, the rust-filling extent remains stable. Using stirrups can decrease the critical crack width. Stirrups can restrict crack propagation and reduce the rust filling. The tangent of the cracking angle increases with increasing corrosion loss. The prediction of corrosion-induced crack is sensitive to the rust-filling extent. PMID:28772367

Metallurgical investigation of wire breakage of tyre bead grade

Directory of Open Access Journals (Sweden)

Piyas Palit

2015-10-01

Full Text Available Tyre bead grade wire is used for tyre making application. The wire is used as reinforcement inside the polymer of tyre. The wire is available in different size/section such as 1.6–0.80 mm thin Cu coated wire. During tyre making operation at tyre manufacturer company, wire failed frequently. In this present study, different broken/defective wire samples were collected from wire mill for detailed investigation of the defect. The natures of the defects were localized and similar in nature. The fracture surface was of finger nail type. Crow feet like defects including button like surface abnormalities were also observed on the broken wire samples. The defect was studied at different directions under microscope. Different advanced metallographic techniques have been used for detail investigation. The analysis revealed that, white layer of surface martensite was formed and it caused the final breakage of wire. In this present study we have also discussed about the possible reason for the formation of such kind of surface martensite (hard-phase.

Nbn and atm cooperate in a tissue and developmental stage-specific manner to prevent double strand breaks and apoptosis in developing brain and eye.

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Paulo M G Rodrigues

Full Text Available Nibrin (NBN or NBS1 and ATM are key factors for DNA Double Strand Break (DSB signaling and repair. Mutations in NBN or ATM result in Nijmegen Breakage Syndrome and Ataxia telangiectasia. These syndromes share common features such as radiosensitivity, neurological developmental defects and cancer predisposition. However, the functional synergy of Nbn and Atm in different tissues and developmental stages is not yet understood. Here, we show in vivo consequences of conditional inactivation of both genes in neural stem/progenitor cells using Nestin-Cre mice. Genetic inactivation of Atm in the central nervous system of Nbn-deficient mice led to reduced life span and increased DSBs, resulting in increased apoptosis during neural development. Surprisingly, the increase of DSBs and apoptosis was found only in few tissues including cerebellum, ganglionic eminences and lens. In sharp contrast, we showed that apoptosis associated with Nbn deletion was prevented by simultaneous inactivation of Atm in developing retina. Therefore, we propose that Nbn and Atm collaborate to prevent DSB accumulation and apoptosis during development in a tissue- and developmental stage-specific manner.

JS-K, a GST-activated nitric oxide generator, induces DNA double-strand breaks, activates DNA damage response pathways, and induces apoptosis in vitro and in vivo in human multiple myeloma cells.

Science.gov (United States)

Kiziltepe, Tanyel; Hideshima, Teru; Ishitsuka, Kenji; Ocio, Enrique M; Raje, Noopur; Catley, Laurence; Li, Chun-Qi; Trudel, Laura J; Yasui, Hiroshi; Vallet, Sonia; Kutok, Jeffery L; Chauhan, Dharminder; Mitsiades, Constantine S; Saavedra, Joseph E; Wogan, Gerald N; Keefer, Larry K; Shami, Paul J; Anderson, Kenneth C

2007-07-15