Toxicological effects of ultraviolet radiation on lymphocyte cells with different manganese superoxide dismutase Ala16Val polymorphism genotypes (original) (raw)
Related papers
DNA damage and oxidative stress in human disease
BioMed research international, 2013
The impact of DNA damage in human diseases is gaining attention since the mid-1990s. DNA damage and oxidative stress are known factors for the origin and progression of cancer. DNA damage and oxidative stress have also been implicated in so diverse diseases such as brain injury, pulmonary diseases, and other chronic inflammation-related disorders.
Mutation Research/DNA Repair, 1998
The extent of the indirect DNA damage generated in mammalian cells by visible light because of the presence of Ž endogenous photosensitizers was studied by means of repair endonucleases. In immortalized human keratinocytes HaCaT. cells exposed to low doses of natural sunlight, the yield of oxidative DNA base modifications sensitive to the repair Ž. endonuclease formamidopyrimidine-DNA glycosylase Fpg protein generated by this indirect mechanism was 10% of that Ž. of pyrimidine dimers generated by direct DNA excitation. A similar yield of Fpg-sensitive modifications, which include 8-hydroxyguanine, was observed in primary keratinocytes. The relative yield of oxidative base modifications decreased at higher light doses, probably as a result of photodecomposition of the endogenous chromophore involved. For the three cell lines tested, viz. HaCaT cells, L1210 mouse leukemia cells and AS52 Chinese hamster cells, the yield of oxidative base modifications generated by a low dose of visible light appeared to be correlated with the basal concentrations of porphyrins in the cells. Induction of cellular porphyrin synthesis by pretreatment with 5-aminolaevulinic acid increased the light-induced oxidative damage in L1210 cells several-fold. In both induced and uninduced cells, the damage was inhibited by more than Ž. 50% in the presence of ascorbic acid 100 mM , while a-tocopherol and the iron chelator o-phenanthroline had no effect and b-carotene even increased the damage. Even high doses of visible light did not significantly increase the numbers of micronuclei in L1210 cells or of gpt mutations in AS52 cells. The negative outcome can be fully explained by the photobleaching of the endogenous photosensitizers, which prevents the generation of sufficiently high levels of oxidative DNA damage. Therefore, the mutagenic risk arising from the indirectly generated oxidative DNA modifications induced by sunlight may be underestimated when results obtained at high doses are extrapolated to low doses or low dose rates.
Persistent oxidative stress after ionizing radiation is involved in inherited radiosensitivity
Free Radical Biology and Medicine, 2003
The SW620IR1 cell line was derived from SW620 human colon cells surviving to ionizing radiations. It shows an increased radiosensitivity and a higher yield of spontaneous chromosomal aberrations. In order to check whether altered reactive oxygen intermediates (ROI) metabolism is involved in this inherited phenotype, we compared the two cell lines for their radiation-induced modifications at the level of ROI production, antioxidant activities, and chromosomal aberrations. Compared to SW620, SW620IR1 cells exhibit a higher and more persistent ROI induction after various doses of ionizing radiations and a higher yield of dicentric chromosomes. They are also characterized by lower basal activities of glutathione peroxidase and manganese-containing superoxide dismutase, and lower ability to induce these antioxidant defenses after irradiation. Resumption of cell growth after irradiation coincides with maximal induction of antioxidant activities and normalization of ROI concentration. However, at that time radiation-induced chromosomal aberrations are not completely eliminated, leading to the proliferation of genetically unstable cells. These results indicate that the inherited sensitivity of SW620IR1 cells is associated with altered antioxidant activities resulting in higher and more prolonged oxidative stress after radiation exposure. They also suggest that the normalization of ROI levels allows these p53 mutant cells to resume proliferation although high levels of DNA damages are still persisting, thereby explaining the chromosomal instability observed as a delayed effect of radiation exposure.
