RESUMO
Benzo[a]pyrene (BaP) is known to form DNA adduct following metabolic activation, which causes phosphorylation of histone H2AX (γ-H2AX). Recent studies have shown that histone deacetylase (HDAC) inhibitors enhanced BaP-induced CYP1A1 gene expression. In this study, we examined the relationship between the HDAC inhibitor-augmented metabolic activation and BaP-induced γ-H2AX. Sodium butyrate (SB), a typical HDAC inhibitor, enhanced BaP-induced γ-H2AX. The enhanced DNA damage was further confirmed by biased sinusoidal field gel electrophoresis, which detects DNA double-strand breaks. SB remarkably augmented BaP-induced CYP1A1 gene expression, and CYP1A1-overexpressing cells showed elevated generation of γ-H2AX. Furthermore, SB enhanced intracellular oxidation after treatment with BaP. These results suggested that SB-induced CYP1A1 upregulation facilitated BaP metabolism, which might result in excess DNA adducts or oxidative DNA damages, leading to augmentation of γ-H2AX.
Assuntos
Benzo(a)pireno , Citocromo P-450 CYP1A1 , Benzo(a)pireno/toxicidade , Benzo(a)pireno/metabolismo , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A1/metabolismo , Inibidores de Histona Desacetilases , Adutos de DNA , Ácido ButíricoRESUMO
Ortho (o)-toluidine is a widely known carcinogenic substance associated with cancers of the human bladder. A study on British chemical factory workers exposed to 2-mercaptobenzothiazole, phenyl-ß-naphthylamine, aniline, and o-toluidine demonstrated the crucial roles of o-toluidine, 2-mercaptobenzothiazole, and phenyl-ß-naphthylamine in the development of bladder cancer. As genotoxic events are crucial steps in the initiation of cancer, in the present study, we aimed to examine the genotoxic potential of the four chemicals using phosphorylated histone H2AX (γ-H2AX), which is a sensitive and reliable marker of DNA damage, in cultured human urothelial cells. Of the four chemicals, 2-mercaptobenzothiazole was a particularly potent DNA-damaging agent. Moreover, mechanistic studies revealed that γ-H2AX generation by 2-mercaptobenzothiazole was mainly associated with the generation of reactive oxygen species via cytochrome P450 2E1-mediated metabolism. The findings of this study may provide information that is important for the assessment of risks associated with chemicals as well as the interpretation of epidemiological studies investigating occupational bladder cancer.
Assuntos
Citocromo P-450 CYP2E1 , Histonas/metabolismo , Neoplasias da Bexiga Urinária , 2-Naftilamina , Benzotiazóis , Citocromo P-450 CYP2E1/metabolismo , Humanos , Espécies Reativas de Oxigênio/metabolismo , Neoplasias da Bexiga Urinária/induzido quimicamenteRESUMO
A typical tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is known as a strong carcinogen. We previously reported that metabolized NNK induced histone H2AX phosphorylation (γ-H2AX), a DNA damage-induced histone modification. In this study, we found that NNK globally acetylated histone H3, which affected γ-H2AX generation. Human lung adenocarcinoma A549 was treated with several doses of NNK. NNK induced dose-dependent global histone H3 acetylation (Ac-H3), at 2 to 12 h after the treatment, independent of the cell cycle. The Ac-H3 pattern was not affected by CYP2A13 overexpression unlike γ-H2AX, indicating no requirement of NNK metabolism to induce Ac-H3. Immunofluorescence staining of Ac-H3 was uniform throughout the nucleus, whereas γ-H2AX was formed as foci and did not coincide with Ac-H3. Nicotinic receptor antagonist methyllycaconitine inhibited Ac-H3 and also γ-H2AX. Phosphoinositide-3-kinase (PI3K)/Akt inhibitors, LY294002, wortmannin, and GSK690693, also suppressed both Ac-H3 and γ-H2AX, whereas KU-55933, an inhibitor of ataxia telangiectasia mutated (ATM) upstream of γ-H2AX, inhibited γ-H2AX but not Ac-H3. These results suggested that binding of NNK to the nicotinic acetylcholine receptor (α7nAChR) activated the PI3K/Akt pathway, resulting in Ac-H3. The activated pathway leading to Ac-H3 enhanced γ-H2AX, suggesting that NNK-induced DNA damage is impacted by the α7nAChR-mediated signal transduction pathway.
Assuntos
Histonas/metabolismo , Nitrosaminas/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor Nicotínico de Acetilcolina alfa7/metabolismo , Células A549 , Acetilação/efeitos dos fármacos , Cromonas/farmacologia , Relação Dose-Resposta a Droga , Histonas/antagonistas & inibidores , Histonas/biossíntese , Humanos , Morfolinas/farmacologia , Oxidiazóis/farmacologia , Pironas/farmacologia , Células Tumorais Cultivadas , Wortmanina/farmacologiaRESUMO
Solar UV radiation consists of both UVA and UVB. The wavelength-specific molecular responses to UV radiation have been studied, but the interaction between UVA and UVB has not been well understood. In this study, we found that long-wavelength UVA, UVA1, augmented UVB-induced cell death, and examined the underlying mechanisms. Human keratinocytes HaCaT were exposed to UVA1, followed by UVB irradiation. Irradiation by UVA1 alone showed no effect on cell survival, whereas the UVA1 pre-irradiation remarkably enhanced UVB-induced cell death. UVA1 delayed the repair of pyrimidine dimers formed by UVB and the accumulation of nucleotide excision repair (NER) proteins to damaged sites. Gap synthesis during NER was also decreased, suggesting that UVA1 delayed NER, and unrepaired pyrimidine dimers and single-strand breaks generated in the process of NER were left behind. Accumulation of this unrepaired DNA damage might have led to the formation of DNA double-strand breaks (DSBs), as was detected using gel electrophoresis analysis and phosphorylated histone H2AX assay. Combined exposure enhanced the ATM-Chk2 signaling pathway, but not the ATR-Chk1 pathway, confirming the enhanced formation of DSBs. Moreover, UVA1 suppressed the UVB-induced phosphorylation of Akt, a survival signal pathway. These results indicated that UVA1 influenced the repair of UVB-induced DNA damage, which resulted in the formation of DSBs and enhanced cell death, suggesting the risk of simultaneous exposure to high doses of UVA1 and UVB.
Assuntos
Queratinócitos/patologia , Raios Ultravioleta , Morte Celular/efeitos da radiação , Sobrevivência Celular/efeitos da radiação , Células Cultivadas , Quebras de DNA de Cadeia Dupla/efeitos da radiação , Humanos , Queratinócitos/efeitos da radiaçãoRESUMO
The Japanese Ministry of Health, Labour, and Welfare recently reported an outbreak of bladder cancer among workers who handled aromatic amines in Japan. 2,4-dimethylaniline (2,4-DMA) is one of the chemicals that workers are considered to have the most opportunities to be exposed. Genotoxic events are known to be crucial steps in the initiation of cancer. However, studies on the genotoxicity of 2,4-DMA are limited, particularly studies investigating the mechanism behind the genotoxicity by 2,4-DMA are completely lacking. We examined genotoxic properties of 2,4-DMA using phosphorylated histone H2AX (γ-H2AX), a sensitive and reliable marker of DNA damage, in cultured human urothelial and hepatic cells. Our results clearly showed that 2,4-DMA at a concentration range of 1-10 mM generates γ-H2AX in both cell lines, indicating that 2,4-DMA is genotoxic. During mechanistic investigation, we found that 2,4-DMA boosts intracellular reactive oxygen species, an effect clearly attenuated by disulfiram, a strong inhibitor of cytochrome P450 2E1 (CYP2E1). In addition, CYP2E1 inhibitors and the antioxidant, N-acetylcysteine, also attenuated γ-H2AX generation following exposure to 2,4-DMA. Collectively, these results suggest that γ-H2AX is formed following exposure to 2,4-DMA via reactive oxygen species produced by CYP2E1-mediated metabolism. Continuous exposure to genotoxic aromatic amines such as 2,4-DMA over a long period of time may have contributed to the development of bladder cancer. Our results provide important insights into the carcinogenicity risk of 2,4-DMA in occupational bladder cancer outbreaks at chemical plants in Japan.