Genet Mol Res, 2013
Environmental contamination by methylmercury (MeHg) is an enormous public health problem in world regions such as Amazonia. MeHg toxic effects seem to be influenced by environmental and genetic factors. However, few studies have evaluated the genetic influences of MeHg toxicity in humans. Therefore, the aim of this study was to evaluate the genetic influence of Ala16Val manganese superoxide dismutase gene polymorphism (Ala16Val-MnSOD) on the cytotoxic effects of in vitro human leukocytes exposed to MeHg. Subjects were selected from 100 individuals aged 26.4 ± 7.3 years genotyped to Ala16Val-MnSOD ©FUNPEC-RP www.funpecrp.com.br Genetics and Molecular Research 12 (4): 5134-5144 Effects of Ala16Val MnSOD polymorphism on MeHg-exposed WBCs polymorphism (AA = 6, VV = 6, and AV = 12) to perform in vitro testing using white blood cells (WBCs). Reactive oxygen species production was measured using 2',7'-dichlorofluorescein diacetate fluorimetric assay, and cell viability was measured using MTT assay on WBC samples from the same subjects that were both exposed and not exposed to MeHg (2.5 µM for 6 h). The results showed that AA-and VV-WBCs exposed to MeHg did not display increased reactive oxygen species levels compared to those in cells that were not exposed. However, AVleukocytes exposed to MeHg displayed increased ROS levels. Cellular viability comparison among genotypes exposed to MeHg showed that the viability of AA-WBCs was lower than that of VV-WBC, with mean values of 3.46 ± 0.13 and 3.08 ± 0.77 (standard error), respectively (P = 0.033), whereas heterozygous cells (AV) displayed intermediate values.
The potency of UVA radiation, representing 90% of solar UV light reaching the earth's surface, to induce human skin cancer is the subject of continuing controversy. This study was undertaken to investigate the role of reactive oxygen species in DNA damage produced by the exposure of human cells to UVA radiation. This knowledge is important for better understanding of UV-induced carcinogenesis. We measured DNA single-strand breaks and alkali-labile sites in human lymphocytes exposed ex vivo to various doses of 365-nm UV photons compared to X-rays and hydrogen peroxide using the comet assay. We demonstrated that the UVA-induced DNA damage increased in a linear dose-dependent manner. The rate of DNA single-strand breaks and alkali-labile sites after exposure to 1 J/cm 2 was similar to the rate induced by exposure to 1 Gy of X-rays or 25 μM hydrogen peroxide. The presence of either the hydroxyl radical scavenger dimethyl sulfoxide or the singlet oxygen quencher sodium azide resulted in a significant reduction in the UVA-induced DNA damage, suggesting a role for these reactive oxygen species in mediating UVA-induced DNA single-strand breaks and alkali-labile sites. We also showed that chromatin relaxation due to hypertonic conditions resulted in increased damage in both untreated and UVA-treated cells. The effect was the most significant in the presence of 0.5 M Na þ , implying a role for histone H1. Our data suggest that the majority of DNA single-strand breaks and alkali-labile sites after exposure of human lymphocytes to UVA are produced by reactive oxygen species (the hydroxyl radical and singlet oxygen) and that the state of chromatin may substantially contribute to the outcome of such exposures. Crown
Environmental and Molecular Mutagenesis, 2006
Nonmelanoma skin cancer (NMSC) is the most frequent type of cancer in humans. Exposure to UV radiation is a major risk factor for NMSC, and oxidative DNA damage, caused either by UV radiation itself or by other agents, may be involved in its induction. Increased sensitivity to oxidative damage and an altered DNA repair capacity (DRC) increase the risk of many types of cancer; however, sensitivity to oxidizing agents has not been evaluated for NMSC, and results regarding DRC in NMSC are inconclusive. In the present study, we evaluated DNA damage and repair in leukocytes from 41 NMSC patients and 45 controls. The Comet assay was used to measure basal and H2O2-induced DNA damage, as well as the DRC, while the cytokinesis-block micronucleus assay was used to measure the basal level of chromosome damage. Although basal DNA damage was higher for the controls than for the patients, this finding was mainly due to sampling more controls in the summer, which was associated with longer comet tails. In contrast, H2O2-induced DNA damage was significantly higher in cases than in controls, and this parameter was not influenced by the season of the year. The DRC for the H2O2-induced damage was similar for cases and controls and unrelated to seasonality. Finally, the frequency of binucleated lymphocytes with micronuclei was similar for cases and controls. The results of this study indicate that NMSC patients are distinguished from controls by an increased sensitivity to oxidative DNA damage. Environ. Mol. Mutagen., 2006. © 2006 Wiley-Liss, Inc.
DNA Repair and Human Health, 2011
Introduction 1.1 Environmental endogenous DNA damages Cancer development is a long-term, multistep process with a complex interplay between genes and environment. The magnitude of environmental effects depends on the presence or absence of genetic susceptibility of the subjects to certain cancer types. Molecular epidemiological studies in cancer have proved that besides target cell genetic instability, the presence of triggering environmental exposure is critical in cancer development [Albertini & Hayes 1997, Newby & Howard 2005]. The biomarker responses, exposure character and the route of exposure of different environmental factors (chemicals, physical agents and biological agents) are also important in causing tumors especially in the cases of occupational cancer [Ward 1995]. The EPA Guidelines for carcinogen Risk Assessment [EPA 2005] is based on the mode of action of chemicals, such as interaction with DNA, cytotoxicity, or binding to the receptors modifying signal pathways. There are several natural compounds-so called chemopreventive agents-which are able to modify the genotoxic or mutagenic response (Ames 1983) in different organisms. These vitamins, antioxidants, phytochemicals, micro nutrients are available on the market without knowing their mode of action. Mutagenesis caused by environmental chemicals or physical agents can be prevented by protection of the cell's DNA replication, increasing the repair capacity or delaying cell replication to allow enough time to make a complete repair of damaged cells. Antioxidants are able to protect the cells from oxidative stress, and stimulate the phase I reactions including oxidation, reduction, and hydrolysis of xenobiotics by the monoxigenase detoxicating key enzymes, such as CYP450 [Xu et al. 1996, Poulsen & Loft 1998]. These changes increase the polarity of these molecules and help to conjugate them in phase II to glucuronic acid, acetic acid and sulfuric acid which are the physiological ways to eliminate active metabolites that are genotoxic to the target cells. The best studied crucial early event in carcinogenesis is chromosomal aberration, including microsatellite instability, abnormal number of chromosomes (aneuploidy), gene amplification or the loss of heterozygosity of tumor suppressor genes. By reducing chromosomal mutation via chemoprevention, the cell may be able to survive the genotoxic effects without any permanent damage, or it is able to go through the physiological pathway of apoptosis, without mutation occurring in the P53 gene [Lowe & Lin 2000]. www.intechopen.com DNA Repair and Human Health 308 1.2 The role of DNA repair in gene-environmental interactions The measurement of UV-induced DNA repair is recommended in the risk assessment of environmental exposure to harmful chemicals (Reg. 440/2008/EC). Data obtained on prokaryota organisms suggest that exposure to chemicals as e.g. free oxygen radicals can interact with UV-induced DNA repair mechanisms (Chandor-Proust et al, 2008). Among the repair mechanisms existing in higher eukaryota, base excision repair (BER) seems to be the main mechanism involved in the removal of lesions produced by alkylation, deamination or oxidation (Rastogi et al, 2010). Orelli et al. (2009) demonstrated recently that nucleotide excision repair (NER) also plays an important role in the development of cisplatin resistance. UV-induced DNA damages can induce the so called three prime exonuclease1 (trex1), as a response to genotoxic stress. Beside thymine dimer production, UV irradiation can also produce reactive oxygen species. Benzo(a)pyrene (BaP) and hydrogen peroxide may, similarly to UV, induce the so-called three prime exonuclease1 (trex1) involved in the repair pathways of UV-induced DNA lesions, and cells deficient in trex1 show reduced recovery from UV and BaP replication inhibition, and increased sensitivity to towards genotoxins compared to the isogenic control (Christmann et al, 2010). These data suggest that both main mechanisms can be involved in the total repair of environmental chemical-induced genotoxic stress. Such mechanisms can probably explain the observed UDS reduction in some of our groups exposed to various chemicals but not UV. A second question is whether decreased UDS can be related to an increase in apoptotic capacity? Cells deficient in the repair of UV-induced DNA damage can be more susceptible to a G1 arrest after UV treatment than cells with normal repair capacity or those cells which have completed their DNA repair prior to movement from G1 to S phase (Geyer et al, 2000). Zampetti-Bosseler and Scott (1981) demonstrated a prolonged mitotic delay in repair deficient ataxia teleangiectasia and retinoblastoma