Assuntos
Compostos de Anilina/toxicidade , Família 2 do Citocromo P450/metabolismo , Hepatócitos/efeitos dos fármacos , Histonas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Ciclo Celular/efeitos dos fármacos , Células Cultivadas , Inibidores do Citocromo P-450 CYP2E1/farmacologia , Quebras de DNA de Cadeia Dupla/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Hepatócitos/metabolismo , Humanos , Fosforilação/efeitos dos fármacos , Ureter/citologiaRESUMO
Trichloroethylene (TCE), a chlorinated hydrocarbon, was recently reclassified as a human carcinogen by the International Agency for Research on Cancer. Genotoxic events are known to be crucial steps in the initiation of cancer. The genotoxic properties of TCE have been examined in many studies using a standard battery of genotoxicity tests both in vitro and in vivo. However, consistent results have not been obtained, and studies investigating the mechanism behind the genotoxicity of this compound are lacking. In the present study, we examined the genotoxicity of TCE by assessing phosphorylated histone H2AX (γ-H2AX), a new sensitive and reliable marker of DNA damage, in WRL-68 cells, cultured human hepatocytes and mouse livers. Our results showed that TCE exposure results in the generation of γ-H2AX, both in vitro and in vivo. By investigating the in vitro mechanism, we found that TCE increases the levels of intracellular reactive oxygen species (ROS) and that this increase in ROS levels is attenuated in the presence of disulfiram, a specific cytochrome P450 2E1 (CYP2E1) inhibitor. Furthermore, γ-H2AX induced by TCE was also attenuated by CYP2E1 inhibitors and the antioxidant N-acetylcysteine. These results suggested that ROS, produced via cytochrome P450 2E1-mediated metabolic processing, is a major causal factor for γ-H2AX generation upon exposure to TCE.
Assuntos
Carcinógenos/toxicidade , Citocromo P-450 CYP2E1/metabolismo , Quebras de DNA de Cadeia Dupla , Hepatócitos/efeitos dos fármacos , Histonas/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Tricloroetileno/toxicidade , Animais , Antioxidantes/farmacologia , Linhagem Celular , Inibidores do Citocromo P-450 CYP2E1/farmacologia , Quebras de DNA de Cadeia Dupla/efeitos dos fármacos , Hepatócitos/enzimologia , Hepatócitos/patologia , Humanos , Masculino , Camundongos , Camundongos da Linhagem 129 , Fosforilação , Espécies Reativas de Oxigênio/metabolismo , Medição de RiscoRESUMO
Post-translational modifications in histones have been associated with cancer. Although cigarette sidestream smoke (CSS) as well as mainstream smoke are carcinogens, the relationship between carcinogenicity and histone modifications has not yet been clarified. Here, we demonstrated that CSS induced phosphorylation of histones, involving a carcinogenic process. Treatment with CSS markedly induced the phosphorylation of histone H3 at serine 10 and 28 residues (H3S10 and H3S28), which was independent from the cell cycle, in the human pulmonary epithelial cell model, A549 and normal human lung fibroblasts, MRC-5 and WI-38. Using specific inhibitors and small interfering RNA, the phosphorylation of H3S10 was found to be mediated by c-jun N-terminal kinase (JNK) and phosphoinositide 3-kinase (PI3K)/Akt pathways. These pathways were different from that of the CSS-induced phosphorylation of histone H2AX (γ-H2AX) mediated by Ataxia telangiectasia-mutated (ATM) and ATM-Rad3-related (ATR) protein kinases. A chromatin immunoprecipitation assay revealed that the phosphorylation of H3S10 was increased in the promoter sites of the proto-oncogenes, c-fos and c-jun, which indicated that CSS plays a role in tumor promotion. Because the phosphorylation of H3S10 was decreased in the aldehyde-removed CSS and was significantly induced by treatment with formaldehyde, aldehydes are suspected to partially contribute to this phosphorylation. These findings suggested that any chemicals in CSS, including aldehydes, phosphorylate H3S10 via JNK and PI3K/Akt pathways, which is different from the DNA damage response, resulting in tumor promotion.
Assuntos
Regulação Neoplásica da Expressão Gênica , Histonas/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proto-Oncogenes/genética , Transdução de Sinais , Poluição por Fumaça de Tabaco/efeitos adversos , Linhagem Celular Tumoral , Dano ao DNA , Humanos , Sistema de Sinalização das MAP Quinases , Neoplasias/etiologia , Fosforilação , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-jun/genética , Ativação TranscricionalRESUMO
Evidence is accumulating indicating that UVA (320-400 nm ultraviolet light) plays an important role in photo-carcinogenesis. UVA is thought to produce reactive oxygen species in irradiated cells through photo-activation of inherent photosensitizers, and was recently reported to cause DNA double-strand breaks (DSBs) in exposed cells. We have investigated the involvement of UVA in mutations and DNA damage in somatic cells using Drosophila melanogaster larvae. Using the Okazaki Large Spectrograph, we previously observed that longer wavelength UVA (>330 nm) was more mutagenic in post-replication repair-deficient D. melanogaster (mei-41) than in the nucleotide excision repair-deficient strain (mei-9). LED-light has recently been developed as a high-dose-rate UVA source. LED-UVA light (365 nm) was also more mutagenic in mei-41 than in mei-9. The mei-41 gene was shown to be an orthologue of the human ATR gene, which is involved in the repair of DSBs through phosphorylation of histone H2AX. In order to estimate the extent to which oxidative damage contributes to mutation, we established a new D. melanogaster strain (urate-null mutant) that is sensitive to oxidative damage and has a marker to detect somatic cell mutations. When somatic cell mutations were examined using this strain, LED-UVA was mutagenic in the urate-null strain at doses that were non-mutagenic in the urate-positive strain. In an effort to investigate the generation of DSBs, we examined the presence of phosphorylated histone H2AvD (H2AX D. melanogaster homologue). At high doses of LED-UVA (>800 kJ m(-2)), levels of phosphorylated H2AvD (γ-H2AvD) increased significantly in the urate-null strain. Moreover, the level of γ-H2AvD increased in the excision repair-deficient strain but not in the ATR-deficient strain following UVA-irradiation. These results supported the notion that the generation of γ-H2AvD was mediated by the function of the mei-41 gene. It was reported that ATR functions on DSB repair in D. melanogaster. Taken together, we propose a possible pathway for UVA-induced mutation, whereby DNA double-strand breaks resulting from oxidative stress might be responsible for UVA-induced mutation in somatic cells of D. melanogaster larvae.
Assuntos
Quebras de DNA de Cadeia Dupla/efeitos da radiação , Raios Ultravioleta , Animais , Proteínas de Ciclo Celular/genética , Reparo do DNA , Proteínas de Drosophila/genética , Drosophila melanogaster/crescimento & desenvolvimento , Histonas/metabolismo , Larva/genética , Larva/efeitos da radiação , Mutação , Proteínas Nucleares/genética , Estresse Oxidativo/efeitos da radiação , Fosforilação/efeitos da radiação , Proteínas Serina-Treonina Quinases/genéticaRESUMO
Nonylphenolpolyethoxylates (NPEOs) are non-ionic surfactants widely used for industrial and household purposes. In actual environments, NPEOs can be biodegraded, but the products are reported to be more persistent and toxic than the parent compounds. NPEOs are also exposed to sunlight and degraded. Studies on the photodegradation of NPEOs have focused mainly on chemical changes after exposure to light. Toxic changes of photodegraded products correlating to the chemical changes are not completely understood. In this study, we examined the genotoxicity of UVB-irradiated NPEOs having ethylene oxide units 15 and 70 in a human breast adenocarcinoma cell line, MCF-7, based on the phosphorylation of histone H2AX (γ-H2AX), a sensitive marker for DNA damage. We clarified that UVB irradiation drastically changed the genotoxic potential of NPEOs: NPEO(15)'s ability to generate γ-H2AX was significantly reduced, whereas non-genotoxic NPEO(70) became able to generate γ-H2AX. Flow cytometric analysis showed that the γ-H2AX generated by UVB-irradiated NPEO(70)was produced independent of cell cycle phases. In addition, its production involved the activation of ATM or DNA-PK, a general signalling pathway in response to DNA double strand breaks. High-performance liquid chromatography analysis indicated that the formation of NPEO intermediates with a short side-chain like NPEO(15) was the cause of the γ-H2AX generation. This study suggests the importance of taking the genotoxicity of photodegraded intermediates into consideration when conducting risk assessments of environmental pollutants.
Assuntos
Histonas/metabolismo , Tensoativos/efeitos da radiação , Tensoativos/toxicidade , Raios Ultravioleta , Biomarcadores/metabolismo , Linhagem Celular Tumoral , Cromatografia Líquida de Alta Pressão , Dano ao DNA/efeitos dos fármacos , Feminino , Citometria de Fluxo , Humanos , Testes de Mutagenicidade/métodos , Fosforilação , Espécies Reativas de Oxigênio/metabolismo , Tensoativos/químicaRESUMO
Medical students and junior residents have five concerns about general medicine training, and senior residents and young academic generalists respond to these concerns. We hope that this paper will help to dispel some common concerns for those who wish to become specialists in general medicine.