fibroblasts after X-ray irradiation compared to normal human fibroblasts, also suggesting a general key role of cell cycle check points beside DNA repair in preservation of genome stability (Kaufman, 1995). Skin fibroblasts from derived ataxia teleangiectasia patients are also more sensitive to UVinduced mutagenesis than those taken from healthy subjects (Hannan et al, 2002), and their results suggested a relationship between cell cycle control and DNA repair pathways in human cells. Genotoxic chemicals can also delay cellular proliferation in DNA repairdeficient cell clones more significantly than in wild type cells, by interfering with DNA replication, thereby inducing DNA damage (Kyunghee et al, 2009). The recently discovered cell cycle checkpoint activation mechanisms are discussed in detail by Rastogi et al (2010). In the present study the so-called premature centromere division (PCD) was used as a cytogenetic indicator of abnormalities in cell cycle regulation (Méhes 1978, Vig, 1981, Major et al, 1999). PCD yields were increased among cytostatic drug producers, anesthesiologists using halothane, and in exposures to formaldehyde, benzene and PAHs. PCD can be involved in the pathomechanism of aneuploidy, it seems to be a possible manifestation of chromosome instability also in human chromosome breakage syndromes and it can be connected with carcinogenesis (for review, c.f. Major et al, 1999). 2. Cancer development and DNA repair We don't know exactly what the cause of cancer is; therefore we have several mechanisms and theories to explain it. One of them is shown in Fig.1. www.intechopen.com Application of UV-Induced Unscheduled DNA-Synthesis Measurements in Human Genotoxicological Risk Assessment 315 6. Analysis of biomarkers in blood samples Biomarkers for DNA damages and risk assessment Considering the basic mechanism of cancer development, the most acceptable predictors of cancer risks are the DNA-damage biomarkers (see Table 3.). These damages can be provoked by exogenous or end o g e n o u s a g e n t s w h e n D N A r e p a i r o r m i s-r e p a i r i s i n dysfunction. The unrepaired DNA damage can reduce the basic cell functions eg. maintenance of genetic integrity, triggering of cell cycle arrest, apoptosis, uncontrolled growth and other functionalities. Ultimately, damaged repair capacity leads to an increase in somatic mutations and cancer. Any living cells Mitochondrial DNA mutation Any living cells Point mutation (HPRT) Any living cells Nuclear p53 Lymphocytes, germ cells DNA-adducts and oxidation, methylation Any living cells Telomere shortening Tumor cells, lymphocytes, germ cells Aneuploidy Lymphocytes, bone marrow cells Micronucleus assay Tumor cells, lymphocytes, germ cells Chromosomal aberrations Any living cells, germ cells DNA strand breaks (SGE or Comet assay) Lymphocytes, hepatocytes Unscheduled DNA synthesis (UDS) Target cells Methods Any living cells Mitochondrial DNA mutation Any living cells Point mutation (HPRT) Any living cells Nuclear p53 Lymphocytes, germ cells DNA-adducts and oxidation, methylation Any living cells Telomere shortening Tumor cells, lymphocytes, germ cells Aneuploidy Lymphocytes, bone marrow cells Micronucleus assay Tumor cells, lymphocytes, germ cells Chromosomal aberrations Any living cells, germ cells DNA strand breaks (SGE or Comet assay) Lymphocytes, hepatocytes Unscheduled DNA synthesis (UDS)
Induction of oxidative DNA base damage in human skin cells by UV and near visible radiation
Carcinogenesis, 1997
The premutagenic oxidative DNA base damage, 7,8dihydro-8-oxoguanine, is induced in human skin fibroblasts by monochromatic radiation ranging from a UVB wavelength (312 nm) up to wavelengths in the near visible (434 nm). The oxidative damage is not generated by absorption of radiation in DNA but rather by activation of photosensitizers generating genotoxic singlet oxygen species. The spectrum for the yield of the oxidative damage in confluent, non-growing, primary skin fibroblasts shows that it is UVA (above 334 nm) and near visible radiations which cause almost all of this guanine oxidation by natural sunlight in the fibroblast model. We estimate that the total amount of oxidation of guanine induced by sunlight in fibroblasts in the epidermis of the skin equals or exceeds the amount of the major type of direct DNA damage, cyclobutane pyrimidine dimers. In rapidly dividing lymphoblastoid cells, no oxidative guanine damage was induced. However, in melanoma cells almost as much damage as in nongrowing fibroblasts (1.1 per 10 4 guanine bases after 1200 kJ/m 2 UVA) was found. We conclude that oxidative DNA base damage can probably contribute to the induction of both non-melanoma and melanoma skin cancer by sunlight.