RESUMO
OBJECTIVES: Crystalline silica, which is a causative agent of silicosis (an occupational disease), is manufactured in a variety of products (particles) with different particle characteristics, such as size and surface properties. In Japan, the products are currently uniformly controlled as crystalline silica, which is a substance subject to labeling and notification requirements. However, since the toxicity of silica particles reportedly varies depending on its characteristics, businesses are encouraged to conduct appropriate risk assessments for each product to prevent silicosis. Recently, silica particles have been reported to induce lysosomal membrane damage, leading to the activation of proinflammatory factors. An indirect method to evaluate lysosomal membrane damage known as the erythrocyte hemolysis assay, in which the erythrocyte membrane is assumed to be the lysosomal membrane, was performed. This study aimed to examine the possibility of constructing a screening system for proinflammatory potential prediction of silica particles based on their erythrocyte hemolytic activity. METHODS: Hemolysis assays were performed on the silica particles with different sizes, crystallinity, and surface functional groups using the erythrocytes from a healthy volunteer. Additionally, the hemolytic activity of other element particles was compared with that of the silica particles, and 27 types of commercially available crystalline silica particle products underwent screening trials. RESULTS: The hemolytic activity of silica particles was higher in crystalline than that in amorphous and increased with the decreasing size. The hemolytic reaction was particular to silica particles and rarely occurred in particles of other elements. Moreover, the hemolytic activity was significantly suppressed if the silica particles surface was modified with metal ions (Fe3+, Al3+). The hemolytic activities of the crystalline silica products used industrially significantly differed. CONCLUSIONS: This study revealed that particle properties, such as size, crystallinity, and surface functional groups, affect the hemolytic activity of silica particles. Particularly, the surface functional groups (silanol groups) that are unique to silica particles were considered to be strongly involved in hemolytic activities. Since grading the commercially available crystalline silica particle products based on the hemolytic rate was possible, hemolytic activity was suggested to be an evaluation index for predicting the proinflammatory potential of silica particles.
Assuntos
Dióxido de Silício , Silicose , Humanos , Dióxido de Silício/toxicidade , Dióxido de Silício/química , Hemólise , Membrana Eritrocítica , Eritrócitos , Tamanho da PartículaRESUMO
A typical antioxidant, N-acetyl-L-cysteine (NAC) generally protects cells from oxidative damage induced by reactive oxygen species (ROS). 9,10-Phenanthrenequinone (9,10-PQ), a major quinone in diesel exhaust particles, produces ROS in redox cycling following two-electron reduction by NAD(P)H:quinone oxidoreductase 1 (NQO1), which has been considered as a cause of its cyto- and genotoxicity. In this study, we show that NAC unexpectedly augments the toxicity of 9,10-PQ in cells with low NQO1 activity. In four human skin cell lines, the expression and the activity of NQO1 were lower than in human adenocarcinoma cell lines, A549 and MCF7. In the skin cells, the cytotoxicity of 9,10-PQ was significantly enhanced by addition of NAC. The formation of DNA double strand breaks accompanying phosphorylation of histone H2AX, was also remarkably augmented. On the other hand, the cyto- and genotoxicity were suppressed by addition of NAC in the adenocarcinoma cells. Two contrasting experiments: overexpression of NQO1 in CHO-K1 cells which originally expressed low NQO1 levels, and knock-down of NQO1 in the adenocarcinoma cell line A549 by transfection of RNAi, also showed that NAC suppressed 9,10-PQ-induced toxicity in cell lines expressing high NQO1 activity and enhanced it in cell lines with low NQO1 activity. The results suggested that dual effects of NAC on the cyto- and genotoxicity of 9,10-PQ were dependent on tissue-specific NQO1 activity.
Assuntos
Acetilcisteína/farmacologia , Fibroblastos/efeitos dos fármacos , Queratinócitos/efeitos dos fármacos , NAD(P)H Desidrogenase (Quinona)/metabolismo , Fenantrenos/toxicidade , Animais , Linhagem Celular , Cricetinae , Fibroblastos/metabolismo , Sequestradores de Radicais Livres/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Humanos , Queratinócitos/metabolismoRESUMO
We recently clarified that the side-scatter(ed) light (SSC) of flow cytometry (FCM) could be used as a guide to measure the uptake potential of nanoparticles [ Suzuki et al. Environ. Sci. Technol. 2007 , 41 , 3018 - 3024 ]. In this paper, the method was improved so as to be able to determine simultaneously the uptake potential of nanoparticles and the production of reactive oxygen species (ROS), and correlations with genotoxicity were evaluated. In the FCM analysis, SSC and fluorescence of 6-carboxy-2,7'-diclorodihydrofluorescein diacetate, di(acetoxy ester) based on ROS production were concurrently detected after treatments with ZnO, CuO, Fe(3)O(4), TiO(2), and Ag nanoparticles. The ZnO and CuO nanoparticles caused high ROS production, which was more significant in the cells with higher SSC intensity. The increase of SSC intensity was more significant for TiO(2) than ZnO and CuO, whereas ROS production was higher for ZnO and CuO than TiO(2), suggesting that the extent of ROS production based on the uptake of nanoparticles differed with each kind of nanoparticle. ROS production was correlated with generation of the phosphorylated histone H2AX (γ-H2AX), a marker of DNA damage, and an antioxidant, n-acetylcysteine, could partially suppress the γ-H2AX. This method makes it possible to predict not only uptake potential but also genotoxicity.
Assuntos
Citometria de Fluxo/métodos , Luz , Mutagênicos/toxicidade , Nanopartículas/toxicidade , Espécies Reativas de Oxigênio/metabolismo , Espalhamento de Radiação , Animais , Células CHO , Cricetinae , Quebras de DNA de Cadeia Dupla/efeitos dos fármacos , Fluoresceína/metabolismo , Fluorescência , Histonas/metabolismo , Testes de Mutagenicidade , Fatores de TempoRESUMO
Nonylphenol polyethoxylates (NPEOs) are non-ionic surfactants widely used for industrial and household purposes. Since biodegraded short chain NPEOs were reported to elicit estrogenic activity in organisms, numerous studies have been carried out to assess the endocrine-disrupting potential of NPEOs; however, the genotoxicity of the compounds is not fully known, let alone the relationship between the genotoxic potential and number of ethylene oxide (EO) units of NPEOs. In this study, we examined the genotoxicity of NPEO(n) having various EO units (n=0, 5, 10, 15, 20, 30, 40 and 70) in a human breast adenocarcinoma cell line, MCF-7, based on the phosphorylation of histone H2AX (γ-H2AX), recently regarded as a sensitive marker for DNA damage. We clarified that NPEOs have the ability to form γ-H2AX via activation of ATM or DNA-PK, a general signaling pathway in response to DSBs, and this ability was strongly dependent on the number of EO units, that is, NPEO(0-15) having smaller numbers of EO units more readily generated γ-H2AX. Flow cytometric analysis revealed that the generation of γ-H2AX was independent of cell cycle phases. Although the mechanism by which the NPEOs generated γ-H2AX was not able to be elucidated in the present study, it was clear that the involvement of reactive oxygen species and apoptotic DNA fragmentation were not causal factors. The generation of γ-H2AX means the formation of DSBs, the worst type of DNA damage. The results indicated that attention should be paid to degradated short chain NPEOs and their genotoxicity.
Assuntos
Apoptose/efeitos dos fármacos , Quebras de DNA de Cadeia Dupla , Etilenoglicóis/toxicidade , Histonas/metabolismo , Mutagênicos/toxicidade , Ciclo Celular , Linhagem Celular Tumoral , Etilenoglicóis/química , Óxido de Etileno , Feminino , Humanos , Fosforilação , Espécies Reativas de OxigênioRESUMO
With the rising use of nano-sized particles in nanotechnology, harmful effects of TiO(2) particles which have been recognized as a safe material, are of increasing concern. In this study, we examined the genotoxicity of two different sized TiO(2) particles in the lung adenocarcinoma epithelial cell line A549 based on the phosphorylation of histone H2AX (γ-H2AX), recently regarded as a sensitive marker for DNA damage. TiO(2) particles generated γ-H2AX, which was more remarkable with the smaller particles. Flow cytometric analysis showed that the generation was independent of cell cycle phases and cells which incorporated larger amounts of TiO(2) particles had more significant γ-H2AX. Although there are some reports that the incorporation of nanomaterials into cells generates reactive oxygen species (ROS), the level of ROS was low even if large amounts of TiO(2) particles were taken-up. In addition, the generation of γ-H2AX by TiO(2) particles was more significant than that after treatment with hydrogen peroxide. On the other hand, the generation of γ-H2AX was attenuated by coating the surface of TiO(2) particles with bovine serum albumin. These results suggested that smaller TiO(2) particles were easy to incorporate into cells and generated cell cycle phase-independent γ-H2AX, which was dependent on the condition of the TiO(2) surface but not on the formation of ROS.