Biochemical and Biophysical Research Communications, 1996
When human respiratory tract epithelial cells were exposed to 100 mM H 2 O 2 , there was rapid induction of DNA strand breakage and chemical modifications to all 4 DNA bases suggestive of attack by OH •. The major products were FAPy-adenine, FAPy-guanine, and 8-OH-guanine. Some of the base modifications were removed very quickly from the DNA (e.g., 8-OH-guanine), whereas others persisted for longer (e.g., thymine glycol), probably due to differential activity of different repair enzymes. By contrast, strand breaks continued to increase over the time course of the experiment, perhaps because strand breakage is also implicated in the repair process. One should therefore be cautious in using strand breakage as a sole measure of oxidative DNA damage, and when drawing conclusions about the pattern and biological significance of oxidative DNA damage in cells the relative persistence of different lesions must be considered. ᭧ 1996 Academic Press, Inc. Oxygen derived species such as the superoxide radical (O • 0 2) and hydrogen peroxide (H 2 O 2) are produced in mammalian cells during normal aerobic metabolism [1,2]. Excess generation of these species in vivo results in damage to many biological molecules including lipids, protein and carbohydrates [2-4]. Reactive oxygen species (ROS) are also thought to contribute to the development of cancer by promoting chemical changes in DNA which are potentially mutagenic [5-9]. Both DNA strand breakage [2,10-13] and modification of DNA bases [14-16] are frequently observed in cells subjected to oxidative stress. Such damage may result by a variety of mechanisms including rises in intracellular free Ca 2/ that are sufficient to activate endonucleases [17] or from direct attack on DNA by highly reactive radicals, such as hydroxyl (OH •) [6,7,18,19]. It is well established that H 2 O 2 may react with transition metal ions bound to DNA to form OH • [10,20-22] and produce a pattern of base modification very similar to that produced by ionizing radiation, an established source of OH • [7,21,14,23,24]. Consistent with direct OH • radical attack on DNA, several groups have reported increases in products of base oxidation, particularly in 8-hydroxyguanine (or 8-hydroxydeoxyguanosine) in mammalian cells exposed to oxidative stress [8,11,12,25-27]. Chemical changes in the DNA bases are of considerable importance if repair of these changes does not occur, or if repair is such that the fidelity of the original code is lost [28]. For example, formation of thymine glycol or formamidopyrimidines in the DNA template is known to cause a block in DNA replication [29,30] and 8-hydroxyguanine causes miscoding (GrC r TrA transversion mutation) [31,32]. H 2 O 2-derived radicals have been shown to cause GrC r TrA transversions in the SupF gene of E. coli [33] and UV light can cause tandem double CC r TT mutations in the p53 gene, implicated in the development of squamous cell
Genome, 2001
Different types of mutations and DNA-damage profiles induced by near-UV radiation and the superoxide anion (O 2 -·) indicate separate lesions and (or) mechanisms of mutagenesis. Despite a wealth of data, it is still unclear whether variations in the activity levels of antioxidant enzymes naturally present in suboptimal concentrations are among the underlying causes of the increase of near UV radiation genotoxicity. We incorporated a low-activity allele of copper-zinc superoxide dismutase (CuZnSOD), recovered from natural populations of Drosophila melanogaster, into standard marked strains and employed a somatic mutation and recombination test (SMART) to compare paraquat and near UV radiation genotoxicity in these strains. Our results show that, although the low-activity CuZnSOD allele of D. melanogaster confers hypersensitivity to paraquat, the near UV radiation damage was not affected.