Assuntos
Dano ao DNA/efeitos dos fármacos , Histonas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Titânio/metabolismo , Biomarcadores/análise , Ciclo Celular , Linhagem Celular Tumoral , Humanos , Nanopartículas , Tamanho da Partícula , Fosforilação , Titânio/químicaRESUMO
OBJECTIVES: Beryllium is primarily used in its metallic form, in alloys, or in beryllium oxide ceramics. Its physical and mechanical properties make it useful for many applications across a range of industries. Because beryllium is recognized as a sensitizing and carcinogenic agent, the management of occupational health for workers who may be occupationally exposed to beryllium has long been an important issue in the world. Under these circumstances, the U.S. Occupational Safety and Health Administration (OSHA) had published a rule in January 2017, to prevent the development of chronic beryllium disease and lung cancer. This rule strengthens the regulations governing the use of beryllium and its compounds. With the announcement of the OSHA rule in January 2017, the purpose of this study is to gain insight into the health problems and industrial hygiene associated with the use of beryllium and share the issues related to the management of occupational health for persons working with beryllium in Japan. METHODS: We collected information regarding the beryllium industry, beryllium exposure, beryllium-induced health disorders, OSHA rule of January 2017, and regulations for beryllium use in Japan. After reviewing them, we discussed the issues concerning occupational health management of workers exposed to beryllium in Japan. RESULTS: It has been reconfirmed that in recent years, the most serious health problem due to beryllium exposure is chronic beryllium disease caused by beryllium sensitization. Management of occupational health that emphasizes reduction of beryllium sensitization and early detection of beryllium-sensitized workers is important. CONCLUSIONS: It was suggested that the following should be considered as the issues of management of occupational health of workers exposed to beryllium in Japan: (1) Collect epidemiologic data on health hazards from beryllium exposure in Japan. (2) Review the diagnostic items of special medical check-ups. (3) Review the definition of beryllium and its compounds in the Ordinance on Prevention of Hazards due to Specified Chemical Substances.
Assuntos
Beriliose/etiologia , Beriliose/prevenção & controle , Berílio/efeitos adversos , Doenças Profissionais/etiologia , Doenças Profissionais/prevenção & controle , Exposição Ocupacional/efeitos adversos , Saúde Ocupacional , Local de Trabalho , Beriliose/diagnóstico , Beriliose/epidemiologia , Berílio/análise , Doença Crônica , Feminino , Humanos , Japão , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/etiologia , Neoplasias Pulmonares/prevenção & controle , Masculino , Doenças Profissionais/diagnóstico , Doenças Profissionais/epidemiologia , Exposição Ocupacional/análise , Saúde Ocupacional/tendênciasRESUMO
Polycyclic aromatic hydrocarbons (PAHs), wide-spread mutagenic and carcinogenic environmental pollutants, are consistently exposed to sunlight in the environment. Our previous paper showed that benzo[a]pyrene (BaP) exposed to solar-simulated light (SSL) induced phosphorylation of histone H2AX (γ-H2AX) [T. Toyooka, G. Ohnuki, Y. Ibuki, Solar-simulated light-exposed benzo[a]pyrene induces phosphorylation of histone H2AX, Mutat. Res. 650 (2008) 132-139], a marker of DNA double strand breaks. In this study, we found the ultraviolet B (UVB) region of SSL to produce photomodified BaP with high cytotoxicity and the ability to generate γ-H2AX. Degradation of BaP by SSL, resulting in an increase in cytotoxicity and the generation of γ-H2AX, was decreased by UVB-masking using a glass plate. Exposure to UVB itself increased the cytotoxicity of BaP and amount of γ-H2AX generated. Other PAHs, 1,2-benzoanthracene and 1,2:5,6-dibenzoanthracene, which absorb UVB, also showed enhanced cytotoxicity and the promoted the generation of γ-H2AX after exposure to SSL, whereas naphthalene and chrysene, which have low absorption in the UVB region, did not. These findings suggested that UVB is important for the degradation of PAHs having absorbance in this region, but that the production of genotoxic intermediates during the degradation process needs to be considered. UVB is a two-edged blade in environments, effectively degrading toxic chemicals but also producing genotoxic compounds as reactive intermediates.
Assuntos
Benzo(a)pireno/efeitos da radiação , Benzo(a)pireno/toxicidade , Histonas/metabolismo , Luz Solar , Raios Ultravioleta/efeitos adversos , Animais , Benzo(a)pireno/metabolismo , Células CHO , Cricetinae , Cricetulus , Histonas/análise , Mutagênicos/metabolismo , FosforilaçãoRESUMO
A variety of chemotherapeutic drugs, e.g., etoposide and bleomycin, are widely used in clinical practice to treat many types of animal malignancies. In the clinical situation, cellular resistance to chemotherapy is a significant component of tumor treatment failure. A variety of DNA repair factors, e.g., Ku80, might be a key contributor to chemoresistance to anticancer agents. In both cancer and normal cells, Ku80 plays a key role as a sensor of DNA double-strand break (DSB) induced by treatment with some chemotherapeutic drugs. Although the localization and mobility of Ku80 play a key role in regulating the physiological function of Ku80, it is not clear whether those of Ku80 are affected after treatment with chemotherapeutic drugs. We examined the localization and mobility of Ku80 in living hamster cells with or without DSBs, which were induced by treatment with chemotherapeutic drugs. Our data showed that Ku80, in contrast to H2AX, is highly mobile in the nuclei. We found that before and after the induction of DNA damage by treatment with etoposide or bleomycin, a major portion of Ku80 is exchanged by the same kinetics in the nuclei of interphase cells. These results suggest that the mobility of a major portion of Ku80 is not affected by DNA DSBs in order to find other DSBs. In addition, the information would be worthy to develop some new chemotherapeutic drugs to treat many types of animal malignancies.
Assuntos
Antígenos Nucleares/metabolismo , Antineoplásicos/farmacologia , Cricetinae/genética , Quebras de DNA de Cadeia Dupla/efeitos dos fármacos , Proteínas de Ligação a DNA/metabolismo , Animais , Antígenos Nucleares/genética , Bleomicina/farmacologia , Células CHO/citologia , Células CHO/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Cricetulus , Dano ao DNA/efeitos dos fármacos , Reparo do DNA , Proteínas de Ligação a DNA/genética , Etoposídeo/farmacologia , Proteínas de Fluorescência Verde/metabolismo , Histonas/imunologia , Cinética , Autoantígeno Ku , Microscopia Confocal/métodosRESUMO
Recently, it was reported that ten cases of bladder cancer occurred among employees, who handled several kinds of aromatic amines, at a Japanese chemical plant. The common aromatic amines were identified as ortho-toluidine, para-toluidine, aniline, ortho-chloroaniline, ortho-anisidine, and 2,4-dimethylaniline. All of these aromatic amines, except ortho-chloroaniline, have been found to be carcinogenic in animals and/or humans. Genotoxic events are known to be crucial steps in the initiation of cancer; information on the genotoxicity of these aromatic amines is insufficient and consistent results have not been obtained. In this study, we examined the genotoxicity of the six different aromatic amines associated with bladder cancer by assessing phosphorylated histone H2AX (γ-H2AX) in a cultured human urothelial cell line, 1T1. We showed that all six aromatic amines generated γ-H2AX. In addition, the γ-H2AX-inducing potential of these six aromatic amines was distinctly different; ortho-chloroaniline and 2,4-dimethylaniline showed particularly high potential, followed by ortho-toluidine, ortho-anisidine, para-toluidine â aniline. The findings of this study may provide important information for the risk assessment of chemicals and for interpreting epidemiological studies on occupational bladder cancer.
Assuntos
Compostos de Anilina/toxicidade , Dano ao DNA , Histonas/metabolismo , Mutagênicos/toxicidade , Urotélio/citologia , Caspase 3/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , HumanosRESUMO
Cigarette sidestream smoke (CSS) is a widespread environmental pollutant having highly genotoxic potency. In spite of the overwhelming evidence that CSS induces a wide range of DNA damage such as oxidative base damage and DNA adducts, evidence that CSS can result in DNA double strand breaks (DSBs) is little. In this study, we showed that CSS generated phosphorylated histone H2AX (gamma-H2AX), recently considered as a sensitive marker of the generation of DSBs, in a human pulmonary epithelial cell model, A549. Treatment with CSS drastically induced discrete foci of gamma-H2AX within the nucleus in a dose-dependent manner. CSS increased intracellular oxidation, and N-acetylcysteine (NAC), an antioxidant, significantly attenuated the formation of gamma-H2AX, suggesting that reactive oxygen species produced from CSS partially contributed to the phosphorylation. The generation of gamma-H2AX is considered to be accompanied the induction of DSBs. CSS in fact induced DSBs, which was also inhibited by NAC. DSBs are the worst type of DNA damage, related to genomic instability and carcinogenesis. Our results would increase the evidence of the strong genotoxicity of passive smoking.